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Sample records for express distinct surface

Distinct regions in the C-Terminus required for GLP-1R cell surface expression, activity and internalisation.

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Thompson, Aiysha; Kanamarlapudi, Venkateswarlu

2015-09-15

The glucagon-like peptide-1 (GLP-1) receptor (GLP-1R), an important drug target in the treatment of type 2 diabetes, is a G-protein coupled receptor (GPCR) that mediates insulin secretion by GLP-1. The N-terminus controls GLP-1R biosynthetic trafficking to the cell surface but the C-terminus involvement in that trafficking is unknown. The aim of this study was to identify distinct regions within the C-terminal domain required for human GLP-1R (hGLP-1R) cell surface expression, activity and internalisation using a number of C-terminal deletions and site-directed mutations. The results of this study revealed that the residues 411-418 within the C-terminal domain of the hGLP-1R are critical in targeting the newly synthesised receptor to the plasma membrane. The residues 419-430 are important for cAMP producing activity of the receptor, most likely by coupling to Gαs. However, the residues 431-450 within the C-terminus are essential for agonist-induced hGLP-1R internalisation. In conclusion, these findings demonstrate the hGLP-1R has distinct regions within the C-terminal domain required for its cell surface expression, activity and agonist-induced internalisation. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
Retention in the Golgi apparatus and expression on the cell surface of Cfr/Esl-1/Glg-1/MG-160 are regulated by two distinct mechanisms.

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Miyaoka, Yuichiro; Kato, Hidenori; Ebato, Kazuki; Saito, Shigeru; Miyata, Naoko; Imamura, Toru; Miyajima, Atsushi

2011-11-15

Cfr (cysteine-rich fibroblast growth factor receptor) is an Fgf (fibroblast growth factor)-binding protein without a tyrosine kinase. We have shown previously that Cfr is involved in Fgf18 signalling via Fgf receptor 3c. However, as Cfr is also known as Glg (Golgi apparatus protein)-1 or MG-160 and occurs in the Golgi apparatus, it remains unknown how the distribution of Cfr is regulated. In the present study, we performed a mutagenic analysis of Cfr to show that two distinct regions contribute to its distribution and stability. First, the C-terminal region retains Cfr in the Golgi apparatus. Secondly, the Cfr repeats in the extracellular juxtamembrane region destabilizes Cfr passed through the Golgi apparatus. This destabilization does not depend on the cleavage and secretion of the extracellular domain of Cfr. Furthermore, we found that Cfr with a GPI (glycosylphosphatidylinositol) anchor was predominantly expressed on the cell surface in Ba/F3 cells and affected Fgf18 signalling in a similar manner to the full-length Cfr, indicating that the interaction of Cfr with Fgfs on the cell surface is important for its function in Fgf signalling. These results suggest that the expression of Cfr in the Golgi apparatus and on the plasma membrane is finely tuned through two distinct mechanisms for exhibiting different functions.
The influence of context on distinct facial expressions of disgust.

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Reschke, Peter J; Walle, Eric A; Knothe, Jennifer M; Lopez, Lukas D

2018-06-11

Face perception is susceptible to contextual influence and perceived physical similarities between emotion cues. However, studies often use structurally homogeneous facial expressions, making it difficult to explore how within-emotion variability in facial configuration affects emotion perception. This study examined the influence of context on the emotional perception of categorically identical, yet physically distinct, facial expressions of disgust. Participants categorized two perceptually distinct disgust facial expressions, "closed" (i.e., scrunched nose, closed mouth) and "open" (i.e., scrunched nose, open mouth, protruding tongue), that were embedded in contexts comprising emotion postures and scenes. Results demonstrated that the effect of nonfacial elements was significantly stronger for "open" disgust facial expressions than "closed" disgust facial expressions. These findings provide support that physical similarity within discrete categories of facial expressions is mutable and plays an important role in affective face perception. (PsycINFO Database Record (c) 2018 APA, all rights reserved).
Number of perceptually distinct surface colors in natural scenes.

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Marín-Franch, Iván; Foster, David H

2010-09-30

The ability to perceptually identify distinct surfaces in natural scenes by virtue of their color depends not only on the relative frequency of surface colors but also on the probabilistic nature of observer judgments. Previous methods of estimating the number of discriminable surface colors, whether based on theoretical color gamuts or recorded from real scenes, have taken a deterministic approach. Thus, a three-dimensional representation of the gamut of colors is divided into elementary cells or points which are spaced at one discrimination-threshold unit intervals and which are then counted. In this study, information-theoretic methods were used to take into account both differing surface-color frequencies and observer response uncertainty. Spectral radiances were calculated from 50 hyperspectral images of natural scenes and were represented in a perceptually almost uniform color space. The average number of perceptually distinct surface colors was estimated as 7.3 × 10(3), much smaller than that based on counting methods. This number is also much smaller than the number of distinct points in a scene that are, in principle, available for reliable identification under illuminant changes, suggesting that color constancy, or the lack of it, does not generally determine the limit on the use of color for surface identification.
CD30 expression defines a novel subgroup of diffuse large B-cell lymphoma with favorable prognosis and distinct gene expression signature

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Hu, Shimin; Xu-Monette, Zijun Y; Balasubramanyam, Aarthi

2013-01-01

CD30, originally identified as a cell-surface marker of Reed-Sternberg and Hodgkin cells of classical Hodgkin lymphoma, is also expressed by several types of non-Hodgkin lymphoma, including a subset of diffuse large B-cell lymphoma (DLBCL). However, the prognostic and biological importance of CD3...... value of CD30 as a therapeutic target for brentuximab vedotin in ongoing successful clinical trials, it seems appropriate to consider CD30(+) DLBCL as a distinct subgroup of DLBCL....
Reconstructing dynamic mental models of facial expressions in prosopagnosia reveals distinct representations for identity and expression.

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Richoz, Anne-Raphaëlle; Jack, Rachael E; Garrod, Oliver G B; Schyns, Philippe G; Caldara, Roberto

2015-04-01

The human face transmits a wealth of signals that readily provide crucial information for social interactions, such as facial identity and emotional expression. Yet, a fundamental question remains unresolved: does the face information for identity and emotional expression categorization tap into common or distinct representational systems? To address this question we tested PS, a pure case of acquired prosopagnosia with bilateral occipitotemporal lesions anatomically sparing the regions that are assumed to contribute to facial expression (de)coding (i.e., the amygdala, the insula and the posterior superior temporal sulcus--pSTS). We previously demonstrated that PS does not use information from the eye region to identify faces, but relies on the suboptimal mouth region. PS's abnormal information use for identity, coupled with her neural dissociation, provides a unique opportunity to probe the existence of a dichotomy in the face representational system. To reconstruct the mental models of the six basic facial expressions of emotion in PS and age-matched healthy observers, we used a novel reverse correlation technique tracking information use on dynamic faces. PS was comparable to controls, using all facial features to (de)code facial expressions with the exception of fear. PS's normal (de)coding of dynamic facial expressions suggests that the face system relies either on distinct representational systems for identity and expression, or dissociable cortical pathways to access them. Interestingly, PS showed a selective impairment for categorizing many static facial expressions, which could be accounted for by her lesion in the right inferior occipital gyrus. PS's advantage for dynamic facial expressions might instead relate to a functionally distinct and sufficient cortical pathway directly connecting the early visual cortex to the spared pSTS. Altogether, our data provide critical insights on the healthy and impaired face systems, question evidence of deficits
Immunophenotyping of Waldenstroms macroglobulinemia cell lines reveals distinct patterns of surface antigen expression: potential biological and therapeutic implications.

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Aneel Paulus

Full Text Available Waldenströms macroglobulinemia (WM is a subtype of Non-Hodgkin's lymphoma in which the tumor cell population is markedly heterogeneous, consisting of immunoglobulin-M secreting B-lymphocytes, plasmacytoid lymphocytes and plasma cells. Due to rarity of disease and scarcity of reliable preclinical models, many facets of WM molecular and phenotypic architecture remain incompletely understood. Currently, there are 3 human WM cell lines that are routinely used in experimental studies, namely, BCWM.1, MWCL-1 and RPCI-WM1. During establishment of RPCI-WM1, we observed loss of the CD19 and CD20 antigens, which are typically present on WM cells. Intrigued by this observation and in an effort to better define the immunophenotypic makeup of this cell line, we conducted a more comprehensive analysis for the presence or absence of other cell surface antigens that are present on the RPCI-WM1 model, as well as those on the two other WM cell lines, BCWM.1 and MWCL-1. We examined expression of 65 extracellular and 4 intracellular antigens, comprising B-cell, plasma cell, T-cell, NK-cell, myeloid and hematopoietic stem cell surface markers by flow cytometry analysis. RPCI-WM1 cells demonstrated decreased expression of CD19, CD20, and CD23 with enhanced expression of CD28, CD38 and CD184, antigens that were differentially expressed on BCWM.1 and MWCL-1 cells. Due to increased expression of CD184/CXCR4 and CD38, RPCI-WM1 represents a valuable model in which to study the effects anti-CXCR4 or anti-CD38 targeted therapies that are actively being developed for treatment of hematologic cancers. Overall, differences in surface antigen expression across the 3 cell lines may reflect the tumor clone population predominant in the index patients, from whom the cell lines were developed. Our analysis defines the utility of the most commonly employed WM cell lines as based on their immunophenotype profiles, highlighting unique differences that can be further studied for
Molecular polymorphism of a cell surface proteoglycan: distinct structures on simple and stratified epithelia.

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Sanderson, R D; Bernfield, M

1988-12-01

Epithelial cells are organized into either a single layer (simple epithelia) or multiple layers (stratified epithelia). Maintenance of these cellular organizations requires distinct adhesive mechanisms involving many cell surface molecules. One such molecule is a cell surface proteoglycan, named syndecan, that contains both heparan sulfate and chondroitin sulfate chains. This proteoglycan binds cells to fibrillar collagens and fibronectin and thus acts as a receptor for interstitial matrix. The proteoglycan is restricted to the basolateral surface of simple epithelial cells, but is located over the entire surface of stratified epithelial cells, even those surfaces not contacting matrix. We now show that the distinct localization in simple and stratified epithelia correlates with a distinct proteoglycan structure. The proteoglycan from simple epithelia (modal molecular size, 160 kDa) is larger than that from stratified epithelia (modal molecular size, 92 kDa), but their core proteins are identical in size and immunoreactivity. The proteoglycan from simple epithelia has more and larger heparan sulfate and chondroitin sulfate chains than the proteoglycan from stratified epithelia. Thus, the cell surface proteoglycan shows a tissue-specific structural polymorphism due to distinct posttranslational modifications. This polymorphism likely reflects distinct proteoglycan functions in simple and stratified epithelia, potentially meeting the different adhesive requirements of the cells in these different organizations.
Elucidating distinct ion channel populations on the surface of hippocampal neurons via single-particle tracking recurrence analysis

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Sikora, Grzegorz; Wyłomańska, Agnieszka; Gajda, Janusz; Solé, Laura; Akin, Elizabeth J.; Tamkun, Michael M.; Krapf, Diego

2017-12-01

Protein and lipid nanodomains are prevalent on the surface of mammalian cells. In particular, it has been recently recognized that ion channels assemble into surface nanoclusters in the soma of cultured neurons. However, the interactions of these molecules with surface nanodomains display a considerable degree of heterogeneity. Here, we investigate this heterogeneity and develop statistical tools based on the recurrence of individual trajectories to identify subpopulations within ion channels in the neuronal surface. We specifically study the dynamics of the K+ channel Kv1.4 and the Na+ channel Nav1.6 on the surface of cultured hippocampal neurons at the single-molecule level. We find that both these molecules are expressed in two different forms with distinct kinetics with regards to surface interactions, emphasizing the complex proteomic landscape of the neuronal surface. Further, the tools presented in this work provide new methods for the analysis of membrane nanodomains, transient confinement, and identification of populations within single-particle trajectories.
Aping expressions? Chimpanzees produce distinct laugh types when responding to laughter of others.

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Davila-Ross, Marina; Allcock, Bethan; Thomas, Chris; Bard, Kim A

2011-10-01

Humans have the ability to replicate the emotional expressions of others even when they undergo different emotions. Such distinct responses of expressions, especially positive expressions, play a central role in everyday social communication of humans and may give the responding individuals important advantages in cooperation and communication. The present work examined laughter in chimpanzees to test whether nonhuman primates also use their expressions in such distinct ways. The approach was first to examine the form and occurrence of laugh replications (laughter after the laughter of others) and spontaneous laughter of chimpanzees during social play and then to test whether their laugh replications represented laugh-elicited laugh responses (laughter triggered by the laughter of others) by using a quantitative method designed to measure responses in natural social settings. The results of this study indicated that chimpanzees produce laugh-elicited laughter that is distinct in form and occurrence from their spontaneous laughter. These findings provide the first empirical evidence that nonhuman primates have the ability to replicate the expressions of others by producing expressions that differ in their underlying emotions and social implications. The data further showed that the laugh-elicited laugh responses of the subjects were closely linked to play maintenance, suggesting that chimpanzees might gain important cooperative and communicative advantages by responding with laughter to the laughter of their social partners. Notably, some chimpanzee groups of this study responded more with laughter than others, an outcome that provides empirical support of a socialization of expressions in great apes similar to that of humans.
Distinct Calcium Signaling Pathways Regulate Calmodulin Gene Expression in Tobacco1

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van der Luit, Arnold H.; Olivari, Claudio; Haley, Ann; Knight, Marc R.; Trewavas, Anthony J.

1999-01-01

Cold shock and wind stimuli initiate Ca2+ transients in transgenic tobacco (Nicotiana plumbaginifolia) seedlings (named MAQ 2.4) containing cytoplasmic aequorin. To investigate whether these stimuli initiate Ca2+ pathways that are spatially distinct, stress-induced nuclear and cytoplasmic Ca2+ transients and the expression of a stress-induced calmodulin gene were compared. Tobacco seedlings were transformed with a construct that encodes a fusion protein between nucleoplasmin (a major oocyte nuclear protein) and aequorin. Immunocytochemical evidence indicated targeting of the fusion protein to the nucleus in these plants, which were named MAQ 7.11. Comparison between MAQ 7.11 and MAQ 2.4 seedlings confirmed that wind stimuli and cold shock invoke separate Ca2+ signaling pathways. Partial cDNAs encoding two tobacco calmodulin genes, NpCaM-1 and NpCaM-2, were identified and shown to have distinct nucleotide sequences that encode identical polypeptides. Expression of NpCaM-1, but not NpCaM-2, responded to wind and cold shock stimulation. Comparison of the Ca2+ dynamics with NpCaM-1 expression after stimulation suggested that wind-induced NpCaM-1 expression is regulated by a Ca2+ signaling pathway operational predominantly in the nucleus. In contrast, expression of NpCaM-1 in response to cold shock is regulated by a pathway operational predominantly in the cytoplasm. PMID:10557218
Signal peptide cleavage is essential for surface expression of a regulatory T cell surface protein, leucine rich repeat containing 32 (LRRC32).

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Chan, Derek V; Somani, Ally-Khan; Young, Andrew B; Massari, Jessica V; Ohtola, Jennifer; Sugiyama, Hideaki; Garaczi, Edina; Babineau, Denise; Cooper, Kevin D; McCormick, Thomas S

2011-05-26

Elevated numbers of regulatory T cells (T(regs)) have been implicated in certain cancers. Depletion of T(regs) has been shown to increase anti-tumor immunity. T(regs) also play a critical role in the suppression of autoimmune responses. The study of T(regs) has been hampered by a lack of adequate surface markers. Leucine Rich Repeat Containing 32 (LRRC32), also known as Glycoprotein A Repetitions Predominant (GARP), has been postulated as a novel surface marker of activated T(regs). However, there is limited information regarding the processing of LRRC32 or the regulatory phenotype and functional activity of T(regs) expressing LRRC32. Using naturally-occurring freshly isolated T(regs), we demonstrate that low levels of LRRC32 are present intracellularly prior to activation and that freshly isolated LRRC32+ T(regs) are distinct from LRRC32- T(regs) with respect to the expression of surface CD62L. Using LRRC32 transfectants of HEK cells, we demonstrate that the N-terminus of LRRC32 is cleaved prior to expression of the protein at the cell surface. Furthermore, we demonstrate using a construct containing a deleted putative signal peptide region that the presence of a signal peptide region is critical to cell surface expression of LRRC32. Finally, mixed lymphocyte assays demonstrate that LRRC32+ T(regs) are more potent suppressors than LRRC32- T(regs). A cleaved signal peptide site in LRRC32 is necessary for surface localization of native LRRC32 following activation of naturally-occurring freshly-isolated regulatory T cells. LRRC32 expression appears to alter the surface expression of activation markers of T cells such as CD62L. LRRC32 surface expression may be useful as a marker that selects for more potent T(reg) populations. In summary, understanding the processing and expression of LRRC32 may provide insight into the mechanism of action of T(regs) and the refinement of immunotherapeutic strategies aimed at targeting these cells.
Signal peptide cleavage is essential for surface expression of a regulatory T cell surface protein, leucine rich repeat containing 32 (LRRC32

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Sugiyama Hideaki

2011-05-01

Full Text Available Abstract Background Elevated numbers of regulatory T cells (Tregs have been implicated in certain cancers. Depletion of Tregs has been shown to increase anti-tumor immunity. Tregs also play a critical role in the suppression of autoimmune responses. The study of Tregs has been hampered by a lack of adequate surface markers. Leucine Rich Repeat Containing 32 (LRRC32, also known as Glycoprotein A Repetitions Predominant (GARP, has been postulated as a novel surface marker of activated Tregs. However, there is limited information regarding the processing of LRRC32 or the regulatory phenotype and functional activity of Tregs expressing LRRC32. Results Using naturally-occurring freshly isolated Tregs, we demonstrate that low levels of LRRC32 are present intracellularly prior to activation and that freshly isolated LRRC32+ Tregs are distinct from LRRC32- Tregs with respect to the expression of surface CD62L. Using LRRC32 transfectants of HEK cells, we demonstrate that the N-terminus of LRRC32 is cleaved prior to expression of the protein at the cell surface. Furthermore, we demonstrate using a construct containing a deleted putative signal peptide region that the presence of a signal peptide region is critical to cell surface expression of LRRC32. Finally, mixed lymphocyte assays demonstrate that LRRC32+ Tregs are more potent suppressors than LRRC32- Tregs. Conclusions A cleaved signal peptide site in LRRC32 is necessary for surface localization of native LRRC32 following activation of naturally-occurring freshly-isolated regulatory T cells. LRRC32 expression appears to alter the surface expression of activation markers of T cells such as CD62L. LRRC32 surface expression may be useful as a marker that selects for more potent Treg populations. In summary, understanding the processing and expression of LRRC32 may provide insight into the mechanism of action of Tregs and the refinement of immunotherapeutic strategies aimed at targeting these cells.
Genetic analysis of the SARS-coronavirus spike glycoprotein functional domains involved in cell-surface expression and cell-to-cell fusion

International Nuclear Information System (INIS)

Petit, Chad M.; Melancon, Jeffrey M.; Chouljenko, Vladimir N.; Colgrove, Robin; Farzan, Michael; Knipe, David M.; Kousoulas, K.G.

2005-01-01

The SARS-coronavirus (SARS-CoV) is the etiological agent of severe acute respiratory syndrome (SARS). The SARS-CoV spike (S) glycoprotein mediates membrane fusion events during virus entry and virus-induced cell-to-cell fusion. To delineate functional domains of the SARS-CoV S glycoprotein, single point mutations, cluster-to-lysine and cluster-to-alanine mutations, as well as carboxyl-terminal truncations were investigated in transient expression experiments. Mutagenesis of either the coiled-coil domain of the S glycoprotein amino terminal heptad repeat, the predicted fusion peptide, or an adjacent but distinct region, severely compromised S-mediated cell-to-cell fusion, while intracellular transport and cell-surface expression were not adversely affected. Surprisingly, a carboxyl-terminal truncation of 17 amino acids substantially increased S glycoprotein-mediated cell-to-cell fusion suggesting that the terminal 17 amino acids regulated the S fusogenic properties. In contrast, truncation of 26 or 39 amino acids eliminating either one or both of the two endodomain cysteine-rich motifs, respectively, inhibited cell fusion in comparison to the wild-type S. The 17 and 26 amino-acid deletions did not adversely affect S cell-surface expression, while the 39 amino-acid truncation inhibited S cell-surface expression suggesting that the membrane proximal cysteine-rich motif plays an essential role in S cell-surface expression. Mutagenesis of the acidic amino-acid cluster in the carboxyl terminus of the S glycoprotein as well as modification of a predicted phosphorylation site within the acidic cluster revealed that this amino-acid motif may play a functional role in the retention of S at cell surfaces. This genetic analysis reveals that the SARS-CoV S glycoprotein contains extracellular domains that regulate cell fusion as well as distinct endodomains that function in intracellular transport, cell-surface expression, and cell fusion
Express saccades in distinct populations: east, west, and in-between.

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Knox, Paul C; Wolohan, Felicity D A; Helmy, Mai S

2017-12-01

Express saccades are low latency (80-130 ms), visually guided saccades. While their occurrence is encouraged by the use of gap tasks (the fixation target is extinguished 200 ms prior to the saccade target appearing) and suppressed by the use of overlap tasks (the fixation target remains present when the saccade target appears), there are some healthy, adult participants, "express saccade makers" (ESMs), who persist in generating high proportions (> 30%) of express saccades in overlap conditions. These participants are encountered much more frequently in Chinese participant groups than amongst the Caucasian participants tested to date. What is not known is whether this high number of ESMs is only a feature of Chinese participant groups. More broadly, there are few comparative studies of saccade behaviour across large participant groups drawn from different populations. We, therefore, tested an independent group of 70 healthy adult Egyptian participants, using the same equipment and procedures as employed in the previous studies. Each participant was exposed to two blocks of 200 gap, and two blocks of 200 overlap trials, with block order counterbalanced. Results from the Schwartz Value Survey were used to confirm that this group of participants was culturally distinct from the Chinese and Caucasian (white British) groups tested previously. Fourteen percent (10/70) of this new group were ESMs, and the pattern of latency distribution in these ESMs was identical to that identified in the other participant groups, with a prominent peak in the express latency range in overlap conditions. Overall, we identified three modes in the distribution of saccade latency in overlap conditions, the timing of which (express peak at 110 ms, subsequent peaks at 160 and 210 ms) were strikingly consistent with our previous observations. That these behavioural patterns of saccade latency are observed consistently in large participant groups, drawn from geographically, ethnically, and
Tumour cells expressing single VEGF isoforms display distinct growth, survival and migration characteristics.

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Chryso Kanthou

Full Text Available Vascular endothelial growth factor-A (VEGF is produced by most cancer cells as multiple isoforms, which display distinct biological activities. VEGF plays an undisputed role in tumour growth, vascularisation and metastasis; nevertheless the functions of individual isoforms in these processes remain poorly understood. We investigated the effects of three main murine isoforms (VEGF188, 164 and 120 on tumour cell behaviour, using a panel of fibrosarcoma cells we developed that express them individually under endogenous promoter control. Fibrosarcomas expressing only VEGF188 (fs188 or wild type controls (fswt were typically mesenchymal, formed ruffles and displayed strong matrix-binding activity. VEGF164- and VEGF120-producing cells (fs164 and fs120 respectively were less typically mesenchymal, lacked ruffles but formed abundant cell-cell contacts. On 3D collagen, fs188 cells remained mesenchymal while fs164 and fs120 cells adopted rounded/amoeboid and a mix of rounded and elongated morphologies respectively. Consistent with their mesenchymal characteristics, fs188 cells migrated significantly faster than fs164 or fs120 cells on 2D surfaces while contractility inhibitors accelerated fs164 and fs120 cell migration. VEGF164/VEGF120 expression correlated with faster proliferation rates and lower levels of spontaneous apoptosis than VEGF188 expression. Nevertheless, VEGF188 was associated with constitutively active/phosphorylated AKT, ERK1/2 and Stat3 proteins. Differences in proliferation rates and apoptosis could be explained by defective signalling downstream of pAKT to FOXO and GSK3 in fs188 and fswt cells, which also correlated with p27/p21 cyclin-dependent kinase inhibitor over-expression. All cells expressed tyrosine kinase VEGF receptors, but these were not active/activatable suggesting that inherent differences between the cell lines are governed by endogenous VEGF isoform expression through complex interactions that are independent of tyrosine
Deep Coherent Vortices and Their Sea Surface Expressions

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Ienna, Federico; Bashmachnikov, Igor; Dias, Joaquim; Peliz, Alvaro

2017-04-01

Mediterranean Water eddies, known as Meddies, are an important dynamic process occurring at depths of 1000-meters in the Northeast Atlantic Ocean. Meddies occur as a direct result of the Mediterranean Outflow exiting through the Gibraltar Strait, and represent a prevalent mechanism that can be found extensively throughout the ocean. Moreover, Meddy cores are known to produce measurable expressions at the sea surface in the form of rotating coherent vortices, not only affecting the sea surface from beneath, but also allowing for the possibility to remotely study these deep phenomena through data gathered at the sea surface. While many past studies have focused on the properties of Meddy cores, only a handful of studies focus on the physical characteristics and behavior of the surface expressions produced. Are Meddy surface expressions different from other like vortices that dominate the physical ocean surface? What are the relationships between deep and surface mechanisms, and do any feedbacks exist? To shed light on these questions, we investigate the relationship between Meddies and their sea-surface expressions through observations using in-situ float and drifter profiles and satellite altimetry. A total of 782 Meddy cores were examined in the Northeast Atlantic using temperature and salinity data obtained by CTD and Argo during the Mecanismos de transporte e de dispersão da Água Mediterrânica no Atlântico Nordeste (MEDTRANS) project, and their corresponding sea-level expressions were geo-temporally matched in satellite altimetry data. We report several statistical properties of the sea-surface expressions of Meddies, including their mean diameter and vertical magnitude, and compare the properties of their surface features to the underlying Meddy cores. We investigate how the deep core affects the surface, and whether surface expressions may in return yield information about the underlying cores. Additionally, we examine the variability of the surface
Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms

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Kober, Daniel L.; Alexander-Brett, Jennifer M.; Karch, Celeste M.; Cruchaga, Carlos; Colonna, Marco; Holtzman, Michael J.; Brett, Thomas J. (WU-MED)

2016-12-20

Genetic variations in the myeloid immune receptor TREM2 are linked to several neurodegenerative diseases. To determine how TREM2 variants contribute to these diseases, we performed structural and functional studies of wild-type and variant proteins. Our 3.1 Å TREM2 crystal structure revealed that mutations found in Nasu-Hakola disease are buried whereas Alzheimer’s disease risk variants are found on the surface, suggesting that these mutations have distinct effects on TREM2 function. Biophysical and cellular methods indicate that Nasu-Hakola mutations impact protein stability and decrease folded TREM2 surface expression, whereas Alzheimer’s risk variants impact binding to a TREM2 ligand. Additionally, the Alzheimer’s risk variants appear to epitope map a functional surface on TREM2 that is unique within the larger TREM family. These findings provide a guide to structural and functional differences among genetic variants of TREM2, indicating that therapies targeting the TREM2 pathway should be tailored to these genetic and functional differences with patient-specific medicine approaches for neurodegenerative disorders.
Multiple ace genes encoding acetylcholinesterases of Caenorhabditis elegans have distinct tissue expression.

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Combes, Didier; Fedon, Yann; Toutant, Jean-Pierre; Arpagaus, Martine

2003-08-01

ace-1 and ace-2 genes encoding acetylcholinesterase in the nematode Caenorhabditis elegans present 35% identity in coding sequences but no homology in noncoding regions (introns, 5'- and 3'-untranslated regions). A 5'-region of ace-2 was defined by rescue of ace-1;ace-2 mutants. When green fluorescent protein (GFP) expression was driven by this regulatory region, the resulting pattern was distinct from that of ace-1. This latter gene is expressed in all body-wall and vulval muscle cells (Culetto et al., 1999), whereas ace-2 is expressed almost exclusively in neurons. ace-3 and ace-4 genes are located in close proximity on chromosome II (Combes et al., 2000). These two genes were first transcribed in vivo as a bicistronic messenger and thus constitute an ace-3;ace-4 operon. However, there was a very low level of monocistronic mRNA of ace-4 (the upstream gene) in vivo, and no ACE-4 enzymatic activity was ever detected. GFP expression driven by a 5' upstream region of the ace-3;ace-4 operon was detected in several muscle cells of the pharynx (pm3, pm4, pm5 and pm7) and in the two canal associated neurons (CAN cells). A dorsal row of body-wall muscle cells was intensively labelled in larval stages but no longer detected in adults. The distinct tissue-specific expression of ace-1, ace-2 and ace-3 (coexpressed only in pm5 cells) indicates that ace genes are not redundant.
Gene expression profiling of two distinct neuronal populations in the rodent spinal cord

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Ryge, Jesper; Westerdahl, Ann Charlotte; Alstøm, Preben

2008-01-01

Background: In the field of neuroscience microarray gene expression profiles on anatomically defined brain structures are being used increasingly to study both normal brain functions as well as pathological states. Fluorescent tracing techniques in brain tissue that identifies distinct neuronal p...

Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans.

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Maria C Ow

2018-02-01

Full Text Available Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite ("seesaw" transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome.
Early experiences mediate distinct adult gene expression and reproductive programs in Caenorhabditis elegans

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Ow, Maria C.; Nichitean, Alexandra M.; Dorus, Steve; Hall, Sarah E.

2018-01-01

Environmental stress during early development in animals can have profound effects on adult phenotypes via programmed changes in gene expression. Using the nematode C. elegans, we demonstrated previously that adults retain a cellular memory of their developmental experience that is manifested by differences in gene expression and life history traits; however, the sophistication of this system in response to different environmental stresses, and how it dictates phenotypic plasticity in adults that contribute to increased fitness in response to distinct environmental challenges, was unknown. Using transcriptional profiling, we show here that C. elegans adults indeed retain distinct cellular memories of different environmental conditions. We identified approximately 500 genes in adults that entered dauer due to starvation that exhibit significant opposite (“seesaw”) transcriptional phenotypes compared to adults that entered dauer due to crowding, and are distinct from animals that bypassed dauer. Moreover, we show that two-thirds of the genes in the genome experience a 2-fold or greater seesaw trend in gene expression, and based upon the direction of change, are enriched in large, tightly linked regions on different chromosomes. Importantly, these transcriptional programs correspond to significant changes in brood size depending on the experienced stress. In addition, we demonstrate that while the observed seesaw gene expression changes occur in both somatic and germline tissue, only starvation-induced changes require a functional GLP-4 protein necessary for germline development, and both programs require the Argonaute CSR-1. Thus, our results suggest that signaling between the soma and the germ line can generate phenotypic plasticity as a result of early environmental experience, and likely contribute to increased fitness in adverse conditions and the evolution of the C. elegans genome. PMID:29447162
Gene expression profiling of two distinct neuronal populations in the rodent spinal cord.

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Jesper Ryge

Full Text Available BACKGROUND: In the field of neuroscience microarray gene expression profiles on anatomically defined brain structures are being used increasingly to study both normal brain functions as well as pathological states. Fluorescent tracing techniques in brain tissue that identifies distinct neuronal populations can in combination with global gene expression profiling potentially increase the resolution and specificity of such studies to shed new light on neuronal functions at the cellular level. METHODOLOGY/PRINCIPAL FINDINGS: We examine the microarray gene expression profiles of two distinct neuronal populations in the spinal cord of the neonatal rat, the principal motor neurons and specific interneurons involved in motor control. The gene expression profiles of the respective cell populations were obtained from amplified mRNA originating from 50-250 fluorescently identified and laser microdissected cells. In the data analysis we combine a new microarray normalization procedure with a conglomerate measure of significant differential gene expression. Using our methodology we find 32 genes to be more expressed in the interneurons compared to the motor neurons that all except one have not previously been associated with this neuronal population. As a validation of our method we find 17 genes to be more expressed in the motor neurons than in the interneurons and of these only one had not previously been described in this population. CONCLUSIONS/SIGNIFICANCE: We provide an optimized experimental protocol that allows isolation of gene transcripts from fluorescent retrogradely labeled cell populations in fresh tissue, which can be used to generate amplified aRNA for microarray hybridization from as few as 50 laser microdissected cells. Using this optimized experimental protocol in combination with our microarray analysis methodology we find 49 differentially expressed genes between the motor neurons and the interneurons that reflect the functional
Expression of variant surface antigens by Plasmodium falciparum parasites in the peripheral blood of clinically immune pregnant women indicates ongoing placental infection

DEFF Research Database (Denmark)

Ofori, Michael F; Staalsoe, Trine; Bam, Victoria

2003-01-01

Placenta-sequestered Plasmodium falciparum parasites that cause pregnancy-associated malaria (PAM) in otherwise clinically immune women express distinct variant surface antigens (VSA(PAM)) not expressed by parasites in nonpregnant individuals. We report here that parasites from the peripheral blood...... of clinically immune pregnant women also express VSA(PAM), making them a convenient source of VSA(PAM) expressors for PAM vaccine research....
Comparative genome and transcriptome analysis reveals distinctive surface characteristics and unique physiological potentials of Pseudomonas aeruginosa ATCC 27853

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Cao, Huiluo

2017-06-12

Pseudomonas aeruginosa ATCC 27853 was isolated from a hospital blood specimen in 1971 and has been widely used as a model strain to survey antibiotics susceptibilities, biofilm development, and metabolic activities of Pseudomonas spp.. Although four draft genomes of P. aeruginosa ATCC 27853 have been sequenced, the complete genome of this strain is still lacking, hindering a comprehensive understanding of its physiology and functional genome.Here we sequenced and assembled the complete genome of P. aeruginosa ATCC 27853 using the Pacific Biosciences SMRT (PacBio) technology and Illumina sequencing platform. We found that accessory genes of ATCC 27853 including prophages and genomic islands (GIs) mainly contribute to the difference between P. aeruginosa ATCC 27853 and other P. aeruginosa strains. Seven prophages were identified within the genome of P. aeruginosa ATCC 27853. Of the predicted 25 GIs, three contain genes that encode monoxoygenases, dioxygenases and hydrolases that could be involved in the metabolism of aromatic compounds. Surveying virulence-related genes revealed that a series of genes that encode the B-band O-antigen of LPS are lacking in ATCC 27853. Distinctive SNPs in genes of cellular adhesion proteins such as type IV pili and flagella biosynthesis were also observed in this strain. Colony morphology analysis confirmed an enhanced biofilm formation capability of ATCC 27853 on solid agar surface compared to Pseudomonas aeruginosa PAO1. We then performed transcriptome analysis of ATCC 27853 and PAO1 using RNA-seq and compared the expression of orthologous genes to understand the functional genome and the genomic details underlying the distinctive colony morphogenesis. These analyses revealed an increased expression of genes involved in cellular adhesion and biofilm maturation such as type IV pili, exopolysaccharide and electron transport chain components in ATCC 27853 compared with PAO1. In addition, distinctive expression profiles of the
Dynamic Changes in Amygdala Psychophysiological Connectivity Reveal Distinct Neural Networks for Facial Expressions of Basic Emotions.

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Diano, Matteo; Tamietto, Marco; Celeghin, Alessia; Weiskrantz, Lawrence; Tatu, Mona-Karina; Bagnis, Arianna; Duca, Sergio; Geminiani, Giuliano; Cauda, Franco; Costa, Tommaso

2017-03-27

The quest to characterize the neural signature distinctive of different basic emotions has recently come under renewed scrutiny. Here we investigated whether facial expressions of different basic emotions modulate the functional connectivity of the amygdala with the rest of the brain. To this end, we presented seventeen healthy participants (8 females) with facial expressions of anger, disgust, fear, happiness, sadness and emotional neutrality and analyzed amygdala's psychophysiological interaction (PPI). In fact, PPI can reveal how inter-regional amygdala communications change dynamically depending on perception of various emotional expressions to recruit different brain networks, compared to the functional interactions it entertains during perception of neutral expressions. We found that for each emotion the amygdala recruited a distinctive and spatially distributed set of structures to interact with. These changes in amygdala connectional patters characterize the dynamic signature prototypical of individual emotion processing, and seemingly represent a neural mechanism that serves to implement the distinctive influence that each emotion exerts on perceptual, cognitive, and motor responses. Besides these differences, all emotions enhanced amygdala functional integration with premotor cortices compared to neutral faces. The present findings thus concur to reconceptualise the structure-function relation between brain-emotion from the traditional one-to-one mapping toward a network-based and dynamic perspective.
A microarray analysis of two distinct lymphatic endothelial cell populations

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Bernhard Schweighofer

2015-06-01

Full Text Available We have recently identified lymphatic endothelial cells (LECs to form two morphologically different populations, exhibiting significantly different surface protein expression levels of podoplanin, a major surface marker for this cell type. In vitro shockwave treatment (IVSWT of LECs resulted in enrichment of the podoplaninhigh cell population and was accompanied by markedly increased cell proliferation, as well as 2D and 3D migration. Gene expression profiles of these distinct populations were established using Affymetrix microarray analyses. Here we provide additional details about our dataset (NCBI GEO accession number GSE62510 and describe how we analyzed the data to identify differently expressed genes in these two LEC populations.
Oxygen Modulates Human Decidual Natural Killer Cell Surface Receptor Expression and Interactions with Trophoblasts1

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Wallace, Alison E.; Goulwara, Sonu S.; Whitley, Guy S.; Cartwright, Judith E.

2014-01-01

Decidual natural killer (dNK) cells have been shown to both promote and inhibit trophoblast behavior important for decidual remodeling in pregnancy and have a distinct phenotype compared to peripheral blood NK cells. We investigated whether different levels of oxygen tension, mimicking the physiological conditions of the decidua in early pregnancy, altered cell surface receptor expression and activity of dNK cells and their interactions with trophoblast. dNK cells were isolated from terminated first-trimester pregnancies and cultured in oxygen tensions of 3%, 10%, and 21% for 24 h. Cell surface receptor expression was examined by flow cytometry, and the effects of secreted factors in conditioned medium (CM) on the trophoblast cell line SGHPL-4 were assessed in vitro. SGHPL-4 cells treated with dNK cell CM incubated in oxygen tensions of 10% were significantly more invasive (P cells treated with dNK cell CM incubated in oxygen tensions of 3% or 21%. After 24 h, a lower percentage of dNK cells expressed CD56 at 21% oxygen (P cells expressed NKG2D at 10% oxygen (P oxygen tensions, with large patient variation. This study demonstrates dNK cell phenotype and secreted factors are modulated by oxygen tension, which induces changes in trophoblast invasion and endovascular-like differentiation. Alterations in dNK cell surface receptor expression and secreted factors at different oxygen tensions may represent regulation of function within the decidua during the first trimester of pregnancy. PMID:25232021
Distinct ASIC currents are expressed in rat putative nociceptors and are modulated by nerve injury.

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Poirot, Olivier; Berta, Temugin; Decosterd, Isabelle; Kellenberger, Stephan

2006-10-01

The H(+)-gated acid-sensing ion channels (ASICs) are expressed in dorsal root ganglion (DRG) neurones. Studies with ASIC knockout mice indicated either a pro-nociceptive or a modulatory role of ASICs in pain sensation. We have investigated in freshly isolated rat DRG neurones whether neurones with different ASIC current properties exist, which may explain distinct cellular roles, and we have investigated ASIC regulation in an experimental model of neuropathic pain. Small-diameter DRG neurones expressed three different ASIC current types which were all preferentially expressed in putative nociceptors. Type 1 currents were mediated by ASIC1a homomultimers and characterized by steep pH dependence of current activation in the pH range 6.8-6.0. Type 3 currents were activated in a similar pH range as type 1, while type 2 currents were activated at pH ASIC current density. Nerve injury induced differential regulation of ASIC subunit expression and selective changes in ASIC function in DRG neurones, suggesting a complex reorganization of ASICs during the development of neuropathic pain. In summary, we describe a basis for distinct cellular functions of different ASIC types in small-diameter DRG neurones.
Nuclear pores and perinuclear expression sites of var and ribosomal DNA genes correspond to physically distinct regions in Plasmodium falciparum.

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Guizetti, Julien; Martins, Rafael Miyazawa; Guadagnini, Stéphanie; Claes, Aurélie; Scherf, Artur

2013-05-01

The human malaria parasite Plasmodium falciparum modifies the erythrocyte it infects by exporting variant proteins to the host cell surface. The var gene family that codes for a large, variant adhesive surface protein called P. falciparum erythrocyte membrane protein 1 (PfEMP1) plays a particular role in this process, which is linked to pathogenesis and immune evasion. A single member of this gene family is highly transcribed while the other 59 members remain silenced. Importantly, var gene transcription occurs at a spatially restricted, but yet undefined, perinuclear site that is distinct from repressed var gene clusters. To advance our understanding of monoallelic expression, we investigated whether nuclear pores associate with the var gene expression site. To this end, we studied the nuclear pore organization during the asexual blood stage using a specific antibody directed against a subunit of the nuclear pore, P. falciparum Nup116 (PfNup116). Ring and schizont stage parasites showed highly polarized nuclear pore foci, whereas in trophozoite stage nuclear pores redistributed over the entire nuclear surface. Colocalization studies of var transcripts and anti-PfNup116 antibodies showed clear dissociation between nuclear pores and the var gene expression site in ring stage. Similar results were obtained for another differentially transcribed perinuclear gene family, the ribosomal DNA units. Furthermore, we show that in the poised state, the var gene locus is not physically linked to nuclear pores. Our results indicate that P. falciparum does form compartments of high transcriptional activity at the nuclear periphery which are, unlike the case in yeast, devoid of nuclear pores.
Distinct types of primary cutaneous large B-cell lymphoma identified by gene expression profiling.

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Hoefnagel, Juliette J; Dijkman, Remco; Basso, Katia; Jansen, Patty M; Hallermann, Christian; Willemze, Rein; Tensen, Cornelis P; Vermeer, Maarten H

2005-05-01

In the European Organization for Research and Treatment of Cancer (EORTC) classification 2 types of primary cutaneous large B-cell lymphoma (PCLBCL) are distinguished: primary cutaneous follicle center cell lymphomas (PCFCCL) and PCLBCL of the leg (PCLBCL-leg). Distinction between both groups is considered important because of differences in prognosis (5-year survival > 95% and 52%, respectively) and the first choice of treatment (radiotherapy or systemic chemotherapy, respectively), but is not generally accepted. To establish a molecular basis for this subdivision in the EORTC classification, we investigated the gene expression profiles of 21 PCLBCLs by oligonucleotide microarray analysis. Hierarchical clustering based on a B-cell signature (7450 genes) classified PCLBCL into 2 distinct subgroups consisting of, respectively, 8 PCFCCLs and 13 PCLBCLsleg. PCLBCLs-leg showed increased expression of genes associated with cell proliferation; the proto-oncogenes Pim-1, Pim-2, and c-Myc; and the transcription factors Mum1/IRF4 and Oct-2. In the group of PCFCCL high expression of SPINK2 was observed. Further analysis suggested that PCFCCLs and PCLBCLs-leg have expression profiles similar to that of germinal center B-cell-like and activated B-cell-like diffuse large B-cell lymphoma, respectively. The results of this study suggest that different pathogenetic mechanisms are involved in the development of PCFCCLs and PCLBCLs-leg and provide molecular support for the subdivision used in the EORTC classification.
Distinctive expression pattern of OCT4 variants in different types of breast cancer.

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Soheili, Saamaaneh; Asadi, Malek Hossein; Farsinejad, Alireza

2017-01-01

OCT4 is a key regulator of self-renewal and pluripotency in embryonic stem cells which can potentially encode three spliced variants designated OCT4A, OCT4B and OCT4B1. Based on cancer stem cell concept, it is suggested that the stemness factors misexpressed in cancer cells and potentially is involved in tumorigenesis. Accordingly, in this study, we investigated the potential expression of OCT4 variants in breast cancer tissues. A total of 94 tumoral and peritumoral breast specimens were evaluated with respect to the expression of OCT4 variants using quantitative RT-PCR and immunohistochemical (IHC) analysis. We detected the expression of OCT4 variants in breast tumor tissues with no or very low levels of expression in peritumoral samples of the same patients. While OCT4B was highly expressed in lobular type of breast cancer, OCT4A and OCTB1 variants are highly expressed in low grade (I and II) ductal tumors. Furthermore, the results of this study revealed a considerable association between the expression level of OCT4 variants and the expression of ER, PR, Her2 and P53 factors. All data demonstrated a distinctive expression pattern of OCT4 spliced variants in different types of breast cancer and provide further evidence for the involvement of embryonic genes in carcinogenesis.
Quantitative expression profile of distinct functional regions in the adult mouse brain.

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Takeya Kasukawa

Full Text Available The adult mammalian brain is composed of distinct regions with specialized roles including regulation of circadian clocks, feeding, sleep/awake, and seasonal rhythms. To find quantitative differences of expression among such various brain regions, we conducted the BrainStars (B* project, in which we profiled the genome-wide expression of ∼50 small brain regions, including sensory centers, and centers for motion, time, memory, fear, and feeding. To avoid confounds from temporal differences in gene expression, we sampled each region every 4 hours for 24 hours, and pooled the samples for DNA-microarray assays. Therefore, we focused on spatial differences in gene expression. We used informatics to identify candidate genes with expression changes showing high or low expression in specific regions. We also identified candidate genes with stable expression across brain regions that can be used as new internal control genes, and ligand-receptor interactions of neurohormones and neurotransmitters. Through these analyses, we found 8,159 multi-state genes, 2,212 regional marker gene candidates for 44 small brain regions, 915 internal control gene candidates, and 23,864 inferred ligand-receptor interactions. We also found that these sets include well-known genes as well as novel candidate genes that might be related to specific functions in brain regions. We used our findings to develop an integrated database (http://brainstars.org/ for exploring genome-wide expression in the adult mouse brain, and have made this database openly accessible. These new resources will help accelerate the functional analysis of the mammalian brain and the elucidation of its regulatory network systems.
Contrasting expression of membrane metalloproteinases, MT1-MMP and MT3-MMP, suggests distinct functions in skeletal development.

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Yang, Maozhou; Zhang, Bingbing; Zhang, Liang; Gibson, Gary

2008-07-01

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is the most ubiquitous and widely studied of the membrane-type metalloproteinases (MT-MMPs). It was thus surprising to find no published data on chicken MT1-MMP. We report here the characterization of the chicken gene. Its low sequence identity with the MT1-MMP genes of other species, high GC content, and divergent catalytic domain explains the absence of data and our difficulties in characterizing the gene. The absence of structural features in the chicken gene that have been suggested to be critical for the activation of MMP-2 by MT1-MMP; for the effect of MT1-MMP on cell migration and for the recycling of MT1-MMP suggest these features are either not essential or that MT1-MMP does not perform these functions in chickens. Comparison of the expression of chicken MT1-MMP with MT3-MMP and with MMP-2 and MMP-13 has confirmed the previously recognized co-expression of MT1-MMP with MMP-2 and MMP-13 in fibrous and vascular tissues, particularly those surrounding the developing long bones in other species. By contrast, MT3-MMP expression differs markedly from that of MT1-MMP and of both MMP-2 and MMP-13. MT3-MMP is expressed by chondrocytes of the developing articular surface. Similar expression patterns of this group of MT-MMPs and MMPs have been observed in mouse embryos and suggest distinct and specific functions for MT1-MMP and MT3-MMP in skeletal development.
Rhinovirus infection induces distinct transcriptome profiles in polarized human macrophages.

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Rajput, Charu; Walsh, Megan P; Eder, Breanna N; Metitiri, Ediri E; Popova, Antonia P; Hershenson, Marc B

2018-05-01

Infections with rhinovirus (RV) cause asthma exacerbations. Recent studies suggest that macrophages play a role in asthmatic airway inflammation and the innate immune response to RV infection. Macrophages exhibit phenotypes based on surface markers and gene expression. We hypothesized that macrophage polarization state alters gene expression in response to RV infection. Cells were derived from human peripheral blood derived monocytes. M1 and M2 polarization was carried out by using IFN-γ and IL-4, respectively, and RNA was extracted for Affymetrix Human Gene ST2.1 exon arrays. Selected genes were validated by quantitative (q)PCR. Treatment of nonactivated (M0) macrophages with IFN-γ and IL-4 induced the expression of 252 and 153 distinct genes, respectively, including previously-identified M1 and M2 markers. RV infection of M0 macrophages induced upregulation of 232 genes; pathway analysis showed significant overrepresentation of genes involved in IFN-α/β signaling and cytokine signaling in the immune system. RV infection induced differential expression of 195 distinct genes in M1-like macrophages but only seven distinct genes in M2-like-polarized cells. In a secondary analysis, comparison between M0-, RV-infected, and M1-like-polarized, RV-infected macrophages revealed differential expression of 227 genes including those associated with asthma and its exacerbation. qPCR demonstrated increased expression of CCL8, CXCL10, TNFSF10, TNFSF18, IL6, NOD2, and GSDMD and reduced expression of VNN1, AGO1, and AGO2. Together, these data show that, in contrast to M2-like-polarized macrophages, gene expression of M1-like macrophages is highly regulated by RV.
Distinct patterns of gene and protein expression elicited by organophosphorus pesticides in Caenorhabditis elegans

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Dennis William E

2009-04-01

Full Text Available Abstract Background The wide use of organophosphorus (OP pesticides makes them an important public health concern. Persistent effects of exposure and the mechanism of neuronal degeneration are continuing issues in OP toxicology. To elucidate early steps in the mechanisms of OP toxicity, we studied alterations in global gene and protein expression in Caenorhabditis elegans exposed to OPs using microarrays and mass spectrometry. We tested two structurally distinct OPs (dichlorvos and fenamiphos and employed a mechanistically different third neurotoxicant, mefloquine, as an out-group for analysis. Treatment levels used concentrations of chemical sufficient to prevent the development of 10%, 50% or 90% of mid-vulval L4 larvae into early gravid adults (EGA at 24 h after exposure in a defined, bacteria-free medium. Results After 8 h of exposure, the expression of 87 genes responded specifically to OP treatment. The abundance of 34 proteins also changed in OP-exposed worms. Many of the genes and proteins affected by the OPs are expressed in neuronal and muscle tissues and are involved in lipid metabolism, cell adhesion, apoptosis/cell death, and detoxification. Twenty-two genes were differentially affected by the two OPs; a large proportion of these genes encode cytochrome P450s, UDP-glucuronosyl/UDP-glucosyltransferases, or P-glycoproteins. The abundance of transcripts and the proteins they encode were well correlated. Conclusion Exposure to OPs elicits a pattern of changes in gene expression in exposed worms distinct from that of the unrelated neurotoxicant, mefloquine. The functional roles and the tissue location of the genes and proteins whose expression is modulated in response to exposure is consistent with the known effects of OPs, including damage to muscle due to persistent hypercontraction, neuronal cell death, and phase I and phase II detoxification. Further, the two different OPs evoked distinguishable changes in gene expression; about half
Paired Expression Analysis of Tumor Cell Surface Antigens

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Rimas J. Orentas

2017-08-01

Full Text Available Adoptive immunotherapy with antibody-based therapy or with T cells transduced to express chimeric antigen receptors (CARs is useful to the extent that the cell surface membrane protein being targeted is not expressed on normal tissues. The most successful CAR-based (anti-CD19 or antibody-based therapy (anti-CD20 in hematologic malignancies has the side effect of eliminating the normal B cell compartment. Targeting solid tumors may not provide a similar expendable marker. Beyond antibody to Her2/NEU and EGFR, very few antibody-based and no CAR-based therapies have seen broad clinical application for solid tumors. To expand the way in which the surfaceome of solid tumors can be analyzed, we created an algorithm that defines the pairwise relative overexpression of surface antigens. This enables the development of specific immunotherapies that require the expression of two discrete antigens on the surface of the tumor target. This dyad analysis was facilitated by employing the Hotelling’s T-squared test (Hotelling–Lawley multivariate analysis of variance for two independent variables in comparison to a third constant entity (i.e., gene expression levels in normal tissues. We also present a unique consensus scoring mechanism for identifying transcripts that encode cell surface proteins. The unique application of our bioinformatics processing pipeline and statistical tools allowed us to compare the expression of two membrane protein targets as a pair, and to propose a new strategy based on implementing immunotherapies that require both antigens to be expressed on the tumor cell surface to trigger therapeutic effector mechanisms. Specifically, we found that, for MYCN amplified neuroblastoma, pairwise expression of ACVR2B or anaplastic lymphoma kinase (ALK with GFRA3, GFRA2, Cadherin 24, or with one another provided the strongest hits. For MYCN, non-amplified stage 4 neuroblastoma, neurotrophic tyrosine kinase 1, or ALK paired with GFRA2, GFRA3, SSK
Contributions of feature shapes and surface cues to the recognition of facial expressions.

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Sormaz, Mladen; Young, Andrew W; Andrews, Timothy J

2016-10-01

Theoretical accounts of face processing often emphasise feature shapes as the primary visual cue to the recognition of facial expressions. However, changes in facial expression also affect the surface properties of the face. In this study, we investigated whether this surface information can also be used in the recognition of facial expression. First, participants identified facial expressions (fear, anger, disgust, sadness, happiness) from images that were manipulated such that they varied mainly in shape or mainly in surface properties. We found that the categorization of facial expression is possible in either type of image, but that different expressions are relatively dependent on surface or shape properties. Next, we investigated the relative contributions of shape and surface information to the categorization of facial expressions. This employed a complementary method that involved combining the surface properties of one expression with the shape properties from a different expression. Our results showed that the categorization of facial expressions in these hybrid images was equally dependent on the surface and shape properties of the image. Together, these findings provide a direct demonstration that both feature shape and surface information make significant contributions to the recognition of facial expressions. Copyright © 2016 Elsevier Ltd. All rights reserved.
Mycoplasma hyopneumoniae and Mycoplasma flocculare differential domains from orthologous surface proteins induce distinct cellular immune responses in mice.

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Leal, Fernanda Munhoz Dos Anjos; Virginio, Veridiana Gomes; Martello, Carolina Lumertz; Paes, Jéssica Andrade; Borges, Thiago J; Jaeger, Natália; Bonorino, Cristina; Ferreira, Henrique Bunselmeyer

2016-07-15

Mycoplasma hyopneumoniae and Mycoplasma flocculare are two genetically close species found in the swine respiratory tract. Despite their similarities, while M. hyopneumoniae is the causative agent of porcine enzootic pneumonia, M. flocculare is a commensal bacterium. Genomic and transcriptional comparative analyses so far failed to explain the difference in pathogenicity between these two species. We then hypothesized that such difference might be, at least in part, explained by amino acid sequence and immunological or functional differences between ortholog surface proteins. In line with that, it was verified that approximately 85% of the ortholog surface proteins from M. hyopneumoniae 7448 and M. flocculare present one or more differential domains. To experimentally assess possible immunological implications of this kind of difference, the extracellular differential domains from one pair of orthologous surface proteins (MHP7448_0612, from M. hyopneumoniae, and MF_00357, from M. flocculare) were expressed in E. coli and used to immunize mice. The recombinant polypeptides (rMHP61267-169 and rMF35767-196, respectively) induced distinct cellular immune responses. While, rMHP61267-169 induced both Th1 and Th2 responses, rMF35767-196 induced just an early pro-inflammatory response. These results indicate that immunological properties determined by differential domains in orthologous surface protein might play a role in pathogenicity, contributing to elicit specific and differential immune responses against each species. Copyright © 2016 Elsevier B.V. All rights reserved.
Do Infants Show Distinct Negative Facial Expressions for Fear and Anger? Emotional Expression in 11-Month-Old European American, Chinese, and Japanese Infants

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Camras, Linda A.; Oster, Harriet; Bakeman, Roger; Meng, Zhaolan; Ujiie, Tatsuo; Campos, Joseph J.

2007-01-01

Do infants show distinct negative facial expressions for different negative emotions? To address this question, European American, Chinese, and Japanese 11-month-olds were videotaped during procedures designed to elicit mild anger or frustration and fear. Facial behavior was coded using Baby FACS, an anatomically based scoring system. Infants'…

Early and long-standing rheumatoid arthritis: distinct molecular signatures identified by gene-expression profiling in synovia

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Lequerré, Thierry; Bansard, Carine; Vittecoq, Olivier; Derambure, Céline; Hiron, Martine; Daveau, Maryvonne; Tron, François; Ayral, Xavier; Biga, Norman; Auquit-Auckbur, Isabelle; Chiocchia, Gilles; Le Loët, Xavier; Salier, Jean-Philippe

2009-01-01

Introduction Rheumatoid arthritis (RA) is a heterogeneous disease and its underlying molecular mechanisms are still poorly understood. Because previous microarray studies have only focused on long-standing (LS) RA compared to osteoarthritis, we aimed to compare the molecular profiles of early and LS RA versus control synovia. Methods Synovial biopsies were obtained by arthroscopy from 15 patients (4 early untreated RA, 4 treated LS RA and 7 controls, who had traumatic or mechanical lesions). Extracted mRNAs were used for large-scale gene-expression profiling. The different gene-expression combinations identified by comparison of profiles of early, LS RA and healthy synovia were linked to the biological processes involved in each situation. Results Three combinations of 719, 116 and 52 transcripts discriminated, respectively, early from LS RA, and early or LS RA from healthy synovia. We identified several gene clusters and distinct molecular signatures specifically expressed during early or LS RA, thereby suggesting the involvement of different pathophysiological mechanisms during the course of RA. Conclusions Early and LS RA have distinct molecular signatures with different biological processes participating at different times during the course of the disease. These results suggest that better knowledge of the main biological processes involved at a given RA stage might help to choose the most appropriate treatment. PMID:19563633
Gene expression profiling in the striatum of inbred mouse strains with distinct opioid-related phenotypes

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Piechota Marcin

2006-06-01

Full Text Available Abstract Background Mouse strains with a contrasting response to morphine provide a unique model for studying the genetically determined diversity of sensitivity to opioid reward, tolerance and dependence. Four inbred strains selected for this study exhibit the most distinct opioid-related phenotypes. C57BL/6J and DBA/2J mice show remarkable differences in morphine-induced antinociception, self-administration and locomotor activity. 129P3/J mice display low morphine tolerance and dependence in contrast to high sensitivity to precipitated withdrawal observed in SWR/J and C57BL/6J strains. In this study, we attempted to investigate the relationships between genetic background and basal gene expression profile in the striatum, a brain region involved in the mechanism of opioid action. Results Gene expression was studied by Affymetrix Mouse Genome 430v2.0 arrays with probes for over 39.000 transcripts. Analysis of variance with the control for false discovery rate (q Khdrbs1 and ATPase Na+/K+ alpha2 subunit (Atp1a2 with morphine self-administration and analgesic effects, respectively. Finally, the examination of transcript structure demonstrated a possible inter-strain variability of expressed mRNA forms as for example the catechol-O-methyltransferase (Comt gene. Conclusion The presented study led to the recognition of differences in the gene expression that may account for distinct phenotypes. Moreover, results indicate strong contribution of genetic background to differences in gene transcription in the mouse striatum. The genes identified in this work constitute promising candidates for further animal studies and for translational genetic studies in the field of addictive and analgesic properties of opioids.
Connectivity from OR37 expressing olfactory sensory neurons to distinct cell types in the hypothalamus

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Andrea eBader

2012-11-01

Full Text Available Olfactory sensory neurons which express a member from the OR37 subfamily of odorant receptor genes are wired to the main olfactory bulb in a unique monoglomerular fashion; from these glomeruli an untypical connectivity into higher brain centers exists. In the present study we have investigated by DiI and transsynaptic tracing approaches how the connection pattern from these glomeruli into distinct hypothalamic nuclei is organized. The application of DiI onto the ventral domain of the bulb which harbors the OR37 glomeruli resulted in the labeling of fibers within the paraventricular and supraoptic nucleus of the hypothalamus; some of these fibers were covered with varicose-like structures. No DiI-labeled cell somata were detectable in these nuclei. The data indicate that projection neurons which originate in the OR37 region of the main olfactory bulb form direct connections into these nuclei. The cells that were labeled by the transsynaptic tracer WGA in these nuclei were further characterized. Their distribution pattern in the paraventricular nucleus was reminiscent of cells which produce distinct neuropeptides. Double labeling experiments confirmed that they contained vasopressin, but not the related neuropeptide oxytocin. Morphological analysis revealed that they comprise of magno- and parvocellular cells. A comparative investigation of the WGA-positive cells in the supraoptic nucleus demonstrated that these were vasopressin-positive, as well, whereas oxytocin-producing cells of this nucleus also contained no transsynaptic tracer. Together, the data demonstrate a connectivity from OR37 expressing sensory neurons to distinct hypothalamic neurons with the same neuropeptide content.
Cognitive, cultural, and linguistic sources of a handshape distinction expressing agentivity.

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Brentari, Diane; Di Renzo, Alessio; Keane, Jonathan; Volterra, Virginia

2015-01-01

In this paper the cognitive, cultural, and linguistic bases for a pattern of conventionalization of two types of iconic handshapes are described. Work on sign languages has shown that handling handshapes (H-HSs: those that represent how objects are handled or manipulated) and object handshapes (O-HSs: those that represent the class, size, or shape of objects) express an agentive/non-agentive semantic distinction in many sign languages. H-HSs are used in agentive event descriptions and O-HSs are used in non-agentive event descriptions. In this work, American Sign Language (ASL) and Italian Sign Language (LIS) productions are compared (adults and children) as well as the corresponding groups of gesturers in each country using "silent gesture." While the gesture groups, in general, did not employ an H-HS/O-HS distinction, all participants (signers and gesturers) used iconic handshapes (H-HSs and O-HSs together) more often in agentive than in no-agent event descriptions; moreover, none of the subjects produced an opposite pattern than the expected one (i.e., H-HSs associated with no-agent descriptions and O-HSs associated with agentive ones). These effects are argued to be grounded in cognition. In addition, some individual gesturers were observed to produce the H-HS/O-HS opposition for agentive and non-agentive event descriptions-that is, more Italian than American adult gesturers. This effect is argued to be grounded in culture. Finally, the agentive/non-agentive handshape opposition is confirmed for signers of ASL and LIS, but previously unreported cross-linguistic differences were also found across both adult and child sign groups. It is, therefore, concluded that cognitive, cultural, and linguistic factors contribute to the conventionalization of this distinction of handshape type. Copyright © 2014 Cognitive Science Society, Inc.
Growth and gene expression are predominantly controlled by distinct regions of the human IL-4 receptor.

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Ryan, J J; McReynolds, L J; Keegan, A; Wang, L H; Garfein, E; Rothman, P; Nelms, K; Paul, W E

1996-02-01

IL-4 causes hematopoietic cells to proliferate and express a series of genes, including CD23. We examined whether IL-4-mediated growth, as measured by 4PS phosphorylation, and gene induction were similarly controlled. Studies of M12.4.1 cells expressing human IL-4R truncation mutants indicated that the region between amino acids 557-657 is necessary for full gene expression, which correlated with Stat6 DNA binding activity. This region was not required for 4PS phosphorylation. Tyrosine-to-phenylalanine mutations in the interval between amino acids 557-657 revealed that as long as one tyrosine remained unmutated, CD23 was fully induced. When all three tyrosines were mutated, the receptor was unable to induce CD23. The results indicate that growth regulation and gene expression are principally controlled by distinct regions of IL-4R.
The IFN Response in Bats Displays Distinctive IFN-Stimulated Gene Expression Kinetics with Atypical RNASEL Induction.

Science.gov (United States)

De La Cruz-Rivera, Pamela C; Kanchwala, Mohammed; Liang, Hanquan; Kumar, Ashwani; Wang, Lin-Fa; Xing, Chao; Schoggins, John W

2018-01-01

Bats host a large number of zoonotic viruses, including several viruses that are highly pathogenic to other mammals. The mechanisms underlying this rich viral diversity are unknown, but they may be linked to unique immunological features that allow bats to act as asymptomatic viral reservoirs. Vertebrates respond to viral infection by inducing IFNs, which trigger antiviral defenses through IFN-stimulated gene (ISG) expression. Although the IFN system of several bats is characterized at the genomic level, less is known about bat IFN-mediated transcriptional responses. In this article, we show that IFN signaling in bat cells from the black flying fox ( Pteropus alecto ) consists of conserved and unique ISG expression profiles. In IFN-stimulated cells, bat ISGs comprise two unique temporal subclusters with similar early induction kinetics but distinct late-phase declines. In contrast, human ISGs lack this decline phase and remained elevated for longer periods. Notably, in unstimulated cells, bat ISGs were expressed more highly than their human counterparts. We also found that the antiviral effector 2-5A-dependent endoribonuclease, which is not an ISG in humans, is highly IFN inducible in black flying fox cells and contributes to cell-intrinsic control of viral infection. These studies reveal distinctive innate immune features that may underlie a unique virus-host relationship in bats. Copyright © 2017 by The American Association of Immunologists, Inc.
Alternate phase variation in expression of two major surface membrane proteins (MBA and UU376) of Ureaplasma parvum serovar 3.

Science.gov (United States)

Zimmerman, Carl-Ulrich R; Stiedl, Thomas; Rosengarten, Renate; Spergser, Joachim

2009-03-01

Ureaplasma urealyticum and Ureaplasma parvum are commensals and pathogens of the human urogenital tract and of newborn infants. There are four distinct U. parvum serovars and 10 distinct U. urealyticum serovars. Both species possess a distinct immunodominant variable surface protein, the multiple banded antigen (MBA), which shows size variability among isolates as a result of changes in the number of C-terminal repeating units. Adjacent to the MBA gene (UU375) lies UU376, which was annotated as 'Ureaplasma-specific conserved hypothetical gene'. In four different strains of U. parvum serovar 3, we demonstrated expression of UU376 by Western blot analysis and phase variation between UU376, here designated Upvmp376 (Ureaplasma phase-variable membrane protein 376), and MBA after application of selective pressure with hyperimmune antisera directed against either protein. By Southern blot analysis, we found that the switch between MBA and Upvmp376 expression is associated with a DNA inversion event in which the nonrepetitive region of the MBA gene and its putative promoter region are opposed to either the repetitive region of MBA or UU376. We propose that in U. parvum serovar 3, and presumably in all U. parvum and U. urealyticum, an inversion event at specific sites effects an alternate ON/OFF switching of the genes UU375 and UU376.
TALE factors use two distinct functional modes to control an essential zebrafish gene expression program.

Science.gov (United States)

Ladam, Franck; Stanney, William; Donaldson, Ian J; Yildiz, Ozge; Bobola, Nicoletta; Sagerström, Charles G

2018-06-18

TALE factors are broadly expressed embryonically and known to function in complexes with transcription factors (TFs) like Hox proteins at gastrula/segmentation stages, but it is unclear if such generally expressed factors act by the same mechanism throughout embryogenesis. We identify a TALE-dependent gene regulatory network (GRN) required for anterior development and detect TALE occupancy associated with this GRN throughout embryogenesis. At blastula stages, we uncover a novel functional mode for TALE factors, where they occupy genomic DECA motifs with nearby NF-Y sites. We demonstrate that TALE and NF-Y form complexes and regulate chromatin state at genes of this GRN. At segmentation stages, GRN-associated TALE occupancy expands to include HEXA motifs near PBX:HOX sites. Hence, TALE factors control a key GRN, but utilize distinct DNA motifs and protein partners at different stages - a strategy that may also explain their oncogenic potential and may be employed by other broadly expressed TFs. © 2018, Ladam et al.
Bacterial feeding, Leishmania infection and distinct infection routes induce differential defensin expression in Lutzomyia longipalpis.

Science.gov (United States)

Telleria, Erich L; Sant'Anna, Maurício R Viana; Alkurbi, Mohammad O; Pitaluga, André N; Dillon, Rod J; Traub-Csekö, Yara M

2013-01-11

Phlebotomine insects harbor bacterial, viral and parasitic pathogens that can cause diseases of public health importance. Lutzomyia longipalpis is the main vector of visceral leishmaniasis in the New World. Insects can mount a powerful innate immune response to pathogens. Defensin peptides take part in this response and are known to be active against Gram-positive and Gram-negative bacteria, and some parasites. We studied the expression of a defensin gene from Lutzomyia longipalpis to understand its role in sand fly immune response. We identified, sequenced and evaluated the expression of a L. longipalpis defensin gene by semi-quantitative RT-PCR. The gene sequence was compared to other vectors defensins and expression was determined along developmental stages and after exposure of adult female L. longipalpis to bacteria and Leishmania. Phylogenetic analysis showed that the L. longipalpis defensin is closely related to a defensin from the Old World sand fly Phlebotomus duboscqi. Expression was high in late L4 larvae and pupae in comparison to early larval stages and newly emerged flies. Defensin expression was modulated by oral infection with bacteria. The Gram-positive Micrococcus luteus induced early high defensin expression, whilst the Gram-negative entomopathogenic Serratia marcescens induced a later response. Bacterial injection also induced defensin expression in adult insects. Female sand flies infected orally with Leishmania mexicana showed no significant difference in defensin expression compared to blood fed insects apart from a lower defensin expression 5 days post Leishmania infection. When Leishmania was introduced into the hemolymph by injection there was no induction of defensin expression until 72 h later. Our results suggest that L. longipalpis modulates defensin expression upon bacterial and Leishmania infection, with patterns of expression that are distinct among bacterial species and routes of infection.
Expression of the cell-surface heparan sulfate proteoglycan syndecan-2 in developing rat anterior pituitary gland.

Science.gov (United States)

Horiguchi, Kotaro; Syaidah, Rahimi; Fujiwara, Ken; Tsukada, Takehiro; Ramadhani, Dini; Jindatip, Depicha; Kikuchi, Motoshi; Yashiro, Takashi

2013-09-01

In the anterior pituitary gland, folliculo-stellate cells and five types of hormone-producing cells are surrounded by an extracellular matrix (ECM) essential for these cells to perform their respective roles. Syndecans-type I transmembrane cell-surface heparan sulfate proteoglycans act as major ECM coreceptors via their respective heparan sulfate chains and efficiently transduce intracellular signals through the convergent action of their transmembrane and cytoplasmic domains. The syndecans comprise four family members in vertebrates: syndecan-1, -2, -3 and -4. However, whether syndecans are produced in the pituitary gland or whether they have a role as a coreceptor is not known. We therefore used (1) reverse transcription plus the polymerase chain reaction to analyze the expression of syndecan genes and (2) immunohistochemical techniques to identify the cells that produce the syndecans in the anterior pituitary gland of adult rat. Syndecan-2 mRNA expression was clearly detected in the corticotropes of the anterior pituitary gland. Moreover, the expression of syndecan-2 in the developing pituitary gland had a distinct temporospatial pattern. To identify the cells expressing syndecan-2 in the developing pituitary gland, we used double-immunohistochemistry for syndecan-2 and the cell markers E-cadherin (immature cells) and Ki-67 (proliferating cells). Some E-cadherin- and Ki-67-immunopositive cells expressed syndecan-2. Therefore, syndecan-2 expression occurs in developmentally regulated patterns and syndecan-2 probably has different roles in adult and developing anterior pituitary glands.
Distinctive Citizenship

DEFF Research Database (Denmark)

Kaur, Ravinder

2009-01-01

The refugee, in India's Partition history, appears as an enigmatic construct - part pitiful, part heroic, though mostly shorn of agency - representing the surface of the human tragedy of Partition. Yet this archetype masks the undercurrent of social distinctions that produced hierarchies of post...
Vasopressin up-regulates the expression of growth-related immediate-early genes via two distinct EGF receptor transactivation pathways

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Fuentes, Lida Q.; Reyes, Carlos E.; Sarmiento, José M.; Villanueva, Carolina I.; Figueroa, Carlos D.; Navarro, Javier; González, Carlos B.

2008-01-01

Activation of V1a receptor triggers the expression of growth-related immediate-early genes (IEGs), including c-Fos and Egr-1. Here we found that pre-treatment of rat vascular smooth muscle A-10 cell line with the EGF receptor inhibitor AG1478 or the over-expression of an EGFR dominant negative mutant (HEBCD533) blocked the vasopressin-induced expression of IEGs, suggesting that activation of these early genes mediated by V1a receptor is via transactivation of the EGF receptor. Importantly, the inhibition of the metalloproteinases, which catalyzed the shedding of the EGF receptor agonist HB-EGF, selectively blocked the vasopressin-induced expression c-Fos. On the other hand, the inhibition of c-Src selectively blocked the vasopressin-induced expression of Egr-1. Interestingly, in contrast to the expression of c-Fos, the expression of Egr-1 was mediated via the Ras/MEK/MAPK-dependent signalling pathway. Vasopressin-triggered expression of both genes required the release of intracellular calcium, activation of PKC and β-arrestin 2. These findings demonstrated that vasopressin up-regulated the expression of c-Fos and Erg-1 via transactivation of two distinct EGF receptor-dependent signalling pathways. PMID:18571897
Gene expression profiling reveals distinct molecular signatures associated with the rupture of intracranial aneurysm.

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Nakaoka, Hirofumi; Tajima, Atsushi; Yoneyama, Taku; Hosomichi, Kazuyoshi; Kasuya, Hidetoshi; Mizutani, Tohru; Inoue, Ituro

2014-08-01

The rupture of intracranial aneurysm (IA) causes subarachnoid hemorrhage associated with high morbidity and mortality. We compared gene expression profiles in aneurysmal domes between unruptured IAs and ruptured IAs (RIAs) to elucidate biological mechanisms predisposing to the rupture of IA. We determined gene expression levels of 8 RIAs, 5 unruptured IAs, and 10 superficial temporal arteries with the Agilent microarrays. To explore biological heterogeneity of IAs, we classified the samples into subgroups showing similar gene expression patterns, using clustering methods. The clustering analysis identified 4 groups: superficial temporal arteries and unruptured IAs were aggregated into their own clusters, whereas RIAs segregated into 2 distinct subgroups (early and late RIAs). Comparing gene expression levels between early RIAs and unruptured IAs, we identified 430 upregulated and 617 downregulated genes in early RIAs. The upregulated genes were associated with inflammatory and immune responses and phagocytosis including S100/calgranulin genes (S100A8, S100A9, and S100A12). The downregulated genes suggest mechanical weakness of aneurysm walls. The expressions of Krüppel-like family of transcription factors (KLF2, KLF12, and KLF15), which were anti-inflammatory regulators, and CDKN2A, which was located on chromosome 9p21 that was the most consistently replicated locus in genome-wide association studies of IA, were also downregulated. We demonstrate that gene expression patterns of RIAs were different according to the age of patients. The results suggest that macrophage-mediated inflammation is a key biological pathway for IA rupture. The identified genes can be good candidates for molecular markers of rupture-prone IAs and therapeutic targets. © 2014 American Heart Association, Inc.
Downregulation of transferrin receptor surface expression by intracellular antibody

International Nuclear Information System (INIS)

Peng Jilin; Wu Sha; Zhao Xiaoping; Wang Min; Li Wenhan; Shen Xin; Liu Jing; Lei Ping; Zhu Huifen; Shen Guanxin

2007-01-01

To deplete cellular iron uptake, and consequently inhibit the proliferation of tumor cells, we attempt to block surface expression of transferrin receptor (TfR) by intracellular antibody technology. We constructed two expression plasmids (scFv-HAK and scFv-HA) coding for intracellular single-chain antibody against TfR with or without endoplasmic reticulum (ER) retention signal, respectively. Then they were transfected tumor cells MCF-7 by liposome. Applying RT-PCR, Western blotting, immunofluorescence microscopy and immunoelectron microscope experiments, we insure that scFv-HAK intrabody was successfully expressed and retained in ER contrasted to the secreted expression of scFv-HA. Flow cytometric analysis confirmed that the TfR surface expression was markedly decreased approximately 83.4 ± 2.5% in scFv-HAK transfected cells, while there was not significantly decrease in scFv-HA transfected cells. Further cell growth and apoptosis characteristics were evaluated by cell cycle analysis, nuclei staining and MTT assay. Results indicated that expression of scFv-HAK can dramatically induce cell cycle G1 phase arrest and apoptosis of tumor cells, and consequently significantly suppress proliferation of tumor cells compared with other control groups. For First time this study demonstrates the potential usage of anti-TfR scFv-intrabody as a growth inhibitor of TfR overexpressing tumors
Distinct interferon-gamma and interleukin-9 expression in cutaneous and oral lichen planus.

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Weber, B; Schlapbach, C; Stuck, M; Simon, H-U; Borradori, L; Beltraminelli, H; Simon, D

2017-05-01

Cutaneous (CLP) and oral lichen planus (OLP) as the main subtypes of lichen planus (LP) present with different clinical manifestation and disease course, although their histopathologic features such as the band-like lymphocyte infiltrate and keratinocyte apoptosis are similar. So far, the underlying cellular and molecular mechanisms remain poorly understood. The aim of this study was to characterize and compare the in situ cellular infiltrates, cytokine expression profiles and apoptosis markers in CLP and OLP. Using immunofluorescence staining and laser scanning microscopy, we evaluated the cellular infiltrate (CD1a, CD3, CD4, CD8, CD21, CD57, CD123), cytokine expression (interleukin (IL)-1, IL-6, IL-9, IL-10, IL-17, IL-22, IL-23, tumour necrosis factor-α, transforming growth factor-β, interferon (IFN)-γ), and apoptosis markers (Fas, Fas ligand, cleaved caspase-3, TUNEL) of 21 anonymized biopsy specimens of LP (11 CLP, 10 OLP). Among infiltrating cells mainly T cells and natural killer (NK) cells as well as plasmacytoid dendritic cells (DC) were observed. A predominance of CD8+ T cells was noted in OLP. In both CLP and OLP, T helper (Th)1, Th9, Th17, and Th22-type cytokines were expressed. The expression of IL-9, IFN-γ and IL-22 was higher in CLP compared to that of OLP (P = 0.0165; P = 0.0016; P = 0.052 respectively). Expression of Fas and Fas ligand as well as cleaved caspase-3-positive cells was observed in the epithelium of all LP samples. The cell and cytokine patterns of CLP and OLP were partially distinct and generally resembled those reported for autoimmune diseases. The presence of CD8+ and NK cells as well as Fas/Fas ligand expression suggested that various pathways involved in keratinocyte apoptosis are relevant for LP. These results might help to establish targeted therapies for LP. © 2016 European Academy of Dermatology and Venereology.
A distinct regulatory region of the Bmp5 locus activates gene expression following adult bone fracture or soft tissue injury.

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Guenther, Catherine A; Wang, Zhen; Li, Emma; Tran, Misha C; Logan, Catriona Y; Nusse, Roel; Pantalena-Filho, Luiz; Yang, George P; Kingsley, David M

2015-08-01

Bone morphogenetic proteins (BMPs) are key signaling molecules required for normal development of bones and other tissues. Previous studies have shown that null mutations in the mouse Bmp5 gene alter the size, shape and number of multiple bone and cartilage structures during development. Bmp5 mutations also delay healing of rib fractures in adult mutants, suggesting that the same signals used to pattern embryonic bone and cartilage are also reused during skeletal regeneration and repair. Despite intense interest in BMPs as agents for stimulating bone formation in clinical applications, little is known about the regulatory elements that control developmental or injury-induced BMP expression. To compare the DNA sequences that activate gene expression during embryonic bone formation and following acute injuries in adult animals, we assayed regions surrounding the Bmp5 gene for their ability to stimulate lacZ reporter gene expression in transgenic mice. Multiple genomic fragments, distributed across the Bmp5 locus, collectively coordinate expression in discrete anatomic domains during normal development, including in embryonic ribs. In contrast, a distinct regulatory region activated expression following rib fracture in adult animals. The same injury control region triggered gene expression in mesenchymal cells following tibia fracture, in migrating keratinocytes following dorsal skin wounding, and in regenerating epithelial cells following lung injury. The Bmp5 gene thus contains an "injury response" control region that is distinct from embryonic enhancers, and that is activated by multiple types of injury in adult animals. Copyright © 2015 Elsevier Inc. All rights reserved.
Transcriptomic landscape of acute promyelocytic leukemia reveals aberrant surface expression of the platelet aggregation agonist Podoplanin.

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Lavallée, Vincent-Philippe; Chagraoui, Jalila; MacRae, Tara; Marquis, Miriam; Bonnefoy, Arnaud; Krosl, Jana; Lemieux, Sébastien; Marinier, Anne; Pabst, Caroline; Rivard, Georges-Étienne; Hébert, Josée; Sauvageau, Guy

2018-02-23

Acute promyelocytic leukemia (APL) is a medical emergency because of associated lethal early bleeding, a condition preventable by prompt diagnosis and therapeutic intervention. The mechanisms underlying the hemostatic anomalies of APL are not completely elucidated. RNA-sequencing-based characterization of APL (n = 30) was performed and compared to that of other acute myeloid leukemia (n = 400) samples and normal promyelocytes. Perturbations in the transcriptome of coagulation and fibrinolysis-related genes in APL extend beyond known culprits and now include Thrombin, Factor X and Urokinase Receptor. Most intriguingly, the Podoplanin (PDPN) gene, involved in platelet aggregation, is aberrantly expressed in APL promyelocytes and is the most distinctive transcript for this disease. Using an antibody panel optimized for AML diagnosis by flow cytometry, we also found that PDPN was the most specific surface marker for APL, and that all-trans retinoic acid therapy rapidly decreases its expression. Functional studies showed that engineered overexpression of this gene in human leukemic cells causes aberrant platelet binding, activation and aggregation. PDPN-expressing primary APL cells, but not PDPN-negative primary leukemias, specifically induce platelet binding, activation and aggregation. Finally, PDPN expression on leukemia cells in a xenograft model was associated with thrombocytopenia and prolonged bleeding time in vivo. Together our results suggest that PDPN may contribute to the hemostatic perturbations found in APL.
Duct- and Acinar-Derived Pancreatic Ductal Adenocarcinomas Show Distinct Tumor Progression and Marker Expression

Directory of Open Access Journals (Sweden)

Rute M.M. Ferreira

2017-10-01

Full Text Available The cell of origin of pancreatic ductal adenocarcinoma (PDAC has been controversial. Here, we show that identical oncogenic drivers trigger PDAC originating from both ductal and acinar cells with similar histology but with distinct pathophysiology and marker expression dependent on cell of origin. Whereas acinar-derived tumors exhibited low AGR2 expression and were preceded by pancreatic intraepithelial neoplasias (PanINs, duct-derived tumors displayed high AGR2 and developed independently of a PanIN stage via non-mucinous lesions. Using orthotopic transplantation and chimera experiments, we demonstrate that PanIN-like lesions can be induced by PDAC as bystanders in adjacent healthy tissues, explaining the co-existence of mucinous and non-mucinous lesions and highlighting the need to distinguish between true precursor PanINs and PanIN-like bystander lesions. Our results suggest AGR2 as a tool to stratify PDAC according to cell of origin, highlight that not all PanIN-like lesions are precursors of PDAC, and add an alternative progression route to the current model of PDAC development.
Distinct temporal changes in host cell lncRNA expression during the course of an adenovirus infection

Energy Technology Data Exchange (ETDEWEB)

Zhao, Hongxing, E-mail: Hongxing.Zhao@igp.uu.se [The Beijer Laboratory, Dept. of Immunology, Genetics and Pathology, Uppsala University, S-751 85 Uppsala (Sweden); Chen, Maoshan [Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria 3086 (Australia); Lind, Sara Bergström [Department of Chemistry-BMC, Analytical Chemistry, Science for Life Laboratory, Uppsala University, Box 599, SE-751 24 Uppsala (Sweden); Pettersson, Ulf [The Beijer Laboratory, Dept. of Immunology, Genetics and Pathology, Uppsala University, S-751 85 Uppsala (Sweden)

2016-05-15

The deregulation of cellular long non-coding RNA (lncRNA) expression during a human adenovirus infection was studied by deep sequencing. Expression of lncRNAs increased substantially following the progression of the infection. Among 645 significantly expressed lncRNAs, the expression of 398 was changed more than 2-fold. More than 80% of them were up-regulated and 80% of them were detected during the late phase. Based on the genomic locations of the deregulated lncRNAs in relation to known mRNAs and miRNAs, they were predicted to be involved in growth, structure, apoptosis and wound healing in the early phase, cell proliferation in the intermediate phase and protein synthesis, modification and transport in the late phase. The most significant functions of cellular RNA-binding proteins, previously shown to interact with the deregulated lncRNAs identified here, are involved in RNA splicing, nuclear export and translation events. We hypothesize that adenoviruses exploit the lncRNA network to optimize their reproduction. - Highlights: • The expression of 398 lncRNAs showed a distinct temporal pattern during Ad2 infection. • 80% of the deregulated lncRNAs were up-regulated during the late phase of infection. • The deregulated lncRNAs potentiallyinteract with 33 cellular RNA binding proteins. • These RBPs are involved in RNA splicing, nuclear export and translation. • Adenovirus exploits the cellular lncRNA network to optimize its replication.
The neural cell adhesion molecule L1 is distinct from the N-CAM related group of surface antigens BSP-2 and D2

DEFF Research Database (Denmark)

Faissner, A; Kruse, J; Goridis, C

1984-01-01

The neural cell adhesion molecule L1 and the group of N-CAM related molecules, BSP-2 and D2 antigen, are immunochemically distinct molecular species. The two groups of surface molecules are also functionally distinct entities, since inhibition of Ca2+-independent adhesion among early post-natal m...

Surface expression of NMDA receptor changes during memory consolidation in the crab Neohelice granulata

Science.gov (United States)

Hepp, Yanil; Salles, Angeles; Carbo-Tano, Martin

2016-01-01

The aim of the present study was to analyze the surface expression of the NMDA-like receptors during the consolidation of contextual learning in the crab Neohelice granulata. Memory storage is based on alterations in the strength of synaptic connections between neurons. The glutamatergic synapses undergo various forms of N-methyl-D aspartate receptor (NMDAR)-dependent changes in strength, a process that affects the abundance of other receptors at the synapse and underlies some forms of learning and memory. Here we propose a direct regulation of the NMDAR. Changes in NMDAR's functionality might be induced by the modification of the subunit's expression or cellular trafficking. This trafficking does not only include NMDAR's movement between synaptic and extra-synaptic localizations but also the cycling between intracellular compartments and the plasma membrane, a process called surface expression. Consolidation of contextual learning affects the surface expression of the receptor without affecting its general expression. The surface expression of the GluN1 subunit of the NMDAR is down-regulated immediately after training, up-regulated 3 h after training and returns to naïve and control levels 24 h after training. The changes in NMDAR surface expression observed in the central brain are not seen in the thoracic ganglion. A similar increment in surface expression of GluN1 in the central brain is observed 3 h after administration of the competitive GABAA receptor antagonist, bicuculline. These consolidation changes are part of a plasticity event that first, during the down-regulation, stabilizes the trace and later, at 3-h post-training, changes the threshold for synapse activation. PMID:27421895
Distinct patterns of ALDH1A1 expression predict metastasis and poor outcome of colorectal carcinoma

Science.gov (United States)

Xu, Sen-Lin; Zeng, Dong-Zu; Dong, Wei-Guo; Ding, Yan-Qing; Rao, Jun; Duan, Jiang-Jie; Liu, Qing; Yang, Jing; Zhan, Na; Liu, Ying; Hu, Qi-Ping; Zhang, Xia; Cui, You-Hong; Kung, Hsiang-Fu; Yu, Shi-Cang; Bian, Xiu-Wu

2014-01-01

Purpose: Aldehyde dehydrogenase 1A1 (ALDH1A1) has been proposed as a candidate biomarker for colorectal carcinoma (CRC). However, the heterogeneity of its expression makes it difficult to predict the outcome of CRC. The aim of this study was to evaluate the diagnostic and prognostic value of this molecule in CRC. Methods and Results: In this study, we examined ALDH1A1 expression by immunohistochemistry including 406 cases of primary CRC with corresponding adjacent mucosa, with confirmation of real-time PCR and Western blotting. We found that the expression patterns of ALDH1A1 were heterogeneous in the CRC and corresponding adjacent tissues. We defined the ratio of ALDH1A1 level in adjacent mucosa to that in tumor tissues as RA/C and found that the capabilities of tumor invasion and metastasis in the tumors with RA/C < 1 were significantly higher than those with RA/C ≥ 1. Follow-up data showed the worse prognoses in the CRC patients with RA/C < 1. For understanding the underlying mechanism, the localization of β-catenin was detected in the CRC tissues with different patterns of ALDH1A1 expression from 221 patients and β-catenin was found preferentially expressed in cell nuclei of the tumors with RA/C < 1 and ALDH1A1high expression of HT29 cell line, indicating that nuclear translocation of β-catenin might contribute to the increased potentials of invasion and metastasis. Conclusion: Our results indicate that RA/C is a novel biomarker to reflect the distinct expression patterns of ALDH1A1 for predicting metastasis and prognosis of CRC. PMID:25031716
Distinct RNA transcriptome patterns are potentially associated with angiogenesis in Tie2-expressing monocytes.

Science.gov (United States)

Wang, Xinjing; Dai, Zhiyuan; Wu, Xiaoli; Wang, Kai; Wang, Xipeng

2016-04-10

Tie2-expressing Monocytes (TEMs) were previously identified as a novel subset of monocytes and were believed to have prominent pro-angiogenesis activities in human tumors. While the molecular mechanism of the angiogenesis promoting capacity of TEMs remains unclear. RNA transcriptome pattern, including non-coding RNAs as microRNA (miRNA) and long non-coding RNA (lncRNA), plays important role in cell differentiation and functions. However, little is known about the transcriptome patterns of TEMs, including those non-coding RNAs. We explore the transcriptome of TEMs and the matched monocytes that do not express Tie2 (Tie2(-)monocytes) isolated from peripheral blood of healthy adults employing the Agilent Human miRNA(8*60K,Design ID: 046064)microarray and the Agilent lncRNA Gene Expression(4*180K, Design ID: 042818)microarray. A total of 141 mRNAs, 142 lncRNAs and 75 miRNAs were found dysregulated in TEMs compared to Tie2(-)monocytes. TEMs have the distinct RNA transcriptome patterns according to the Hierarchical clustering and then the gene expression patterns were confirmed by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Functional annotation by Gene Ontology (GO) analyses showed that the up-regulated mRNAs in TEMs were associated to blood vessel remodeling and positive regulation of epithelial cell proliferation, and the up-regulated insulin like growth factor 1(IGF1) mRNA was involved in both pathways. For functional analysis of those dysregulated non-coding RNAs, target genes of the miRNAs were predicted and cis/trans-regulation analysis of the lncRNAs were performed. Copyright © 2016 Elsevier B.V. All rights reserved.
Glutamic acid decarboxylase 67 expression by a distinct population of mouse vestibular supporting cells.

Science.gov (United States)

Tavazzani, Elisa; Tritto, Simona; Spaiardi, Paolo; Botta, Laura; Manca, Marco; Prigioni, Ivo; Masetto, Sergio; Russo, Giancarlo

2014-01-01

The function of the enzyme glutamate decarboxylase (GAD) is to convert glutamate in γ-aminobutyric acid (GABA). Glutamate decarboxylase exists as two major isoforms, termed GAD65 and GAD67, that are usually expressed in GABA-containing neurons in the central nervous system. GAD65 has been proposed to be associated with GABA exocytosis whereas GAD67 with GABA metabolism. In the present immunofluorescence study, we have investigated the presence of the two GAD isoforms in the semicircular canal cristae of wild type and GAD67-GFP knock-in mice. While no evidence for GAD65 expression was found, GAD67 was detected in a distinct population of peripherally-located supporting cells, but not in hair cells or in centrally-located supporting cells. GABA, on the other hand, was found in all supporting cells. The present result indicate that only a discrete population of supporting cells use GAD67 to synthesize GABA. This is the first report of a marker that allows to distinguish two populations of supporting cells in the vestibular epithelium. On the other hand, the lack of GABA and GAD enzymes in hair cells excludes its involvement in afferent transmission.
Glutamic acid decarboxylase 67 expression by a distinct population of mouse vestibular supporting cells

Directory of Open Access Journals (Sweden)

Giancarlo eRusso

2014-12-01

Full Text Available The function of the enzyme glutamate decarboxylase (GAD is to convert glutamate in -aminobutyric acid (GABA.GAD exists as two major isoforms, termed GAD65 and GAD67,.that are usually expressed in GABA-containing neurons in the central nervous system. GAD65 has been proposed to be associated with GABA exocytosis whereas GAD67 with GABA metabolism. In the present immunofluorescence study, we have investigated the presence of the two GAD isoforms in the semicircular canal cristae of wild type and GAD67-GFP knock-in mice. While no evidence for GAD65 expression was found, GAD67 was detected in a distinct population of peripherally-located supporting cells, but not in hair cells or in centrally-located supporting cells. GABA, on the other hand, was found in all supporting cells. The present result indicate that only a discrete population of supporting cells use GAD67 to synthesize GABA. This is the first report of a marker that allows to distinguish two populations of supporting cells in the vestibular epithelium. On the other hand, the lack of GABA and GAD enzymes in hair cells excludes its involvement in afferent transmission.
Distinct UV-B and UV-A/blue light signal transduction pathways induce chalcone synthase gene expression in Arabidopsis cells

International Nuclear Information System (INIS)

Christie, J.M.; Jenkins, G.I.

1996-01-01

UV and blue light control the expression of flavonoid biosynthesis genes in a range of higher plants. To investigate the signal transduction processes involved in the induction of chalcone synthase (CHS) gene expression by UV-B and UV-A/blue light, we examined the, effects of specific agonists and inhibitors of known signaling components in mammalian systems in a photomixotrophic Arabidopsis cell suspension culture. CHS expression is induced specifically by these wavelengths in the cell culture, in a manner similar to that in mature Arabidopsis leaf tissue. Both the UV-B and UV-A/blue phototransduction processes involve calcium, although the elevation of cytosolic calcium is insufficient on its own to stimulate CHS expression. The UV-A/blue light induction of CHS expression does not appear to involve calmodulin, whereas the UV-B response does; this difference indicates that the signal transduction pathways are, at least in part, distinct. We provide evidence that both pathways involve reversible protein phosphorylation and require protein synthesis. The UV-B and UV-A/blue light signaling pathways are therefore different from the phytochrome signal transduction pathway regulating CHS expression in other species
Predicting the minimum liquid surface tension activity of pseudomonads expressing biosurfactants.

Science.gov (United States)

Mohammed, I U; Deeni, Y; Hapca, S M; McLaughlin, K; Spiers, A J

2015-01-01

Bacteria produce a variety of biosurfactants capable of significantly reducing liquid (aqueous) surface tension (γ) with a range of biological roles and biotechnological uses. To determine the lowest achievable surface tension (γMin ), we tested a diverse collection of Pseudomonas-like isolates from contaminated soil and activated sludge and identified those expressing biosurfactants by drop-collapse assay. Liquid surface tension-reducing ability was quantitatively determined by tensiometry, with 57 isolates found to significantly lower culture supernatant surface tensions to 24·5-49·1 mN m(-1) . Differences in biosurfactant behaviour determined by foaming, emulsion and oil-displacement assays were also observed amongst isolates producing surface tensions of 25-27 mN m(-1) , suggesting that a range of structurally diverse biosurfactants were being expressed. Individual distribution identification (IDI) analysis was used to identify the theoretical probability distribution that best fitted the surface tension data, which predicted a γMin of 24·24 mN m(-1) . This was in agreement with predictions based on earlier work of published mixed bacterial spp. data, suggesting a fundamental limit to the ability of bacterial biosurfactants to reduce surface tensions in aqueous systems. This implies a biological restriction on the synthesis and export of these agents or a physical-chemical restriction on their functioning once produced. Numerous surveys of biosurfactant-producing bacteria have been conducted, but only recently has an attempt been made to predict the minimum liquid surface tension these surface-active agents can achieve. Here, we determine a theoretical minimum of 24 mN m(-1) by statistical analysis of tensiometry data, suggesting a fundamental limit for biosurfactant activity in bacterial cultures incubated under standard growth conditions. This raises a challenge to our understanding of biosurfactant expression, secretion and function, as well as
Pentameric ligand-gated ion channels exhibit distinct transmembrane domain archetypes for folding/expression and function.

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Therien, J P Daniel; Baenziger, John E

2017-03-27

Although transmembrane helix-helix interactions must be strong enough to drive folding, they must still permit the inter-helix movements associated with conformational change. Interactions between the outermost M4 and adjacent M1 and M3 α-helices of pentameric ligand-gated ion channels have been implicated in folding and function. Here, we evaluate the role of different physical interactions at this interface in the function of two prokaryotic homologs, GLIC and ELIC. Strikingly, disruption of most interactions in GLIC lead to either a reduction or a complete loss of expression and/or function, while analogous disruptions in ELIC often lead to gains in function. Structural comparisons suggest that GLIC and ELIC represent distinct transmembrane domain archetypes. One archetype, exemplified by GLIC, the glycine and GABA receptors and the glutamate activated chloride channel, has extensive aromatic contacts that govern M4-M1/M3 interactions and that are essential for expression and function. The other archetype, exemplified by ELIC and both the nicotinic acetylcholine and serotonin receptors, has relatively few aromatic contacts that are detrimental to function. These archetypes likely have evolved different mechanisms to balance the need for strong M4 "binding" to M1/M3 to promote folding/expression, and the need for weaker interactions that allow for greater conformational flexibility.
The pursuit of optimal distinctiveness and consumer preferences.

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He, Lingnan; Cong, Feng; Liu, Yanping; Zhou, Xinyue

2010-10-01

This article investigates the effect of optimal distinctiveness on consumer product consumption. The authors argue that consumers acquire and display material possessions to restore their optimal levels of distinctiveness. Results showed that placing consumers in a state of low distinctiveness increased desire to acquire distinctive products, whereas perceptions of high distinctiveness reduced desire to acquire such products. Consumers' desire for distinctiveness-related products held true for various consumer choices, including willingness to pay more for limited-edition products and preference for unpopular gifts. This finding has implications for understanding consumer choice in expressing identity. © 2010 The Authors. Scandinavian Journal of Psychology © 2010 The Scandinavian Psychological Associations.
Processing vertical size disparities in distinct depth planes.

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Duke, Philip A; Howard, Ian P

2012-08-17

A textured surface appears slanted about a vertical axis when the image in one eye is horizontally enlarged relative to the image in the other eye. The surface appears slanted in the opposite direction when the same image is vertically enlarged. Two superimposed textured surfaces with different horizontal size disparities appear as two surfaces that differ in slant. Superimposed textured surfaces with equal and opposite vertical size disparities appear as a single frontal surface. The vertical disparities are averaged. We investigated whether vertical size disparities are averaged across two superimposed textured surfaces in different depth planes or whether they induce distinct slants in the two depth planes. In Experiment 1, two superimposed textured surfaces with different vertical size disparities were presented in two depth planes defined by horizontal disparity. The surfaces induced distinct slants when the horizontal disparity was more than ±5 arcmin. Thus, vertical size disparities are not averaged over surfaces with different horizontal disparities. In Experiment 2 we confirmed that vertical size disparities are processed in surfaces away from the horopter, so the results of Experiment 1 cannot be explained by the processing of vertical size disparities in a fixated surface only. Together, these results show that vertical size disparities are processed separately in distinct depth planes. The results also suggest that vertical size disparities are not used to register slant globally by their effect on the registration of binocular direction of gaze.
Temporal regulation of HTLV-2 expression in infected cell lines and patients: evidence for distinct expression kinetics with nuclear accumulation of APH-2 mRNA

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Bender Cecilia

2012-09-01

Full Text Available Abstract Background Human T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2 are delta retroviruses with similar genetic organization. Although both viruses immortalize T-cells in vitro, they exhibit distinct pathogenic potential in vivo. To search for possible differences in its expression strategy with respect to HTLV-1, we investigated the pattern of HTLV-2 expression in infected cell lines and peripheral blood mononuclear cells (PBMCs from infected patients using splice site-specific quantitative RT-PCR. Findings A novel alternative splice acceptor site for exon 2 was identified; its usage in env transcripts was found to be subtype-specific. Time-course analysis revealed a two-phase expression kinetics in an infected cell line and in PBMCs of two of the three patients examined; this pattern was reminiscent of HTLV-1. In addition, the minus-strand APH2 transcript was mainly detected in the nucleus, a feature that was similar to its HTLV-1 orthologue HBZ. In contrast to HTLV-1, expression of the mRNA encoding the main regulatory proteins Tax and Rex and that of the mRNAs encoding the p28 and truncated Rex inhibitors is skewed towards p28/truncated Rex inhibitors in HTLV-2. Conclusion Our data suggest a general converging pattern of expression of HTLV-2 and HTLV-1 and highlight peculiar differences in the expression of regulatory proteins that might influence the pathobiology of these viruses.
Interactions between β-catenin and the HSlo potassium channel regulates HSlo surface expression.

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Shumin Bian

Full Text Available The large conductance calcium-activated potassium channel alpha-subunit (Slo is widely distributed throughout the body and plays an important role in a number of diseases. Prior work has shown that Slo, through its S10 region, interacts with β-catenin, a key component of the cytoskeleton framework and the Wnt signaling pathway. However, the physiological significance of this interaction was not clear.Using a combination of proteomic and cell biology tools we show the existence of additional multiple binding sites in Slo, and explore in detail β-catenin interactions with the S10 region. We demonstrate that deletion of this region reduces Slo surface expression in HEK cells, which indicates that interaction with beta-catenin is important for Slo surface expression. This is confirmed by reduced expression of Slo in HEK cells and chicken (Gallus gallus domesticus leghorn white hair cells treated with siRNA to β-catenin. HSlo reciprocally co-immunoprecipitates with β-catenin, indicating a stable binding between these two proteins, with the S10 deletion mutant having reduced binding with β-catenin. We also observed that mutations of the two putative GSK phosphorylation sites within the S10 region affect both the surface expression of Slo and the channel's voltage and calcium sensitivities. Interestingly, expression of exogenous Slo in HEK cells inhibits β-catenin-dependent canonical Wnt signaling.These studies identify for the first time a central role for β-catenin in mediating Slo surface expression. Additionally we show that Slo overexpression can lead to downregulation of Wnt signaling.
Cell surface expression of single chain antibodies with applications to imaging of gene expression in vivo

International Nuclear Information System (INIS)

Northrop, Jeffrey P.; Bednarski, Mark; Li, King C.; Barbieri, Susan O.; Lu, Amy T.; Nguyen, Dee; Varadarajan, John; Osen, Maureen; Star-Lack, Josh

2003-01-01

Imaging of gene expression in vivo has many potential uses for biomedical research and drug discovery, ranging from the study of gene regulation and cancer to the non-invasive assessment of gene therapies. To streamline the development of imaging marker gene technologies for nuclear medicine, we propose a new approach to the design of reporter/probe pairs wherein the reporter is a cell surface-expressed single chain antibody variable fragment that has been raised against a low molecular weight imaging probe with optimized pharmacokinetic properties. Proof of concept of the approach was achieved using a single chain antibody variable fragment that binds with high affinity to fluorescein and an imaging probe consisting of fluorescein isothiocyanate coupled to the chelator diethylene triamine penta-acetic acid labeled with the gamma-emitter 111 In. We demonstrate specific high-affinity binding of this probe to the cell surface-expressed reporter in vitro and assess the in vivo biodistribution of the probe both in wild-type mice and in mice harboring tumor xenografts expressing the reporter. Specific uptake of the probe by, and in vivo imaging of, tumors expressing the reporter are shown. Since ScFvs with high affinities can be raised to almost any protein or small molecule, the proposed methodology may offer a new flexibility in the design of imaging tracer/reporter pairs wherein both probe pharmacokinetics and binding affinities can be readily optimized. (orig.)
Dietary soy and meat proteins induce distinct physiological and gene expression changes in rats

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Song, Shangxin; Hooiveld, Guido J.; Li, Mengjie; Zhao, Fan; Zhang, Wei; Xu, Xinglian; Muller, Michael; Li, Chunbao; Zhou, Guanghong

2016-01-01

This study reports on a comprehensive comparison of the effects of soy and meat proteins given at the recommended level on physiological markers of metabolic syndrome and the hepatic transcriptome. Male rats were fed semi-synthetic diets for 1 wk that differed only regarding protein source, with casein serving as reference. Body weight gain and adipose tissue mass were significantly reduced by soy but not meat proteins. The insulin resistance index was improved by soy, and to a lesser extent by meat proteins. Liver triacylglycerol contents were reduced by both protein sources, which coincided with increased plasma triacylglycerol concentrations. Both soy and meat proteins changed plasma amino acid patterns. The expression of 1571 and 1369 genes were altered by soy and meat proteins respectively. Functional classification revealed that lipid, energy and amino acid metabolic pathways, as well as insulin signaling pathways were regulated differently by soy and meat proteins. Several transcriptional regulators, including NFE2L2, ATF4, Srebf1 and Rictor were identified as potential key upstream regulators. These results suggest that soy and meat proteins induce distinct physiological and gene expression responses in rats and provide novel evidence and suggestions for the health effects of different protein sources in human diets. PMID:26857845
Dopamine receptors play distinct roles in sexual behavior expression of rats with a different sexual motivational tone.

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Guadarrama-Bazante, Irma L; Canseco-Alba, Ana; Rodríguez-Manzo, Gabriela

2014-10-01

Dopamine (DA) plays a central role in the expression of male sexual behavior. The effects of DA-enhancing drugs on copulation seem to vary depending on the dose of the agonist used, the type of DA receptor activated, and the sexual condition of the animals. The aim of the present study was to carry out a systematic analysis of the effects of dopaminergic agonists on the expression of male sexual behavior by sexually competent rats in different sexual motivational states, that is when sexually active (sexually experienced) and when temporarily inhibited (sexually exhausted). To this end, the same doses of the nonselective DA receptor agonist apomorphine, the selective D2-like DA receptor agonist quinpirole, and the selective D1-like DA receptor agonist SKF38393 were injected intraperitoneally to sexually experienced or sexually exhausted male rats and their sexual behavior was recorded. Low apomorphine doses induced expression of sexual behavior in sexually satiated rats, but only reduced the intromission latency of sexually experienced rats. SKF38393 facilitated the expression of sexual behavior by sexually exhausted rats, but not that of sexually experienced males and quinpirole did not exert an effect in both types of animal. In line with these results, the apomorphine-induced reversal of sexual exhaustion was blocked by the D1-like receptor antagonist SCH23390. The data suggest that DA receptors play distinct roles in the expression of sexual behavior by male rats depending on their motivational state and that activation of D1-like receptors promotes the expression of sexual behavior in satiated rats.
Single Particle Tracking reveals two distinct environments for CD4 receptors at the surface of living T lymphocytes

International Nuclear Information System (INIS)

Mascalchi, Patrice; Lamort, Anne Sophie; Salomé, Laurence; Dumas, Fabrice

2012-01-01

Highlights: ► We studied the diffusion of single CD4 receptors on living lymphocytes. ► This study reveals that CD4 receptors have either a random or confined diffusion. ► The dynamics of unconfined CD4 receptors was accelerated by a temperature raise. ► The dynamics of confined CD4 receptors was unchanged by a temperature raise. ► Our results suggest the existence of two different environments for CD4 receptors. -- Abstract: We investigated the lateral diffusion of the HIV receptor CD4 at the surface of T lymphocytes at 20 °C and 37 °C by Single Particle Tracking using Quantum Dots. We found that the receptors presented two major distinct behaviors that were not equally affected by temperature changes. About half of the receptors showed a random diffusion with a diffusion coefficient increasing upon raising the temperature. The other half of the receptors was permanently or transiently confined with unchanged dynamics on raising the temperature. These observations suggest that two distinct subpopulations of CD4 receptors with different environments are present at the surface of living T lymphocytes.
Expression, immunogenicity and variation of iron-regulated surface protein A from bovine isolates of Staphylococcus aureus

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Misra, Neha; Wines, Tyler F.; Knopp, Colton L.; McGuire, Mark A.; Tinker, Juliette K.

2017-01-01

Abstract Staphylococcus aureus iron-regulated surface protein A (IsdA) is a fibrinogen and fibronectin adhesin that also contributes to iron sequestration and resistance to innate immunity. IsdA is conserved in human isolates and has been investigated as a human vaccine candidate. Here we report the expression of isdA, the efficacy of anti-IsdA responses and the existence of IsdA sequence variants from bovine Staphylococcus. Clinical staphylococci were obtained from US dairy farms and assayed by PCR for the presence and expression of isdA. isdA-positive species from bovines included S. aureus, S. haemolyticus and S. chromogenes. Immunoassays on bovine milk and serum confirmed the induction and opsonophagocytic activity of anti-IsdA humoral responses. The variable region of isdA was sequenced and protein alignments predicted the presence of two main variants consistent with those from human S. aureus. Mouse antibodies against one IsdA variant reduced staphylococcal binding to fibronectin in vitro in an isotype-dependent manner. Purified IsdA variants bound distinctly to fibronectin and fibrinogen. Our findings demonstrate that variability within the C-terminus of this adhesin affects immune reactivity and binding specificity, but are consistent with the significance of IsdA in bovine disease and relevant for vaccine development. PMID:28430959
Expression and display of UreA of Helicobacter acinonychis on the surface of Bacillus subtilis spores

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De Felice Maurilio

2010-01-01

Full Text Available Abstract Background The bacterial endospore (spore has recently been proposed as a new surface display system. Antigens and enzymes have been successfully exposed on the surface layers of the Bacillus subtilis spore, but only in a few cases the efficiency of expression and the effective surface display and have been determined. We used this heterologous expression system to produce the A subunit of the urease of the animal pathogen Helicobater acinonychis. Ureases are multi-subunit enzymes with a central role in the virulence of various bacterial pathogens and necessary for colonization of the gastric mucosa by the human pathogen H. pylori. The urease subunit UreA has been recognized as a major antigen, able to induce high levels of protection against challenge infections. Results We expressed UreA from H. acinonychis on the B. subtilis spore coat by using three different spore coat proteins as carriers and compared the efficiency of surface expression and surface display obtained with the three carriers. A combination of western-, dot-blot and immunofluorescence microscopy allowed us to conclude that, when fused to CotB, UreA is displayed on the spore surface (ca. 1 × 103 recombinant molecules per spore, whereas when fused to CotC, although most efficiently expressed (7-15 × 103 recombinant molecules per spore and located in the coat layer, it is not displayed on the surface. Experiments with CotG gave results similar to those with CotC, but the CotG-UreA recombinant protein appeared to be partially processed. Conclusion UreA was efficiently expressed on the spore coat of B. subtilis when fused to CotB, CotC or CotG. Of these three coat proteins CotC allows the highest efficiency of expression, whereas CotB is the most appropriate for the display of heterologous proteins on the spore surface.
Expression and display of UreA of Helicobacter acinonychis on the surface of Bacillus subtilis spores.

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Hinc, Krzysztof; Isticato, Rachele; Dembek, Marcin; Karczewska, Joanna; Iwanicki, Adam; Peszyńska-Sularz, Grazyna; De Felice, Maurilio; Obuchowski, Michał; Ricca, Ezio

2010-01-18

The bacterial endospore (spore) has recently been proposed as a new surface display system. Antigens and enzymes have been successfully exposed on the surface layers of the Bacillus subtilis spore, but only in a few cases the efficiency of expression and the effective surface display and have been determined. We used this heterologous expression system to produce the A subunit of the urease of the animal pathogen Helicobater acinonychis. Ureases are multi-subunit enzymes with a central role in the virulence of various bacterial pathogens and necessary for colonization of the gastric mucosa by the human pathogen H. pylori. The urease subunit UreA has been recognized as a major antigen, able to induce high levels of protection against challenge infections. We expressed UreA from H. acinonychis on the B. subtilis spore coat by using three different spore coat proteins as carriers and compared the efficiency of surface expression and surface display obtained with the three carriers. A combination of western-, dot-blot and immunofluorescence microscopy allowed us to conclude that, when fused to CotB, UreA is displayed on the spore surface (ca. 1 x 10(3) recombinant molecules per spore), whereas when fused to CotC, although most efficiently expressed (7-15 x 10(3) recombinant molecules per spore) and located in the coat layer, it is not displayed on the surface. Experiments with CotG gave results similar to those with CotC, but the CotG-UreA recombinant protein appeared to be partially processed. UreA was efficiently expressed on the spore coat of B. subtilis when fused to CotB, CotC or CotG. Of these three coat proteins CotC allows the highest efficiency of expression, whereas CotB is the most appropriate for the display of heterologous proteins on the spore surface.
Surface-expressed enolases of Plasmodium and other pathogens

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Anil Kumar Ghosh

2011-08-01

Full Text Available Enolase is the eighth enzyme in the glycolytic pathway, a reaction that generates ATP from phosphoenol pyruvate in cytosolic compartments. Enolase is essential, especially for organisms devoid of the Krebs cycle that depend solely on glycolysis for energy. Interestingly, enolase appears to serve a separate function in some organisms, in that it is also exported to the cell surface via a poorly understood mechanism. In these organisms, surface enolase assists in the invasion of their host cells by binding plasminogen, an abundant plasma protease precursor. Binding is mediated by the interaction between a lysine motif of enolase with Kringle domains of plasminogen. The bound plasminogen is then cleaved by specific proteases to generate active plasmin. Plasmin is a potent serine protease that is thought to function in the degradation of the extracellular matrix surrounding the targeted host cell, thereby facilitating pathogen invasion. Recent work revealed that the malaria parasite Plasmodium also expresses surface enolase, and that this feature may be essential for completion of its life cycle. The therapeutic potential of targeting surface enolases of pathogens is discussed.

B7h-expressing dendritic cells and plasma B cells mediate distinct outcomes of ICOS costimulation in T cell-dependent antibody responses

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Larimore Kevin

2012-06-01

Full Text Available Abstract Background The ICOS-B7h costimulatory receptor-ligand pair is required for germinal center formation, the production of isotype-switched antibodies, and antibody affinity maturation in response to T cell-dependent antigens. However, the potentially distinct roles of regulated B7h expression on B cells and dendritic cells in T cell-dependent antibody responses have not been defined. Results We generated transgenic mice with lineage-restricted B7h expression to assess the cell-type specific roles of B7h expression on B cells and dendritic cells in regulating T cell-dependent antibody responses. Our results show that endogenous B7h expression is reduced on B cells after activation in vitro and is also reduced in vivo on antibody-secreting plasma B cells in comparison to both naïve and germinal center B cells from which they are derived. Increasing the level of B7h expression on activated and plasma B cells in B-B7hTg mice led to an increase in the number of antibody-secreting plasma cells generated after immunization and a corresponding increase in the concentration of antigen-specific high affinity serum IgG antibodies of all isotypes, without affecting the number of responding germinal center B cells. In contrast, ICOS costimulation mediated by dendritic cells in DC-B7hTg mice contributed to germinal center formation and selectively increased IgG2a production without affecting the overall magnitude of antibody responses. Conclusions Using transgenic mice with lineage-restricted B7h expression, we have revealed distinct roles of ICOS costimulation mediated by dendritic cells and B cells in the regulation of T cell-dependent antibody responses.
Distinct mutations in yeast TAF(II)25 differentially affect the composition of TFIID and SAGA complexes as well as global gene expression patterns.

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Kirschner, Doris B; vom Baur, Elmar; Thibault, Christelle; Sanders, Steven L; Gangloff, Yann-Gaël; Davidson, Irwin; Weil, P Anthony; Tora, Làszlò

2002-05-01

The RNA polymerase II transcription factor TFIID, composed of the TATA-binding protein (TBP) and TBP-associated factors (TAF(II)s), nucleates preinitiation complex formation at protein-coding gene promoters. SAGA, a second TAF(II)-containing multiprotein complex, is involved in transcription regulation in Saccharomyces cerevisiae. One of the essential protein components common to SAGA and TFIID is yTAF(II)25. We define a minimal evolutionarily conserved 91-amino-acid region of TAF(II)25 containing a histone fold domain that is necessary and sufficient for growth in vivo. Different temperature-sensitive mutations of yTAF(II)25 or chimeras with the human homologue TAF(II)30 arrested cell growth at either the G(1) or G(2)/M cell cycle phase and displayed distinct phenotypic changes and gene expression patterns. Immunoprecipitation studies revealed that TAF(II)25 mutation-dependent gene expression and phenotypic changes correlated at least partially with the integrity of SAGA and TFIID. Genome-wide expression analysis revealed that the five TAF(II)25 temperature-sensitive mutant alleles individually affect the expression of between 18 and 33% of genes, whereas taken together they affect 64% of all class II genes. Thus, different yTAF(II)25 mutations induce distinct phenotypes and affect the regulation of different subsets of genes, demonstrating that no individual TAF(II) mutant allele reflects the full range of its normal functions.
Role of Site-Specific N-Glycans Expressed on GluA2 in the Regulation of Cell Surface Expression of AMPA-Type Glutamate Receptors.

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Yusuke Takeuchi

Full Text Available The AMPA-type glutamate receptor (AMPAR, which is a tetrameric complex composed of four subunits (GluA1-4 with several combinations, mediates the majority of rapid excitatory synaptic transmissions in the nervous system. Cell surface expression levels of AMPAR modulate synaptic plasticity, which is considered one of the molecular bases for learning and memory formation. To date, a unique trisaccharide (HSO3-3GlcAβ1-3Galβ1-4GlcNAc, human natural killer-1 (HNK-1 carbohydrate, was found expressed specifically on N-linked glycans of GluA2 and regulated the cell surface expression of AMPAR and the spine maturation process. However, evidence that the HNK-1 epitope on N-glycans of GluA2 directly affects these phenomena is lacking. Moreover, it is thought that other N-glycans on GluA2 also have potential roles in the regulation of AMPAR functions. In the present study, using a series of mutants lacking potential N-glycosylation sites (N256, N370, N406, and N413 within GluA2, we demonstrated that the mutant lacking the N-glycan at N370 strongly suppressed the intracellular trafficking of GluA2 from the endoplasmic reticulum (ER in HEK293 cells. Cell surface expression of GluA1, which is a major subunit of AMPAR in neurons, was also suppressed by co-expression of the GluA2 N370S mutant. The N370S mutant and wild-type GluA2 were co-immunoprecipitated with GluA1, suggesting that N370S was properly associated with GluA1. Moreover, we found that N413 was the main potential site of the HNK-1 epitope that promoted the interaction of GluA2 with N-cadherin, resulting in enhanced cell surface expression of GluA2. The HNK-1 epitope on N-glycan at the N413 of GluA2 was also involved in the cell surface expression of GluA1. Thus, our data suggested that site-specific N-glycans on GluA2 regulate the intracellular trafficking and cell surface expression of AMPAR.
A sensitive electrochemiluminescence cytosensor for quantitative evaluation of epidermal growth factor receptor expressed on cell surfaces

Energy Technology Data Exchange (ETDEWEB)

Tang, Yanjuan; Zhang, Shaolian; Wen, Qingqing; Huang, Hongxing; Yang, Peihui, E-mail: typh@jnu.edu.cn

2015-06-30

Highlights: • EGF-cytosensor was used for evaluating EGFR expression level on cell surfaces. • CdSQDs and EGF were coated on magnetic beads (MBs) for ECL-probe. • Good sensitivity was achieved due to the signal amplification of ECL-probe. - Abstract: A sensitive electrochemiluminescence (ECL) strategy for evaluating the epidermal growth factor receptor (EGFR) expression level on cell surfaces was designed by integrating the specific recognition of EGFR expressed on MCF-7 cell surfaces with an epidermal growth factor (EGF)-funtionalized CdS quantum dots (CdSQDs)-capped magnetic bead (MB) probe. The high sensitivity of ECL probe of EGF-funtionalized CdSQD-capped-MB was used for competitive recognition with EGFR expressed on cell surfaces with recombinant EGFR protein. The changes of ECL intensity depended on both the cell number and the expression level of EGFR receptor on cell surfaces. A wide linear response to cells ranging from 80 to 4 × 10{sup 6} cells mL{sup −1} with a detection limit of 40 cells mL{sup −1} was obtained. The EGF-cytosensor was used to evaluate EGFR expression levels on MCF-7 cells, and the average number of EGFR receptor on single MCF-7 cells was 1.35 × 10{sup 5} with the relative standard deviation of 4.3%. This strategy was further used for in-situ and real-time evaluating EGFR receptor expressed on cell surfaces in response to drugs stimulation at different concentration and incubation time. The proposed method provided potential applications in the detection of receptors on cancer cells and anticancer drugs screening.
The establishment of Saccharomyces boulardii surface display system using a single expression vector.

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Wang, Tiantian; Sun, Hui; Zhang, Jie; Liu, Qing; Wang, Longjiang; Chen, Peipei; Wang, Fangkun; Li, Hongmei; Xiao, Yihong; Zhao, Xiaomin

2014-03-01

In the present study, an a-agglutinin-based Saccharomyces boulardii surface display system was successfully established using a single expression vector. Based on the two protein co-expression vector pSP-G1 built by Partow et al., a S. boulardii surface display vector-pSDSb containing all the display elements was constructed. The display results of heterologous proteins were confirmed by successfully displaying enhanced green fluorescent protein (EGFP) and chicken Eimeria tenella Microneme-2 proteins (EtMic2) on the S. boulardii cell surface. The DNA sequence of AGA1 gene from S. boulardii (SbAGA1) was determined and used as the cell wall anchor partner. This is the first time heterologous proteins have been displayed on the cell surface of S. boulardii. Because S. boulardii is probiotic and eukaryotic, its surface display system would be very valuable, particularly in the development of a live vaccine against various pathogenic organisms especially eukaryotic pathogens such as protistan parasites. Copyright © 2013 Elsevier Inc. All rights reserved.
Bipotential precursors of putative fibrous astrocytes and oligodendrocytes in rat cerebellar cultures express distinct surface features and neuron-like γ-aminobutyric acid transport

International Nuclear Information System (INIS)

Levi, G.; Gallo, V.; Ciotti, T.

1986-01-01

When postnatal rat cerebellar cells were cultured in a chemically defined, serum-free medium, the only type of astrocyte present was unable to accumulate γ-[ 3 H]aminobutyric acid (GABA), did not express surface antigens recognized by two monoclonal antibodies, A2B5 and LB1, and showed minimal proliferation. In these cultures, nonneuronal A2B5 + , LB1 + stellate cells exhibiting neuron-like [ 3 H]GABA uptake formed cell colonies of increasing size and were GFAP - . After about one week of culturing, the A2B5 + , LB1 + , GABA-uptake positive cell groups became galactocerebroside (GalCer) positive. Immunocytolysis of the A2B5 + cells at 3 and 4 days in vitro prevented the appearance of the A2B5 + , LB1 + , GABA-uptake positive cell colonies, and also of the GalCer + cell groups. If 10% (vol/vol) fetal calf serum was added to 6-day cultures, the A2B5 + , LB1 + , GABA-uptake positive cell groups expressed GFAP and not GalCer. If the serum was added to the cultures 2 days after lysing the A2B5 + cells, only A2B5 - , LB1 - , GABA-uptake negative astrocytes proliferated. It is concluded that the putative fibrous astrocytes previously described in serum-containing cultures derive from bipotential precursors that differentiate into oligodendrocytes (GalCer + ) in serum-free medium or into astrocytes (GFAP + ) in the presence of serum, while the epithelioid A2B5 - , LB1 - , GABA-uptake negative astrocytes originate from a different precursor not yet identified
Divalent cations and the protein surface co-ordinate the intensity of human platelet adhesion and P-selectin surface expression.

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Whiss, P A; Andersson, R G G

2002-07-01

At sites of blood vessel injury, platelets adhere to exposed vessel components, such as collagen, or immobilized fibrinogen derived from plasma or activated platelets. The divalent cations Mg(2+) and Ca(2+) are essential for platelet adhesion and activation, but Mg(2+) can also inhibit platelet activation. The present study evaluates, by an enzymatic method, the effects of various divalent cations on the adhesion of isolated human platelets to collagen, fibrinogen, albumin or plastic in vitro. By enzyme-linked immunosorbent assay, platelet surface expression of P-selectin was measured to estimate the state of activation on adherence. Mg(2+) increased platelet adhesion exclusively to collagen and fibrinogen at physiologically relevant concentrations. At higher concentrations, the adhesion declined. Ca(2+) induced a weak adhesion only to fibrinogen at physiological doses and a peak of increased adhesion to all protein-coated surfaces at 10 mmol/l. Mn(2+) elicited dose-dependent adhesion only to collagen and fibrinogen. Zn(2+), Ni(2+) and Cu(2+) increased the adhesion of platelets independently of the surface. Ca(2+) dose-dependently inhibited adhesion elicited by Mg(2+) to collagen and fibrinogen. No other combination of divalent cations elicited such an effect. Mg(2+)-dependent platelet adhesion to collagen and Ca(2+)-dependent adhesion to fibrinogen increased P-selectin expression. Thus, the present study shows that the outcome of the platelet adhesion depends on the surface and the access of divalent cations, which co-ordinate the intensity of platelet adhesion and P-selectin surface expression.
Expressions of the radius and the surface tension of surface of tension in terms of the pressure distribution for nanoscale liquid threads

International Nuclear Information System (INIS)

Yan Hong; Wei Jiu-An; Cui Shu-Wen; Zhu Ru-Zeng

2013-01-01

The expressions of the radius and the surface tension of surface of tension R s and γ s in terms of the pressure distribution for nanoscale liquid threads are of great importance for molecular dynamics (MD) simulations of the interfacial phenomena of nanoscale fluids; these two basic expressions are derived in this paper. Although these expressions were derived first in the literature [Kim B G, Lee J S, Han M H, and Park S, 2006 Nanoscale and Microscale Thermophysical Engineering, 10, 283] and used widely thereafter, the derivation is wrong both in logical structure and physical thought. In view of the importance of these basic expressions, the logic and physical mistakes appearing in that derivation are pointed out. (condensed matter: structural, mechanical, and thermal properties)
Expression, immunogenicity and variation of iron-regulated surface protein A from bovine isolates of Staphylococcus aureus.

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Misra, Neha; Wines, Tyler F; Knopp, Colton L; McGuire, Mark A; Tinker, Juliette K

2017-05-01

Staphylococcus aureus iron-regulated surface protein A (IsdA) is a fibrinogen and fibronectin adhesin that also contributes to iron sequestration and resistance to innate immunity. IsdA is conserved in human isolates and has been investigated as a human vaccine candidate. Here we report the expression of isdA, the efficacy of anti-IsdA responses and the existence of IsdA sequence variants from bovine Staphylococcus. Clinical staphylococci were obtained from US dairy farms and assayed by PCR for the presence and expression of isdA. isdA-positive species from bovines included S. aureus, S. haemolyticus and S. chromogenes. Immunoassays on bovine milk and serum confirmed the induction and opsonophagocytic activity of anti-IsdA humoral responses. The variable region of isdA was sequenced and protein alignments predicted the presence of two main variants consistent with those from human S. aureus. Mouse antibodies against one IsdA variant reduced staphylococcal binding to fibronectin in vitro in an isotype-dependent manner. Purified IsdA variants bound distinctly to fibronectin and fibrinogen. Our findings demonstrate that variability within the C-terminus of this adhesin affects immune reactivity and binding specificity, but are consistent with the significance of IsdA in bovine disease and relevant for vaccine development. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Whole genome expression profiling using DNA microarray for determining biocompatibility of polymeric surfaces

DEFF Research Database (Denmark)

Stangegaard, Michael; Wang, Zhenyu; Kutter, Jörg Peter

2006-01-01

There is an ever increasing need to find surfaces that are biocompatible for applications like medical implants and microfluidics-based cell culture systems. The biocompatibility of five different surfaces with different hydrophobicity was determined using gene expression profiling as well as more...
The expression pattern of small nucleolar and small Cajal body-specific RNAs characterizes distinct molecular subtypes of multiple myeloma

International Nuclear Information System (INIS)

Ronchetti, D; Todoerti, K; Tuana, G; Agnelli, L; Mosca, L; Lionetti, M; Fabris, S; Colapietro, P; Miozzo, M; Ferrarini, M; Tassone, P; Neri, A

2012-01-01

Small nucleolar RNAs (snoRNAs) and small Cajal body-specific RNAs (scaRNAs) are non-coding RNAs involved in the maturation of other RNA molecules and generally located in the introns of host genes. It is now emerging that altered sno/scaRNAs expression may have a pathological role in cancer. This study elucidates the patterns of sno/scaRNAs expression in multiple myeloma (MM) by profiling purified malignant plasma cells from 55 MMs, 8 secondary plasma cell leukemias (sPCLs) and 4 normal controls. Overall, a global sno/scaRNAs downregulation was found in MMs and, even more, in sPCLs compared with normal plasma cells. Whereas SCARNA22 resulted the only sno/scaRNA characterizing the translocation/cyclin D4 (TC4) MM, TC2 group displayed a distinct sno/scaRNA signature overexpressing members of SNORD115 and SNORD116 families located in a region finely regulated by an imprinting center at 15q11, which, however, resulted overall hypomethylated in MMs independently of the SNORD115 and SNORD116 expression levels. Finally, integrative analyses with available gene expression and genome-wide data revealed the occurrence of significant sno/scaRNAs/host genes co-expression and the putative influence of allelic imbalances on specific snoRNAs expression. Our data extend the current view of sno/scaRNAs deregulation in cancer and add novel information to the bio-molecular complexity of plasma cell dyscrasias
Distinct effects of Cr bulk doping and surface deposition on the chemical environment and electronic structure of the topological insulator Bi{sub 2}Se{sub 3}

Energy Technology Data Exchange (ETDEWEB)

Yilmaz, Turgut, E-mail: yilmaz@phys.uconn.edu [Department of Physics, University of Connecticut, Storrs, CT 06269 (United States); Hines, William [Department of Physics, University of Connecticut, Storrs, CT 06269 (United States); Sun, Fu-Chang [Department of Materials Science and Engineering, University of Connecticut, Storrs, CT 06269 (United States); Pletikosić, Ivo [Department of Physics, Princeton University, Princeton, NJ 08544 (United States); Budnick, Joseph [Department of Physics, University of Connecticut, Storrs, CT 06269 (United States); Valla, Tonica [Condensed Matter Physics and Materials Science Department, Brookhaven National Laboratory, Upton, NY 11973 (United States); Sinkovic, Boris [Department of Physics, University of Connecticut, Storrs, CT 06269 (United States)

2017-06-15

Highlights: • Cr doping into the bulk of Bi{sub 2}Se{sub 3} opens an energy gap at the Dirac point which is observable in the non-magnetic state. • Cr surface deposition does not lead to open an energy gap at the Dirac point of Bi{sub 2}Se{sub 3}. • Formation of two distinct Bi and Cr core level peaks was observed upon the deposition of Cr on the surface of Bi{sub 2}Se{sub 3}. - Abstract: In this report, it is shown that Cr doped into the bulk and Cr deposited on the surface of Bi{sub 2}Se{sub 3} films produced by molecular beam epitaxy (MBE) have strikingly different effects on both the electronic structure and chemical environment. Angle resolved photoemission spectroscopy (ARPES) shows that Cr doped into the bulk opens a surface state energy gap which can be seen at room temperature; much higher than the measured ferromagnetic transition temperature of ≈10 K. On the other hand, similar ARPES measurements show that the surface states remain gapless down to 15 K for films with Cr surface deposition. In addition, core-level photoemission spectroscopy of the Bi 5d, Se 3d, and Cr 3p core levels show distinct differences in the chemical environment for the two methods of Cr introduction. Surface deposition of Cr results in the formation of shoulders on the lower binding energy side for the Bi 5d peaks and two distinct Cr 3p peaks indicative of two Cr sites. These striking differences suggests an interesting possibility that better control of doping at only near surface region may offer a path to quantum anomalous Hall states at higher temperatures than reported in the literature.
The promoter for a variant surface glycoprotein gene expression site in Trypanosoma brucei

NARCIS (Netherlands)

Zomerdijk, J. C.; Ouellette, M.; ten Asbroek, A. L.; Kieft, R.; Bommer, A. M.; Clayton, C. E.; Borst, P.

1990-01-01

The variant-specific surface glycoprotein (VSG) gene 221 of Trypanosoma brucei is transcribed as part of a 60 kb expression site (ES). We have identified the promoter controlling this multigene transcription unit by the use of 221 chromosome-enriched DNA libraries and VSG gene 221 expression site
Uropathogenic Escherichia coli Express Type 1 Fimbriae Only in Surface Adherent Populations Under Physiological Growth Conditions

DEFF Research Database (Denmark)

Stærk, Kristian; Kolmos, Hans Jørn; Khandige, Surabhi

2016-01-01

were correlated with the ability to adhere to and invade cultured human bladder cells. RESULTS: Although inactive during planktonic growth in urine, T1F expression occurs when UPEC settles on and infects bladder epithelial cells or colonizes catheters. As a result, UPEC in these sessile populations...... with increased expression during surface growth adaptation and infection of uroepithelial cells. This leads to separation of UPEC into low-expression planktonic populations and high-expression sessile populations....... enhances bladder cell adhesion and invasion potential. Only T1F-negative UPEC are subsequently released to the urine, thus limiting T1F expression to surface-associated UPEC alone. CONCLUSION: Our results demonstrate that T1F expression is strictly regulated under physiological growth conditions...
Expression and surface display of Cellulomonas endoglucanase in the ethanologenic bacterium Zymobacter palmae

Energy Technology Data Exchange (ETDEWEB)

Kojima, Motoki; Akahoshi, Tomohiro; Okamoto, Kenji; Yanase, Hideshi [Tottori Univ. (Japan). Dept. of Chemistry and Biotechnology

2012-11-15

In order to reduce the cost of bioethanol production from lignocellulosic biomass, we developed a tool for cell surface display of cellulolytic enzymes on the ethanologenic bacterium Zymobacter palmae. Z. palmae is a novel ethanol-fermenting bacterium capable of utilizing a broad range of sugar substrates, but not cellulose. Therefore, to express and display heterologous cellulolytic enzymes on the Z. palmae cell surface, we utilized the cell-surface display motif of the Pseudomonas ice nucleation protein Ina. The gene encoding Ina from Pseudomonas syringae IFO3310 was cloned, and its product was comprised of three functional domains: an N-terminal domain, a central domain with repeated amino acid residues, and a C-terminal domain. The N-terminal domain of Ina was shown to function as the anchoring motif for a green fluorescence protein fusion protein in Escherichia coli. To express a heterologous cellulolytic enzyme extracellularly in Z. palmae, we fused the N-terminal coding sequence of Ina to the coding sequence of an N-terminal-truncated Cellulomonas endoglucanase. Z. palmae cells carrying the fusion endoglucanase gene were shown to degrade carboxymethyl cellulose. Although a portion of the expressed fusion endoglucanase was released from Z. palmae cells into the culture broth, we confirmed the display of the protein on the cell surface by immunofluorescence microscopy. The results indicate that the N-terminal anchoring motif of Ina from P. syringae enabled the translocation and display of the heterologous cellulase on the cell surface of Z. palmae. (orig.)
Correlation of Cell Surface Biomarker Expression Levels with Adhesion Contact Angle Measured by Lateral Microscopy.

Science.gov (United States)

Walz, Jenna A; Mace, Charles R

2018-06-05

Immunophenotyping is typically achieved using flow cytometry, but any influence a biomarker may have on adhesion or surface recognition cannot be determined concurrently. In this manuscript, we demonstrate the utility of lateral microscopy for correlating cell surface biomarker expression levels with quantitative descriptions of cell morphology. With our imaging system, we observed single cells from two T cell lines and two B cell lines adhere to antibody-coated substrates and quantified this adhesion using contact angle measurements. We found that SUP-T1 and CEM CD4+ cells, both of which express similar levels of CD4, experienced average changes in contact angle that were not statistically different from one another on surfaces coated in anti-CD4. However, MAVER-1 and BJAB K20 cells, both of which express different levels of CD20, underwent average changes in contact angle that were significantly different from one another on surfaces coated in anti-CD20. Our results indicate that changes in cell contact angles on antibody-coated substrates reflect the expression levels of corresponding antigens on the surfaces of cells as determined by flow cytometry. Our lateral microscopy approach offers a more reproducible and quantitative alternative to evaluate adhesion compared to commonly used wash assays and can be extended to many additional immunophenotyping applications to identify cells of interest within heterogeneous populations.
Utility of LRF/Pokemon and NOTCH1 protein expression in the distinction between nodular lymphocyte-predominant Hodgkin lymphoma and classical Hodgkin lymphoma.

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Bohn, Olga; Maeda, Takahiro; Filatov, Alexander; Lunardi, Andrea; Pandolfi, Pier Paolo; Teruya-Feldstein, Julie

2014-02-01

Classical Hodgkin lymphoma (CHL) and nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) are considered separate entities with different prognosis and treatment. However, morphologic features can be similar and immunohistochemical studies are essential in the distinction; thus, determination of additional biomarkers is of utmost importance. LRF/Pokemon is a proto-oncogene, an interacting partner co-expressed with BCL6 in germinal centers and highly expressed in diffuse large B-cell lymphoma and follicular lymphoma. Conversely, loss of the LRF gene in mouse hematopoietic stem cells results in complete block of early B cell development with concomitant Notch de-repression, indicating its critical role in B versus T cell fate decision at the hematopoietic stem cell stage. For the first time, we show that LRF/Pokemon is predominantly expressed in NLPHL cases as is BCL6 with low to absent NOTCH1 protein expression; while Hodgkin Reed-Sternberg (HRS) cells in CHL show low to absent BCL6 and LRF/Pokemon expression with higher NOTCH1 expression. We illustrate a potential functional interaction between LRF and BCL6 in NLPHL pathogenesis, and differential expression of LRF/Pokemon and NOTCH1 proteins in CHL thus showing differential expression, making for an additional diagnostic marker and therapeutic target.
Utility of LRF/Pokemon and NOTCH1 Protein Expression in the Distinction of Nodular Lymphocyte-Predominant Hodgkin Lymphoma and Classical Hodgkin Lymphoma

Science.gov (United States)

Bohn, Olga; Maeda, Takahiro; Filatov, Alexander; Lunardi, Andrea; Pandolfi, Pier Paolo; Teruya-Feldstein, Julie

2014-01-01

Classical Hodgkin lymphoma (CHL) and nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) are considered separate entities with different prognosis and treatment. However, morphologic features can be similar and immunohistochemical studies are essential in the distinction; thus, determination of additional biomarkers is of utmost importance. LRF/Pokemon is a protooncogene, an interacting partner co-expressed with BCL6 in germinal centers and highly expressed in diffuse large B-cell lymphoma and follicular lymphoma. Conversely, loss of the LRF gene in mouse hematopoietic stem cells results in complete block of early B cell development with concomitant Notch derepression, indicating its critical role in B versus T cell fate decision at the hematopoietic stem cell stage. For the first time, we show that LRF/Pokemon is predominantly expressed in NLPHL cases as is BCL6 with low to absent NOTCH1 protein expression; while Hodgkin Reed-Sternberg (HRS) cells in CHL show low to absent BCL6 and LRF/Pokemon expression with higher NOTCH1 expression. We illustrate a potential functional interaction between LRF and BCL6 in NLPHL pathogenesis, and differential expression of LRF/Pokemon and NOTCH1 proteins in CHL thus showing differential expression, making for an additional diagnostic marker and therapeutic target. PMID:24326827
Distinctive pattern of expression of spermatogenic molecular markers in testes of azoospermic men with non-mosaic Klinefelter syndrome.

Science.gov (United States)

Kleiman, Sandra E; Yogev, Leah; Lehavi, Ofer; Yavetz, Haim; Hauser, Ron

2016-06-01

Mature sperm cells can be found in testicular specimens extracted from azoospermic men with non-mosaic Klinefelter syndrome (KS). The present study evaluates the expression of various known molecular markers of spermatogenesis in a population of men with KS and assesses the ability of those markers to predict spermatogenesis. Two groups of men with non-obstructive azoospermia who underwent testicular sperm-retrieval procedures were included in the study: 31 had non-mosaic KS (KS group) and 91 had normal karyotype (NK group). Each group was subdivided into mixed atrophy (containing some mature sperm cells) or Sertoli cell only syndrome according to testicular histology and cytology observations. Semi-quantitative histological morphometric analysis (interstitial hyperplasia and hyalinization, tubules with cells and abnormal thickness of the basement membrane) and expression of spermatogenetic markers (DAZ, RBM, BOLL, and CDY1) were evaluated and compared among those subgroups. Clear differences in the histological morphometry and spermatogenetic marker expression were noted between the KS and NK groups. There was a significant difference in the expression of spermatogenetic markers between the subgroups of the NK group (as expected), while no difference could be discerned between the two subgroups in the KS group. We conclude that molecular spermatogenetic markers have a pattern of expression in men with KS that is distinctively different from that of men with NK, and that it precludes and limits their use for predicting spermatogenesis in the former. It is suggested that this difference might be due to the specific highly abnormal histological morphometric parameters in KS specimens.
Peeling back the lithosphere: Controlling parameters, surface expressions and the future directions in delamination modeling

Science.gov (United States)

Göğüş, Oğuz H.; Ueda, Kosuke

2018-06-01

Geodynamical models investigate the rheological and physical properties of the lithosphere that peels back (delaminates) from the upper-middle crust. Meanwhile, model predictions are used to relate to a set of observations in the geological context to the test the validity of delamination. Here, we review numerical and analogue models of delamination from these perspectives and provide a number of first-order topics which future modeling studies may address. Models suggest that the presence of the weak lower crust that resides between the strong mantle lithosphere (at least 100 times more viscous/stronger) and the strong upper crust is necessary to develop delamination. Lower crustal weakening may be induced by melt infiltration, shear heating or it naturally occurs through the jelly sandwich type strength profile of the continental lithosphere. The negative buoyancy of the lithosphere required to facilitate the delamination is induced by the pre-existing ocean subduction and/or the lower crustal eclogitization. Surface expression of the peeling back lithosphere has a distinct transient and migratory imprint on the crust, resulting in rapid surface uplift/subsidence, magmatism, heating and shortening/extension. New generation of geodynamical experiments can explain how different types of melting (e.g hydrated, dry melting) occurs with delamination. Reformation of the lithosphere after removal, three dimensional aspects, and the termination of the process are key investigation areas for future research. The robust model predictions, as with other geodynamic modeling studies should be reconciled with observations.

Cardiac protein expression patterns are associated with distinct inborn exercise capacity in non-selectively bred rats

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L.P. Ribeiro

2018-01-01

Full Text Available In the present study, we successfully demonstrated for the first time the existence of cardiac proteomic differences between non-selectively bred rats with distinct intrinsic exercise capacities. A proteomic approach based on two-dimensional gel electrophoresis coupled to mass spectrometry was used to study the left ventricle (LV tissue proteome of rats with distinct intrinsic exercise capacity. Low running performance (LRP and high running performance (HRP rats were categorized by a treadmill exercise test, according to distance run to exhaustion. The running capacity of HRPs was 3.5-fold greater than LRPs. Protein profiling revealed 29 differences between HRP and LRP rats (15 proteins were identified. We detected alterations in components involved in metabolism, antioxidant and stress response, microfibrillar and cytoskeletal proteins. Contractile proteins were upregulated in the LVs of HRP rats (α-myosin heavy chain-6, myosin light chain-1 and creatine kinase, whereas the LVs of LRP rats exhibited upregulation in proteins associated with stress response (aldehyde dehydrogenase 2, α-crystallin B chain and HSPβ-2. In addition, the cytoskeletal proteins desmin and α-actin were upregulated in LRPs. Taken together, our results suggest that the increased contractile protein levels in HRP rats partly accounted for their improved exercise capacity, and that proteins considered risk factors to the development of cardiovascular disease were expressed in higher amounts in LRP animals.
Diverse Brain Myeloid Expression Profiles Reveal Distinct Microglial Activation States and Aspects of Alzheimer’s Disease Not Evident in Mouse Models

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Brad A. Friedman

2018-01-01

Full Text Available Microglia, the CNS-resident immune cells, play important roles in disease, but the spectrum of their possible activation states is not well understood. We derived co-regulated gene modules from transcriptional profiles of CNS myeloid cells of diverse mouse models, including new tauopathy model datasets. Using these modules to interpret single-cell data from an Alzheimer’s disease (AD model, we identified microglial subsets—distinct from previously reported “disease-associated microglia”—expressing interferon-related or proliferation modules. We then analyzed whole-tissue RNA profiles from human neurodegenerative diseases, including a new AD dataset. Correcting for altered cellular composition of AD tissue, we observed elevated expression of the neurodegeneration-related modules, but also modules not implicated using expression profiles from mouse models alone. We provide a searchable, interactive database for exploring gene expression in all these datasets (http://research-pub.gene.com/BrainMyeloidLandscape. Understanding the dimensions of CNS myeloid cell activation in human disease may reveal opportunities for therapeutic intervention.
Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

DEFF Research Database (Denmark)

Jensen, Helle

frequently express Hsp70 on their cell surface, whereas the corresponding normal tissues do not. In addition, several clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 cell surface expression on cancer...
Regulation of CD93 cell surface expression by protein kinase C isoenzymes.

Science.gov (United States)

Ikewaki, Nobunao; Kulski, Jerzy K; Inoko, Hidetoshi

2006-01-01

Human CD93, also known as complement protein 1, q subcomponent, receptor (C1qRp), is selectively expressed by cells with a myeloid lineage, endothelial cells, platelets, and microglia and was originally reported to be involved in the complement protein 1, q subcomponent (C1q)-mediated enhancement of phagocytosis. The intracellular molecular events responsible for the regulation of its expression on the cell surface, however, have not been determined. In this study, the effect of protein kinases in the regulation of CD93 expression on the cell surface of a human monocyte-like cell line (U937), a human NK-like cell line (KHYG-1), and a human umbilical vein endothelial cell line (HUV-EC-C) was investigated using four types of protein kinase inhibitors, the classical protein kinase C (cPKC) inhibitor Go6976, the novel PKC (nPKC) inhibitor Rottlerin, the protein kinase A (PKA) inhibitor H-89 and the protein tyrosine kinase (PTK) inhibitor herbimycin A at their optimum concentrations for 24 hr. CD93 expression was analyzed using flow cytometry and glutaraldehyde-fixed cellular enzyme-linked immunoassay (EIA) techniques utilizing a CD93 monoclonal antibody (mAb), mNI-11, that was originally established in our laboratory as a CD93 detection probe. The nPKC inhibitor Rottlerin strongly down-regulated CD93 expression on the U937 cells in a dose-dependent manner, whereas the other inhibitors had little or no effect. CD93 expression was down-regulated by Go6976, but not by Rottlerin, in the KHYG-1 cells and by both Rottlerin and Go6976 in the HUV-EC-C cells. The PKC stimulator, phorbol myristate acetate (PMA), strongly up-regulated CD93 expression on the cell surface of all three cell-lines and induced interleukin-8 (IL-8) production by the U937 cells and interferon-gamma (IFN-gamma) production by the KHYG-1 cells. In addition, both Go6976 and Rottlerin inhibited the up-regulation of CD93 expression induced by PMA and IL-8 or IFN-gamma production in the respective cell
Human decidual macrophages and NK cells differentially express Toll-like receptors and display distinct cytokine profiles upon TLR stimulation.

Directory of Open Access Journals (Sweden)

Marion eDuriez

2014-07-01

Full Text Available Maternofetal pathogen transmission is partially controlled at the level of the maternal uterine mucosa at the fetal implantation site (the decidua basalis, where maternal and fetal cells are in close contact. Toll-like receptors (TLRs may play an important role in initiating rapid immune responses against pathogens in the decidua basalis, however the tolerant microenvironment should be preserved in order to allow fetal development. Here we investigated the expression and functionality of TLRs expressed by decidual macrophages (dMs and NK cells (dNKs, the major decidual immune cell populations.We report for the first time that both human dMs and dNK cells express mRNAs encoding TLRs 1-9, albeit with a higher expression level in dMs. TLR2, TLR3 and TLR4 protein expression checked by flow cytometry was positive for both dMs and dNK cells. In vitro treatment of primary dMs and dNK cells with specific TLR2, TLR3, TLR4, TLR7/8 and TLR9 agonists enhanced their secretion of pro- and anti-inflammatory cytokines, as well as cytokines and chemokines involved in immune cell crosstalk. Only dNK cells released IFN-γ, whereas only dMs released IL-1β, IL-10 and IL-12. TLR9 activation of dMs resulted in a distinct pattern of cytokine expression compared to the other TLRs. The cytokine profiles expressed by dMs and dNK cells upon TLR activation are compatible with maintenance of the fetotolerant immune environment during initiation of immune responses to pathogens at the maternofetal interface.
Surface expression, single-channel analysis and membrane topology of recombinant Chlamydia trachomatis Major Outer Membrane Protein

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McClafferty Heather

2005-01-01

Full Text Available Abstract Background Chlamydial bacteria are obligate intracellular pathogens containing a cysteine-rich porin (Major Outer Membrane Protein, MOMP with important structural and, in many species, immunity-related roles. MOMP forms extensive disulphide bonds with other chlamydial proteins, and is difficult to purify. Leaderless, recombinant MOMPs expressed in E. coli have yet to be refolded from inclusion bodies, and although leadered MOMP can be expressed in E. coli cells, it often misfolds and aggregates. We aimed to improve the surface expression of correctly folded MOMP to investigate the membrane topology of the protein, and provide a system to display native and modified MOMP epitopes. Results C. trachomatis MOMP was expressed on the surface of E. coli cells (including "porin knockout" cells after optimizing leader sequence, temperature and medium composition, and the protein was functionally reconstituted at the single-channel level to confirm it was folded correctly. Recombinant MOMP formed oligomers even in the absence of its 9 cysteine residues, and the unmodified protein also formed inter- and intra-subunit disulphide bonds. Its topology was modeled as a (16-stranded β-barrel, and specific structural predictions were tested by removing each of the four putative surface-exposed loops corresponding to highly immunogenic variable sequence (VS domains, and one or two of the putative transmembrane strands. The deletion of predicted external loops did not prevent folding and incorporation of MOMP into the E. coli outer membrane, in contrast to the removal of predicted transmembrane strands. Conclusions C. trachomatis MOMP was functionally expressed on the surface of E. coli cells under newly optimized conditions. Tests of its predicted membrane topology were consistent with β-barrel oligomers in which major immunogenic regions are displayed on surface-exposed loops. Functional surface expression, coupled with improved understanding of MOMP
The Effect of Laminin-1-Doped Nanoroughened Implant Surfaces: Gene Expression and Morphological Evaluation

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Humberto Osvaldo Schwartz-Filho

2012-01-01

Full Text Available Aim. This study aimed to observe the morphological and molecular effect of laminin-1 doping to nanostructured implant surfaces in a rabbit model. Materials and Methods. Nanostructured implants were coated with laminin-1 (test; dilution, 100 μg/mL and inserted into the rabbit tibiae. Noncoated implants were used as controls. After 2 weeks of healing, the implants were removed and subjected to morphological analysis using scanning electron microscopy (SEM and gene expression analysis using the real-time reverse transcriptase-polymerase chain reaction (RT-PCR. Results. SEM revealed bony tissue attachment for both control and test implants. Real-time RT-PCR analysis showed that the expression of osteoblast markers RUNX-2, osteocalcin, alkaline phosphatase, and collagen I was higher (1.62-fold, 1.53-fold, 1.97-fold, and 1.04-fold, resp. for the implants modified by laminin-1 relative to the control. All osteoclast markers investigated in the study presented higher expression on the test implants than controls as follows: tartrate-resistant acid phosphatase (1.67-fold, calcitonin receptor (1.35-fold, and ATPase (1.25-fold. The test implants demonstrated higher expression of inflammatory markers interleukin-10 (1.53-fold and tumour necrosis factor-α (1.61-fold relative to controls. Conclusion. The protein-doped surface showed higher gene expression of typical genes involved in the osseointegration cascade than the control surface.
Testis-expressed profilins 3 and 4 show distinct functional characteristics and localize in the acroplaxome-manchette complex in spermatids

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Rothkegel Martin

2009-05-01

Full Text Available Abstract Background Multiple profilin isoforms exist in mammals; at least four are expressed in the mammalian testis. The testis-specific isoforms profilin-3 (PFN3 and profilin-4 (PFN4 may have specialized roles in spermatogenic cells which are distinct from known functions fulfilled by the "somatic" profilins, profilin-1 (PFN1 and profilin-2 (PFN2. Results Ligand interactions and spatial distributions of PFN3 and PFN4 were compared by biochemical, molecular and immunological methods; PFN1 and PFN2 were employed as controls. β-actin, phosphoinositides, poly-L-proline and mDia3, but not VASP, were confirmed as in vitro interaction partners of PFN3. In parallel experiments, PFN4 bound to selected phosphoinositides but not to poly-L-proline, proline-rich proteins, or actin. Immunofluorescence microscopy of PFN3 and PFN4 revealed distinct subcellular locations in differentiating spermatids. Both were associated first with the acroplaxome and later with the transient manchette. Predicted 3D structures indicated that PFN3 has the actin-binding site conserved, but retains only approximately half of the common poly-L-proline binding site. PFN4, in comparison, has lost both, polyproline and actin binding sites completely, which is well in line with the experimental data. Conclusion The testis-specific isoform PFN3 showed major hallmarks of the well characterized "somatic" profilin isoforms, albeit with distinct binding affinities. PFN4, on the other hand, did not interact with actin or polyproline in vitro. Rather, it seemed to be specialized for phospholipid binding, possibly providing cellular functions which are distinct from actin dynamics regulation.
Profiling of wheat class III peroxidase genes derived from powdery mildew-attacked epidermis reveals distinct sequence-associated expression patterns.

Science.gov (United States)

Liu, Guosheng; Sheng, Xiaoyan; Greenshields, David L; Ogieglo, Adam; Kaminskyj, Susan; Selvaraj, Gopalan; Wei, Yangdou

2005-07-01

A cDNA library was constructed from leaf epidermis of diploid wheat (Triticum monococcum) infected with the powdery mildew fungus (Blumeria graminis f. sp. tritici) and was screened for genes encoding peroxidases. From 2,500 expressed sequence tags (ESTs), 36 cDNAs representing 10 peroxidase genes (designated TmPRX1 to TmPRX10) were isolated and further characterized. Alignment of the deduced amino acid sequences and phylogenetic clustering with peroxidases from other plant species demonstrated that these peroxidases fall into four distinct groups. Differential expression and tissue-specific localization among the members were observed during the B. graminis f. sp. tritici attack using Northern blots and reverse-transcriptase polymerase chain reaction analyses. Consistent with its abundance in the EST collection, TmPRX1 expression showed the highest induction during pathogen attack and fluctuated in response to the fungal parasitic stages. TmPRX1 to TmPRX6 were expressed predominantly in mesophyll cells, whereas TmPRX7 to TmPRX10, which feature a putative C-terminal propeptide, were detectable mainly in epidermal cells. Using TmPRX8 as a representative, we demonstrated that its C-terminal propeptide was sufficient to target a green fluorescent protein fusion protein to the vacuoles in onion cells. Finally, differential expression profiles of the TmPRXs after abiotic stresses and signal molecule treatments were used to dissect the potential role of these peroxidases in multiple stress and defense pathways.
Integrative genome-wide expression profiling identifies three distinct molecular subgroups of renal cell carcinoma with different patient outcome

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Beleut Manfred

2012-07-01

Full Text Available Abstract Background Renal cell carcinoma (RCC is characterized by a number of diverse molecular aberrations that differ among individuals. Recent approaches to molecularly classify RCC were based on clinical, pathological as well as on single molecular parameters. As a consequence, gene expression patterns reflecting the sum of genetic aberrations in individual tumors may not have been recognized. In an attempt to uncover such molecular features in RCC, we used a novel, unbiased and integrative approach. Methods We integrated gene expression data from 97 primary RCC of different pathologic parameters, 15 RCC metastases as well as 34 cancer cell lines for two-way nonsupervised hierarchical clustering using gene groups suggested by the PANTHER Classification System. We depicted the genomic landscape of the resulted tumor groups by means of Single Nuclear Polymorphism (SNP technology. Finally, the achieved results were immunohistochemically analyzed using a tissue microarray (TMA composed of 254 RCC. Results We found robust, genome wide expression signatures, which split RCC into three distinct molecular subgroups. These groups remained stable even if randomly selected gene sets were clustered. Notably, the pattern obtained from RCC cell lines was clearly distinguishable from that of primary tumors. SNP array analysis demonstrated differing frequencies of chromosomal copy number alterations among RCC subgroups. TMA analysis with group-specific markers showed a prognostic significance of the different groups. Conclusion We propose the existence of characteristic and histologically independent genome-wide expression outputs in RCC with potential biological and clinical relevance.
cDNA cloning and expression analysis of two distinct Sox8 genes in Paramisgurnus dabryanus (Cypriniformes).

Science.gov (United States)

Xia, Xiaohua; Zhao, Jie; Du, Qiyan; Chang, Zhongjie

2010-08-01

The Sox9 gene attracts a lot of attention because of its connection with gonadal development and differentiation. However, Sox8, belonging to the same subgroup SoxE, has rarely been studied. To investigate the function as well as the evolutionary origin of SOXE subgroup, we amplified the genomic DNA of Paramisgurnus dabryanu using a pair of degenerate primers. Using rapid amplification of the cDNA ends (RACE), it was discovered that P. dabryanu has two duplicates: Sox8a and Sox8b. Each has an intron of different length in the conserved HMG-box region. The overall sequence similarity of the deduced amino acid of PdSox8a and PdSox8b was 46.26%, and only two amino acids changed in the HMG-box. This is the first evidence showing that there are two distinct duplications of Sox8 genes in Cypriniformes. Southern blot analysis showed only one hybrid band, with lengths 7.4 or 9.2 kb. Both semi-quantitative RT-PCR and real-time quantitative PCR assay displayed that both PdSox8a and PdSox8b are downregulated during early embryonic development. In adult tissues, the two Sox8 genes expressed ubiquitously, and expression levels are particularly high in the gonads and brain. In gonads, both PdSox8a and PdSox8b are expressed at a higher level in the tesis than in the ovary. PdSox8a and PdSox8b may have functional overlaps and are essential for the neuronal development and differentiation of gonads.
Population heterogeneity in the surface expression of Ulex europaeus I-lectin (UEA I)-binding sites in cultured malignant and transformed cells

Energy Technology Data Exchange (ETDEWEB)

Virtanen, I.; Lehtonen, E.; Naervaenen, O.; Leivo, I.; Lehto, V.P.

1985-11-01

We studied the binding of fluorochrome-coupled Ulex europaeus I-lectin (UEA-I) to cultured malignant cells: all human malignant and transformed cells and also mouse teratocarcinoma cells examined gave a homogeneous cell membrane-type of surface staining only in some of the cells. Such a population heterogeneity appeared to be independent of the cell cycle. Instead, other lectin conjugates used bound homogeneously to all cell. In permeabilized cells, a juxtanuclear reticular staining of the Golgi apparatus was seen in the UEA-I-positive cells. No staining of the pericellular matrix components, produced by malignant cells grown in serum-free culture medium, could be obtained with TRITC-UEA-I. UEA-I-lectin recognized most polypeptides from A8387 fibrosarcoma cells and HeLa cells, metabolically labelled with (/sup 3/H)fucose. Furthermore, surface labelling of these cells with the neuraminidase-galactose oxidase/sodium borohydride method disclosed that both UEA-I and Ricinus communis agglutinin I revealed the same major surface glycoproteins. Results with metabolically labelled cells showed, in addition, that UEA-I-lectin did not bind to secreted glycoproteins produced by A8387 cells and recognized by other lectins. The results indicate that transformed and malignant cells show a distinct population heterogeneity in their expression of some cell surface-associated fucosyl glycoconjugates. The results also suggest that malignant cells can glycosylate their membrane and secreted glycoproteins in a different manner.
Intermediate monocytes in ANCA vasculitis: increased surface expression of ANCA autoantigens and IL-1β secretion in response to anti-MPO antibodies.

LENUS (Irish Health Repository)

O'Brien, Eóin C

2015-01-01

ANCA vasculitis encompasses several autoimmune conditions characterised by destruction of small vessels, inflammation of the respiratory tract and glomerulonephritis. Most patients harbour autoantibodies to myeloperoxidase (MPO) or proteinase 3 (PR3). Clinical and experimental data suggest that pathogenesis is driven by ANCA-mediated activation of neutrophils and monocytes. We investigated a potential role for distinct monocyte subsets. We found that the relative proportion of intermediate monocytes is increased in patients versus control individuals, and both MPO and PR3 are preferentially expressed on these cells. We demonstrate that MPO and PR3 are expressed independently of each other on monocytes and that PR3 is not associated with CD177. MPO expression correlates with that of Fc receptor CD16 on intermediate monocytes. Monocyte subsets respond differently to antibodies directed against MPO and PR3, with anti-MPO but not anti-PR3 leading to increased IL-1β, IL-6 and IL-8 production. In concordance with the observed higher surface expression of MPO on intermediate monocytes, this subset produces the highest quantity of IL-1β in response to anti-MPO stimulation. These data suggest that monocytes, specifically, the intermediate subset, may play a role in ANCA vasculitis, and also indicate that substantial differences exist between the effect of anti-MPO and anti-PR3 antibodies on these cells.
Distinctive properties and expression profiles of glutamine synthetase from a plant symbiotic fungus.

Science.gov (United States)

Montanini, Barbara; Betti, Marco; Márquez, Antonio J; Balestrini, Raffaella; Bonfante, Paola; Ottonello, Simone

2003-01-01

The nucleotide sequences reported in this paper have been submitted to the GenBank(R)/EBI Nucleotide Sequence Databases with accession numbers AF462037 (glutamine synthetase) and AF462032 (glutamate synthase). Nitrogen retrieval and assimilation by symbiotic ectomycorrhizal fungi is thought to play a central role in the mutualistic interaction between these organisms and their plant hosts. Here we report on the molecular characterization of the key N-assimilation enzyme glutamine synthetase from the mycorrhizal ascomycete Tuber borchii (TbGS). TbGS displayed a strong positive co-operativity ( n =1.7+/-0.29) and an unusually high S(0.5) value (54+/-16 mM; S(0.5) is the substrate concentration value at which v =(1/2) V (max)) for glutamate, and a correspondingly low sensitivity towards inhibition by the glutamate analogue herbicide phosphinothricin. The TbGS mRNA, which is encoded by a single-copy gene in the Tuber genome, was up-regulated in N-starved mycelia and returned to basal levels upon resupplementation of various forms of N, the most effective of which was nitrate. Both responses were accompanied by parallel variations of TbGS protein amount and glutamine synthetase activity, thus indicating that TbGS levels are primarily controlled at the pre-translational level. As revealed by a comparative analysis of the TbGS mRNA and of the mRNAs for the metabolically related enzymes glutamate dehydrogenase and glutamate synthase, TbGS is not only the sole messenger that positively responds to N starvation, but also the most abundant under N-limiting conditions. A similar, but even more discriminating expression pattern, with practically undetectable glutamate dehydrogenase mRNA levels, was observed in fruitbodies. The TbGS mRNA was also found to be expressed in symbiosis-engaged hyphae, with distinctively higher hybridization signals in hyphae that were penetrating among and within root cells. PMID:12683951
Lhx2 expression promotes self-renewal of a distinct multipotential hematopoietic progenitor cell in embryonic stem cell-derived embryoid bodies.

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Lina Dahl

Full Text Available The molecular mechanisms regulating the expansion of the hematopoietic system including hematopoietic stem cells (HSCs in the fetal liver during embryonic development are largely unknown. The LIM-homeobox gene Lhx2 is a candidate regulator of fetal hematopoiesis since it is expressed in the fetal liver and Lhx2(-/- mice die in utero due to severe anemia. Moreover, expression of Lhx2 in embryonic stem (ES cell-derived embryoid bodies (EBs can lead to the generation of HSC-like cell lines. To further define the role of this transcription factor in hematopoietic regulation, we generated ES cell lines that enabled tet-inducible expression of Lhx2. Using this approach we observed that Lhx2 expression synergises with specific signalling pathways, resulting in increased frequency of colony forming cells in developing EB cells. The increase in growth factor-responsive progenitor cells directly correlates to the efficiency in generating HSC-like cell lines, suggesting that Lhx2 expression induce self-renewal of a distinct multipotential hematopoietic progenitor cell in EBs. Signalling via the c-kit tyrosine kinase receptor and the gp130 signal transducer by IL-6 is necessary and sufficient for the Lhx2 induced self-renewal. While inducing self-renewal of multipotential progenitor cells, expression of Lhx2 inhibited proliferation of primitive erythroid precursor cells and interfered with early ES cell commitment, indicating striking lineage specificity of this effect.
Large-scale expression analysis reveals distinct microRNA profiles at different stages of human neurodevelopment.

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Brandon Smith

Full Text Available BACKGROUND: MicroRNAs (miRNAs are short non-coding RNAs predicted to regulate one third of protein coding genes via mRNA targeting. In conjunction with key transcription factors, such as the repressor REST (RE1 silencing transcription factor, miRNAs play crucial roles in neurogenesis, which requires a highly orchestrated program of gene expression to ensure the appropriate development and function of diverse neural cell types. Whilst previous studies have highlighted select groups of miRNAs during neural development, there remains a need for amenable models in which miRNA expression and function can be analyzed over the duration of neurogenesis. PRINCIPAL FINDINGS: We performed large-scale expression profiling of miRNAs in human NTera2/D1 (NT2 cells during retinoic acid (RA-induced transition from progenitors to fully differentiated neural phenotypes. Our results revealed dynamic changes of miRNA patterns, resulting in distinct miRNA subsets that could be linked to specific neurodevelopmental stages. Moreover, the cell-type specific miRNA subsets were very similar in NT2-derived differentiated cells and human primary neurons and astrocytes. Further analysis identified miRNAs as putative regulators of REST, as well as candidate miRNAs targeted by REST. Finally, we confirmed the existence of two predicted miRNAs; pred-MIR191 and pred-MIR222 associated with SLAIN1 and FOXP2, respectively, and provided some evidence of their potential co-regulation. CONCLUSIONS: In the present study, we demonstrate that regulation of miRNAs occurs in precise patterns indicative of their roles in cell fate commitment, progenitor expansion and differentiation into neurons and glia. Furthermore, the similarity between our NT2 system and primary human cells suggests their roles in molecular pathways critical for human in vivo neurogenesis.
SiglecF+Gr1hi eosinophils are a distinct subpopulation within the lungs of allergen-challenged mice.

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Percopo, Caroline M; Brenner, Todd A; Ma, Michelle; Kraemer, Laura S; Hakeem, Reem M A; Lee, James J; Rosenberg, Helene F

2017-01-01

Although eosinophils as a group are readily identified by their unique morphology and staining properties, flow cytometry provides an important means for identification of subgroups based on differential expression of distinct surface Ags. Here, we characterize an eosinophil subpopulation defined by high levels of expression of the neutrophil Ag Gr1 (CD45 + CD11c - SiglecF + Gr1 hi ). SiglecF + Gr1 hi eosinophils, distinct from the canonical SiglecF + Gr1 - eosinophil population, were detected in allergen-challenged wild-type and granule protein-deficient (EPX -/- and MBP-1 -/- ) mice, but not in the eosinophil-deficient ΔdblGATA strain. In contrast to Gr1 + neutrophils, which express both cross-reacting Ags Ly6C and Ly6G, SiglecF + Gr1 hi eosinophils from allergen-challenged lung tissue are uniquely Ly6G + Although indistinguishable from the more-numerous SiglecF + Gr1 - eosinophils under light microscopy, FACS-isolated populations revealed prominent differences in cytokine contents. The lymphocyte-targeting cytokines CXCL13 and IL-27 were identified only in the SiglecF + Gr1 hi eosinophil population (at 3.9 and 4.8 pg/10 6 cells, respectively), as was the prominent proinflammatory mediator IL-13 (72 pg/10 6 cells). Interestingly, bone marrow-derived (SiglecF + ), cultured eosinophils include a more substantial Gr1 + subpopulation (∼50%); Gr1 + bmEos includes primarily a single Ly6C + and a smaller, double-positive (Ly6C + Ly6G + ) population. Taken together, our findings characterize a distinct SiglecF + Gr1 hi eosinophil subset in lungs of allergen-challenged, wild-type and granule protein-deficient mice. SiglecF + Gr1 hi eosinophils from wild-type mice maintain a distinct subset of cytokines, including those active on B and T lymphocytes. These cytokines may facilitate eosinophil-mediated immunomodulatory responses in the allergen-challenged lung as well as in other distinct microenvironments. © Society for Leukocyte Biology.
Mars Express radar collects first surface data

Science.gov (United States)

2005-08-01

the middle of August, when the night-time portion of the observations will have almost ended. After that, observation priority will be given to other Mars Express instruments that are best suited to operating in daytime, such as the HRSC camera and Omega mapping spectrometer. However, Marsis will continue its surface and ionospheric investigations in daytime, with ionospheric sounding being reserved for more than 20% of all Mars Express orbits, under all possible Sun illumination conditions. In December, the Mars Express orbit pericentre will enter night-time again. By then, the pericentre will have moved closer to the south pole, allowing Marsis to carry out optimal probing of the subsurface once again, this time in the southern hemisphere. Note to editors The first commissioning phase was given over to testing the Marsis electronics and software and the two 20m-long antennas (dipole). The second commissioning phase, lasting about ten days, will be spent calibrating the 7m ‘monopole’ antenna. This antenna is to be used in conjunction with the Marsis dipole to correct any surface roughness effects caused by the radio waves emitted by the dipole and reflected by an irregular surface. The monopole will find its best use during investigations of areas where surface roughness is greater. The Marsis instrument was developed within the framework of a Memorandum of Understanding between the Italian Space Agency (ASI) and NASA. It was developed by Alenia Spazio under ASI management and the scientific supervision of University of Rome ‘La Sapienza’, in partnership with the Jet Propulsion Laboratory (JPL) and the University of Iowa. JPL provided the antenna manufactured by Astro Aerospace. It is the first instrument designed to actually look below the surface of Mars. Its major goals are to characterise the subsurface layers of sediments and possibly detect underground water or ice, conduct large-scale altimetry mapping and provide data on the planet’s ionosphere. For
The surface diffusion coefficient for an arbitrarily curved fluid-fluid interface. (I). General expression

Science.gov (United States)

M. C. Sagis, Leonard

2001-03-01

In this paper, we develop a theory for the calculation of the surface diffusion coefficient for an arbitrarily curved fluid-fluid interface. The theory is valid for systems in hydrodynamic equilibrium, with zero mass-averaged velocities in the bulk and interfacial regions. We restrict our attention to systems with isotropic bulk phases, and an interfacial region that is isotropic in the plane parallel to the dividing surface. The dividing surface is assumed to be a simple interface, without memory effects or yield stresses. We derive an expression for the surface diffusion coefficient in terms of two parameters of the interfacial region: the coefficient for plane-parallel diffusion D (AB)aa(ξ) , and the driving force d(B)I||(ξ) . This driving force is the parallel component of the driving force for diffusion in the interfacial region. We derive an expression for this driving force using the entropy balance.
Tamoxifen-inducible gene deletion reveals a distinct cell type associated with trabecular bone, and direct regulation of PTHrP expression and chondrocyte morphology by Ihh in growth region cartilage.

Science.gov (United States)

Hilton, Matthew J; Tu, Xiaolin; Long, Fanxin

2007-08-01

Indian hedgehog (Ihh) controls multiple aspects of endochondral skeletal development by signaling to both chondrocytes and perichondrial cells. Previous efforts to delineate direct effects of Ihh on chondrocytes by Col2-Cre-mediated ablation of Smoothened (Smo, encoding a transmembrane protein indispensable for Ihh signaling) has been only partially successful, due to the inability to discriminate between chondrocytes and perichondrial cells. Here we report a transgenic line (Col2-Cre) expressing under the control of the Colalpha1(II) promoter an inert form of Cre that is activatable by exogenous tamoxifen (TM); TM administration at proper times during embryogenesis induced Cre activity in chondrocytes but not in the perichondrium. By using this mouse line, we deleted Smo within subsets of chondrocytes without affecting the perichondrium and found that Smo removal led to localized disruption of the expression of parathyroid hormone-related protein (PTHrP) and the morphology of chondrocytes. Unexpectedly, TM invariably induced Cre activity in a subset of cells associated with the trabecular bone surface of long bones. These cells, when genetically marked and cultured in vitro, were capable of producing bone nodules. Expression of the Col2-Cre transgene in these cells likely reflected the endogenous Colalpha1(II) promoter activity as similar cells were found to express the IIA isoform of Colalpha1(II) mRNA endogenously. In summary, the present study has not only provided evidence that Ihh signaling directly controls PTHrP expression and chondrocyte morphology in the growth region cartilage, but has also uncovered a distinct cell type associated with the trabecular bone that appears to possess osteogenic potential.

Influence of HLA-C Expression Level on HIV Control

Science.gov (United States)

Apps, Richard; Qi, Ying; Carlson, Jonathan M.; Chen, Haoyan; Gao, Xiaojiang; Thomas, Rasmi; Yuki, Yuko; Del Prete, Greg Q.; Goulder, Philip; Brumme, Zabrina L.; Brumme, Chanson J.; John, Mina; Mallal, Simon; Nelson, George; Bosch, Ronald; Heckerman, David; Stein, Judy L.; Soderberg, Kelly A.; Moody, M. Anthony; Denny, Thomas N.; Zeng, Xue; Fang, Jingyuan; Moffett, Ashley; Lifson, Jeffrey D.; Goedert, James J.; Buchbinder, Susan; Kirk, Gregory D.; Fellay, Jacques; McLaren, Paul; Deeks, Steven G.; Pereyra, Florencia; Walker, Bruce; Michael, Nelson L.; Weintrob, Amy; Wolinsky, Steven; Liao, Wilson; Carrington, Mary

2013-01-01

A variant upstream of human leukocyte antigen C (HLA-C) shows the most significant genome-wide effect on HIV control in European Americans and is also associated with the level of HLA-C expression. We characterized the differential cell surface expression levels of all common HLA-C allotypes and tested directly for effects of HLA-C expression on outcomes of HIV infection in 5243 individuals. Increasing HLA-C expression was associated with protection against multiple outcomes independently of individual HLA allelic effects in both African and European Americans, regardless of their distinct HLA-C frequencies and linkage relationships with HLA-B and HLA-A. Higher HLA-C expression was correlated with increased likelihood of cytotoxic T lymphocyte responses and frequency of viral escape mutation. In contrast, high HLA-C expression had a deleterious effect in Crohn’s disease, suggesting a broader influence of HLA expression levels in human disease. PMID:23559252
Hsc70 regulates cell surface ASIC2 expression and vascular smooth muscle cell migration.

Science.gov (United States)

Grifoni, Samira C; McKey, Susan E; Drummond, Heather A

2008-05-01

Recent studies suggest members of the degenerin (DEG)/epithelial Na(+) channel (ENaC)/acid-sensing ion channel (ASIC) protein family play an important role in vascular smooth muscle cell (VSMC) migration. In a previous investigation, we found suppression of a certain DEG/ENaC/ASIC member, ASIC2, increased VSMC chemotactic migration, raising the possibility that ASIC2 may play an inhibitory role. Because ASIC2 protein was retained in the cytoplasm, we reasoned increasing surface expression of ASIC2 might unmask the inhibitory role of ASIC2 in VSMC migration so we could test the hypothesis that ASIC2 inhibits VSMC migration. Therefore, we used the chemical chaperone glycerol to enhance ASIC2 expression. Glycerol 1) increased cytoplasm ASIC2 expression, 2) permitted detection of ASIC2 at the cell surface, and 3) inhibited platelet-derived growth factor (PDGF)-bb mediated VSMC migration. Furthermore, ASIC2 silencing completely abolished the inhibitory effect of glycerol on migration, suggesting upregulation of ASIC2 is responsible for glycerol-induced inhibition of VSMC migration. Because other investigators have shown that glycerol regulates ENaC/ASIC via interactions with a certain heat shock protein, heat shock protein 70 (Hsc70), we wanted to determine the importance of Hsc70 on ASIC2 expression in VSMCs. We found that Hsc70 silencing increases ASIC2 cell surface expression and inhibits VSMC migration, which is abolished by cosilencing ASIC2. These data demonstrate that Hsc70 inhibits ASIC2 expression, and, when the inhibitory effect of Hsc70 is removed, ASIC2 expression increases, resulting in reduced VSMC migration. Because VSMC migration contributes to vasculogenesis and remodeling following vascular injury, our findings raise the possibility that ASIC2-Hsc70 interactions may play a role in these processes.
Homo sapiens exhibit a distinct pattern of CNV genes regulation: an important role of miRNAs and SNPs in expression plasticity.

Science.gov (United States)

Dweep, Harsh; Kubikova, Nada; Gretz, Norbert; Voskarides, Konstantinos; Felekkis, Kyriacos

2015-07-16

Gene expression regulation is a complex and highly organized process involving a variety of genomic factors. It is widely accepted that differences in gene expression can contribute to the phenotypic variability between species, and that their interpretation can aid in the understanding of the physiologic variability. CNVs and miRNAs are two major players in the regulation of expression plasticity and may be responsible for the unique phenotypic characteristics observed in different lineages. We have previously demonstrated that a close interaction between these two genomic elements may have contributed to the regulation of gene expression during evolution. This work presents the molecular interactions between CNV and non CNV genes with miRNAs and other genomic elements in eight different species. A comprehensive analysis of these interactions indicates a unique nature of human CNV genes regulation as compared to other species. By using genes with short 3' UTR that abolish the "canonical" miRNA-dependent regulation, as a model, we demonstrate a distinct and tight regulation of human genes that might explain some of the unique features of human physiology. In addition, comparison of gene expression regulation between species indicated that there is a significant difference between humans and mice possibly questioning the effectiveness of the latest as experimental models of human diseases.
Distinct projection targets define subpopulations of mouse brainstem vagal neurons that express the autism-associated MET receptor tyrosine kinase.

Science.gov (United States)

Kamitakahara, Anna; Wu, Hsiao-Huei; Levitt, Pat

2017-12-15

Detailed anatomical tracing and mapping of the viscerotopic organization of the vagal motor nuclei has provided insight into autonomic function in health and disease. To further define specific cellular identities, we paired information based on visceral connectivity with a cell-type specific marker of a subpopulation of neurons in the dorsal motor nucleus of the vagus (DMV) and nucleus ambiguus (nAmb) that express the autism-associated MET receptor tyrosine kinase. As gastrointestinal disturbances are common in children with autism spectrum disorder (ASD), we sought to define the relationship between MET-expressing (MET+) neurons in the DMV and nAmb, and the gastrointestinal tract. Using wholemount tissue staining and clearing, or retrograde tracing in a MET EGFP transgenic mouse, we identify three novel subpopulations of EGFP+ vagal brainstem neurons: (a) EGFP+ neurons in the nAmb projecting to the esophagus or laryngeal muscles, (b) EGFP+ neurons in the medial DMV projecting to the stomach, and (b) EGFP+ neurons in the lateral DMV projecting to the cecum and/or proximal colon. Expression of the MET ligand, hepatocyte growth factor (HGF), by tissues innervated by vagal motor neurons during fetal development reveal potential sites of HGF-MET interaction. Furthermore, similar cellular expression patterns of MET in the brainstem of both the mouse and nonhuman primate suggests that MET expression at these sites is evolutionarily conserved. Together, the data suggest that MET+ neurons in the brainstem vagal motor nuclei are anatomically positioned to regulate distinct portions of the gastrointestinal tract, with implications for the pathophysiology of gastrointestinal comorbidities of ASD. © 2017 Wiley Periodicals, Inc.
Designed Surface Topographies Control ICAM-1 Expression in Tonsil-Derived Human Stromal Cells

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Aliaksei S. Vasilevich

2018-06-01

Full Text Available Fibroblastic reticular cells (FRCs, the T-cell zone stromal cell subtype in the lymph nodes, create a scaffold for adhesion and migration of immune cells, thus allowing them to communicate. Although known to be important for the initiation of immune responses, studies about FRCs and their interactions have been impeded because FRCs are limited in availability and lose their function upon culture expansion. To circumvent these limitations, stromal cell precursors can be mechanotranduced to form mature FRCs. Here, we used a library of designed surface topographies to trigger FRC differentiation from tonsil-derived stromal cells (TSCs. Undifferentiated TSCs were seeded on a TopoChip containing 2176 different topographies in culture medium without differentiation factors, then monitored cell morphology and the levels of ICAM-1, a marker of FRC differentiation. We identified 112 and 72 surfaces that upregulated and downregulated, respectively, ICAM-1 expression. By monitoring cell morphology, and expression of the FRC differentiation marker ICAM-1 via image analysis and machine learning, we discovered correlations between ICAM-1 expression, cell shape and design of surface topographies and confirmed our findings by using flow cytometry. Our findings confirmed that TSCs are mechano-responsive cells and identified particular topographies that can be used to improve FRC differentiation protocols.
Differential Expression of Osteo-Modulatory Molecules in Periodontal Ligament Stem Cells in Response to Modified Titanium Surfaces

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So Yeon Kim

2014-01-01

Full Text Available This study assessed differential gene expression of signaling molecules involved in osteogenic differentiation of periodontal ligament stem cells (PDLSCs subjected to different titanium (Ti surface types. PDLSCs were cultured on tissue culture polystyrene (TCPS, and four types of Ti discs (PT, SLA, hydrophilic PT (pmodPT, and hydrophilic SLA (modSLA with no osteoinductive factor and then osteogenic activity, including alkaline phosphatase (ALP activity, mRNA expression of runt-related gene 2, osterix, FOSB, FRA1, and protein levels of osteopontin and collagen type IA, were examined. The highest osteogenic activity appeared in PDLSCs cultured on SLA, compared with the TCPS and other Ti surfaces. The role of surface properties in affecting signaling molecules to modulate PDLSC behavior was determined by examining the regulation of Wnt pathways. mRNA expression of the canonical Wnt signaling molecules, Wnt3a and β-catenin, was higher on SLA and modSLA than on smooth surfaces, but gene expression of the calcium-dependent Wnt signaling molecules Wnt5a, calmodulin, and NFATc1 was increased significantly on PT and pmodPT. Moreover, integrin α2/β1, sonic hedgehog, and Notch signaling molecules were affected differently by each surface modification. In conclusion, surface roughness and hydrophilicity can affect differential Wnt pathways and signaling molecules, targeting the osteogenic differentiation of PDLSCs.
Kv7.1 surface expression is regulated by epithelial cell polarization

DEFF Research Database (Denmark)

Andersen, Martin N; Olesen, Søren-Peter; Rasmussen, Hanne Borger

2011-01-01

The potassium channel K(V)7.1 is expressed in the heart where it contributes to the repolarization of the cardiac action potential. In addition, K(V)7.1 is expressed in epithelial tissues where it plays a role in salt and water transport. Mutations in the kcnq1 gene can lead to long QT syndrome...... and deafness, and several mutations have been described as trafficking mutations. To learn more about the basic mechanisms that regulate K(V)7.1 surface expression, we have investigated the trafficking of K(V)7.1 during the polarization process of the epithelial cell line Madin-Darby Canine Kidney (MDCK) using...... is regulated by signaling mechanisms involved in epithelial cell polarization in particular signaling cascades involving protein kinase C and PI3K....
Surface modification of nanoparticles enables selective evasion of phagocytic clearance by distinct macrophage phenotypes

Science.gov (United States)

Qie, Yaqing; Yuan, Hengfeng; von Roemeling, Christina A.; Chen, Yuanxin; Liu, Xiujie; Shih, Kevin D.; Knight, Joshua A.; Tun, Han W.; Wharen, Robert E.; Jiang, Wen; Kim, Betty Y. S.

2016-05-01

Nanomedicine is a burgeoning industry but an understanding of the interaction of nanomaterials with the immune system is critical for clinical translation. Macrophages play a fundamental role in the immune system by engulfing foreign particulates such as nanoparticles. When activated, macrophages form distinct phenotypic populations with unique immune functions, however the mechanism by which these polarized macrophages react to nanoparticles is unclear. Furthermore, strategies to selectively evade activated macrophage subpopulations are lacking. Here we demonstrate that stimulated macrophages possess higher phagocytic activities and that classically activated (M1) macrophages exhibit greater phagocytic capacity than alternatively activated (M2) macrophages. We show that modification of nanoparticles with polyethylene-glycol results in decreased clearance by all macrophage phenotypes, but importantly, coating nanoparticles with CD47 preferentially lowers phagocytic activity by the M1 phenotype. These results suggest that bio-inspired nanoparticle surface design may enable evasion of specific components of the immune system and provide a rational approach for developing immune tolerant nanomedicines.
Cell surface expression of channel catfish leukocyte immune-type receptors (IpLITRs) and recruitment of both Src homology 2 domain-containing protein tyrosine phosphatase (SHP)-1 and SHP-2.

Science.gov (United States)

Montgomery, Benjamin C S; Mewes, Jacqueline; Davidson, Chelsea; Burshtyn, Deborah N; Stafford, James L

2009-04-01

Channel catfish leukocyte immune-type receptors (IpLITRs) are immunoglobulin superfamily (IgSF) members believed to play a role in the control and coordination of cellular immune responses in teleost. Putative stimulatory and inhibitory IpLITRs are co-expressed by different types of catfish immune cells (e.g. NK cells, T cells, B cells, and macrophages) but their signaling potential has not been determined. Following cationic polymer-mediated transfections into human cell lines we examined the surface expression, tyrosine phosphorylation, and phosphatase recruitment potential of two types of putative inhibitory IpLITRs using 'chimeric' expression constructs and an epitope-tagged 'native' IpLITR. We also cloned and expressed the teleost Src homology 2 domain-containing protein tyrosine phosphatases (SHP)-1 and SHP-2 and examined their expression in adult tissues and developing zebrafish embryos. Co-immunoprecipitation experiments support the inhibitory signaling potential of distinct IpLITR-types that bound both SHP-1 and SHP-2 following the phosphorylation of tyrosine residues within their cytoplasmic tail (CYT) regions. Phosphatase recruitment by IpLITRs represents an important first step in understanding their influence on immune cell effector functions and suggests that certain inhibitory signaling pathways are conserved among vertebrates.
Expression and Purification of Glycosyltransferases in Pichia Pastoris: Towards Improving the Migration of Stem Cells by Enhancing Surface Expression of Sialyl Lewis X

KAUST Repository

Al-Amoodi, Asma S.

2017-05-01

Recruitment of circulating cells towards target sites is primarily dependent on E-selectin receptor/ligand adhesive interactions. Glycosyltransferase (GTs) are involved in the creation of E-selectin ligands. A sialofucosylated terminal tetrasaccharide like glycan structure known as sialyl Lewis x (sLex), is the most recognized ligand by selectins. This structure is found on the surface of cancer cells and leukocytes but is often absent on the surface of many adult stem cell populations. In order to synthesize sLex, GTs must be endogenously expressed and remain active within the cells. Generally, these stem cells express terminal sialylated lactosamine structures on their glycoproteins which require the addition of alpha-(1,3)-fucose to be converted into an E-selectin ligand. There are a number of fucosyltransferases (FUTs) that are able to modify terminal lactosamine structures to create sLex such as FUT6. In this work we focused on expressing and purifying active recombinant FUTs as a tool to help create sLex structures on the surface of adult stem cells in order to enhance their migration.
Allelic imbalance modulates surface expression of the tolerance-inducing HLA-G molecule on primary trophoblast cells.

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Djurisic, S; Teiblum, S; Tolstrup, C K; Christiansen, O B; Hviid, T V F

2015-03-01

The HLA-G molecule is expressed on trophoblast cells at the feto-maternal interface, where it interacts with local immune cells, and upholds tolerance against the semi-allogeneic fetus. Aberrant HLA-G expression in the placenta and reduced soluble HLA-G levels are observed in pregnancy complications, partly explained by HLA-G polymorphisms which are associated with differences in the alternative splicing pattern and of the stability of HLA-G mRNA. Of special importance is a 14 bp insertion/deletion polymorphism located in the 3'-untranslated region of the HLA-G gene. In the current study, we present novel evidence for allelic imbalance of the 14 bp insertion/deletion polymorphism, using a very accurate and sensitive Digital droplet PCR technique. Allelic imbalance in heterozygous samples was observed as differential expression levels of 14 bp insertion/deletion allele-specific mRNA transcripts, which was further associated with low levels of HLA-G surface expression on primary trophoblast cells. Full gene sequencing of HLA-G allowed us to study correlations between HLA-G extended haplotypes and single-nucleotide polymorphisms and HLA-G surface expression. We found that a 1:1 expression (allelic balance) of the 14 bp insertion/deletion mRNA alleles was associated with high surface expression of HLA-G and with a specific HLA-G extended haplotype. The 14 bp del/del genotype was associated with a significantly lower abundance of the G1 mRNA isoform, and a higher abundance of the G3 mRNA isoform. Overall, the present study provides original evidence for allelic imbalance of the 14 bp insertion/deletion polymorphism, which influences HLA-G surface expression on primary trophoblast cells, considered to be important in the pathogenesis of pre-eclampsia and other pregnancy complications. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.
Two Silene vulgaris copper transporters residing in different cellular compartments confer copper hypertolerance by distinct mechanisms when expressed in Arabidopsis thaliana.

Science.gov (United States)

Li, Yanbang; Iqbal, Mazhar; Zhang, Qianqian; Spelt, Cornelis; Bliek, Mattijs; Hakvoort, Henk W J; Quattrocchio, Francesca M; Koes, Ronald; Schat, Henk

2017-08-01

Silene vulgaris is a metallophyte of calamine, cupriferous and serpentine soils all over Europe. Its metallicolous populations are hypertolerant to zinc (Zn), cadmium (Cd), copper (Cu) or nickel (Ni), compared with conspecific nonmetallicolous populations. These hypertolerances are metal-specific, but the underlying mechanisms are poorly understood. We investigated the role of HMA5 copper transporters in Cu-hypertolerance of a S. vulgaris copper mine population. Cu-hypertolerance in Silene is correlated and genetically linked with enhanced expression of two HMA5 paralogs, SvHMA5I and SvHMA5II, each of which increases Cu tolerance when expressed in Arabidopsis thaliana. Most Spermatophytes, except Brassicaceae, possess homologs of SvHMA5I and SvHMA5II, which originate from an ancient duplication predating the appearance of spermatophytes. SvHMA5II and the A. thaliana homolog AtHMA5 localize in the endoplasmic reticulum and upon Cu exposure move to the plasma membrane, from where they are internalized and degraded in the vacuole. This resembles trafficking of mammalian homologs and is apparently an extremely ancient mechanism. SvHMA5I, instead, neofunctionalized and always resides on the tonoplast, likely sequestering Cu in the vacuole. Adaption of Silene to a Cu-polluted soil is at least in part due to upregulation of two distinct HMA5 transporters, which contribute to Cu hypertolerance by distinct mechanisms. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.
3D facial expression recognition based on histograms of surface differential quantities

KAUST Repository

Li, Huibin

2011-01-01

3D face models accurately capture facial surfaces, making it possible for precise description of facial activities. In this paper, we present a novel mesh-based method for 3D facial expression recognition using two local shape descriptors. To characterize shape information of the local neighborhood of facial landmarks, we calculate the weighted statistical distributions of surface differential quantities, including histogram of mesh gradient (HoG) and histogram of shape index (HoS). Normal cycle theory based curvature estimation method is employed on 3D face models along with the common cubic fitting curvature estimation method for the purpose of comparison. Based on the basic fact that different expressions involve different local shape deformations, the SVM classifier with both linear and RBF kernels outperforms the state of the art results on the subset of the BU-3DFE database with the same experimental setting. © 2011 Springer-Verlag.
Expression of cytokeratins in odontogenic jaw cysts: monoclonal antibodies reveal distinct variation between different cyst types.

Science.gov (United States)

Hormia, M; Ylipaavalniemi, P; Nagle, R B; Virtanen, I

1987-08-01

Immunostaining with monoclonal antibodies was used to study and compare the cytokeratin content of odontogenic cysts and normal gingival epithelium. Two monoclonal antibodies, PKK2 and KA1, stained the whole epithelium in all cyst samples. In gingiva, PKK2 gave a suprabasal staining and KA1 reacted with all epithelial cell layers. Antibodies PKK1, KM 4.62 and KS 8.12 gave a heterogeneous staining in follicular and radicular cysts. In keratocysts and in gingiva PKK1 and KM 4.62 reacted mainly with basal cells and KS 8.12 gave a suprabasal staining. Antibodies reacting with the simple epithelial cytokeratin polypeptide No. 18 (PKK3, KS 18.18) recognized in gingiva only solitary cells compatible with Merkel cells. In a case of follicular ameloblastoma a distinct staining of tumor epithelium was revealed with these antibodies. In 2 follicular cysts, but not in other cyst types, a layer of cytokeratin 18-positive cells was revealed. KA5 and KK 8.60 antibodies, reacting exclusively with keratinizing epithelia, including normal gingiva, gave no reaction in radicular cysts, keratocysts and ameloblastoma. Two of the follicular cysts, were negative for PKK3 and KS 18.18, but reacted strongly with KA5 and KK 8.60. The present results show that odontogenic jaw cysts have distinct differences in their cytokeratin content. With the exception of some follicular cysts, they lack signs of keratinizing epithelial differentiation. Only follicular cysts appear to share with some types of ameloblastoma the expression of cytokeratin polypeptide No. 18.
Sense and antisense transcripts of the developmentally regulated murine hsp70.2 gene are expressed in distinct and only partially overlapping areas in the adult brain

Science.gov (United States)

Murashov, A. K.; Wolgemuth, D. J.

1996-01-01

We have examined the spatial pattern of expression of a member of the hsp70 gene family, hsp70.2, in the mouse central nervous system. Surprisingly, RNA blot analysis and in situ hybridization revealed abundant expression of an 'antisense' hsp70.2 transcript in several areas of adult mouse brain. Two different transcripts recognized by sense and antisense riboprobes for the hsp70.2 gene were expressed in distinct and only partially overlapping neuronal populations. RNA blot analysis revealed low levels of the 2.7 kb transcript of hsp70.2 in several areas of the brain, with highest signal in the hippocampus. Abundant expression of a slightly larger (approximately 2.8 kb) 'antisense' transcript was detected in several brain regions, notably in the brainstem, cerebellum, mesencephalic tectum, thalamus, cortex, and hippocampus. In situ hybridization revealed that the sense and antisense transcripts were both predominantly neuronal and localized to the same cell types in the granular layer of the cerebellum, trapezoid nucleus of the superior olivary complex, locus coeruleus and hippocampus. The hsp70.2 antisense transcripts were particularly abundant in the frontal cortex, dentate gyrus, subthalamic nucleus, zona incerta, superior and inferior colliculi, central gray, brainstem, and cerebellar Purkinje cells. Our findings have revealed a distinct cellular and spatial localization of both sense and antisense transcripts, demonstrating a new level of complexity in the function of the heat shock genes.
Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

DEFF Research Database (Denmark)

Jensen, Helle; Andresen, Lars; Hansen, Karen Aagaard

Heat shock proteins (HSPs) are highly conserved molecules, which support folding of proteins under physiological conditions and mediate protection against lethal damage after various stress stimuli. Five HSP families exist defined by their molecular size (i.e. HSP100, HSP90, HSP70, HSP60, and the......Heat shock proteins (HSPs) are highly conserved molecules, which support folding of proteins under physiological conditions and mediate protection against lethal damage after various stress stimuli. Five HSP families exist defined by their molecular size (i.e. HSP100, HSP90, HSP70, HSP60...... clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 surface expression on cancer cells. We have found that inhibition of histone deacetylase (HDAC) activity leads to surface expression of Hsp70 on various...... hematopoietic cancer cells, an occurance that was not observed on naïve or activated peripheral blood cells. HDAC-inhibitor mediated Hsp70 surface expression was confined to the apoptotic Annexin V positive cells and blocked by inhibition of apoptosis. Other chemotherapeutic inducers of apoptosis...
Distinct expression of muscle-specific microRNAs (myomirs) in brown adipocytes

DEFF Research Database (Denmark)

Walden, Tomas B; Timmons, James A; Keller, Pernille

2009-01-01

MicroRNAs, a novel class of post-transcriptional gene regulators, have been demonstrated to be involved in several cellular processes regulating the expression of protein-coding genes. Here we examine murine white and brown primary cell cultures for differential expression of miRNAs. The adipogen......MicroRNAs, a novel class of post-transcriptional gene regulators, have been demonstrated to be involved in several cellular processes regulating the expression of protein-coding genes. Here we examine murine white and brown primary cell cultures for differential expression of mi...
Developmental expression of a cell surface protein involved in sea urchin skeleton formation

International Nuclear Information System (INIS)

Farach, M.C.; Valdizan, M.; Park, H.R.; Decker, G.L.; Lennarz, W.J.

1986-01-01

The authors have previously used a monoclonal antibody (1223) to identify a 130 Kd cell surface protein involved in skeleton formation is sea urchin embryos. In the current study the authors have examined the expression of the 1223 antigen over the course of development of embryos of two species, Strongylocentrotus purpuratus and Lytechinus pictus. The 130 Kd protein is detected in S. purp eggs on immunoblots. Labeling with [ 3 H] leucine and immunoaffinity chromatography show that it also is synthesized shortly after fertilization. Immunofluroescence reveals that at this early stage the 1223 antigen is uniformly distributed on all of the cells. Synthesis decreases to a minimum by the time of hatching (18 h), as does the total amount of antigen present in the embryo. A second period of synthesis commences at the mesenchyme blastula stage, when the spicule-forming primary mesenchyme cells (PMCs) have appeared. During this later stage, synthesis and cell surface expression are restricted to the PMCs. In contrast to S. purp., in L. pictus the 130 Kd protein does not appear until the PMCs are formed. Hybrid embryos demonstrate a pattern of expression of the maternal species. These results suggest that early expression of 1223 antigen in S. purp. is due to utilization of maternal transcripts present in the egg. In both species later expression in PMCs appears to be the result of cell-type specific synthesis, perhaps encoded by embryonic transcripts
Development of a porcine skeletal muscle cDNA microarray: analysis of differential transcript expression in phenotypically distinct muscles

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Stear Michael

2003-03-01

Full Text Available Abstract Background Microarray profiling has the potential to illuminate the molecular processes that govern the phenotypic characteristics of porcine skeletal muscles, such as hypertrophy or atrophy, and the expression of specific fibre types. This information is not only important for understanding basic muscle biology but also provides underpinning knowledge for enhancing the efficiency of livestock production. Results We report on the de novo development of a composite skeletal muscle cDNA microarray, comprising 5500 clones from two developmentally distinct cDNA libraries (longissimus dorsi of a 50-day porcine foetus and the gastrocnemius of a 3-day-old pig. Clones selected for the microarray assembly were of low to moderate abundance, as indicated by colony hybridisation. We profiled the differential expression of genes between the psoas (red muscle and the longissimus dorsi (white muscle, by co-hybridisation of Cy3 and Cy5 labelled cDNA derived from these two muscles. Results from seven microarray slides (replicates correctly identified genes that were expected to be differentially expressed, as well as a number of novel candidate regulatory genes. Quantitative real-time RT-PCR on selected genes was used to confirm the results from the microarray. Conclusion We have developed a porcine skeletal muscle cDNA microarray and have identified a number of candidate genes that could be involved in muscle phenotype determination, including several members of the casein kinase 2 signalling pathway.
Glycoproteins of mouse vaginal epithelium: differential expression related to estrous cyclicity

DEFF Research Database (Denmark)

Horvat, B; Multhaupt, H A; Damjanov, I

1993-01-01

We used lectin overlay blotting and SDS-PAGE to analyze the estrous cycle-specific expression of mouse vaginal epithelial glycoproteins. Seven lectins chosen for their differential carbohydrate-binding specificity revealed 15 glycoproteins that showed cycle-related expression. Each lectin had...... in proestrus, coincident with the transformation of two superficial layers of vaginal squamous epithelium into mucinous cuboidal cells. Electron microscopic lectin histochemistry revealed the glycoproteins in the mucinous granules of surface cuboidal cells and in the lumen of the vagina. Our results illustrate...... the complexity of glycoconjugate synthesis in mouse vagina and reveal the distinct cycle-specific patterns of individual glycoprotein expression. These cyclic glycoproteins could serve as vaginal biochemical markers for the specific phases of the estrous cycle....

Two distinct genes for ADP/ATP translocase are expressed at the mRNA level in adult human liver

International Nuclear Information System (INIS)

Houldsworth, J.; Attardi, G.

1988-01-01

Several clones hybridizing with a bovine ADP/ATP translocase cDNA were isolated from an adult human liver cDNA library in the vector pEX1. DNA sequence analysis revealed that these clones encode two distinct forms of translocase. In particular, two clones specifying the COOH-end-proximal five-sixths of the protein exhibit a 9% amino acid sequence divergence and totally dissimilar 3' untranslated regions. One of these cDNAs is nearly identical in sequence to an ADP/ATP translocase clone (hp2F1) recently isolated from a human fibroblast cDNA library with three amino acid changes and a few differences in the 3' untranslated region. Another clone isolated from the pEX1 library contains a reading frame encoding the remaining, NH 2 -end-proximal, 37 amino acids of the translocase. This sequence differs significantly (14% amino acid sequence divergence) from the corresponding segment of hp2F1, and the 5' untranslated regions of the two clones are totally dissimilar. RNA transfer hybridization experiments utilizing the clones isolated from the pEX1 library revealed the presence in HeLa cells of three distinct mRNA species. The pattern of hybridization and the sizes of these mRNAs suggest a greater complexity of organization and expression of the ADP/ATP translocase genes in human cells than indicated by the analysis of the cDNA clones
Pathogenic Leptospira species express surface-exposed proteins belonging to the bacterial immunoglobulin superfamily

Science.gov (United States)

Matsunaga, James; Barocchi, Michele A.; Croda, Julio; Young, Tracy A.; Sanchez, Yolanda; Siqueira, Isadora; Bolin, Carole A.; Reis, Mitermayer G.; Riley, Lee W.; Haake, David A.; Ko, Albert I.

2005-01-01

Summary Proteins with bacterial immunoglobulin-like (Big) domains, such as the Yersinia pseudotuberculosis invasin and Escherichia coli intimin, are surface-expressed proteins that mediate host mammalian cell invasion or attachment. Here, we report the identification and characterization of a new family of Big domain proteins, referred to as Lig (leptospiral Ig-like) proteins, in pathogenic Leptospira. Screening of L. interrogans and L. kirschneri expression libraries with sera from leptospirosis patients identified 13 lambda phage clones that encode tandem repeats of the 90 amino acid Big domain. Two lig genes, designated ligA and ligB, and one pseudo-gene, ligC, were identified. The ligA and ligB genes encode amino-terminal lipoprotein signal peptides followed by 10 or 11 Big domain repeats and, in the case of ligB, a unique carboxy-terminal non-repeat domain. The organization of ligC is similar to that of ligB but contains mutations that disrupt the reading frame. The lig sequences are present in pathogenic but not saprophytic Leptospira species. LigA and LigB are expressed by a variety of virulent leptospiral strains. Loss of Lig protein and RNA transcript expression is correlated with the observed loss of virulence during culture attenuation of pathogenic strains. High-pressure freeze substitution followed by immunocytochemical electron microscopy confirmed that the Lig proteins were localized to the bacterial surface. Immunoblot studies with patient sera found that the Lig proteins are a major antigen recognized during the acute host infection. These observations demonstrate that the Lig proteins are a newly identified surface protein of pathogenic Leptospira, which by analogy to other bacterial immunoglobulin superfamily virulence factors, may play a role in host cell attachment and invasion during leptospiral pathogenesis. PMID:12890019
Distinct Fiber Type Signature in Mouse Muscles Expressing a Mutant Lamin A Responsible for Congenital Muscular Dystrophy in a Patient

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Alice Barateau

2017-04-01

Full Text Available Specific mutations in LMNA, which encodes nuclear intermediate filament proteins lamins A/C, affect skeletal muscle tissues. Early-onset LMNA myopathies reveal different alterations of muscle fibers, including fiber type disproportion or prominent dystrophic and/or inflammatory changes. Recently, we identified the p.R388P LMNA mutation as responsible for congenital muscular dystrophy (L-CMD and lipodystrophy. Here, we asked whether viral-mediated expression of mutant lamin A in murine skeletal muscles would be a pertinent model to reveal specific muscle alterations. We found that the total amount and size of muscle fibers as well as the extent of either inflammation or muscle regeneration were similar to wildtype or mutant lamin A. In contrast, the amount of fast oxidative muscle fibers containing myosin heavy chain IIA was lower upon expression of mutant lamin A, in correlation with lower expression of genes encoding transcription factors MEF2C and MyoD. These data validate this in vivo model for highlighting distinct muscle phenotypes associated with different lamin contexts. Additionally, the data suggest that alteration of muscle fiber type identity may contribute to the mechanisms underlying physiopathology of L-CMD related to R388P mutant lamin A.
Based on Molecular Profiling of Gene Expression, Palmoplantar Pustulosis and Palmoplantar Pustular Psoriasis Are Highly Related Diseases that Appear to Be Distinct from Psoriasis Vulgaris.

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Robert Bissonnette

Full Text Available There is a controversy surrounding the existence of palmoplantar pustulosis (PPP and palmoplantar pustular psoriasis (PPPP as separate clinical entities or as variants of the same clinical entity. We used gene expression microarray to compare gene expression in PPP and PPPP.Skin biopsies from subjects with PPP (3, PPPP (6, psoriasis vulgaris (10 and acral skin from normal subjects (7 were analyzed using gene expression microarray. Principal component analysis showed that PPP and PPPP were different from psoriasis vulgaris and normal acral skin. However gene expression of PPP and PPPP clustered together and could not be used to differentiate PPP from PPPP. Gene-wise comparison between PPP and PPPP found no gene to be differentially expressed at a false discovery rate lower than 0.05. Surprisingly we found a higher expression of several genes involved in neural pathways (e.g. GPRIN and ADAM23 in PPP/PPPP as compared to psoriasis vulgaris and normal acral skin. Immunohistochemistry confirmed those findings and showed a keratinocyte localization for those proteins.PPP and PPPP could not be differentiated using gene expression microarray suggesting that they are not distinct clinical entities. Increased expression of GPRIN1, and ADAM23 in keratinocytes suggests that these proteins could be new therapeutic targets for PPP/PPPP.
Spontaneously immortalised bovine mammary epithelial cells exhibit a distinct gene expression pattern from the breast cancer cells

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Li Qianqian

2010-10-01

exhibit non-malignant transformation. Although this cell line displays altered patterns of gene expression, it is clearly distinct from malignant breast cancer cell line. It showed that co-inhibition of cellular senescence and mitochondrial apoptosis pathways coordinates BME65Cs cells immortalisation. Additionally, mechanisms other than gene mutation are likely to be involved in regulation of cellular functions. This study provides an insight into the relationship between cell senescence and immortalisation. BME65Cs cells will be useful in future studies of cellular senescence and tumorigenesis.
Organization of Mitochondrial Gene Expression in Two Distinct Ribosome-Containing Assemblies

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Kirsten Kehrein

2015-02-01

Full Text Available Mitochondria contain their own genetic system that provides subunits of the complexes driving oxidative phosphorylation. A quarter of the mitochondrial proteome participates in gene expression, but how all these factors are orchestrated and spatially organized is currently unknown. Here, we established a method to purify and analyze native and intact complexes of mitochondrial ribosomes. Quantitative mass spectrometry revealed extensive interactions of ribosomes with factors involved in all the steps of posttranscriptional gene expression. These interactions result in large expressosome-like assemblies that we termed mitochondrial organization of gene expression (MIOREX complexes. Superresolution microscopy revealed that most MIOREX complexes are evenly distributed throughout the mitochondrial network, whereas a subset is present as nucleoid-MIOREX complexes that unite the whole spectrum of organellar gene expression. Our work therefore provides a conceptual framework for the spatial organization of mitochondrial protein synthesis that likely developed to facilitate gene expression in the organelle.
Distinct populations of GABAergic neurons in mouse rhombomere 1 express but do not require the homeodomain transcription factor PITX2.

Science.gov (United States)

Waite, Mindy R; Skaggs, Kaia; Kaviany, Parisa; Skidmore, Jennifer M; Causeret, Frédéric; Martin, James F; Martin, Donna M

2012-01-01

Hindbrain rhombomere 1 (r1) is located caudal to the isthmus, a critical organizer region, and rostral to rhombomere 2 in the developing mouse brain. Dorsal r1 gives rise to the cerebellum, locus coeruleus, and several brainstem nuclei, whereas cells from ventral r1 contribute to the trochlear and trigeminal nuclei as well as serotonergic and GABAergic neurons of the dorsal raphe. Recent studies have identified several molecular events controlling dorsal r1 development. In contrast, very little is known about ventral r1 gene expression and the genetic mechanisms regulating its formation. Neurons with distinct neurotransmitter phenotypes have been identified in ventral r1 including GABAergic, serotonergic, and cholinergic neurons. Here we show that PITX2 marks a distinct population of GABAergic neurons in mouse embryonic ventral r1. This population appears to retain its GABAergic identity even in the absence of PITX2. We provide a comprehensive map of markers that places these PITX2-positive GABAergic neurons in a region of r1 that intersects and is potentially in communication with the dorsal raphe. Copyright © 2011 Elsevier Inc. All rights reserved.
Biological and immunological characterization of recombinant Yellow Fever 17D Viruses expressing a Trypanosoma cruzi Amastigote Surface Protein-2 CD8+ T cell epitope at two distinct regions of the genome

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Bonaldo Myrna C

2011-03-01

Full Text Available Abstract Background The attenuated Yellow fever (YF 17D vaccine virus is one of the safest and most effective viral vaccines administered to humans, in which it elicits a polyvalent immune response. Herein, we used the YF 17D backbone to express a Trypanosoma cruzi CD8+ T cell epitope from the Amastigote Surface Protein 2 (ASP-2 to provide further evidence for the potential of this virus to express foreign epitopes. The TEWETGQI CD8+ T cell epitope was cloned and expressed based on two different genomic insertion sites: in the fg loop of the viral Envelope protein and the protease cleavage site between the NS2B and NS3. We investigated whether the site of expression had any influence on immunogenicity of this model epitope. Results Recombinant viruses replicated similarly to vaccine virus YF 17D in cell culture and remained genetically stable after several serial passages in Vero cells. Immunogenicity studies revealed that both recombinant viruses elicited neutralizing antibodies to the YF virus as well as generated an antigen-specific gamma interferon mediated T-cell response in immunized mice. The recombinant viruses displayed a more attenuated phenotype than the YF 17DD vaccine counterpart in mice. Vaccination of a mouse lineage highly susceptible to infection by T. cruzi with a homologous prime-boost regimen of recombinant YF viruses elicited TEWETGQI specific CD8+ T cells which might be correlated with a delay in mouse mortality after a challenge with a lethal dose of T. cruzi. Conclusions We conclude that the YF 17D platform is useful to express T. cruzi (Protozoan antigens at different functional regions of its genome with minimal reduction of vector fitness. In addition, the model T. cruzi epitope expressed at different regions of the YF 17D genome elicited a similar T cell-based immune response, suggesting that both expression sites are useful. However, the epitope as such is not protective and it remains to be seen whether expression
Kirchhoff approximation and closed-form expressions for atom-surface scattering

International Nuclear Information System (INIS)

Marvin, A.M.

1980-01-01

In this paper an approximate solution for atom-surface scattering is presented beyond the physical optics approximation. The potential is well represented by a hard corrugated surface but includes an attractive tail in front. The calculation is carried out analytically by two different methods, and the limit of validity of our formulas is well established in the text. In contrast with other workers, I find those expressions to be exact in both limits of small (Rayleigh region) and large momenta (classical region), with the correct behavior at the threshold. The result is attained through a particular use of the extinction theorem in writing the scattered amplitudes, hitherto not employed, and not for particular boundary values of the field. An explicit evaluation of the field on the surface shows in fact the present formulas to be simply related to the well known Kirchhoff approximation (KA) or more generally to an ''extended'' KA fit to the potential model above. A possible application of the theory to treat strong resonance-overlapping effects is suggested in the last part of the work
Processing OMEGA/Mars Express hyperspectral imagery from radiance-at-sensor to surface reflectance

NARCIS (Netherlands)

Bakker, W.H.; Ruitenbeek, F.J.A. van; Werff, H.M.A. van der; Zegers, T.E.; Oosthoek, J.H.P.; Marsh, S.H.; Meer, F.D. van der

2014-01-01

OMEGA/Mars Express hyperspectral imagery is an excellent source of data for exploring the surface composition of the planet Mars. Compared to terrestrial hyperspectral imagery, the data are challenging to work with; scene-specific transmission models are lacking, spectral features are shallow making
Distinct iris gene expression profiles of primary angle closure glaucoma and primary open angle glaucoma and their interaction with ocular biometric parameters.

Science.gov (United States)

Seet, Li-Fong; Narayanaswamy, Arun; Finger, Sharon N; Htoon, Hla M; Nongpiur, Monisha E; Toh, Li Zhen; Ho, Henrietta; Perera, Shamira A; Wong, Tina T

2016-11-01

This study aimed to evaluate differences in iris gene expression profiles between primary angle closure glaucoma (PACG) and primary open angle glaucoma (POAG) and their interaction with biometric characteristics. Prospective study. Thirty-five subjects with PACG and thirty-three subjects with POAG who required trabeculectomy were enrolled at the Singapore National Eye Centre, Singapore. Iris specimens, obtained by iridectomy, were analysed by real-time polymerase chain reaction for expression of type I collagen, vascular endothelial growth factor (VEGF)-A, -B and -C, as well as VEGF receptors (VEGFRs) 1 and 2. Anterior segment optical coherence tomography (ASOCT) imaging for biometric parameters, including anterior chamber depth (ACD), anterior chamber volume (ACV) and lens vault (LV), was also performed pre-operatively. Relative mRNA levels between PACG and POAG irises, biometric measurements, discriminant analyses using genes and biometric parameters. COL1A1, VEGFB, VEGFC and VEGFR2 mRNA expression was higher in PACG compared to POAG irises. LV, ACD and ACV were significantly different between the two subgroups. Discriminant analyses based on gene expression, biometric parameters or a combination of both gene expression and biometrics (LV and ACV), correctly classified 94.1%, 85.3% and 94.1% of the original PACG and POAG cases, respectively. The discriminant function combining genes and biometrics demonstrated the highest accuracy in cross-validated classification of the two glaucoma subtypes. Distinct iris gene expression supports the pathophysiological differences that exist between PACG and POAG. Biometric parameters can combine with iris gene expression to more accurately define PACG from POAG. © 2016 The Authors. Clinical & Experimental Ophthalmology published by John Wiley & Sons Australia, Ltd on behalf of Royal Australian and New Zealand College of Ophthalmologists.
Ionizing radiation modulates the surface expression of human leukocyte antigen-G in a human melanoma cell line

Energy Technology Data Exchange (ETDEWEB)

Michelin, S.; Gallegos, C.E.; Dubner, D. [Radiopathology Laboratory, Nuclear Regulatory Authority, Buenos Aires (Argentina); Favier, B.; Carosella, E.D. [CEA, I2BM, Hopital Saint-Louis, IUH, Service de Recherches en Hemato-Immunologie, Paris (France)

2009-07-01

Human leukocyte antigen G (HLA-G) is a nonclassical HLA class I molecule involved in fetus protection from the maternal immune system, transplant tolerance, and viral and tumoral immune escape. Tumor-specific HLA-G expression has been described for a wide variety of malignancies, including melanomas. The aim of this study was to evaluate whether ionizing radiation (IR) could modulate the surface expression of HLA-G1 in a human melanoma cell line that expresses endogenously membrane-bound HLA-G1. For this purpose, cells were exposed to increasing doses of {gamma}-irradiation (0-20 Gy) and HLA-G1 levels at the plasma membrane were analyzed at different times postirradiation by flow cytometry. HLA-G total expression and the presence of the soluble form of HLA-G1 (sHLA-G1) in the culture medium of irradiated cells were also evaluated. IR was capable of down regulating cell surface and total HLA-G levels, with a concomitant increase of sHLA-G1 in the medium. These results could indicate that {gamma}-irradiation decreases HLA-G1 surface levels by enhancing the proteolytic cleavage of this molecule. (authors)
Ionizing radiation modulates the surface expression of human leukocyte antigen-G in a human melanoma cell line

International Nuclear Information System (INIS)

Michelin, S.; Gallegos, C.E.; Dubner, D.; Favier, B.; Carosella, E.D.

2009-01-01

Human leukocyte antigen G (HLA-G) is a nonclassical HLA class I molecule involved in fetus protection from the maternal immune system, transplant tolerance, and viral and tumoral immune escape. Tumor-specific HLA-G expression has been described for a wide variety of malignancies, including melanomas. The aim of this study was to evaluate whether ionizing radiation (IR) could modulate the surface expression of HLA-G1 in a human melanoma cell line that expresses endogenously membrane-bound HLA-G1. For this purpose, cells were exposed to increasing doses of γ-irradiation (0-20 Gy) and HLA-G1 levels at the plasma membrane were analyzed at different times postirradiation by flow cytometry. HLA-G total expression and the presence of the soluble form of HLA-G1 (sHLA-G1) in the culture medium of irradiated cells were also evaluated. IR was capable of down regulating cell surface and total HLA-G levels, with a concomitant increase of sHLA-G1 in the medium. These results could indicate that γ-irradiation decreases HLA-G1 surface levels by enhancing the proteolytic cleavage of this molecule. (authors)
Synergetic effect of yeast cell-surface expression of cellulase and expansin-like protein on direct ethanol production from cellulose

Science.gov (United States)

2013-01-01

Background Numerous studies have examined the direct fermentation of cellulosic materials by cellulase-expressing yeast; however, ethanol productivity in these systems has not yet reached an industrial level. Certain microorganisms, such as the cellulolytic fungus Trichoderma reesei, produce expansin-like proteins, which have a cellulose-loosening effect that may increase the breakdown of cellulose. Here, to improve the direct conversion of cellulose to ethanol, yeast Saccharomyces cerevisiae co-displaying cellulase and expansin-like protein on the cell surface were constructed and examined for direct ethanol fermentation performance. Results The cellulase and expansin-like protein co-expressing strain showed 246 mU/g-wet cell of phosphoric acid swollen cellulose (PASC) degradation activity, which corresponded to 2.9-fold higher activity than that of a cellulase-expressing strain. This result clearly demonstrated that yeast cell-surface expressed cellulase and expansin-like protein act synergistically to breakdown cellulose. In fermentation experiments examining direct ethanol production from PASC, the cellulase and expansin-like protein co-expressing strain produced 3.4 g/L ethanol after 96 h of fermentation, a concentration that was 1.4-fold higher than that achieved by the cellulase-expressing strain (2.5 g/L). Conclusions The PASC degradation and fermentation ability of an engineered yeast strain was markedly improved by co-expressing cellulase and expansin-like protein on the cell surface. To our knowledge, this is the first report to demonstrate the synergetic effect of co-expressing cellulase and expansin-like protein on a yeast cell surface, which may be a promising strategy for constructing direct ethanol fermenting yeast from cellulose. PMID:23835302
ER-mitochondria contacts control surface glycan expression and sensitivity to killer lymphocytes in glioma stem-like cells.

Science.gov (United States)

Bassoy, Esen Yonca; Kasahara, Atsuko; Chiusolo, Valentina; Jacquemin, Guillaume; Boydell, Emma; Zamorano, Sebastian; Riccadonna, Cristina; Pellegatta, Serena; Hulo, Nicolas; Dutoit, Valérie; Derouazi, Madiha; Dietrich, Pierre Yves; Walker, Paul R; Martinvalet, Denis

2017-06-01

Glioblastoma is a highly heterogeneous aggressive primary brain tumor, with the glioma stem-like cells (GSC) being more sensitive to cytotoxic lymphocyte-mediated killing than glioma differentiated cells (GDC). However, the mechanism behind this higher sensitivity is unclear. Here, we found that the mitochondrial morphology of GSCs modulates the ER-mitochondria contacts that regulate the surface expression of sialylated glycans and their recognition by cytotoxic T lymphocytes and natural killer cells. GSCs displayed diminished ER-mitochondria contacts compared to GDCs. Forced ER-mitochondria contacts in GSCs increased their cell surface expression of sialylated glycans and reduced their susceptibility to cytotoxic lymphocytes. Therefore, mitochondrial morphology and dynamism dictate the ER-mitochondria contacts in order to regulate the surface expression of certain glycans and thus play a role in GSC recognition and elimination by immune effector cells. Targeting the mitochondrial morphology, dynamism, and contacts with the ER could be an innovative strategy to deplete the cancer stem cell compartment to successfully treat glioblastoma. © 2017 The Authors.
Major aging-associated RNA expressions change at two distinct age-positions

NARCIS (Netherlands)

Gheorghe, M.; Snoeck, M.; Emmerich, M.; Back, T.; Goeman, J.J.; Raz, V.

2014-01-01

BACKGROUND: Genome-wide expression profiles are altered during biological aging and can describe molecular regulation of tissue degeneration. Age-regulated mRNA expression trends from cross-sectional studies could describe how aging progresses. We developed a novel statistical methodology to
The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status.

Science.gov (United States)

Horváthová, Mira; Ilavská, Silvia; Štefíková, Kornélia; Szabová, Michaela; Krivošíková, Zora; Jahnová, Eva; Tulinská, Jana; Spustová, Viera; Gajdoš, Martin

2017-07-11

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs) were in postmenopausal women versus fertile women. Body mass index (BMI) affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells ( p < 0.05). The confounding factors such as women age, BMI, bone mineral density (BMD), waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause.
The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status

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Mira Horváthová

2017-07-01

Full Text Available The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The significant changes of cytotoxic, naive, and memory T-lymphocytes, plasmacytoid dendritic cells (DCs were in postmenopausal women versus fertile women. Body mass index (BMI affected markedly the cell surface expression of CD265/RANK. Osteoporosis is linked to reduced percentage of plasmacytoid DCs, and elevated natural Treg cells (p < 0.05. The confounding factors such as women age, BMI, bone mineral density (BMD, waist size and tissue fat affect the expression of RANK on myeloid DCs and CD40L on T-lymphocytes that might be the immunophenotypic modulators after menopause.
Orf virus interferes with MHC class I surface expression by targeting vesicular transport and Golgi

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Rohde Jörg

2012-07-01

Full Text Available Abstract Background The Orf virus (ORFV, a zoonotic Parapoxvirus, causes pustular skin lesions in small ruminants (goat and sheep. Intriguingly, ORFV can repeatedly infect its host, despite the induction of a specific immunity. These immune modulating and immune evading properties are still unexplained. Results Here, we describe that ORFV infection of permissive cells impairs the intracellular transport of MHC class I molecules (MHC I as a result of structural disruption and fragmentation of the Golgi apparatus. Depending on the duration of infection, we observed a pronounced co-localization of MHC I and COP-I vesicular structures as well as a reduction of MHC I surface expression of up to 50%. These subversion processes are associated with early ORFV gene expression and are accompanied by disturbed carbohydrate trimming of post-ER MHC I. The MHC I population remaining on the cell surface shows an extended half-life, an effect that might be partially controlled also by late ORFV genes. Conclusions The presented data demonstrate that ORFV down-regulates MHC I surface expression in infected cells by targeting the late vesicular export machinery and the structure and function of the Golgi apparatus, which might aid to escape cellular immune recognition.
Activation of p44/42 in Human Natural Killer Cells Decreases Cell-surface Protein Expression: Relationship to Tributyltin-induced alterations of protein expression

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Dudimah, Fred D.; Abraha, Abraham; Wang, Xiaofei; Whalen, Margaret M.

2010-01-01

Tributyltin (TBT) activates the mitogen activated protein kinase (MAPK), p44/42 in human natural killer (NK) cells. TBT also reduces NK cytotoxic function and decreases the expression of several NK-cell proteins. To understand the role that p44/42 activation plays in TBT-induced loss of NK cell function, we have investigated how selective activation of p44/42 by phorbol 12-myristate 13-acetate (PMA) affects NK cells. Previously we showed that PMA caused losses of lytic function similar to those seen with TBT exposures. Here we examined activation of p44/42 in the regulation of NK-cell protein expression and how this regulation may explain the protein expression changes seen with TBT exposures. NK cells exposed to PMA were examined for levels of cell-surface proteins, granzyme mRNA, and perforin mRNA expression. The expression of CD11a, CD16, CD18, and CD56 were reduced, perforin mRNA levels were unchanged and granzyme mRNA levels were increased. To verify that activation of p44/42 was responsible for the alterations seen in CD11a, CD16, CD18, and CD56 with PMA, NK cells were treated with the p44/42 pathway inhibitor (PD98059) prior to PMA exposures. In the presence of PD98059, PMA caused no decreases in the expression of the cell-surface proteins. Results of these studies indicate that the activation of p44/42 may lead to the loss of NK cell cytotoxic function by decreasing the expression of CD11a, CD16, CD18, and CD56. Further, activation of p44/42 appears to be at least in part responsible for the TBT-induced decreases in expression of CD16, CD18, and CD56. PMID:20883105

Fab is the most efficient format to express functional antibodies by yeast surface display.

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Sivelle, Coline; Sierocki, Raphaël; Ferreira-Pinto, Kelly; Simon, Stéphanie; Maillere, Bernard; Nozach, Hervé

2018-04-30

Multiple formats are available for engineering of monoclonal antibodies (mAbs) by yeast surface display, but they do not all lead to efficient expression of functional molecules. We therefore expressed four anti-tumor necrosis factor and two anti-IpaD mAbs as single-chain variable fragment (scFv), antigen-binding fragment (Fab) or single-chain Fabs and compared their expression levels and antigen-binding efficiency. Although the scFv and scFab formats are widely used in the literature, 2 of 6 antibodies were either not or weakly expressed. In contrast, all 6 antibodies expressed as Fab revealed strong binding and high affinity, comparable to that of the soluble form. We also demonstrated that the variations in expression did not affect Fab functionality and were due to variations in light chain display and not to misfolded dimers. Our results suggest that Fab is the most versatile format for the engineering of mAbs.
Distinctive subdomains in the resorbing surface of osteoclasts.

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Kinga A Szewczyk

Full Text Available We employed a novel technique to inspect the substrate-apposed surface of activated osteoclasts, the cells that resorb bone, in the scanning electron microscope. The surface revealed unexpected complexity. At the periphery of the cells were circles and crescents of individual or confluent nodules. These corresponded to the podosomes and actin rings that form a 'sealing zone', encircling the resorptive hemivacuole into which protons and enzymes are secreted. Inside these rings and crescents the osteoclast surface was covered with strips and patches of membrane folds, which were flattened against the substrate surface and surrounded by fold-free membrane in which many orifices could be seen. Corresponding regions of folded and fold-free membrane were found by transmission electron microscopy in osteoclasts incubated on bone. We correlated these patterns with the distribution of several proteins crucial to resorption. The strips and patches of membrane folds corresponded in distribution to vacuolar H+-ATPase, and frequently co-localized with F-actin. Cathepsin K localized to F-actin-free foci towards the center of cells with circular actin rings, and at the retreating pole of cells with actin crescents. The chloride/proton antiporter ClC-7 formed a sharply-defined band immediately inside the actin ring, peripheral to vacuolar H+-ATPase. The sealing zone of osteoclasts is permeable to molecules with molecular mass up to 10,000. Therefore, ClC-7 might be distributed at the periphery of the resorptive hemivacuole in order to prevent protons from escaping laterally from the hemivacuole into the sealing zone, where they would dissolve the bone mineral. Since the activation of resorption is attributable to recognition of the αVβ3 ligands bound to bone mineral, such leakage would, by dissolving bone mineral, release the ligands and so terminate resorption. Therefore, ClC-7 might serve not only to provide the counter-ions that enable proton pumping, but
Distinct expression profile in fumarate-hydratase-deficient uterine fibroids

DEFF Research Database (Denmark)

Vanharanta, S; Pollard, PJ; Lehtonen, HJ

2006-01-01

-related genes. Other significantly up-regulated gene categories in FH mutants were, for example, iron ion homeostasis and oxidoreduction. Genes with lower expression in FH-mutant fibroids belonged to groups such as extracellular matrix, cell adhesion, muscle development and cell contraction. We show that FH...
Computational exploration of cis-regulatory modules in rhythmic expression data using the "Exploration of Distinctive CREs and CRMs" (EDCC) and "CRM Network Generator" (CNG) programs.

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Bekiaris, Pavlos Stephanos; Tekath, Tobias; Staiger, Dorothee; Danisman, Selahattin

2018-01-01

Understanding the effect of cis-regulatory elements (CRE) and clusters of CREs, which are called cis-regulatory modules (CRM), in eukaryotic gene expression is a challenge of computational biology. We developed two programs that allow simple, fast and reliable analysis of candidate CREs and CRMs that may affect specific gene expression and that determine positional features between individual CREs within a CRM. The first program, "Exploration of Distinctive CREs and CRMs" (EDCC), correlates candidate CREs and CRMs with specific gene expression patterns. For pairs of CREs, EDCC also determines positional preferences of the single CREs in relation to each other and to the transcriptional start site. The second program, "CRM Network Generator" (CNG), prioritizes these positional preferences using a neural network and thus allows unbiased rating of the positional preferences that were determined by EDCC. We tested these programs with data from a microarray study of circadian gene expression in Arabidopsis thaliana. Analyzing more than 1.5 million pairwise CRE combinations, we found 22 candidate combinations, of which several contained known clock promoter elements together with elements that had not been identified as relevant to circadian gene expression before. CNG analysis further identified positional preferences of these CRE pairs, hinting at positional information that may be relevant for circadian gene expression. Future wet lab experiments will have to determine which of these combinations confer daytime specific circadian gene expression.
Correlation of cell surface proteins of distinct Beauveria bassiana cell types and adaption to varied environment and interaction with the host insect.

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Yang, Zhi; Jiang, Hongyan; Zhao, Xin; Lu, Zhuoyue; Luo, Zhibing; Li, Xuebing; Zhao, Jing; Zhang, Yongjun

2017-02-01

The insect fungal pathogen Beauveria bassiana produces a number of distinct cell types that include aerial conidia, blastospores and haemolymph-derived cells, termed hyphal bodies, to adapt varied environment niches and within the host insect. These cells display distinct biochemical properties and surface structures, and a highly ordered outermost brush-like structure uniquely present on hyphal bodies, but not on any in vitro cells. Here, we found that the outermost structure on the hyphal bodies mainly consisted of proteins associated to structural wall components in that most of it could be removed by dithiothreitol (DTT) or proteinase K. DTT-treatment also caused delayed germination, decreased tolerance to ultraviolet irradiation and virulence of conidia or blastospores, with decreased adherence and alternated carbohydrate epitopes, suggesting involvement in fungal development, stress responses and virulence. To characterize these cell surface molecules, proteins were released from the living cells using DTT, and identified and quantitated using label-free quantitative mass spectrometry. Thereafter, a series of bioinformatics programs were used to predict cell surface-associated proteins (CSAPs), and 96, 166 and 54 CSAPs were predicted from the identified protein pools of conidia, blastospores and hyphal bodies, respectively, which were involved in utilization of carbohydrate, nitrogen, and lipid, detoxification, pathogen-host interaction, and likely other cellular processes. Thirteen, sixty-nine and six CSAPs were exclusive in conidia, blastospores and hyphal bodies, respectively, which were verified by eGFP-tagged proteins at their N-terminus. Our data provide a crucial cue to understand mechanism of B. bassiana to adapt to varied environment and interaction with insect host. Copyright © 2016 Elsevier Inc. All rights reserved.
Different gene-expression profiles for the poorly differentiated carcinoma and the highly differentiated papillary adenocarcinoma in mammary glands support distinct metabolic pathways

International Nuclear Information System (INIS)

Eilon, Tali; Barash, Itamar

2008-01-01

Deregulation of Stat5 in the mammary gland of transgenic mice causes tumorigenesis. Poorly differentiated carcinoma and highly differentiated papillary adenocarcinoma tumors evolve. To distinguish the genes and elucidate the cellular processes and metabolic pathways utilized to preserve these phenotypes, gene-expression profiles were analyzed. Mammary tumors were excised from transgenic mice carrying a constitutively active variant of Stat5, or a Stat5 variant lacking s transactivation domain. These tumors displayed either the carcinoma or the papillary adenocarcinoma phenotypes. cRNAs, prepared from each tumor were hybridized to an Affymetrix GeneChip ® Mouse Genome 430A 2.0 array. Gene-ontology analysis, hierarchical clustering and biological-pathway analysis were performed to distinct the two types of tumors. Histopathology and immunofluorescence staining complemented the comparison between the tumor phenotypes. The nucleus-cytoskeleton-plasma membrane axis is a major target for differential gene expression between phenotypes. In the carcinoma, stronger expression of genes coding for specific integrins, cytoskeletal proteins and calcium-binding proteins highlight cell-adhesion and motility features of the tumor cells. This is supported by the higher expression of genes involved in O-glycan synthesis, TGF-β, activin, their receptors and Smad3, as well as the Notch ligands and members of the γ-secretase complex that enable Notch nuclear localization. The Wnt pathway was also a target for differential gene expression. Higher expression of genes encoding the degradation complex of the canonical pathway and limited TCF expression in the papillary adenocarcinoma result in membranal accumulation of β-catenin, in contrast to its nuclear translocation in the carcinoma. Genes involved in cell-cycle arrest at G1 and response to DNA damage were more highly expressed in the papillary adenocarcinomas, as opposed to favored G2/M regulation in the carcinoma tumors. At least
Are Titan's radial Labyrinth terrains surface expressions of large laccoliths?

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Schurmeier, L.; Dombard, A. J.; Malaska, M.; Radebaugh, J.

2017-12-01

The Labyrinth terrain unit may be the one of the best examples of the surface expression of Titan's complicated history. They are characterized as highly eroded, dissected, and elevated plateaus and remnant ridges, with an assumed composition that is likely organic-rich based on radar emissivity. How these features accumulated organic-rich sediments and formed topographic highs by either locally uplifting or surviving pervasive regional deflation or erosion is an important question for understanding the history of Titan. There are several subsets of Labyrinth terrains, presumably with differing evolutionary histories and formation processes. We aim to explain the formation of a subset of Labyrinth terrain units informally referred to as "radial Labyrinth terrains." They are elevated and appear dome-like, circular in planform, have a strong radial dissection pattern, are bordered by Undifferentiated Plains units, and are found in the mid-latitudes. Based on their shape, clustering, and dimensions, we suggest that they may be the surface expression of large subsurface laccoliths. A recent study by Manga and Michaut (Icarus, 2017) explained Europa's lenticulae (pits, domes, spots) with the formation of saucer-shaped sills that form laccoliths around the brittle-ductile transition depth within the ice shell (1-5 km). Here, we apply the same scaling relationships and find that the larger size of radial labyrinth terrains with Titan's higher gravity implies deeper intrusion depths of around 20-40 km. This intrusion depth matches the expected brittle-ductile transition on Titan based on our finite element simulations and yield strength envelope analyses. We hypothesize that Titan's radial labyrinth terrains formed as cryovolcanic (water) intrusions that rose to the brittle-ductile transition within the ice shell where they spread horizontally, and uplifted the overlying ice. The organic-rich sedimentary cover also uplifted, becoming more susceptible to pluvial and fluvial
Modified expression of surface glyconjugates in stored human platelets

International Nuclear Information System (INIS)

Dhar, A.; Ganguly, P.

1987-01-01

Platelets are anucleated cells which play an important part in blood coagulation and thrombosis. These cells may be stored in the blood bank for only 4/5 days. In order to improve the storage of platelets, it is essential to first understand the changes in these cells due to storage. In this work, human platelets were stored in autologous plasma at 4 0 or 22 0 and their surface changes were monitored with three lectins - wheat germ afflutinin (WGA), concanavalin A (Con A) and lentil lectin (LL). Blood was drawn from healthy donors and platelet rich plasma (PRP) was collected by slow speed centrifugation. Platelets stored at either temperature for different times showed increased sensitivity to agglutination by WGA after 34-48 hrs. Lectins, Con A and LL, which were not agglutinating to fresh platelets readily caused agglutination after 48-72 hrs. The platelets stored for 25 hrs or longer period were insensitive to thrombin but showed enhanced aggregation with WGA. Labelling of surface glycoconjugates of stored platelets with 3 H-boro-hydride revealed progressive loss of a glycoprotein of Mr 150,000 (GPIb infinity) together with the appearance of components of Mr 69,000; Mr 60,000; Mr 25,000. New high molecular weight glycoproteins were also detected only in stored platelets. The author studies clearly indicate that modification or altered expression of platelets surface glycoproteins may be one factor of storage related dysfunction of platelets
Modified expression of surface glyconjugates in stored human platelets

Energy Technology Data Exchange (ETDEWEB)

Dhar, A.; Ganguly, P.

1987-05-01

Platelets are anucleated cells which play an important part in blood coagulation and thrombosis. These cells may be stored in the blood bank for only 4/5 days. In order to improve the storage of platelets, it is essential to first understand the changes in these cells due to storage. In this work, human platelets were stored in autologous plasma at 4/sup 0/ or 22/sup 0/ and their surface changes were monitored with three lectins - wheat germ afflutinin (WGA), concanavalin A (Con A) and lentil lectin (LL). Blood was drawn from healthy donors and platelet rich plasma (PRP) was collected by slow speed centrifugation. Platelets stored at either temperature for different times showed increased sensitivity to agglutination by WGA after 34-48 hrs. Lectins, Con A and LL, which were not agglutinating to fresh platelets readily caused agglutination after 48-72 hrs. The platelets stored for 25 hrs or longer period were insensitive to thrombin but showed enhanced aggregation with WGA. Labelling of surface glycoconjugates of stored platelets with /sup 3/H-boro-hydride revealed progressive loss of a glycoprotein of Mr 150,000 (GPIb infinity) together with the appearance of components of Mr 69,000; Mr 60,000; Mr 25,000. New high molecular weight glycoproteins were also detected only in stored platelets. The author studies clearly indicate that modification or altered expression of platelets surface glycoproteins may be one factor of storage related dysfunction of platelets.
Basement to surface expressions and critical factors in the genesis of unconformity-related deposits

International Nuclear Information System (INIS)

Potter, Eric

2014-01-01

Two subprojects: 1) Basement to surface expressions of deep mineralization and refinement of critical factors leading to the genesis of unconformity-related uranium deposits; and 2) Recognition of uranium ore system alteration signatures in complex terranes: IOCG vs albite-hosted uranium vs volcanic-hosted uranium.
Distinct functional characteristics of levocabastine sensitive rat neurotensin NT2 receptor expressed in Chinese hamster ovary cells.

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Yamada, M; Yamada, M; Lombet, A; Forgez, P; Rostène, W

1998-01-01

Neurotensin has been shown to produce pharmacological effects both in brain and periphery. Several of these effects are mediated by a high-affinity neurotensin NT1 receptor. On the other hand, a low-affinity levocabastine-sensitive neurotensin NT2 receptor was molecularly cloned from rodent brain recently. In this study, in contrast to NT1 receptor, levocabastine (a histamine H1 receptor antagonist) and SR48692 (an antagonist for NT1 receptor) strongly stimulated intracellular Ca2+ mobilization in transfected Chinese hamster ovary cells expressing rat NT2 receptor, thus acting as potent NT2 receptor. Furthermore, despite of their affinities for NT2 receptor, the Ca2+ responses to potent NT1 agonists, neurotensin or JMV449 ([Lys8-(CH2NH)-Lys9]Pro-Tyr-Ile-Leu, a peptidase resistant analogue of neurotensin) were much smaller than that observed with SR48692. These findings suggest that NT1 and NT2 receptors present distinct functional characteristics and that SR48692 may act as a potent agonist for NT2 receptor.
Influence of dietary nicotinic acid supplementation on lipid metabolism and related gene expression in two distinct broiler breeds of female chickens.

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Jiang, R R; Zhao, G P; Zhao, J P; Chen, J L; Zheng, M Q; Liu, R R; Wen, J

2014-10-01

This study aimed to evaluate the influence of supplemental dietary nicotinic acid (NA) on lipid metabolism and hepatic expression of related genes in female chickens of two distinct broiler strains [Arbor Acres (AA) and Beijing-You (BJY)]. The treatments were arranged in a 2 × 4 factorial in a completely randomized design. Day-old females (n = 384) were allocated to four treatments with six cages per treatment and fed diets (basal contained approximately 25 mg NA/kg) supplemented with 0, 30, 60 and 120 mg NA/kg. A sample of 72 birds from each breed was slaughtered and sampled at their different market times (8 week for AA and 16 week for BJY). Arbor Acres broilers had thickness of subcutaneous fat plus the skin (SFS), and plasma concentration of low-density lipoprotein cholesterol (LDLC) and lower percentage of abdominal fat (PAF), plasma concentrations of TG, NEFA and adiponectin than the BJY line. The hepatic transcription of apolipoprotein A-I (ApoA-I), apolipoproteinB (ApoB), and adiponectin was significantly higher in AA broilers than in BJY broilers. In both breeds, BW, PAF, SFS, NEFA and TG were increased with increasing supplementation from 0 to 60 mg NA/kg, but then decreased slightly with 120 mg added NA/kg. With increasing supplementation, hepatic expression and plasma concentrations of adiponectin decreased from 0 to 60 mg added NA/kg and then increased with 120 mg added NA/kg. The expression of ApoA-I and ApoB mRNA showed linear response to dietary supplementation with NA. These findings indicate that: (i) supplementation of NA influenced the lipid metabolism and related gene expression; (ii) when supplemented with 120 mg NA/kg, some pharmacologic actions on lipid metabolism appeared; and (iii) changes in BW and fat deposition appeared to be associated with hepatic expression of adiponectin.
High-risk human papillomavirus E7 expression reduces cell-surface MHC class I molecules and increases susceptibility to natural killer cells

DEFF Research Database (Denmark)

Bottley, G; Watherston, O G; Hiew, Y-L

2007-01-01

a role for E7 in tumour immune evasion. We show that knockdown of E7 expression in HPV16- and HPV18-transformed cervical carcinoma cells by RNA interference increased expression of major histocompatibility complex (MHC) class I at the cell surface and reduced susceptibility of these cells to natural...... killer (NK) cells. Tetracycline-regulated induction of HPV16 E7 resulted in reduced expression of cell surface MHC class I molecules and increased NK cell killing. Our results suggest that, for HPV-associated malignancies, reduced MHC class I expression is the result of an active immune evasion strategy...
The SnSAG merozoite surface antigens of Sarcocystis neurona are expressed differentially during the bradyzoite and sporozoite life cycle stages.

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Gautam, A; Dubey, J P; Saville, W J; Howe, D K

2011-12-29

Sarcocystis neurona is a two-host coccidian parasite whose complex life cycle progresses through multiple developmental stages differing at morphological and molecular levels. The S. neurona merozoite surface is covered by multiple, related glycosylphosphatidylinositol-linked proteins, which are orthologous to the surface antigen (SAG)/SAG1-related sequence (SRS) gene family of Toxoplasma gondii. Expression of the SAG/SRS proteins in T. gondii and another related parasite Neospora caninum is life-cycle stage specific and seems necessary for parasite transmission and persistence of infection. In the present study, the expression of S. neurona merozoite surface antigens (SnSAGs) was evaluated in the sporozoite and bradyzoite stages. Western blot analysis was used to compare SnSAG expression in merozoites versus sporozoites, while immunocytochemistry was performed to examine expression of the SnSAGs in merozoites versus bradyzoites. These analyses revealed that SnSAG2, SnSAG3 and SnSAG4 are expressed in sporozoites, while SnSAG5 was appeared to be downregulated in this life cycle stage. In S. neurona bradyzoites, it was found that SnSAG2, SnSAG3, SnSAG4 and SnSAG5 were either absent or expression was greatly reduced. As shown for T. gondii, stage-specific expression of the SnSAGs may be important for the parasite to progress through its developmental stages and complete its life cycle successfully. Thus, it is possible that the SAG switching mechanism by these parasites could be exploited as a point of intervention. As well, the alterations in surface antigen expression during different life cycle stages may need to be considered when designing prospective approaches for protective vaccination. Copyright © 2011 Elsevier B.V. All rights reserved.
Commentary Relative to the Emission Spectrum of the Solar Atmosphere: Further Evidence for a Distinct Solar Surface

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Robitaille P.-M.

2013-07-01

Full Text Available The chromosphere and corona of the Sun represent tenuous regions which are characterized by numerous optically thin emission lines in the ultraviolet and X-ray bands. When observed from the center of the solar disk outward, these emission lines experience modest brightening as the limb is approached. The intensity of many ultraviolet and X-ray emission lines nearly doubles when observation is extended just beyond the edge of the disk. These ﬁndings indicate that the solar body is opaque in this frequency range and that an approximately two fold greater region of the solar atmosphere is being sampled outside the limb. These observations provide strong support for the presence of a distinct solar surface. Therefore, the behavior of the emission lines in this frequency range constitutes the twenty ﬁfth line of evidence that the Sun is comprised of condensed matter
A distinct bacterial dysbiosis associated skin inflammation in ovine footrot

Science.gov (United States)

Maboni, Grazieli; Blanchard, Adam; Frosth, Sara; Stewart, Ceri; Emes, Richard; Tötemeyer, Sabine

2017-03-01

Ovine footrot is a highly prevalent bacterial disease caused by Dichelobacter nodosus and characterised by the separation of the hoof horn from the underlying skin. The role of innate immune molecules and other bacterial communities in the development of footrot lesions remains unclear. This study shows a significant association between the high expression of IL1β and high D. nodosus load in footrot samples. Investigation of the microbial population identified distinct bacterial populations in the different disease stages and also depending on the level of inflammation. Treponema (34%), Mycoplasma (29%) and Porphyromonas (15%) were the most abundant genera associated with high levels of inflammation in footrot. In contrast, Acinetobacter (25%), Corynebacteria (17%) and Flavobacterium (17%) were the most abundant genera associated with high levels of inflammation in healthy feet. This demonstrates for the first time there is a distinct microbial community associated with footrot and high cytokine expression.
Age-dependent brain gene expression and copy number anomalies in autism suggest distinct pathological processes at young versus mature ages.

Science.gov (United States)

Chow, Maggie L; Pramparo, Tiziano; Winn, Mary E; Barnes, Cynthia Carter; Li, Hai-Ri; Weiss, Lauren; Fan, Jian-Bing; Murray, Sarah; April, Craig; Belinson, Haim; Fu, Xiang-Dong; Wynshaw-Boris, Anthony; Schork, Nicholas J; Courchesne, Eric

2012-01-01

Autism is a highly heritable neurodevelopmental disorder, yet the genetic underpinnings of the disorder are largely unknown. Aberrant brain overgrowth is a well-replicated observation in the autism literature; but association, linkage, and expression studies have not identified genetic factors that explain this trajectory. Few studies have had sufficient statistical power to investigate whole-genome gene expression and genotypic variation in the autistic brain, especially in regions that display the greatest growth abnormality. Previous functional genomic studies have identified possible alterations in transcript levels of genes related to neurodevelopment and immune function. Thus, there is a need for genetic studies involving key brain regions to replicate these findings and solidify the role of particular functional pathways in autism pathogenesis. We therefore sought to identify abnormal brain gene expression patterns via whole-genome analysis of mRNA levels and copy number variations (CNVs) in autistic and control postmortem brain samples. We focused on prefrontal cortex tissue where excess neuron numbers and cortical overgrowth are pronounced in the majority of autism cases. We found evidence for dysregulation in pathways governing cell number, cortical patterning, and differentiation in young autistic prefrontal cortex. In contrast, adult autistic prefrontal cortex showed dysregulation of signaling and repair pathways. Genes regulating cell cycle also exhibited autism-specific CNVs in DNA derived from prefrontal cortex, and these genes were significantly associated with autism in genome-wide association study datasets. Our results suggest that CNVs and age-dependent gene expression changes in autism may reflect distinct pathological processes in the developing versus the mature autistic prefrontal cortex. Our results raise the hypothesis that genetic dysregulation in the developing brain leads to abnormal regional patterning, excess prefrontal neurons
Age-dependent brain gene expression and copy number anomalies in autism suggest distinct pathological processes at young versus mature ages.

Directory of Open Access Journals (Sweden)

Maggie L Chow

Full Text Available Autism is a highly heritable neurodevelopmental disorder, yet the genetic underpinnings of the disorder are largely unknown. Aberrant brain overgrowth is a well-replicated observation in the autism literature; but association, linkage, and expression studies have not identified genetic factors that explain this trajectory. Few studies have had sufficient statistical power to investigate whole-genome gene expression and genotypic variation in the autistic brain, especially in regions that display the greatest growth abnormality. Previous functional genomic studies have identified possible alterations in transcript levels of genes related to neurodevelopment and immune function. Thus, there is a need for genetic studies involving key brain regions to replicate these findings and solidify the role of particular functional pathways in autism pathogenesis. We therefore sought to identify abnormal brain gene expression patterns via whole-genome analysis of mRNA levels and copy number variations (CNVs in autistic and control postmortem brain samples. We focused on prefrontal cortex tissue where excess neuron numbers and cortical overgrowth are pronounced in the majority of autism cases. We found evidence for dysregulation in pathways governing cell number, cortical patterning, and differentiation in young autistic prefrontal cortex. In contrast, adult autistic prefrontal cortex showed dysregulation of signaling and repair pathways. Genes regulating cell cycle also exhibited autism-specific CNVs in DNA derived from prefrontal cortex, and these genes were significantly associated with autism in genome-wide association study datasets. Our results suggest that CNVs and age-dependent gene expression changes in autism may reflect distinct pathological processes in the developing versus the mature autistic prefrontal cortex. Our results raise the hypothesis that genetic dysregulation in the developing brain leads to abnormal regional patterning, excess
Age-Dependent Brain Gene Expression and Copy Number Anomalies in Autism Suggest Distinct Pathological Processes at Young Versus Mature Ages

Science.gov (United States)

Winn, Mary E.; Barnes, Cynthia Carter; Li, Hai-Ri; Weiss, Lauren; Fan, Jian-Bing; Murray, Sarah; April, Craig; Belinson, Haim; Fu, Xiang-Dong; Wynshaw-Boris, Anthony; Schork, Nicholas J.; Courchesne, Eric

2012-01-01

Autism is a highly heritable neurodevelopmental disorder, yet the genetic underpinnings of the disorder are largely unknown. Aberrant brain overgrowth is a well-replicated observation in the autism literature; but association, linkage, and expression studies have not identified genetic factors that explain this trajectory. Few studies have had sufficient statistical power to investigate whole-genome gene expression and genotypic variation in the autistic brain, especially in regions that display the greatest growth abnormality. Previous functional genomic studies have identified possible alterations in transcript levels of genes related to neurodevelopment and immune function. Thus, there is a need for genetic studies involving key brain regions to replicate these findings and solidify the role of particular functional pathways in autism pathogenesis. We therefore sought to identify abnormal brain gene expression patterns via whole-genome analysis of mRNA levels and copy number variations (CNVs) in autistic and control postmortem brain samples. We focused on prefrontal cortex tissue where excess neuron numbers and cortical overgrowth are pronounced in the majority of autism cases. We found evidence for dysregulation in pathways governing cell number, cortical patterning, and differentiation in young autistic prefrontal cortex. In contrast, adult autistic prefrontal cortex showed dysregulation of signaling and repair pathways. Genes regulating cell cycle also exhibited autism-specific CNVs in DNA derived from prefrontal cortex, and these genes were significantly associated with autism in genome-wide association study datasets. Our results suggest that CNVs and age-dependent gene expression changes in autism may reflect distinct pathological processes in the developing versus the mature autistic prefrontal cortex. Our results raise the hypothesis that genetic dysregulation in the developing brain leads to abnormal regional patterning, excess prefrontal neurons
Distinct expression profile of HCMV encoded miRNAs in plasma from oral lichen planus patients.

Science.gov (United States)

Ding, Meng; Wang, Xiang; Wang, Cheng; Liu, Xiaoshuang; Zen, Ke; Wang, Wenmei; Zhang, Chen-Yu; Zhang, Chunni

2017-06-07

Oral lichen planus (OLP) is a T cell-mediated autoimmune disease. The aetiology and molecular mechanisms of OLP remain unclear. Human cytomegalovirus (HCMV) infection is a causal factor in the development of various diseases, but the clinical relevance of HCMV in OLP has not been thoroughly investigated. In the present study, we firstly examined twenty-three HCMV-encoded microRNA (miRNA) expression profiles in plasma from training set that including 21 OLP patients and 18 healthy controls using RT-qPCR technology. Dysregulated miRNAs were subsequently confirmed in another larger cohort refereed as validation set consisting of 40 OLP patients and 33 healthy controls. HCMV DNA in peripheral blood leukocytes (PBLs) was also measured in an additional cohort of 13 OLP patients and 12 control subjects. Furthermore, bioinformatics analyses, luciferase reporter assay and western blotting were also performed to predict and verify the direct potential targets of HCMV-encoded miRNAs. The RT-qPCR results showed that the plasma levels of five HCMV-encoded miRNAs including hcmv-miR-UL112-3p, hcmv-miR-UL22a-5p, hcmv-miR-UL148d, hcmv-miR-UL36-5p and hcmv-miR-UL59 were significantly increased in OLP patients in both training and validation sets. HCMV DNA in PBLs was also significantly higher in OLP patients than in control subjects. Additionally, by using a combination of luciferase reporter assay and western blotting, we demonstrated that cytomegalovirus UL16-binding protein 1, a molecule that mediates the killing of virus-infected cells by natural killer cells, is a direct target of hcmv-miR-UL59. Our results demonstrate a distinct expression pattern of HCMV-encoded miRNAs in OLP patients, which may provide insight into the relationship between HCMV infection and OLP, and warrants additional study in the diagnosis and aetiology of OLP.

Whole-Genome Expression Analysis of Human Mesenchymal Stromal Cells Exposed to Ultrasmooth Tantalum vs. Titanium Oxide Surfaces

DEFF Research Database (Denmark)

Stiehler, C.; Bunger, C.; Overall, R. W.

2013-01-01

to titanium (Ti) surface. The aim of this study was to extend the previous investigation of biocompatibility by monitoring temporal gene expression of MSCs on topographically comparable smooth Ta and Ti surfaces using whole-genome gene expression analysis. Total RNA samples from telomerase-immortalized human...... MSCs cultivated on plain sputter-coated surfaces of Ta or Ti for 1, 2, 4, and 8 days were hybridized to n = 16 U133 Plus 2.0 arrays (Affymetrix(A (R))). Functional annotation, cluster and pathway analyses were performed. The vast majority of genes were differentially regulated after 4 days...... of cultivation and genes upregulated by MSCs exposed to Ta and Ti were predominantly related to the processes of differentiation and transcription, respectively. Functional annotation analysis of the 1,000 temporally most significantly regulated genes suggests earlier cellular differentiation on Ta compared...
Nitrogen transporter and assimilation genes exhibit developmental stage-selective expression in maize (Zea mays L.) associated with distinct cis-acting promoter motifs.

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Liseron-Monfils, Christophe; Bi, Yong-Mei; Downs, Gregory S; Wu, Wenqing; Signorelli, Tara; Lu, Guangwen; Chen, Xi; Bondo, Eddie; Zhu, Tong; Lukens, Lewis N; Colasanti, Joseph; Rothstein, Steven J; Raizada, Manish N

2013-10-01

Nitrogen is considered the most limiting nutrient for maize (Zea mays L.), but there is limited understanding of the regulation of nitrogen-related genes during maize development. An Affymetrix 82K maize array was used to analyze the expression of ≤ 46 unique nitrogen uptake and assimilation probes in 50 maize tissues from seedling emergence to 31 d after pollination. Four nitrogen-related expression clusters were identified in roots and shoots corresponding to, or overlapping, juvenile, adult, and reproductive phases of development. Quantitative real time PCR data was consistent with the existence of these distinct expression clusters. Promoters corresponding to each cluster were screened for over-represented cis-acting elements. The 8-bp distal motif of the Arabidopsis 43-bp nitrogen response element (NRE) was over-represented in nitrogen-related maize gene promoters. This conserved motif, referred to here as NRE43-d8, was previously shown to be critical for nitrate-activated transcription of nitrate reductase (NIA1) and nitrite reductase (NIR1) by the NIN-LIKE PROTEIN 6 (NLP6) in Arabidopsis. Here, NRE43-d8 was over-represented in the promoters of maize nitrate and ammonium transporter genes, specifically those that showed peak expression during early-stage vegetative development. This result predicts an expansion of the NRE-NLP6 regulon and suggests that it may have a developmental component in maize. We also report leaf expression of putative orthologs of nitrite transporters (NiTR1), a transporter not previously reported in maize. We conclude by discussing how each of the four transcriptional modules may be responsible for the different nitrogen uptake and assimilation requirements of leaves and roots at different stages of maize development.
Low Constitutive Cell Surface Expression of HLA-B Is Caused by a Posttranslational Mechanism Involving Glu180 and Arg239

DEFF Research Database (Denmark)

Dellgren, Christoffer; Ekwelum, Vanessa A. C.; Ormhøj, Maria

2016-01-01

HLA class I cell surface expression is crucial for normal immune responses, and variability in HLA expression may influence the course of infections. We have previously shown that classical HLA class I expression on many human cell types is biased with greatly reduced expression of HLA-B compared...... by affecting HLA processing in the Ag presentation pathway....
Suppression of tumor growth and angiogenesis by a specific antagonist of the cell-surface expressed nucleolin.

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Damien Destouches

Full Text Available BACKGROUND: Emerging evidences suggest that nucleolin expressed on the cell surface is implicated in growth of tumor cells and angiogenesis. Nucleolin is one of the major proteins of the nucleolus, but it is also expressed on the cell surface where is serves as a binding protein for variety of ligands implicated in cell proliferation, differentiation, adhesion, mitogenesis and angiogenesis. METHODOLOGY/PRINCIPAL FINDINGS: By using a specific antagonist that binds the C-terminal tail of nucleolin, the HB-19 pseudopeptide, here we show that the growth of tumor cells and angiogenesis are suppressed in various in vitro and in vivo experimental models. HB-19 inhibited colony formation in soft agar of tumor cell lines, impaired migration of endothelial cells and formation of capillary-like structures in collagen gel, and reduced blood vessel branching in the chick embryo chorioallantoic membrane. In athymic nude mice, HB-19 treatment markedly suppressed the progression of established human breast tumor cell xenografts in nude mice, and in some cases eliminated measurable tumors while displaying no toxicity to normal tissue. This potent antitumoral effect is attributed to the direct inhibitory action of HB-19 on both tumor and endothelial cells by blocking and down regulating surface nucleolin, but without any apparent effect on nucleolar nucleolin. CONCLUSION/SIGNIFICANCE: Our results illustrate the dual inhibitory action of HB-19 on the tumor development and the neovascularization process, thus validating the cell-surface expressed nucleolin as a strategic target for an effective cancer drug. Consequently, the HB-19 pseudopeptide provides a unique candidate to consider for innovative cancer therapy.
Chronic zinc exposure decreases the surface expression of NR2A-containing NMDA receptors in cultured hippocampal neurons.

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Jia Zhu

Full Text Available Zinc distributes widely in the central nervous system, especially in the hippocampus, amygdala and cortex. The dynamic balance of zinc is critical for neuronal functions. Zinc modulates the activity of N-methyl-D-aspartate receptors (NMDARs through the direct inhibition and various intracellular signaling pathways. Abnormal NMDAR activities have been implicated in the aetiology of many brain diseases. Sustained zinc accumulation in the extracellular fluid is known to link to pathological conditions. However, the mechanism linking this chronic zinc exposure and NMDAR dysfunction is poorly understood.We reported that chronic zinc exposure reduced the numbers of NR1 and NR2A clusters in cultured hippocampal pyramidal neurons. Whole-cell and synaptic NR2A-mediated currents also decreased. By contrast, zinc did not affect NR2B, suggesting that chronic zinc exposure specifically influences NR2A-containg NMDARs. Surface biotinylation indicated that zinc exposure attenuated the membrane expression of NR1 and NR2A, which might arise from to the dissociation of the NR2A-PSD-95-Src complex.Chronic zinc exposure perturbs the interaction of NR2A to PSD-95 and causes the disorder of NMDARs in hippocampal neurons, suggesting a novel action of zinc distinct from its acute effects on NMDAR activity.
Radiation-associated breast tumors display a distinct gene expression profile

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Broeks, Annegien; Braaf, Linde M; Wessels, Lodewyk F A

2010-01-01

PURPOSE: Women who received irradiation for Hodgkin's lymphoma have a strong increased risk for developing breast cancer. Approximately 90% of the breast cancers in these patients can be attributed to their radiation treatment, rendering such series extremely useful to determine whether a common...... radiation-associated cause underlies the carcinogenic process. METHODS AND MATERIALS: In this study we used gene expression profiling technology to assess gene expression changes in radiation-associated breast tumors compared with a set of control breast tumors of women unexposed to radiation, diagnosed...... at the same age. RNA was obtained from fresh frozen tissue samples from 22 patients who developed breast cancer after Hodgkin's lymphoma (BfHL) and from 20 control breast tumors. RESULTS: Unsupervised hierarchical clustering of the profile data resulted in a clustering of the radiation-associated tumors...
The expressive stance: intentionality, expression, and machine art

OpenAIRE

Linson, Adam

2013-01-01

This paper proposes a new interpretive stance for interpreting artistic works and performances that is relevant to artificial intelligence research but also has broader implications. Termed the expressive stance, this stance makes intelligible a critical distinction between present-day machine art and human art, but allows for the possibility that future machine art could find a place alongside our own. The expressive stance is elaborated as a response to Daniel Dennett's notion of the intent...
Cell surface carbohydrates as prognostic markers in human carcinomas

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Dabelsteen, Erik

1996-01-01

Tumour development is usually associated with changes in cell surface carbohydrates. These are often divided into changes related to terminal carbohydrate structures, which include incomplete synthesis and modification of normally existing carbohydrates, and changes in the carbohydrate core...... structure. The latter includes chain elongation of both glycolipids and proteins, increased branching of carbohydrates in N-linked glycoproteins, and blocked synthesis of carbohydrates in O-linked mucin-like glycoproteins. In mature organisms, expression of distinct carbohydrates is restricted to specific...... cell types; within a given tissue, variation in expression may be related to cell maturation. Tumour-associated carbohydrate structures often reflect a certain stage of cellular development; most of these moieties are structures normally found in other adult or embryonic tissues. There is no unique...
Gene expression profiling supports the hypothesis that human ovarian surface epithelia are multipotent and capable of serving as ovarian cancer initiating cells

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Matyunina Lilya V

2009-12-01

Full Text Available Abstract Background Accumulating evidence suggests that somatic stem cells undergo mutagenic transformation into cancer initiating cells. The serous subtype of ovarian adenocarcinoma in humans has been hypothesized to arise from at least two possible classes of progenitor cells: the ovarian surface epithelia (OSE and/or an as yet undefined class of progenitor cells residing in the distal end of the fallopian tube. Methods Comparative gene expression profiling analyses were carried out on OSE removed from the surface of normal human ovaries and ovarian cancer epithelial cells (CEPI isolated by laser capture micro-dissection (LCM from human serous papillary ovarian adenocarcinomas. The results of the gene expression analyses were randomly confirmed in paraffin embedded tissues from ovarian adenocarcinoma of serous subtype and non-neoplastic ovarian tissues using immunohistochemistry. Differentially expressed genes were analyzed using gene ontology, molecular pathway, and gene set enrichment analysis algorithms. Results Consistent with multipotent capacity, genes in pathways previously associated with adult stem cell maintenance are highly expressed in ovarian surface epithelia and are not expressed or expressed at very low levels in serous ovarian adenocarcinoma. Among the over 2000 genes that are significantly differentially expressed, a number of pathways and novel pathway interactions are identified that may contribute to ovarian adenocarcinoma development. Conclusions Our results are consistent with the hypothesis that human ovarian surface epithelia are multipotent and capable of serving as the origin of ovarian adenocarcinoma. While our findings do not rule out the possibility that ovarian cancers may also arise from other sources, they are inconsistent with claims that ovarian surface epithelia cannot serve as the origin of ovarian cancer initiating cells.
Mendelian and non-mendelian mutations affecting surface antigen expression in Paramecium tetraurelia

International Nuclear Information System (INIS)

Epstein, L.M.; Forney, J.D.

1984-01-01

A screening procedure was devised for the isolation of X-ray-induced mutations affecting the expression of the A immobilization antigen (i-antigen) in Paramecium tetraurelia. Two of the mutations isolated by this procedure proved to be in modifier genes. The two genes are unlinked to each other and unlinked to the structural A i-antigen gene. These are the first modifier genes identified in a Paramecium sp. that affect surface antigen expression. Another mutation was found to be a deletion of sequences just downstream from the A i-antigen gene. In cells carrying this mutation, the A i-antigen gene lies in close proximity to the end of a macronuclear chromosome. The expression of the A i-antigen is not affected in these cells, demonstrating that downstream sequences are not important for the regulation and expression of the A i-antigen gene. A stable cell line was also recovered which shows non-Mendelian inheritance of a macronuclear deletion of the A i-antigen gene. This mutant does not contain the gene in its macronucleus, but contains a complete copy of the gene in its micronucleus. In the cytoplasm of wild-type animals, the micronuclear gene is included in the developing macronucleus; in the cytoplasm of the mutant, the incorporation of the A i-antigen gene into the macronucleus is inhibited. This is the first evidence that a mechanism is available in ciliates to control the expression of a gene by regulating its incorporation into developing macronuclei
Natural Microbial Assemblages Reflect Distinct Organismal and Functional Partitioning

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Wilmes, P.; Andersson, A.; Kalnejais, L. H.; Verberkmoes, N. C.; Lefsrud, M. G.; Wexler, M.; Singer, S. W.; Shah, M.; Bond, P. L.; Thelen, M. P.; Hettich, R. L.; Banfield, J. F.

2007-12-01

The ability to link microbial community structure to function has long been a primary focus of environmental microbiology. With the advent of community genomic and proteomic techniques, along with advances in microscopic imaging techniques, it is now possible to gain insights into the organismal and functional makeup of microbial communities. Biofilms growing within highly acidic solutions inside the Richmond Mine (Iron Mountain, Redding, California) exhibit distinct macro- and microscopic morphologies. They are composed of microorganisms belonging to the three domains of life, including archaea, bacteria and eukarya. The proportion of each organismal type depends on sampling location and developmental stage. For example, mature biofilms floating on top of acid mine drainage (AMD) pools exhibit layers consisting of a densely packed bottom layer of the chemoautolithotroph Leptospirillum group II, a less dense top layer composed mainly of archaea, and fungal filaments spanning across the entire biofilm. The expression of cytochrome 579 (the most highly abundant protein in the biofilm, believed to be central to iron oxidation and encoded by Leptospirillum group II) is localized at the interface of the biofilm with the AMD solution, highlighting that biofilm architecture is reflected at the functional gene expression level. Distinct functional partitioning is also apparent in a biological wastewater treatment system that selects for distinct polyphosphate accumulating organisms. Community genomic data from " Candidatus Accumulibacter phosphatis" dominated activated sludge has enabled high mass-accuracy shotgun proteomics for identification of key metabolic pathways. Comprehensive genome-wide alignment of orthologous proteins suggests distinct partitioning of protein variants involved in both core-metabolism and specific metabolic pathways among the dominant population and closely related species. In addition, strain- resolved proteogenomic analysis of the AMD biofilms
Viral DNA Sensors IFI16 and Cyclic GMP-AMP Synthase Possess Distinct Functions in Regulating Viral Gene Expression, Immune Defenses, and Apoptotic Responses during Herpesvirus Infection.

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Diner, Benjamin A; Lum, Krystal K; Toettcher, Jared E; Cristea, Ileana M

2016-11-15

The human interferon-inducible protein IFI16 is an important antiviral factor that binds nuclear viral DNA and promotes antiviral responses. Here, we define IFI16 dynamics in space and time and its distinct functions from the DNA sensor cyclic dinucleotide GMP-AMP synthase (cGAS). Live-cell imaging reveals a multiphasic IFI16 redistribution, first to viral entry sites at the nuclear periphery and then to nucleoplasmic puncta upon herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV) infections. Optogenetics and live-cell microscopy establish the IFI16 pyrin domain as required for nuclear periphery localization and oligomerization. Furthermore, using proteomics, we define the signature protein interactions of the IFI16 pyrin and HIN200 domains and demonstrate the necessity of pyrin for IFI16 interactions with antiviral proteins PML and cGAS. We probe signaling pathways engaged by IFI16, cGAS, and PML using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-mediated knockouts in primary fibroblasts. While IFI16 induces cytokines, only cGAS activates STING/TBK-1/IRF3 and apoptotic responses upon HSV-1 and HCMV infections. cGAS-dependent apoptosis upon DNA stimulation requires both the enzymatic production of cyclic dinucleotides and STING. We show that IFI16, not cGAS or PML, represses HSV-1 gene expression, reducing virus titers. This indicates that regulation of viral gene expression may function as a greater barrier to viral replication than the induction of antiviral cytokines. Altogether, our findings establish coordinated and distinct antiviral functions for IFI16 and cGAS against herpesviruses. How mammalian cells detect and respond to DNA viruses that replicate in the nucleus is poorly understood. Here, we decipher the distinct functions of two viral DNA sensors, IFI16 and cGAS, during active immune signaling upon infection with two herpesviruses, herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV). We show that IFI16
Autophagosome Proteins LC3A, LC3B and LC3C Have Distinct Subcellular Distribution Kinetics and Expression in Cancer Cell Lines.

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Michael I Koukourakis

Full Text Available LC3s (MAP1-LC3A, B and C are structural proteins of autophagosomal membranes, widely used as biomarkers of autophagy. Whether these three LC3 proteins have a similar biological role in autophagy remains obscure. We examine in parallel the subcellular expression patterns of the three LC3 proteins in a panel of human cancer cell lines, as well as in normal MRC5 fibroblasts and HUVEC, using confocal microscopy and western blot analysis of cell fractions. In the cytoplasm, there was a minimal co-localization between LC3A, B and C staining, suggesting that the relevant autophagosomes are formed by only one out of the three LC3 proteins. LC3A showed a perinuclear and nuclear localization, while LC3B was equally distributed throughout the cytoplasm and localized in the nucleolar regions. LC3C was located in the cytoplasm and strongly in the nuclei (excluding nucleoli, where it extensively co-localized with the LC3A and the Beclin-1 autophagy initiating protein. Beclin 1 is known to contain a nuclear trafficking signal. Blocking nuclear export function by Leptomycin B resulted in nuclear accumulation of all LC3 and Beclin-1 proteins, while Ivermectin that blocks nuclear import showed reduction of accumulation, but not in all cell lines. Since endogenous LC3 proteins are used as major markers of autophagy in clinical studies and cell lines, it is essential to check the specificity of the antibodies used, as the kinetics of these molecules are not identical and may have distinct biological roles. The distinct subcellular expression patterns of LC3s provide a basis for further studies.
Translational analysis of mouse and human placental protein and mRNA reveals distinct molecular pathologies in human preeclampsia.

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Cox, Brian; Sharma, Parveen; Evangelou, Andreas I; Whiteley, Kathie; Ignatchenko, Vladimir; Ignatchenko, Alex; Baczyk, Dora; Czikk, Marie; Kingdom, John; Rossant, Janet; Gramolini, Anthony O; Adamson, S Lee; Kislinger, Thomas

2011-12-01

Preeclampsia (PE) adversely impacts ~5% of pregnancies. Despite extensive research, no consistent biomarkers or cures have emerged, suggesting that different molecular mechanisms may cause clinically similar disease. To address this, we undertook a proteomics study with three main goals: (1) to identify a panel of cell surface markers that distinguish the trophoblast and endothelial cells of the placenta in the mouse; (2) to translate this marker set to human via the Human Protein Atlas database; and (3) to utilize the validated human trophoblast markers to identify subgroups of human preeclampsia. To achieve these goals, plasma membrane proteins at the blood tissue interfaces were extracted from placentas using intravascular silica-bead perfusion, and then identified using shotgun proteomics. We identified 1181 plasma membrane proteins, of which 171 were enriched at the maternal blood-trophoblast interface and 192 at the fetal endothelial interface with a 70% conservation of expression in humans. Three distinct molecular subgroups of human preeclampsia were identified in existing human microarray data by using expression patterns of trophoblast-enriched proteins. Analysis of all misexpressed genes revealed divergent dysfunctions including angiogenesis (subgroup 1), MAPK signaling (subgroup 2), and hormone biosynthesis and metabolism (subgroup 3). Subgroup 2 lacked expected changes in known preeclampsia markers (sFLT1, sENG) and uniquely overexpressed GNA12. In an independent set of 40 banked placental specimens, GNA12 was overexpressed during preeclampsia when co-incident with chronic hypertension. In the current study we used a novel translational analysis to integrate mouse and human trophoblast protein expression with human microarray data. This strategy identified distinct molecular pathologies in human preeclampsia. We conclude that clinically similar preeclampsia patients exhibit divergent placental gene expression profiles thus implicating divergent
The Staphylococcus aureus Global Regulator MgrA Modulates Clumping and Virulence by Controlling Surface Protein Expression.

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Heidi A Crosby

2016-05-01

Full Text Available Staphylococcus aureus is a human commensal and opportunistic pathogen that causes devastating infections in a wide range of locations within the body. One of the defining characteristics of S. aureus is its ability to form clumps in the presence of soluble fibrinogen, which likely has a protective benefit and facilitates adhesion to host tissue. We have previously shown that the ArlRS two-component regulatory system controls clumping, in part by repressing production of the large surface protein Ebh. In this work we show that ArlRS does not directly regulate Ebh, but instead ArlRS activates expression of the global regulator MgrA. Strains lacking mgrA fail to clump in the presence of fibrinogen, and clumping can be restored to an arlRS mutant by overexpressing either arlRS or mgrA, indicating that ArlRS and MgrA constitute a regulatory pathway. We used RNA-seq to show that MgrA represses ebh, as well as seven cell wall-associated proteins (SraP, Spa, FnbB, SasG, SasC, FmtB, and SdrD. EMSA analysis showed that MgrA directly represses expression of ebh and sraP. Clumping can be restored to an mgrA mutant by deleting the genes for Ebh, SraP and SasG, suggesting that increased expression of these proteins blocks clumping by steric hindrance. We show that mgrA mutants are less virulent in a rabbit model of endocarditis, and virulence can be partially restored by deleting the genes for the surface proteins ebh, sraP, and sasG. While mgrA mutants are unable to clump, they are known to have enhanced biofilm capacity. We demonstrate that this increase in biofilm formation is partially due to up-regulation of SasG, a surface protein known to promote intercellular interactions. These results confirm that ArlRS and MgrA constitute a regulatory cascade, and that they control expression of a number of genes important for virulence, including those for eight large surface proteins.
Functional Elements on SIRPα IgV domain Mediate Cell Surface Binding to CD47

OpenAIRE

Liu, Yuan; Tong, Qiao; Zhou, Yubin; Lee, Hsiau-Wei; Yang, Jenny J.; Bühring, Hans-Jörg; Chen, Yi-Tien; Ha, Binh; Chen, Celia X-J.; Zen, Ke

2006-01-01

SIRPα and SIRPβ1, the two major isoforms of the signal regulatory protein (SIRP) family, are co-expressed in human leukocytes but mediate distinct extracellular binding interactions and divergent cell signaling responses. Previous studies have demonstrated that binding of SIRPα with CD47, another important cell surface molecule, through the extracellular IgV domain regulates important leukocyte functions including macrophage recognition, leukocyte adhesion and transmigration. Although SIRPβ1 ...
Identification of an evolutionarily conserved extracellular threonine residue critical for surface expression and its potential coupling of adjacent voltage-sensing and gating domains in voltage-gated potassium channels.

Science.gov (United States)

Mckeown, Lynn; Burnham, Matthew P; Hodson, Charlotte; Jones, Owen T

2008-10-31

The dynamic expression of voltage-gated potassium channels (Kvs) at the cell surface is a fundamental factor controlling membrane excitability. In exploring possible mechanisms controlling Kv surface expression, we identified a region in the extracellular linker between the first and second of the six (S1-S6) transmembrane-spanning domains of the Kv1.4 channel, which we hypothesized to be critical for its biogenesis. Using immunofluorescence microscopy, flow cytometry, patch clamp electrophysiology, and mutagenesis, we identified a single threonine residue at position 330 within the Kv1.4 S1-S2 linker that is absolutely required for cell surface expression. Mutation of Thr-330 to an alanine, aspartate, or lysine prevented surface expression. However, surface expression occurred upon co-expression of mutant and wild type Kv1.4 subunits or mutation of Thr-330 to a serine. Mutation of the corresponding residue (Thr-211) in Kv3.1 to alanine also caused intracellular retention, suggesting that the conserved threonine plays a generalized role in surface expression. In support of this idea, sequence comparisons showed conservation of the critical threonine in all Kv families and in organisms across the evolutionary spectrum. Based upon the Kv1.2 crystal structure, further mutagenesis, and the partial restoration of surface expression in an electrostatic T330K bridging mutant, we suggest that Thr-330 hydrogen bonds to equally conserved outer pore residues, which may include a glutamate at position 502 that is also critical for surface expression. We propose that Thr-330 serves to interlock the voltage-sensing and gating domains of adjacent monomers, thereby yielding a structure competent for the surface expression of functional tetramers.
Expression and immunological characterisation of Eimeria tenella glycosylphosphatidylinositol-anchored surface antigen-5

Science.gov (United States)

Ho, Sue-Kim; Nathan, Sheila; Wan, Kiew-Lian

2016-11-01

Eimeria tenella is the most pathogenic of the Eimeria species that infect chickens and causes huge economic losses to the poultry industry. The glycosylphosphatidylinositol-anchored surface antigen-5 (SAG5) found on the surface of the parasite has been shown to activate the chicken's immune system. In this study, recombinant SAG5 was expressed, purified and used to investigate the immune-inducing characteristics of the molecule. Chickens were immunized with purified recombinant SAG5 and sera were subjected to Enzyme-linked Immunosorbant Assay (ELISA). Results indicated that specific antibodies against rSAG5 were produced, with IgG detected at a higher level compared to IgA and IgM. Information on the immunological responses elicited by SAG5 provides essential knowledge that will contribute towards the effort to develop more effective strategies against coccidiosis.
A comparative study of small RNAs in Toxoplasma gondii of distinct genotypes

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Wang Jielin

2012-09-01

Full Text Available Abstract Background Toxoplasma gondii is an intracellular parasite with a significant impact on human health. Inside the mammalian and avian hosts, the parasite can undergo rapid development or remain inactive in the cysts. The mechanism that regulates parasite proliferation has not been fully understood. Small noncoding RNAs (sncRNA such as microRNAs (miRNAs are endogenous regulatory factors that can modulate cell differentiation and development. It is anticipated that hundreds of miRNAs regulate the expression of thousands of genes in a single organism. SncRNAs have been identified in T. gondii, however the profiles of sncRNAs expression and their potential regulatory function in parasites of distinct genotypes has largely been unknown. Methods The transcription profiles of miRNAs in the two genetically distinct strains, RH and ME49, of T. gondii were investigated and compared by a high-through-put RNA sequencing technique and systematic bioinformatics analysis. The expression of some of the miRNAs was confirmed by Northern blot analysis. Results 1,083,320 unique sequences were obtained. Of which, 17 conserved miRNAs related to 2 metazoan miRNA families and 339 novel miRNAs were identified. A total of 175 miRNAs showed strain-specific expression, of which 155 miRNAs were up-regulated in RH strain and 20 miRNAs were up-regulated in ME49 strain. Strain-specific expression of miRNAs in T. gondii could be due to activation of specific genes at different genomic loci or due to arm-switching of the same pre-miRNA duplex. Conclusions Evidence for the differential expression of miRNAs in the two genetically distinct strains of T. gondii has been identified and defined. MiRNAs of T. gondii are more species-specific as compared to other organisms, which can be developed as diagnostic biomarkers for toxoplasmosis. The data also provide a framework for future studies on RNAi-dependent regulatory mechanisms in the zoonotic parasite.
False memories in children and adults: age, distinctiveness, and subjective experience.

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Ghetti, Simona; Qin, Jianjian; Goodman, Gail S

2002-09-01

This study investigated developmental trends associated with the Deese/Roediger-McDermott false-memory effect, the role of distinctive information in false-memory formation, and participants' subjective experience of true and false memories. Children (5- and 7-year-olds) and adults studied lists of semantically associated words. Half of the participants studied words alone, and half studied words accompanied by pictures. There were significant age differences in recall (5-year-olds evinced more false memories than did adults) but not in recognition of critical lures. Distinctive information reduced false memory for all age groups. Younger children provided with distinctive information, and older children and adults regardless of whether they viewed distinctive information, expressed higher levels of confidence in true than in false memories. Source attributions did not significantly differ between true and false memories. Implications for theories of false memory and memory development are discussed.

Generation of functional scFv intrabody to abate the expression of CD147 surface molecule of 293A cells

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Mai Sabine

2008-01-01

Full Text Available Abstract Background Expression of intracellular antibodies (intrabodies has become a broadly applicable technology for generation of phenotypic knockouts in vivo. The method uses surface depletion of cellular membrane proteins to examine their biological function. In this study, we used this strategy to block the transport of cell surface molecule CD147 to the cell membrane. Phage display technology was introduced to generate the functional antibody fragment to CD147, and we subsequently constructed a CD147-specific scFv that was expressed intracellularly and retained in the endoplasmic reticulum by adenoviral gene transfer. Results The recombinant antibody fragments, Fab and scFv, of the murine monoclonal antibody (clone M6-1B9 reacted specifically to CD147 by indirect enzyme-linked immunosorbent assays (ELISA using a recombinant CD147-BCCP as a target. This indicated that the Fab- and scFv-M6-1B9 displaying on phage surfaces were correctly folded and functionally active. We subsequently constructed a CD147-specific scFv, scFv-M6-1B9-intrabody, in 293A cells. The expression of CD147 on 293A cell surface was monitored at 36 h after transduction by flow cytometry and demonstrated remarkable reduction. Colocalization of scFv-M6-1B9 intrabody with CD147 in the ER network was depicted using a 3D deconvolution microscopy system. Conclusion The results suggest that our approach can generate antibody fragments suitable for decreasing the expression of CD147 on 293A cells. This study represents a step toward understanding the role of the cell surface protein, CD147.
The amygdalo-motor pathway and the control of facial expressions

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Katalin M Gothard

2014-03-01

Full Text Available Facial expressions reflect decisions about the perceived meaning of social stimuli emitted by others and the expected socio-emotional outcome of the reciprocating expression. The decision to produce a facial expression emerges from the joint activity of a network of structures that include the amygdala and multiple, interconnected cortical and subcortical motor areas. Reciprocal transformations between sensory and motor signals give rise to distinct brain states that promote, or impede the production of facial expressions. The muscles of the upper and lower face are controlled by anatomically distinct motor areas and thus require distinct patterns of motor commands. Concomitantly multiple areas, including the amygdala, monitor the ongoing overt behavior (the expression of self and the covert, autonomic responses that accompany emotional expressions. Interoceptive signals and visceral states, therefore, should be incorporated into the formalisms of decision making in order account for decisions that govern the receiving-emitting cycle of facial expressions.
Cell Surface Trafficking of TLR1 Is Differentially Regulated by the Chaperones PRAT4A and PRAT4B*

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Hart, Bryan E.; Tapping, Richard I.

2012-01-01

The subcellular localization of Toll-like receptors (TLRs) is critical to their ability to function as innate immune sensors of microbial infection. We previously reported that an I602S polymorphism of human TLR1 is associated with aberrant trafficking of the receptor to the cell surface, loss of responses to TLR1 agonists, and differential susceptibility to diseases caused by pathogenic mycobacteria. Through an extensive analysis of receptor deletion and point mutants we have discovered that position 602 resides within a short 6 amino acid cytoplasmic region that is required for TLR1 surface expression. This short trafficking motif, in conjunction with the adjacent transmembrane domain, is sufficient to direct TLR1 to the cell surface. A serine at position 602 interrupts this trafficking motif and prevents cell surface expression of TLR1. Additionally, we have found that ER-resident TLR chaperones, PRAT4A and PRAT4B, act as positive and negative regulators of TLR1 surface trafficking, respectively. Importantly, either over-expression of PRAT4A or knock-down of PRAT4B rescues cell surface expression of the TLR1 602S variant. We also report that IFN-γ treatment of primary human monocytes derived from homozygous 602S individuals rescues TLR1 cell surface trafficking and cellular responses to soluble agonists. This event appears to be mediated by PRAT4A whose expression is strongly induced in human monocytes by IFN-γ. Collectively, these results provide a mechanism for the differential trafficking of TLR1 I602S variants, and highlight the distinct roles for PRAT4A and PRAT4B in the regulation of TLR1 surface expression. PMID:22447933
Temporomandibular joint formation requires two distinct hedgehog-dependent steps.

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Purcell, Patricia; Joo, Brian W; Hu, Jimmy K; Tran, Pamela V; Calicchio, Monica L; O'Connell, Daniel J; Maas, Richard L; Tabin, Clifford J

2009-10-27

We conducted a genetic analysis of the developing temporo-mandibular or temporomandi-bular joint (TMJ), a highly specialized synovial joint that permits movement and function of the mammalian jaw. First, we used laser capture microdissection to perform a genome-wide expression analysis of each of its developing components. The expression patterns of genes identified in this screen were examined in the TMJ and compared with those of other synovial joints, including the shoulder and the hip joints. Striking differences were noted, indicating that the TMJ forms via a distinct molecular program. Several components of the hedgehog (Hh) signaling pathway are among the genes identified in the screen, including Gli2, which is expressed specifically in the condyle and in the disk of the developing TMJ. We found that mice deficient in Gli2 display aberrant TMJ development such that the condyle loses its growth-plate-like cellular organization and no disk is formed. In addition, we used a conditional strategy to remove Smo, a positive effector of the Hh signaling pathway, from chondrocyte progenitors. This cell autonomous loss of Hh signaling allows for disk formation, but the resulting structure fails to separate from the condyle. Thus, these experiments establish that Hh signaling acts at two distinct steps in disk morphogenesis, condyle initiation, and disk-condyle separation and provide a molecular framework for future studies of the TMJ.
SRC Inhibition Reduces NR2B Surface Expression and Synaptic Plasticity in the Amygdala

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Sinai, Laleh; Duffy, Steven; Roder, John C.

2010-01-01

The Src protein tyrosine kinase plays a central role in the regulation of N-methyl-d-aspartate receptor (NMDAR) activity by regulating NMDAR subunit 2B (NR2B) surface expression. In the amygdala, NMDA-dependent synaptic plasticity resulting from convergent somatosensory and auditory inputs contributes to emotional memory; however, the role of Src…
Inflammation promotes oral squamous carcinoma immune evasion via induced programmed death ligand-1 surface expression.

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Lu, Wanlu; Lu, Libing; Feng, Yun; Chen, Jiao; Li, Yan; Kong, Xiangli; Chen, Sixiu; Li, Xiaoyu; Chen, Qianming; Zhang, Ping

2013-05-01

The association between inflammation and cancer provides a new target for tumor biotherapy. The inflammatory cells and molecules within the tumor microenvironment have decisive dual roles in antitumor immunity and immune evasion. In the present study, phytohemagglutinin (PHA) was used to stimulate peripheral blood mononuclear cells (PBMCs) to simulate the tumor inflammatory microenvironment. The effect of immune cells and inflammatory cytokines on the surface expression of programmed cell death-1 ligand 1 (PD-L1) and tumor immune evasion was investigated using flow cytometry (FCM) and an in vivo xenotransplantation model. Based on the data, PHA-activated, but not resting, immune cells were able to promote the surface expression of PD-L1 in Tca8113 oral squamous carcinoma cells via the secretion of inflammatory cytokines, but not by cell-cell contact. The majority of the inflammatory cytokines had no significant effect on the proliferation, cell cycle progression and apoptosis of the Tca8113 cells, although they each induced the expression of PD-L1 in a dose-dependent manner. In total, 99% of the Tca8113 cells expressed PD-L1 following treatment with the supernatant of PHA-stimulated PBMCs. The PHA-supernatant pretreated Tca8113 cells unusually induced Tca8113 antigen-specific CD8 + T cell apoptosis in vitro and the evasion of antigen-specific T cell attraction in a nude mouse tumor-bearing model. These results indicate a new mechanism for the promotion of tumor immune evasion by the tumor inflammatory microenvironment.
Surface TRAIL decoy receptor-4 expression is correlated with TRAIL resistance in MCF7 breast cancer cells

International Nuclear Information System (INIS)

Sanlioglu, Ahter D; Dirice, Ercument; Aydin, Cigdem; Erin, Nuray; Koksoy, Sadi; Sanlioglu, Salih

2005-01-01

Tumor Necrosis Factor (TNF)-Related Apoptosis-Inducing Ligand (TRAIL) selectively induces apoptosis in cancer cells but not in normal cells. Despite this promising feature, TRAIL resistance observed in cancer cells seriously challenged the use of TRAIL as a death ligand in gene therapy. The current dispute concerns whether or not TRAIL receptor expression pattern is the primary determinant of TRAIL sensitivity in cancer cells. This study investigates TRAIL receptor expression pattern and its connection to TRAIL resistance in breast cancer cells. In addition, a DcR2 siRNA approach and a complementary gene therapy modality involving IKK inhibition (AdIKKβKA) were also tested to verify if these approaches could sensitize MCF7 breast cancer cells to adenovirus delivery of TRAIL (Ad5hTRAIL). TRAIL sensitivity assays were conducted using Molecular Probe's Live/Dead Cellular Viability/Cytotoxicity Kit following the infection of breast cancer cells with Ad5hTRAIL. The molecular mechanism of TRAIL induced cell death under the setting of IKK inhibition was revealed by Annexin V binding. Novel quantitative Real Time RT-PCR and flow cytometry analysis were performed to disclose TRAIL receptor composition in breast cancer cells. MCF7 but not MDA-MB-231 breast cancer cells displayed strong resistance to adenovirus delivery of TRAIL. Only the combinatorial use of Ad5hTRAIL and AdIKKβKA infection sensitized MCF7 breast cancer cells to TRAIL induced cell death. Moreover, novel quantitative Real Time RT-PCR assays suggested that while the level of TRAIL Decoy Receptor-4 (TRAIL-R4) expression was the highest in MCF7 cells, it was the lowest TRAIL receptor expressed in MDA-MB-231 cells. In addition, conventional flow cytometry analysis demonstrated that TRAIL resistant MCF7 cells exhibited substantial levels of TRAIL-R4 expression but not TRAIL decoy receptor-3 (TRAIL-R3) on surface. On the contrary, TRAIL sensitive MDA-MB-231 cells displayed very low levels of surface TRAIL-R4
Plasmodium falciparum parasites expressing pregnancy-specific variant surface antigens adhere strongly to the choriocarcinoma cell line BeWo

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Haase, Rikke N; Megnekou, Rosette; Lundquist, Maja

2006-01-01

Placenta-sequestering Plasmodium falciparum parasites causing pregnancy-associated malaria express pregnancy-specific variant surface antigens (VSA(PAM)). We report here that VSA(PAM)-expressing patient isolates adhere strongly to the choriocarcinoma cell line BeWo and that the BeWo line can...... be used to efficiently select for VSA(PAM) expression in vitro....
Identification of a regulatory T cell specific cell surface molecule that mediates suppressive signals and induces Foxp3 expression.

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Wang, Rui; Wan, Qi; Kozhaya, Lina; Fujii, Hodaka; Unutmaz, Derya

2008-07-16

Regulatory T (T(reg)) cells control immune activation and maintain tolerance. How T(regs) mediate their suppressive function is unclear. Here we identified a cell surface molecule, called GARP, (or LRRC32), which within T cells is specifically expressed in T(regs) activated through the T cell receptor (TCR). Ectopic expression of GARP in human naïve T (T(N)) cells inhibited their proliferation and cytokine secretion upon TCR activation. Remarkably, GARP over-expression in T(N) cells induced expression of T(reg) master transcription factor Foxp3 and endowed them with a partial suppressive function. The extracellular but not the cytoplasmic region of GARP, was necessary for these functions. Silencing Foxp3 in human T(reg) cells reduced expression of GARP and attenuated their suppressive function. However, GARP function was not affected when Foxp3 was downregulated in GARP-overexpressing cells, while silencing GARP in Foxp3-overexpressing cells reduced their suppressive activity. These findings reveal a novel cell surface molecule-mediated regulatory mechanism, with implications for modulating aberrant immune responses.
Identification of a regulatory T cell specific cell surface molecule that mediates suppressive signals and induces Foxp3 expression.

Directory of Open Access Journals (Sweden)

Rui Wang

2008-07-01

Full Text Available Regulatory T (T(reg cells control immune activation and maintain tolerance. How T(regs mediate their suppressive function is unclear. Here we identified a cell surface molecule, called GARP, (or LRRC32, which within T cells is specifically expressed in T(regs activated through the T cell receptor (TCR. Ectopic expression of GARP in human naïve T (T(N cells inhibited their proliferation and cytokine secretion upon TCR activation. Remarkably, GARP over-expression in T(N cells induced expression of T(reg master transcription factor Foxp3 and endowed them with a partial suppressive function. The extracellular but not the cytoplasmic region of GARP, was necessary for these functions. Silencing Foxp3 in human T(reg cells reduced expression of GARP and attenuated their suppressive function. However, GARP function was not affected when Foxp3 was downregulated in GARP-overexpressing cells, while silencing GARP in Foxp3-overexpressing cells reduced their suppressive activity. These findings reveal a novel cell surface molecule-mediated regulatory mechanism, with implications for modulating aberrant immune responses.
The neural signatures of distinct psychopathic traits.

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Carré, Justin M; Hyde, Luke W; Neumann, Craig S; Viding, Essi; Hariri, Ahmad R

2013-01-01

Recent studies suggest that psychopathy may be associated with dysfunction in the neural circuitry supporting both threat- and reward-related processes. However, these studies have involved small samples and often focused on extreme groups. Thus, it is unclear to what extent current findings may generalize to psychopathic traits in the general population. Furthermore, no studies have systematically and simultaneously assessed associations between distinct psychopathy facets and both threat- and reward-related brain function in the same sample of participants. Here, we examined the relationship between threat-related amygdala reactivity and reward-related ventral striatum (VS) reactivity and variation in four facets of self-reported psychopathy in a sample of 200 young adults. Path models indicated that amygdala reactivity to fearful facial expressions is negatively associated with the interpersonal facet of psychopathy, whereas amygdala reactivity to angry facial expressions is positively associated with the lifestyle facet. Furthermore, these models revealed that differential VS reactivity to positive versus negative feedback is negatively associated with the lifestyle facet. There was suggestive evidence for gender-specific patterns of association between brain function and psychopathy facets. Our findings are the first to document differential associations between both threat- and reward-related neural processes and distinct facets of psychopathy and thus provide a more comprehensive picture of the pattern of neural vulnerabilities that may predispose to maladaptive outcomes associated with psychopathy.
Canine Distemper Virus Infects Canine Keratinocytes and Immune Cells by Using Overlapping and Distinct Regions Located on One Side of the Attachment Protein▿

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Langedijk, Johannes P. M.; Janda, Jozef; Origgi, Francesco C.; Örvell, Claes; Vandevelde, Marc; Zurbriggen, Andreas; Plattet, Philippe

2011-01-01

The morbilliviruses measles virus (MeV) and canine distemper virus (CDV) both rely on two surface glycoproteins, the attachment (H) and fusion proteins, to promote fusion activity for viral cell entry. Growing evidence suggests that morbilliviruses infect multiple cell types by binding to distinct host cell surface receptors. Currently, the only known in vivo receptor used by morbilliviruses is CD150/SLAM, a molecule expressed in certain immune cells. Here we investigated the usage of multiple receptors by the highly virulent and demyelinating CDV strain A75/17. We based our study on the assumption that CDV-H may interact with receptors similar to those for MeV, and we conducted systematic alanine-scanning mutagenesis on CDV-H throughout one side of the β-propeller documented in MeV-H to contain multiple receptor-binding sites. Functional and biochemical assays performed with SLAM-expressing cells and primary canine epithelial keratinocytes identified 11 residues mutation of which selectively abrogated fusion in keratinocytes. Among these, four were identical to amino acids identified in MeV-H as residues contacting a putative receptor expressed in polarized epithelial cells. Strikingly, when mapped on a CDV-H structural model, all residues clustered in or around a recessed groove located on one side of CDV-H. In contrast, reported CDV-H mutants with SLAM-dependent fusion deficiencies were characterized by additional impairments to the promotion of fusion in keratinocytes. Furthermore, upon transfer of residues that selectively impaired fusion induction in keratinocytes into the CDV-H of the vaccine strain, fusion remained largely unaltered. Taken together, our results suggest that a restricted region on one side of CDV-H contains distinct and overlapping sites that control functional interaction with multiple receptors. PMID:21849439
Distinct molecular subtypes of uterine leiomyosarcoma respond differently to chemotherapy treatment.

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An, Yang; Wang, Shuzhen; Li, Songlin; Zhang, Lulu; Wang, Dayong; Wang, Haojie; Zhu, Shibai; Zhu, Wan; Li, Yongqiang; Chen, Wenwu; Ji, Shaoping; Guo, Xiangqian

2017-09-11

Uterine leiomyosarcoma (ULMS) is an aggressive form of soft tissue tumors. The molecular heterogeneity and pathogenesis of ULMS are not well understood. Expression profiling data were used to determine the possibility and optimal number of ULMS molecular subtypes. Next, clinicopathological characters and molecular pathways were analyzed in each subtype to prospect the clinical applications and progression mechanisms of ULMS. Two distinct molecular subtypes of ULMS were defined based on different gene expression signatures. Subtype I ULMS recapitulated low-grade ULMS, the gene expression pattern of which resembled normal smooth muscle cells, characterized by overexpression of smooth muscle function genes such as LMOD1, SLMAP, MYLK, MYH11. In contrast, subtype II ULMS recapitulated high-grade ULMS with higher tumor weight and invasion rate, and was characterized by overexpression of genes involved in the pathway of epithelial to mesenchymal transition and tumorigenesis, such as CDK6, MAPK13 and HOXA1. We identified two distinct molecular subtypes of ULMS responding differently to chemotherapy treatment. Our findings provide a better understanding of ULMS intrinsic molecular subtypes, and will potentially facilitate the development of subtype-specific diagnosis biomarkers and therapy strategies for these tumors.
Sarcocystis neurona merozoites express a family of immunogenic surface antigens that are orthologues of the Toxoplasma gondii surface antigens (SAGs) and SAG-related sequences.

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Howe, Daniel K; Gaji, Rajshekhar Y; Mroz-Barrett, Meaghan; Gubbels, Marc-Jan; Striepen, Boris; Stamper, Shelby

2005-02-01

Sarcocystis neurona is a member of the Apicomplexa that causes myelitis and encephalitis in horses but normally cycles between the opossum and small mammals. Analysis of an S. neurona expressed sequence tag (EST) database revealed four paralogous proteins that exhibit clear homology to the family of surface antigens (SAGs) and SAG-related sequences of Toxoplasma gondii. The primary peptide sequences of the S. neurona proteins are consistent with the two-domain structure that has been described for the T. gondii SAGs, and each was predicted to have an amino-terminal signal peptide and a carboxyl-terminal glycolipid anchor addition site, suggesting surface localization. All four proteins were confirmed to be membrane associated and displayed on the surface of S. neurona merozoites. Due to their surface localization and homology to T. gondii surface antigens, these S. neurona proteins were designated SnSAG1, SnSAG2, SnSAG3, and SnSAG4. Consistent with their homology, the SnSAGs elicited a robust immune response in infected and immunized animals, and their conserved structure further suggests that the SnSAGs similarly serve as adhesins for attachment to host cells. Whether the S. neurona SAG family is as extensive as the T. gondii SAG family remains unresolved, but it is probable that additional SnSAGs will be revealed as more S. neurona ESTs are generated. The existence of an SnSAG family in S. neurona indicates that expression of multiple related surface antigens is not unique to the ubiquitous organism T. gondii. Instead, the SAG gene family is a common trait that presumably has an essential, conserved function(s).
Sarcocystis neurona Merozoites Express a Family of Immunogenic Surface Antigens That Are Orthologues of the Toxoplasma gondii Surface Antigens (SAGs) and SAG-Related Sequences†

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Howe, Daniel K.; Gaji, Rajshekhar Y.; Mroz-Barrett, Meaghan; Gubbels, Marc-Jan; Striepen, Boris; Stamper, Shelby

2005-01-01

Sarcocystis neurona is a member of the Apicomplexa that causes myelitis and encephalitis in horses but normally cycles between the opossum and small mammals. Analysis of an S. neurona expressed sequence tag (EST) database revealed four paralogous proteins that exhibit clear homology to the family of surface antigens (SAGs) and SAG-related sequences of Toxoplasma gondii. The primary peptide sequences of the S. neurona proteins are consistent with the two-domain structure that has been described for the T. gondii SAGs, and each was predicted to have an amino-terminal signal peptide and a carboxyl-terminal glycolipid anchor addition site, suggesting surface localization. All four proteins were confirmed to be membrane associated and displayed on the surface of S. neurona merozoites. Due to their surface localization and homology to T. gondii surface antigens, these S. neurona proteins were designated SnSAG1, SnSAG2, SnSAG3, and SnSAG4. Consistent with their homology, the SnSAGs elicited a robust immune response in infected and immunized animals, and their conserved structure further suggests that the SnSAGs similarly serve as adhesins for attachment to host cells. Whether the S. neurona SAG family is as extensive as the T. gondii SAG family remains unresolved, but it is probable that additional SnSAGs will be revealed as more S. neurona ESTs are generated. The existence of an SnSAG family in S. neurona indicates that expression of multiple related surface antigens is not unique to the ubiquitous organism T. gondii. Instead, the SAG gene family is a common trait that presumably has an essential, conserved function(s). PMID:15664946
Distinct mechanisms underlie adaptation of proximal tubule Na+/H+ exchanger isoform 3 in response to chronic metabolic and respiratory acidosis.

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Silva, Pedro Henrique Imenez; Girardi, Adriana Castello Costa; Neri, Elida Adalgisa; Rebouças, Nancy Amaral

2012-04-01

The Na(+/)H(+) exchanger isoform 3 (NHE3) is essential for HCO(3)(-) reabsorption in renal proximal tubules. The expression and function of NHE3 must adapt to acid-base conditions. The goal of this study was to elucidate the mechanisms responsible for higher proton secretion in proximal tubules during acidosis and to evaluate whether there are differences between metabolic and respiratory acidosis with regard to NHE3 modulation and, if so, to identify the relevant parameters that may trigger these distinct adaptive responses. We achieved metabolic acidosis by lowering HCO(3)(-) concentration in the cell culture medium and respiratory acidosis by increasing CO(2) tension in the incubator chamber. We found that cell-surface NHE3 expression was increased in response to both forms of acidosis. Mild (pH 7.21 ± 0.02) and severe (6.95 ± 0.07) metabolic acidosis increased mRNA levels, at least in part due to up-regulation of transcription, whilst mild (7.11 ± 0.03) and severe (6.86 ± 0.01) respiratory acidosis did not up-regulate NHE3 expression. Analyses of the Nhe3 promoter region suggested that the regulatory elements sensitive to metabolic acidosis are located between -466 and -153 bp, where two consensus binding sites for SP1, a transcription factor up-regulated in metabolic acidosis, were localised. We conclude that metabolic acidosis induces Nhe3 promoter activation, which results in higher mRNA and total protein level. At the plasma membrane surface, NHE3 expression was increased in metabolic and respiratory acidosis alike, suggesting that low pH is responsible for NHE3 displacement to the cell surface.
A global RNA-seq-driven analysis of CHO host and production cell lines reveals distinct differential expression patterns of genes contributing to recombinant antibody glycosylation.

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Könitzer, Jennifer D; Müller, Markus M; Leparc, Germán; Pauers, Martin; Bechmann, Jan; Schulz, Patrick; Schaub, Jochen; Enenkel, Barbara; Hildebrandt, Tobias; Hampel, Martin; Tolstrup, Anne B

2015-09-01

Boehringer Ingelheim uses two CHO-DG44 lines for manufacturing biotherapeutics, BI-HEX-1 and BI-HEX-2, which produce distinct cell type-specific antibody glycosylation patterns. A recently established CHO-K1 descended host, BI-HEX-K1, generates antibodies with glycosylation profiles differing from CHO-DG44. Manufacturing process development is significantly influenced by these unique profiles. To investigate the underlying glycosylation related gene expression, we leveraged our CHO host and production cell RNA-seqtranscriptomics and product quality database together with the CHO-K1 genome. We observed that each BI-HEX host and antibody producing cell line has a unique gene expression fingerprint. CHO-DG44 cells only transcribe Fut10, Gfpt2 and ST8Sia6 when expressing antibodies. BI-HEX-K1 cells express ST8Sia6 at host cell level. We detected a link between BI-HEX-1/BI-HEX-2 antibody galactosylation and mannosylation and the gene expression of the B4galt gene family and genes controlling mannose processing. Furthermore, we found major differences between the CHO-DG44 and CHO-K1 lineages in the expression of sialyl transferases and enzymes synthesizing sialic acid precursors, providing a rationale for the lack of immunogenic NeuGc/NGNA synthesis in CHO. Our study highlights the value of systems biotechnology to understand glycoprotein synthesis and product glycoprofiles. Such data improve future production clone selection and process development strategies for better steering of biotherapeutic product quality. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Distinct choline metabolic profiles are associated with differences in gene expression for basal-like and luminal-like breast cancer xenograft models

International Nuclear Information System (INIS)

Moestue, Siver A; Borgan, Eldrid; Huuse, Else M; Lindholm, Evita M; Sitter, Beathe; Børresen-Dale, Anne-Lise; Engebraaten, Olav; Mælandsmo, Gunhild M; Gribbestad, Ingrid S

2010-01-01

concentrations corresponded well with differences in gene expression, demonstrating distinct metabolic profiles in the xenograft models representing basal-like and luminal-like breast cancer. The same characteristics of choline metabolite profiles were also observed in patient material from ER+/PgR+ and triple-negative breast cancer, suggesting that the xenografts are relevant model systems for studies of choline metabolism in luminal-like and basal-like breast cancer
The fibroblast surface markers FAP, anti-fibroblast, and FSP are expressed by cells of epithelial origin and may be altered during epithelial-to-mesenchymal transition.

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Kahounová, Zuzana; Kurfürstová, Daniela; Bouchal, Jan; Kharaishvili, Gvantsa; Navrátil, Jiří; Remšík, Ján; Šimečková, Šárka; Študent, Vladimír; Kozubík, Alois; Souček, Karel

2017-04-06

The identification of fibroblasts and cancer-associated fibroblasts from human cancer tissue using surface markers is difficult, especially because the markers used currently are usually not expressed solely by fibroblasts, and the identification of fibroblast-specific surface molecules is still under investigation. It was aimed to compare three commercially available antibodies in the detection of different surface epitopes of fibroblasts (anti-fibroblast, fibroblast activation protein α, and fibroblast surface protein). The specificity of their expression, employing fibroblast cell lines and tumor-derived fibroblasts from breast and prostate tissues was investigated. Both the established fibroblast cell line HFF-1 and ex vivo primary fibroblasts isolated from breast and prostate cancer tissues expressed the tested surface markers to different degrees. Surprisingly, those markers were expressed also by permanent cell lines of epithelial origin, both benign and cancer-derived (breast-cell lines MCF 10A, HMLE and prostate-cell lines BPH-1, DU 145, and PC-3). The expression of fibroblast activation protein α increased on the surface of previously described models of epithelial cells undergoing epithelial-to-mesenchymal transition in response to treatment with TGF-β1. To prove the co-expression of the fibroblast markers on cells of epithelial origin, we used freshly dissociated human prostate and breast cancer tissues. The results confirmed the co-expression of anti-fibroblast and fibroblast surface protein on CD31/CD45-negative/EpCAM-positive epithelial cells. In summary, our data support the findings that the tested fibroblast markers are not fibroblast specific and may be expressed also by cells of epithelial origin (e.g., cells undergoing EMT). Therefore, the expression of these markers should be interpreted with caution, and the combination of several epitopes for both positive (anti-fibroblast or fibroblast activation protein α) and negative (Ep
Mucosal Immunogenicity of Genetically Modified Lactobacillus acidophilus Expressing an HIV-1 Epitope within the Surface Layer Protein.

Directory of Open Access Journals (Sweden)

Akinobu Kajikawa

Full Text Available Surface layer proteins of probiotic lactobacilli are theoretically efficient epitope-displaying scaffolds for oral vaccine delivery due to their high expression levels and surface localization. In this study, we constructed genetically modified Lactobacillus acidophilus strains expressing the membrane proximal external region (MPER from human immunodeficiency virus type 1 (HIV-1 within the context of the major S-layer protein, SlpA. Intragastric immunization of mice with the recombinants induced MPER-specific and S-layer protein-specific antibodies in serum and mucosal secretions. Moreover, analysis of systemic SlpA-specific cytokines revealed that the responses appeared to be Th1 and Th17 dominant. These findings demonstrated the potential use of the Lactobacillus S-layer protein for development of oral vaccines targeting specific peptides.

Site-directed mutagenesis of HIV-1 vpu gene demonstrates two clusters of replication-defective mutants with distinct ability to down-modulate cell surface CD4 and tetherin

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Masako Nomaguchi

2010-11-01

Full Text Available HIV-1 Vpu acts positively on viral infectivity by mediating CD4 degradation in endoplasmic reticulum and enhances virion release by counteracting a virion release restriction factor, tetherin. In order to define the impact of Vpu activity on HIV-1 replication, we have generated a series of site-specific proviral vpu mutants. Of fifteen mutants examined, seven exhibited a replication-defect similar to that of a vpu-deletion mutant in a lymphocyte cell line H9. These mutations clustered in narrow regions within transmembrane domain (TMD and cytoplasmic domain (CTD. Replication-defective mutants displayed the reduced ability to enhance virion release from a monolayer cell line HEp2 without exception. Upon transfection with Vpu expression vectors, neither TMD mutants nor CTD mutants blocked CD4 expression at the cell surface in another monolayer cell line MAGI. While TMD mutants were unable to down-modulate cell surface tetherin in HEp2 cells, CTD mutants did quite efficiently. Confocal microscopy analysis revealed the difference of intracellular localization between TMD and CTD mutants. In total, replication capability of HIV-1 carrying vpu mutations correlates well with the ability of Vpu to enhance virion release and to impede the cell surface expression of CD4 but not with the ability to down-modulate cell surface tetherin. Our results here suggest that efficient viral replication requires not only down-regulation of cell surface tetherin but also its degradation.
Impaired cell surface expression of HLA-B antigens on mesenchymal stem cells and muscle cell progenitors

DEFF Research Database (Denmark)

Isa, Adiba; Nehlin, Jan; Sabir, Hardee Jawad

2010-01-01

HLA class-I expression is weak in embryonic stem cells but increases rapidly during lineage progression. It is unknown whether all three classical HLA class-I antigens follow the same developmental program. In the present study, we investigated allele-specific expression of HLA-A, -B, and -C...... at the mRNA and protein levels on human mesenchymal stem cells from bone marrow and adipose tissue as well as striated muscle satellite cells and lymphocytes. Using multicolour flow cytometry, we found high cell surface expression of HLA-A on all stem cells and PBMC examined. Surprisingly, HLA-B was either...... undetectable or very weakly expressed on all stem cells protecting them from complement-dependent cytotoxicity (CDC) using relevant human anti-B and anti-Cw sera. IFNgamma stimulation for 48-72 h was required to induce full HLA-B protein expression. Quantitative real-time RT-PCR showed that IFNgamma induced...
Surface expression and limited proteolysis of ADAM10 are increased by a dominant negative inhibitor of dynamin

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Slack Barbara E

2011-05-01

Full Text Available Abstract Background The amyloid precursor protein (APP is cleaved by β- and γ-secretases to generate toxic amyloid β (Aβ peptides. Alternatively, α-secretases cleave APP within the Aβ domain, precluding Aβ formation and releasing the soluble ectodomain, sAPPα. We previously showed that inhibition of the GTPase dynamin reduced APP internalization and increased release of sAPPα, apparently by prolonging the interaction between APP and α-secretases at the plasma membrane. This was accompanied by a reduction in Aβ generation. In the present study, we investigated whether surface expression of the α-secretase ADAM (a disintegrin and metalloprotease10 is also regulated by dynamin-dependent endocytosis. Results Transfection of human embryonic kidney (HEK cells stably expressing M3 muscarinic receptors with a dominant negative dynamin I mutant (dyn I K44A, increased surface expression of both immature, and mature, catalytically active forms of co-expressed ADAM10. Surface levels of ADAM10 were unaffected by activation of protein kinase C (PKC or M3 receptors, indicating that receptor-coupled shedding of the ADAM substrate APP is unlikely to be mediated by inhibition of ADAM10 endocytosis in this cell line. Dyn I K44A strongly increased the formation of a C-terminal fragment of ADAM10, consistent with earlier reports that the ADAM10 ectodomain is itself a target for sheddases. The abundance of this fragment was increased in the presence of a γ-secretase inhibitor, but was not affected by M3 receptor activation. The dynamin mutant did not affect the distribution of ADAM10 and its C-terminal fragment between raft and non-raft membrane compartments. Conclusions Surface expression and limited proteolysis of ADAM10 are regulated by dynamin-dependent endocytosis, but are unaffected by activation of signaling pathways that upregulate shedding of ADAM substrates such as APP. Modulation of ADAM10 internalization could affect cellular behavior in two
Distinct uptake mechanisms but similar intracellular processing of two different toll-like receptor ligand-peptide conjugates in dendritic cells.

Science.gov (United States)

Khan, Selina; Bijker, Martijn S; Weterings, Jimmy J; Tanke, Hans J; Adema, Gosse J; van Hall, Thorbald; Drijfhout, Jan W; Melief, Cornelis J M; Overkleeft, Hermen S; van der Marel, Gijsbert A; Filippov, Dmitri V; van der Burg, Sjoerd H; Ossendorp, Ferry

2007-07-20

Covalent conjugation of Toll-like receptor ligands (TLR-L) to synthetic antigenic peptides strongly improves antigen presentation in vitro and T lymphocyte priming in vivo. These molecularly well defined TLR-L-peptide conjugates, constitute an attractive vaccination modality, sharing the peptide antigen and a defined adjuvant in one single molecule. We have analyzed the intracellular trafficking and processing of two TLR-L conjugates in dendritic cells (DCs). Long synthetic peptides containing an ovalbumin cytotoxic T-cell epitope were chemically conjugated to two different TLR-Ls the TLR2 ligand, Pam(3)CysSK(4) (Pam) or the TLR9 ligand CpG. Rapid and enhanced uptake of both types of TLR-L-conjugated peptide occurred in DCs. Moreover, TLR-L conjugation greatly enhanced antigen presentation, a process that was dependent on endosomal acidification, proteasomal cleavage, and TAP translocation. The uptake of the CpG approximately conjugate was independent of endosomally-expressed TLR9 as reported previously. Unexpectedly, we found that Pam approximately conjugated peptides were likewise internalized independently of the expression of cell surface-expressed TLR2. Further characterization of the uptake mechanisms revealed that TLR2-L employed a different uptake route than TLR9-L. Inhibition of clathrin- or caveolin-dependent endocytosis greatly reduced uptake and antigen presentation of the Pam-conjugate. In contrast, internalization and antigen presentation of CpG approximately conjugates was independent of clathrin-coated pits but partly dependent on caveolae formation. Importantly, in contrast to the TLR-independent uptake of the conjugates, TLR expression and downstream TLR signaling was required for dendritic cell maturation and for priming of naïve CD8(+) T-cells. Together, our data show that targeting to two distinct TLRs requires distinct uptake mechanism but follows similar trafficking and intracellular processing pathways leading to optimal antigen
Investigation of integrin expression on the surface of osteoblast-like cells by atomic force microscopy

International Nuclear Information System (INIS)

Caneva Soumetz, Federico; Saenz, Jose F.; Pastorino, Laura; Ruggiero, Carmelina; Nosi, Daniele; Raiteri, Roberto

2010-01-01

The transforming growth factor β1 (TGF-β1) is a human cytokine which has been demonstrated to modulate cell surface integrin repertoire. In this work integrin expression in response to TGF-β1 stimulation has been investigated on the surface of human osteoblast-like cells. We used atomic force microscopy (AFM) and confocal laser scanning microscopy to assess integrin expression and to evaluate their distribution over the dorsal side of the plasma membrane. AFM probes have been covalently functionalised with monoclonal antibodies specific to the β1 integrin subunit. Force curves have been collected in order to obtain maps of the interaction between the immobilized antibody and the respective cell membrane receptors. Adhesion peaks have been automatically detected by means of an ad hoc developed data analysis software. The specificity of the detected interactions has been assessed by adding free antibody in the solution and monitoring the dramatic decrease in the recorded interactions. In addition, the effect of TGF-β1 treatment on both the fluorescence signal and the adhesion events has been tested. The level of expression of the β1 integrin subunit was enhanced by TGF-β1. As a further analysis, the adhesion force of the single living cells to the substrate was measured by laterally pushing the cell with the AFM tip and measuring the force necessary to displace it. The treatment with TGF-β1 resulted in a decrease of the cell/substrate adhesion force. Results obtained by AFM have been validated by confocal laser scanning microscopy thus demonstrating the high potential of the AFM technique for the investigation of cell surface receptors distribution and trafficking at the nanoscale.
Investigation of integrin expression on the surface of osteoblast-like cells by atomic force microscopy

Energy Technology Data Exchange (ETDEWEB)

Caneva Soumetz, Federico [Department of Communication, Computer and System Sciences, University of Genova, Via Opera Pia, 13-16145 Genova (Italy); Saenz, Jose F. [Biophysical and Electronic Engineering Department, University of Genova, Via All' Opera Pia 11a, 16145 Genova (Italy); Pastorino, Laura; Ruggiero, Carmelina [Department of Communication, Computer and System Sciences, University of Genova, Via Opera Pia, 13-16145 Genova (Italy); Nosi, Daniele [Department of Anatomy, Histology and Forensic Medicine, Bio-photonic Laboratory, University of Florence, viale Morgagni, 85 Firenze, CAP 50134 Florence (Italy); Raiteri, Roberto, E-mail: rr@unige.it [Biophysical and Electronic Engineering Department, University of Genova, Via All' Opera Pia 11a, 16145 Genova (Italy)

2010-03-15

The transforming growth factor {beta}1 (TGF-{beta}1) is a human cytokine which has been demonstrated to modulate cell surface integrin repertoire. In this work integrin expression in response to TGF-{beta}1 stimulation has been investigated on the surface of human osteoblast-like cells. We used atomic force microscopy (AFM) and confocal laser scanning microscopy to assess integrin expression and to evaluate their distribution over the dorsal side of the plasma membrane. AFM probes have been covalently functionalised with monoclonal antibodies specific to the {beta}1 integrin subunit. Force curves have been collected in order to obtain maps of the interaction between the immobilized antibody and the respective cell membrane receptors. Adhesion peaks have been automatically detected by means of an ad hoc developed data analysis software. The specificity of the detected interactions has been assessed by adding free antibody in the solution and monitoring the dramatic decrease in the recorded interactions. In addition, the effect of TGF-{beta}1 treatment on both the fluorescence signal and the adhesion events has been tested. The level of expression of the {beta}1 integrin subunit was enhanced by TGF-{beta}1. As a further analysis, the adhesion force of the single living cells to the substrate was measured by laterally pushing the cell with the AFM tip and measuring the force necessary to displace it. The treatment with TGF-{beta}1 resulted in a decrease of the cell/substrate adhesion force. Results obtained by AFM have been validated by confocal laser scanning microscopy thus demonstrating the high potential of the AFM technique for the investigation of cell surface receptors distribution and trafficking at the nanoscale.
A conserved WW domain-like motif regulates invariant chain-dependent cell-surface transport of the NKG2D ligand ULBP2.

Science.gov (United States)

Uhlenbrock, Franziska; van Andel, Esther; Andresen, Lars; Skov, Søren

2015-08-01

Malignant cells expressing NKG2D ligands on their cell surface can be directly sensed and killed by NKG2D-bearing lymphocytes. To ensure this immune recognition, accumulating evidence suggests that NKG2D ligands are trafficed via alternative pathways to the cell surface. We have previously shown that the NKG2D ligand ULBP2 traffics over an invariant chain (Ii)-dependent pathway to the cell surface. This study set out to elucidate how Ii regulates ULBP2 cell-surface transport: We discovered conserved tryptophan (Trp) residues in the primary protein sequence of ULBP1-6 but not in the related MICA/B. Substitution of Trp to alanine resulted in cell-surface inhibition of ULBP2 in different cancer cell lines. Moreover, the mutated ULBP2 constructs were retained and not degraded inside the cell, indicating a crucial role of this conserved Trp-motif in trafficking. Finally, overexpression of Ii increased surface expression of wt ULBP2 while Trp-mutants could not be expressed, proposing that this Trp-motif is required for an Ii-dependent cell-surface transport of ULBP2. Aberrant soluble ULBP2 is immunosuppressive. Thus, targeting a distinct protein module on the ULBP2 sequence could counteract this abnormal expression of ULBP2. Copyright © 2015 Elsevier Ltd. All rights reserved.
Eukaryotic translation initiation factor 3 subunit e controls intracellular calcium homeostasis by regulation of cav1.2 surface expression.

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Pawel Buda

Full Text Available Inappropriate surface expression of voltage-gated Ca(2+channels (CaV in pancreatic ß-cells may contribute to the development of type 2 diabetes. First, failure to increase intracellular Ca(2+ concentrations at the sites of exocytosis impedes insulin release. Furthermore, excessive Ca(2+ influx may trigger cytotoxic effects. The regulation of surface expression of CaV channels in the pancreatic β-cells remains unknown. Here, we used real-time 3D confocal and TIRFM imaging, immunocytochemistry, cellular fractionation, immunoprecipitation and electrophysiology to study trafficking of L-type CaV1.2 channels upon β-cell stimulation. We found decreased surface expression of CaV1.2 and a corresponding reduction in L-type whole-cell Ca(2+ currents in insulin-secreting INS-1 832/13 cells upon protracted (15-30 min stimulation. This internalization occurs by clathrin-dependent endocytosis and could be prevented by microtubule or dynamin inhibitors. eIF3e (Eukaryotic translation initiation factor 3 subunit E is part of the protein translation initiation complex, but its effect on translation are modest and effects in ion channel trafficking have been suggested. The factor interacted with CaV1.2 and regulated CaV1.2 traffic bidirectionally. eIF3e silencing impaired CaV1.2 internalization, which resulted in an increased intracellular Ca(2+ load upon stimulation. These findings provide a mechanism for regulation of L-type CaV channel surface expression with consequences for β-cell calcium homeostasis, which will affect pancreatic β-cell function and insulin production.
Surface Expression of TGF-β Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer

OpenAIRE

Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-01-01

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGF-β which is expressed naturally by platelets and regulatory T cells. Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGF-β in the tumor microenvironment. We found that human breast, lung and colon cancers expres...
Biofilm-Forming Staphylococcus epidermidis Expressing Vancomycin Resistance Early after Adhesion to a Metal Surface

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Toshiyuki Sakimura

2015-01-01

Full Text Available We investigated biofilm formation and time of vancomycin (VCM resistance expression after adhesion to a metal surface in Staphylococcus epidermidis. Biofilm-forming Staphylococcus epidermidis with a VCM MIC of 1 μg/mL was used. The bacteria were made to adhere to a stainless steel washer and treated with VCM at different times and concentrations. VCM was administered 0, 2, 4, and 8 hours after adhesion. The amount of biofilm formed was evaluated based on the biofilm coverage rates (BCRs before and after VCM administration, bacterial viability in biofilm was visually observed using the fluorescence staining method, and the viable bacterial count in biofilm was measured. The VCM concentration required to decrease BCR significantly compared with that of VCM-untreated bacteria was 4 μg/mL, even in the 0 hr group. In the 4 and 8 hr groups, VCM could not inhibit biofilm growth even at 1,024 μg/mL. In the 8 hr group, viable bacteria remained in biofilm at a count of 104 CFU even at a high VCM concentration (1,024 μg/mL. It was suggested that biofilm-forming Staphylococcus epidermidis expresses resistance to VCM early after adhesion to a metal surface. Resistance increased over time after adhesion as the biofilm formed, and strong resistance was expressed 4–8 hours after adhesion.
Signaling profiling at the single-cell level identifies a distinct signaling signature in murine hematopoietic stem cells.

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Du, Juan; Wang, Jinyong; Kong, Guangyao; Jiang, Jing; Zhang, Jingfang; Liu, Yangang; Tong, Wei; Zhang, Jing

2012-07-01

Hematopoietic stem cell (HSC) function is tightly regulated by cytokine signaling. Although phospho-flow cytometry allows us to study signaling in defined populations of cells, there has been tremendous hurdle to carry out this study in rare HSCs due to unrecoverable critical HSC markers, low HSC number, and poor cell recovery rate. Here, we overcame these difficulties and developed a "HSC phospho-flow" method to analyze cytokine signaling in murine HSCs at the single-cell level and compare HSC signaling profile to that of multipotent progenitors (MPPs), a cell type immediately downstream of HSCs, and commonly used Lin(-) cKit(+) cells (LK cells, enriched for myeloid progenitors). We chose to study signaling evoked from three representative cytokines, stem cell factor (SCF) and thrombopoietin (TPO) that are essential for HSC function and granulocyte macrophage-colony-stimulating factor (GM-CSF) that is dispensable for HSCs. HSCs display a distinct TPO and GM-CSF signaling signature from MPPs and LK cells, which highly correlates with receptor surface expression. In contrast, although majority of LK cells express lower levels of cKit than HSCs and MPPs, SCF-evoked ERK1/2 activation in LK cells shows a significantly increased magnitude for a prolonged period. These results suggest that specific cellular context plays a more important role than receptor surface expression in SCF signaling. Our study of HSC signaling at the homeostasis stage paves the way to investigate signaling changes in HSCs under conditions of stress, aging, and hematopoietic diseases. Copyright © 2012 AlphaMed Press.
Distinctive receptor binding properties of the surface glycoprotein of a natural Feline Leukemia Virus isolate with unusual disease spectrum

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Albritton Lorraine M

2011-05-01

Full Text Available Abstract Background Feline leukemia virus (FeLV-945, a member of the FeLV-A subgroup, was previously isolated from a cohort of naturally infected cats. An unusual multicentric lymphoma of non-T-cell origin was observed in natural and experimental infection with FeLV-945. Previous studies implicated the FeLV-945 surface glycoprotein (SU as a determinant of disease outcome by an as yet unknown mechanism. The present studies demonstrate that FeLV-945 SU confers distinctive properties of binding to the cell surface receptor. Results Virions bearing the FeLV-945 Env protein were observed to bind the cell surface receptor with significantly increased efficiency, as was soluble FeLV-945 SU protein, as compared to the corresponding virions or soluble protein from a prototype FeLV-A isolate. SU proteins cloned from other cohort isolates exhibited increased binding efficiency comparable to or greater than FeLV-945 SU. Mutational analysis implicated a domain containing variable region B (VRB to be the major determinant of increased receptor binding, and identified a single residue, valine 186, to be responsible for the effect. Conclusions The FeLV-945 SU protein binds its cell surface receptor, feTHTR1, with significantly greater efficiency than does that of prototype FeLV-A (FeLV-A/61E when present on the surface of virus particles or in soluble form, demonstrating a 2-fold difference in the relative dissociation constant. The results implicate a single residue, valine 186, as the major determinant of increased binding affinity. Computational modeling suggests a molecular mechanism by which residue 186 interacts with the receptor-binding domain through residue glutamine 110 to effect increased binding affinity. Through its increased receptor binding affinity, FeLV-945 SU might function in pathogenesis by increasing the rate of virus entry and spread in vivo, or by facilitating entry into a novel target cell with a low receptor density.
Confluence for classical logic through the distinction between values and computations

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José Espírito Santo

2014-09-01

Full Text Available We apply an idea originated in the theory of programming languages - monadic meta-language with a distinction between values and computations - in the design of a calculus of cut-elimination for classical logic. The cut-elimination calculus we obtain comprehends the call-by-name and call-by-value fragments of Curien-Herbelin's lambda-bar-mu-mu-tilde-calculus without losing confluence, and is based on a distinction of "modes" in the proof expressions and "mode" annotations in types. Modes resemble colors and polarities, but are quite different: we give meaning to them in terms of a monadic meta-language where the distinction between values and computations is fully explored. This meta-language is a refinement of the classical monadic language previously introduced by the authors, and is also developed in the paper.
Distinct Expression Profiles and Novel Targets of MicroRNAs in Human Spermatogonia, Pachytene Spermatocytes, and Round Spermatids between OA Patients and NOA Patients

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Chencheng Yao

2017-12-01

Full Text Available Human spermatogenesis includes three main stages, namely, the mitosis of spermatogonia, meiosis of spermatocytes, and spermiogenesis of spermatids, which are precisely regulated by epigenetic and genetic factors. Abnormality of epigenetic and genetic factors can result in aberrant spermatogenesis and eventual male infertility. However, epigenetic regulators in controlling each stage of normal and abnormal human spermatogenesis remain unknown. Here, we have revealed for the first time the distinct microRNA profiles in human spermatogonia, pachytene spermatocytes, and round spermatids between obstructive azoospermia (OA patients and non-obstructive azoospermia (NOA patients. Human spermatogonia, pachytene spermatocytes, and round spermatids from OA patients and NOA patients were isolated using STA-PUT velocity sedimentation and identified by numerous hallmarks for these cells. RNA deep sequencing showed that 396 microRNAs were differentially expressed in human spermatogonia between OA patients and NOA patients and 395 differentially expressed microRNAs were found in human pachytene spermatocytes between OA patients and NOA patients. Moreover, 378 microRNAs were differentially expressed in human round spermatids between OA patients and NOA patients. The differential expression of numerous microRNAs identified by RNA deep sequencing was verified by real-time PCR. Moreover, a number of novel targeting genes for microRNAs were predicted using various kinds of software and further verified by real-time PCR. This study thus sheds novel insights into epigenetic regulation of human normal spermatogenesis and the etiology of azoospermia, and it could offer new targets for molecular therapy to treat male infertility.
N-glycosylation of asparagine 8 regulates surface expression of major histocompatibility complex class I chain-related protein A (MICA) alleles dependent on threonine 24

DEFF Research Database (Denmark)

Pedersen, Maiken Mellergaard; Skovbakke, Sarah Line; Schneider, Christine L.

2014-01-01

for cell-surface expression and sought to identify the essential residues. We found that a single N-glycosylation site (N8) was important for MICA018 surface expression. The frequently expressed MICA allele 008, with an altered transmembrane and intracellular domain, was not affected by mutation of this N......-glycosylation site. Mutational analysis revealed that a single amino acid (T24) in the extracellular domain of MICA018 was essential for the N-glycosylation dependency, while the intracellular domain was not involved. The HHV7 immunoevasin, U21, was found to inhibit MICA018 surface expression by affecting N......-glycosylation and the retention was rescued by T24A substitution. Our study reveals N-glycosylation as an allele-specific regulatory mechanism important for regulation of surface expression of MICA018 and we pinpoint the residues essential for this N-glycosylation dependency. In addition we show that this regulatory mechanism...
Effector and regulatory dendritic cells display distinct patterns of miRNA expression

OpenAIRE

Lombardi, Vincent; Luce, Sonia; Moussu, H?l?ne; Morizur, Lise; Gueguen, Claire; Neukirch, Catherine; Chollet?Martin, Sylvie; Mascarell, Laurent; Aubier, Michel; Baron?Bodo, V?ronique; Moingeon, Philippe

2017-01-01

Abstract Introduction MicroRNAs (miRNAs) contribute to the regulation of dendritic cell (DC) polarization, thereby influencing the balance of adaptive immune responses. Herein, we studied the expression of miRNAs in polarized DCs and analyzed whether expression of these miRNAs could be associated with allergic rhinitis and allergen immunotherapy (AIT) outcome. Method Using specific culture conditions, we differentiated immature human monocyte?derived DCs into DC1, DC2, and DCreg subsets (supp...
PAF-receptor is preferentially expressed in a distinct synthetic phenotype of smooth muscle cells cloned from human internal thoracic artery: Functional implications in cell migration

International Nuclear Information System (INIS)

Stengel, Dominique; O'Neil, Caroline; Brocheriou, Isabelle; Karabina, Sonia-Athina; Durand, Herve; Caplice, Noel M.; Pickering, J. Geoffrey; Ninio, Ewa

2006-01-01

Platelet-activating-Factor (PAF) and its structural analogues formed upon low density lipoprotein oxidation are involved in atherosclerotic plaque formation and may signal through PAF-receptor (PAF-R) expressed in human macrophages and in certain smooth muscle cells (SMCs) in the media, but rarely in the intima of human plaques. Our aim was to determine which SMC phenotype expresses PAF-R and whether this receptor is functional in cell migration. Circulating SMC progenitors and two phenotypically distinct clones of proliferative, epithelioid phenotype vs contractile, spindle-shaped SMCs from the media of adult internal thoracic artery were studied for the presence of PAF-receptor (PAF-R). The levels of specific mRNA were obtained by reverse transcription/real-time PCR, the protein expression was deduced from immunohistochemistry staining, and the functional transmigration assay was performed by Boyden chamber-type chemotaxis assay. Only SMCs of spindle-shape and synthetic phenotype expressed both mRNA and PAF-R protein and in the functional test migrated at low concentrations of PAF. Two unrelated, specific PAF-R antagonists inhibited PAF-induced migration, but did not modify the migration initiated by PDGF. The presence of functional PAF-R in arterial spindle-shaped SMCs of synthetic phenotype may be important for their migration from the media into the intima and atherosclerotic plaques formation
Distinct gene subsets in pterygia formation and recurrence: dissecting complex biological phenomenon using genome wide expression data

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Ang Leonard PK

2009-03-01

Full Text Available Abstract Background Pterygium is a common ocular surface disease characterized by fibrovascular invasion of the cornea and is sight-threatening due to astigmatism, tear film disturbance, or occlusion of the visual axis. However, the mechanisms for formation and post-surgical recurrence of pterygium are not understood, and a valid animal model does not exist. Here, we investigated the possible mechanisms of pterygium pathogenesis and recurrence. Methods First we performed a genome wide expression analysis (human Affymetrix Genechip, >22000 genes with principal component analysis and clustering techniques, and validated expression of key molecules with PCR. The controls for this study were the un-involved conjunctival tissue of the same eye obtained during the surgical resection of the lesions. Interesting molecules were further investigated with immunohistochemistry, Western blots, and comparison with tear proteins from pterygium patients. Results Principal component analysis in pterygium indicated a signature of matrix-related structural proteins, including fibronectin-1 (both splice-forms, collagen-1A2, keratin-12 and small proline rich protein-1. Immunofluorescence showed strong expression of keratin-6A in all layers, especially the superficial layers, of pterygium epithelium, but absent in the control, with up-regulation and nuclear accumulation of the cell adhesion molecule CD24 in the pterygium epithelium. Western blot shows increased protein expression of beta-microseminoprotein, a protein up-regulated in human cutaneous squamous cell carcinoma. Gene products of 22 up-regulated genes in pterygium have also been found by us in human tears using nano-electrospray-liquid chromatography/mass spectrometry after pterygium surgery. Recurrent disease was associated with up-regulation of sialophorin, a negative regulator of cell adhesion, and never in mitosis a-5, known to be involved in cell motility. Conclusion Aberrant wound healing is therefore
Remodelling of Membrane Rafts Expression in Lung Cells as an Early Sign of Mechanotransduction-Signalling in Pulmonary Edema

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Paola Palestini

2011-01-01

Full Text Available Membrane rafts (MRs are clusters of lipids, organized in a “quasicrystalline” liquid-order phase, organized on the cell surface and whose pattern of molecules and physicochemical properties are distinct from those of the surrounding plasma membrane. MRs may be considered an efficient and fairly rapid cell-activated mechanism to express or mask surface receptors aimed at triggering specific response pathways. This paper reports observations concerning the role of MRs in the control of lung extravascular water that ought to be kept at minimum to assure gas diffusion, supporting the hypothesis that MRs expression is a potential mechanism of sensing minor changes in the volume of extravascular water. We present the evidence that MRs expression specifically relates to signal-transduction processes evoked by mechanical stimuli arising in the interstitial lung compartment when a small increase in extravascular volume occurs. We further hypothesize that a differential expression of MRs might also reflect the damage to precise components of the extracellular matrix caused by the perturbation in water balance and thus can trigger a molecule-oriented specific matrix remodelling.
In-surface confinement of topological insulator nanowire surface states

International Nuclear Information System (INIS)

Chen, Fan W.; Jauregui, Luis A.; Tan, Yaohua; Manfra, Michael; Klimeck, Gerhard; Chen, Yong P.; Kubis, Tillmann

2015-01-01

The bandstructures of [110] and [001] Bi 2 Te 3 nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects

In-surface confinement of topological insulator nanowire surface states

Energy Technology Data Exchange (ETDEWEB)

Chen, Fan W., E-mail: fanchen@purdue.edu [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); Jauregui, Luis A. [School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Tan, Yaohua [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Manfra, Michael [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); School of Materials Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Klimeck, Gerhard [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Chen, Yong P. [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Kubis, Tillmann [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States)

2015-09-21

The bandstructures of [110] and [001] Bi{sub 2}Te{sub 3} nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects.
In-surface confinement of topological insulator nanowire surface states

Science.gov (United States)

Chen, Fan W.; Jauregui, Luis A.; Tan, Yaohua; Manfra, Michael; Klimeck, Gerhard; Chen, Yong P.; Kubis, Tillmann

2015-09-01

The bandstructures of [110] and [001] Bi2Te3 nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects.
A conserved WW domain-like motif regulates invariant chain-dependent cell-surface transport of the NKG2D ligand ULBP2

DEFF Research Database (Denmark)

Uhlenbrock, Franziska Katharina; van Andel, Esther; Andresen, Lars

2015-01-01

that the NKG2D ligand ULBP2 traffics over an invariant chain (Ii)-dependent pathway to the cell surface. This study set out to elucidate how Ii regulates ULBP2 cell-surface transport: We discovered conserved tryptophan (Trp) residues in the primary protein sequence of ULBP1-6 but not in the related MICA....../B. Substitution of Trp to alanine resulted in cell-surface inhibition of ULBP2 in different cancer cell lines. Moreover, the mutated ULBP2 constructs were retained and not degraded inside the cell, indicating a crucial role of this conserved Trp-motif in trafficking. Finally, overexpression of Ii increased...... surface expression of wt ULBP2 while Trp-mutants could not be expressed, proposing that this Trp-motif is required for an Ii-dependent cell-surface transport of ULBP2. Aberrant soluble ULBP2 is immunosuppressive. Thus, targeting a distinct protein module on the ULBP2 sequence could counteract...
Effector and regulatory dendritic cells display distinct patterns of miRNA expression.

Science.gov (United States)

Lombardi, Vincent; Luce, Sonia; Moussu, Hélène; Morizur, Lise; Gueguen, Claire; Neukirch, Catherine; Chollet-Martin, Sylvie; Mascarell, Laurent; Aubier, Michel; Baron-Bodo, Véronique; Moingeon, Philippe

2017-09-01

MicroRNAs (miRNAs) contribute to the regulation of dendritic cell (DC) polarization, thereby influencing the balance of adaptive immune responses. Herein, we studied the expression of miRNAs in polarized DCs and analyzed whether expression of these miRNAs could be associated with allergic rhinitis and allergen immunotherapy (AIT) outcome. Using specific culture conditions, we differentiated immature human monocyte-derived DCs into DC1, DC2, and DCreg subsets (supporting the differentiation of T H 1, T H 2 or regulatory T cells, respectively). Profiling of miRNA expression was performed in these DC subpopulations using microarrays. Levels of miRNAs specific for polarized DCs were then evaluated in a cohort of 58 patients with allergic rhinitis and 25 non-allergic controls, as well as in samples from 30 subjects treated with sublingual grass pollen tablets or placebo for four months. We successfully identified 16 miRNAs differentially regulated between immature DCs, DC1, DC2, and DCreg cells. In allergic rhinoconjunctivitis patients, the expression of two of those miRNAs (miR-132 and miR-155), was down-regulated compared to non-allergic individuals. However, the levels of these miRNAs were not significantly modified following four months of grass pollen immunotherapy. Studying polarized DCs and clinical samples from subjects with or without allergic rhinoconjunctivitis, we demonstrated that the expression of two miRNAs linked to effector DCs (i.e., DC1 and/or DC2 cells), was reduced in the blood of patients with allergic rhinoconjunctivitis. Nevertheless, these miRNAs did not represent relevant biomarkers to predict or follow-up AIT efficacy. © 2017 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd.
Novel CLCNKB mutations causing Bartter syndrome affect channel surface expression.

Science.gov (United States)

Keck, Mathilde; Andrini, Olga; Lahuna, Olivier; Burgos, Johanna; Cid, L Pablo; Sepúlveda, Francisco V; L'hoste, Sébastien; Blanchard, Anne; Vargas-Poussou, Rosa; Lourdel, Stéphane; Teulon, Jacques

2013-09-01

Mutations in the CLCNKB gene encoding the ClC-Kb Cl(-) channel cause Bartter syndrome, which is a salt-losing renal tubulopathy. Here, we investigate the functional consequences of seven mutations. When expressed in Xenopus laevis oocytes, four mutants carried no current (c.736G>C, p.Gly246Arg; c.1271G>A, p.Gly424Glu; c.1313G>A, p.Arg438His; c.1316T>C, p.Leu439Pro), whereas others displayed a 30%-60% reduction in conductance as compared with wild-type ClC-Kb (c.242T>C, p.Leu81Pro; c.274C>T, p.Arg92Trp; c.1052G>C, p.Arg351Pro). Anion selectivity and sensitivity to external Ca(2+) and H(+), typical of the ClC-Kb channel, were not modified in the partially active mutants. In oocytes, we found that all the mutations reduced surface expression with a profile similar to that observed for currents. In HEK293 cells, the currents in the mutants had similar profiles to those obtained in oocytes, except for p.Leu81Pro, which produced no current. Furthermore, p.Arg92Trp and p.Arg351Pro mutations did not modify the unit-conductance of closely related ClC-K1. Western blot analysis in HEK293 cells showed that ClC-Kb protein abundance was lower for the nonconducting mutants but similar to wild-type for other mutants. Overall, two classes of mutants can be distinguished: nonconducting mutants associated with low total protein expression, and partially conducting mutants with unaltered channel properties and ClC-Kb protein abundance. © 2013 WILEY PERIODICALS, INC.
Heat stress causes spatially-distinct membrane re-modelling in K562 leukemia cells.

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Gábor Balogh

Full Text Available Cellular membranes respond rapidly to various environmental perturbations. Previously we showed that modulations in membrane fluidity achieved by heat stress (HS resulted in pronounced membrane organization alterations which could be intimately linked to the expression and cellular distribution of heat shock proteins. Here we examine heat-induced membrane changes using several visualisation methods. With Laurdan two-photon microscopy we demonstrate that, in contrast to the enhanced formation of ordered domains in surface membranes, the molecular disorder is significantly elevated within the internal membranes of cells preexposed to mild HS. These results were compared with those obtained by anisotropy, fluorescence lifetime and electron paramagnetic resonance measurements. All probes detected membrane changes upon HS. However, the structurally different probes revealed substantially distinct alterations in membrane heterogeneity. These data call attention to the careful interpretation of results obtained with only a single label. Subtle changes in membrane microstructure in the decision-making of thermal cell killing could have potential application in cancer therapy.
Retinal ganglion cells with distinct directional preferences differ in molecular identity, structure, and central projections.

Science.gov (United States)

Kay, Jeremy N; De la Huerta, Irina; Kim, In-Jung; Zhang, Yifeng; Yamagata, Masahito; Chu, Monica W; Meister, Markus; Sanes, Joshua R

2011-05-25

The retina contains ganglion cells (RGCs) that respond selectively to objects moving in particular directions. Individual members of a group of ON-OFF direction-selective RGCs (ooDSGCs) detect stimuli moving in one of four directions: ventral, dorsal, nasal, or temporal. Despite this physiological diversity, little is known about subtype-specific differences in structure, molecular identity, and projections. To seek such differences, we characterized mouse transgenic lines that selectively mark ooDSGCs preferring ventral or nasal motion as well as a line that marks both ventral- and dorsal-preferring subsets. We then used the lines to identify cell surface molecules, including Cadherin 6, CollagenXXVα1, and Matrix metalloprotease 17, that are selectively expressed by distinct subsets of ooDSGCs. We also identify a neuropeptide, CART (cocaine- and amphetamine-regulated transcript), that distinguishes all ooDSGCs from other RGCs. Together, this panel of endogenous and transgenic markers distinguishes the four ooDSGC subsets. Patterns of molecular diversification occur before eye opening and are therefore experience independent. They may help to explain how the four subsets obtain distinct inputs. We also demonstrate differences among subsets in their dendritic patterns within the retina and their axonal projections to the brain. Differences in projections indicate that information about motion in different directions is sent to different destinations.
Differential expression of syndecan isoforms during mouse incisor amelogenesis.

Science.gov (United States)

Muto, Taro; Miyoshi, Keiko; Munesue, Seiichi; Nakada, Hiroshi; Okayama, Minoru; Matsuo, Takashi; Noma, Takafumi

2007-08-01

Syndecans are transmembranous heparan sulfate proteoglycans (HSPGs) with covalently attached glycosaminoglycan side-chains located on the cell surface. The mammalian syndecan family is composed of four types of syndecans (syndecan-1 to -4). Syndecans interact with the intracellular cytoskeleton through the cytoplasmic domains of their core proteins and membrane proteins, extracellular enzymes, growth factors, and matrix components, through their heparan-sulfate chains, to regulate developmental processes.Here, as a first step to assess the possible roles of syndecan proteins in amelogenesis, we examined the expression patterns of all syndecan isoforms in continuously growing mouse incisors, in which we can overview major differentiation stages of amelogenesis at a glance. Understanding the expression domain of each syndecan isoform during specific developmental stages seems useful for investigating their physiological roles in amelogenesis. Immunohistochemical analysis of syndecan core proteins in the lower incisors from postnatal day 1 mice revealed spatially and temporally specific expression patterns, with syndecan-1 expressed in undifferentiated epithelial and mesenchymal cells, and syndecan-2, -3, and -4 in more differentiated cells. These findings suggest that each syndecan isoform functions distinctly during the amelogenesis of the incisors of mice.
Fragile X Proteins FMRP and FXR2P Control Synaptic GluA1 Expression and Neuronal Maturation via Distinct Mechanisms

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Weixiang Guo

2015-06-01

Full Text Available Fragile X mental retardation protein (FMRP and its autosomal paralog FXR2P are selective neuronal RNA-binding proteins, and mice that lack either protein exhibit cognitive deficits. Although double-mutant mice display more severe learning deficits than single mutants, the molecular mechanism behind this remains unknown. In the present study, we discovered that FXR2P (also known as FXR2 is important for neuronal dendritic development. FMRP and FXR2P additively promote the maturation of new neurons by regulating a common target, the AMPA receptor GluA1, but they do so via distinct mechanisms: FXR2P binds and stabilizes GluA1 mRNA and enhances subsequent protein expression, whereas FMRP promotes GluA1 membrane delivery. Our findings unveil important roles for FXR2P and GluA1 in neuronal development, uncover a regulatory mechanism of GluA1, and reveal a functional convergence between fragile X proteins in neuronal development.
Gene expression in Pseudomonas aeruginosa swarming motility

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Déziel Eric

2010-10-01

Full Text Available Abstract Background The bacterium Pseudomonas aeruginosa is capable of three types of motilities: swimming, twitching and swarming. The latter is characterized by a fast and coordinated group movement over a semi-solid surface resulting from intercellular interactions and morphological differentiation. A striking feature of swarming motility is the complex fractal-like patterns displayed by migrating bacteria while they move away from their inoculation point. This type of group behaviour is still poorly understood and its characterization provides important information on bacterial structured communities such as biofilms. Using GeneChip® Affymetrix microarrays, we obtained the transcriptomic profiles of both bacterial populations located at the tip of migrating tendrils and swarm center of swarming colonies and compared these profiles to that of a bacterial control population grown on the same media but solidified to not allow swarming motility. Results Microarray raw data were corrected for background noise with the RMA algorithm and quantile normalized. Differentially expressed genes between the three conditions were selected using a threshold of 1.5 log2-fold, which gave a total of 378 selected genes (6.3% of the predicted open reading frames of strain PA14. Major shifts in gene expression patterns are observed in each growth conditions, highlighting the presence of distinct bacterial subpopulations within a swarming colony (tendril tips vs. swarm center. Unexpectedly, microarrays expression data reveal that a minority of genes are up-regulated in tendril tip populations. Among them, we found energy metabolism, ribosomal protein and transport of small molecules related genes. On the other hand, many well-known virulence factors genes were globally repressed in tendril tip cells. Swarm center cells are distinct and appear to be under oxidative and copper stress responses. Conclusions Results reported in this study show that, as opposed to
Expression of cell adhesion and differentiation related genes in MC3T3 osteoblasts plated on titanium alloys: role of surface properties

International Nuclear Information System (INIS)

Sista, Subhash; Wen, Cuie; Hodgson, Peter D.; Pande, Gopal

2013-01-01

It is important to understand the cellular and molecular events that take place at the cell–material interface of implants used for bone repair. An understanding of the mechanisms involved in the initial stages of osteoblast interactions with the surface of the implant material is fundamental in deciding the fate of the cells that come in contact with it. In this study, we compared the relative gene expression of markers that are known to be associated with cell adhesion and differentiation in MC3T3 osteoblast cells, at various time points after plating the cells on surfaces of titanium (Ti) and its two alloys, titanium–zirconium (TiZr) and titanium–niobium (TiNb) by using Quantitative Real Time Polymerase Chain Reaction (RT-PCR). Our analysis indicated that expression of adhesion supporting genes was higher on TiZr surface as compared to Ti and TiNb. The behavior of these genes is possibly driven by a higher surface energy of TiZr. However no significant difference in the expression of differentiation related genes could be seen between the two alloys, although on both substrates it was higher as compared to unalloyed Ti. We propose that substrate composition of the alloys can influence the adhesion and differentiation related gene expression and that Ti alloys are better substrates for inducing osteogenesis as compared to unalloyed Ti. - Highlights: ► Methodology for comparing gene expression in osteoblasts plated on Ti, TiZr or TiNb ► Alloys with higher surface energy (TiZr) induce cell adhesion genes more efficiently ► Alloyed Ti is superior to unalloyed Ti to induce osteoblast differentiation genes
Identification of differentially expressed genes in cucumber (Cucumis sativus L.) root under waterlogging stress by digital gene expression profile.

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Qi, Xiao-Hua; Xu, Xue-Wen; Lin, Xiao-Jian; Zhang, Wen-Jie; Chen, Xue-Hao

2012-03-01

High-throughput tag-sequencing (Tag-seq) analysis based on the Solexa Genome Analyzer platform was applied to analyze the gene expression profiling of cucumber plant at 5 time points over a 24h period of waterlogging treatment. Approximately 5.8 million total clean sequence tags per library were obtained with 143013 distinct clean tag sequences. Approximately 23.69%-29.61% of the distinct clean tags were mapped unambiguously to the unigene database, and 53.78%-60.66% of the distinct clean tags were mapped to the cucumber genome database. Analysis of the differentially expressed genes revealed that most of the genes were down-regulated in the waterlogging stages, and the differentially expressed genes mainly linked to carbon metabolism, photosynthesis, reactive oxygen species generation/scavenging, and hormone synthesis/signaling. Finally, quantitative real-time polymerase chain reaction using nine genes independently verified the tag-mapped results. This present study reveals the comprehensive mechanisms of waterlogging-responsive transcription in cucumber. Copyright Â© 2011 Elsevier Inc. All rights reserved.
High wind speeds prevent formation of a distinct bacterioneuston community in the sea-surface microlayer.

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Rahlff, Janina; Stolle, Christian; Giebel, Helge-Ansgar; Brinkhoff, Thorsten; Ribas-Ribas, Mariana; Hodapp, Dorothee; Wurl, Oliver

2017-05-01

The sea-surface microlayer (SML) at the boundary between atmosphere and hydrosphere represents a demanding habitat for bacteria. Wind speed is a crucial but poorly studied factor for its physical integrity. Increasing atmospheric burden of CO2, as suggested for future climate scenarios, may particularly act on this habitat at the air-sea interface. We investigated the effect of increasing wind speeds and different pCO2 levels on SML microbial communities in a wind-wave tunnel, which offered the advantage of low spatial and temporal variability. We found that enrichment of bacteria in the SML occurred solely at a U10 wind speed of ≤5.6 m s-1 in the tunnel and ≤4.1 m s-1 in the Baltic Sea. High pCO2 levels further intensified the bacterial enrichment in the SML during low wind speed. In addition, low wind speed and pCO2 induced the formation of a distinctive bacterial community as revealed by 16S rRNA gene fingerprints and influenced the presence or absence of individual taxonomic units within the SML. We conclude that physical stability of the SML below a system-specific wind speed threshold induces specific bacterial communities in the SML entailing strong implications for ecosystem functioning by wind-driven impacts on habitat properties, gas exchange and matter cycling processes. © FEMS 2017.
Characterization of a distinct population of circulating human non-adherent endothelial forming cells and their recruitment via intercellular adhesion molecule-3.

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Sarah L Appleby

Full Text Available Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133(+ population of non-adherent endothelial forming cells (naEFCs which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38 together with mature endothelial cell markers (VEGFR2, CD144 and CD31. These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8 or myeloid markers (CD11b and CD14 which distinguishes them from 'early' endothelial progenitor cells (EPCs. Functional studies demonstrated that these naEFCs (i bound Ulex europaeus lectin, (ii demonstrated acetylated-low density lipoprotein uptake, (iii increased vascular cell adhesion molecule (VCAM-1 surface expression in response to tumor necrosis factor and (iv in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs. Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis.
Characterization of a Distinct Population of Circulating Human Non-Adherent Endothelial Forming Cells and Their Recruitment via Intercellular Adhesion Molecule-3

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Thompson, Emma J.; Barrett, Jeffrey M.; Tooley, Katie; Sen, Shaundeep; Sun, Wai Yan; Grose, Randall; Nicholson, Ian; Levina, Vitalina; Cooke, Ira; Talbo, Gert; Lopez, Angel F.; Bonder, Claudine S.

2012-01-01

Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133+ population of non-adherent endothelial forming cells (naEFCs) which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38) together with mature endothelial cell markers (VEGFR2, CD144 and CD31). These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8) or myeloid markers (CD11b and CD14) which distinguishes them from ‘early’ endothelial progenitor cells (EPCs). Functional studies demonstrated that these naEFCs (i) bound Ulex europaeus lectin, (ii) demonstrated acetylated-low density lipoprotein uptake, (iii) increased vascular cell adhesion molecule (VCAM-1) surface expression in response to tumor necrosis factor and (iv) in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs). Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM)-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis. PMID:23144795
Specific Regional and Age-Related Small Noncoding RNA Expression Patterns Within Superior Temporal Gyrus of Typical Human Brains Are Less Distinct in Autism Brains.

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Stamova, Boryana; Ander, Bradley P; Barger, Nicole; Sharp, Frank R; Schumann, Cynthia M

2015-12-01

Small noncoding RNAs play a critical role in regulating messenger RNA throughout brain development and when altered could have profound effects leading to disorders such as autism spectrum disorders (ASD). We assessed small noncoding RNAs, including microRNA and small nucleolar RNA, in superior temporal sulcus association cortex and primary auditory cortex in typical and ASD brains from early childhood to adulthood. Typical small noncoding RNA expression profiles were less distinct in ASD, both between regions and changes with age. Typical micro-RNA coexpression associations were absent in ASD brains. miR-132, miR-103, and miR-320 micro-RNAs were dysregulated in ASD and have previously been associated with autism spectrum disorders. These diminished region- and age-related micro-RNA expression profiles are in line with previously reported findings of attenuated messenger RNA and long noncoding RNA in ASD brain. This study demonstrates alterations in superior temporal sulcus in ASD, a region implicated in social impairment, and is the first to demonstrate molecular alterations in the primary auditory cortex. © The Author(s) 2015.
Ovarian Surface Epithelium in Patients with Severe Ovarian Infertility: A Potential Source of Cells Expressing Markers of Pluripotent/Multipotent Stem Cells

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Irma Virant-Klun

2011-01-01

Full Text Available The aim of this study was to confirm the presence of stem cells in the ovarian surface epithelium of patients with premature ovarian failure and no mature follicles and oocytes. In these patients, small round cells of unknown origin expressing SOX-2 marker of pluripotency were observed among the epithelial cells just after the ovarian surface epithelium scraping. These cells were an integral part of the ovarian surface epithelium. When the scraped cells were cultured in a medium with added follicular fluid to provide some ovarian niche, primitive oocyte-like cells and typical round-shaped cell clusters positively stained on alkaline phosphatase, and markers of pluripotency, such as SOX-2 and SSEA-4, were developed. These markers were expressed early and also later in the culture. Single oocyte-like cells expressed genes OCT4A, SOX-2, NANOG, NANOS, STELLA, CD9, LIN28, KLF4, GDF3, and MYC, characteristic for pluripotent stem cells. The results of this study confirmed the presence of putative stem cells in the ovarian surface epithelium of these patients and provided some basis to create a stem cell line in the future.
Internalization of interleukin 1 (IL 1) correlates with IL 1-induced IL 2 receptor expression and IL 2 secretion of EL4 thymoma cells

OpenAIRE

Von Hoegen, I.; Falk, Werner; Kojouharoff, G.; Krammer, P. H.

1989-01-01

The cytokine interleukin 1 (IL 1) plays an important role in the induction of IL 2 secretion and high-affinity IL 2 receptor (IL 2R) expression by T cells. The events that follow binding of IL 1 to IL 1R, however, are still unknown. In this study we describe two variants of the murine thymoma EL4 (5D3 and D6/76) that express comparable numbers of cell surface IL 1 receptors and bind IL 1 with the same affinity, but show distinct IL 1-dependent IL 2 secretion and IL 2R expression. In the prese...
Viral DNA Sensors IFI16 and Cyclic GMP-AMP Synthase Possess Distinct Functions in Regulating Viral Gene Expression, Immune Defenses, and Apoptotic Responses during Herpesvirus Infection

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Benjamin A. Diner

2016-11-01

Full Text Available The human interferon-inducible protein IFI16 is an important antiviral factor that binds nuclear viral DNA and promotes antiviral responses. Here, we define IFI16 dynamics in space and time and its distinct functions from the DNA sensor cyclic dinucleotide GMP-AMP synthase (cGAS. Live-cell imaging reveals a multiphasic IFI16 redistribution, first to viral entry sites at the nuclear periphery and then to nucleoplasmic puncta upon herpes simplex virus 1 (HSV-1 and human cytomegalovirus (HCMV infections. Optogenetics and live-cell microscopy establish the IFI16 pyrin domain as required for nuclear periphery localization and oligomerization. Furthermore, using proteomics, we define the signature protein interactions of the IFI16 pyrin and HIN200 domains and demonstrate the necessity of pyrin for IFI16 interactions with antiviral proteins PML and cGAS. We probe signaling pathways engaged by IFI16, cGAS, and PML using clustered regularly interspaced short palindromic repeat (CRISPR/Cas9-mediated knockouts in primary fibroblasts. While IFI16 induces cytokines, only cGAS activates STING/TBK-1/IRF3 and apoptotic responses upon HSV-1 and HCMV infections. cGAS-dependent apoptosis upon DNA stimulation requires both the enzymatic production of cyclic dinucleotides and STING. We show that IFI16, not cGAS or PML, represses HSV-1 gene expression, reducing virus titers. This indicates that regulation of viral gene expression may function as a greater barrier to viral replication than the induction of antiviral cytokines. Altogether, our findings establish coordinated and distinct antiviral functions for IFI16 and cGAS against herpesviruses.
Thy-1 attenuates TNF-alpha-activated gene expression in mouse embryonic fibroblasts via Src family kinase.

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Bin Shan

Full Text Available Heterogeneous surface expression of Thy-1 in fibroblasts modulates inflammation and may thereby modulate injury and repair. As a paradigm, patients with idiopathic pulmonary fibrosis, a disease with pathologic features of chronic inflammation, demonstrate an absence of Thy-1 immunoreactivity within areas of fibrotic activity (fibroblast foci in contrast to the predominant Thy-1 expressing fibroblasts in the normal lung. Likewise, Thy-1 deficient mice display more severe lung fibrosis in response to an inflammatory injury than wildtype littermates. We investigated the role of Thy-1 in the response of fibroblasts to the pro-inflammatory cytokine TNF-alpha. Our study demonstrates distinct profiles of TNF-alpha-activated gene expression in Thy-1 positive (Thy-1+ and negative (Thy-1- subsets of mouse embryonic fibroblasts (MEF. TNF-alpha induced a robust activation of MMP-9, ICAM-1, and the IL-8 promoter driven reporter in Thy-1- MEFs, in contrast to only a modest increase in Thy-1+ counterparts. Consistently, ectopic expression of Thy-1 in Thy-1- MEFs significantly attenuated TNF-alpha-activated gene expression. Mechanistically, TNF-alpha activated Src family kinase (SFK only in Thy-1- MEFs. Blockade of SFK activation abrogated TNF-alpha-activated gene expression in Thy-1- MEFs, whereas restoration of SFK activation rescued the TNF-alpha response in Thy-1+ MEFs. Our findings suggest that Thy-1 down-regulates TNF-alpha-activated gene expression via interfering with SFK- and NF-kappaB-mediated transactivation. The current study provides a novel mechanistic insight to the distinct roles of fibroblast Thy-1 subsets in inflammation.

Campylobacter jejuni cocultured with epithelial cells reduces surface capsular polysaccharide expression.

LENUS (Irish Health Repository)

Corcionivoschi, N

2012-02-01

The host cell environment can alter bacterial pathogenicity. We employed a combination of cellular and molecular techniques to study the expression of Campylobacter jejuni polysaccharides cocultured with HCT-8 epithelial cells. After two passages, the amount of membrane-bound high-molecular-weight polysaccharide was considerably reduced. Microarray profiling confirmed significant downregulation of capsular polysaccharide (CPS) locus genes. Experiments using conditioned media showed that sugar depletion occurred only when the bacterial and epithelial cells were cocultured. CPS depletion occurred when C. jejuni organisms were exposed to conditioned media from a different C. jejuni strain but not when exposed to conditioned media from other bacterial species. Proteinase K or heat treatment of conditioned media under coculture conditions abrogated the effect on the sugars, as did formaldehyde fixation and cycloheximide treatment of host cells or chloramphenicol treatment of the bacteria. However, sugar depletion was not affected in flagellar export (fliQ) and quorum-sensing (luxS) gene mutants. Passaged C. jejuni showed reduced invasiveness and increased serum sensitivity in vitro. C. jejuni alters its surface polysaccharides when cocultured with epithelial cells, suggesting the existence of a cross talk mechanism that modulates CPS expression during infection.
Potential costs of bacterial infection on storage protein gene expression and reproduction in queenless Apis mellifera worker bees on distinct dietary regimes.

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Lourenço, Anete Pedro; Martins, Juliana Ramos; Guidugli-Lazzarini, Karina Rosa; Macedo, Liliane Maria Fróes; Bitondi, Márcia Maria Gentile; Simões, Zilá Luz Paulino

2012-09-01

Insects are able to combat infection by initiating an efficient immune response that involves synthesizing antimicrobial peptides and a range of other defense molecules. These responses may be costly to the organism, resulting in it exploiting endogenous resources to maintain homeostasis or support defense to the detriment of other physiological needs. We used queenless worker bees on distinct dietary regimes that may alter hemolymph protein storage and ovary activation to investigate the physiological costs of infection with Serratia marcescens. The expression of the genes encoding the storage proteins vitellogenin and hexamerin 70a, the vitellogenin receptor, and vasa (which has a putative role in reproduction), was impaired in the infected bees. This impairment was mainly evident in the bees fed beebread, which caused significantly higher expression of these genes than did royal jelly or syrup, and this was confirmed at the vitellogenin and hexamerin 70a protein levels. Beebread was also the only diet that promoted ovary activation in the queenless bees, but this activation was significantly impaired by the infection. The expression of the genes encoding the storage proteins apolipophorins-I and -III and the lipophorin receptor was not altered by infection regardless the diet provided to the bees. Similarly, the storage of apolipophorin-I in the hemolymph was only slightly impaired by the infection, independently of the supplied diet. Taken together these results indicate that, infection demands a physiological cost from the transcription of specific protein storage-related genes and from the reproductive capacity. Copyright © 2012 Elsevier Ltd. All rights reserved.
Exposure to nickel, chromium, or cadmium causes distinct changes in the gene expression patterns of a rat liver derived cell line.

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Matthew G Permenter

Full Text Available Many heavy metals, including nickel (Ni, cadmium (Cd, and chromium (Cr are toxic industrial chemicals with an exposure risk in both occupational and environmental settings that may cause harmful outcomes. While these substances are known to produce adverse health effects leading to disease or health problems, the detailed mechanisms remain unclear. To elucidate the processes involved in the toxicity of nickel, cadmium, and chromium at the molecular level and to perform a comparative analysis, H4-II-E-C3 rat liver-derived cell lines were treated with soluble salts of each metal using concentrations derived from viability assays, and gene expression patterns were determined with DNA microarrays. We identified both common and unique biological responses to exposure to the three metals. Nickel, cadmium, chromium all induced oxidative stress with both similar and unique genes and pathways responding to this stress. Although all three metals are known to be genotoxic, evidence for DNA damage in our study only exists in response to chromium. Nickel induced a hypoxic response as well as inducing genes involved in chromatin structure, perhaps by replacing iron in key proteins. Cadmium distinctly perturbed genes related to endoplasmic reticulum stress and invoked the unfolded protein response leading to apoptosis. With these studies, we have completed the first gene expression comparative analysis of nickel, cadmium, and chromium in H4-II-E-C3 cells.
Distinct expression profiles and different functions of odorant binding proteins in Nilaparvata lugens Stål.

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Peng He

Full Text Available BACKGROUND: Odorant binding proteins (OBPs play important roles in insect olfaction. The brown planthopper (BPH, Nilaparvata lugens Stål (Delphacidae, Auchenorrhyncha, Hemiptera is one of the most important rice pests. Its monophagy (only feeding on rice, wing form (long and short wing variation, and annual long distance migration (seeking for rice plants of high nutrition imply that the olfaction would play a central role in BPH behavior. However, the olfaction related proteins have not been characterized in this insect. METHODOLOGY/PRINCIPAL FINDINGS: Full length cDNA of three OBPs were obtained and distinct expression profiles were revealed regarding to tissue, developmental stage, wing form and gender for the first time for the species. The results provide important clues in functional differentiation of these genes. Binding assays with 41 compounds demonstrated that NlugOBP3 had markedly higher binding ability and wider binding spectrum than the other two OBPs. Terpenes and Ketones displayed higher binding while Alkanes showed no binding to the three OBPs. Focused on NlugOBP3, RNA interference experiments showed that NlugOBP3 not only involved in nymph olfaction on rice seedlings, but also had non-olfactory functions, as it was closely related to nymph survival. CONCLUSIONS: NlugOBP3 plays important roles in both olfaction and survival of BPH. It may serve as a potential target for developing behavioral disruptant and/or lethal agent in N. lugens.
Host gene expression profiles in ferrets infected with genetically distinct henipavirus strains.

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Alberto J Leon

2018-03-01

Full Text Available Henipavirus infection causes severe respiratory and neurological disease in humans that can be fatal. To characterize the pathogenic mechanisms of henipavirus infection in vivo, we performed experimental infections in ferrets followed by genome-wide gene expression analysis of lung and brain tissues. The Hendra, Nipah-Bangladesh, and Nipah-Malaysia strains caused severe respiratory and neurological disease with animals succumbing around 7 days post infection. Despite the presence of abundant viral shedding, animal-to-animal transmission did not occur. The host gene expression profiles of the lung tissue showed early activation of interferon responses and subsequent expression of inflammation-related genes that coincided with the clinical deterioration. Additionally, the lung tissue showed unchanged levels of lymphocyte markers and progressive downregulation of cell cycle genes and extracellular matrix components. Infection in the brain resulted in a limited breadth of the host responses, which is in accordance with the immunoprivileged status of this organ. Finally, we propose a model of the pathogenic mechanisms of henipavirus infection that integrates multiple components of the host responses.
Distinct abscisic acid signaling pathways for modulation of guard cell versus mesophyll cell potassium channels revealed by expression studies in Xenopus laevis oocytes

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Sutton, F.; Paul, S. S.; Wang, X. Q.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

2000-01-01

Regulation of guard cell ion transport by abscisic acid (ABA) and in particular ABA inhibition of a guard cell inward K(+) current (I(Kin)) is well documented. However, little is known concerning ABA effects on ion transport in other plant cell types. Here we applied patch clamp techniques to mesophyll cell protoplasts of fava bean (Vicia faba cv Long Pod) plants and demonstrated ABA inhibition of an outward K(+) current (I(Kout)). When mesophyll cell protoplast mRNA (mesophyll mRNA) was expressed in Xenopus laevis oocytes, I(Kout) was generated that displayed similar properties to I(Kout) observed from direct analysis of mesophyll cell protoplasts. I(Kout) expressed by mesophyll mRNA-injected oocytes was inhibited by ABA, indicating that the ABA signal transduction pathway observed in mesophyll cells was preserved in the frog oocytes. Co-injection of oocytes with guard cell protoplast mRNA and cRNA for KAT1, an inward K(+) channel expressed in guard cells, resulted in I(Kin) that was similarly inhibited by ABA. However, oocytes co-injected with mesophyll mRNA and KAT1 cRNA produced I(Kin) that was not inhibited by ABA. These results demonstrate that the mesophyll-encoded signaling mechanism could not substitute for the guard cell pathway. These findings indicate that mesophyll cells and guard cells use distinct and different receptor types and/or signal transduction pathways in ABA regulation of K(+) channels.
The Plasmodium serine-type SERA proteases display distinct expression patterns and non-essential in vivo roles during life cycle progression of the malaria parasite.

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Putrianti, Elyzana D; Schmidt-Christensen, Anja; Arnold, Iris; Heussler, Volker T; Matuschewski, Kai; Silvie, Olivier

2010-06-01

Parasite proteases play key roles in several fundamental steps of the Plasmodium life cycle, including haemoglobin degradation, host cell invasion and parasite egress. Plasmodium exit from infected host cells appears to be mediated by a class of papain-like cysteine proteases called 'serine repeat antigens' (SERAs). A SERA subfamily, represented by Plasmodium falciparum SERA5, contains an atypical active site serine residue instead of a catalytic cysteine. Members of this SERAser subfamily are abundantly expressed in asexual blood stages, rendering them attractive drug and vaccine targets. In this study, we show by antibody localization and in vivo fluorescent tagging with the red fluorescent protein mCherry that the two P. berghei serine-type family members, PbSERA1 and PbSERA2, display differential expression towards the final stages of merozoite formation. Via targeted gene replacement, we generated single and double gene knockouts of the P. berghei SERAser genes. These loss-of-function lines progressed normally through the parasite life cycle, suggesting a specialized, non-vital role for serine-type SERAs in vivo. Parasites lacking PbSERAser showed increased expression of the cysteine-type PbSERA3. Compensatory mechanisms between distinct SERA subfamilies may thus explain the absence of phenotypical defect in SERAser disruptants, and challenge the suitability to develop potent antimalarial drugs based on specific inhibitors of Plasmodium serine-type SERAs.
Distinct spatiotemporal expression of ISM1 during mouse and chick development.

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Osório, Liliana; Wu, Xuewei; Zhou, Zhongjun

2014-01-01

Isthmin 1 (ISM1) constitutes the founder of a new family of secreted proteins characterized by the presence of 2 functional domains: thrombospondin type 1 repeat (TSR1) and adhesion-associated domain in MUC4 and other proteins (AMOP). ISM1 was identified in the frog embryo as a member of the FGF8 synexpression group due to its expression in the brain midbrain-hindbrain boundary (MHB) or isthmus. In zebrafish, ISM1 was described as a WNT- and NODAL-regulated gene. The function of ISM1 remains largely elusive. So far, ISM1 has been described as an angiogenesis inhibitor that has a dual function in endothelial cell survival and cell death. For a better understanding of ISM1 function, we examined its spatiotemporal distribution in mouse and chick using RT-PCR, ISH, and IHC analyses. In the mouse, ISM1 transcripts are found in tissues such as the anterior mesendoderm, paraxial and lateral plate mesoderm, MHB and trunk neural tube, as well as in the somites and dermomyotome. In the newborn and adult, ISM1 is prominently expressed in the lung and brain. In addition to its putative role during embryonic and postnatal development, ISM1 may also be important for organ homeostasis in the adult. In the chick embryo, ISM1 transcripts are strongly detected in the ear, eye, and spinal cord primordia. Remarkable differences in ISM1 spatiotemporal expression were found during mouse and chick development, despite the high homology of ISM1 orthologs in these species.
Expressed var gene repertoire and variant surface antigen diversity in a shrinking Plasmodium falciparum population.

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Carlos, Bianca C; Fotoran, Wesley L; Menezes, Maria J; Cabral, Fernanda J; Bastos, Marcele F; Costa, Fabio T M; Sousa-Neto, Jayme A; Ribolla, Paulo E M; Wunderlich, Gerhard; Ferreira, Marcelo U

2016-11-01

The var gene-encoded erythrocyte membrane protein-1 of Plasmodium falciparum (PfEMP-1) is the main variant surface antigen (VSA) expressed on infected erythrocytes. The rate at which antibody responses to VSA expressed by circulating parasites are acquired depends on the size of the local VSA repertoire and the frequency of exposure to new VSA. Because parasites from areas with declining malaria endemicity, such as the Amazon, typically express a restricted PfEMP-1 repertoire, we hypothesized that Amazonians would rapidly acquire antibodies to most locally circulating VSA. Consistent with our expectations, the analysis of 5878 sequence tags expressed by 10 local P. falciparum samples revealed little PfEMP-1 DBL1α domain diversity. Among the most commonly expressed DBL1α types, 45% were shared by two or more independent parasite lines. Nevertheless, Amazonians displayed major gaps in their repertoire of anti-VSA antibodies, although the breadth of anti-VSA antibody responses correlated positively with their cumulative exposure to malaria. We found little antibody cross-reactivity even when testing VSA from related parasites expressing the same dominant DBL1α types. We conclude that variant-specific immunity to P. falciparum VSAs develops slowly despite the relatively restricted PfEMP-1 repertoire found in low-endemicity settings. Copyright © 2016 Elsevier Inc. All rights reserved.
RABA Members Act in Distinct Steps of Subcellular Trafficking of the FLAGELLIN SENSING2 Receptor[W

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Choi, Seung-won; Tamaki, Takayuki; Ebine, Kazuo; Uemura, Tomohiro; Ueda, Takashi; Nakano, Akihiko

2013-01-01

Cell surface proteins play critical roles in the perception of environmental stimuli at the plasma membrane (PM) and ensuing signal transduction. Intracellular localization of such proteins must be strictly regulated, which requires elaborate integration of exocytic and endocytic trafficking pathways. Subcellular localization of Arabidopsis thaliana FLAGELLIN SENSING2 (FLS2), a receptor that recognizes bacterial flagellin, also depends on membrane trafficking. However, our understanding about the mechanisms involved is still limited. In this study, we visualized ligand-induced endocytosis of FLS2 using green fluorescent protein (GFP)-tagged FLS2 expressed in Nicotiana benthamiana. Upon treatment with the flg22 peptide, internalized FLS2-GFP from the PM was transported to a compartment with properties intermediate between the trans-Golgi network (TGN) and the multivesicular endosome. This compartment gradually discarded the TGN characteristics as it continued along the trafficking pathway. We further found that FLS2 endocytosis involves distinct RABA/RAB11 subgroups at different steps. Moreover, we demonstrated that transport of de novo–synthesized FLS2 to the PM also involves a distinct RABA/RAB11 subgroup. Our results demonstrate the complex regulatory system for properly localizing FLS2 and functional differentiation in RABA members in endo- and exocytosis. PMID:23532067
Somatostatin receptor subtype expression in human thyroid tumours.

Science.gov (United States)

Klagge, A; Krause, K; Schierle, K; Steinert, F; Dralle, H; Fuhrer, D

2010-04-01

Somatostatin receptors (SSTR) are expressed in various endocrine tumours. The expression of SSTR at the tumour cell surface confers the possibility for diagnostic imaging and therapy of tumours using radiolabeled somatostatin analogues. The majority of currently available somatostatin analogues show a higher binding affinity for the SSTR2 subtype. To date, the precise expression pattern of the SSTR subtypes 1-5 in thyroid epithelial tumours remains to be determined. We investigated the mRNA expression of SSTR1-5 in benign and malignant epithelial thyroid tumours [20 cold thyroid nodules (CTNs), 20 toxic thyroid nodules (TTNs), 20 papillary, 20 follicular, and 5 anaplastic carcinomas (PTCs, FTCs, ATCs, respectively)] and compared them to normal surrounding thyroid tissues. Four out of five SSTR subtypes were detected in malignant thyroid tumours, benign neoplasia, and normal surrounding tissue with a predominant expression of SSTR2 and SSTR5, and a weak expression of SSTR1 and SSTR3. Weak SSTR4 mRNA expression was detected in some PTCs. Compared to normal thyroid tissue, SSTR2 was significantly upregulated in PTC and ATC. In addition significant upregulation of SSTR3 was found in PTC. SSTR5 mRNA expression was increased in PTC and FTC and significantly decreased in CTN and TTN compared to normal thyroid tissue. SSTR2 is the predominant subtype in thyroid epithelial tumours with a high expression pattern, in particular, in PTC . Perspectively, the expression of distinct SSTR in thyroid epithelial tumours might represent a promising avenue for diagnostics and therapy of advanced thyroid cancer with somatostatin analogues. Georg Thieme Verlag KG Stuttgart New York.
Geographical and temporal conservation of antibody recognition of Plasmodium falciparum variant surface antigens

DEFF Research Database (Denmark)

Nielsen, Morten A; Vestergaard, Lasse S; Lusingu, John

2004-01-01

The slow acquisition of protection against Plasmodium falciparum malaria probably reflects the extensive diversity of important antigens. The variant surface antigens (VSA) that mediate parasite adhesion to a range of host molecules are regarded as important targets of acquired protective immunity......, but their diversity makes them questionable vaccine candidates. We determined levels of VSA-specific immunoglobulin G (IgG) in human plasma collected at four geographically distant and epidemiologically distinct localities with specificity for VSA expressed by P. falciparum isolates from three African countries...
Anticancer Properties of Distinct Antimalarial Drug Classes

Science.gov (United States)

Hooft van Huijsduijnen, Rob; Guy, R. Kiplin; Chibale, Kelly; Haynes, Richard K.; Peitz, Ingmar; Kelter, Gerhard; Phillips, Margaret A.; Vennerstrom, Jonathan L.; Yuthavong, Yongyuth; Wells, Timothy N. C.

2013-01-01

We have tested five distinct classes of established and experimental antimalarial drugs for their anticancer potential, using a panel of 91 human cancer lines. Three classes of drugs: artemisinins, synthetic peroxides and DHFR (dihydrofolate reductase) inhibitors effected potent inhibition of proliferation with IC50s in the nM- low µM range, whereas a DHODH (dihydroorotate dehydrogenase) and a putative kinase inhibitor displayed no activity. Furthermore, significant synergies were identified with erlotinib, imatinib, cisplatin, dasatinib and vincristine. Cluster analysis of the antimalarials based on their differential inhibition of the various cancer lines clearly segregated the synthetic peroxides OZ277 and OZ439 from the artemisinin cluster that included artesunate, dihydroartemisinin and artemisone, and from the DHFR inhibitors pyrimethamine and P218 (a parasite DHFR inhibitor), emphasizing their shared mode of action. In order to further understand the basis of the selectivity of these compounds against different cancers, microarray-based gene expression data for 85 of the used cell lines were generated. For each compound, distinct sets of genes were identified whose expression significantly correlated with compound sensitivity. Several of the antimalarials tested in this study have well-established and excellent safety profiles with a plasma exposure, when conservatively used in malaria, that is well above the IC50s that we identified in this study. Given their unique mode of action and potential for unique synergies with established anticancer drugs, our results provide a strong basis to further explore the potential application of these compounds in cancer in pre-clinical or and clinical settings. PMID:24391728
Anticancer properties of distinct antimalarial drug classes.

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Rob Hooft van Huijsduijnen

Full Text Available We have tested five distinct classes of established and experimental antimalarial drugs for their anticancer potential, using a panel of 91 human cancer lines. Three classes of drugs: artemisinins, synthetic peroxides and DHFR (dihydrofolate reductase inhibitors effected potent inhibition of proliferation with IC50s in the nM- low µM range, whereas a DHODH (dihydroorotate dehydrogenase and a putative kinase inhibitor displayed no activity. Furthermore, significant synergies were identified with erlotinib, imatinib, cisplatin, dasatinib and vincristine. Cluster analysis of the antimalarials based on their differential inhibition of the various cancer lines clearly segregated the synthetic peroxides OZ277 and OZ439 from the artemisinin cluster that included artesunate, dihydroartemisinin and artemisone, and from the DHFR inhibitors pyrimethamine and P218 (a parasite DHFR inhibitor, emphasizing their shared mode of action. In order to further understand the basis of the selectivity of these compounds against different cancers, microarray-based gene expression data for 85 of the used cell lines were generated. For each compound, distinct sets of genes were identified whose expression significantly correlated with compound sensitivity. Several of the antimalarials tested in this study have well-established and excellent safety profiles with a plasma exposure, when conservatively used in malaria, that is well above the IC50s that we identified in this study. Given their unique mode of action and potential for unique synergies with established anticancer drugs, our results provide a strong basis to further explore the potential application of these compounds in cancer in pre-clinical or and clinical settings.
Morphogenesis of respiratory syncytial virus in human primary nasal ciliated epithelial cells occurs at surface membrane microdomains that are distinct from cilia

International Nuclear Information System (INIS)

Jumat, Muhammad Raihan; Yan, Yan; Ravi, Laxmi Iyer; Wong, Puisan; Huong, Tra Nguyen; Li, Chunwei; Tan, Boon Huan; Wang, De Yun; Sugrue, Richard J.

2015-01-01

The distribution of cilia and the respiratory syncytial virus (RSV) nucleocapsid (N) protein, fusion (F) protein, attachment (G) protein, and M2-1 protein in human ciliated nasal epithelial cells was examined at between 1 and 5 days post-infection (dpi). All virus structural proteins were localized at cell surface projections that were distinct from cilia. The F protein was also trafficked into the cilia, and while its presence increased as the infection proceeded, the N protein was not detected in the cilia at any time of infection. The presence of the F protein in the cilia correlated with cellular changes in the cilia and reduced cilia function. At 5 dpi extensive cilia loss and further reduced cilia function was noted. These data suggested that although RSV morphogenesis occurs at non-cilia locations on ciliated nasal epithelial cells, RSV infection induces changes in the cilia body that leads to extensive cilia loss. - Highlights: • Respiratory syncytial virus (RSV) infects nasal ciliated epithelial cells. • Virus morphogenesis occurs within filamentous projections distinct from cilia. • The RSV N protein was not detected in the cilia at any time during infection. • Trafficking of the F protein into the cilia occurred early in infection. • Presence of the F protein in cilia correlated with impaired cilia function
Morphogenesis of respiratory syncytial virus in human primary nasal ciliated epithelial cells occurs at surface membrane microdomains that are distinct from cilia

Energy Technology Data Exchange (ETDEWEB)

Jumat, Muhammad Raihan [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); Yan, Yan [Department of Otolaryngology, Yong Loo Lin School of Medicine, National University Health System, National University of Singapore, Singapore 119228 (Singapore); Ravi, Laxmi Iyer [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); Wong, Puisan [Detection and Diagnostics Laboratory, DSO National Laboratories, 27 Medical Drive, Singapore 117510 (Singapore); Huong, Tra Nguyen [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); Li, Chunwei [Department of Otolaryngology, Yong Loo Lin School of Medicine, National University Health System, National University of Singapore, Singapore 119228 (Singapore); Tan, Boon Huan [Detection and Diagnostics Laboratory, DSO National Laboratories, 27 Medical Drive, Singapore 117510 (Singapore); Wang, De Yun [Department of Otolaryngology, Yong Loo Lin School of Medicine, National University Health System, National University of Singapore, Singapore 119228 (Singapore); Sugrue, Richard J., E-mail: rjsugrue@ntu.edu.sg [School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore)

2015-10-15

The distribution of cilia and the respiratory syncytial virus (RSV) nucleocapsid (N) protein, fusion (F) protein, attachment (G) protein, and M2-1 protein in human ciliated nasal epithelial cells was examined at between 1 and 5 days post-infection (dpi). All virus structural proteins were localized at cell surface projections that were distinct from cilia. The F protein was also trafficked into the cilia, and while its presence increased as the infection proceeded, the N protein was not detected in the cilia at any time of infection. The presence of the F protein in the cilia correlated with cellular changes in the cilia and reduced cilia function. At 5 dpi extensive cilia loss and further reduced cilia function was noted. These data suggested that although RSV morphogenesis occurs at non-cilia locations on ciliated nasal epithelial cells, RSV infection induces changes in the cilia body that leads to extensive cilia loss. - Highlights: • Respiratory syncytial virus (RSV) infects nasal ciliated epithelial cells. • Virus morphogenesis occurs within filamentous projections distinct from cilia. • The RSV N protein was not detected in the cilia at any time during infection. • Trafficking of the F protein into the cilia occurred early in infection. • Presence of the F protein in cilia correlated with impaired cilia function.
Cathelicidin LL-37 Affects Surface and Intracellular Toll-Like Receptor Expression in Tissue Mast Cells

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Justyna Agier

2018-01-01

Full Text Available Undoubtedly, mast cells take part in host defense against microorganisms as they are numerous at the portal of infection, they release many proinflammatory and antimicrobial mediators, and they express pattern recognition receptors, such as TLRs. These receptors play a key role in recognition and binding molecules associated with microorganisms and molecules associated with damage. Cathelicidins exhibit direct antimicrobial activities against a broad spectrum of microbes by perturbing their cell membranes. Accumulating evidence suggests a role for these molecules in supporting cell activation. We examined the impact of human cathelicidin LL-37 on tissue mast cell TLR expression and distribution. Depending on context, we show that LL-37 stimulation resulted in minor to major effects on TLR2, TLR3, TLR4, TLR5, TLR7, and TLR9 expression. Confocal microscopy analysis showed that, upon stimulation, TLRs may translocate from the cell interior to the surface and conversely. FPR2 and EGFR inhibitors reduced the increase in expression of selected receptors. We also established that LL-37 acts as a powerful inducer of CCL3 and ROS generation. These results showed that in response to LL-37, mast cells enhance the capability to detect invading pathogens by modulation of TLR expression in what may be involved FPR2 or EGFR molecules.
CD45RC isoform expression identifies functionally distinct T cell subsets differentially distributed between healthy individuals and AAV patients.

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Laurence Ordonez

Full Text Available In animal models of anti-neutrophil cytoplasmic antibody (ANCA-associated vasculitis (AAV, the proportion of CD45RC T cell subsets is important for disease susceptibility. Their human counterparts are, however, functionally ill defined. In this report, we studied their distribution in healthy controls (HC, AAV patients and in Systemic lupus erythematous (SLE patients as disease controls. We showed that CD45RC expression level on human CD4 and CD8 T cells identifies subsets that are highly variable among individuals. Interestingly, AAV patients exhibit an increased proportion of CD45RC(low CD4 T cells as compared to HC and SLE patients. This increase is stable over time and independent of AAV subtype, ANCA specificity, disease duration, or number of relapses. We also analyzed the cytokine profile of purified CD4 and CD8 CD45RC T cell subsets from HC, after stimulation with anti-CD3 and anti-CD28 mAbs. The CD45RC subsets exhibit different cytokine profiles. Type-1 cytokines (IL-2, IFN-gamma and TNF-alpha were produced by all CD45RC T cell subsets, while the production of IL-17, type-2 (IL-4, IL-5 and regulatory (IL-10 cytokines was restricted to the CD45RC(low subset. In conclusion, we have shown that CD45RC expression divides human T cells in functionally distinct subsets that are imbalanced in AAV. Since this imbalance is stable over time and independent of several disease parameters, we hypothesize that this is a pre-existing immune abnormality involved in the etiology of AAV.
Distinct cardiac transcriptional profiles defining pregnancy and exercise.

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Eunhee Chung

Full Text Available BACKGROUND: Although the hypertrophic responses of the heart to pregnancy and exercise are both considered to be physiological processes, they occur in quite different hormonal and temporal settings. In this study, we have compared the global transcriptional profiles of left ventricular tissues at various time points during the progression of hypertrophy in exercise and pregnancy. METHODOLOGY/PRINCIPAL FINDINGS: The following groups of female mice were analyzed: non-pregnant diestrus cycle sedentary control, mid-pregnant, late-pregnant, and immediate-postpartum, and animals subjected to 7 and 21 days of voluntary wheel running. Hierarchical clustering analysis shows that while mid-pregnancy and both exercise groups share the closest relationship and similar gene ontology categories, late pregnancy and immediate post-partum are quite different with high representation of secreted/extracellular matrix-related genes. Moreover, pathway-oriented ontological analysis shows that metabolism regulated by cytochrome P450 and chemokine pathways are the most significant signaling pathways regulated in late pregnancy and immediate-postpartum, respectively. Finally, increases in expression of components of the proteasome observed in both mid-pregnancy and immediate-postpartum also result in enhanced proteasome activity. Interestingly, the gene expression profiles did not correlate with the degree of cardiac hypertrophy observed in the animal groups, suggesting that distinct pathways are employed to achieve similar amounts of cardiac hypertrophy. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that cardiac adaptation to the later stages of pregnancy is quite distinct from both mid-pregnancy and exercise. Furthermore, it is very dynamic since, by 12 hours post-partum, the heart has already initiated regression of cardiac growth, and 50 genes have changed expression significantly in the immediate-postpartum compared to late-pregnancy. Thus, pregnancy
Temporal expression and localization patterns of variant surface antigens in clinical Plasmodium falciparum isolates during erythrocyte schizogony.

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Anna Bachmann

Full Text Available Avoidance of antibody-mediated immune recognition allows parasites to establish chronic infections and enhances opportunities for transmission. The human malaria parasite Plasmodium falciparum possesses a number of multi-copy gene families, including var, rif, stevor and pfmc-2tm, which encode variant antigens believed to be expressed on the surfaces of infected erythrocytes. However, most studies of these antigens are based on in vitro analyses of culture-adapted isolates, most commonly the laboratory strain 3D7, and thus may not be representative of the unique challenges encountered by P. falciparum in the human host. To investigate the expression of the var, rif-A, rif-B, stevor and pfmc-2tm family genes under conditions that mimic more closely the natural course of infection, ex vivo clinical P. falciparum isolates were analyzed using a novel quantitative real-time PCR approach. Expression patterns in the clinical isolates at various time points during the first intraerythrocytic developmental cycle in vitro were compared to those of strain 3D7. In the clinical isolates, in contrast to strain 3D7, there was a peak of expression of the multi-copy gene families rif-A, stevor and pfmc-2tm at the young ring stage, in addition to the already known expression peak in trophozoites. Furthermore, most of the variant surface antigen families were overexpressed in the clinical isolates relative to 3D7, with the exception of the pfmc-2tm family, expression of which was higher in 3D7 parasites. Immunofluorescence analyses performed in parallel revealed two stage-dependent localization patterns of RIFIN, STEVOR and PfMC-2TM. Proteins were exported into the infected erythrocyte at the young trophozoite stage, whereas they remained inside the parasite membrane during schizont stage and were subsequently observed in different compartments in the merozoite. These results reveal a complex pattern of expression of P. falciparum multi-copy gene families during

Inhibitory effects of two G protein-coupled receptor kinases on the cell surface expression and signaling of the human adrenomedullin receptor

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Kuwasako, Kenji, E-mail: kuwasako@med.miyazaki-u.ac.jp [Frontier Science Research Center, University of Miyazaki, Miyazaki, 889-1692 (Japan); Sekiguchi, Toshio [Noto Marine Laboratory, Division of Marine Environmental Studies, Institute of Nature and Environmental Technology, Kanazawa University, Ishikawa, 927-0553 (Japan); Nagata, Sayaka [Division of Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Miyazaki, 889-1692 (Japan); Jiang, Danfeng; Hayashi, Hidetaka [Frontier Science Research Center, University of Miyazaki, Miyazaki, 889-1692 (Japan); Murakami, Manabu [Department of Pharmacology, Hirosaki University, Graduate School of Medicine, Hirosaki, 036-8562 (Japan); Hattori, Yuichi [Department of Molecular and Medical Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, 930-0194 (Japan); Kitamura, Kazuo [Division of Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Miyazaki, 889-1692 (Japan); Kato, Johji [Frontier Science Research Center, University of Miyazaki, Miyazaki, 889-1692 (Japan)

2016-02-19

Receptor activity-modifying protein 2 (RAMP2) enables the calcitonin receptor-like receptor (CLR, a family B GPCR) to form the type 1 adrenomedullin receptor (AM{sub 1} receptor). Here, we investigated the effects of the five non-visual GPCR kinases (GRKs 2 through 6) on the cell surface expression of the human (h)AM{sub 1} receptor by cotransfecting each of these GRKs into HEK-293 cells that stably expressed hRAMP2. Flow cytometric analysis revealed that when coexpressed with GRK4 or GRK5, the cell surface expression of the AM{sub 1} receptor was markedly decreased prior to stimulation with AM, thereby attenuating both the specific [{sup 125}I]AM binding and AM-induced cAMP production. These inhibitory effects of both GRKs were abolished by the replacement of the cytoplasmic C-terminal tail (C-tail) of CLR with that of the calcitonin receptor (a family B GPCR) or β{sub 2}-adrenergic receptor (a family A GPCR). Among the sequentially truncated CLR C-tail mutants, those lacking the five residues 449–453 (Ser-Phe-Ser-Asn-Ser) abolished the inhibition of the cell surface expression of CLR via the overexpression of GRK4 or GRK5. Thus, we provided new insight into the function of GRKs in agonist-unstimulated GPCR trafficking using a recombinant AM{sub 1} receptor and further determined the region of the CLR C-tail responsible for this GRK function. - Highlights: • We discovered a novel function of GRKs in GPCR trafficking using human CLR/RAMP2. • GRKs 4 and 5 markedly inhibited the cell surface expression of human CLR/RAMP2. • Both GRKs exhibited highly significant receptor signaling inhibition. • Five residues of the C-terminal tail of CLR govern this function of GRKs.
Inhibitory effects of two G protein-coupled receptor kinases on the cell surface expression and signaling of the human adrenomedullin receptor

International Nuclear Information System (INIS)

Kuwasako, Kenji; Sekiguchi, Toshio; Nagata, Sayaka; Jiang, Danfeng; Hayashi, Hidetaka; Murakami, Manabu; Hattori, Yuichi; Kitamura, Kazuo; Kato, Johji

2016-01-01

Receptor activity-modifying protein 2 (RAMP2) enables the calcitonin receptor-like receptor (CLR, a family B GPCR) to form the type 1 adrenomedullin receptor (AM_1 receptor). Here, we investigated the effects of the five non-visual GPCR kinases (GRKs 2 through 6) on the cell surface expression of the human (h)AM_1 receptor by cotransfecting each of these GRKs into HEK-293 cells that stably expressed hRAMP2. Flow cytometric analysis revealed that when coexpressed with GRK4 or GRK5, the cell surface expression of the AM_1 receptor was markedly decreased prior to stimulation with AM, thereby attenuating both the specific ["1"2"5I]AM binding and AM-induced cAMP production. These inhibitory effects of both GRKs were abolished by the replacement of the cytoplasmic C-terminal tail (C-tail) of CLR with that of the calcitonin receptor (a family B GPCR) or β_2-adrenergic receptor (a family A GPCR). Among the sequentially truncated CLR C-tail mutants, those lacking the five residues 449–453 (Ser-Phe-Ser-Asn-Ser) abolished the inhibition of the cell surface expression of CLR via the overexpression of GRK4 or GRK5. Thus, we provided new insight into the function of GRKs in agonist-unstimulated GPCR trafficking using a recombinant AM_1 receptor and further determined the region of the CLR C-tail responsible for this GRK function. - Highlights: • We discovered a novel function of GRKs in GPCR trafficking using human CLR/RAMP2. • GRKs 4 and 5 markedly inhibited the cell surface expression of human CLR/RAMP2. • Both GRKs exhibited highly significant receptor signaling inhibition. • Five residues of the C-terminal tail of CLR govern this function of GRKs.
Classification, expression pattern and comparative analysis of sugarcane expressed sequences tags (ESTs encoding glycine-rich proteins (GRPs

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Fusaro Adriana

2001-01-01

Full Text Available Since the isolation of the first glycine-rich proteins (GRPs in plants a wealth of new GRPs have been identified. The highly specific but diverse expression pattern of grp genes, taken together with the distinct sub-cellular localization of some GRP groups, clearly indicate that these proteins are involved in several independent physiological processes. Notwithstanding the absence of a clear definition of the role of GRPs in plant cells, studies conducted with these proteins have provided new and interesting insights into the molecular biology and cell biology of plants. Complexly regulated promoters and distinct mechanisms for the regulation of gene expression have been demonstrated and new protein targeting pathways, as well as the exportation of GRPs from different cell types have been discovered. These data show that GRPs can be useful as markers and/or models to understand distinct aspects of plant biology. In this paper, the structural and functional features of these proteins in sugarcane (Saccharum officinarum L. are summarized. Since this is the first description of GRPs in sugarcane, special emphasis has been given to the expression pattern of these GRP genes by studying their abundance and prevalence in the different cDNA-libraries of the Sugarcane Expressed Sequence Tag (SUCEST project . The comparison of sugarcane GRPs with GRPs from other species is also discussed.
Generation of murine tumor cell lines deficient in MHC molecule surface expression using the CRISPR/Cas9 system.

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Krishna Das

Full Text Available In this study, the CRISPR/Cas9 technology was used to establish murine tumor cell lines, devoid of MHC I or MHC II surface expression, respectively. The melanoma cell line B16F10 and the murine breast cancer cell line EO-771, the latter stably expressing the tumor antigen NY-BR-1 (EO-NY, were transfected with an expression plasmid encoding a β2m-specific single guide (sgRNA and Cas9. The resulting MHC I negative cells were sorted by flow cytometry to obtain single cell clones, and loss of susceptibility of peptide pulsed MHC I negative clones to peptide-specific CTL recognition was determined by IFNγ ELISpot assay. The β2m knockout (KO clones did not give rise to tumors in syngeneic mice (C57BL/6N, unless NK cells were depleted, suggesting that outgrowth of the β2m KO cell lines was controlled by NK cells. Using sgRNAs targeting the β-chain encoding locus of the IAb molecule we also generated several B16F10 MHC II KO clones. Peptide loaded B16F10 MHC II KO cells were insusceptible to recognition by OT-II cells and tumor growth was unaltered compared to parental B16F10 cells. Thus, in our hands the CRISPR/Cas9 system has proven to be an efficient straight forward strategy for the generation of MHC knockout cell lines. Such cell lines could serve as parental cells for co-transfection of compatible HLA alleles together with human tumor antigens of interest, thereby facilitating the generation of HLA matched transplantable tumor models, e.g. in HLAtg mouse strains of the newer generation, lacking cell surface expression of endogenous H2 molecules. In addition, our tumor cell lines established might offer a useful tool to investigate tumor reactive T cell responses that function independently from MHC molecule surface expression by the tumor.
A novel thromboxane A2 receptor N42S variant results in reduced surface expression and platelet dysfunction.

Science.gov (United States)

Nisar, Shaista P; Lordkipanidzé, Marie; Jones, Matthew L; Dawood, Ban; Murden, Sherina; Cunningham, Margaret R; Mumford, Andrew D; Wilde, Jonathan T; Watson, Steve P; Mundell, Stuart J; Lowe, Gillian C

2014-05-05

A small number of thromboxane receptor variants have been described in patients with a bleeding history that result in platelet dysfunction. We have identified a patient with a history of significant bleeding, who expresses a novel heterozygous thromboxane receptor variant that predicts an asparagine to serine substitution (N42S). This asparagine is conserved across all class A GPCRs, suggesting a vital role for receptor structure and function.We investigated the functional consequences of the TP receptor heterozygous N42S substitution by performing platelet function studies on platelet-rich plasma taken from the patient and healthy controls. We investigated the N42S mutation by expressing the wild-type (WT) and mutant receptor in human embryonic kidney (HEK) cells. Aggregation studies showed an ablation of arachidonic acid responses in the patient, whilst there was right-ward shift of the U46619 concentration response curve (CRC). Thromboxane generation was unaffected. Calcium mobilisation studies in cells lines showed a rightward shift of the U46619 CRC in N42S-expressing cells compared to WT. Radioligand binding studies revealed a reduction in BMax in platelets taken from the patient and in N42S-expressing cells, whilst cell studies confirmed poor surface expression. We have identified a novel thromboxane receptor variant, N42S, which results in platelet dysfunction due to reduced surface expression. It is associated with a significant bleeding history in the patient in whom it was identified. This is the first description of a naturally occurring variant that results in the substitution of this highly conserved residue and confirms the importance of this residue for correct GPCR function.
The Cell Surface Markers Expression in Postmenopausal Women and Relation to Obesity and Bone Status

OpenAIRE

Horv?thov?, Mira; Ilavsk?, Silvia; ?tef?kov?, Korn?lia; Szabov?, Michaela; Krivo??kov?, Zora; Jahnov?, Eva; Tulinsk?, Jana; Spustov?, Viera; Gajdo?, Martin

2017-01-01

The age-related changes and hormonal deprivation in postmenopausal women are associated with the immune response alteration. The excessive fat accumulation, local and systemic inflammation may lead to dysregulation in immune function and relevant health problems, including obesity and osteoporosis. We analyzed the expression of cell surface markers in the venous blood specimens, stained with fluorophores-conjugated monoclonal antibodies and analysed by multicolour flow cytometry. The signific...
Distinct spatiotemporal expression of serine proteases Prss23 and Prss35 in periimplantation mouse uterus and dispensable function of Prss35 in fertility.

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Honglu Diao

Full Text Available PRSS23 and PRSS35 are homologous proteases originally identified in mouse ovaries. In the periimplantation mouse uterus, Prss23 was highly expressed in the preimplantation gestation day 3.5 (D3.5 uterine luminal epithelium (LE. It disappeared from the postimplantation LE and reappeared in the stromal compartment next to the myometrium on D6.5. It was undetectable in the embryo from D4.5 to D6.5 but highly expressed in the embryo on D7.5. Prss35 became detectable in the uterine stromal compartment surrounding the embryo on D4.5 and shifted towards the mesometrial side of the stromal compartment next to the embryo from D5.5 to D7.5. In the ovariectomized uterus, Prss23 was moderately and Prss35 was dramatically downregulated by progesterone and 17β-estradiol. Based on the expression of Prss35 in granulosa cells and corpus luteum of the ovary and the early pregnant uterus, we hypothesized that PRSS35 might play a role in female reproduction, especially in oocyte development, ovulation, implantation, and decidualization. This hypothesis was tested in Prss35((-/- mice, which proved otherwise. Between wild type (WT and Prss35((-/- mice, superovulation of immature females produced comparable numbers of cumulus-oocyte complexes; there were comparable numbers of implantation sites detected on D4.5 and D7.5; there were no obvious differences in the expression of implantation and decidualization marker genes in D4.5 or D7.5 uteri. Comparable mRNA expression levels of a few known protease-related genes in the WT and Prss35((-/- D4.5 uteri indicated no compensatory upregulation. Comparable litter sizes from WT × WT and Prss35((-/-× Prss35((-/- crosses suggested that Prss35 gene was unessential for fertility and embryo development. Prss35 gene has been linked to cleft lip/palate in humans. However, no obvious such defects were observed in Prss35((-/- mice. This study demonstrates the distinct expression of Prss23 and Prss35 in the periimplantation uterus
Spleen-dependent regulation of antigenic variation in malaria parasites: Plasmodium knowlesi SICAvar expression profiles in splenic and asplenic hosts.

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Stacey A Lapp

Full Text Available Antigenic variation by malaria parasites was first described in Plasmodium knowlesi, which infects humans and macaque monkeys, and subsequently in P. falciparum, the most virulent human parasite. The schizont-infected cell agglutination (SICA variant proteins encoded by the SICAvar multigene family in P. knowlesi, and Erythrocyte Membrane Protein-1 (EMP-1 antigens encoded by the var multigene family in P. falciparum, are expressed at the surface of infected erythrocytes, are associated with virulence, and serve as determinants of naturally acquired immunity. A parental P. knowlesi clone, Pk1(A+, and a related progeny clone, Pk1(B+1+, derived by an in vivo induced variant antigen switch, were defined by the expression of distinct SICA variant protein doublets of 210/190 and 205/200 kDa, respectively. Passage of SICA[+] infected erythrocytes through splenectomized rhesus monkeys results in the SICA[-] phenotype, defined by the lack of surface expression and agglutination with variant specific antisera.We have investigated SICAvar RNA and protein expression in Pk1(A+, Pk1(B+1+, and SICA[-] parasites. The Pk1(A+ and Pk1(B+1+ parasites express different distinct SICAvar transcript and protein repertoires. By comparison, SICA[-] parasites are characterized by a vast reduction in SICAvar RNA expression, the lack of full-length SICAvar transcript signals on northern blots, and correspondingly, the absence of any SICA protein detected by mass spectrometry.SICA protein expression may be under transcriptional as well as post-transcriptional control, and we show for the first time that the spleen, an organ central to blood-stage immunity in malaria, exerts an influence on these processes. Furthermore, proteomics has enabled the first in-depth characterization of SICA[+] protein phenotypes and we show that the in vivo switch from Pk1(A+ to Pk1(B+1+ parasites resulted in a complete change in SICA profiles. These results emphasize the importance of studying
Phenotype specific analyses reveal distinct regulatory mechanism for chronically activated p53.

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Kristina Kirschner

2015-03-01

Full Text Available The downstream functions of the DNA binding tumor suppressor p53 vary depending on the cellular context, and persistent p53 activation has recently been implicated in tumor suppression and senescence. However, genome-wide information about p53-target gene regulation has been derived mostly from acute genotoxic conditions. Using ChIP-seq and expression data, we have found distinct p53 binding profiles between acutely activated (through DNA damage and chronically activated (in senescent or pro-apoptotic conditions p53. Compared to the classical 'acute' p53 binding profile, 'chronic' p53 peaks were closely associated with CpG-islands. Furthermore, the chronic CpG-island binding of p53 conferred distinct expression patterns between senescent and pro-apoptotic conditions. Using the p53 targets seen in the chronic conditions together with external high-throughput datasets, we have built p53 networks that revealed extensive self-regulatory 'p53 hubs' where p53 and many p53 targets can physically interact with each other. Integrating these results with public clinical datasets identified the cancer-associated lipogenic enzyme, SCD, which we found to be directly repressed by p53 through the CpG-island promoter, providing a mechanistic link between p53 and the 'lipogenic phenotype', a hallmark of cancer. Our data reveal distinct phenotype associations of chronic p53 targets that underlie specific gene regulatory mechanisms.
Comparison of Forced ENSO-Like Hydrological Expressions in Simulations of the Preindustrial and Mid-Holocene

Science.gov (United States)

Lewis, Sophie C.; LeGrande, Allegra N.; Schmidt, Gavin A.; Kelley, Maxwell

2014-01-01

Using the water isotope- and vapor source distribution (VSD) tracer-enabled Goddard Institute for Space Studies ModelE-R, we examine changing El Nino-Southern Oscillation (ENSO)-like expressions in the hydrological cycle in a suite of model experiments. We apply strong surface temperature anomalies associated with composite observed El Nino and La Nina events as surface boundary conditions to preindustrial and mid-Holocene model experiments in order to investigate ENSO-like expressions in the hydrological cycle under varying boundary conditions. We find distinct simulated hydrological anomalies associated with El Nino-like ("ENSOWARM") and La Nina-like ("ENSOCOOL") conditions, and the region-specific VSD tracers show hydrological differences across the Pacific basin between El Nino-like and La Nina-like events. The application of ENSOCOOL forcings does not produce climatological anomalies that represent the equal but opposite impacts of the ENSOWARM experiment, as the isotopic anomalies associated with ENSOWARM conditions are generally stronger than with ENSOCOOL and the spatial patterns of change distinct. Also, when the same ENSO-like surface temperature anomalies are imposed on the mid-Holocene, the hydrological response is muted, relative to the preindustrial. Mid-Holocene changes in moisture sources to the analyzed regions across the Pacific reveal potentially complex relationships between ENSO-like conditions and boundary conditions. Given the complex impacts of ENSO-like conditions on various aspects of the hydrological cycle, we suggest that proxy record insights into paleo-ENSO variability are most likely to be robust when synthesized from a network of many spatially diverse archives, which can account for the potential nonstationarity of ENSO teleconnections under different boundary conditions.
Proteomic plasma membrane profiling reveals an essential role for gp96 in the cell surface expression of LDLR family members, including the LDL receptor and LRP6.

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Weekes, Michael P; Antrobus, Robin; Talbot, Suzanne; Hör, Simon; Simecek, Nikol; Smith, Duncan L; Bloor, Stuart; Randow, Felix; Lehner, Paul J

2012-03-02

The endoplasmic reticulum chaperone gp96 is required for the cell surface expression of a narrow range of proteins, including toll-like receptors (TLRs) and integrins. To identify a more comprehensive repertoire of proteins whose cell surface expression is dependent on gp96, we developed plasma membrane profiling (PMP), a technique that combines SILAC labeling with selective cell surface aminooxy-biotinylation. This approach allowed us to compare the relative abundance of plasma membrane (PM) proteins on gp96-deficient versus gp96-reconstituted murine pre-B cells. Analysis of unfractionated tryptic peptides initially identified 113 PM proteins, which extended to 706 PM proteins using peptide prefractionation. We confirmed a requirement for gp96 in the cell surface expression of certain TLRs and integrins and found a marked decrease in cell surface expression of four members of the extended LDL receptor family (LDLR, LRP6, Sorl1 and LRP8) in the absence of gp96. Other novel gp96 client proteins included CD180/Ly86, important in the B-cell response to lipopolysaccharide. We highlight common structural motifs in these client proteins that may be recognized by gp96, including the beta-propeller and leucine-rich repeat. This study therefore identifies the extended LDL receptor family as an important new family of proteins whose cell surface expression is regulated by gp96.
Surface Expression of TGFβ Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer.

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Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip H; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-12-15

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGFβ, which is expressed naturally by platelets and regulatory T cells (Treg). Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here, we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGFβ in the tumor microenvironment. We found that human breast, lung, and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGFβ bioactivity and promoted malignant transformation in immunodeficient mice. In breast carcinoma-bearing mice that were immunocompetent, GARP overexpression promoted Foxp3 + Treg activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a GARP-specific mAb limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP-TGFβ axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. Cancer Res; 76(24); 7106-17. ©2016 AACR. ©2016 American Association for Cancer Research.
Altered expression of epithelial cell surface glycoconjugates and intermediate filaments at the margins of mucosal wounds

DEFF Research Database (Denmark)

Dabelsteen, Erik; Grøn, B.; Mandel, U.

1998-01-01

Alterations in cell to cell adhesion are necessary to enable the type of cell movements that are associated with epithelial wound healing and malignant invasion. Several studies of transformed cells have related epithelial cell movement to changes in the cell surface expression of the carbohydrate......-T antigen. The changes induced by wounding in the expression of collagen IV, laminin gamma2-chain (laminin-5), and laminin alpha5-chain were similar to those found in skin wounds and served to define the region of epithelial movement. This region was found to show a marked increase in staining for both...... epithelium, a pattern of expression similar to K16, which was also strongly upregulated in both the outgrowth and the adjacent nonwounded epithelium. These findings provide further support for an influence of such carbohydrate structures on the migratory behavior of epithelial cells....
Mesenchymal stromal cells express GARP/LRRC32 on their surface: effects on their biology and immunomodulatory capacity.

Science.gov (United States)

Carrillo-Galvez, Ana Belén; Cobo, Marién; Cuevas-Ocaña, Sara; Gutiérrez-Guerrero, Alejandra; Sánchez-Gilabert, Almudena; Bongarzone, Pierpaolo; García-Pérez, Angélica; Muñoz, Pilar; Benabdellah, Karim; Toscano, Miguel G; Martín, Francisco; Anderson, Per

2015-01-01

Mesenchymal stromal cells (MSCs) represent a promising tool for therapy in regenerative medicine, transplantation, and autoimmune disease due to their trophic and immunomodulatory activities. However, we are still far from understanding the mechanisms of action of MSCs in these processes. Transforming growth factor (TGF)-β1 is a pleiotropic cytokine involved in MSC migration, differentiation, and immunomodulation. Recently, glycoprotein A repetitions predominant (GARP) was shown to bind latency-associated peptide (LAP)/TGF-β1 to the cell surface of activated Foxp3(+) regulatory T cells (Tregs) and megakaryocytes/platelets. In this manuscript, we show that human and mouse MSCs express GARP which presents LAP/TGF-β1 on their cell surface. Silencing GARP expression in MSCs increased their secretion and activation of TGF-β1 and reduced their proliferative capacity in a TGF-β1-independent manner. Importantly, we showed that GARP expression on MSCs contributed to their ability to inhibit T-cell responses in vitro. In summary, we have found that GARP is an essential molecule for MSC biology, regulating their immunomodulatory and proliferative activities. We envision GARP as a new target for improving the therapeutic efficacy of MSCs and also as a novel MSC marker. © 2014 AlphaMed Press.
Kinetoplastid membrane protein-11 is present in promastigotes and amastigotes of Leishmania amazonensis and its surface expression increases during metacyclogenesis

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Denise CS Matos

2010-05-01

Full Text Available Kinetoplastid membrane protein-11 (KMP-11, a protein present in all kinetoplastid protozoa, is considered a potential candidate for a leishmaniasis vaccine. A suitable leishmaniasis vaccine candidate molecule must be expressed in amastigotes, the infective stage for mammals. However, the expression of KMP-11 in Leishmania amastigotes has been a subject of controversy. We evaluated the expression of this molecule in logarithmic and stationary growth phase promastigotes, as well as in amastigotes, of Leishmania amazonensis by immunoblotting, flow cytometry and immunocytochemistry, using a monoclonal antibody against KMP-11. We found that KMP-11 is present in promastigotes and amastigotes. In both stages, the protein was found in association with membrane structures (at the cell surface, flagellar pocket and intracellular vesicles. More importantly, its surface expression is higher in amastigotes than in promastigotes and increases during metacyclogenesis. The increased expression of KMP-11 in metacyclic promastigotes, and especially in amastigotes, indicates a role for this molecule in the parasite relationship with the mammalian host. The presence of this molecule in amastigotes is consistent with the previously demonstrated immunoprotective capacity of vaccine prototypes based on the KMP-11-coding gene and the presence of humoral and cellular immune responses to KMP-11 in Leishmania-infected humans and animals.
N-linked glycans are required on epithelial Na+ channel subunits for maturation and surface expression.

Science.gov (United States)

Kashlan, Ossama B; Kinlough, Carol L; Myerburg, Michael M; Shi, Shujie; Chen, Jingxin; Blobner, Brandon M; Buck, Teresa M; Brodsky, Jeffrey L; Hughey, Rebecca P; Kleyman, Thomas R

2018-03-01

Epithelial Na + channel (ENaC) subunits undergo N-linked glycosylation in the endoplasmic reticulum where they assemble into an αβγ complex. Six, 13, and 5 consensus sites (Asn-X-Ser/Thr) for N-glycosylation reside in the extracellular domains of the mouse α-, β-, and γ-subunits, respectively. Because the importance of ENaC N-linked glycans has not been fully addressed, we examined the effect of preventing N-glycosylation of specific subunits on channel function, expression, maturation, and folding. Heterologous expression in Xenopus oocytes or Fischer rat thyroid cells with αβγ-ENaC lacking N-linked glycans on a single subunit reduced ENaC activity as well as the inhibitory response to extracellular Na + . The lack of N-linked glycans on the β-subunit also precluded channel activation by trypsin. However, channel activation by shear stress was N-linked glycan independent, regardless of which subunit was modified. We also discovered that the lack of N-linked glycans on any one subunit reduced the total and surface levels of cognate subunits. The lack of N-linked glycans on the β-subunit had the largest effect on total levels, with the lack of N-linked glycans on the γ- and α-subunits having intermediate and modest effects, respectively. Finally, channels with wild-type β-subunits were more sensitive to limited trypsin proteolysis than channels lacking N-linked glycans on the β-subunit. Our results indicate that N-linked glycans on each subunit are required for proper folding, maturation, surface expression, and function of the channel.
Analysis of gene expression data from non-small cell lung carcinoma cell lines reveals distinct sub-classes from those identified at the phenotype level.

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Andrew R Dalby

Full Text Available Microarray data from cell lines of Non-Small Cell Lung Carcinoma (NSCLC can be used to look for differences in gene expression between the cell lines derived from different tumour samples, and to investigate if these differences can be used to cluster the cell lines into distinct groups. Dividing the cell lines into classes can help to improve diagnosis and the development of screens for new drug candidates. The micro-array data is first subjected to quality control analysis and then subsequently normalised using three alternate methods to reduce the chances of differences being artefacts resulting from the normalisation process. The final clustering into sub-classes was carried out in a conservative manner such that sub-classes were consistent across all three normalisation methods. If there is structure in the cell line population it was expected that this would agree with histological classifications, but this was not found to be the case. To check the biological consistency of the sub-classes the set of most strongly differentially expressed genes was be identified for each pair of clusters to check if the genes that most strongly define sub-classes have biological functions consistent with NSCLC.
Cloning and expression of the receptor for human urokinase plasminogen activator, a central molecule in cell surface, plasmin dependent proteolysis

DEFF Research Database (Denmark)

Roldan, A.L.; Cubellis, M.V.; Masucci, M.T.

1990-01-01

, and therefore the capacity of cells to migrate and invade neighboring tissues. We have isolated a 1.4 kb cDNA clone coding for the entire human uPAR. An oligonucleotide synthesized on the basis of the N-terminal sequence of the purified protein was used to screen a cDNA library made from SV40 transformed human......, a size very close to that of the cloned cDNA. Expression of the uPAR cDNA in mouse cells confirms that the clone is complete and expresses a functional uPA binding protein, located on the cell surface and with properties similar to the human uPAR. Caseinolytic plaque assay, immunofluorescence analysis......The surface receptor for urokinase plasminogen activator (uPAR) has been recognized in recent years as a key molecule in regulating plasminogen mediated extracellular proteolysis. Surface plasminogen activation controls the connections between cells, basement membrane and extracellular matrix...
Immunization of dogs with a canine herpesvirus vector expressing Neospora caninum surface protein, NcSRS2.

Science.gov (United States)

Nishikawa, Y; Ikeda, H; Fukumoto, S; Xuan, X; Nagasawa, H; Otsuka, H; Mikami, T

2000-10-01

In order to develop a vaccine against Neospora caninum in dogs, we constructed recombinant canine herpesvirus (CHV) expressing N. caninum surface protein, NcSRS2. Indirect immunofluorescence indicated that the antigenic structure of the recombinant NcSRS2 was similar to the authentic parasite protein. The dogs immunised with recombinant virus produced IgG antibody to N. caninum, and their sera recognised the parasite protein on Western blot. The dogs inoculated with recombinant virus showed no clinical symptoms and infectious CHV was not recovered from the dogs, suggesting that recombinant CHV expressing N. caninum proteins may lead to a vaccine against neosporosis in dogs.
Transmembrane and ubiquitin-like domain-containing protein 1 (Tmub1/HOPS facilitates surface expression of GluR2-containing AMPA receptors.

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Hyunjeong Yang

Full Text Available Some ubiquitin-like (UBL domain-containing proteins are known to play roles in receptor trafficking. Alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs undergo constitutive cycling between the intracellular compartment and the cell surface in the central nervous system. However, the function of UBL domain-containing proteins in the recycling of the AMPARs to the synaptic surface has not yet been reported.Here, we report that the Transmembrane and ubiquitin-like domain-containing 1 (Tmub1 protein, formerly known as the Hepatocyte Odd Protein Shuttling (HOPS protein, which is abundantly expressed in the brain and which exists in a synaptosomal membrane fraction, facilitates the recycling of the AMPAR subunit GluR2 to the cell surface. Neurons transfected with Tmub1/HOPS-RNAi plasmids showed a significant reduction in the AMPAR current as compared to their control neurons. Consistently, the synaptic surface expression of GluR2, but not of GluR1, was significantly decreased in the neurons transfected with the Tmub1/HOPS-RNAi and increased in the neurons overexpressing EGFP-Tmub1/HOPS. The altered surface expression of GluR2 was speculated to be due to the altered surface-recycling of the internalized GluR2 in our recycling assay. Eventually, we found that GluR2 and glutamate receptor interacting protein (GRIP were coimmunoprecipitated by the anti-Tmub1/HOPS antibody from the mouse brain. Taken together, these observations show that the Tmub1/HOPS plays a role in regulating basal synaptic transmission; it contributes to maintain the synaptic surface number of the GluR2-containing AMPARs by facilitating the recycling of GluR2 to the plasma membrane.

Integrative Genomic Analysis of Cholangiocarcinoma Identifies Distinct IDH-Mutant Molecular Profiles

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Farshad Farshidfar

2017-03-01

Full Text Available Cholangiocarcinoma (CCA is an aggressive malignancy of the bile ducts, with poor prognosis and limited treatment options. Here, we describe the integrated analysis of somatic mutations, RNA expression, copy number, and DNA methylation by The Cancer Genome Atlas of a set of predominantly intrahepatic CCA cases and propose a molecular classification scheme. We identified an IDH mutant-enriched subtype with distinct molecular features including low expression of chromatin modifiers, elevated expression of mitochondrial genes, and increased mitochondrial DNA copy number. Leveraging the multi-platform data, we observed that ARID1A exhibited DNA hypermethylation and decreased expression in the IDH mutant subtype. More broadly, we found that IDH mutations are associated with an expanded histological spectrum of liver tumors with molecular features that stratify with CCA. Our studies reveal insights into the molecular pathogenesis and heterogeneity of cholangiocarcinoma and provide classification information of potential therapeutic significance.
Quantum Distinction: Quantum Distinctiones!

OpenAIRE

Zeps, Dainis

2009-01-01

10 pages; How many distinctions, in Latin, quantum distinctiones. We suggest approach of anthropic principle based on anthropic reference system which should be applied equally both in theoretical physics and in mathematics. We come to principle that within reference system of life subject of mathematics (that of thinking) should be equated with subject of physics (that of nature). For this reason we enter notions of series of distinctions, quantum distinction, and argue that quantum distinct...
Initial attachment, subsequent cell proliferation/viability and gene expression of epithelial cells related to attachment and wound healing in response to different titanium surfaces.

Science.gov (United States)

An, Na; Rausch-fan, Xiaohui; Wieland, Marco; Matejka, Michael; Andrukhov, Oleh; Schedle, Andreas

2012-12-01

A tight seal between the epithelium and the dental implant surface is required to prevent bacterial inflammation and soft tissue recession and therefore to demonstrate a long-term success. Surface hydrophilicity was recently shown to promote osseointegration. The aim of this study was to investigate the influence of surface hydrophilicity in combination with surface topography of Ti implant surfaces on the behavior and activation/differentiation of epithelial cells using a set of in vitro experiments mimicking the implant-soft tissue contact. Hydrophobic acid-etched (A) and coarse-grit-blasted, acid-etched (SLA) surfaces and hydrophilic acid-etched (modA) and modSLA surfaces were produced. The behavior of an oral squamous cell carcinoma cell line (HSC-2) grown on all surfaces was compared through determination of cell attachment and proliferation/viability (CCK-8 and MTT assay), time-lapse microscopy of fluorescence labeled cells and determination of gene expression by real time polymerase chain reaction. Within the surfaces with similar wettability cell spreading and cell movements observed by time-lapse microscopy after one day of incubation were most pronounced on smoother (A and modA) surfaces compared to rougher (SLA and modSLA) surfaces. Within the surfaces with similar roughness the hydrophilic surfaces (modA and modSLA) showed more cell spreading and cell activity compared to the hydrophobic surfaces (A and SLA). The relative gene expressions of cytokeratin14, integrin α6, integrin β4, vinculin, transforming growth factor (TGF)-β, TGF-β1, and TGF-β3 were decreased in HSC-2 on all four types of Ti surfaces compared to control surfaces (tissue culture polystyrene; pmodA). These results suggest that surface hydrophilicity might positively influence the epithelial seal around dental implants. All tested titanium surfaces downregulate cell attachment, cell proliferation, expression of adhesion promoters, and cytokines involved in wound healing in HSC-2
Molecular subsets in the gene expression signatures of scleroderma skin.

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Ausra Milano

2008-07-01

Full Text Available Scleroderma is a clinically heterogeneous disease with a complex phenotype. The disease is characterized by vascular dysfunction, tissue fibrosis, internal organ dysfunction, and immune dysfunction resulting in autoantibody production.We analyzed the genome-wide patterns of gene expression with DNA microarrays in skin biopsies from distinct scleroderma subsets including 17 patients with systemic sclerosis (SSc with diffuse scleroderma (dSSc, 7 patients with SSc with limited scleroderma (lSSc, 3 patients with morphea, and 6 healthy controls. 61 skin biopsies were analyzed in a total of 75 microarray hybridizations. Analysis by hierarchical clustering demonstrates nearly identical patterns of gene expression in 17 out of 22 of the forearm and back skin pairs of SSc patients. Using this property of the gene expression, we selected a set of 'intrinsic' genes and analyzed the inherent data-driven groupings. Distinct patterns of gene expression separate patients with dSSc from those with lSSc and both are easily distinguished from normal controls. Our data show three distinct patient groups among the patients with dSSc and two groups among patients with lSSc. Each group can be distinguished by unique gene expression signatures indicative of proliferating cells, immune infiltrates and a fibrotic program. The intrinsic groups are statistically significant (p<0.001 and each has been mapped to clinical covariates of modified Rodnan skin score, interstitial lung disease, gastrointestinal involvement, digital ulcers, Raynaud's phenomenon and disease duration. We report a 177-gene signature that is associated with severity of skin disease in dSSc.Genome-wide gene expression profiling of skin biopsies demonstrates that the heterogeneity in scleroderma can be measured quantitatively with DNA microarrays. The diversity in gene expression demonstrates multiple distinct gene expression programs in the skin of patients with scleroderma.
Modulation of expression of HLA components at the cell surface induced by anti-β2m reagents

International Nuclear Information System (INIS)

Ceppellini, R.; Malavasi, F.; Garotta, G.; Trucco, M.

1981-01-01

Antibodies against lymphocyte surface components are able to rearrange profoundly the topography of the cell membrane with a different modulation of surface antigens. Of particular interest is the effect of anti-β2m reagents, which are able to suppress completely the reactivity of epitopes carried by the two chains of the ABC dimers, while the expressivity of other antigens, such as DR, is significantly increased. These results have been obtained with immunoradiobinding under a varity of conditions, thus confirming the validity of the ''bb'' (β2m blanketing) test. (author)
Polarized axonal surface expression of neuronal KCNQ potassium channels is regulated by calmodulin interaction with KCNQ2 subunit.

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John P Cavaretta

Full Text Available KCNQ potassium channels composed of KCNQ2 and KCNQ3 subunits give rise to the M-current, a slow-activating and non-inactivating voltage-dependent potassium current that limits repetitive firing of action potentials. KCNQ channels are enriched at the surface of axons and axonal initial segments, the sites for action potential generation and modulation. Their enrichment at the axonal surface is impaired by mutations in KCNQ2 carboxy-terminal tail that cause benign familial neonatal convulsion and myokymia, suggesting that their correct surface distribution and density at the axon is crucial for control of neuronal excitability. However, the molecular mechanisms responsible for regulating enrichment of KCNQ channels at the neuronal axon remain elusive. Here, we show that enrichment of KCNQ channels at the axonal surface of dissociated rat hippocampal cultured neurons is regulated by ubiquitous calcium sensor calmodulin. Using immunocytochemistry and the cluster of differentiation 4 (CD4 membrane protein as a trafficking reporter, we demonstrate that fusion of KCNQ2 carboxy-terminal tail is sufficient to target CD4 protein to the axonal surface whereas inhibition of calmodulin binding to KCNQ2 abolishes axonal surface expression of CD4 fusion proteins by retaining them in the endoplasmic reticulum. Disruption of calmodulin binding to KCNQ2 also impairs enrichment of heteromeric KCNQ2/KCNQ3 channels at the axonal surface by blocking their trafficking from the endoplasmic reticulum to the axon. Consistently, hippocampal neuronal excitability is dampened by transient expression of wild-type KCNQ2 but not mutant KCNQ2 deficient in calmodulin binding. Furthermore, coexpression of mutant calmodulin, which can interact with KCNQ2/KCNQ3 channels but not calcium, reduces but does not abolish their enrichment at the axonal surface, suggesting that apo calmodulin but not calcium-bound calmodulin is necessary for their preferential targeting to the axonal
Distinctive Features of Surface Winds over Indian Ocean Between Strong and Weak Indian Summer Monsoons: Implications With Respect To Regional Rainfall Change in India

Science.gov (United States)

Zheng, Y.; Bourassa, M. A.; Ali, M. M.

2017-12-01

This observational study focuses on characterizing the surface winds in the Arabian Sea (AS), the Bay of Bengal (BoB), and the southern Indian Ocean (SIO) with special reference to the strong and weak Indian summer monsoon rainfall (ISMR) using the latest daily gridded rainfall dataset provided by the Indian Meteorological Department (IMD) and the Cross-Calibrated Multi-Platform (CCMP) gridded wind product version 2.0 produced by Remote Sensing System (RSS) over the overlapped period 1991-2014. The potential links between surface winds and Indian regional rainfall are also examined. Results indicate that the surface wind speeds in AS and BoB during June-August are almost similar during strong ISMRs and weak ISMRs, whereas significant discrepancies are observed during September. By contrast, the surface wind speeds in SIO during June-August are found to be significantly different between strong and weak ISMRs, where they are similar during September. The significant differences in monthly mean surface wind convergence between strong and weak ISMRs are not coherent in space in the three regions. However, the probability density function (PDF) distributions of daily mean area-averaged values are distinctive between strong and weak ISMRs in the three regions. The correlation analysis indicates the area-averaged surface wind speeds in AS and the area-averaged wind convergence in BoB are highly correlated with regional rainfall for both strong and weak ISMRs. The wind convergence in BoB during strong ISMRs is relatively better correlated with regional rainfall than during weak ISMRs. The surface winds in SIO do not greatly affect Indian rainfall in short timescales, however, they will ultimately affect the strength of monsoon circulation by modulating Indian Ocean Dipole (IOD) mode via atmosphere-ocean interactions.
Lipopolysaccharide-induced expression of cell surface receptors and cell activation of neutrophils and monocytes in whole human blood

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N.E. Gomes

2010-09-01

Full Text Available Lipopolysaccharide (LPS activates neutrophils and monocytes, inducing a wide array of biological activities. LPS rough (R and smooth (S forms signal through Toll-like receptor 4 (TLR4, but differ in their requirement for CD14. Since the R-form LPS can interact with TLR4 independent of CD14 and the differential expression of CD14 on neutrophils and monocytes, we used the S-form LPS from Salmonella abortus equi and the R-form LPS from Salmonella minnesota mutants to evaluate LPS-induced activation of human neutrophils and monocytes in whole blood from healthy volunteers. Expression of cell surface receptors and reactive oxygen species (ROS and nitric oxide (NO generation were measured by flow cytometry in whole blood monocytes and neutrophils. The oxidative burst was quantified by measuring the oxidation of 2',7'-dichlorofluorescein diacetate and the NO production was quantified by measuring the oxidation of 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate. A small increase of TLR4 expression by monocytes was observed after 6 h of LPS stimulation. Monocyte CD14 modulation by LPS was biphasic, with an initial 30% increase followed by a 40% decrease in expression after 6 h of incubation. Expression of CD11b was rapidly up-regulated, doubling after 5 min on monocytes, while down-regulation of CXCR2 was observed on neutrophils, reaching a 50% reduction after 6 h. LPS induced low production of ROS and NO. This study shows a complex LPS-induced cell surface receptor modulation on human monocytes and neutrophils, with up- and down-regulation depending on the receptor. R- and S-form LPS activate human neutrophils similarly, despite the low CD14 expression, if the stimulation occurs in whole blood.
The cell surface expressed nucleolin is a glycoprotein that triggers calcium entry into mammalian cells

International Nuclear Information System (INIS)

Losfeld, Marie-Estelle; Khoury, Diala El; Mariot, Pascal; Carpentier, Mathieu; Krust, Bernard; Briand, Jean-Paul; Mazurier, Joel; Hovanessian, Ara G.; Legrand, Dominique

2009-01-01

Nucleolin is an ubiquitous nucleolar phosphoprotein involved in fundamental aspects of transcription regulation, cell proliferation and growth. It has also been described as a shuttling molecule between nucleus, cytosol and the cell surface. Several studies have demonstrated that surface nucleolin serves as a receptor for various extracellular ligands implicated in cell proliferation, differentiation, adhesion, mitogenesis and angiogenesis. Previously, we reported that nucleolin in the extranuclear cell compartment is a glycoprotein containing N- and O-glycans. In the present study, we show that glycosylation is an essential requirement for surface nucleolin expression, since it is prevented when cells are cultured in the presence of tunicamycin, an inhibitor of N-glycosylation. Accordingly, surface but not nuclear nucleolin is radioactively labeled upon metabolic labeling of cells with [ 3 H]glucosamine. Besides its well-demonstrated role in the internalization of specific ligands, here we show that ligand binding to surface nucleolin could also induce Ca 2+ entry into cells. Indeed, by flow cytometry, microscopy and patch-clamp experiments, we show that the HB-19 pseudopeptide, which binds specifically surface nucleolin, triggers rapid and intense membrane Ca 2+ fluxes in various types of cells. The use of several drugs then indicated that Store-Operated Ca 2+ Entry (SOCE)-like channels are involved in the generation of these fluxes. Taken together, our findings suggest that binding of an extracellular ligand to surface nucleolin could be involved in the activation of signaling pathways by promoting Ca 2+ entry into cells
Heterologous Expression of Three Plant Serpins with Distinct Inhibitory Specificities

DEFF Research Database (Denmark)

Dahl, Søren Weis; Rasmussen, Søren Kjærsgård; Hejgaard, Jørn

1996-01-01

For the first time, inhibitory plant serpins, including WSZ1 from wheat, BSZ4, and the previously unknown protein BSZx from barley, have been expressed in Escherichia coli, and a procedure for fast purification of native plant serpins has been developed, BSZx, BSZ4, and WSZ1 were assayed...... favorable P-2 Leu. BSZ4 inhibited cathepsin G (k(a) = 2.7 x 10(4) M(-1) s(-1)) at P-1 Met but was hydrolyzed by trypsin and chymotrypsin. The three plant serpins formed stable SDS-resistant complexes with the proteinases in accordance with the kinetic data....
The nicotinic acetylcholine receptors of the parasitic nematode Ascaris suum: formation of two distinct drug targets by varying the relative expression levels of two subunits.

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Sally M Williamson

2009-07-01

Full Text Available Parasitic nematodes are of medical and veterinary importance, adversely affecting human health and animal welfare. Ascaris suum is a gastrointestinal parasite of pigs; in addition to its veterinary significance it is a good model of the human parasite Ascaris lumbricoides, estimated to infect approximately 1.4 billion people globally. Anthelmintic drugs are essential to control nematode parasites, and nicotinic acetylcholine receptors (nAChRs on nerve and muscle are the targets of cholinergic anthelmintics such as levamisole and pyrantel. Previous genetic analyses of nematode nAChRs have been confined to Caenorhabditis elegans, which is phylogenetically distinct from Ascaris spp. and many other important parasites. Here we report the cloning and expression of two nAChR subunit cDNAs from A. suum. The subunits are very similar in sequence to C. elegans UNC-29 and UNC-38, are expressed on muscle cells and can be expressed robustly in Xenopus oocytes to form acetylcholine-, nicotine-, levamisole- and pyrantel-sensitive channels. We also demonstrate that changing the stoichiometry of the receptor by injecting different ratios of the subunit cRNAs can reproduce two of the three pharmacological subtypes of nAChR present in A. suum muscle cells. When the ratio was 5:1 (Asu-unc-38ratioAsu-unc-29, nicotine was a full agonist and levamisole was a partial agonist, and oocytes responded to oxantel, but not pyrantel. At the reverse ratio (1:5 Asu-unc-38ratioAsu-unc-29, levamisole was a full agonist and nicotine was a partial agonist, and the oocytes responded to pyrantel, but not oxantel. These results represent the first in vitro expression of any parasitic nicotinic receptor and show that their properties are substantially different from those of C. elegans. The results also show that changing the expression level of a single receptor subunit dramatically altered the efficacy of some anthelmintic drugs. In vitro expression of these subunits may permit the
Insect-cell expression, crystallization and X-ray data collection of the bradyzoite-specific antigen BSR4 from Toxoplasma gondii

International Nuclear Information System (INIS)

Grujic, Ognjen; Grigg, Michael E.; Boulanger, Martin J.

2008-01-01

Preliminary X-ray diffraction studies of the bradyzoite-specific surface antigen BSR4 from T. gondii are described. Toxoplasma gondii is an important global pathogen that infects nearly one third of the world’s adult population. A family of developmentally expressed structurally related surface-glycoprotein adhesins (SRSs) mediate attachment to and are utilized for entry into host cells. The latent bradyzoite form of T. gondii persists for the life of the host and expresses a distinct family of SRS proteins, of which the bradyzoite-specific antigen BSR4 is a prototypical member. Structural studies of BSR4 were initiated by first recombinantly expressing BSR4 in insect cells, which was followed by crystallization and preliminary X-ray data collection to 1.95 Å resolution. Data processing showed that BSR4 crystallized with one molecule in the asymmetric unit of the P4 1 2 1 2 or P4 3 2 1 2 space group, with a solvent content of 60% and a corresponding Matthews coefficient of 2.98 Å 3 Da −1
Cell-surface glycoproteins of human sarcomas: differential expression in normal and malignant tissues and cultured cells

International Nuclear Information System (INIS)

Rettig, W.F.; Garin-Chesa, P.; Beresford, H.R.; Oettgen, H.F.; Melamed, M.R.; Old, L.J.

1988-01-01

Normal differentiation and malignant transformation of human cells are characterized by specific changes in surface antigen phenotype. In the present study, the authors have defined six cell-surface antigens of human sarcomas and normal mesenchymal cells, by using mixed hemadsorption assays and immunochemical methods for the analysis of cultured cells and immunohistochemical staining for the analysis of normal tissues and > 200 tumor specimens. Differential patterns of F19, F24, G171, G253, S5, and Thy-1 antigen expression were found to characterize (i) subsets of cultured sarcoma cell lines, (ii) cultured fibroblasts derived from various organs, (iii) normal resting and activated mesenchymal tissues, and (iv) sarcoma and nonmesenchymal tumor tissues. These results provide a basic surface antigenic map for cultured mesenchymal cells and mesenchymal tissues and permit the classification of human sarcomas according to their antigenic phenotypes
Gene Expression Commons: an open platform for absolute gene expression profiling.

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Jun Seita

Full Text Available Gene expression profiling using microarrays has been limited to comparisons of gene expression between small numbers of samples within individual experiments. However, the unknown and variable sensitivities of each probeset have rendered the absolute expression of any given gene nearly impossible to estimate. We have overcome this limitation by using a very large number (>10,000 of varied microarray data as a common reference, so that statistical attributes of each probeset, such as the dynamic range and threshold between low and high expression, can be reliably discovered through meta-analysis. This strategy is implemented in a web-based platform named "Gene Expression Commons" (https://gexc.stanford.edu/ which contains data of 39 distinct highly purified mouse hematopoietic stem/progenitor/differentiated cell populations covering almost the entire hematopoietic system. Since the Gene Expression Commons is designed as an open platform, investigators can explore the expression level of any gene, search by expression patterns of interest, submit their own microarray data, and design their own working models representing biological relationship among samples.
Distinct neutrophil subpopulations phenotype by flow cytometry in myelodysplastic syndromes.

Science.gov (United States)

Vikentiou, Myrofora; Psarra, Katerina; Kapsimali, Violetta; Liapis, Konstantinos; Michael, Michalis; Tsionos, Konstantinos; Lianidou, Evi; Papasteriades, Chryssa

2009-03-01

The cardinal feature of myelodysplastic syndromes (MDS) is dysplasia involving one or more myeloid cell lineages. In the present study, we used 4-color flow cytometric analysis to investigate dysgranulopoiesis in bone marrow specimens from 65 patients with MDS. The antigen expression patterns of total neutrophil granulocytes (TNG) and of the two distinct neutrophil granulocytic subpopulations (NGSs), NGS-1 (dimmer CD45 expression) and NGS-2 (stronger CD45 expression) identified on the side scatter (SS) vs. CD45-intensity plot, were studied. The neutrophil granulocytes from patients with MDS showed characteristic antigen expression aberrancies which were more pronounced in NGS-2 subpopulation. Studying separately the NGS-2 subpopulation with the CD16/MPO/LF combination, the low CD16(+)/MPO(+) and low CD16(+)/LF(+) percentages seemed to discriminate between lower-risk and higher-risk patients with MDS in most occasions. Furthermore, a detailed assessment of the NGS-1 and NGS-2 immunophenotypic patterns revealed early dysplastic changes, not otherwise observed by standard TNG analysis, especially in cases of lower-risk MDS.
Motivation to hide emotion and children's understanding of the distinction between real and apparent emotions.

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Gosselin, Pierre; Warren, Madeleine; Diotte, Michèle

2002-12-01

The authors investigated the extent to which children's understanding of the distinction between real and apparent emotions varied according to the motivation to hide emotions. Children, aged 6-7 and 10-11 years, were read stories designed to elicit either prosocial or self-protective motivated display rules and were asked to predict the facial expressions the protagonists would make to hide felt emotions. Children were found to understand the distinction between real and apparent emotions very well, independently of the type of motivation. Contrary to predictions, boys understood this distinction better than did girls when the motivation to hide positive emotions was prosocial. Children perceived neutralization as the most appropriate strategy to hide felt emotions, followed by masking.
Monoclonal antibody DS6 detects a tumor-associated sialoglycotope expressed on human serous ovarian carcinomas.

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Kearse, K P; Smith, N L; Semer, D A; Eagles, L; Finley, J L; Kazmierczak, S; Kovacs, C J; Rodriguez, A A; Kellogg-Wennerberg, A E

2000-12-15

A newly developed murine monoclonal antibody, DS6, immunohistochemically reacts with an antigen, CA6, that is expressed by human serous ovarian carcinomas but not by normal ovarian surface epithelium or mesothelium. CA6 has a limited distribution in normal adult tissues and is most characteristically detected in fallopian tube epithelium, inner urothelium and type 2 pneumocytes. Pre-treatment of tissue sections with either periodic acid or neuraminidase from Vibrio cholerae abolishes immunoreactivity with DS6, indicating that CA6 is a neuraminidase-sensitive and periodic acid-sensitive sialic acid glycoconjugate ("sialoglycotope"). SDS-PAGE of OVCAR5 cell lysates has revealed that the CA6 epitope is expressed on an 80 kDa non-disulfide-linked glycoprotein containing N-linked oligosaccharides. Two-dimensional non-equilibrium pH gradient gel electrophoresis indicates an isoelectric point of approximately 6.2 to 6.5. Comparison of the immunohistochemical distribution of CA6 in human serous ovarian adenocarcinomas has revealed similarities to that of CA125; however, distinct differences and some complementarity of antigen expression were revealed by double-label, 2-color immunohistochemical studies. The DS6-detected CA6 antigen appears to be distinct from other well-characterized tumor-associated antigens, including MUC1, CA125 and the histo-blood group-related antigens sLea, sLex and sTn. Copyright 2000 Wiley-Liss, Inc.
The mitochondrial elongation factors MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics

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Liu, Tong; Yu, Rong [Department of Oncology–Pathology, Karolinska Institutet, CCK R8:05, Karolinska University Hospital Solna, SE-171 76 Stockholm (Sweden); Jin, Shao-Bo [Department of Cell and Molecular Biology, Karolinska Institutet, SE-171 77 Stockholm (Sweden); Han, Liwei [Department of Oncology–Pathology, Karolinska Institutet, CCK R8:05, Karolinska University Hospital Solna, SE-171 76 Stockholm (Sweden); Lendahl, Urban [Department of Cell and Molecular Biology, Karolinska Institutet, SE-171 77 Stockholm (Sweden); Zhao, Jian, E-mail: Jian.Zhao@ki.se [Department of Oncology–Pathology, Karolinska Institutet, CCK R8:05, Karolinska University Hospital Solna, SE-171 76 Stockholm (Sweden); Nistér, Monica [Department of Oncology–Pathology, Karolinska Institutet, CCK R8:05, Karolinska University Hospital Solna, SE-171 76 Stockholm (Sweden)

2013-11-01

Mitochondria are dynamic organelles whose morphology is regulated by a complex balance of fission and fusion processes, and we still know relatively little about how mitochondrial dynamics is regulated. MIEF1 (also called MiD51) has recently been characterized as a key regulator of mitochondrial dynamics and in this report we explore the functions of its paralog MIEF2 (also called MiD49), to learn to what extent MIEF2 is functionally distinct from MIEF1. We show that MIEF1 and MIEF2 have many functions in common. Both are anchored in the mitochondrial outer membrane, recruit Drp1 from the cytoplasm to the mitochondrial surface and cause mitochondrial fusion, and MIEF2, like MIEF1, can interact with Drp1 and hFis1. MIEF1 and MIEF2, however, also differ in certain aspects. MIEF1 and MIEF2 are differentially expressed in human tissues during development. When overexpressed, MIEF2 exerts a stronger fusion-promoting effect than MIEF1, and in line with this, hFis1 and Mff can only partially revert the MIEF2-induced fusion phenotype, whereas MIEF1-induced fusion is reverted to a larger extent by hFis1 and Mff. MIEF2 forms high molecular weight oligomers, while MIEF1 is largely present as a dimer. Furthermore, MIEF1 and MIEF2 use distinct domains for oligomerization: in MIEF1, the region from amino acid residues 109–154 is required, whereas oligomerization of MIEF2 depends on amino acid residues 1 to 49, i.e. the N-terminal end. We also show that oligomerization of MIEF1 is not required for its mitochondrial localization and interaction with Drp1. In conclusion, our data suggest that the mitochondrial regulators MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics. - Highlights: • MIEF1 and MIEF2 recruit Drp1 to mitochondria and cause mitochondrial fusion. • MIEF2, like MIEF1, can interact with Drp1 and hFis1. • MIEF1 and MIEF2 are differentially expressed in human tissues during development. • MIEF2 exerts a stronger fusion
The mitochondrial elongation factors MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics

International Nuclear Information System (INIS)

Liu, Tong; Yu, Rong; Jin, Shao-Bo; Han, Liwei; Lendahl, Urban; Zhao, Jian; Nistér, Monica

2013-01-01

Mitochondria are dynamic organelles whose morphology is regulated by a complex balance of fission and fusion processes, and we still know relatively little about how mitochondrial dynamics is regulated. MIEF1 (also called MiD51) has recently been characterized as a key regulator of mitochondrial dynamics and in this report we explore the functions of its paralog MIEF2 (also called MiD49), to learn to what extent MIEF2 is functionally distinct from MIEF1. We show that MIEF1 and MIEF2 have many functions in common. Both are anchored in the mitochondrial outer membrane, recruit Drp1 from the cytoplasm to the mitochondrial surface and cause mitochondrial fusion, and MIEF2, like MIEF1, can interact with Drp1 and hFis1. MIEF1 and MIEF2, however, also differ in certain aspects. MIEF1 and MIEF2 are differentially expressed in human tissues during development. When overexpressed, MIEF2 exerts a stronger fusion-promoting effect than MIEF1, and in line with this, hFis1 and Mff can only partially revert the MIEF2-induced fusion phenotype, whereas MIEF1-induced fusion is reverted to a larger extent by hFis1 and Mff. MIEF2 forms high molecular weight oligomers, while MIEF1 is largely present as a dimer. Furthermore, MIEF1 and MIEF2 use distinct domains for oligomerization: in MIEF1, the region from amino acid residues 109–154 is required, whereas oligomerization of MIEF2 depends on amino acid residues 1 to 49, i.e. the N-terminal end. We also show that oligomerization of MIEF1 is not required for its mitochondrial localization and interaction with Drp1. In conclusion, our data suggest that the mitochondrial regulators MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics. - Highlights: • MIEF1 and MIEF2 recruit Drp1 to mitochondria and cause mitochondrial fusion. • MIEF2, like MIEF1, can interact with Drp1 and hFis1. • MIEF1 and MIEF2 are differentially expressed in human tissues during development. • MIEF2 exerts a stronger fusion
Ultraviolet radiation (UVR) induces cell-surface Ro/SSA antigen expression by human keratinocytes in vitro: a possible mechanism for the UVR induction of cutaneous lupus lesions

International Nuclear Information System (INIS)

Jones, S.K.

1992-01-01

Antinuclear antibodies are useful markers of connective tissue disease. In this study, UVB but not UVA induced the expression of Ro/SSA antigen on keratinocyte surfaces in vitro. This expression was also found with the extractable nuclear antigens RnP and Sm, but not with single or double-stranded DNA. The expression was prevented by blocking protein synthesis, suggesting that it was an active process. The results suggest that UVB exposure may result in the expression of Ro/SSA antigen on the surfaces of basal keratinocytes in vivo. This antigen could then bind circulating antibody leading to the cutaneous lesions in neonatal and subacute cutaneous lupus erythematosus. (Author)

What is shared, what is different? Core relational themes and expressive displays of eight positive emotions.

Science.gov (United States)

Campos, Belinda; Shiota, Michelle N; Keltner, Dacher; Gonzaga, Gian C; Goetz, Jennifer L

2013-01-01

Understanding positive emotions' shared and differentiating features can yield valuable insight into the structure of positive emotion space and identify emotion states, or aspects of emotion states, that are most relevant for particular psychological processes and outcomes. We report two studies that examined core relational themes (Study 1) and expressive displays (Study 2) for eight positive emotion constructs--amusement, awe, contentment, gratitude, interest, joy, love, and pride. Across studies, all eight emotions shared one quality: high positive valence. Distinctive core relational theme and expressive display patterns were found for four emotions--amusement, awe, interest, and pride. Gratitude was associated with a distinct core relational theme but not an expressive display. Joy and love were each associated with a distinct expressive display but their core relational themes also characterised pride and gratitude, respectively. Contentment was associated with a distinct expressive display but not a core relational theme. The implications of this work for the study of positive emotion are discussed.
Gene Expression Differences in Peripheral Blood of Parkinson's Disease Patients with Distinct Progression Profiles.

Directory of Open Access Journals (Sweden)

Raquel Pinho

Full Text Available The prognosis of neurodegenerative disorders is clinically challenging due to the inexistence of established biomarkers for predicting disease progression. Here, we performed an exploratory cross-sectional, case-control study aimed at determining whether gene expression differences in peripheral blood may be used as a signature of Parkinson's disease (PD progression, thereby shedding light into potential molecular mechanisms underlying disease development. We compared transcriptional profiles in the blood from 34 PD patients who developed postural instability within ten years with those of 33 patients who did not develop postural instability within this time frame. Our study identified >200 differentially expressed genes between the two groups. The expression of several of the genes identified was previously found deregulated in animal models of PD and in PD patients. Relevant genes were selected for validation by real-time PCR in a subset of patients. The genes validated were linked to nucleic acid metabolism, mitochondria, immune response and intracellular-transport. Interestingly, we also found deregulation of these genes in a dopaminergic cell model of PD, a simple paradigm that can now be used to further dissect the role of these molecular players on dopaminergic cell loss. Altogether, our study provides preliminary evidence that expression changes in specific groups of genes and pathways, detected in peripheral blood samples, may be correlated with differential PD progression. Our exploratory study suggests that peripheral gene expression profiling may prove valuable for assisting in prediction of PD prognosis, and identifies novel culprits possibly involved in dopaminergic cell death. Given the exploratory nature of our study, further investigations using independent, well-characterized cohorts will be essential in order to validate our candidates as predictors of PD prognosis and to definitively confirm the value of gene expression
Dimensional Information-Theoretic Measurement of Facial Emotion Expressions in Schizophrenia

Directory of Open Access Journals (Sweden)

Jihun Hamm

2014-01-01

Full Text Available Altered facial expressions of emotions are characteristic impairments in schizophrenia. Ratings of affect have traditionally been limited to clinical rating scales and facial muscle movement analysis, which require extensive training and have limitations based on methodology and ecological validity. To improve reliable assessment of dynamic facial expression changes, we have developed automated measurements of facial emotion expressions based on information-theoretic measures of expressivity of ambiguity and distinctiveness of facial expressions. These measures were examined in matched groups of persons with schizophrenia (n=28 and healthy controls (n=26 who underwent video acquisition to assess expressivity of basic emotions (happiness, sadness, anger, fear, and disgust in evoked conditions. Persons with schizophrenia scored higher on ambiguity, the measure of conditional entropy within the expression of a single emotion, and they scored lower on distinctiveness, the measure of mutual information across expressions of different emotions. The automated measures compared favorably with observer-based ratings. This method can be applied for delineating dynamic emotional expressivity in healthy and clinical populations.
Identification of new TSGA10 transcript variants in human testis with conserved regulatory RNA elements in 5'untranslated region and distinct expression in breast cancer.

Science.gov (United States)

Salehipour, Pouya; Nematzadeh, Mahsa; Mobasheri, Maryam Beigom; Afsharpad, Mandana; Mansouri, Kamran; Modarressi, Mohammad Hossein

2017-09-01

Testis specific gene antigen 10 (TSGA10) is a cancer testis antigen involved in the process of spermatogenesis. TSGA10 could also play an important role in the inhibition of angiogenesis by preventing nuclear localization of HIF-1α. Although it has been shown that TSGA10 messenger RNA (mRNA) is mainly expressed in testis and some tumors, the transcription pattern and regulatory mechanisms of this gene remain largely unknown. Here, we report that human TSGA10 comprises at least 22 exons and generates four different transcript variants. It was identified that using two distinct promoters and splicing of exons 4 and 7 produced these transcript variants, which have the same coding sequence, but the sequence of 5'untanslated region (5'UTR) is different between them. This is significant because conserved regulatory RNA elements like upstream open reading frame (uORF) and putative internal ribosome entry site (IRES) were found in this region which have different combinations in each transcript variant and it may influence translational efficiency of them in normal or unusual environmental conditions like hypoxia. To indicate the transcription pattern of TSGA10 in breast cancer, expression of identified transcript variants was analyzed in 62 breast cancer samples. We found that TSGA10 tends to express variants with shorter 5'UTR and fewer uORF elements in breast cancer tissues. Our study demonstrates for the first time the expression of different TSGA10 transcript variants in testis and breast cancer tissues and provides a first clue to a role of TSGA10 5'UTR in regulation of translation in unusual environmental conditions like hypoxia. Copyright © 2017. Published by Elsevier B.V.
Comparison of distinct transcriptional expression patterns of flavonoid biosynthesis in Cabernet Sauvignon grapes from east and west China.

Science.gov (United States)

Li, Qiang; He, Fei; Zhu, Bao-Qing; Liu, Bin; Sun, Run-Ze; Duan, Chang-Qing; Reeves, Malcolm J; Wang, Jun

2014-11-01

Flavonoids make a very important contribution to the organoleptic qualities of grapes and wines. In this work these were analyzed in Cabernet Sauvignon grown in Changli, Hebei Province in east China and Gaotai, Gansu Province in west China. These regions have distinctly different climates contributing to their different 'terroir'. RNA sequencing was performed to trace transcriptome changes in Cabernet Sauvignon berries at pea size, veraison and ripening, corresponding to E-L 31, 35 and 38. The accumulation of flavonols, flavan-3-ols and anthocyanins together with the expression of relevant genes were analyzed and compared between the two regions. The biosynthesis patterns were similar between two regions, but more flavonols, anthocyanins, and tri-hydroxylated flavonoids accumulated in grapes from Gaotai before berry harvest, possibly due to the higher transcript levels of the genes that encode biosynthetic enzymes and their potential candidate transcription factors. The lower levels of flavan-3-ols, mainly (-)-epigallocatechin, in the pre-veraison grapes from Changli, might be due to limited flow of carbon to the F3'5'H branch pathway, as the ratio of F3'5'H to F3'H was lower in these berries from Changli. It is suggested that the combination of climatic factors profoundly affect the flavonoid pathway in grapes from China, providing regionally specific metabolism patterns. Copyright © 2014 Elsevier Masson SAS. All rights reserved.
Effects of the nanotopographic surface structure of commercially pure titanium following anodization-hydrothermal treatment on gene expression and adhesion in gingival epithelial cells.

Science.gov (United States)

Takebe, J; Miyata, K; Miura, S; Ito, S

2014-09-01

The long-term stability and maintenance of endosseous implants with anodized-hydrothermally treated commercially pure titanium surfaces and a nanotopographic structure (SA-treated c.p.Ti) depend on the barrier function provided by the interface between the transmucosal portion of the implant surface and the peri-implant epithelium. This study investigated the effects of extracellular and intracellular gene expression in adherent gingival epithelial cells cultured for 1-7 days on SA-treated c.p.Ti implant surfaces compared to anodic oxide (AO) c.p.Ti and c.p.Ti disks. Scanning electron microscopy (SEM) showed filopodium-like extensions bound closely to the nanotopographic structure of SA-treated c.p.Ti at day 7 of culture. Gene expressions of focal adhesion kinase, integrin-α6β4, and laminin-5 (α3, β3, γ2) were significantly higher on SA-treated c.p.Ti than on c.p.Ti or AO c.p.Ti after 7 days (Pcells adhere to SA-treated c.p.Ti as the transmucosal portion of an implant, and that this interaction markedly improves expression of focal adhesion molecules and enhances the epithelial cell phenotype. The cellular gene expression responses driving extracellular and intracellular molecular interactions thus play an important role in maintenance at the interface between SA-treated c.p.Ti implant surfaces and the gingival epithelial cells. Copyright © 2014 Elsevier B.V. All rights reserved.
Host gene expression profiles in ferrets infected with genetically distinct henipavirus strains

NARCIS (Netherlands)

Leon, A.J. (Alberto J.); Borisevich, V. (Viktoriya); Boroumand, N. (Nahal); Seymour, R. (Robert); Nusbaum, R. (Rebecca); Escaffre, O. (Olivier); Xu, L. (Luoling); Kelvin, D.J. (David J.); B. Rockx (Barry)

2018-01-01

textabstractHenipavirus infection causes severe respiratory and neurological disease in humans that can be fatal. To characterize the pathogenic mechanisms of henipavirus infection in vivo, we performed experimental infections in ferrets followed by genome-wide gene expression analysis of lung and
Agitation down-regulates immunoglobulin binding protein EibG expression in Shiga toxin-producing Escherichia coli (STEC.

Directory of Open Access Journals (Sweden)

Thorsten Kuczius

Full Text Available Shiga toxin (Stx-producing Escherichia coli (STEC carrying eibG synthesize Escherichia coli immunoglobulin binding protein (EibG. EibG nonspecifically binds to immunoglobulins and tends to aggregate in multimers but is poorly expressed in wild-type strains. To study synthesis of the proteins and their regulation in the pathogens, we identified natural growth conditions that increased EibG synthesis. EibG proteins as well as corresponding mRNA were highly expressed under static growth conditions while shearing stress created by agitation during growth repressed protein synthesis. Further regulation effects were driven by reduced oxygen tension, and pH up-regulated EibG expression, but to a lesser extent than growth conditions while decreased temperature down-regulated EibG. Bacteria with increased EibG expression during static growth conditions showed a distinct phenotype with chain formation and biofilm generation, which disappeared with motion. High and low EibG expression was reversible indicating a process with up- and down-regulation of the protein expression. Our findings indicate that shear stress represses EibG expression and might reduce bacterial attachments to cells and surfaces.
[Optimization of prokaryotic expression conditions of Leptospira interrogans trigeminy genus-specific protein antigen based on surface response analysis].

Science.gov (United States)

Wang, Jiang; Luo, Dongjiao; Sun, Aihua; Yan, Jie

2008-07-01

Lipoproteins LipL32 and LipL21 and transmembrane protein OMPL1 have been confirmed as the superficial genus-specific antigens of Leptospira interrogans, which can be used as antigens for developing a universal genetic engineering vaccine. In order to obtain high expression of an artificial fusion gene lipL32/1-lipL21-ompL1/2, we optimized prokaryotic expression conditions. We used surface response analysis based on the central composite design to optimize culture conditions of a new antigen protein by recombinant Escherichia coli DE3.The culture conditions included initial pH, induction start time, post-induction time, Isopropyl beta-D-thiogalactopyranoside (IPTG) concentration, and temperature. The maximal production of antigen protein was 37.78 mg/l. The optimal culture conditions for high recombinant fusion protein was determined: initial pH 7.9, induction start time 2.5 h, a post-induction time of 5.38 h, 0.20 mM IPTG, and a post-induction temperature of 31 degrees C. Surface response analysis based on CCD increased the target production. This statistical method reduced the number of experiments required for optimization and enabled rapid identification and integration of the key culture condition parameters for optimizing recombinant protein expression.
Effects of charge and surface ligand properties of nanoparticles on oxidative stress and gene expression within the gut of Daphnia magna

Energy Technology Data Exchange (ETDEWEB)

Dominguez, Gustavo A.; Lohse, Samuel E.; Torelli, Marco; Murphy, Catherine; Hamers, Robert J.; Orr, Galya; Klaper, Rebecca D.

2015-05-01

Concern has been raised regarding the current and future release of engineered nanomaterials into aquatic environments from industry and other sources. However, not all nanomaterials may cause an environ-mental impact and identifying which nanomaterials may be of greatest concern has been difficult. It is thought that the surface groups of a functionalized nanoparticles (NPs) may play a significant role in determining their interactions with aquatic organisms, but the way in which surface properties of NPs impact their toxicity in whole organisms has been minimally explored. A major point of interaction of NPs with aquatic organisms is in the gastrointestinal tract as they ingest particulates from the water column or from the sediment. The main goal of this study was to use model gold NP (AuNPs) to evaluate the potential effects of the different surfaces groups on NPs on the gut of an aquatic model organism, Daphnia magna. In this study, we exposed daphnids to a range of AuNPs concentrations and assessed the impact of AuNP exposure in the daphnid gut by measuring reactive oxygen species (ROS) production and expression of genes associated with oxidative stress and general cellular stress: glutathione S-transferase(gst), catalase (cat), heat shock protein 70 (hsp70), and metallothionein1 (mt1). We found ROS formation and gene expression were impacted by both charge and the specific surface ligand used. We detected some degree of ROS production in all NP exposures, but positively charged AuNPs induced a greater ROS response. Similarly, we observed that, compared to controls, both positively charged AuNPs and only one negatively AuNP impacted expression of genes associated with cellular stress. Finally, ligand-AuNP exposures showed a different toxicity and gene expression profile than the ligand alone, indicating a NP specific effect.
Distinct Developmental Origins Manifest in the Specialized Encoding of Movement by Adult Neurons of the External Globus Pallidus

Science.gov (United States)

Dodson, Paul D.; Larvin, Joseph T.; Duffell, James M.; Garas, Farid N.; Doig, Natalie M.; Kessaris, Nicoletta; Duguid, Ian C.; Bogacz, Rafal; Butt, Simon J.B.; Magill, Peter J.

2015-01-01

Summary Transcriptional codes initiated during brain development are ultimately realized in adulthood as distinct cell types performing specialized roles in behavior. Focusing on the mouse external globus pallidus (GPe), we demonstrate that the potential contributions of two GABAergic GPe cell types to voluntary action are fated from early life to be distinct. Prototypic GPe neurons derive from the medial ganglionic eminence of the embryonic subpallium and express the transcription factor Nkx2-1. These neurons fire at high rates during alert rest, and encode movements through heterogeneous firing rate changes, with many neurons decreasing their activity. In contrast, arkypallidal GPe neurons originate from lateral/caudal ganglionic eminences, express the transcription factor FoxP2, fire at low rates during rest, and encode movements with robust increases in firing. We conclude that developmental diversity positions prototypic and arkypallidal neurons to fulfil distinct roles in behavior via their disparate regulation of GABA release onto different basal ganglia targets. PMID:25843402
MUC19 expression in human ocular surface and lacrimal gland and its alteration in Sjögren syndrome patients.

Science.gov (United States)

Yu, D F; Chen, Y; Han, J M; Zhang, H; Chen, X P; Zou, W J; Liang, L Y; Xu, C C; Liu, Z G

2008-02-01

This study investigated the expression of MUC19, a newly discovered gel-forming mucin gene, in normal human lacrimal functional unit components and its alteration in Sjögren syndrome patients. Real-time PCR and immunohistochemistry were performed to determine the expression of MUC19 and MUC5AC in human cornea, conjunctiva, and lacrimal gland tissues. Conjunctival impression cytology specimens were collected from normal control subjects and Sjögren syndrome patients for Real-time PCR, PAS staining, and immunohistochemistry assays. In addition, conjunctiva biopsy specimens from both groups were examined for the expression differences of MUC19 and MUC5AC at both mRNA and protein level. The MUC19 mRNA was found to be present in cornea, conjunctiva and lacrimal gland tissues. The immunohistochemical staining of mucins showed that MUC19 was expressed in epithelial cells from corneal, conjunctival, and lacrimal gland tissues. In contrast, MUC5AC mRNA was only present in conjunctiva and lacrimal gland tissues, but not in cornea. Immunostaining demonstrates the co-staining of MUC19 and MUC5AC in conjunctival goblet cells. Consistent with the significant decrease of mucous secretion, both MUC19 and MUC5AC were decreased in conjunctiva of Sjögren syndrome patients compared to normal subjects. Considering the contribution of gel-forming mucins to the homeostasis of the ocular surface, the decreased expression of MUC19 and MUC5AC in Sjögren syndrome patients suggested that these mucins may be involved in the disruption of the ocular surface homeostasis in this disease.
Surface Expression of TGF-β Docking Receptor GARP Promotes Oncogenesis and Immune Tolerance in Breast Cancer

Science.gov (United States)

Metelli, Alessandra; Wu, Bill X; Fugle, Caroline W; Rachidi, Saleh; Sun, Shaoli; Zhang, Yongliang; Wu, Jennifer; Tomlinson, Stephen; Howe, Philip; Yang, Yi; Garrett-Mayer, Elizabeth; Liu, Bei; Li, Zihai

2016-01-01

GARP encoded by the Lrrc32 gene is the cell surface docking receptor for latent TGF-β which is expressed naturally by platelets and regulatory T cells. Although Lrrc32 is amplified frequently in breast cancer, the expression and relevant functions of GARP in cancer have not been explored. Here we report that GARP exerts oncogenic effects, promoting immune tolerance by enriching and activating latent TGF-β in the tumor microenvironment. We found that human breast, lung and colon cancers expressed GARP aberrantly. In genetic studies in normal mammary gland epithelial and carcinoma cells, GARP expression increased TGF-β bioactivity and promoted malignant transformation in immune deficient mice. In breast carcinoma-bearing mice that were immune competent, GARP overexpression promoted Foxp3+ regulatory T cell activity, which in turn contributed to enhancing cancer progression and metastasis. Notably, administration of a panel of GARP-specific monoclonal antibodies limited metastasis in an orthotopic model of human breast cancer. Overall, these results define the oncogenic effects of the GARP-TGF-β axis in the tumor microenvironment and suggest mechanisms that might be exploited for diagnostic and therapeutic purposes. PMID:27913437
Expression robust 3D face recognition via mesh-based histograms of multiple order surface differential quantities

KAUST Repository

Li, Huibin

2011-09-01

This paper presents a mesh-based approach for 3D face recognition using a novel local shape descriptor and a SIFT-like matching process. Both maximum and minimum curvatures estimated in the 3D Gaussian scale space are employed to detect salient points. To comprehensively characterize 3D facial surfaces and their variations, we calculate weighted statistical distributions of multiple order surface differential quantities, including histogram of mesh gradient (HoG), histogram of shape index (HoS) and histogram of gradient of shape index (HoGS) within a local neighborhood of each salient point. The subsequent matching step then robustly associates corresponding points of two facial surfaces, leading to much more matched points between different scans of a same person than the ones of different persons. Experimental results on the Bosphorus dataset highlight the effectiveness of the proposed method and its robustness to facial expression variations. © 2011 IEEE.
An investigation of interactions between hypocretin/orexin signaling and glutamate receptor surface expression in the rat nucleus accumbens under basal conditions and after cocaine exposure.

Science.gov (United States)

Plaza-Zabala, Ainhoa; Li, Xuan; Milovanovic, Mike; Loweth, Jessica A; Maldonado, Rafael; Berrendero, Fernando; Wolf, Marina E

2013-12-17

Hypocretin peptides are critical for the effects of cocaine on excitatory synaptic strength in the ventral tegmental area (VTA). However, little is known about their role in cocaine-induced synaptic plasticity in the nucleus accumbens (NAc). First, we tested whether hypocretin-1 by itself could acutely modulate glutamate receptor surface expression in the NAc, given that hypocretin-1 in the VTA reproduces cocaine's effects on glutamate transmission. We found no effect of hypocretin-1 infusion on AMPA or NMDA receptor surface expression in the NAc, measured by biotinylation, either 30 min or 3h after the infusion. Second, we were interested in whether changes in hypocretin receptor-2 (Hcrtr-2) expression contribute to cocaine-induced plasticity in the NAc. As a first step towards addressing this question, Hcrtr-2 surface expression was compared in the NAc after withdrawal from extended-access self-administration of saline (control) versus cocaine. We found that surface Hcrtr-2 levels remain unchanged following 14, 25 or 48 days of withdrawal from cocaine, a time period in which high conductance GluA2-lacking AMPA receptors progressively emerge in the NAc. Overall, our results fail to support a role for hypocretins in acute modulation of glutamate receptor levels in the NAc or a role for altered Hcrtr-2 expression in withdrawal-dependent synaptic adaptations in the NAc following cocaine self-administration. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.
Tributyltin Differentially Promotes Development of a Phenotypically Distinct Adipocyte

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Regnier, Shane M.; El-Hashani, Essam; Kamau, Wakanene; Zhang, Xiaojie; Massad, Nicole L.; Sargis, Robert M.

2015-01-01

Objective Environmental endocrine disrupting chemicals (EDCs) are increasingly implicated in the pathogenesis of obesity. Evidence implicates various EDCs as being pro-adipogenic, including tributyltin (TBT), which activates the peroxisome proliferator activated receptor-γ (PPARγ). However, the conditions required for TBT-induced adipogenesis and its functional consequences are incompletely known. Methods The co-stimulatory conditions necessary for preadipocyte-to-adipocyte differentiation were compared between TBT and the pharmacological PPARγ agonist troglitazone (Trog) in the 3T3-L1 cell line; basal and insulin-stimulated glucose uptake were assessed using radiolabeled 2-deoxyglucose. Results TBT enhanced expression of the adipocyte marker C/EBPα with co-exposure to either isobutylmethylxanthine or insulin in the absence of other adipogenic stimuli. Examination of several adipocyte-specific proteins revealed that TBT and Trog differentially affected protein expression despite comparable PPARγ stimulation. In particular, TBT reduced adiponectin expression upon maximal adipogenic stimulation. Under submaximal stimulation, TBT and Trog differentially promoted adipocyte-specific gene expression despite similar lipid accumulation. Moreover, TBT attenuated Trog-induced adipocyte gene expression under conditions of co-treatment. Finally, TBT-induced adipocytes exhibited altered glucose metabolism, with increased basal glucose uptake. Conclusions TBT-induced adipocytes are functionally distinct from those generated by a pharmacological PPARγ agonist, suggesting that obesogen-induced adipogenesis may generate dysfunctional adipocytes with the capacity to deleteriously affect global energy homeostasis. PMID:26243053
Tributyltin differentially promotes development of a phenotypically distinct adipocyte.

Science.gov (United States)

Regnier, Shane M; El-Hashani, Essam; Kamau, Wakanene; Zhang, Xiaojie; Massad, Nicole L; Sargis, Robert M

2015-09-01

Environmental endocrine disrupting chemicals (EDCs) are increasingly implicated in the pathogenesis of obesity. Evidence implicates various EDCs as being proadipogenic, including tributyltin (TBT), which activates the peroxisome proliferator activated receptor-γ (PPARγ). However, the conditions required for TBT-induced adipogenesis and its functional consequences are incompletely known. The costimulatory conditions necessary for preadipocyte-to-adipocyte differentiation were compared between TBT and the pharmacological PPARγ agonist troglitazone (Trog) in the 3T3-L1 cell line; basal and insulin-stimulated glucose uptake were assessed using radiolabeled 2-deoxyglucose. TBT enhanced expression of the adipocyte marker C/EBPα with coexposure to either isobutylmethylxanthine or insulin in the absence of other adipogenic stimuli. Examination of several adipocyte-specific proteins revealed that TBT and Trog differentially affected protein expression despite comparable PPARγ stimulation. In particular, TBT reduced adiponectin expression upon maximal adipogenic stimulation. Under submaximal stimulation, TBT and Trog differentially promoted adipocyte-specific gene expression despite similar lipid accumulation. Moreover, TBT attenuated Trog-induced adipocyte gene expression under conditions of cotreatment. Finally, TBT-induced adipocytes exhibited altered glucose metabolism, with increased basal glucose uptake. TBT-induced adipocytes are functionally distinct from those generated by a pharmacological PPARγ agonist, suggesting that obesogen-induced adipogenesis may generate dysfunctional adipocytes with the capacity to deleteriously affect global energy homeostasis. © 2015 The Obesity Society.
Understanding the biological responses of nanostructured metals and surfaces

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Lowe, Terry C.; Reiss, Rebecca A.

2014-08-01

Metals produced by Severe Plastic Deformation (SPD) offer distinct advantages for medical applications such as orthopedic devices, in part because of their nanostructured surfaces. We examine the current theoretical foundations and state of knowledge for nanostructured biomaterials surface optimization within the contexts that apply to bulk nanostructured metals, differentiating how their microstructures impact osteogenesis, in particular, for Ultrafine Grained (UFG) titanium. Then we identify key gaps in the research to date, pointing out areas which merit additional focus within the scientific community. For example, we highlight the potential of next-generation DNA sequencing techniques (NGS) to reveal gene and non-coding RNA (ncRNA) expression changes induced by nanostructured metals. While our understanding of bio-nano interactions is in its infancy, nanostructured metals are already being marketed or developed for medical devices such as dental implants, spinal devices, and coronary stents. Our ability to characterize and optimize the biological response of cells to SPD metals will have synergistic effects on advances in materials, biological, and medical science.
Understanding the biological responses of nanostructured metals and surfaces

International Nuclear Information System (INIS)

Lowe, Terry C; A Reiss, Rebecca

2014-01-01

Metals produced by Severe Plastic Deformation (SPD) offer distinct advantages for medical applications such as orthopedic devices, in part because of their nanostructured surfaces. We examine the current theoretical foundations and state of knowledge for nanostructured biomaterials surface optimization within the contexts that apply to bulk nanostructured metals, differentiating how their microstructures impact osteogenesis, in particular, for Ultrafine Grained (UFG) titanium. Then we identify key gaps in the research to date, pointing out areas which merit additional focus within the scientific community. For example, we highlight the potential of next-generation DNA sequencing techniques (NGS) to reveal gene and non-coding RNA (ncRNA) expression changes induced by nanostructured metals. While our understanding of bio-nano interactions is in its infancy, nanostructured metals are already being marketed or developed for medical devices such as dental implants, spinal devices, and coronary stents. Our ability to characterize and optimize the biological response of cells to SPD metals will have synergistic effects on advances in materials, biological, and medical science
Surface-stabilized gold nanocatalysts

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Dai, Sheng [Knoxville, TN; Yan, Wenfu [Oak Ridge, TN

2009-12-08

A surface-stabilized gold nanocatalyst includes a solid support having stabilizing surfaces for supporting gold nanoparticles, and a plurality of gold nanoparticles having an average particle size of less than 8 nm disposed on the stabilizing surfaces. The surface-stabilized gold nanocatalyst provides enhanced stability, such as at high temperature under oxygen containing environments. In one embodiment, the solid support is a multi-layer support comprising at least a first layer having a second layer providing the stabilizing surfaces disposed thereon, the first and second layer being chemically distinct.

Distinct Grammars and their Role in Interlanguage Development

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Aparecida de Araújo Oliveira

2016-07-01

Full Text Available The expression of temporality encompasses the concepts of tense and aspect, typically conveyed by lexical and inflectional morphemes that usually vary from language to language. This kind of cross-linguistic distinctions often affects L2 learning. In this paper I discuss the role of the Portuguese grammar in the acquisition of the English present perfect tense by eighteen Brazilian EFL learners. I compared patterns found in interlanguage data to L2 patterns, and, as expected, L1 phonology affected learners’ production of regular past tenses, especially in early interlanguage. As for form-function mappings, besides L2 patterns, learners created two major meaning categories of current relevance, based on persisting / non-persisting situations, which were systematically conveyed by forms that resembled their L1 grammar. Durative events and states (imperfective situations persisting into the present were expressed by present tenses, while all aspectual categories of concluded (perfective situations were encoded by past tenses. Specific adverbials were used in each category
Distinct GAGE and MAGE-A expression during early human development indicate specific roles in lineage differentiation

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Gjerstorff, Morten; Harkness, Linda; Kassem, Moustapha

2008-01-01

BACKGROUND: Expression of cancer/testis-associated proteins (CTAs) has traditionally been considered to be restricted to germ cells in normal tissues and to different types of malignancies. We have evaluated the potential role of CTAs in early human development. METHODS: Using immunohistochemistry...... and RT-PCR, we investigated the expression of CTAs in differentiated human embryonic stem cells (hESC) and in late embryos and early fetuses. RESULTS: We found that melanoma antigen A (MAGE-A) family members were expressed during differentiation of hESC to embryoid bodies and in teratomas, and overlapped...... with expression of the neuroectodermal markers beta-tubulin 3, Pax6 and nestin. A widespread expression of MAGE-A was also observed in neurons of the early developing central nervous system and peripheral nerves. G antigen (GAGE) expression was present in the early ectoderm of embryos, including cells...
Distinct lithium-induced gene expression effects in lymphoblastoid cell lines from patients with bipolar disorder.

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Fries, Gabriel R; Colpo, Gabriela D; Monroy-Jaramillo, Nancy; Zhao, Junfei; Zhao, Zhongming; Arnold, Jodi G; Bowden, Charles L; Walss-Bass, Consuelo

2017-11-01

Lithium is the most commonly prescribed medication for the treatment of bipolar disorder (BD), yet the mechanisms underlying its beneficial effects are still unclear. We aimed to compare the effects of lithium treatment in lymphoblastoid cell lines (LCLs) from BD patients and controls. LCLs were generated from sixty-two BD patients (based on DSM-IV) and seventeen healthy controls matched for age, sex, and ethnicity. Patients were recruited from outpatient clinics from February 2012 to October 2014. LCLs were treated with 1mM lithium for 7 days followed by microarray gene expression assay and validation by real-time quantitative PCR. Baseline differences between groups, as well as differences between vehicle- and lithium-treated cells within each group were analyzed. The biological significance of differentially expressed genes was examined by pathway enrichment analysis. No significant differences in baseline gene expression (adjusted p-value < 0.05) were detected between groups. Lithium treatment of LCLs from controls did not lead to any significant differences. However, lithium altered the expression of 236 genes in LCLs from patients; those genes were enriched for signaling pathways related to apoptosis. Among those genes, the alterations in the expression of PIK3CG, SERP1 and UPP1 were validated by real-time PCR. A significant correlation was also found between circadian functioning and CEBPG and FGF2 expression levels. In summary, our results suggest that lithium treatment induces expression changes in genes associated with the apoptosis pathway in BD LCLs. The more pronounced effects of lithium in patients compared to controls suggest a disease-specific effect of this drug. Copyright © 2017 Elsevier B.V. and ECNP. All rights reserved.
Effects of the nanotopographic surface structure of commercially pure titanium following anodization–hydrothermal treatment on gene expression and adhesion in gingival epithelial cells

International Nuclear Information System (INIS)

Takebe, J.; Miyata, K.; Miura, S.; Ito, S.

2014-01-01

The long-term stability and maintenance of endosseous implants with anodized–hydrothermally treated commercially pure titanium surfaces and a nanotopographic structure (SA-treated c.p.Ti) depend on the barrier function provided by the interface between the transmucosal portion of the implant surface and the peri-implant epithelium. This study investigated the effects of extracellular and intracellular gene expression in adherent gingival epithelial cells cultured for 1–7 days on SA-treated c.p.Ti implant surfaces compared to anodic oxide (AO) c.p.Ti and c.p.Ti disks. Scanning electron microscopy (SEM) showed filopodium-like extensions bound closely to the nanotopographic structure of SA-treated c.p.Ti at day 7 of culture. Gene expressions of focal adhesion kinase, integrin-α6β4, and laminin-5 (α3, β3, γ2) were significantly higher on SA-treated c.p.Ti than on c.p.Ti or AO c.p.Ti after 7 days (P < 0.05). Our results confirmed that gingival epithelial cells adhere to SA-treated c.p.Ti as the transmucosal portion of an implant, and that this interaction markedly improves expression of focal adhesion molecules and enhances the epithelial cell phenotype. The cellular gene expression responses driving extracellular and intracellular molecular interactions thus play an important role in maintenance at the interface between SA-treated c.p.Ti implant surfaces and the gingival epithelial cells. - Highlights: • SA-treated Ti provides a nanotopographic structure for clinical oral implants. • This could regulate integrin-mediated epithelial cell adhesion and gene expression. • FAK mRNA was significantly higher on SA-treated Ti. • Integrin-α6β4 and laminin-5 mRNA were significantly higher on SA-treated Ti. • Extracellular/intracellular molecular interactions play a key role on SA-treated Ti
Expression of surface markers on the human monocytic leukaemia cell line, THP-1, as indicators for the sensitizing potential of chemicals.

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An, Susun; Kim, Seoyoung; Huh, Yong; Lee, Tae Ryong; Kim, Han-Kon; Park, Kui-Lea; Eun, Hee Chul

2009-04-01

Evaluation of skin sensitization potential is an important part of the safety assessment of cosmetic ingredients and topical drugs. Recently, evaluation of changes in surface marker expression induced in dendritic cells (DC) or DC surrogate cell lines following exposure to chemicals represents one approach for in vitro test methods. The study aimed to test the change of expression patterns of surface markers on THP-1 cells by chemicals as a predictive in vitro method for contact sensitization. We investigated the expression of CD54, CD86, CD83, CD80, and CD40 after a 1-day exposure to sensitizers (1-chloro-2,4-dinitrobenzene; 2,4-dinitrofluorobenzene; benzocaine; 5-chloro-2-methyl-4-isothiazolin-3-one; hexyl cinnamic aldehyde; eugenol; nickel sulfate hexahydrate; potassium dichromate; cobalt sulfate; 2-mercaptobenzothiazole; and ammonium tetrachloroplatinate) and non-sensitizers (sodium lauryl sulfate, benzalkonium chloride, lactic acid, salicylic acid, isopropanol, and dimethyl sulphoxide). The test concentrations were 0.1x, 0.5x, and 1x of the 50% inhibitory concentration, and the relative fluorescence intensity was used as an expression indicator. By evaluating the expression patterns of CD54, CD86, and CD40, we could classify the chemicals as sensitizers or non-sensitizers, but CD80 and CD83 showed non-specific patterns of expression. These data suggest that the THP-1 cells are good model for screening contact sensitizers and CD40 could be a useful marker complementary to CD54 and CD86.
Heterologous expression of carcinoembryonic antigen in Lactococcus lactis via LcsB-mediated surface displaying system for oral vaccine development.

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Zhang, Xiaowei; Hu, Shumin; Du, Xue; Li, Tiejun; Han, Lanlan; Kong, Jian

2016-12-01

Carcinoembryonic antigen (CEA) is an attractive target for immunotherapy because it is expressed minimally in normal tissue, but is overexpressed in a wide variety of malignant epithelial tissues. Lactic acid bacteria (LABs), widely used in food processes, are attractive candidates for oral vaccination. Thus, we examined whether LABs could be used as a live vaccine vector to deliver CEA antigen. CEA was cloned into an Escherichia coli/Lactococcus lactis shuttle vector pSEC:LEISS under the control of a nisin promoter. For displaying the CEA on the cell surface of the L. lactis strain, the anchor motif LcsB from the S-layer protein of Lactobacillus crispatus was fused with CEA. Intracellular and cell surface expression of the CEA-LcsB fusion was confirmed by western blot analysis. Significantly higher levels of CEA-specific secretory immunoglobulin A in the sera of mice were observed upon oral administration of strain cultures containing the CEA-LcsB fused protein. In addition, the CEA-LcsB antigen group showed a higher spleen index compared to the CEA antigen alone or negative control, demonstrating that surface-displayed CEA antigen could induce a higher immune response. These results provided the first evidence for displaying CEA antigen on the cell surfaces of LABs as oral vaccines against cancer or infectious diseases. Copyright © 2014. Published by Elsevier B.V.
Hydroxy decenoic acid down regulates gtfB and gtfC expression and prevents Streptococcus mutans adherence to the cell surfaces.

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Yousefi, Behnam; Ghaderi, Shahrooz; Rezapoor-Lactooyi, Alireza; Amiri, Niusha; Verdi, Javad; Shoae-Hassani, Alireza

2012-07-28

10-Hydroxy-2-decenoic acid, an unsaturated fatty acid is the most active and unique component to the royal jelly that has antimicrobial properties. Streptococcus mutans is associated with pathogenesis of oral cavity, gingivoperiodontal diseases and bacteremia following dental manipulations. In the oral cavity, S. mutans colonize the soft tissues including tongue, palate, and buccal mucosa. When considering the role of supragingival dental plaque in caries, the proportion of acid producing bacteria (particularly S. mutans), has direct relevance to the pathogenicity of the plaque. The genes that encode glucosyltransferases (gtfs) especially gtfB and gtfC are important in S. mutans colonization and pathogenesis. This study investigated the hydroxy-decenoic acid (HDA) effects on gtfB and gtfC expression and S. mutans adherence to cells surfaces. Streptococcus mutans was treated by different concentrations of HPLC purified HDA supplied by Iran Beekeeping and Veterinary Association. Real time RT-PCR and western blot assays were conducted to evaluate gtfB and gtfC genes transcription and translation before and after HDA treatment. The bacterial attachment to the cell surfaces was evaluated microscopically. 500 μg ml-1 of HDA inhibited gtfB and gtfC mRNA transcription and its expression. The same concentration of HDA decreased 60% the adherence of S. mutans to the surface of P19 cells. Hydroxy-decenoic acid prevents gtfB and gtfC expression efficiently in the bactericide sub-concentrations and it could effectively reduce S. mutans adherence to the cell surfaces. In the future, therapeutic approaches to affecting S. mutans could be selective and it's not necessary to put down the oral flora completely.
Expression

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Wang-Xia Wang

2014-02-01

Full Text Available The miR-15/107 family comprises a group of 10 paralogous microRNAs (miRNAs, sharing a 5′ AGCAGC sequence. These miRNAs have overlapping targets. In order to characterize the expression of miR-15/107 family miRNAs, we employed customized TaqMan Low-Density micro-fluid PCR-array to investigate the expression of miR-15/107 family members, and other selected miRNAs, in 11 human tissues obtained at autopsy including the cerebral cortex, frontal cortex, primary visual cortex, thalamus, heart, lung, liver, kidney, spleen, stomach and skeletal muscle. miR-103, miR-195 and miR-497 were expressed at similar levels across various tissues, whereas miR-107 is enriched in brain samples. We also examined the expression patterns of evolutionarily conserved miR-15/107 miRNAs in three distinct primary rat brain cell preparations (enriched for cortical neurons, astrocytes and microglia, respectively. In primary cultures of rat brain cells, several members of the miR-15/107 family are enriched in neurons compared to other cell types in the central nervous system (CNS. In addition to mature miRNAs, we also examined the expression of precursors (pri-miRNAs. Our data suggested a generally poor correlation between the expression of mature miRNAs and their precursors. In summary, we provide a detailed study of the tissue and cell type-specific expression profile of this highly expressed and phylogenetically conserved family of miRNA genes.
Ancestral amphibian v2rs are expressed in the main olfactory epithelium

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Syed, Adnan S.; Sansone, Alfredo; Nadler, Walter; Manzini, Ivan; Korsching, Sigrun I.

2013-01-01

Mammalian olfactory receptor families are segregated into different olfactory organs, with type 2 vomeronasal receptor (v2r) genes expressed in a basal layer of the vomeronasal epithelium. In contrast, teleost fish v2r genes are intermingled with all other olfactory receptor genes in a single sensory surface. We report here that, strikingly different from both lineages, the v2r gene family of the amphibian Xenopus laevis is expressed in the main olfactory as well as the vomeronasal epithelium. Interestingly, late diverging v2r genes are expressed exclusively in the vomeronasal epithelium, whereas “ancestral” v2r genes, including the single member of v2r family C, are restricted to the main olfactory epithelium. Moreover, within the main olfactory epithelium, v2r genes are expressed in a basal zone, partially overlapping, but clearly distinct from an apical zone of olfactory marker protein and odorant receptor-expressing cells. These zones are also apparent in the spatial distribution of odor responses, enabling a tentative assignment of odor responses to olfactory receptor gene families. Responses to alcohols, aldehydes, and ketones show an apical localization, consistent with being mediated by odorant receptors, whereas amino acid responses overlap extensively with the basal v2r-expressing zone. The unique bimodal v2r expression pattern in main and accessory olfactory system of amphibians presents an excellent opportunity to study the transition of v2r gene expression during evolution of higher vertebrates. PMID:23613591
Distinct physiological roles for the two L-asparaginase isozymes of Escherichia coli

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Srikhanta, Yogitha N. [Department of Microbiology and Immunology, The University of Melbourne, Victoria 3010 (Australia); Atack, John M.; Beacham, Ifor R. [Institute for Glycomics, Griffith University, Gold Coast, QLD 4222 (Australia); Jennings, Michael P., E-mail: m.jennings@griffith.edu.au [Institute for Glycomics, Griffith University, Gold Coast, QLD 4222 (Australia)

2013-07-05

Highlights: •Escherichia coli contains two L-asparaginase isozymes with distinct localization, kinetics and regulation. •Mutant strains were used to examine the roles of these enzymes in L-asparagine utilization. •We report that L-asparaginase II permits growth on asparagine and glycerol under anaerobic conditions. •We propose that this enzyme is the first step in a co-regulated pathway leading to fumarate. •The pathway is regulated by anaerobiosis and cAMP and provides a terminal elector acceptor. -- Abstract: Escherichia coli expresses two L-asparaginase (EC 3.5.1.1) isozymes: L-asparaginse I, which is a low affinity, cytoplasmic enzyme that is expressed constitutively, and L-asparaginase II, a high affinity periplasmic enzyme that is under complex co-transcriptional regulation by both Fnr and Crp. The distinct localisation and regulation of these enzymes suggest different roles. To define these roles, a set of isogenic mutants was constructed that lacked either or both enzymes. Evidence is provided that L-asparaginase II, in contrast to L-asparaginase I, can be used in the provision of an anaerobic electron acceptor when using a non-fermentable carbon source in the presence of excess nitrogen.
The MASP family of Trypanosoma cruzi: changes in gene expression and antigenic profile during the acute phase of experimental infection.

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Sara Lopes dos Santos

Full Text Available Trypanosoma cruzi is the etiological agent of Chagas disease, a debilitating illness that affects millions of people in the Americas. A major finding of the T. cruzi genome project was the discovery of a novel multigene family composed of approximately 1,300 genes that encode mucin-associated surface proteins (MASPs. The high level of polymorphism of the MASP family associated with its localization at the surface of infective forms of the parasite suggests that MASP participates in host-parasite interactions. We speculate that the large repertoire of MASP sequences may contribute to the ability of T. cruzi to infect several host cell types and/or participate in host immune evasion mechanisms.By sequencing seven cDNA libraries, we analyzed the MASP expression profile in trypomastigotes derived from distinct host cells and after sequential passages in acutely infected mice. Additionally, to investigate the MASP antigenic profile, we performed B-cell epitope prediction on MASP proteins and designed a MASP-specific peptide array with 110 putative epitopes, which was screened with sera from acutely infected mice.We observed differential expression of a few MASP genes between trypomastigotes derived from epithelial and myoblast cell lines. The more pronounced MASP expression changes were observed between bloodstream and tissue-culture trypomastigotes and between bloodstream forms from sequential passages in acutely infected mice. Moreover, we demonstrated that different MASP members were expressed during the acute T. cruzi infection and constitute parasite antigens that are recognized by IgG and IgM antibodies. We also found that distinct MASP peptides could trigger different antibody responses and that the antibody level against a given peptide may vary after sequential passages in mice. We speculate that changes in the large repertoire of MASP antigenic peptides during an infection may contribute to the evasion of host immune responses during the
Lentivirus display: stable expression of human antibodies on the surface of human cells and virus particles.

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Ran Taube

Full Text Available BACKGROUND: Isolation of human antibodies using current display technologies can be limited by constraints on protein expression, folding and post-translational modifications. Here we describe a discovery platform that utilizes self-inactivating (SIN lentiviral vectors for the surface display of high-affinity single-chain variable region (scFv antibody fragments on human cells and lentivirus particles. METHODOLOGY/PRINCIPAL FINDINGS: Bivalent scFvFc human antibodies were fused in frame with different transmembrane (TM anchoring moieties to allow efficient high-level expression on human cells and the optimal TM was identified. The addition of an eight amino acid HIV-1 gp41 envelope incorporation motif further increased scFvFc expression on human cells and incorporation into lentiviral particles. Both antibody-displaying human cells and virus particles bound antigen specifically. Sulfation of CDR tyrosine residues, a property recently shown to broaden antibody binding affinity and antigen recognition was also demonstrated. High level scFvFc expression and stable integration was achieved in human cells following transduction with IRES containing bicistronic SIN lentivectors encoding ZsGreen when scFvFc fusion proteins were expressed from the first cassette. Up to 10(6-fold enrichment of antibody expressing cells was achieved with one round of antigen coupled magnetic bead pre-selection followed by FACS sorting. Finally, the scFvFc displaying human cells could be used directly in functional biological screens with remarkable sensitivity. CONCLUSIONS/SIGNIFICANCE: This antibody display platform will complement existing technologies by virtue of providing properties unique to lentiviruses and antibody expression in human cells, which, in turn, may aid the discovery of novel therapeutic human mAbs.
Selection of conformational states in surface self-assembly for a molecule with eight possible pairs of surface enantiomers

DEFF Research Database (Denmark)

Nuermaimaiti, Ajiguli; Schultz-Falk, Vickie; Lind Cramer, Jacob

2016-01-01

Self-assembly of a molecule with many distinct conformational states, resulting in eight possible pairs of surface enantiomers, is investigated on a Au(111) surface under UHV conditions. The complex molecule is equipped with alkyl and carboxyl moieties to promote controlled self-assembly of lamel......Self-assembly of a molecule with many distinct conformational states, resulting in eight possible pairs of surface enantiomers, is investigated on a Au(111) surface under UHV conditions. The complex molecule is equipped with alkyl and carboxyl moieties to promote controlled self......-assembly of lamellae structures. From statistical analysis of Scanning Tunnelling Microscopy (STM) data we observe a clear selection of specific conformational states after self-assembly. Using Density Functional Theory (DFT) calculations we rationalise how this selection is correlated to the orientation of the alkyl...
Post-Spaceflight (STS-135 Mouse Splenocytes Demonstrate Altered Activation Properties and Surface Molecule Expression.

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Shen-An Hwang

Full Text Available Alterations in immune function have been documented during or post-spaceflight and in ground based models of microgravity. Identification of immune parameters that are dysregulated during spaceflight is an important step in mitigating crew health risks during deep space missions. The in vitro analysis of leukocyte activity post-spaceflight in both human and animal species is primarily focused on lymphocytic function. This report completes a broader spectrum analysis of mouse lymphocyte and monocyte changes post 13 days orbital flight (mission STS-135. Analysis includes an examination in surface markers for cell activation, and antigen presentation and co-stimulatory molecules. Cytokine production was measured after stimulation with T-cell mitogen or TLR-2, TLR-4, or TLR-5 agonists. Splenocyte surface marker analysis immediate post-spaceflight and after in vitro culture demonstrated unique changes in phenotypic populations between the flight mice and matched treatment ground controls. Post-spaceflight splenocytes (flight splenocytes had lower expression intensity of CD4+CD25+ and CD8+CD25+ cells, lower percentage of CD11c+MHC II+ cells, and higher percentage of CD11c+MHC I+ populations compared to ground controls. The flight splenocytes demonstrated an increase in phagocytic activity. Stimulation with ConA led to decrease in CD4+ population but increased CD4+CD25+ cells compared to ground controls. Culturing with TLR agonists led to a decrease in CD11c+ population in splenocytes isolated from flight mice compared to ground controls. Consequently, flight splenocytes with or without TLR-agonist stimulation showed a decrease in CD11c+MHC I+, CD11c+MHC II+, and CD11c+CD86+ cells compared to ground controls. Production of IFN-γ was decreased and IL-2 was increased from ConA stimulated flight splenocytes. This study demonstrated that expression of surface molecules can be affected by conditions of spaceflight and impaired responsiveness persists under
Glutathione Transferases Superfamily: Cold-Inducible Expression of Distinct GST Genes in Brassica oleracea

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Harshavardhanan Vijayakumar

2016-07-01

Full Text Available Plants, as sessile organisms, can suffer serious growth and developmental consequences under cold stress conditions. Glutathione transferases (GSTs, EC 2.5.1.18 are ubiquitous and multifunctional conjugating proteins, which play a major role in stress responses by preventing oxidative damage by reactive oxygen species (ROS. Currently, understanding of their function(s during different biochemical and signaling pathways under cold stress condition remain unclear. In this study, using combined computational strategy, we identified 65 Brassica oleracea glutathione transferases (BoGST and characterized them based on evolutionary analysis into 11 classes. Inter-species and intra-species duplication was evident between BoGSTs and Arabidopsis GSTs. Based on localization analyses, we propose possible pathways in which GST genes are involved during cold stress. Further, expression analysis of the predicted putative functions for GST genes were investigated in two cold contrasting genotypes (cold tolerance and susceptible under cold condition, most of these genes were highly expressed at 6 h and 1 h in the cold tolerant (CT and cold susceptible (CS lines, respectively. Overall, BoGSTU19, BoGSTU24, BoGSTF10 are candidate genes highly expressed in B. oleracea. Further investigation of GST superfamily in B. oleracea will aid in understanding complex mechanism underlying cold tolerance in plants.
The Plasmodium falciparum transcriptome in severe malaria reveals altered expression of genes involved in important processes including surface antigen–encoding var genes

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Tonkin-Hill, Gerry Q.; Trianty, Leily; Noviyanti, Rintis; Nguyen, Hanh H. T.; Sebayang, Boni F.; Lampah, Daniel A.; Marfurt, Jutta; Cobbold, Simon A.; Rambhatla, Janavi S.; McConville, Malcolm J.; Rogerson, Stephen J.; Brown, Graham V.; Day, Karen P.; Price, Ric N.; Anstey, Nicholas M.

2018-01-01

Within the human host, the malaria parasite Plasmodium falciparum is exposed to multiple selection pressures. The host environment changes dramatically in severe malaria, but the extent to which the parasite responds to—or is selected by—this environment remains unclear. From previous studies, the parasites that cause severe malaria appear to increase expression of a restricted but poorly defined subset of the PfEMP1 variant, surface antigens. PfEMP1s are major targets of protective immunity. Here, we used RNA sequencing (RNAseq) to analyse gene expression in 44 parasite isolates that caused severe and uncomplicated malaria in Papuan patients. The transcriptomes of 19 parasite isolates associated with severe malaria indicated that these parasites had decreased glycolysis without activation of compensatory pathways; altered chromatin structure and probably transcriptional regulation through decreased histone methylation; reduced surface expression of PfEMP1; and down-regulated expression of multiple chaperone proteins. Our RNAseq also identified novel associations between disease severity and PfEMP1 transcripts, domains, and smaller sequence segments and also confirmed all previously reported associations between expressed PfEMP1 sequences and severe disease. These findings will inform efforts to identify vaccine targets for severe malaria and also indicate how parasites adapt to—or are selected by—the host environment in severe malaria. PMID:29529020
Intermodulation distortion and surface resistance in impurity-doped YBCO and MgB{sub 2}

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Agassi, Y.D. [Naval Surface Warfare Center, Carderock Division, Bethesda, MD 20817 (United States); Oates, D.E., E-mail: oates@ll.mit.edu [MIT-Lincoln Laboratory, Lexington, MA 02420 (United States)

2014-11-15

Highlights: • Calculations of impurity-doping effects on surface resistance and intermodulation distortion. • The calculations are compared with previously published measurements in YBCO and MgB{sub 2}. • Excellent agreement between calculations and experiments are shown. • The effects of the symmetry of the energy gap are presented and discussed. - Abstract: Calculations of the microwave intermodulation distortion (IMD) and surface resistance of impurity-doped YBCO, MgB{sub 2} and Nb are presented. These are qualitatively distinct superconductors due to their energy-gap symmetries, d-wave (ℓ = 2), i-wave (ℓ = 6) and s-wave (ℓ = 0), respectively. The calculations are compared with previously published IMD and surface-resistance measurements of impurity-doped YBCO and Nb. The agreement between the data and fitted calculations is excellent in all cases. In the absence of IMD and surface-resistance measurements for doped MgB{sub 2}, we present representative predictions. The calculations are based on a Green’s-function approach that yields analytical expressions for the penetration depth and the nonlinear kernel in the constitutive relation. This penetration-depth expression reproduces the measured T{sup 2} low-temperature variation for doped superconductors and the surface-resistance reduction over that of the pure material. Regarding the IMD in superconductors with a nodal energy gap, the effect of doping is to enhance its magnitude and suppress its low-temperature 1/T{sup 2} divergence predicted by the nonlinear Meissner effect.
Development of a flagellin surface display expression system in a moderate thermophile, Bacillus halodurans Alk36

CSIR Research Space (South Africa)

Crampton, Michael C

2007-06-01

Full Text Available BIOTECHNOLOGY The development of a flagellin surface display expression system in a moderate thermophile, Bacillus halodurans Alk36 Michael Crampton & Eldie Berger & Sharon Reid & Maureen Louw Received: 3 October 2006 /Revised: 29 January 2007 /Accepted... techniques Plasmid DNA was isolated using a Plasmid Midi Kit (Qiagen). Restriction enzymes were used as specified by the manufacturer (Fermentas and Roche Diagnostics). All mini-preps were done using Perfectprep Plasmid Mini Kit (Eppendorf). All DNA...
Surface structure of AU3Cu(001)

DEFF Research Database (Denmark)

Eckstein, G.A.; Maupai, S.; Dakkouri, A.S.

1999-01-01

The surface morphology, composition, and structure of Au3Cu(001) as determined by scanning tunneling microscopy and surface x-ray diffraction are presented. Atomic resolution STM images reveal distinctive geometric features. The analysis of the surface x-ray diffraction data provides clear evidence...... for the surface structure. [S0163-1829(99)04535-X]....
Distinctive features of Phox2b-expressing neurons in the rat reticular formation dorsal to the trigeminal motor nucleus.

Science.gov (United States)

Nagoya, Kouta; Nakamura, Shiro; Ikeda, Keiko; Onimaru, Hiroshi; Yoshida, Atsushi; Nakayama, Kiyomi; Mochizuki, Ayako; Kiyomoto, Masaaki; Sato, Fumihiko; Kawakami, Kiyoshi; Takahashi, Koji; Inoue, Tomio

2017-09-01

Phox2b encodes a paired-like homeodomain-containing transcription factor essential for development of the autonomic nervous system. Phox2b-expressing (Phox2b + ) neurons are present in the reticular formation dorsal to the trigeminal motor nucleus (RdV) as well as the nucleus of the solitary tract and parafacial respiratory group. However, the nature of Phox2b + RdV neurons is still unclear. We investigated the physiological and morphological properties of Phox2b + RdV neurons using postnatal day 2-7 transgenic rats expressing yellow fluorescent protein under the control of Phox2b. Almost all of Phox2b + RdV neurons were glutamatergic, whereas Phox2b-negative (Phox2b - ) RdV neurons consisted of a few glutamatergic, many GABAergic, and many glycinergic neurons. The majority (48/56) of Phox2b + neurons showed low-frequency firing (LF), while most of Phox2b - neurons (35/42) exhibited high-frequency firing (HF) in response to intracellularly injected currents. All, but one, Phox2b + neurons (55/56) did not fire spontaneously, whereas three-fourths of the Phox2b - neurons (31/42) were spontaneously active. K + channel and persistent Na + current blockers affected the firing of LF and HF neurons. The majority of Phox2b + (35/46) and half of the Phox2b - neurons (19/40) did not respond to stimulations of the mesencephalic trigeminal nucleus, the trigeminal tract, and the principal sensory trigeminal nucleus. Biocytin labeling revealed that about half of the Phox2b + (5/12) and Phox2b - RdV neurons (5/10) send their axons to the trigeminal motor nucleus. These results suggest that Phox2b + RdV neurons have distinct neurotransmitter phenotypes and firing properties from Phox2b - RdV neurons and might play important roles in feeding-related functions including suckling and possibly mastication. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

Accumulation of distinct prelamin A variants in human diploid fibroblasts differentially affects cell homeostasis

International Nuclear Information System (INIS)

Candelario, Jose; Borrego, Stacey; Reddy, Sita; Comai, Lucio

2011-01-01

Lamin A is a component of the nuclear lamina that plays a major role in the structural organization and function of the nucleus. Lamin A is synthesized as a prelamin A precursor which undergoes four sequential post-translational modifications to generate mature lamin A. Significantly, a large number of point mutations in the LMNA gene cause a range of distinct human disorders collectively known as laminopathies. The mechanisms by which mutations in lamin A affect cell function and cause disease are unclear. Interestingly, recent studies have suggested that alterations in the normal lamin A pathway can contribute to cellular dysfunction. Specifically, we and others have shown, at the cellular level, that in the absence of mutations or altered splicing events, increased expression of wild-type prelamin A results in a growth defective phenotype that resembles that of cells expressing the mutant form of lamin A, termed progerin, associated with Hutchinson-Gilford Progeria syndrome (HGPS). Remarkably, the phenotypes of cells expressing elevated levels of wild-type prelamin A can be reversed by either treatment with farnesyltransferase inhibitors or overexpression of ZMPSTE24, a critical prelamin A processing enzyme, suggesting that minor increases in the steady-state levels of one or more prelamin A intermediates is sufficient to induce cellular toxicity. Here, to investigate the molecular basis of the lamin A pathway toxicity, we characterized the phenotypic changes occurring in cells expressing distinct prelamin A variants mimicking specific prelamin A processing intermediates. This analysis demonstrates that distinct prelamin A variants differentially affect cell growth, nuclear membrane morphology, nuclear distribution of lamin A and the fundamental process of transcription. Expression of prelamin A variants that are constitutively farnesylated induced the formation of lamin A aggregates and dramatic changes in nuclear membrane morphology, which led to reduced
Interleukin-1β modulates smooth muscle cell phenotype to a distinct inflammatory state relative to PDGF-DD via NF-κB-dependent mechanisms.

Science.gov (United States)

Alexander, Matthew R; Murgai, Meera; Moehle, Christopher W; Owens, Gary K

2012-04-02

Smooth muscle cell (SMC) phenotypic modulation in atherosclerosis and in response to PDGF in vitro involves repression of differentiation marker genes and increases in SMC proliferation, migration, and matrix synthesis. However, SMCs within atherosclerotic plaques can also express a number of proinflammatory genes, and in cultured SMCs the inflammatory cytokine IL-1β represses SMC marker gene expression and induces inflammatory gene expression. Studies herein tested the hypothesis that IL-1β modulates SMC phenotype to a distinct inflammatory state relative to PDGF-DD. Genome-wide gene expression analysis of IL-1β- or PDGF-DD-treated SMCs revealed that although both stimuli repressed SMC differentiation marker gene expression, IL-1β distinctly induced expression of proinflammatory genes, while PDGF-DD primarily induced genes involved in cell proliferation. Promoters of inflammatory genes distinctly induced by IL-1β exhibited over-representation of NF-κB binding sites, and NF-κB inhibition in SMCs reduced IL-1β-induced upregulation of proinflammatory genes as well as repression of SMC differentiation marker genes. Interestingly, PDGF-DD-induced SMC marker gene repression was not NF-κB dependent. Finally, immunofluorescent staining of mouse atherosclerotic lesions revealed the presence of cells positive for the marker of an IL-1β-stimulated inflammatory SMC, chemokine (C-C motif) ligand 20 (CCL20), but not the PDGF-DD-induced gene, regulator of G protein signaling 17 (RGS17). Results demonstrate that IL-1β- but not PDGF-DD-induced phenotypic modulation of SMC is characterized by NF-κB-dependent activation of proinflammatory genes, suggesting the existence of a distinct inflammatory SMC phenotype. In addition, studies provide evidence for the possible utility of CCL20 and RGS17 as markers of inflammatory and proliferative state SMCs within atherosclerotic plaques in vivo.
Distinct cell clusters touching islet cells induce islet cell replication in association with over-expression of Regenerating Gene (REG protein in fulminant type 1 diabetes.

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Kaoru Aida

Full Text Available BACKGROUND: Pancreatic islet endocrine cell-supporting architectures, including islet encapsulating basement membranes (BMs, extracellular matrix (ECM, and possible cell clusters, are unclear. PROCEDURES: The architectures around islet cell clusters, including BMs, ECM, and pancreatic acinar-like cell clusters, were studied in the non-diabetic state and in the inflamed milieu of fulminant type 1 diabetes in humans. RESULT: Immunohistochemical and electron microscopy analyses demonstrated that human islet cell clusters and acinar-like cell clusters adhere directly to each other with desmosomal structures and coated-pit-like structures between the two cell clusters. The two cell-clusters are encapsulated by a continuous capsule composed of common BMs/ECM. The acinar-like cell clusters have vesicles containing regenerating (REG Iα protein. The vesicles containing REG Iα protein are directly secreted to islet cells. In the inflamed milieu of fulminant type 1 diabetes, the acinar-like cell clusters over-expressed REG Iα protein. Islet endocrine cells, including beta-cells and non-beta cells, which were packed with the acinar-like cell clusters, show self-replication with a markedly increased number of Ki67-positive cells. CONCLUSION: The acinar-like cell clusters touching islet endocrine cells are distinct, because the cell clusters are packed with pancreatic islet clusters and surrounded by common BMs/ECM. Furthermore, the acinar-like cell clusters express REG Iα protein and secrete directly to neighboring islet endocrine cells in the non-diabetic state, and the cell clusters over-express REG Iα in the inflamed milieu of fulminant type 1 diabetes with marked self-replication of islet cells.
Distinct spatiotemporal expression of ISM1 during mouse and chick development

OpenAIRE

Os?rio, Liliana; Wu, Xuewei; Zhou, Zhongjun

2014-01-01

Isthmin 1 (ISM1) constitutes the founder of a new family of secreted proteins characterized by the presence of 2 functional domains: thrombospondin type 1 repeat (TSR1) and adhesion-associated domain in MUC4 and other proteins (AMOP). ISM1 was identified in the frog embryo as a member of the FGF8 synexpression group due to its expression in the brain midbrain-hindbrain boundary (MHB) or isthmus. In zebrafish, ISM1 was described as a WNT- and NODAL-regulated gene. The function of ISM1 remains ...
The concentration, gene expression, and spatial distribution of aggrecan in canine articular cartilage, meniscus, and anterior and posterior cruciate ligaments: a new molecular distinction between hyaline cartilage and fibrocartilage in the knee joint.

Science.gov (United States)

Valiyaveettil, Manojkumar; Mort, John S; McDevitt, Cahir A

2005-01-01

The concentration, spatial distribution, and gene expression of aggrecan in meniscus, articular cartilage, and the anterior and posterior cruciate ligaments (ACL and PCL) was determined in the knee joints of five mature dogs. An anti-serum against peptide sequences specific to the G1 domain of aggrecan was employed in competitive-inhibition ELISA of guanidine HCl extracts and immunofluorescence microscopy. Gene expression was determined by Taqman real-time PCR. The concentration of aggrecan in articular cartilage (240.1 +/- 32 nMol/g dry weight) was higher than that in meniscus (medial meniscus: 33.4 +/- 4.3 nMol/g) and ligaments (ACL: 6.8 +/- 0.9 nMol/g). Aggrecan was more concentrated in the inner than the outer zone of the meniscus. Aggrecan in meniscus showed an organized, spatial network, in contrast to its diffuse distribution in articular cartilage. Thus, differences in the concentration, gene expression, and spatial distribution of aggrecan constitute another molecular distinction between hyaline cartilage and fibrocartilage of the knee.
Spatial and Single-Cell Transcriptional Profiling Identifies Functionally Distinct Human Dermal Fibroblast Subpopulations.

Science.gov (United States)

Philippeos, Christina; Telerman, Stephanie B; Oulès, Bénédicte; Pisco, Angela O; Shaw, Tanya J; Elgueta, Raul; Lombardi, Giovanna; Driskell, Ryan R; Soldin, Mark; Lynch, Magnus D; Watt, Fiona M

2018-04-01

Previous studies have shown that mouse dermis is composed of functionally distinct fibroblast lineages. To explore the extent of fibroblast heterogeneity in human skin, we used a combination of comparative spatial transcriptional profiling of human and mouse dermis and single-cell transcriptional profiling of human dermal fibroblasts. We show that there are at least four distinct fibroblast populations in adult human skin, not all of which are spatially segregated. We define markers permitting their isolation and show that although marker expression is lost in culture, different fibroblast subpopulations retain distinct functionality in terms of Wnt signaling, responsiveness to IFN-γ, and ability to support human epidermal reconstitution when introduced into decellularized dermis. These findings suggest that ex vivo expansion or in vivo ablation of specific fibroblast subpopulations may have therapeutic applications in wound healing and diseases characterized by excessive fibrosis. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.
Specific gene expression profiles and chromosomal abnormalities are associated with infant disseminated neuroblastoma

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Kushner Brian

2009-02-01

Full Text Available Abstract Background Neuroblastoma (NB tumours have the highest incidence of spontaneous remission, especially among the stage 4s NB subgroup affecting infants. Clinical distinction of stage 4s from lethal stage 4 can be difficult, but critical for therapeutic decisions. The aim of this study was to investigate chromosomal alterations and differential gene expression amongst infant disseminated NB subgroups. Methods Thirty-five NB tumours from patients diagnosed at Results All stage 4s patients underwent spontaneous remission, only 48% stage 4 patients survived despite combined modality therapy. Stage 4 tumours were 90% near-diploid/tetraploid, 44% MYCN amplified, 77% had 1p LOH (50% 1p36, 23% 11q and/or 14q LOH (27% and 47% had 17q gain. Stage 4s were 90% near-triploid, none MYCN amplified and LOH was restricted to 11q. Initial comparison analyses between stage 4s and 4 P P = 0.0054, 91% with higher expression in stage 4. Less definite expression profiles were observed between stage 4s and 4 P P = 0.005 was maintained. Distinct gene expression profiles but no significant association with specific chromosomal region localization was observed between stage 4s and stage 4 Conclusion Specific chromosomal aberrations are associated with distinct gene expression profiles which characterize spontaneously regressing or aggressive infant NB, providing the biological basis for the distinct clinical behaviour.
Hydroxy decenoic acid down regulates gtfB and gtfC expression and prevents Streptococcus mutans adherence to the cell surfaces

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Yousefi Behnam

2012-07-01

Full Text Available Abstract Background 10-Hydroxy-2-decenoic acid, an unsaturated fatty acid is the most active and unique component to the royal jelly that has antimicrobial properties. Streptococcus mutans is associated with pathogenesis of oral cavity, gingivoperiodontal diseases and bacteremia following dental manipulations. In the oral cavity, S. mutans colonize the soft tissues including tongue, palate, and buccal mucosa. When considering the role of supragingival dental plaque in caries, the proportion of acid producing bacteria (particularly S. mutans, has direct relevance to the pathogenicity of the plaque. The genes that encode glucosyltransferases (gtfs especially gtfB and gtfC are important in S. mutans colonization and pathogenesis. This study investigated the hydroxy-decenoic acid (HDA effects on gtfB and gtfC expression and S. mutans adherence to cells surfaces. Methods Streptococcus mutans was treated by different concentrations of HPLC purified HDA supplied by Iran Beekeeping and Veterinary Association. Real time RT-PCR and western blot assays were conducted to evaluate gtfB and gtfC genes transcription and translation before and after HDA treatment. The bacterial attachment to the cell surfaces was evaluated microscopically. Results 500 μg ml-1 of HDA inhibited gtfB and gtfC mRNA transcription and its expression. The same concentration of HDA decreased 60% the adherence of S. mutans to the surface of P19 cells. Conclusion Hydroxy-decenoic acid prevents gtfB and gtfC expression efficiently in the bactericide sub-concentrations and it could effectively reduce S. mutans adherence to the cell surfaces. In the future, therapeutic approaches to affecting S. mutans could be selective and it’s not necessary to put down the oral flora completely.
PKG in honey bees: spatial expression, Amfor gene expression, sucrose responsiveness, and division of labor.

Science.gov (United States)

Thamm, Markus; Scheiner, Ricarda

2014-06-01

Division of labor is a hallmark of social insects. In honey bees, division of labor involves transition of female workers from one task to the next. The most distinct tasks are nursing (providing food for the brood) and foraging (collecting pollen and nectar). The brain mechanisms regulating this form of behavioral plasticity have largely remained elusive. Recently, it was suggested that division of labor is based on nutrition-associated signaling pathways. One highly conserved gene associated with food-related behavior across species is the foraging gene, which encodes a cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG). Our analysis of this gene reveals the presence of alternative splicing in the honey bee. One isoform is expressed in the brain. Expression of this isoform is most pronounced in the mushroom bodies, the subesophageal ganglion, and the corpora allata. Division of labor and sucrose responsiveness in honey bees correlate significantly with foraging gene expression in distinct brain regions. Activating PKG selectively increases sucrose responsiveness in nurse bees to the level of foragers, whereas the same treatment does not affect responsiveness to light. These findings demonstrate a direct link between PKG signaling in distinct brain areas and division of labor. Furthermore, they demonstrate that the difference in sensory responsiveness between nurse bees and foragers can be compensated for by activating PKG. Our findings on the function of PKG in regulating specific sensory responsiveness and social organization offer valuable indications for the function of the cGMP/PKG pathway in many other insects and vertebrates. Copyright © 2013 Wiley Periodicals, Inc.
MicroRNA profiling reveals distinct signatures in B cell chronic lymphocytic leukemias

Science.gov (United States)

Calin, George Adrian; Liu, Chang-Gong; Sevignani, Cinzia; Ferracin, Manuela; Felli, Nadia; Dumitru, Calin Dan; Shimizu, Masayoshi; Cimmino, Amelia; Zupo, Simona; Dono, Mariella; Dell'Aquila, Marie L.; Alder, Hansjuerg; Rassenti, Laura; Kipps, Thomas J.; Bullrich, Florencia; Negrini, Massimo; Croce, Carlo M.

2004-01-01

Little is known about the expression levels or function of micro-RNAs (miRNAs) in normal and neoplastic cells, although it is becoming clear that miRNAs play important roles in the regulation of gene expression during development [Ambros, V. (2003) Cell 113, 673–676; McManus, M. T. (2003) Semin. Cancer Biol. 13, 253–258]. We now report the genomewide expression profiling of miRNAs in human B cell chronic lymphocytic leukemia (CLL) by using a microarray containing hundreds of human precursor and mature miRNA oligonucleotide probes. This approach allowed us to identify significant differences in miRNome expression between CLL samples and normal CD5+ B cells; data were confirmed by Northern blot analyses and real-time RT-PCR. At least two distinct clusters of CLL samples can be identified that were associated with the presence or absence of Zap-70 expression, a predictor of early disease progression. Two miRNA signatures were associated with the presence or absence of mutations in the expressed Ig variableregion genes or with deletions at 13q14, respectively. These data suggest that miRNA expression patterns have relevance to the biological and clinical behavior of this leukemia. PMID:15284443
Distinct roles for dystroglycan, beta1 integrin and perlecan in cell surface laminin organization

DEFF Research Database (Denmark)

Henry, M D; Satz, J S; Brakebusch, C

2001-01-01

Dystroglycan (DG) is a cell surface receptor for several extracellular matrix (ECM) molecules including laminins, agrin and perlecan. Recent data indicate that DG function is required for the formation of basement membranes in early development and the organization of laminin on the cell surface...... integrin-deficient ES cells, laminin-1 binds to the cell surface, but fails to organize into more morphologically complex structures. This result indicates that beta1 integrin function is required after DG function in the cell surface-mediated laminin assembly process. In perlecan-deficient ES cells......, the formation of complex laminin-1 structures is defective, implicating perlecan in the laminin matrix assembly process. Moreover, laminin and perlecan reciprocally modulate the organization of the other on the cell surface. Taken together, the data support a model whereby DG serves as a receptor essential...
Relationship of CD86 surface marker expression and cytotoxicity on dendritic cells exposed to chemical allergen

International Nuclear Information System (INIS)

Hulette, Ben C.; Ryan, Cindy A.; Gildea, Lucy A.; Gerberick, G. Frank

2005-01-01

Human peripheral blood-derived dendritic cells (DC) respond to a variety of chemical allergens by up-regulating expression of the co-stimulatory molecule CD86. It has been postulated that this measure might provide the basis for an in vitro alternative approach for the identification of skin sensitizing chemicals. We recently reported that DC, exposed in culture to the highest non-cytotoxic concentrations of various chemical allergens, displayed marginal up-regulation of membrane CD86 expression; the interpretation being that such changes were insufficiently sensitive for the purposes of hazard identification. For the work presented here, immature DC were derived from human monocytes and treated with the chemical allergens 2,4-dinitrobenzenesulfonic acid (DNBS), nickel sulfate (NiSO 4 ), p-phenylenediamine (PPD), Bandrowski's base (BB), hydroquinone (HQ) and propyl gallate (PG) for 48 h at concentrations which induced both no to slight to moderate cytotoxicity. For comparison, DC were treated with the irritants sodium dodecyl sulfate (SDS), benzoic acid (BA), and benzalkonium chloride (BZC) at concentrations resulting in comparable levels of cytotoxicity. CD86 expression, as measured by flow cytometry, was consistently up-regulated (ranging from 162 to 386% control) on DC treated with concentrations of chemical allergens that induced approximately 10-15% cytotoxicity. The irritants BA and BZC did not induce up-regulation of CD86 expression when tested at concentrations that induced similar levels of cytotoxicity. SDS, however, up-regulated CD86 expression to 125-138% of control in 2/4 preparations when tested at concentrations which induced similar toxicity. Our results confirm that chemical allergens up-regulate CD86 expression on blood-derived DC and illustrate further that up-regulation of CD86 surface marker expression is more robust when DC are treated with concentrations of chemical allergen that induce slight to moderate cytotoxicity
Distribution of cellular HSV-1 receptor expression in human brain.

Science.gov (United States)

Lathe, Richard; Haas, Juergen G

2017-06-01

Herpes simplex virus type 1 (HSV-1) is a neurotropic virus linked to a range of acute and chronic neurological disorders affecting distinct regions of the brain. Unusually, HSV-1 entry into cells requires the interaction of viral proteins glycoprotein D (gD) and glycoprotein B (gB) with distinct cellular receptor proteins. Several different gD and gB receptors have been identified, including TNFRSF14/HVEM and PVRL1/nectin 1 as gD receptors and PILRA, MAG, and MYH9 as gB receptors. We investigated the expression of these receptor molecules in different areas of the adult and developing human brain using online transcriptome databases. Whereas all HSV-1 receptors showed distinct expression patterns in different brain areas, the Allan Brain Atlas (ABA) reported increased expression of both gD and gB receptors in the hippocampus. Specifically, for PVRL1, TNFRFS14, and MYH9, the differential z scores for hippocampal expression, a measure of relative levels of increased expression, rose to 2.9, 2.9, and 2.5, respectively, comparable to the z score for the archetypical hippocampus-enriched mineralocorticoid receptor (NR3C2, z = 3.1). These data were confirmed at the Human Brain Transcriptome (HBT) database, but HBT data indicate that MAG expression is also enriched in hippocampus. The HBT database allowed the developmental pattern of expression to be investigated; we report that all HSV1 receptors markedly increase in expression levels between gestation and the postnatal/adult periods. These results suggest that differential receptor expression levels of several HSV-1 gD and gB receptors in the adult hippocampus are likely to underlie the susceptibility of this brain region to HSV-1 infection.
Object-location training elicits an overlapping but temporally distinct transcriptional profile from contextual fear conditioning.

Science.gov (United States)

Poplawski, Shane G; Schoch, Hannah; Wimmer, Mathieu; Hawk, Joshua D; Walsh, Jennifer L; Giese, Karl P; Abel, Ted

2014-12-01

Hippocampus-dependent learning is known to induce changes in gene expression, but information on gene expression differences between different learning paradigms that require the hippocampus is limited. The bulk of studies investigating RNA expression after learning use the contextual fear conditioning task, which couples a novel environment with a footshock. Although contextual fear conditioning has been useful in discovering gene targets, gene expression after spatial memory tasks has received less attention. In this study, we used the object-location memory task and studied gene expression at two time points after learning in a high-throughput manner using a microfluidic qPCR approach. We found that expression of the classic immediate-early genes changes after object-location training in a fashion similar to that observed after contextual fear conditioning. However, the temporal dynamics of gene expression are different between the two tasks, with object-location memory producing gene expression changes that last at least 2 hours. Our findings indicate that different training paradigms may give rise to distinct temporal dynamics of gene expression after learning. Copyright © 2014 Elsevier Inc. All rights reserved.
Object-Location Training Elicits an Overlapping but Temporally Distinct Transcriptional Profile from Contextual Fear Conditioning

Science.gov (United States)

Wimmer, Mathieu; Hawk, Joshua D.; Walsh, Jennifer L.; Giese, Karl P.; Abel, Ted

2014-01-01

Hippocampus-dependent learning is known to induce changes in gene expression, but information on gene expression differences between different learning paradigms that require the hippocampus is limited. The bulk of studies investigating RNA expression after learning use the contextual fear conditioning task, which couples a novel environment with a footshock. Although contextual fear conditioning has been useful in discovering gene targets, gene expression after spatial memory tasks has received less attention. In this study, we used the object-location memory task and studied gene expression at two time points after learning in a high-throughput manner using a microfluidic qPCR approach. We found that expression of the classic immediate-early genes changes after object-location training in a fashion similar to that observed after contextual fear conditioning. However, the temporal dynamics of gene expression are different between the two tasks, with object-location memory producing gene expression changes that last at least 2 hours. Our findings indicate that different training paradigms may give rise to distinct temporal dynamics of gene expression after learning. PMID:25242102
Optimization of Mechanical Expression of Castor Seeds Oil (Ricinus communis using Response Surface Methodology

Directory of Open Access Journals (Sweden)

J. O. Olaoye

2017-12-01

Full Text Available The effect of the processing parameters of Castor seed on its oil yield was investigated. The castor seeds were passed through drying, crushing and separation into seeds and shells. These processing conditions were further succeeded by seed roasting and subsequent mechanical expression of the roasted nut by means of screw press in the course of its preparation for oil expression. Seed samples were conditioned by adding calculated amount of distilled water to obtain different moisture levels from the initial moisture content of the seeds. Samples were roasted at the temperatures of 83.18, 90.00, 100.00, 110.00 and 116.82°C, over periods of 6.59, 10.00, 15.00, 20.00 and 23.41min, seed moisture content of 6.32, 7.00, 8.00, 9.00 and 9.68 % wb, respectively and the oil was expressed using a screw roaster-expeller. Optimization of the oil expression process was achieved by applying Central Composite Rotatable Design of Response Surface Methodology. The optimal conditions for oil yield within the experimental range of the studied variables were 7%, 110°C and 20 min; moisture content, roasting temperature and roasting duration respectively. These values of the optimum process conditions were used to predict optimum value of oil yield to be 25.77%. A second-order model was obtained to predict oil yield as a function of moisture content, heating temperature and duration. Thus the result from this research work has established the optimal conditions for mechanical extraction of oil from castor seed. Closed agreement between experimental and predicted yield was obtained.
Loss of ATP-Sensitive Potassium Channel Surface Expression in Heart Failure Underlies Dysregulation of Action Potential Duration and Myocardial Vulnerability to Injury.

Directory of Open Access Journals (Sweden)

Zhan Gao

Full Text Available The search for new approaches to treatment and prevention of heart failure is a major challenge in medicine. The adenosine triphosphate-sensitive potassium (KATP channel has been long associated with the ability to preserve myocardial function and viability under stress. High surface expression of membrane KATP channels ensures a rapid energy-sparing reduction in action potential duration (APD in response to metabolic challenges, while cellular signaling that reduces surface KATP channel expression blunts APD shortening, thus sacrificing energetic efficiency in exchange for greater cellular calcium entry and increased contractile force. In healthy hearts, calcium/calmodulin-dependent protein kinase II (CaMKII phosphorylates the Kir6.2 KATP channel subunit initiating a cascade responsible for KATP channel endocytosis. Here, activation of CaMKII in a transaortic banding (TAB model of heart failure is coupled with a 35-40% reduction in surface expression of KATP channels compared to hearts from sham-operated mice. Linkage between KATP channel expression and CaMKII is verified in isolated cardiomyocytes in which activation of CaMKII results in downregulation of KATP channel current. Accordingly, shortening of monophasic APD is slowed in response to hypoxia or heart rate acceleration in failing compared to non-failing hearts, a phenomenon previously shown to result in significant increases in oxygen consumption. Even in the absence of coronary artery disease, failing myocardium can be further injured by ischemia due to a mismatch between metabolic supply and demand. Ischemia-reperfusion injury, following ischemic preconditioning, is diminished in hearts with CaMKII inhibition compared to wild-type hearts and this advantage is largely eliminated when myocardial KATP channel expression is absent, supporting that the myocardial protective benefit of CaMKII inhibition in heart failure may be substantially mediated by KATP channels. Recognition of Ca
Fowler's approximation for the surface tension and surface energy of Lennard-Jones fluids revisited

International Nuclear Information System (INIS)

Mulero, A; Galan, C; Cuadros, F

2003-01-01

We present a detailed study of the validity of Fowler's approximation for calculating the surface tension and the surface energy of Lennard-Jones fluids. To do so, we consider three different explicit analytical expressions for the radial distribution function (RDF), including one proposed by our research group, together with very accurate expressions for the liquid and vapour densities, also proposed by our group. The calculation of the surface tension from the direct correlation function using both the Percus-Yevick and the hypernetted-chain approximations is also considered. Finally, our results are compared with those obtained by other authors by computer simulations or through relevant theoretical approximations. In particular, we consider the analytical expression proposed by Kalikmanov and Hofmans (1994 J. Phys.: Condens. Matter 6 2207-14) for the surface tension. Our results indicate that the values for the surface energy in Fowler's approximation obtained by other authors are adequate, and can be calculated from the RDF models. For the surface tension, however, the values considered as valid in previous works seem to be incorrect. The correct values can be obtained from our model for the RDF or from the Kalikmanov and Hofmans expression with suitable inputs
Genetic Variants Contribute to Gene Expression Variability in Humans

Science.gov (United States)

Hulse, Amanda M.; Cai, James J.

2013-01-01

Expression quantitative trait loci (eQTL) studies have established convincing relationships between genetic variants and gene expression. Most of these studies focused on the mean of gene expression level, but not the variance of gene expression level (i.e., gene expression variability). In the present study, we systematically explore genome-wide association between genetic variants and gene expression variability in humans. We adapt the double generalized linear model (dglm) to simultaneously fit the means and the variances of gene expression among the three possible genotypes of a biallelic SNP. The genomic loci showing significant association between the variances of gene expression and the genotypes are termed expression variability QTL (evQTL). Using a data set of gene expression in lymphoblastoid cell lines (LCLs) derived from 210 HapMap individuals, we identify cis-acting evQTL involving 218 distinct genes, among which 8 genes, ADCY1, CTNNA2, DAAM2, FERMT2, IL6, PLOD2, SNX7, and TNFRSF11B, are cross-validated using an extra expression data set of the same LCLs. We also identify ∼300 trans-acting evQTL between >13,000 common SNPs and 500 randomly selected representative genes. We employ two distinct scenarios, emphasizing single-SNP and multiple-SNP effects on expression variability, to explain the formation of evQTL. We argue that detecting evQTL may represent a novel method for effectively screening for genetic interactions, especially when the multiple-SNP influence on expression variability is implied. The implication of our results for revealing genetic mechanisms of gene expression variability is discussed. PMID:23150607
Genes expressed in specific areas of the human fetal cerebral cortex display distinct patterns of evolution.

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Nelle Lambert

2011-03-01

Full Text Available The developmental mechanisms through which the cerebral cortex increased in size and complexity during primate evolution are essentially unknown. To uncover genetic networks active in the developing cerebral cortex, we combined three-dimensional reconstruction of human fetal brains at midgestation and whole genome expression profiling. This novel approach enabled transcriptional characterization of neurons from accurately defined cortical regions containing presumptive Broca and Wernicke language areas, as well as surrounding associative areas. We identified hundreds of genes displaying differential expression between the two regions, but no significant difference in gene expression between left and right hemispheres. Validation by qRTPCR and in situ hybridization confirmed the robustness of our approach and revealed novel patterns of area- and layer-specific expression throughout the developing cortex. Genes differentially expressed between cortical areas were significantly associated with fast-evolving non-coding sequences harboring human-specific substitutions that could lead to divergence in their repertoires of transcription factor binding sites. Strikingly, while some of these sequences were accelerated in the human lineage only, many others were accelerated in chimpanzee and/or mouse lineages, indicating that genes important for cortical development may be particularly prone to changes in transcriptional regulation across mammals. Genes differentially expressed between cortical regions were also enriched for transcriptional targets of FoxP2, a key gene for the acquisition of language abilities in humans. Our findings point to a subset of genes with a unique combination of cortical areal expression and evolutionary patterns, suggesting that they play important roles in the transcriptional network underlying human-specific neural traits.

BMPs regulate msx gene expression in the dorsal neuroectoderm of Drosophila and vertebrates by distinct mechanisms.

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Esteves, Francisco F; Springhorn, Alexander; Kague, Erika; Taylor, Erika; Pyrowolakis, George; Fisher, Shannon; Bier, Ethan

2014-09-01

In a broad variety of bilaterian species the trunk central nervous system (CNS) derives from three primary rows of neuroblasts. The fates of these neural progenitor cells are determined in part by three conserved transcription factors: vnd/nkx2.2, ind/gsh and msh/msx in Drosophila melanogaster/vertebrates, which are expressed in corresponding non-overlapping patterns along the dorsal-ventral axis. While this conserved suite of "neural identity" gene expression strongly suggests a common ancestral origin for the patterning systems, it is unclear whether the original regulatory mechanisms establishing these patterns have been similarly conserved during evolution. In Drosophila, genetic evidence suggests that Bone Morphogenetic Proteins (BMPs) act in a dosage-dependent fashion to repress expression of neural identity genes. BMPs also play a dose-dependent role in patterning the dorsal and lateral regions of the vertebrate CNS, however, the mechanism by which they achieve such patterning has not yet been clearly established. In this report, we examine the mechanisms by which BMPs act on cis-regulatory modules (CRMs) that control localized expression of the Drosophila msh and zebrafish (Danio rerio) msxB in the dorsal central nervous system (CNS). Our analysis suggests that BMPs act differently in these organisms to regulate similar patterns of gene expression in the neuroectoderm: repressing msh expression in Drosophila, while activating msxB expression in the zebrafish. These findings suggest that the mechanisms by which the BMP gradient patterns the dorsal neuroectoderm have reversed since the divergence of these two ancient lineages.
BMPs regulate msx gene expression in the dorsal neuroectoderm of Drosophila and vertebrates by distinct mechanisms.

Directory of Open Access Journals (Sweden)

Francisco F Esteves

2014-09-01

Full Text Available In a broad variety of bilaterian species the trunk central nervous system (CNS derives from three primary rows of neuroblasts. The fates of these neural progenitor cells are determined in part by three conserved transcription factors: vnd/nkx2.2, ind/gsh and msh/msx in Drosophila melanogaster/vertebrates, which are expressed in corresponding non-overlapping patterns along the dorsal-ventral axis. While this conserved suite of "neural identity" gene expression strongly suggests a common ancestral origin for the patterning systems, it is unclear whether the original regulatory mechanisms establishing these patterns have been similarly conserved during evolution. In Drosophila, genetic evidence suggests that Bone Morphogenetic Proteins (BMPs act in a dosage-dependent fashion to repress expression of neural identity genes. BMPs also play a dose-dependent role in patterning the dorsal and lateral regions of the vertebrate CNS, however, the mechanism by which they achieve such patterning has not yet been clearly established. In this report, we examine the mechanisms by which BMPs act on cis-regulatory modules (CRMs that control localized expression of the Drosophila msh and zebrafish (Danio rerio msxB in the dorsal central nervous system (CNS. Our analysis suggests that BMPs act differently in these organisms to regulate similar patterns of gene expression in the neuroectoderm: repressing msh expression in Drosophila, while activating msxB expression in the zebrafish. These findings suggest that the mechanisms by which the BMP gradient patterns the dorsal neuroectoderm have reversed since the divergence of these two ancient lineages.
Improving the performance of industrial ethanol-producing yeast by expressing the aspartyl protease on the cell surface.

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Guo, Zhong-peng; Zhang, Liang; Ding, Zhong-yang; Wang, Zheng-Xiang; Shi, Gui-Yang

2010-12-01

The yeasts used in fuel ethanol manufacture are unable to metabolize soluble proteins. The PEP4 gene, encoding a vacuolar aspartyl protease in Saccharomyces cerevisiae, was either secretively or cell-surface anchored expressed in industrial ethanol-producing S. cerevisiae. The obtained recombinant strains APA (expressing the protease secretively) and APB (expressing the protease on the cell wall) were studied under ethanol fermentation conditions in feed barley cultures. The effects of expression of the protease on product formation, growth and cell protein content were measured. The biomass yield of the wild-type was clearly lower than that of the recombinant strains (0.578 ± 0.12 g biomass/g glucose for APA and 0.582 ± 0.08 g biomass/g glucose for APB). In addition, nearly 98-99% of the theoretical maximum level of ethanol yield was achieved (relative to the amount of substrate consumed) for the recombinant strains, while limiting the nitrogen source resulted in dissatisfactory fermentation for the wild-type and more than 30 g/l residual sugar was detected at the end of fermentation. In addition, higher growth rate, viability and lower yields of byproducts such as glycerol and pyruvic acid for recombinant strains were observed. Expressing acid protease can be expected to lead to a significant increase in ethanol productivity. Copyright © 2010 John Wiley & Sons, Ltd.
Distinct gene expression profile of Xanthomonas retroflexus engaged in synergistic multispecies biofilm formation

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Hansen, Lea Benedicte Skov; Ren, Dawei; Burmølle, Mette

2017-01-01

biofilms. This study presents a comparative gene expression analysis of the Xanthomonas retroflexus transcriptome when grown in a single-species biofilm and in dual-and four-species consortia with Stenotrophomonas rhizophila, Microbacterium oxydans and Paenibacillus amylolyticus. The results revealed...
Multiple distinct subtypes of GABAergic neurons in mouse visual cortex identified by triple immunostaining

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Yuri Gonchar

2008-03-01

Full Text Available The majority of cortical interneurons use GABA (gamma amino butyric acid as inhibitory neurotransmitter. GABAergic neurons are morphologically, connectionally, electrically and chemically heterogeneous. In rat cerebral cortex three distinct groups of GABAergic interneurons have been identifi ed by the expression of parvalbumin (PV, calretinin (CR and somatostatin (SOM. Recent studies in mouse cerebral cortex have revealed a different organization in which the CR and SOM populations are partially overlapping. Because CR and SOM neurons derive from different progenitors located in different embryonic structures, the coexpression of CR + SOM suggests that the chemical differentiation of interneurons is regulated postmitotically. Here, we have taken an important fi rst step towards understanding this process by triple immunostaining mouse visual cortex with a panel of antibodies, which has been used extensively for classifying developing interneurons. We have found at least 13 distinct groups of GABAergic neurons which include PV, CR, SOM, CCK (cholecystokinin, CR + SOM, CR + NPY (neuropeptide Y, CR + VIP (vasointestinal polypeptide, SOM + NPY, SOM + VIP, VIP + ChAT (choline acetyltransferase, CCK + NPY, CR + SOM + NPY and CR + SOM + VIP expressing cells. Triple immunostaining with PV, CR and SOM antibodies during postnatal development further showed that PV is never colocalized with CR and SOM. Importantly, expression of SOM and CR + SOM developed after the percentage of CR cells that do not express SOM has reached the mature level, suggesting that the chemical differentiation of SOM and CR + SOM neurons is a postnatal event, which may be controlled by transcriptional regulation.
Distinct sperm nucleus behaviors between genotypic and temperature-dependent sex determination males are associated with replication and expression-related pathways in a gynogenetic fish.

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Zhu, Yao-Jun; Li, Xi-Yin; Zhang, Jun; Li, Zhi; Ding, Miao; Zhang, Xiao-Juan; Zhou, Li; Gui, Jian-Fang

2018-06-05

Coexistence and transition of diverse sex determination strategies have been revealed in some ectothermic species, but the variation between males caused by different sex determination strategies and the underlying mechanism remain unclear. Here, we used the gynogenetic gibel carp (Carassius gibelio) with both genotypic sex determination (GSD) and temperature-dependent sex determination (TSD) strategies to illustrate this issue. We found out that males of GSD and TSD in gibel carp had similar morphology, testicular histology, sperm structure and sperm vitality. However, when maternal individuals were mated with males of GSD, sperm nucleus swelling and fusing with the female pronucleus were observed in the fertilized eggs. On the contrary, when maternal individuals were mated with males of TSD, sperm nucleus remained in the condensed status throughout the whole process. Subsequently, semen proteomics analysis unveiled that DNA replication and gene expression-related pathways were inhibited in the sperm from males of TSD compared to males of GSD, and most differentially expressed proteins associated with DNA replication, transcription and translation were down-regulated. Moreover, via BrdU incorporation and immunofluorescence detection, male nucleus replication was revealed to be present in the fertilized eggs by the sperm from males of GSD, but absent in the fertilized eggs by the sperm from males of TSD. These findings indicate that DNA replication and gene expression-related pathways are associated with the distinct sperm nucleus development behaviors in fertilized eggs in response to the sperm from males of GSD and TSD. And this study is the first attempt to screen the differences between males determined via GSD and TSD in gynogenetic species, which might give a hint for understanding evolutionary adaption of diverse sex determination mechanisms in unisexual vertebrates.
The distinct proteome of placental malaria parasites.

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Fried, Michal; Hixson, Kim K.; Anderson, Lori; Ogata, Yuko; Mutabingwa, Theonest K.; Duffy, Patrick E.

2007-09-01

Malaria proteins expressed on the surface of Plasmodium falciparum infected erythrocytes (IE) mediate adhesion and are targeted by protective immune responses. During pregnancy, IE sequester in the placenta. Placental IE bind to the molecule chondroitin sulfate A (CSA) and preferentially transcribe the gene that encodes VAR2CSA, a member of the PfEMP1 variant surface antigen family. Over successive pregnancies women develop specific immunity to CSA-binding IE and antibodies to VAR2CSA. We used tandem mass spectrometry together with accurate mass and time tag technology to study IE membrane fractions of placental parasites. VAR2CSA peptides were detected in placental IE and in IE from children, but the MC variant of VAR2CSA was specifically associated with placental IE. We identified six conserved hypothetical proteins with putative TM or signal peptides that were exclusively expressed by the placental IE, and 11 such proteins that were significantly more abundant in placental IE. One of these hypothetical proteins, PFI1785w, is a 42kDa molecule detected by Western blot in parasites infecting pregnant women but not those infecting children.
Nanoroughened adhesion-based capture of circulating tumor cells with heterogeneous expression and metastatic characteristics

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Chen, Weiqiang; Allen, Steven G.; Reka, Ajaya Kumar; Qian, Weiyi; Han, Shuo; Zhao, Jianing; Bao, Liwei; Keshamouni, Venkateshwar G.; Merajver, Sofia D.; Fu, Jianping

2016-01-01

Circulating tumor cells (CTCs) have shown prognostic relevance in many cancer types. However, the majority of current CTC capture methods rely on positive selection techniques that require a priori knowledge about the surface protein expression of disseminated CTCs, which are known to be a dynamic population. We developed a microfluidic CTC capture chip that incorporated a nanoroughened glass substrate for capturing CTCs from blood samples. Our CTC capture chip utilized the differential adhesion preference of cancer cells to nanoroughened etched glass surfaces as compared to normal blood cells and thus did not depend on the physical size or surface protein expression of CTCs. The microfluidic CTC capture chip was able to achieve a superior capture yield for both epithelial cell adhesion molecule positive (EpCAM+) and EpCAM- cancer cells in blood samples. Additionally, the microfluidic CTC chip captured CTCs undergoing transforming growth factor beta-induced epithelial-to-mesenchymal transition (TGF-β-induced EMT) with dynamically down-regulated EpCAM expression. In a mouse model of human breast cancer using EpCAM positive and negative cell lines, the number of CTCs captured correlated positively with the size of the primary tumor and was independent of their EpCAM expression. Furthermore, in a syngeneic mouse model of lung cancer using cell lines with differential metastasis capability, CTCs were captured from all mice with detectable primary tumors independent of the cell lines’ metastatic ability. The microfluidic CTC capture chip using a novel nanoroughened glass substrate is broadly applicable to capturing heterogeneous CTC populations of clinical interest independent of their surface marker expression and metastatic propensity. We were able to capture CTCs from a non-metastatic lung cancer model, demonstrating the potential of the chip to collect the entirety of CTC populations including subgroups of distinct biological and phenotypical properties. Further
Expression of the Ly-6 family proteins Lynx1 and Ly6H in the rat brain is compartmentalized, cell-type specific, and developmentally regulated

DEFF Research Database (Denmark)

Thomsen, Morten Skøtt; Cinar, Betül; Jensen, Majbrit Myrup

2014-01-01

regarding the distribution and developmental regulation of these proteins in the brain. We use protein cross-linking and synaptosomal fractions to demonstrate that the Ly-6 proteins Lynx1 and Ly6H are membrane-bound proteins in the brain, which are present on the cell surface and localize to synaptic...... demonstrate that Lynx1 and Ly6H are expressed in cultured neurons, but not cultured micro- or astroglial cultures. In addition, Lynx1, but not Ly6H was detected in the CSF. Finally, we show that the Ly-6 proteins Lynx1, Lynx2, Ly6H, and PSCA, display distinct expression patterns during postnatal development...
Postnatal Expression of V2 Vasopressin Receptor Splice Variants in the Rat Cerebellum

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Vargas, Karina J.; Sarmiento, José M.; Ehrenfeld, Pamela; Añazco, Carolina C.; Villanueva, Carolina I.; Carmona, Pamela L.; Brenet, Marianne; Navarro, Javier; Müller-Esterl, Werner; Figueroa, Carlos D.; González, Carlos B.

2010-01-01

The V2 vasopressin receptor gene contains an alternative splice site in exon-3, which leads to the generation of two splice variants (V2a and V2b) first identified in the kidney. The open reading frame of the alternatively spliced V2b transcripten codes a truncated receptor, showing the same amino acid sequence as the canonical V2a receptor up to the 6th transmembrane segment, but displaying a distinct sequence to the corresponding 7th transmembrane segment and C-terminal domain relative to the V2a receptor. Here, we demonstrate the postnatal expression of V2a and V2b variants in the rat cerebellum. Most importantly, we showed by in situ hybridization and immunocytochemistry that both V2 splice variants were preferentially expressed in Purkinje cells, from early to late postnatal development. In addition, both variants were transiently expressed in the neuroblastic external granule cells and Bergmann fibers. These results indicate that the cellular distributions of both splice variants are developmentally regulated, and suggest that the transient expression of the V2 receptor is involved in the mechanisms of cerebellar cytodifferentiation by AVP. Finally, transfected CHO-K1 .expressing similar amounts of both V2 splice variants, as that found in the cerebellum, showed a significant reduction in the surface expression of V2a receptors, suggesting that the differential expression of the V2 splice variants regulate the vasopressin signaling in the cerebellum. PMID:19281786
Surface morphology of some articulated corallines from India

Digital Repository Service at National Institute of Oceanography (India)

Kerkar, V.; Iyer, S.D.

Surface structures of seven species belonging to four genera of subfamily Corallinoidae (Fly: Corallinaceae) were microscopically investigated Two distinct surface morphologies were revealed namely a 'Corallina type' (c-type) with round to irregular...
Ray-map migration of transmitted surface waves

KAUST Repository

Li, Jing; Schuster, Gerard T.

2016-01-01

Near-surface normal faults can sometimes separate two distinct zones of velocity heterogeneity, where the medium on one side of the fault has a faster velocity than on the other side. Therefore, the slope of surface-wave arrivals in a common
Distinct mechanisms of axonal globule formation in mice expressing human wild type α-synuclein or dementia with Lewy bodies-linked P123H ß-synuclein

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Sekigawa Akio

2012-09-01

Full Text Available Abstract Background Axonopathy is critical in the early pathogenesis of neurodegenerative diseases, including Parkinson’s disease (PD and dementia with Lewy bodies (DLB. Axonal swellings such as globules and spheroids are a distinct feature of axonopathy and our recent study showed that transgenic (tg mice expressing DLB-linked P123H β-synuclein (P123H βS were characterized by P123H βS-immunoreactive axonal swellings (P123H βS-globules. Therefore, the objectives of this study were to evaluate α-synuclein (αS-immunoreactive axonal swellings (αS-globules in the brains of tg mice expressing human wild-type αS and to compare them with the globules in P123H βS tg mice. Results In αS tg mice, αS-globules were formed in an age-dependent manner in various brain regions, including the thalamus and basal ganglia. These globules were composed of autophagosome-like membranous structures and were reminiscent of P123H βS-globules in P123H βS tg mice. In the αS-globules, frequent clustering and deformation of mitochondria were observed. These changes were associated with oxidative stress, based on staining of nitrated αS and 4-hydroxy-2-nonenal (4-HNE. In accord with the absence of mitochondria in the P123H βS-globules, staining of nitrated αS and 4-HNE in these globules was weaker than that for αS-globules. Leucine-rich repeat kinase 2 (LRRK2, the PARK8 of familial PD, was detected exclusively in αS-globules, suggesting a specific role of this molecule in these globules. Conclusions Lysosomal pathology was similarly observed for both αS- and P123H βS-globules, while oxidative stress was associated with the αS-globules, and to a lesser extent with the P123H βS-globules. Other pathologies, such as mitochondrial alteration and LRRK2 accumulation, were exclusively detected for αS-globules. Collectively, both αS- and P123H βS-globules were formed through similar but distinct pathogenic mechanisms. Our findings suggest that synuclein
Cardiomyocyte expression and cell-specific processing of procholecystokinin

DEFF Research Database (Denmark)

Gøtze, Jens P.; Johnsen, Anders H.; Kistorp, Caroline

2015-01-01

has only been suggested using transcriptional measures or methods, with the post-translational phase of gene expression unaddressed. In this study, we examined the cardiac expression of the CCK gene in adult mammals and its expression at the protein level. Using quantitative PCR, a library of sequence......-specific pro-CCK assays, peptide purification, and mass spectrometry, we demonstrate that the mammalian heart expresses pro-CCK in amounts comparable to natriuretic prohormones and processes it to a unique, triple-sulfated, and N-terminally truncated product distinct from intestinal and cerebral CCK peptides...
CD40 expression in Wehi-164 cell line.

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Karimi, Mohammad Hossein; Ebadi, Padideh; Pourfathollah, Ali Akbar; Soheili, Zahra Soheila; Moazzeni, Seyed Mohammad

2010-07-01

CD40-CD154 interaction is an important process for cellular and humoral immunity regulation and can be effective in the body's defense against tumors. In the present study, we evaluated the expression of CD40 in Wehi-164 cell line. CD40 expressions on the cell surface and in the cytoplasm were assessed by flow cytometry and intracellular staining assay, respectively. Also, the mRNA expression was identified by real time-PCR. The obtained results showed the high mRNA and cytoplasmic protein expression of CD40 but no surface expression. These results suggest that the Wehi-164 cell line down regulates expression of CD40 on the surface for evasion of immune system.
CXCR3 surface expression in human airway epithelial cells: cell cycle dependence and effect on cell proliferation.

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Aksoy, Mark O; Yang, Yi; Ji, Rong; Reddy, P J; Shahabuddin, Syed; Litvin, Judith; Rogers, Thomas J; Kelsen, Steven G

2006-05-01

We recently demonstrated that human bronchial epithelial cells (HBEC) constitutively express the CXC chemokine receptor CXCR3, which when activated, induces directed cell migration. The present study in HBEC examined the relative expression of the CXCR3 splice variants CXCR3-A and -B, cell cycle dependence of CXCR3 expression, and the effects of the CXCR3 ligand, the interferon-gamma-inducible CXC chemokine I-TAC/CXCL11, on DNA synthesis and cell proliferation. Both CXCR3-A and -B mRNA, assessed by real-time RT-PCR, were expressed in normal HBEC (NHBEC) and the HBEC line 16-HBE. However, CXCR3-B mRNA was 39- and 6-fold greater than CXCR3-A mRNA in NHBEC and 16-HBE, respectively. Although most HBEC (>80%) assessed by flow cytometry and immunofluorescence microscopy contained intracellular CXCR3, only a minority (75%) were in the S + G(2)/M phases of the cell cycle. Stimulation of CXCR3 with I-TAC enhanced thymidine incorporation and cell proliferation and increased p38 and ERK1/2 phosphorylation. These data indicate that 1) human airway epithelial cells primarily express CXCR3-B mRNA, 2) surface expression of CXCR3 is largely confined to the S + G(2)/M phases of the cell cycle, and 3) activation of CXCR3 induces DNA synthesis, cell proliferation, and activation of MAPK pathways. We speculate that activation of CXCR3 exerts a mitogenic effect in HBEC, which may be important during airway mucosal injury in obstructive airway diseases such as asthma and chronic obstructive pulmonary disease.
Distinct interneuron types express m2 muscarinic receptor immunoreactivity on their dendrites or axon terminals in the hippocampus.

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Hájos, N; Papp, E C; Acsády, L; Levey, A I; Freund, T F

1998-01-01

In previous studies m2 muscarinic acetylcholine receptor-immunoreactive interneurons and various types of m2-positive axon terminals have been described in the hippocampal formation. The aim of the present study was to identify the types of interneurons expressing m2 receptor and to examine whether the somadendritic and axonal m2 immunostaining labels the same or distinct cell populations. In the CA1 subfield, neurons immunoreactive for m2 have horizontal dendrites, they are located at the stratum oriens/alveus border and have an axon that project to the dendritic region of pyramidal cells. In the CA3 subfield and the hilus, m2-positive neurons are multipolar and are scattered in all layers except stratum lacunosum-moleculare. In stratum pyramidale of the CA1 and CA3 regions, striking axon terminal staining for m2 was observed, surrounding the somata and axon initial segments of pyramidal cells in a basket-like manner. The co-localization of m2 with neurochemical markers and GABA was studied using the "mirror" technique and fluorescent double-immunostaining at the light microscopic level and with double-labelling using colloidal gold-conjugated antisera and immunoperoxidase reaction (diaminobenzidine) at the electron microscopic level. GABA was shown to be present in the somata of most m2-immunoreactive interneurons, as well as in the majority of m2-positive terminals in all layers. The calcium-binding protein parvalbumin was absent from practically all m2-immunoreactive cell bodies and dendrites. In contrast, many of the terminals synapsing on pyramidal cell somata and axon initial segments co-localized parvalbumin and m2, suggesting a differential distribution of m2 receptor immunoreactivity on the axonal and somadendritic membrane of parvalbumin-containing basket and axo-axonic cells. The co-existence of m2 receptors with the calcium-binding protein calbindin and the neuropeptides cholecystokinin and vasoactive intestinal polypeptide was rare throughout the
Characterization of Trichome-Expressed BAHD Acyltransferases in Petunia axillaris Reveals Distinct Acylsugar Assembly Mechanisms within the Solanaceae.

Science.gov (United States)

Nadakuduti, Satya Swathi; Uebler, Joseph B; Liu, Xiaoxiao; Jones, A Daniel; Barry, Cornelius S

2017-09-01

Acylsugars are synthesized in the glandular trichomes of the Solanaceae family and are implicated in protection against abiotic and biotic stress. Acylsugars are composed of either sucrose or glucose esterified with varying numbers of acyl chains of differing length. In tomato ( Solanum lycopersicum ), acylsugar assembly requires four acylsugar acyltransferases (ASATs) of the BAHD superfamily. Tomato ASATs catalyze the sequential esterification of acyl-coenzyme A thioesters to the R4, R3, R3', and R2 positions of sucrose, yielding a tetra-acylsucrose. Petunia spp. synthesize acylsugars that are structurally distinct from those of tomato. To explore the mechanisms underlying this chemical diversity, a Petunia axillaris transcriptome was mined for trichome preferentially expressed BAHDs. A combination of phylogenetic analyses, gene silencing, and biochemical analyses coupled with structural elucidation of metabolites revealed that acylsugar assembly is not conserved between tomato and petunia. In P. axillaris , tetra-acylsucrose assembly occurs through the action of four ASATs, which catalyze sequential addition of acyl groups to the R2, R4, R3, and R6 positions. Notably, in P. axillaris , PaxASAT1 and PaxASAT4 catalyze the acylation of the R2 and R6 positions of sucrose, respectively, and no clear orthologs exist in tomato. Similarly, petunia acylsugars lack an acyl group at the R3' position, and congruently, an ortholog of SlASAT3, which catalyzes acylation at the R3' position in tomato, is absent in P. axillaris Furthermore, where putative orthologous relationships of ASATs are predicted between tomato and petunia, these are not supported by biochemical assays. Overall, these data demonstrate the considerable evolutionary plasticity of acylsugar biosynthesis. © 2017 American Society of Plant Biologists. All Rights Reserved.
Streptococcus pneumoniae and Pseudomonas aeruginosa pneumonia induce distinct host responses.

Science.gov (United States)

McConnell, Kevin W; McDunn, Jonathan E; Clark, Andrew T; Dunne, W Michael; Dixon, David J; Turnbull, Isaiah R; Dipasco, Peter J; Osberghaus, William F; Sherman, Benjamin; Martin, James R; Walter, Michael J; Cobb, J Perren; Buchman, Timothy G; Hotchkiss, Richard S; Coopersmith, Craig M

2010-01-01

Pathogens that cause pneumonia may be treated in a targeted fashion by antibiotics, but if this therapy fails, then treatment involves only nonspecific supportive measures, independent of the inciting infection. The purpose of this study was to determine whether host response is similar after disparate infections with similar mortalities. Prospective, randomized controlled study. Animal laboratory in a university medical center. Pneumonia was induced in FVB/N mice by either Streptococcus pneumoniae or two different concentrations of Pseudomonas aeruginosa. Plasma and bronchoalveolar lavage fluid from septic animals was assayed by a microarray immunoassay measuring 18 inflammatory mediators at multiple time points. The host response was dependent on the causative organism as well as kinetics of mortality, but the pro-inflammatory and anti-inflammatory responses were independent of inoculum concentration or degree of bacteremia. Pneumonia caused by different concentrations of the same bacteria, Pseudomonas aeruginosa, also yielded distinct inflammatory responses; however, inflammatory mediator expression did not directly track the severity of infection. For all infections, the host response was compartmentalized, with markedly different concentrations of inflammatory mediators in the systemic circulation and the lungs. Hierarchical clustering analysis resulted in the identification of five distinct clusters of the host response to bacterial infection. Principal components analysis correlated pulmonary macrophage inflammatory peptide-2 and interleukin-10 with progression of infection, whereas elevated plasma tumor necrosis factor sr2 and macrophage chemotactic peptide-1 were indicative of fulminant disease with >90% mortality within 48 hrs. Septic mice have distinct local and systemic responses to Streptococcus pneumoniae and Pseudomonas aeruginosa pneumonia. Targeting specific host inflammatory responses induced by distinct bacterial infections could represent a
Distinct differences in global gene expression profiles in non-implanted blastocysts and blastocysts resulting in live birth

DEFF Research Database (Denmark)

Kirkegaard, Kirstine Kjær; Fredsted, Palle Villesen; Jensen, Jacob Malte

2015-01-01

Results from animal models points towards the existence of a gene expression profile that is distinguishably different in viable embryos compared with non-viable embryos. Knowledge of human embryo transcripts is however limited, in particular with regard to how gene expression is related...... to clinical outcome. The purpose of the present study was therefore to determine the global gene expression profiles of human blastocysts. Next Generation Sequencing was used to identify genes that were differentially expressed in non-implanted embryos and embryos resulting in live birth. Three trophectoderm...

Mapping surface charge density of lipid bilayers by quantitative surface conductivity microscopy

DEFF Research Database (Denmark)

Klausen, Lasse Hyldgaard; Fuhs, Thomas; Dong, Mingdong

2016-01-01

Local surface charge density of lipid membranes influences membrane-protein interactions leading to distinct functions in all living cells, and it is a vital parameter in understanding membrane-binding mechanisms, liposome design and drug delivery. Despite the significance, no method has so far...
CHARACTERIZATION OF THE OLFACTORY RECEPTORS EXPRESSED IN HUMAN SPERMATOZOA

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Caroline eFlegel

2016-01-01

Full Text Available The detection of external cues is fundamental for human spermatozoa to locate the oocyte in the female reproductive tract. This task requires a specific chemoreceptor repertoire that is expressed on the surface of human spermatozoa, which is not fully identified to date. Olfactory receptors (ORs are candidate molecules and have been attributed to be involved in sperm chemotaxis and chemokinesis, indicating an important role in mammalian spermatozoa. An increasing importance has been suggested for spermatozoal RNA, which led us to investigate the expression of all 387 OR genes. This study provides the first comprehensive analysis of OR transcripts in human spermatozoa of several individuals by RNA-Seq. We detected 91 different transcripts in the spermatozoa samples that could be aligned to annotated OR genes. Using stranded mRNA-Seq, we detected a class of these putative OR transcripts in an antisense orientation, indicating a different function, rather than coding for a functional OR protein. Nevertheless, we were able to detect OR proteins in various compartments of human spermatozoa, indicating distinct functions in human sperm. A panel of various OR ligands induced Ca2+ signals in human spermatozoa, which could be inhibited by mibefradil. This study indicated that a variety of ORs are expressed at the mRNA and protein level in human spermatozoa and demonstrates that ORs are involved in the physiological processes.
STUDIES IN DYNAMICS OF APOPTOSIS-RELATED SURFACE ANTIGEN (CD95 EXPRESSION ON NEUTROPHILS FROM CERVICAL AND VAGINAL SECRETIONS IN WOMEN WITH CHLAMIDIA INFECTION

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O. A. Giesinger

2010-01-01

Full Text Available CD95 (Fas/APO-1 antigen expression was studied on the surface of neutrophil granulocytes from cervical secretions. Sixty-five female patients with established Chlamydia infection were found to have an increased CD95+ antigen expression following basic therapy. CD95+ receptors on neutrophils in the patients with Chlamydia infection have been shown to return to normal levels following a combined magnetic laser treatment.
MicroRNA expression data reveals a signature of kidney damage following ischemia reperfusion injury.

Directory of Open Access Journals (Sweden)

Michael D Shapiro

Full Text Available Ischemia reperfusion injury (IRI is a leading cause of acute kidney injury, a common problem worldwide associated with significant morbidity and mortality. We have recently examined the role of microRNAs (miRs in renal IRI using expression profiling. Here we conducted mathematical analyses to determine if differential expression of miRs can be used to define a biomarker of renal IRI. Principal component analysis (PCA was combined with spherical geometry to determine whether samples that underwent renal injury as a result of IRI can be distinguished from controls based on alterations in miR expression using our data set consisting of time series measuring 571 miRs. Using PCA, we examined whether changes in miR expression in the kidney following IRI have a distinct direction when compared to controls based on the trajectory of the first three principal components (PCs for our time series. We then used Monte Carlo methods and spherical geometry to assess the statistical significance of these directions. We hypothesized that if IRI and control samples exhibit distinct directions, then miR expression can be used as a biomarker of injury. Our data reveal that the pattern of miR expression in the kidney following IRI has a distinct direction based on the trajectory of the first three PCs and can be distinguished from changes observed in sham controls. Analyses of samples from immunodeficient mice indicated that the changes in miR expression observed following IRI were lymphocyte independent, and therefore represent a kidney intrinsic response to injury. Together, these data strongly support the notion that IRI results in distinct changes in miR expression that can be used as a biomarker of injury.
Gene expression changes governing extreme dehydration tolerance in an Antarctic insect

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Teets, Nicholas M.; Peyton, Justin T.; Colinet, Herve; Renault, David; Kelley, Joanna L.; Kawarasaki, Yuta; Lee, Richard E.; Denlinger, David L.

2012-01-01

Among terrestrial organisms, arthropods are especially susceptible to dehydration, given their small body size and high surface area to volume ratio. This challenge is particularly acute for polar arthropods that face near-constant desiccating conditions, as water is frozen and thus unavailable for much of the year. The molecular mechanisms that govern extreme dehydration tolerance in insects remain largely undefined. In this study, we used RNA sequencing to quantify transcriptional mechanisms of extreme dehydration tolerance in the Antarctic midge, Belgica antarctica, the world’s southernmost insect and only insect endemic to Antarctica. Larvae of B. antarctica are remarkably tolerant of dehydration, surviving losses up to 70% of their body water. Gene expression changes in response to dehydration indicated up-regulation of cellular recycling pathways including the ubiquitin-mediated proteasome and autophagy, with concurrent down-regulation of genes involved in general metabolism and ATP production. Metabolomics results revealed shifts in metabolite pools that correlated closely with changes in gene expression, indicating that coordinated changes in gene expression and metabolism are a critical component of the dehydration response. Finally, using comparative genomics, we compared our gene expression results with a transcriptomic dataset for the Arctic collembolan, Megaphorura arctica. Although B. antarctica and M. arctica are adapted to similar environments, our analysis indicated very little overlap in expression profiles between these two arthropods. Whereas several orthologous genes showed similar expression patterns, transcriptional changes were largely species specific, indicating these polar arthropods have developed distinct transcriptional mechanisms to cope with similar desiccating conditions. PMID:23197828
Vascular Gene Expression in Nonneoplastic and Malignant Brain

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Madden, Stephen L.; Cook, Brian P.; Nacht, Mariana; Weber, William D.; Callahan, Michelle R.; Jiang, Yide; Dufault, Michael R.; Zhang, Xiaoming; Zhang, Wen; Walter-Yohrling, Jennifer; Rouleau, Cecile; Akmaev, Viatcheslav R.; Wang, Clarence J.; Cao, Xiaohong; St. Martin, Thia B.; Roberts, Bruce L.; Teicher, Beverly A.; Klinger, Katherine W.; Stan, Radu-Virgil; Lucey, Brenden; Carson-Walter, Eleanor B.; Laterra, John; Walter, Kevin A.

2004-01-01

Malignant gliomas are uniformly lethal tumors whose morbidity is mediated in large part by the angiogenic response of the brain to the invading tumor. This profound angiogenic response leads to aggressive tumor invasion and destruction of surrounding brain tissue as well as blood-brain barrier breakdown and life-threatening cerebral edema. To investigate the molecular mechanisms governing the proliferation of abnormal microvasculature in malignant brain tumor patients, we have undertaken a cell-specific transcriptome analysis from surgically harvested nonneoplastic and tumor-associated endothelial cells. SAGE-derived endothelial cell gene expression patterns from glioma and nonneoplastic brain tissue reveal distinct gene expression patterns and consistent up-regulation of certain glioma endothelial marker genes across patient samples. We define the G-protein-coupled receptor RDC1 as a tumor endothelial marker whose expression is distinctly induced in tumor endothelial cells of both brain and peripheral vasculature. Further, we demonstrate that the glioma-induced gene, PV1, shows expression both restricted to endothelial cells and coincident with endothelial cell tube formation. As PV1 provides a framework for endothelial cell caveolar diaphragms, this protein may serve to enhance glioma-induced disruption of the blood-brain barrier and transendothelial exchange. Additional characterization of this extensive brain endothelial cell gene expression database will provide unique molecular insights into vascular gene expression. PMID:15277233
GARP (LRRC32) is essential for the surface expression of latent TGF-β on platelets and activated FOXP3+ regulatory T cells

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Tran, Dat Q.; Andersson, John; Wang, Rui; Ramsey, Heather; Unutmaz, Derya; Shevach, Ethan M.

2009-01-01

TGF-β family members are highly pleiotropic cytokines with diverse regulatory functions. TGF-β is normally found in the latent form associated with latency-associated peptide (LAP). This latent complex can associate with latent TGFβ-binding protein (LTBP) to produce a large latent form. Latent TGF-β is also found on the surface of activated FOXP3+ regulatory T cells (Tregs), but it is unclear how it is anchored to the cell membrane. We show that GARP or LRRC32, a leucine-rich repeat molecule of unknown function, is critical for tethering TGF-β to the cell surface. We demonstrate that platelets and activated Tregs co-express latent TGF-β and GARP on their membranes. The knockdown of GARP mRNA with siRNA prevented surface latent TGF-β expression on activated Tregs and recombinant latent TGF-β1 is able to bind directly with GARP. Confocal microscopy and immunoprecipitation strongly support their interactions. The role of TGF-β on Tregs appears to have dual functions, both for Treg-mediated suppression and infectious tolerance mechanism. PMID:19651619
GARP (LRRC32) is essential for the surface expression of latent TGF-beta on platelets and activated FOXP3+ regulatory T cells.

Science.gov (United States)

Tran, Dat Q; Andersson, John; Wang, Rui; Ramsey, Heather; Unutmaz, Derya; Shevach, Ethan M

2009-08-11

TGF-beta family members are highly pleiotropic cytokines with diverse regulatory functions. TGF-beta is normally found in the latent form associated with latency-associated peptide (LAP). This latent complex can associate with latent TGFbeta-binding protein (LTBP) to produce a large latent form. Latent TGF-beta is also found on the surface of activated FOXP3(+) regulatory T cells (Tregs), but it is unclear how it is anchored to the cell membrane. We show that GARP or LRRC32, a leucine-rich repeat molecule of unknown function, is critical for tethering TGF-beta to the cell surface. We demonstrate that platelets and activated Tregs co-express latent TGF-beta and GARP on their membranes. The knockdown of GARP mRNA with siRNA prevented surface latent TGF-beta expression on activated Tregs and recombinant latent TGF-beta1 is able to bind directly with GARP. Confocal microscopy and immunoprecipitation strongly support their interactions. The role of TGF-beta on Tregs appears to have dual functions, both for Treg-mediated suppression and infectious tolerance mechanism.
Adiponectin and Its Receptors Are Differentially Expressed in Human Tissues and Cell Lines of Distinct Origin

Directory of Open Access Journals (Sweden)

Simon Jasinski-Bergner

2017-12-01

Full Text Available Background: Adiponectin is secreted by adipose tissue and exerts high abundance and an anti-inflammatory potential. However, only little information exists about the expression profiles of adiponectin and its recently identified receptor CDH13 in non-tumorous human tissues and their association to clinical parameters. Methods: The expression levels of adiponectin and CDH13 were analyzed in heart, liver, kidney, spleen, skin, blood vessels, peripheral nerve and bone marrow of 21 human body donors, in 12 human cell lines, and in purified immune effector cell populations of healthy blood donors by immunohistochemistry, Western-blot, and semi-quantitative PCR. The obtained results were then correlated to clinical parameters, including age, sex and known diseases like cardiovascular and renal diseases. Results: Adiponectin expression in renal corpuscles was significantly higher in humans with known renal diseases. A coordinated expression of adiponectin and CDH13 was observed in the myocard. High levels of adiponectin could be detected in the bone marrow, in certain lymphoid tumor cell lines and in purified immune effector cell populations of healthy donors, in particular in cytotoxic T cells. Conclusion: For the first time, the expression profiles of adiponectin and CDH13 are analyzed in many human tissues in correlation to each other and to clinical parameters.
First comparative characterization of three distinct ferritin subunits from a teleost: Evidence for immune-responsive mRNA expression and iron depriving activity of seahorse (Hippocampus abdominalis) ferritins.

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Oh, Minyoung; Umasuthan, Navaneethaiyer; Elvitigala, Don Anushka Sandaruwan; Wan, Qiang; Jo, Eunyoung; Ko, Jiyeon; Noh, Gyeong Eon; Shin, Sangok; Rho, Sum; Lee, Jehee

2016-02-01

Ferritins play an indispensable role in iron homeostasis through their iron-withholding function in living beings. In the current study, cDNA sequences of three distinct ferritin subunits, including a ferritin H, a ferritin M, and a ferritin L, were identified from big belly seahorse, Hippocampus abdominalis, and molecularly characterized. Complete coding sequences (CDS) of seahorse ferritin H (HaFerH), ferritin M (HaFerM), and ferritin L (HaFerL) subunits were comprised of 531, 528, and 522 base pairs (bp), respectively, which encode polypeptides of 177, 176, and 174 amino acids, respectively, with molecular masses of ∼20-21 kDa. Our in silico analyses demonstrate that these three ferritin subunits exhibit the typical characteristics of ferritin superfamily members including iron regulatory elements, domain signatures, and reactive centers. The coding sequences of HaFerH, M, and L were cloned and the corresponding proteins were overexpressed in a bacterial system. Recombinantly expressed HaFer proteins demonstrated detectable in vivo iron sequestrating (ferroxidase) activity, consistent with their putative iron binding capability. Quantification of the basal expression of these three HaFer sequences in selected tissues demonstrated a gene-specific ubiquitous spatial distribution pattern, with abundance of mRNA in HaFerM in the liver and predominant expression of HaFerH and HaFerL in blood. Interestingly, the basal expression of all three ferritin genes was found to be significantly modulated against pathogenic stress mounted by lipopolysaccharides (LPS), poly I:C, Streptococcus iniae, and Edwardsiella tarda. Collectively, our findings suggest that the three HaFer subunits may be involved in iron (II) homeostasis in big belly seahorse and that they are important in its host defense mechanisms. Copyright © 2016 Elsevier Ltd. All rights reserved.
Cre recombinase expression can result in phenotypic aberrations in plants

NARCIS (Netherlands)

Coppoolse, E.; Vroomen, de M.J.; Roelofs, D.; Smit, J.; Gennip, van F.; Hersmus, B.J.M.; Nijkamp, H.J.J.; Haaren, van M.J.

2003-01-01

The cre recombinase gene was stably introduced and expressed in tomato, petunia and Nicotiana tabacum. Some plants expressing the cre gene driven by a CaMV 35S promoter displayed growth retardation and a distinct pattern of chlorosis in their leaves. Although no direct relation can be proven between
Cre recombinase expression can result in phenotypic aberrations in plants

NARCIS (Netherlands)

Coppoolse, Eric R; de Vroomen, Marianne J; Roelofs, Dick; Smit, Jaap; van Gennip, Femke; Hersmus, Bart J M; Nijkamp, H John J; van Haaren, Mark J J

The cre recombinase gene was stably introduced and expressed in tomato, petunia and Nicotiana tabacum. Some plants expressing the cre gene driven by a CaMV 35S promoter displayed growth retardation and a distinct pattern of chlorosis in their leaves. Although no direct relation can be proven between
Developmental expression of membrane type 4-matrix metalloproteinase (Mt4-mmp/Mmp17) in the mouse embryo

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Clemente, Cristina; Montalvo, María Gregoria; Seiki, Motoharu; Arroyo, Alicia G.

2017-01-01

Matrix metalloproteinases (MMPs) constitute a large group of endoproteases that play important functions during embryonic development, tumor metastasis and angiogenesis by degrading components of the extracellular matrix. Within this family, we focused our study on Mt4-mmp (also called Mmp17) that belongs to a distinct subset that is anchored to the cell surface via a glycosylphosphatidylinositol (GPI) moiety and with the catalytic site exposed to the extracellular space. Information about its function and substrates is very limited to date, and little has been reported on its role in the developing embryo. Here, we report a detailed expression analysis of Mt4-mmp during mouse embryonic development by using a LacZ reporter transgenic mouse line. We showed that Mt4-mmp is detected from early stages of development to postnatal stages following a dynamic and restricted pattern of expression. Mt4-mmp was first detected at E8.5 limited to the intersomitic vascularization, the endocardial endothelium and the dorsal aorta. Mt4-mmpLacZ/+ cells were also observed in the neural crest cells, somites, floor plate and notochord at early stages. From E10.5, expression localized in the limb buds and persists during limb development. A strong expression in the brain begins at E12.5 and continues to postnatal stages. Specifically, staining was observed in the olfactory bulb, cerebral cortex, hippocampus, striatum, septum, dorsal thalamus and the spinal cord. In addition, LacZ-positive cells were also detected during eye development, initially at the hyaloid artery and later on located in the lens and the neural retina. Mt4-mmp expression was confirmed by quantitative RT-PCR and western blot analysis in some embryonic tissues. Our data point to distinct functions for this metalloproteinase during embryonic development, particularly during brain formation, angiogenesis and limb development. PMID:28926609
Distinct gene expression profiles in ovarian cancer linked to Lynch syndrome

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Jönsson, Jenny-Maria; Bartuma, Katarina; Dominguez-Valentin, Mev

2014-01-01

Ovarian cancer linked to Lynch syndrome represents a rare subset that typically presents at young age as early-stage tumors with an overrepresentation of endometrioid and clear cell histologies. We investigated the molecular profiles of Lynch syndrome-associated and sporadic ovarian cancer...... with the aim to identify key discriminators and central tumorigenic mechanisms in hereditary ovarian cancer. Global gene expression profiling using whole-genome c-DNA-mediated Annealing, Selection, extension, and Ligation was applied to 48 histopathologically matched Lynch syndrome-associated and sporadic...... ovarian cancers. Lynch syndrome-associated and sporadic ovarian cancers differed by 349 significantly deregulated genes, including PTPRH, BIRC3, SHH and TNFRSF6B. The genes involved were predominantly linked to cell growth, proliferation, and cell-to-cell signaling and interaction. When stratified...
Parvalbumin+ Neurons and Npas1+ Neurons Are Distinct Neuron Classes in the Mouse External Globus Pallidus.

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Hernández, Vivian M; Hegeman, Daniel J; Cui, Qiaoling; Kelver, Daniel A; Fiske, Michael P; Glajch, Kelly E; Pitt, Jason E; Huang, Tina Y; Justice, Nicholas J; Chan, C Savio

2015-08-26

Compelling evidence suggests that pathological activity of the external globus pallidus (GPe), a nucleus in the basal ganglia, contributes to the motor symptoms of a variety of movement disorders such as Parkinson's disease. Recent studies have challenged the idea that the GPe comprises a single, homogenous population of neurons that serves as a simple relay in the indirect pathway. However, we still lack a full understanding of the diversity of the neurons that make up the GPe. Specifically, a more precise classification scheme is needed to better describe the fundamental biology and function of different GPe neuron classes. To this end, we generated a novel multicistronic BAC (bacterial artificial chromosome) transgenic mouse line under the regulatory elements of the Npas1 gene. Using a combinatorial transgenic and immunohistochemical approach, we discovered that parvalbumin-expressing neurons and Npas1-expressing neurons in the GPe represent two nonoverlapping cell classes, amounting to 55% and 27% of the total GPe neuron population, respectively. These two genetically identified cell classes projected primarily to the subthalamic nucleus and to the striatum, respectively. Additionally, parvalbumin-expressing neurons and Npas1-expressing neurons were distinct in their autonomous and driven firing characteristics, their expression of intrinsic ion conductances, and their responsiveness to chronic 6-hydroxydopamine lesion. In summary, our data argue that parvalbumin-expressing neurons and Npas1-expressing neurons are two distinct functional classes of GPe neurons. This work revises our understanding of the GPe, and provides the foundation for future studies of its function and dysfunction. Until recently, the heterogeneity of the constituent neurons within the external globus pallidus (GPe) was not fully appreciated. We addressed this knowledge gap by discovering two principal GPe neuron classes, which were identified by their nonoverlapping expression of the
Parvalbumin+ Neurons and Npas1+ Neurons Are Distinct Neuron Classes in the Mouse External Globus Pallidus

Science.gov (United States)

Hernández, Vivian M.; Hegeman, Daniel J.; Cui, Qiaoling; Kelver, Daniel A.; Fiske, Michael P.; Glajch, Kelly E.; Pitt, Jason E.; Huang, Tina Y.; Justice, Nicholas J.

2015-01-01

Compelling evidence suggests that pathological activity of the external globus pallidus (GPe), a nucleus in the basal ganglia, contributes to the motor symptoms of a variety of movement disorders such as Parkinson's disease. Recent studies have challenged the idea that the GPe comprises a single, homogenous population of neurons that serves as a simple relay in the indirect pathway. However, we still lack a full understanding of the diversity of the neurons that make up the GPe. Specifically, a more precise classification scheme is needed to better describe the fundamental biology and function of different GPe neuron classes. To this end, we generated a novel multicistronic BAC (bacterial artificial chromosome) transgenic mouse line under the regulatory elements of the Npas1 gene. Using a combinatorial transgenic and immunohistochemical approach, we discovered that parvalbumin-expressing neurons and Npas1-expressing neurons in the GPe represent two nonoverlapping cell classes, amounting to 55% and 27% of the total GPe neuron population, respectively. These two genetically identified cell classes projected primarily to the subthalamic nucleus and to the striatum, respectively. Additionally, parvalbumin-expressing neurons and Npas1-expressing neurons were distinct in their autonomous and driven firing characteristics, their expression of intrinsic ion conductances, and their responsiveness to chronic 6-hydroxydopamine lesion. In summary, our data argue that parvalbumin-expressing neurons and Npas1-expressing neurons are two distinct functional classes of GPe neurons. This work revises our understanding of the GPe, and provides the foundation for future studies of its function and dysfunction. SIGNIFICANCE STATEMENT Until recently, the heterogeneity of the constituent neurons within the external globus pallidus (GPe) was not fully appreciated. We addressed this knowledge gap by discovering two principal GPe neuron classes, which were identified by their nonoverlapping
Two distinct E3 ubiquitin ligases have complementary functions in the regulation of delta and serrate signaling in Drosophila.

Directory of Open Access Journals (Sweden)

Roland Le Borgne

2005-04-01

Full Text Available Signaling by the Notch ligands Delta (Dl and Serrate (Ser regulates a wide variety of essential cell-fate decisions during animal development. Two distinct E3 ubiquitin ligases, Neuralized (Neur and Mind bomb (Mib, have been shown to regulate Dl signaling in Drosophila melanogaster and Danio rerio, respectively. While the neur and mib genes are evolutionarily conserved, their respective roles in the context of a single organism have not yet been examined. We show here that the Drosophila mind bomb (D-mib gene regulates a subset of Notch signaling events, including wing margin specification, leg segmentation, and vein determination, that are distinct from those events requiring neur activity. D-mib also modulates lateral inhibition, a neur- and Dl-dependent signaling event, suggesting that D-mib regulates Dl signaling. During wing development, expression of D-mib in dorsal cells appears to be necessary and sufficient for wing margin specification, indicating that D-mib also regulates Ser signaling. Moreover, the activity of the D-mib gene is required for the endocytosis of Ser in wing imaginal disc cells. Finally, ectopic expression of neur in D-mib mutant larvae rescues the wing D-mib phenotype, indicating that Neur can compensate for the lack of D-mib activity. We conclude that D-mib and Neur are two structurally distinct proteins that have similar molecular activities but distinct developmental functions in Drosophila.
Expression of Cellular Isoform of Prion Protein on the Surface of Peripheral Blood Lymphocytes Among Women Exposed to Low Doses of Ionizing Radiation

International Nuclear Information System (INIS)

Klucinski, P.; Martirosian, G.; Mazur, B.; Kaufman, J.; Hrycek, A.; Masluch, E.; Cieslik, P.

2007-01-01

Ionizing radiation affect the expression of adhesive and co-stimulation molecules in lymphocytes. The objective of this study was to determinate the effect of low doses of ionizing radiation on the expression of prion protein PrPc on the surface peripheral blood lymphocytes in the women operating X-ray equipment. In female workers and persons of the control group the PrPc expression on CD3 (T-lymphocytes), Cd4 (T-helper), CD8 (T-cytotoxic) and CD19 (B- lymphocytes), were tested. We conclude that in women operating X-ray equipment the relationship between low doses of ionizing radiation and expression of PrPc on lymphocytes does exist concerning CD3, CD4 and CD lymphocytes. (author)
Different culture media affect growth characteristics, surface marker distribution and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells.

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Hagmann, Sebastien; Moradi, Babak; Frank, Sebastian; Dreher, Thomas; Kämmerer, Peer Wolfgang; Richter, Wiltrud; Gotterbarm, Tobias

2013-07-30

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) play an important role in modern tissue engineering, while distinct variations of culture media compositions and supplements have been reported. Because MSCs are heterogeneous regarding their regenerative potential and their surface markers, these parameters were compared in four widely used culture media compositions. MSCs were isolated from bone marrow and expanded in four established cell culture media. MSC yield/1000 MNCs, passage time and growth index were observed. In P4, typical MSC surface markers were analysed by fluorescence cytometry. Additionally, chondrogenic, adipogenic and osteogenic differentiation potential were evaluated. Growth index and P0 cell yield varied importantly between the media. The different expansion media had a significant influence on the expression of CD10, CD90, CD105, CD140b CD146 and STRO-1. While no significant differences were observed regarding osteogenic and adipogenic differentiation, chondrogenic differentiation was superior in medium A as reflected by GAG/DNA content. The choice of expansion medium can have a significant influence on growth, differentiation potential and surface marker expression of mesenchymal stromal cells, which is of fundamental importance for tissue engineering procedures.
Genomic profiling of rice sperm cell transcripts reveals conserved and distinct elements in the flowering plant male germ lineage.

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Russell, Scott D; Gou, Xiaoping; Wong, Chui E; Wang, Xinkun; Yuan, Tong; Wei, Xiaoping; Bhalla, Prem L; Singh, Mohan B

2012-08-01

Genomic assay of sperm cell RNA provides insight into functional control, modes of regulation, and contributions of male gametes to double fertilization. Sperm cells of rice (Oryza sativa) were isolated from field-grown, disease-free plants and RNA was processed for use with the full-genome Affymetrix microarray. Comparison with Gene Expression Omnibus (GEO) reference arrays confirmed expressionally distinct gene profiles. A total of 10,732 distinct gene sequences were detected in sperm cells, of which 1668 were not expressed in pollen or seedlings. Pathways enriched in male germ cells included ubiquitin-mediated pathways, pathways involved in chromatin modeling including histones, histone modification and nonhistone epigenetic modification, and pathways related to RNAi and gene silencing. Genome-wide expression patterns in angiosperm sperm cells indicate common and divergent themes in the male germline that appear to be largely self-regulating through highly up-regulated chromatin modification pathways. A core of highly conserved genes appear common to all sperm cells, but evidence is still emerging that another class of genes have diverged in expression between monocots and dicots since their divergence. Sperm cell transcripts present at fusion may be transmitted through plasmogamy during double fertilization to effect immediate post-fertilization expression of early embryo and (or) endosperm development. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

Role of protein kinase A and class II phosphatidylinositol 3-kinase C2β in the downregulation of KCa3.1 channel synthesis and membrane surface expression by lyso-globotriaosylceramide

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Choi, Ju Yeon; Park, Seonghee, E-mail: sp@ewha.ac.kr

2016-02-19

The intermediate conductance calcium-activated potassium channel (KCa3.1) mediates proliferation of many cell types including fibroblasts, and is a molecular target for intervention in various cell proliferative diseases. Our previous study showed that reduction of KCa3.1 channel expression by lyso-globotriaosylceramide (lyso-Gb3) inhibits differentiation into myofibroblasts and collagen synthesis, which might lead to development of ascending thoracic aortic aneurysm secondary to Fabry disease. However, how lyso-Gb3 downregulates KCa3.1 channel expression is unknown. Therefore, we aimed to investigate the underlying mechanisms of lyso-Gb3-mediated KCa3.1 channel downregulation, focusing on the cAMP signaling pathway. We found that lyso-Gb3 increased the intracellular cAMP concentration by upregulation of adenylyl cyclase 6 and inhibited ERK 1/2 phosphorylation through the protein kinase A (PKA) pathway, leading to the inhibition of KCa3.1 channel synthesis, not the exchange protein directly activated by cAMP (Epac) pathway. Moreover, lyso-Gb3 suppressed expression of class II phosphatidylinositol 3-kinase C2β (PI3KC2β) by PKA activation, which reduces the production of phosphatidylinositol 3-phosphate [PI(3)P], and the reduced membrane surface expression of KCa3.1 channel was recovered by increasing the intracellular levels of PI(3)P. Consequently, our findings that lyso-Gb3 inhibited both KCa3.1 channel synthesis and surface expression by increasing intracellular cAMP, and controlled surface expression through changes in PI3KC2β-mediated PI(3)P production, suggest that modulation of PKA and PI3KC2β activity to control of KCa3.1 channel expression can be an alternative important target to attenuate ascending thoracic aortic aneurysms in Fabry disease. - Highlights: • Lyso-Gb3 causes elevation of intracellular cAMP. • Lyso-Gb3 inhibits the ERK 1/2 phosphorylation through PKA, thereby reducing KCa3.1 channel synthesis. • Lyso-Gb3 reduces PI3KC2�
β-Secretase BACE1 Promotes Surface Expression and Function of Kv3.4 at Hippocampal Mossy Fiber Synapses.

Science.gov (United States)

Hartmann, Stephanie; Zheng, Fang; Kyncl, Michele C; Karch, Sandra; Voelkl, Kerstin; Zott, Benedikt; D'Avanzo, Carla; Lomoio, Selene; Tesco, Giuseppina; Kim, Doo Y; Alzheimer, Christian; Huth, Tobias

2018-04-04

The β-secretase β-site APP-cleaving enzyme 1 (BACE1) is deemed a major culprit in Alzheimer's disease, but accumulating evidence indicates that there is more to the enzyme than driving the amyloidogenic processing of the amyloid precursor protein. For example, BACE1 has emerged as an important regulator of neuronal activity through proteolytic and, most unexpectedly, also through nonproteolytic interactions with several ion channels. Here, we identify and characterize the voltage-gated K + channel 3.4 (Kv3.4) as a new and functionally relevant interaction partner of BACE1. Kv3.4 gives rise to A-type current with fast activating and inactivating kinetics and serves to repolarize the presynaptic action potential. We found that BACE1 and Kv3.4 are highly enriched and remarkably colocalized in hippocampal mossy fibers (MFs). In BACE1 -/- mice of either sex, Kv3.4 surface expression was significantly reduced in the hippocampus and, in synaptic fractions thereof, Kv3.4 was specifically diminished, whereas protein levels of other presynaptic K + channels such as K Ca 1.1 and K Ca 2.3 remained unchanged. The apparent loss of presynaptic Kv3.4 affected the strength of excitatory transmission at the MF-CA3 synapse in hippocampal slices of BACE1 -/- mice when probed with the Kv3 channel blocker BDS-I. The effect of BACE1 on Kv3.4 expression and function should be bidirectional, as predicted from a heterologous expression system, in which BACE1 cotransfection produced a concomitant upregulation of Kv3.4 surface level and current based on a physical interaction between the two proteins. Our data show that, by targeting Kv3.4 to presynaptic sites, BACE1 endows the terminal with a powerful means to regulate the strength of transmitter release. SIGNIFICANCE STATEMENT The β-secretase β-site APP-cleaving enzyme 1 (BACE1) is infamous for its crucial role in the pathogenesis of Alzheimer's disease, but its physiological functions in the intact nervous system are only gradually
Chemistry of Frozen NaCl and MgSO4 Brines - Implications for Surface Expression of Europa's Ocean Composition

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Johnson, P. V.; Hodyss, R. P.; Choukroun, M.; Vu, T. H.

2015-12-01

The composition of Europa's subsurface ocean is a critical determinant of its habitability, but current analysis of the ocean composition is limited to its expression on the Europan surface. While there is observational evidence indicating that ocean materials make their way to the surface, our understanding of the chemical processes that can alter this material under Europan surface conditions is limited. We present experimental data on the chemistry of mixed solutions of NaCl and MgSO4 as they are frozen to 100 K, replicating the conditions that may occur when subsurface ocean fluids are emplaced onto Europa's surface. Confocal micro-Raman spectroscopy is used to study the formation of salts during the freezing process, and the interaction of ions in the frozen brines. Our data indicate that mixed aqueous solutions of NaCl and MgSO4 form Na2SO4 and MgCl2 preferentially when frozen, rather than making NaCl and MgSO4 precipitates. The detection of epsomite (MgSO4•7H2O) on Europa's surface may therefore imply an ocean composition relatively low in sodium, unless radiolytic chemistry converts MgCl2 to MgSO4 as suggested by Hand and Brown 2013 (ApJ 145 110). These results have important implications for the interpretation of remote sensing data of Europa's surface.
The structure of a conserved piezo channel domain reveals a topologically distinct β sandwich fold.

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Kamajaya, Aron; Kaiser, Jens T; Lee, Jonas; Reid, Michelle; Rees, Douglas C

2014-10-07

Piezo has recently been identified as a family of eukaryotic mechanosensitive channels composed of subunits containing over 2,000 amino acids, without recognizable sequence similarity to other channels. Here, we present the crystal structure of a large, conserved extramembrane domain located just before the last predicted transmembrane helix of C. elegans PIEZO, which adopts a topologically distinct β sandwich fold. The structure was also determined of a point mutation located on a conserved surface at the position equivalent to the human PIEZO1 mutation found in dehydrated hereditary stomatocytosis patients (M2225R). While the point mutation does not change the overall domain structure, it does alter the surface electrostatic potential that may perturb interactions with a yet-to-be-identified ligand or protein. The lack of structural similarity between this domain and any previously characterized fold, including those of eukaryotic and bacterial channels, highlights the distinctive nature of the Piezo family of eukaryotic mechanosensitive channels. Copyright © 2014 Elsevier Ltd. All rights reserved.
Effects of DDT and Triclosan on Tumor-cell Binding Capacity and Cell-Surface Protein Expression of Human Natural Killer Cells

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Hurd-Brown, Tasia; Udoji, Felicia; Martin, Tamara; Whalen, Margaret M.

2012-01-01

1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT) and triclosan (TCS) are organochlorine (OC) compounds that contaminate the environment, are found in human blood, and have been shown to decrease the tumor-cell killing (lytic) function of human natural killer (NK) cells. NK cells defend against tumor cells and virally infected cells. They bind to these targets, utilizing a variety of cell surface proteins. This study examined concentrations of DDT and TCS that decrease lytic function for alteration of NK binding to tumor targets. Levels of either compound that caused loss of binding function were then examined for effects on expression of cell-surface proteins needed for binding. NK cells exposed to 2.5 μM DDT for 24 h (which caused a greater than 55% loss of lytic function) showed a decrease in NK binding function of about 22%, and a decrease in CD16 cell-surface protein of 20%. NK cells exposed to 5 μM TCS for 24 h showed a decrease in ability to bind tumor cells of 37% and a decrease in expression of CD56 of about 34%. This same treatment caused a decrease in lytic function of greater than 87%. These results indicated that only a portion of the loss of NK lytic function seen with exposures to these compounds could be accounted for by loss of binding function. They also showed that loss of binding function is accompanied by a loss cell-surface proteins important in binding function. PMID:22729613
Distinct roles of synaptic and extrasynaptic GABAA receptors in striatal inhibition dynamics

Directory of Open Access Journals (Sweden)

Ruixi eLuo

2013-11-01

Full Text Available Striatonigral and striatopallidal projecting medium spiny neurons (MSNs express dopamine D1 (D1+ and D2 receptors (D2+, respectively. Both classes receive extensive GABAergic input via expression of synaptic, perisynaptic and extrasynaptic GABAA receptors. The activation patterns of different presynaptic GABAergic neurons produce transient and sustained GABAA receptor-mediated conductance that fulfill distinct physiological roles. We performed single and dual whole cell recordings from striatal neurons in mice expressing fluorescent proteins in interneurons and MSNs. We report specific inhibitory dynamics produced by distinct activation patterns of presynaptic GABAergic neurons as source of synaptic, perisynaptic and extrasynaptic inhibition. Synaptic GABAA receptors in MSNs contain the α2, γ2 and a β subunit. In addition, there is evidence for the developmental increase of the α1 subunit that contributes to faster inhibitory postsynaptic current (IPSC. Tonic GABAergic currents in MSNs from adult mice are carried by extrasynaptic receptors containing the α4 and δ subunit, while in younger mice this current is mediated by receptors that contain the α5 subunit. Both forms of tonic currents are differentially expressed in D1+ and D2+ MSNs. This study extends these findings by relating presynaptic activation with pharmacological analysis of inhibitory conductance in mice where the β3 subunit is conditionally removed in fluorescently labeled D2+ MSNs and in mice with global deletion of the δ subunit. Our results show that responses to low doses of gaboxadol (2μM, a GABAA receptor agonist with preference to δ subunit, are abolished in the δ but not the β3 subunit knock out mice. This suggests that the β3 subunit is not a component of the adult extrasynaptic receptor pool, in contrast to what has been shown for tonic current in young mice. Deletion of the β3 subunit from D2+ MSNs however, removed slow spontaneous IPSCs, implicating its
Regulatory CD4 T cells inhibit HIV-1 expression of other CD4 T cell subsets via interactions with cell surface regulatory proteins.

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Zhang, Mingce; Robinson, Tanya O; Duverger, Alexandra; Kutsch, Olaf; Heath, Sonya L; Cron, Randy Q

2018-03-01

During chronic HIV-1 infection, regulatory CD4 T cells (Tregs) frequently represent the largest subpopulation of CD4 T cell subsets, implying relative resistant to HIV-1. When HIV-1 infection of CD4 T cells was explored in vitro and ex vivo from patient samples, Tregs possessed lower levels of HIV-1 DNA and RNA in comparison with conventional effector and memory CD4 T cells. Moreover, Tregs suppressed HIV-1 expression in other CD4 T cells in an in vitro co-culture system. This suppression was mediated in part via multiple inhibitory surface proteins expressed on Tregs. Antibody blockade of CTLA-4, PD-1, and GARP on Tregs resulted in increased HIV-1 DNA integration and mRNA expression in neighboring CD4 T cells. Moreover, antibody blockade of Tregs inhibitory proteins resulted in increased HIV-1 LTR transcription in co-cultured CD4 T cells. Thus, Tregs inhibit HIV-1 infection of other CD4 T cell subsets via interactions with inhibitory cell surface proteins. Copyright © 2018 Elsevier Inc. All rights reserved.
Distinct cis regulatory elements govern the expression of TAG1 in embryonic sensory ganglia and spinal cord.

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Yoav Hadas

Full Text Available Cell fate commitment of spinal progenitor neurons is initiated by long-range, midline-derived, morphogens that regulate an array of transcription factors that, in turn, act sequentially or in parallel to control neuronal differentiation. Included among these are transcription factors that regulate the expression of receptors for guidance cues, thereby determining axonal trajectories. The Ig/FNIII superfamily molecules TAG1/Axonin1/CNTN2 (TAG1 and Neurofascin (Nfasc are co-expressed in numerous neuronal cell types in the CNS and PNS - for example motor, DRG and interneurons - both promote neurite outgrowth and both are required for the architecture and function of nodes of Ranvier. The genes encoding TAG1 and Nfasc are adjacent in the genome, an arrangement which is evolutionarily conserved. To study the transcriptional network that governs TAG1 and Nfasc expression in spinal motor and commissural neurons, we set out to identify cis elements that regulate their expression. Two evolutionarily conserved DNA modules, one located between the Nfasc and TAG1 genes and the second directly 5' to the first exon and encompassing the first intron of TAG1, were identified that direct complementary expression to the CNS and PNS, respectively, of the embryonic hindbrain and spinal cord. Sequential deletions and point mutations of the CNS enhancer element revealed a 130bp element containing three conserved E-boxes required for motor neuron expression. In combination, these two elements appear to recapitulate a major part of the pattern of TAG1 expression in the embryonic nervous system.
Differential surface phenotype and context-dependent reactivity of functionally diverse NKT cells.

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Cameron, Garth; Godfrey, Dale I

2018-03-05

Natural Killer T (NKT) cells are a functionally diverse population that recognizes lipid-based antigens in association with the antigen-presenting molecule CD1d. Here, we define a technique to separate the functionally distinct thymic NKT1, NKT2 and NKT17 cell subsets by their surface expression of CD278 (ICOS) and the activation-associated glycoform of CD43, enabling the investigation of subset-specific effector-functions. We report that all three subsets express the transcription factor GATA-3 and the potential to produce IL-4 and IL-10 following activation. This questions the notion that NKT2 cells are the predominant source of IL-4 within the NKT cell pool, and suggests that IL-10-production may be more indicative of NKT cell plasticity than the existence of a distinct regulatory lineage or subset. We also show that many NKT17 cells are CD4 + and are biased toward Vβ8.3 TCR gene usage. Lastly, we demonstrate that the toll-like receptor (TLR) ligand lipopolysaccharide (LPS) can induce a NKT17 cell-biased response, even in the absence of exogenous antigen, and that combining LPS with α-GalCer resulted in enhanced IL-17A-production, and reduced levels of the immunosuppressive cytokine IL-10. This study provides a novel means to examine the context-dependent reactivity of the functionally heterogeneous NKT cell population and provides important new insight into the functional biology of these subsets. © 2018 Australasian Society for Immunology Inc.
MicroRNAs define distinct human neuroblastoma cell phenotypes and regulate their differentiation and tumorigenicity

International Nuclear Information System (INIS)

Samaraweera, Leleesha; Grandinetti, Kathryn B; Huang, Ruojun; Spengler, Barbara A; Ross, Robert A

2014-01-01

Neuroblastoma (NB) is the most common extracranial solid tumor in children. NB tumors and derived cell lines are phenotypically heterogeneous. Cell lines are classified by phenotype, each having distinct differentiation and tumorigenic properties. The neuroblastic phenotype is tumorigenic, has neuronal features and includes stem cells (I-cells) and neuronal cells (N-cells). The non-neuronal phenotype (S-cell) comprises cells that are non-tumorigenic with features of glial/smooth muscle precursor cells. This study identified miRNAs associated with each distinct cell phenotypes and investigated their role in regulating associated differentiation and tumorigenic properties. A miRNA microarray was performed on the three cell phenotypes and expression verified by qRT-PCR. miRNAs specific for certain cell phenotypes were modulated using miRNA inhibitors or stable transfection. Neuronal differentiation was induced by RA; non-neuronal differentiation by BrdU. Changes in tumorigenicity were assayed by soft agar colony forming ability. N-myc binding to miR-375 promoter was assayed by chromatin-immunoprecipitation. Unsupervised hierarchical clustering of miRNA microarray data segregated neuroblastic and non-neuronal cell lines and showed that specific miRNAs define each phenotype. qRT-PCR validation confirmed that increased levels of miR-21, miR-221 and miR-335 are associated with the non-neuronal phenotype, whereas increased levels of miR-124 and miR-375 are exclusive to neuroblastic cells. Downregulation of miR-335 in non-neuronal cells modulates expression levels of HAND1 and JAG1, known modulators of neuronal differentiation. Overexpression of miR-124 in stem cells induces terminal neuronal differentiation with reduced malignancy. Expression of miR-375 is exclusive for N-myc-expressing neuroblastic cells and is regulated by N-myc. Moreover, miR-375 downregulates expression of the neuronal-specific RNA binding protein HuD. Thus, miRNAs define distinct NB cell phenotypes
Distinction of gastric cancer tissue based on surface-enhanced Raman spectroscopy

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Ma, Jun; Zhou, Hanjing; Gong, Longjing; Liu, Shu; Zhou, Zhenghua; Mao, Weizheng; Zheng, Rong-er

2012-12-01

Gastric cancer is one of the most common malignant tumors with high recurrence rate and mortality rate in China. This study aimed to evaluate the diagnostic capability of Surface-enhanced Raman spectroscopy (SERS) based on gold colloids for distinguishing gastric tissues. Gold colloids were directly mixed with the supernatant of homogenized tissues to heighten the Raman signal of various biomolecule. A total of 56 samples were collected from normal (30) and cancer (26). Raman spectra were obtained with a 785nm excitation in the range of 600-1800 cm-1. Significant spectral differences in SERS mainly belong to nucleic acid, proteins and lipids, particularly in the range of 653, 726, 828, 963, 1004, 1032, 1088, 1130, 1243, 1369, 1474, 1596, 1723 cm-1. PCA-LDA algorithms with leave-one-patient-out cross validation yielded diagnostic sensitivities of 90% (27/30), specificities of 88.5% (23/26), and accuracy of 89.3% (50/56), for classification of normal and cancer tissues. The receiver operating characteristic (ROC) surface is 0.917, illustrating the diagnostic utility of SERS together with PCA-LDA to identify gastric cancer from normal tissue. This work demonstrated the SERS techniques can be useful for gastric cancer detection, and it is also a potential technique for accurately identifying cancerous tumor, which is of considerable clinical importance to real-time diagnosis.
Gene expression profile data for mouse facial development

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Sonia M. Leach

2017-08-01

Full Text Available This article contains data related to the research articles "Spatial and Temporal Analysis of Gene Expression during Growth and Fusion of the Mouse Facial Prominences" (Feng et al., 2009 [1] and “Systems Biology of facial development: contributions of ectoderm and mesenchyme” (Hooper et al., 2017 In press [2]. Embryonic mammalian craniofacial development is a complex process involving the growth, morphogenesis, and fusion of distinct facial prominences into a functional whole. Aberrant gene regulation during this process can lead to severe craniofacial birth defects, including orofacial clefting. As a means to understand the genes involved in facial development, we had previously dissected the embryonic mouse face into distinct prominences: the mandibular, maxillary or nasal between E10.5 and E12.5. The prominences were then processed intact, or separated into ectoderm and mesenchyme layers, prior analysis of RNA expression using microarrays (Feng et al., 2009, Hooper et al., 2017 in press [1,2]. Here, individual gene expression profiles have been built from these datasets that illustrate the timing of gene expression in whole prominences or in the separated tissue layers. The data profiles are presented as an indexed and clickable list of the genes each linked to a graphical image of that gene׳s expression profile in the ectoderm, mesenchyme, or intact prominence. These data files will enable investigators to obtain a rapid assessment of the relative expression level of any gene on the array with respect to time, tissue, prominence, and expression trajectory.
Enteroendocrine cells are specifically marked by cell surface expression of claudin-4 in mouse small intestine.

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Takahiro Nagatake

Full Text Available Enteroendocrine cells are solitary epithelial cells scattered throughout the gastrointestinal tract and produce various types of hormones, constituting one of the largest endocrine systems in the body. The study of these rare epithelial cells has been hampered by the difficulty in isolating them because of the lack of specific cell surface markers. Here, we report that enteroendocrine cells selectively express a tight junction membrane protein, claudin-4 (Cld4, and are efficiently isolated with the use of an antibody specific for the Cld4 extracellular domain and flow cytometry. Sorted Cld4+ epithelial cells in the small intestine exclusively expressed a chromogranin A gene (Chga and other enteroendocrine cell-related genes (Ffar1, Ffar4, Gpr119, and the population was divided into two subpopulations based on the activity of binding to Ulex europaeus agglutinin-1 (UEA-1. A Cld4+UEA-1- cell population almost exclusively expressed glucose-dependent insulinotropic polypeptide gene (Gip, thus representing K cells, whereas a Cld4+UEA-1+ cell population expressed other gut hormone genes, including glucagon-like peptide 1 (Gcg, pancreatic polypeptide-like peptide with N-terminal tyrosine amide (Pyy, cholecystokinin (Cck, secretin (Sct, and tryptophan hydroxylase 1 (Tph1. In addition, we found that orally administered luminal antigens were taken up by the solitary Cld4+ cells in the small intestinal villi, raising the possibility that enteroendocrine cells might also play a role in initiation of mucosal immunity. Our results provide a useful tool for the cellular and functional characterization of enteroendocrine cells.
Expression of Pluripotency and Oocyte-Related Genes in Single Putative Stem Cells from Human Adult Ovarian Surface Epithelium Cultured In Vitro in the Presence of Follicular Fluid

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Irma Virant-Klun

2013-01-01

Full Text Available The aim of this study was to trigger the expression of genes related to oocytes in putative ovarian stem cells scraped from the ovarian surface epithelium of women with premature ovarian failure and cultured in vitro in the presence of follicular fluid, rich in substances for oocyte growth and maturation. Ovarian surface epithelium was scraped and cell cultures were set up by scrapings in five women with nonfunctional ovaries and with no naturally present mature follicles or oocytes. In the presence of donated follicular fluid putative stem cells grew and developed into primitive oocyte-like cells. A detailed single-cell gene expression profiling was performed to elucidate their genetic status in comparison to human embryonic stem cells, oocytes, and somatic fibroblasts. The ovarian cell cultures depleted/converted reproductive hormones from the culture medium. Estradiol alone or together with other substances may be involved in development of these primitive oocyte-like cells. The majority of primitive oocyte-like cells was mononuclear and expressed several genes related to pluripotency and oocytes, including genes related to meiosis, although they did not express some important oocyte-specific genes. Our work reveals the presence of putative stem cells in the ovarian surface epithelium of women with premature ovarian failure.
Distinct cell-specific expression of homospermidine synthase involved in pyrrolizidine alkaloid biosynthesis in three species of the boraginales.

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Niemüller, Daniel; Reimann, Andreas; Ober, Dietrich

2012-07-01

Homospermidine synthase (HSS) is the first specific enzyme in pyrrolizidine alkaloid (PA) biosynthesis, a pathway involved in the plant's chemical defense. HSS has been shown to be recruited repeatedly by duplication of a gene involved in primary metabolism. Within the lineage of the Boraginales, only one gene duplication event gave rise to HSS. Here, we demonstrate that the tissue-specific expression of HSS in three boraginaceous species, Heliotropium indicum, Symphytum officinale, and Cynoglossum officinale, is unique with respect to plant organ, tissue, and cell type. Within H. indicum, HSS is expressed exclusively in nonspecialized cells of the lower epidermis of young leaves and shoots. In S. officinale, HSS expression has been detected in the cells of the root endodermis and in leaves directly underneath developing inflorescences. In young roots of C. officinale, HSS is detected only in cells of the endodermis, but in a later developmental stage, additionally in the pericycle. The individual expression patterns are compared with those within the Senecioneae lineage (Asteraceae), where HSS expression is reproducibly found in specific cells of the endodermis and the adjacent cortex parenchyma of the roots. The individual expression patterns within the Boraginales species are discussed as being a requirement for the successful recruitment of HSS after gene duplication. The diversity of HSS expression within this lineage adds a further facet to the already diverse patterns of expression that have been observed for HSS in other PA-producing plant lineages, making this PA-specific enzyme one of the most diverse expressed proteins described in the literature.
Bodily Movement and Facial Actions in Expressive Musical Performance by Solo and Duo Instrumentalists: Two Distinctive Case Studies

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Davidson, Jane W.

2012-01-01

The research literature concerning gesture in musical performance increasingly reports that musically communicative and meaningful performances contain highly expressive bodily movements. These movements are involved in the generation of the musically expressive performance, but enquiry into the development of expressive bodily movement has been…
Fowler's approximation for the surface tension and surface energy of Lennard-Jones fluids revisited

Energy Technology Data Exchange (ETDEWEB)

Mulero, A [Departamento de Fisica, Universidad de Extremadura, 06071-Badajoz (Spain); Galan, C [Departamento de Fisica, Universidad de Extremadura, 06071-Badajoz (Spain); Cuadros, F [Departamento de Fisica, Universidad de Extremadura, 06071-Badajoz (Spain)

2003-04-16

We present a detailed study of the validity of Fowler's approximation for calculating the surface tension and the surface energy of Lennard-Jones fluids. To do so, we consider three different explicit analytical expressions for the radial distribution function (RDF), including one proposed by our research group, together with very accurate expressions for the liquid and vapour densities, also proposed by our group. The calculation of the surface tension from the direct correlation function using both the Percus-Yevick and the hypernetted-chain approximations is also considered. Finally, our results are compared with those obtained by other authors by computer simulations or through relevant theoretical approximations. In particular, we consider the analytical expression proposed by Kalikmanov and Hofmans (1994 J. Phys.: Condens. Matter 6 2207-14) for the surface tension. Our results indicate that the values for the surface energy in Fowler's approximation obtained by other authors are adequate, and can be calculated from the RDF models. For the surface tension, however, the values considered as valid in previous works seem to be incorrect. The correct values can be obtained from our model for the RDF or from the Kalikmanov and Hofmans expression with suitable inputs.
Expression of Lipid Peroxidation Markers in the Tear Film and Ocular Surface of Patients with Non-Sjogren Syndrome: Potential Biomarkers for Dry Eye Disease.

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Choi, Won; Lian, Cui; Ying, Li; Kim, Ga Eon; You, In Cheon; Park, Soo Hyun; Yoon, Kyung Chul

2016-09-01

To investigate the expression of lipid peroxidation markers in the tear film and ocular surface and their correlation with disease severity in patients with dry eye disease. The concentrations of hexanoyl-lysine (HEL), 4-hydroxy-2-nonenal (HNE), and malondialdehyde (MDA) were measured with enzyme-linked immunosorbent assays in tears obtained from 44 patients with non-Sjogren syndrome dry eye and 33 control subjects. The correlations between the marker levels and the tear film and ocular surface parameters, including tear film break-up time (BUT), Schirmer tear value, tear clearance rate, keratoepitheliopathy scores, corneal sensitivity, conjunctival goblet cell density, and symptom score, were analyzed. The expression of the lipid peroxidation markers HEL, 4-HNE, and MDA in the conjunctiva was evaluated using immunohistochemistry. The concentrations of HEL, 4-HNE, and MDA were 279.84 ± 69.98 nmol/L, 0.02 ± 0.01 μg/mL, and 3.80 ± 1.05 pmol/mg in control subjects and 283.21 ± 89.67 nmol/L (p = 0.97), 0.20 ± 0.03 μg/mL (p dry eye patients. 4-HNE and MDA levels significantly correlated with BUT, Schirmer tear value, tear clearance rate, keratoepitheliopathy scores, conjunctival goblet cell density, and symptom score (p dry eye patients. The expression of late lipid peroxidation markers, 4-HNE and MDA, increases in the tear film and ocular surface of patients with dry eye. The levels correlate with various tear film and ocular surface parameters and may reflect the severity of dry eye disease.
Characterization, cell-surface expression and ligand-binding properties of different truncated N-terminal extracellular domains of the ionotropic glutamate receptor subunit GluR1.

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McIlhinney, R A; Molnár, E

1996-04-01

To identify the location of the first transmembrane segment of the GluR1 glutamate receptor subunit artificial stop codons have been introduced into the N-terminal domain at amino acid positions 442, 510, and 563, namely just before and spanning the proposed first two transmembrane regions. The resultant truncated N-terminal fragments of GluR1, termed NT1, NT2, and NT3 respectively were expressed in Cos-7 cells and their cellular distribution and cell-surface expression analysed using an N-terminal antibody to GluR1. All of the fragments were fully glycosylated and were found to be associated with cell membranes but none was secreted. Differential extraction of the cell membranes indicated that both NT1 and NT2 behave as peripheral membrane proteins. In contrast NT3, like the full subunit, has integral membrane protein properties. Furthermore only NT3 is expressed at the cell surface as determined by immunofluorescence and cell-surface biotinylation. Protease protection assays indicated that only NT3 had a cytoplasmic tail. Binding studies using the selective ligand [(3)H]alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate ([(3)H]AMPA) demonstrated that NT3 does not bind ligand. Together these results indicate that the first transmembrane domain of the GluR1 subunit lies between residues 509 and 562, that the N-terminal domain alone cannot form a functional ligand-binding site and that this domain can be targeted to the cell surface provided that it has a transmembrane-spanning region.
Expression variation in connected recombinant populations of Arabidopsis thaliana highlights distinct transcriptome architectures

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Cubillos Francisco A

2012-03-01

Full Text Available Abstract Background Expression traits can vary quantitatively between individuals and have a complex inheritance. Identification of the genetics underlying transcript variation can help in the understanding of phenotypic variation due to genetic factors regulating transcript abundance and shed light into divergence patterns. So far, only a limited number of studies have addressed this subject in Arabidopsis, with contrasting results due to dissimilar statistical power. Here, we present the transcriptome architecture in leaf tissue of two RIL sets obtained from a connected-cross design involving 3 commonly used accessions. We also present the transcriptome architecture observed in developing seeds of a third independent cross. Results The utilisation of the novel R/eqtl package (which goal is to automatize and extend functions from the R/qtl package allowed us to map 4,290 and 6,534 eQTLs in the Cvi-0 × Col-0 and Bur-0 × Col-0 recombinant populations respectively. In agreement with previous studies, we observed a larger phenotypic variance explained by eQTLs in linkage with the controlled gene (potentially cis-acting, compared to distant loci (acting necessarily indirectly or in trans. Distant eQTLs hotspots were essentially not conserved between crosses, but instead, cross-specific. Accounting for confounding factors using a probabilistic approach (VBQTL increased the mapping resolution and the number of significant associations. Moreover, using local eQTLs obtained from this approach, we detected evidence for a directional allelic effect in genes with related function, where significantly more eQTLs than expected by chance were up-regulated from one of the accessions. Primary experimental data, analysis parameters, eQTL results and visualisation of LOD score curves presented here are stored and accessible through the QTLstore service database http://qtlstore.versailles.inra.fr/. Conclusions Our results demonstrate the extensive diversity and

Development of Gene Expression Fingerprints for Identification of Environmental Contaminants Using cDNA Arrays

National Research Council Canada - National Science Library

Inouye, L

2004-01-01

...) to develop cDNA array-based assays that map gene expression from contaminant exposures. Results substantiate that distinct gene expression profiles exist for major contaminant classes such as PARs, PCBs, and PCDD/Fs...
Surface engineered magnetic nanoparticles for specific immunotargeting of cadherin expressing cells

International Nuclear Information System (INIS)

Moros, Maria; Puertas, Sara; Saez, Berta; Grazú, Valeria; Delhaes, Flavien; Feracci, Helene; De la Fuente, Jesús M

2016-01-01

In spite of historic advances in cancer biology and recent development of sophisticated chemotherapeutics, the outlook for patients with advanced cancer is still grim. In this sense nanoparticles (NPs), through their unique physical properties, enable the development of new approaches for cancer diagnosis and treatment. Thus far the most used active targeting scheme involves NPs functionalization with antibodies specific to molecules overexpressed on cancer cell’s surface. Therefore, such active targeting relies on differences in NPs uptake kinetics rates between tumor and healthy cells. Many cancers of epithelial origin are associated with the inappropriate expression of non-epithelial cadherins (e.g. N-, P-, -11) with concomitant loss of E-cadherin. Such phenomenon named cadherin switching favors tumor development and metastasis via interactions of tumor cells with stromal components. That is why we optimized the oriented functionalization of fluorescently labelled magnetic NPs with a novel antibody specific for the extracellular domain of cadherin-11. The obtained Ab-NPs exhibited high specificity when incubated with two cell lines used as models of tumor and healthy cells. Thus, cadherin switching offers a great opportunity for the development of active targeting strategies aimed to improve the early detection and treatment of cancer. (paper)
Distinct gene expression signatures in human embryonic stem cells differentiated towards definitive endoderm at single-cell level

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Norrman, Karin; Strömbeck, Anna; Semb, Henrik

2013-01-01

for the three activin A based protocols applied. Our data provide novel insights in DE gene expression at the cellular level of in vitro differentiated human embryonic stem cells, and illustrate the power of using single-cell gene expression profiling to study differentiation heterogeneity and to characterize...... of anterior definitive endoderm (DE). Here, we differentiated human embryonic stem cells towards DE using three different activin A based treatments. Differentiation efficiencies were evaluated by gene expression profiling over time at cell population level. A panel of key markers was used to study DE...... formation. Final DE differentiation was also analyzed with immunocytochemistry and single-cell gene expression profiling. We found that cells treated with activin A in combination with sodium butyrate and B27 serum-free supplement medium generated the most mature DE cells. Cell population studies were...
MicroRNA transfection and AGO-bound CLIP-seq data sets reveal distinct determinants of miRNA action

DEFF Research Database (Denmark)

Wen, Jiayu; Parker, Brian J; Jacobsen, Anders

2011-01-01

the predictive effect of target flanking features. We observe distinct target determinants between expression-based and CLIP-based data. Target flanking features such as flanking region conservation are an important AGO-binding determinant-we hypothesize that CLIP experiments have a preference for strongly bound......Microarray expression analyses following miRNA transfection/inhibition and, more recently, Argonaute cross-linked immunoprecipitation (CLIP)-seq assays have been used to detect miRNA target sites. CLIP and expression approaches measure differing stages of miRNA functioning-initial binding of the mi...... miRNP-target interactions involving adjacent RNA-binding proteins that increase the strength of cross-linking. In contrast, seed-related features are major determinants in expression-based studies, but less so for CLIP-seq studies, and increased miRNA concentrations typical of transfection studies...
Distinct mechanisms of loss of IFN-gamma mediated HLA class I inducibility in two melanoma cell lines

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Rodríguez, Teresa; Méndez, Rosa; Del Campo, Ana; Jiménez, Pilar; Aptsiauri, Natalia; Garrido, Federico; Ruiz-Cabello, Francisco

2007-01-01

The inability of cancer cells to present antigen on the cell surface via MHC class I molecules is one of the mechanisms by which tumor cells evade anti-tumor immunity. Alterations of Jak-STAT components of interferon (IFN)-mediated signaling can contribute to the mechanism of cell resistance to IFN, leading to lack of MHC class I inducibility. Hence, the identification of IFN-γ-resistant tumors may have prognostic and/or therapeutic relevance. In the present study, we investigated a mechanism of MHC class I inducibility in response to IFN-γ treatment in human melanoma cell lines. Basal and IFN-induced expression of HLA class I antigens was analyzed by means of indirect immunofluorescence flow cytometry, Western Blot, RT-PCR, and quantitative real-time RT-PCR (TaqMan ® Gene Expression Assays). In demethylation studies cells were cultured with 5-aza-2'-deoxycytidine. Electrophoretic Mobility Shift Assay (EMSA) was used to assay whether IRF-1 promoter binding activity is induced in IFN-γ-treated cells. Altered IFN-γ mediated HLA-class I induction was observed in two melanoma cells lines (ESTDAB-004 and ESTDAB-159) out of 57 studied, while treatment of these two cell lines with IFN-α led to normal induction of HLA class I antigen expression. Examination of STAT-1 in ESTDAB-004 after IFN-γ treatment demonstrated that the STAT-1 protein was expressed but not phosphorylated. Interestingly, IFN-α treatment induced normal STAT-1 phosphorylation and HLA class I expression. In contrast, the absence of response to IFN-γ in ESTDAB-159 was found to be associated with alterations in downstream components of the IFN-γ signaling pathway. We observed two distinct mechanisms of loss of IFN-γ inducibility of HLA class I antigens in two melanoma cell lines. Our findings suggest that loss of HLA class I induction in ESTDAB-004 cells results from a defect in the earliest steps of the IFN-γ signaling pathway due to absence of STAT-1 tyrosine-phosphorylation, while absence
Dynamic facial expressions evoke distinct activation in the face perception network: a connectivity analysis study.

Science.gov (United States)

Foley, Elaine; Rippon, Gina; Thai, Ngoc Jade; Longe, Olivia; Senior, Carl

2012-02-01

Very little is known about the neural structures involved in the perception of realistic dynamic facial expressions. In the present study, a unique set of naturalistic dynamic facial emotional expressions was created. Through fMRI and connectivity analysis, a dynamic face perception network was identified, which is demonstrated to extend Haxby et al.'s [Haxby, J. V., Hoffman, E. A., & Gobbini, M. I. The distributed human neural system for face perception. Trends in Cognitive Science, 4, 223-233, 2000] distributed neural system for face perception. This network includes early visual regions, such as the inferior occipital gyrus, which is identified as insensitive to motion or affect but sensitive to the visual stimulus, the STS, identified as specifically sensitive to motion, and the amygdala, recruited to process affect. Measures of effective connectivity between these regions revealed that dynamic facial stimuli were associated with specific increases in connectivity between early visual regions, such as the inferior occipital gyrus and the STS, along with coupling between the STS and the amygdala, as well as the inferior frontal gyrus. These findings support the presence of a distributed network of cortical regions that mediate the perception of different dynamic facial expressions.
Distinct effects of calorie restriction on adipose tissue cytokine and angiogenesis profiles in obese and lean mice

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Kurki Eveliina

2012-06-01

Full Text Available Abstract Background Obesity associates with low-grade inflammation and adipose tissue remodeling. Using sensitive high-throughput protein arrays we here investigated adipose tissue cytokine and angiogenesis-related protein profiles from obese and lean mice, and in particular, the influence of calorie restriction (CR. Methods Tissue samples from visceral fat were harvested from obese mice fed with a high-fat diet (60% of energy, lean controls receiving low-fat control diet as well as from obese and lean mice kept under CR (energy intake 70% of ad libitum intake for 50 days. Protein profiles were analyzed using mouse cytokine and angiogenesis protein array kits. Results In obese and lean mice, CR was associated with 11.3% and 15.6% reductions in body weight, as well as with 4.0% and 4.6% reductions in body fat percentage, respectively. Obesity induced adipose tissue cytokine expressions, the most highly upregulated cytokines being IL-1ra, IL-2, IL-16, MCP-1, MIG, RANTES, C5a, sICAM-1 and TIMP-1. CR increased sICAM-1 and TIMP-1 expression both in obese and lean mice. Overall, CR showed distinct effects on cytokine expressions; in obese mice CR largely decreased but in lean mice increased adipose tissue cytokine expressions. Obesity was also associated with increased expressions of angiogenesis-related proteins, in particular, angiogenin, endoglin, endostatin, endothelin-1, IGFBP-3, leptin, MMP-3, PAI-1, TIMP-4, CXCL16, platelet factor 4, DPPIV and coagulation factor III. CR increased endoglin, endostatin and platelet factor 4 expressions, and decreased IGFBP-3, NOV, MMP-9, CXCL16 and osteopontin expressions both in obese and lean mice. Interestingly, in obese mice, CR decreased leptin and TIMP-4 expressions, whereas in lean mice their expressions were increased. CR decreased MMP-3 and PAI-1 only in obese mice, whereas CR decreased FGF acidic, FGF basic and coagulation factor III, and increased angiogenin and DPPIV expression only in lean mice
Distinctive CD8+ T cell and MHC class I signatures in polycythemia vera patients.

Science.gov (United States)

Cardoso, Elsa M; Esgalhado, André J; Patrão, Luís; Santos, Mónica; Neves, Vasco Pinto; Martinez, Jorge; Patto, Maria Assunção Vaz; Silva, Helena; Arosa, Fernando A

2018-05-22

Polycythemia vera (PV) is a myeloproliferative neoplasm characterized by overproduction of red blood cells. We have performed a comprehensive characterization of blood immune cells for expression of naïve and memory receptors as well as β 2 m-associated and β 2 m-free MHC class I heavy chains, also known as closed and open conformers, respectively, in PV patients and age-matched controls (CTR). We show that the peripheral CD3 + CD8 + T cell pool in PV patients is clearly divided into two discrete populations, a more granular CD3 + CD8 high T cell population enriched in effector-memory CD45RA + T cells (CD8 + TEMRA) when compared to CTR (P e., CD3 + CD8 int CD28 int . While the percentage of CD3 + CD8 int TN cells correlated positively with the number of erythrocytes, the percentage of CD3 + CD8 int TEMRA correlated negatively with the number of platelets. Finally, we report that PV patients' lymphocytes and monocytes display lower levels of closed (W6/32 + ) MHC-I conformers at the cell surface while exhibiting increased amounts of open (HC-10 + ) MHC-I conformers. The implications of this distinctive immune signature are discussed.
Diverse Kir expression contributes to distinct bimodal distribution of resting potentials and vasotone responses of arterioles.

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Yuqin Yang

Full Text Available The resting membrane potential (RP of vascular smooth muscle cells (VSMCs is a major determinant of cytosolic calcium concentration and vascular tone. The heterogeneity of RPs and its underlying mechanism among different vascular beds remain poorly understood. We compared the RPs and vasomotion properties between the guinea pig spiral modiolar artery (SMA, brain arterioles (BA and mesenteric arteries (MA. We found: 1 RPs showed a robust bimodal distribution peaked at -76 and -40 mV evenly in the SMA, unevenly at -77 and -51 mV in the BA and ~-71 and -52 mV in the MA. Ba(2+ 0.1 mM eliminated their high RP peaks ~-75 mV. 2 Cells with low RP (~-45 mV hyperpolarized in response to 10 mM extracellular K(+, while cells with a high RP depolarized, and cells with intermediate RP (~-58 mV displayed an initial hyperpolarization followed by prolonged depolarization. Moderate high K(+ typically induced dilation, constriction and a dilation followed by constriction in the SMA, MA and BA, respectively. 3 Boltzmann-fit analysis of the Ba(2+-sensitive inward rectifier K(+ (Kir whole-cell current showed that the maximum Kir conductance density significantly differed among the vessels, and the half-activation voltage was significantly more negative in the MA. 4 Corresponding to the whole-cell data, computational modeling simulated the three RP distribution patterns and the dynamics of RP changes obtained experimentally, including the regenerative swift shifts between the two RP levels after reaching a threshold. 5 Molecular works revealed strong Kir2.1 and Kir2.2 transcripts and Kir2.1 immunolabeling in all 3 vessels, while Kir2.3 and Kir2.4 transcript levels varied. We conclude that a dense expression of functional Kir2.X channels underlies the more negative RPs in endothelial cells and a subset of VSMC in these arterioles, and the heterogeneous Kir function is primarily responsible for the distinct bimodal RPs among these arterioles. The fast Kir
Layer 5 Callosal Parvalbumin-Expressing Neurons: A Distinct Functional Group of GABAergic Neurons.

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Zurita, Hector; Feyen, Paul L C; Apicella, Alfonso Junior

2018-01-01

Previous studies have shown that parvalbumin-expressing neurons (CC-Parv neurons) connect the two hemispheres of motor and sensory areas via the corpus callosum, and are a functional part of the cortical circuit. Here we test the hypothesis that layer 5 CC-Parv neurons possess anatomical and molecular mechanisms which dampen excitability and modulate the gating of interhemispheric inhibition. In order to investigate this hypothesis we use viral tracing to determine the anatomical and electrophysiological properties of layer 5 CC-Parv and parvalbumin-expressing (Parv) neurons of the mouse auditory cortex (AC). Here we show that layer 5 CC-Parv neurons had larger dendritic fields characterized by longer dendrites that branched farther from the soma, whereas layer 5 Parv neurons had smaller dendritic fields characterized by shorter dendrites that branched nearer to the soma. The layer 5 CC-Parv neurons are characterized by delayed action potential (AP) responses to threshold currents, lower firing rates, and lower instantaneous frequencies compared to the layer 5 Parv neurons. Kv1.1 containing K + channels are the main source of the AP repolarization of the layer 5 CC-Parv and have a major role in determining both the spike delayed response, firing rate and instantaneous frequency of these neurons.
Heterogeneous nuclear ribonucleoproteins H, H', and F are members of a ubiquitously expressed subfamily of related but distinct proteins encoded by genes mapping to different chromosomes

DEFF Research Database (Denmark)

Honoré, B; Rasmussen, H H; Vorum, H

1995-01-01

Molecular cDNA cloning, two-dimensional gel immunoblotting, and amino acid microsequencing identified three sequence-unique and distinct proteins that constitute a subfamily of ubiquitously expressed heterogeneous nuclear ribonucleoproteins corresponding to hnRNPs H, H', and F. These proteins share...... epitopes and sequence identity with two other proteins, isoelectric focusing sample spot numbers 2222 (37.6 kDa; pI 6.5) and 2326 (39.5 kDa; pI 6.6), indicating that the subfamily may contain additional members. The identity between hnRNPs H and H' is 96%, between H and F 78%, and between H' and F 75......%, respectively. The three proteins contain three repeats, which we denote quasi-RRMs (qRRMs) since they have a remote similarity to the RNA recognition motif (RRM). The three qRRMs of hnRNP H, with a few additional NH2-terminal amino acids, were constructed by polymerase chain reaction amplification and used...
Syndecan expressions in the human amnion and chorionic plate

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T. Lorenzi

2010-10-01

Full Text Available The syndecan family consists of four distinct membrane glycoproteins in mammals. Syndecans control cell proliferation, differentiation, adhesion and migration through participation in cell-cell interactions, anchorage of cells to the extracellular environment, and modulation of multiple growth factors. Therefore, syndecans may play a pivotal role in the regulation of cell behaviour depending on the cellular microenvironment. Here, we demonstrate that syndecan-1, syndecan-2 and syndecan-4 are expressed in fetal membrane tissue with different immunolocalizations. Syndecan-1 is expressed in the amniotic epithelium, localizing at basolateral cell surfaces. Syndecan-2 and syndecan-4, in contrast, are mostly localized in intracellular compartments, in the extravillous cytotrophoblastic cells and in some fibroblasts of the chorionic plate as well as in the amniotic epithelial cells. In the latter, syndecan-4 is mainly localized in the apical part of the cells. Our results strongly suggest a key role of syndecan-1, syndecan-2 and syndecan-4 in the determination of structural and functional characteristics of human amnion and chorionic plate. Since the solute exchanges between fetus and mother take place in fetal membranes, our data suggest that syndecans are important players in the placenta for the establishment of the fetal-maternal inter-communication.
Methylation and in vivo expression of the surface-exposed Leptospira interrogans outer-membrane protein OmpL32.

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Eshghi, Azad; Pinne, Marija; Haake, David A; Zuerner, Richard L; Frank, Ami; Cameron, Caroline E

2012-03-01

Recent studies have revealed that bacterial protein methylation is a widespread post-translational modification that is required for virulence in selected pathogenic bacteria. In particular, altered methylation of outer-membrane proteins has been shown to modulate the effectiveness of the host immune response. In this study, 2D gel electrophoresis combined with MALDI-TOF MS identified a Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 protein, corresponding to ORF LIC11848, which undergoes extensive and differential methylation of glutamic acid residues. Immunofluorescence microscopy implicated LIC11848 as a surface-exposed outer-membrane protein, prompting the designation OmpL32. Indirect immunofluorescence microscopy of golden Syrian hamster liver and kidney sections revealed expression of OmpL32 during colonization of these organs. Identification of methylated surface-exposed outer-membrane proteins, such as OmpL32, provides a foundation for delineating the role of this post-translational modification in leptospiral virulence.
Interaction of VLA-5 Molecule With Rheumatoid Articular Cartilage Surface : An Electron Microscopic Evidence of Expression of VLA-5 on Pannus Invading Cells

OpenAIRE

Ishikawa, Hitoshi; Hirata, Souichirou; Saura, Ryuuichi; Andoh, Yoshihiro; Mizuno, Kosaku

1998-01-01

Pannus is made up mainly of fibroblasts, macrophages and lymphocytes. VLA-5 positive cells are present in the pannus in large numbers. It is likely that the tissue distribution of infiltrated cells derived from post-capillary venules is influenced by the ECM of the pannus and the ability of these cells to interact with the ECM through surface receptor expression. VLA-5 molecules are the predominant (31 integrins expressed by synovial pannus. Since the VLA integrins function as fibronectin rec...
Adult-type hypolactasia and regulation of lactase expression

DEFF Research Database (Denmark)

Troelsen, Jesper Thorvald

2005-01-01

, the main carbohydrate in milk. Individuals with adult-type hypolactasia lose their lactase expression before adulthood and consequently often become lactose intolerant with associated digestive problems (e.g. diarrhoea). In contrast, lactase persistent individuals have a lifelong lactase expression......A common genetically determined polymorphism in the human population leads to two distinct phenotypes in adults, lactase persistence and adult-type hypolactasia (lactase non-persistence). All healthy newborn children express high levels of lactase and are able to digest large quantities of lactose...... and are able to digest lactose as adults. Lactase persistence can be regarded as the mutant phenotype since other mammals down-regulate their lactase expression after weaning (the postweaning decline). This phenomenon does not occur in lactase persistent individuals. The regulation of lactase expression...
Counselor Identity: Conformity or Distinction?

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McLaughlin, Jerry E.; Boettcher, Kathryn

2009-01-01

The authors explore 3 debates in other disciplines similar to counseling's identity debate in order to learn about common themes and outcomes. Conformity, distinction, and cohesion emerged as common themes. They conclude that counselors should retain their distinctive, humanistic approach rather than conforming to the dominant, medical approach.
Data-driven asthma endotypes defined from blood biomarker and gene expression data.

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Barbara Jane George

Full Text Available The diagnosis and treatment of childhood asthma is complicated by its mechanistically distinct subtypes (endotypes driven by genetic susceptibility and modulating environmental factors. Clinical biomarkers and blood gene expression were collected from a stratified, cross-sectional study of asthmatic and non-asthmatic children from Detroit, MI. This study describes four distinct asthma endotypes identified via a purely data-driven method. Our method was specifically designed to integrate blood gene expression and clinical biomarkers in a way that provides new mechanistic insights regarding the different asthma endotypes. For example, we describe metabolic syndrome-induced systemic inflammation as an associated factor in three of the four asthma endotypes. Context provided by the clinical biomarker data was essential in interpreting gene expression patterns and identifying putative endotypes, which emphasizes the importance of integrated approaches when studying complex disease etiologies. These synthesized patterns of gene expression and clinical markers from our research may lead to development of novel serum-based biomarker panels.
Imbalanced expression of functional surface molecules in regulatory and effector T cells in systemic lupus erythematosus

International Nuclear Information System (INIS)

Mesquita Júnior, D.; Cruvinel, W.M.; Araujo, J.A.P.; Salmazi, K.C.; Kallas, E.G.; Andrade, L.E.C.

2014-01-01

Regulatory T (TREG) cells play an important role in maintaining immune tolerance and avoiding autoimmunity. We analyzed the expression of membrane molecules in TREG and effector T cells in systemic lupus erythematosus (SLE). TREG and effector T cells were analyzed for the expression of CTLA-4, PD1, CD28, CD95, GITR, HLA-DR, OX40, CD40L, and CD45RO in 26 patients with active disease, 31 with inactive disease, and 26 healthy controls. TREG cells were defined as CD25 +/high CD127 Ø/low FoxP3 + , and effector T cells were defined as CD25 + CD127 + FoxP3 Ø . The ratio of TREG to effector T cells expressing GITR, PD1, HLA-DR, OX40, CD40L, and CD45RO was determined in the three groups. The frequency of TREG cells was similar in patients with SLE and controls. However, SLE patients had a decreased frequency of CTLA-4 + TREG and CD28 + TREG cells and an increased frequency of CD40L + TREG cells. There was a decrease in the TREG/effector-T ratio for GITR + , HLA-DR + , OX40 + , and CD45RO + cells, and an increased ratio of TREG/effector-T CD40L + cells in patients with SLE. In addition, CD40L + TREG cell frequency correlated with the SLE disease activity index (P=0.0163). In conclusion, our findings showed several abnormalities in the expression of functionally critical surface molecules in TREG and effector T cells in SLE that may be relevant to the pathogenesis of this disease
Imbalanced expression of functional surface molecules in regulatory and effector T cells in systemic lupus erythematosus

Energy Technology Data Exchange (ETDEWEB)

Mesquita Júnior, D. [Disciplina de Reumatologia, Departamento de Medicina, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Cruvinel, W.M. [Disciplina de Reumatologia, Departamento de Medicina, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Departamento de Biomedicina, Universidade Católica de Goiás, Goiânia, GO (Brazil); Araujo, J.A.P. [Disciplina de Reumatologia, Departamento de Medicina, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil); Salmazi, K.C.; Kallas, E.G. [Disciplina de Imunologia Clínica e Alergia, Departamento de Clínica Médica, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP (Brazil); Andrade, L.E.C. [Disciplina de Reumatologia, Departamento de Medicina, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP (Brazil)

2014-08-22

Regulatory T (TREG) cells play an important role in maintaining immune tolerance and avoiding autoimmunity. We analyzed the expression of membrane molecules in TREG and effector T cells in systemic lupus erythematosus (SLE). TREG and effector T cells were analyzed for the expression of CTLA-4, PD1, CD28, CD95, GITR, HLA-DR, OX40, CD40L, and CD45RO in 26 patients with active disease, 31 with inactive disease, and 26 healthy controls. TREG cells were defined as CD25{sup +/high}CD127{sup Ø/low}FoxP3{sup +}, and effector T cells were defined as CD25{sup +}CD127{sup +}FoxP3{sup Ø}. The ratio of TREG to effector T cells expressing GITR, PD1, HLA-DR, OX40, CD40L, and CD45RO was determined in the three groups. The frequency of TREG cells was similar in patients with SLE and controls. However, SLE patients had a decreased frequency of CTLA-4{sup +}TREG and CD28{sup +}TREG cells and an increased frequency of CD40L{sup +}TREG cells. There was a decrease in the TREG/effector-T ratio for GITR{sup +}, HLA-DR{sup +}, OX40{sup +}, and CD45RO{sup +} cells, and an increased ratio of TREG/effector-T CD40L{sup +} cells in patients with SLE. In addition, CD40L{sup +}TREG cell frequency correlated with the SLE disease activity index (P=0.0163). In conclusion, our findings showed several abnormalities in the expression of functionally critical surface molecules in TREG and effector T cells in SLE that may be relevant to the pathogenesis of this disease.
Characterization of Trichome-Expressed BAHD Acyltransferases in Petunia axillaris Reveals Distinct Acylsugar Assembly Mechanisms within the Solanaceae1[OPEN

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Uebler, Joseph B.; Liu, Xiaoxiao

2017-01-01

Acylsugars are synthesized in the glandular trichomes of the Solanaceae family and are implicated in protection against abiotic and biotic stress. Acylsugars are composed of either sucrose or glucose esterified with varying numbers of acyl chains of differing length. In tomato (Solanum lycopersicum), acylsugar assembly requires four acylsugar acyltransferases (ASATs) of the BAHD superfamily. Tomato ASATs catalyze the sequential esterification of acyl-coenzyme A thioesters to the R4, R3, R3ʹ, and R2 positions of sucrose, yielding a tetra-acylsucrose. Petunia spp. synthesize acylsugars that are structurally distinct from those of tomato. To explore the mechanisms underlying this chemical diversity, a Petunia axillaris transcriptome was mined for trichome preferentially expressed BAHDs. A combination of phylogenetic analyses, gene silencing, and biochemical analyses coupled with structural elucidation of metabolites revealed that acylsugar assembly is not conserved between tomato and petunia. In P. axillaris, tetra-acylsucrose assembly occurs through the action of four ASATs, which catalyze sequential addition of acyl groups to the R2, R4, R3, and R6 positions. Notably, in P. axillaris, PaxASAT1 and PaxASAT4 catalyze the acylation of the R2 and R6 positions of sucrose, respectively, and no clear orthologs exist in tomato. Similarly, petunia acylsugars lack an acyl group at the R3ʹ position, and congruently, an ortholog of SlASAT3, which catalyzes acylation at the R3ʹ position in tomato, is absent in P. axillaris. Furthermore, where putative orthologous relationships of ASATs are predicted between tomato and petunia, these are not supported by biochemical assays. Overall, these data demonstrate the considerable evolutionary plasticity of acylsugar biosynthesis. PMID:28701351

Comparative pharmacology of a new recombinant FSH expressed by a human cell line

DEFF Research Database (Denmark)

Koechling, Wolfgang; Plaksin, Daniel; Croston, Glenn E.

2017-01-01

Recombinant FSH proteins are important therapeutic agents for the treatment of infertility, including follitropin alfa expressed in Chinese Hamster Ovary (CHO) cells and, more recently, follitropin delta expressed in the human cell line PER.C6. These recombinant FSH proteins have distinct glycosy...
Distinction of heterogeneity on Au nanostructured surface based on phase contrast imaging of atomic force microscopy

International Nuclear Information System (INIS)

Jung, Mi; Choi, Jeong-Woo

2010-01-01

The discrimination of the heterogeneity of different materials on nanostructured surfaces has attracted a great deal of interest in biotechnology as well as nanotechnology. Phase imaging through tapping mode of atomic force microscopy (TMAFM) can be used to distinguish the heterogeneity on a nanostructured surface. Nanostructures were fabricated using anodic aluminum oxide (AAO). An 11-mercaptoundecanoic acid (11-MUA) layer adsorbed onto the Au nanodots through self-assembly to improve the bio-compatibility. The Au nanostructures that were modified with 11-MUA and the concave surfaces were investigated using the TMAFM phase images to compare the heterogeneous and homogeneous nanostructured surfaces. Although the topography and phase images were taken simultaneously, the images were different. Therefore, the contrast in the TMAFM phase images revealed the different compositional materials on the heterogeneous nanostructure surface.
Developmental and adult characterization of secretagogin expressing amacrine cells in zebrafish retina.

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Stefanie Dudczig

Full Text Available Calcium binding proteins show stereotypical expression patterns within diverse neuron types across the central nervous system. Here, we provide a characterization of developmental and adult secretagogin-immunolabelled neurons in the zebrafish retina with an emphasis on co-expression of multiple calcium binding proteins. Secretagogin is a recently identified and cloned member of the F-hand family of calcium binding proteins, which labels distinct neuron populations in the retinas of mammalian vertebrates. Both the adult distribution of secretagogin labeled retinal neurons as well as the developmental expression indicative of the stage of neurogenesis during which this calcium binding protein is expressed was quantified. Secretagogin expression was confined to an amacrine interneuron population in the inner nuclear layer, with monostratified neurites in the center of the inner plexiform layer and a relatively regular soma distribution (regularity index > 2.5 across central-peripheral areas. However, only a subpopulation (~60% co-labeled with gamma-aminobutyric acid as their neurotransmitter, suggesting that possibly two amacrine subtypes are secretagogin immunoreactive. Quantitative co-labeling analysis with other known amacrine subtype markers including the three main calcium binding proteins parvalbumin, calbindin and calretinin identifies secretagogin immunoreactive neurons as a distinct neuron population. The highest density of secretagogin cells of ~1800 cells / mm2 remained relatively evenly along the horizontal meridian, whilst the density dropped of to 125 cells / mm2 towards the dorsal and ventral periphery. Thus, secretagogin represents a new amacrine label within the zebrafish retina. The developmental expression suggests a possible role in late stage differentiation. This characterization forms the basis of functional studies assessing how the expression of distinct calcium binding proteins might be regulated to compensate for the loss
Expression profiles of aquaporins in rat conjunctiva, cornea, lacrimal gland and Meibomian gland.

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Yu, Dongfang; Thelin, William R; Randell, Scott H; Boucher, Richard C

2012-10-01

The aim of the study was to elucidate aquaporin (AQP) family member mRNA expression and protein expression/localization in the rat lacrimal functional unit. The mRNA expression of all rat AQPs (AQP0-9, 11-12) in palpebral, fornical, and bulbar conjunctiva, cornea, lacrimal gland, and Meibomian gland was measured by Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) and real time RT-PCR. Antibodies against AQP1, 3, 4, 5, 9, and 11 were used in Western blotting and immunohistochemistry to determine protein expression and distribution. Our study demonstrated characteristic AQP expression profiles in rat ocular tissues. AQP1, 3, 4, 5, 8, 9, 11, and 12 mRNA were detected in conjunctiva. AQP0, 1, 2, 3, 4, 5, 6, 11, and 12 mRNA were expressed in cornea. AQP0, 1, 2, 3, 4, 5, 7, 8, and 11 mRNA were detected in lacrimal gland. AQP1, 3, 4, 5, 7, 8, 9, 11, and 12 mRNA were identified in Meibomian gland. By Western blot, AQP1, 3, 5, and 11 were detected in conjunctiva; AQP1, 3, 5, and 11 were identified in cornea; AQP1, 3, 4, 5, and 11 were detected in lacrimal gland; and AQP1, 3, 4, 5, 9, and 11 were present in Meibomian gland. Immunohistochemistry localized AQPs to distinct sites in the various tissues. This study rigorously analyzed AQPs expression and localization in rat conjunctiva, cornea, lacrimal gland, and Meibomian gland tissues. Our findings provide a comprehensive platform for further investigation into the physiological or pathophysiological relevance of AQPs in ocular surface. Copyright © 2012 Elsevier Ltd. All rights reserved.
Astragalus polysaccharides lowers plasma cholesterol through mechanisms distinct from statins.

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Yunjiu Cheng

Full Text Available To determine the efficacy and underlying mechanism of Astragalus polysaccharides (APS on plasma lipids in hypercholesterolemia hamsters. The effect of APS (0.25 g/kg/d on plasma and liver lipids, fecal bile acids and neutral sterol, cholesterol absorption and synthesis, HMG-CoA reductase activity, and gene and protein expressions in the liver and small intestine was investigated in twenty-four hypercholesterolemia hamsters. Treatment periods lasted for three months. APS significantly lowered plasma total cholesterol by 45.8%, triglycerides by 30%, and low-density lipoprotein-cholesterol by 47.4%, comparable to simvastatin. Further examinations revealed that APS reduced total cholesterol and triglycerides in the liver, increased fecal bile acid and neutral sterol excretion, inhibited cholesterol absorption, and by contrast, increased hepatic cholesterol synthesis and HMG-CoA reductase activity. Plasma total cholesterol or low-density lipoprotein-cholesterol levels were significantly correlated with cholesterol absorption rates. APS up-regulated cholesterol-7α-hydroxylase and LDL-receptor gene expressions. These new findings identify APS as a potential natural cholesterol lowering agent, working through mechanisms distinct from statins.
Na+/H+ Exchanger 3 Is Expressed in Two Distinct Types of Ionocyte, and Probably Augments Ammonia Excretion in One of Them, in the Gills of the Climbing Perch Exposed to Seawater

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Xiu L. Chen

2017-11-01

Full Text Available The freshwater climbing perch, Anabas testudineus, is an euryhaline teleost and an obligate air-breather with the ability to actively excrete ammonia. Members of the Na+/H+ exchanger (NHE family help maintain intracellular pH homeostasis and ionic balance through the electroneutral exchange of Na+ and H+. This study aimed to obtain, from the gills of A. testudineus, the full cDNA coding sequence of nhe3, and to determine the effects of exposure to seawater or 100 mmol l−1 of NH4Cl in fresh water on its mRNA and protein expression levels. Efforts were also made to elucidate the type of ionocyte that Nhe3 was associated with in the branchial epithelium of A. testudineus. The transcript level and protein abundance of nhe3/Nhe3 were very low in the gills of freshwater A. testudineus, but they increased significantly in the gills of fish acclimated to seawater. In the gills of fish exposed to seawater, Nhe3 was expressed in two distinct types of seawater-inducible Na+/K+-ATPase (Nka-immunoreactive ionocytes. In Nkaα1b-immunoreactive ionocytes, Nhe3 had an apical localization. As these ionocytes also expressed apical Rhcg1 and basolateral Rhcg2, which are known to transport ammonia, they probably participated in proton-facilitated ammonia excretion in A. testudineus during seawater acclimation. In Nkaα1c-immunoreactive ionocytes, Nhe3 was atypically expressed in the basolateral membrane, and its physiological function is uncertain. For A. testudineus exposed to NH4Cl in fresh water, the transcript and protein expression levels of nhe3/Nhe3 remained low. In conclusion, the branchial Nhe3 of A. testudineus plays a greater physiological role in passive ammonia transport and acid-base balance during seawater acclimation than in active ammonia excretion during environmental ammonia exposure.
Progress in the understanding of surface structure and surfactant influence on the electrocatalytic activity of gold nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Ferreira, V.C. [CQB, Departamento de Quimica e Bioquimica, Faculdade de Ciencias da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa (Portugal); CIQ-UP, Linha 4, Departamento de Quimica, Faculdade de Ciencias da Universidade do Porto, Rua do Campo Alegre 687, 4169-007 Porto (Portugal); Solla-Gullon, J.; Aldaz, A. [Instituto Universitario de Electroquimica, Universidad de Alicante, Apartado 99, 03080 Alicante (Spain); Silva, F. [CIQ-UP, Linha 4, Departamento de Quimica, Faculdade de Ciencias da Universidade do Porto, Rua do Campo Alegre 687, 4169-007 Porto (Portugal); Abrantes, L.M., E-mail: luisa.abrantes@fc.ul.pt [CQB, Departamento de Quimica e Bioquimica, Faculdade de Ciencias da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa (Portugal)

2011-11-01

Research highlights: > Electrochemical behaviour of AuNPs depends on the facets present in their surface. > Cubic and rod shaped nanoparticles comprise distinct crystallographic orientation. > Key rule of surfactant on the selective availability of specific surface domains. > AA oxidation at less stabilised (1 1 1) rod facets displays single-crystal response. - Abstract: The preparation of gold nanoparticles (Au-NPs) displaying specific shape, size and surface crystallographic domains has been investigated aiming to clarify the effect of the surface crystallographic orientation, of the synthesised nanoparticles, and surfactant influence on the electrochemical response of the ITO/Au-NPs modified electrodes. Polymorphic and nanorod-shaped Au-NPs have been obtained using distinct synthetic procedures in the presence of cetyltrimethylammonium bromide (CTAB), through seed-mediated growth methods, displaying distinct surface crystallographic domains confirmed by transmission electron microscopy, X-ray diffraction analysis and under potential deposition (UPD) of lead. The nanoparticles have been physically immobilised by casting on indium tin oxide (ITO) surfaces and the electrocatalytic activity of the Au-NPs evaluated using the ascorbic acid (AA) oxidation reaction, by cyclic voltammetry. The polymorphic and distinct surface crystallographic orientations of the Au-NPs were reflected in an irreproducible electrochemical response. Using gold nanorods comprising (1 1 1) and (1 1 0) facets and gold nanocubes consisting of faces displaying (1 0 0) surface domains, by contrasting the behaviour of CTAB-stabilised and clean particles, it has been possible to verify that the distinct voltammetric results are due to the exposure of specific crystallographic orientations owing to dissimilar interaction strength of CTAB with those facets.
Stage 3 immature human natural killer cells found in secondary lymphoid tissue constitutively and selectively express the TH17 cytokine interleukin-22

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Hughes, Tiffany; Becknell, Brian; McClory, Susan; Briercheck, Edward; Freud, Aharon G.; Zhang, Xiaoli; Mao, Hsiaoyin; Nuovo, Gerard; Yu, Jianhua

2009-01-01

Considerable functional heterogeneity within human natural killer (NK) cells has been revealed through the characterization of distinct NK-cell subsets. Accordingly, a small subset of CD56+NKp44+NK cells, termed NK-22 cells, was recently described within secondary lymphoid tissue (SLT) as IL-22− when resting, with a minor fraction of this population becoming IL-22+ when activated. Here we discover that the vast majority of stage 3 immature NK (iNK) cells in SLT constitutively and selectively express IL-22, a TH17 cytokine important for mucosal immunity, whereas earlier and later stages of NK developmental intermediates do not express IL-22. These iNK cells have a surface phenotype of CD34−CD117+CD161+CD94−, largely lack expression of NKp44 and CD56, and do not produce IFN-γ or possess cytolytic activity. In summary, stage 3 iNK cells are highly enriched for IL-22 and IL-26 messenger RNA, and IL-22 protein production, but do not express IL-17A or IL-17F. PMID:19244159
Gene Expression and the Diversity of Identified Neurons

OpenAIRE

Buck, L.; Stein, R.; Palazzolo, M.; Anderson, D. J.; Axel, R.

1983-01-01

Nervous systems consist of diverse populations of neurons that are anatomically and functionally distinct. The diversity of neurons and the precision with which they are interconnected suggest that specific genes or sets of genes are activated in some neurons but not expressed in others. Experimentally, this problem may be considered at two levels. First, what is the total number of genes expressed in the brain, and how are they distributed among the different populations of neurons? Second, ...
Ion surface collisions on surfaces relevant for fusion devices

International Nuclear Information System (INIS)

Rasul, B.; Endstrasser, N.; Zappa, F.; Grill, V.; Scheier, P.; Mark, T.

2006-01-01

Full text: One of the great challenges of fusion research is the compatibility of reactor grade plasmas with plasma facing materials coating the inner walls of a fusion reactor. The question of which surface coating should be used is of particular interest for the design of ITER. The impact of energetic plasma particles leads to sputtering of wall material into the plasma. A possible solution for the coating of plasma facing walls would be the use of special carbon surfaces. Investigations of these various surfaces have been started at BESTOF ion-surface collision apparatus. Experiment beam of singly charged molecular ions of hydrocarbon molecules, i.e. C 2 H + 4 , is generated in a Nier-type electron impact ionization source at an electron energy of about 70 eV. In the first double focusing mass spectrometer the ions are mass and energy analyzed and afterwards refocused onto a surface. The secondary reaction products are monitored using a Time Of Flight mass spectrometer. The secondary ion mass spectra are recorded as a function of the collision energy for different projectile ions and different surfaces. A comparison of these spectra show for example distinct changes in the survival probability of the same projectile ion C 2 H + 4 for different surfaces. (author)
A single point in protein trafficking by Plasmodium falciparum determines the expression of major antigens on the surface of infected erythrocytes targeted by human antibodies.

Science.gov (United States)

Chan, Jo-Anne; Howell, Katherine B; Langer, Christine; Maier, Alexander G; Hasang, Wina; Rogerson, Stephen J; Petter, Michaela; Chesson, Joanne; Stanisic, Danielle I; Duffy, Michael F; Cooke, Brian M; Siba, Peter M; Mueller, Ivo; Bull, Peter C; Marsh, Kevin; Fowkes, Freya J I; Beeson, James G

2016-11-01

Antibodies to blood-stage antigens of Plasmodium falciparum play a pivotal role in human immunity to malaria. During parasite development, multiple proteins are trafficked from the intracellular parasite to the surface of P. falciparum-infected erythrocytes (IEs). However, the relative importance of different proteins as targets of acquired antibodies, and key pathways involved in trafficking major antigens remain to be clearly defined. We quantified antibodies to surface antigens among children, adults, and pregnant women from different malaria-exposed regions. We quantified the importance of antigens as antibody targets using genetically engineered P. falciparum with modified surface antigen expression. Genetic deletion of the trafficking protein skeleton-binding protein-1 (SBP1), which is involved in trafficking the surface antigen PfEMP1, led to a dramatic reduction in antibody recognition of IEs and the ability of human antibodies to promote opsonic phagocytosis of IEs, a key mechanism of parasite clearance. The great majority of antibody epitopes on the IE surface were SBP1-dependent. This was demonstrated using parasite isolates with different genetic or phenotypic backgrounds, and among antibodies from children, adults, and pregnant women in different populations. Comparisons of antibody reactivity to parasite isolates with SBP1 deletion or inhibited PfEMP1 expression suggest that PfEMP1 is the dominant target of acquired human antibodies, and that other P. falciparum IE surface proteins are minor targets. These results establish SBP1 as part of a critical pathway for the trafficking of major surface antigens targeted by human immunity, and have key implications for vaccine development, and quantifying immunity in populations.
Herbivore Oral Secreted Bacteria Trigger Distinct Defense Responses in Preferred and Non-Preferred Host Plants.

Science.gov (United States)

Wang, Jie; Chung, Seung Ho; Peiffer, Michelle; Rosa, Cristina; Hoover, Kelli; Zeng, Rensen; Felton, Gary W

2016-06-01

Insect symbiotic bacteria affect host physiology and mediate plant-insect interactions, yet there are few clear examples of symbiotic bacteria regulating defense responses in different host plants. We hypothesized that plants would induce distinct defense responses to herbivore- associated bacteria. We evaluated whether preferred hosts (horsenettle) or non-preferred hosts (tomato) respond similarly to oral secretions (OS) from the false potato beetle (FPB, Leptinotarsa juncta), and whether the induced defense triggered by OS was due to the presence of symbiotic bacteria in OS. Both horsenettle and tomato damaged by antibiotic (AB) treated larvae showed higher polyphenol oxidase (PPO) activity than those damaged by non-AB treated larvae. In addition, application of OS from AB treated larvae induced higher PPO activity compared with OS from non-AB treated larvae or water treatment. False potato beetles harbor bacteria that may provide abundant cues that can be recognized by plants and thus mediate corresponding defense responses. Among all tested bacterial isolates, the genera Pantoea, Acinetobacter, Enterobacter, and Serratia were found to suppress PPO activity in tomato, while only Pantoea sp. among these four isolates was observed to suppress PPO activity in horsenettle. The distinct PPO suppression caused by symbiotic bacteria in different plants was similar to the pattern of induced defense-related gene expression. Pantoea inoculated FPB suppressed JA-responsive genes and triggered a SA-responsive gene in both tomato and horsenettle. However, Enterobacter inoculated FPB eliminated JA-regulated gene expression and elevated SA-regulated gene expression in tomato, but did not show evident effects on the expression levels of horsenettle defense-related genes. These results indicate that suppression of plant defenses by the bacteria found in the oral secretions of herbivores may be a more widespread phenomenon than previously indicated.
Optimization of the Expression of DT386-BR2 Fusion Protein in Escherichia coli using Response Surface Methodology.

Science.gov (United States)

Shafiee, Fatemeh; Rabbani, Mohammad; Jahanian-Najafabadi, Ali

2017-01-01

The aim of this study was to determine the best condition for the production of DT386-BR2 fusion protein, an immunotoxin consisting of catalytic and translocation domains of diphtheria toxin fused to BR2, a cancer specific cell penetrating peptide, for targeted eradication of cancer cells, in terms of the host, cultivation condition, and culture medium. Recombinant pET28a vector containing the codons optimized for the expression of the DT386-BR2 gene was transformed to different strains of Escherichia coli ( E. coli BL21 DE3, E. coli Rosetta DE3 and E. coli Rosetta-gami 2 DE3), followed by the induction of expression using 1 mM IPTG. Then, the strain with the highest ability to produce recombinant protein was selected and used to determine the best expression condition using response surface methodology (RSM). Finally, the best culture medium was selected. Densitometry analysis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the expressed fusion protein showed that E. coli Rosetta DE3 produced the highest amounts of the recombinant fusion protein when quantified by 1 mg/ml bovine serum albumin (178.07 μg/ml). Results of RSM also showed the best condition for the production of the recombinant fusion protein was induction with 1 mM IPTG for 2 h at 37°C. Finally, it was established that terrific broth could produce higher amounts of the fusion protein when compared to other culture media. In this study, we expressed the recombinant DT386-BR2 fusion protein in large amounts by optimizing the expression host, cultivation condition, and culture medium. This fusion protein will be subjected to purification and evaluation of its cytotoxic effects in future studies.
Optimization of the Expression of DT386-BR2 Fusion Protein in Escherichia coli using Response Surface Methodology

Directory of Open Access Journals (Sweden)

Fatemeh Shafiee

2017-01-01

Full Text Available Background: The aim of this study was to determine the best condition for the production of DT386-BR2 fusion protein, an immunotoxin consisting of catalytic and translocation domains of diphtheria toxin fused to BR2, a cancer specific cell penetrating peptide, for targeted eradication of cancer cells, in terms of the host, cultivation condition, and culture medium. Materials and Methods: Recombinant pET28a vector containing the codons optimized for the expression of the DT386-BR2 gene was transformed to different strains of Escherichia coli (E. coli BL21 DE3, E. coli Rosetta DE3 and E. coli Rosetta-gami 2 DE3, followed by the induction of expression using 1 mM IPTG. Then, the strain with the highest ability to produce recombinant protein was selected and used to determine the best expression condition using response surface methodology (RSM. Finally, the best culture medium was selected. Results: Densitometry analysis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the expressed fusion protein showed that E. coli Rosetta DE3 produced the highest amounts of the recombinant fusion protein when quantified by 1 mg/ml bovine serum albumin (178.07 μg/ml. Results of RSM also showed the best condition for the production of the recombinant fusion protein was induction with 1 mM IPTG for 2 h at 37°C. Finally, it was established that terrific broth could produce higher amounts of the fusion protein when compared to other culture media. Conclusion: In this study, we expressed the recombinant DT386-BR2 fusion protein in large amounts by optimizing the expression host, cultivation condition, and culture medium. This fusion protein will be subjected to purification and evaluation of its cytotoxic effects in future studies.
Endocrine disrupting chemicals (bisphenol A, 4-nonylphenol, 4-tert-octylphenol) modulate expression of two distinct cytochrome P450 aromatase genes differently in gender types of the hermaphroditic fish Rivulus marmoratus.

Science.gov (United States)

Lee, Young-Mi; Seo, Jung Soo; Kim, Il-Chan; Yoon, Yong-Dal; Lee, Jae-Seong

2006-06-30

To understand the effect of endocrine-disrupting chemicals (EDCs) on cytochrome P450 aromatase (rm-cyp19) gene expression between gender types in the hermaphroditic fish Rivulus marmoratus, we cloned two distinct rm-cyp19 genes using RT-PCR with degenerative primers, obtained full-length cDNAs using 5'- and 3'-RACE-PCR methods, and completely sequenced them. The brain aromatase (rm-cyp19b) cDNA consisted of 2,124 bp including the open reading frame (ORF), which encoded a putative protein of 505 amino acids. The ovarian aromatase (rm-cyp19a) cDNA consisted of 2,075 bp, including the ORF encoding a putative protein of 516 amino acids. Expression patterns of rm-cyp19b and rm-cyp19a mRNAs were investigated in embryos of different developmental stages and in seven different tissues of adult fish. The rm-cyp19b gene in hermaphrodite and secondary male R. marmoratus was predominantly expressed in the brain, while the rm-cyp19a gene was expressed gender-specifically in the gonad. The expression of rm-cyp19b mRNA increased from stage 1 (2 d post fertilization) to stage 4 (12 d post fertilization) in a developmental stage-dependent manner but steeply decreased in the hatching stage. Compared to the rm-cyp19b gene, the abundance of ovarian aromatase rm-cyp19a transcripts was very low, and its expression was first detected at stage 3 and then decreased gradually to the hatching stage. Alteration of rm-cyp19b and rm-cyp19a gene expression was further analyzed in the brain and gonad by real-time RT-PCR 96 h after EDC exposure in hermaphrodites and secondary males. The brain aromatase rm-cyp19b gene was up-regulated in the brain after 4-nonylphenol (4-NP)-exposure, while the ovarian aromatase rm-cyp19a gene was significantly down-regulated in the gonad. In 300 microg/L 4-tert octylphenol (4-tert-OP), or 600 microg/L bisphenol A-exposed brain and gonad, both rm-cyp19b and rm-cyp19a genes were up-regulated. In the case of secondary males, the rm-cyp19b gene was highly expressed in
Quantitative rather than qualitative differences in gene expression predominate in intestinal cell maturation along distinct cell lineages

International Nuclear Information System (INIS)

Velcich, Anna; Corner, Georgia; Paul, Doru; Zhuang Min; Mariadason, John M.; Laboisse, Christian; Augenlicht, Leonard

2005-01-01

Several cell types are present in the intestinal epithelium that likely arise from a common precursor, the stem cell, and each mature cell type expresses a unique set of genes that characterizes its functional phenotype. Although the process of differentiation is intimately linked to the cessation of proliferation, the mechanisms that dictate intestinal cell fate determination are not well characterized. To investigate the reprogramming of gene expression during the cell lineage allocation/differentiation process, we took advantage of a unique system of two clonal derivatives of HT29 cells, Cl16E and Cl19A cells, which spontaneously differentiate as mucus producing goblet and chloride-secreting cells, respectively, as a function of time. By profiling gene expression, we found that these two cell lines show remarkably similar kinetics of change in gene expression and common clusters of coordinately regulated genes. This demonstrates that lineage-specific differentiation of intestinal epithelial cells is characterized overall by the sequential recruitment of functionally similar gene sets independent of the final phenotype of the mature cells
Overexpressed Genes/ESTs and Characterization of Distinct Amplicons on 17823 in Breast Cancer Cells

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Ayse E. Erson

2001-01-01

Full Text Available 17823 is a frequent site of gene amplification in breast cancer. Several lines of evidence suggest the presence of multiple amplicons on 17823. To characterize distinct amplicons on 17823 and localize putative oncogenes, we screened genes and expressed sequence tags (ESTs in existing physical and radiation hybrid maps for amplification and overexpression in breast cancer cell lines by semiquantitative duplex PCR, semiquantitative duplex RT-PCR, Southern blot, Northern blot analyses. We identified two distinct amplicons on 17823, one including TBX2 and another proximal region including RPS6KB1 (PS6K and MUL. In addition to these previously reported overexpressed genes, we also identified amplification and overexpression of additional uncharacterized genes and ESTs, some of which suggest potential oncogenic activity. In conclusion, we have further defined two distinct regions of gene amplification and overexpression on 17823 with identification of new potential oncogene candidates. Based on the amplification and overexpression patterns of known and as of yet unrecognized genes on 17823, it is likely that some of these genes mapping to the discrete amplicons function as oncogenes and contribute to tumor progression in breast cancer cells.
The surface brightness of 1550 galaxies in Fornax: automated galaxy surface photometry: Pt. 2

International Nuclear Information System (INIS)

Phillipps, S.; Disney, M.J.; Kibblewhite, E.J.; Cawson, M.G.M.

1987-01-01

A survey of a complete sample of galaxies in the region of the Fornax cluster is presented. Measurements with the Automatic Plate Measuring machine are used to derive the observed distribution of galaxy surface brightness for 1550 objects. Corrections for surface brightness dependent selection effects are then made in order to estimate the true distribution. It is found that the sample (with 16.6 ≤ Msub(APM) ≤ 19.1) is divided into two distinct populations. The 'normal' galaxies with extrapolated central surface brightness Ssub(x) ≤ 22.5 Bμ form a uniformly distributed background of field galaxies. Low surface brightness galaxies (Ssub(x) ≥ 22.5 Bμ), on the other hand, are strongly clumped about the cluster centre. There appear to be few low surface brightness field galaxies. (author)
Glucose transporter expression in human skeletal muscle fibers

DEFF Research Database (Denmark)

Gaster, M; Handberg, A; Beck-Nielsen, H

2000-01-01

, but its expression is markedly reduced around birth and is further reduced to undetectable levels within the first year of life; 2) GLUT-3 protein expression appears at 18 wk of gestation and disappears after birth; and 3) GLUT-4 protein is diffusely expressed in muscle cells throughout gestation, whereas...... after birth, the characteristic subcellular localization is as seen in adult muscle fibers. Our results show that GLUT-1, GLUT-3, and GLUT-4 seem to be of importance during muscle fiber growth and development. GLUT-5 protein was undetectable in fetal and adult skeletal muscle fibers. In adult muscle...... amplification (TSA) technique to detect the localization of glucose transporter expression in human skeletal muscle. We found expression of GLUT-1, GLUT-3, and GLUT-4 in developing human muscle fibers showing a distinct expression pattern. 1) GLUT-1 is expressed in human skeletal muscle cells during gestation...
Ray-map migration of transmitted surface waves

KAUST Repository

Li, Jing

2016-08-25

Near-surface normal faults can sometimes separate two distinct zones of velocity heterogeneity, where the medium on one side of the fault has a faster velocity than on the other side. Therefore, the slope of surface-wave arrivals in a common-shot gather should abruptly change near the surface projection of the fault. We present ray-map imaging method that migrates transmitted surface waves to the fault plane, and therefore it roughly estimates the orientation, depth, and location of the near-surface fault. The main benefits of this method are that it is computationally inexpensive and robust in the presence of noise.

Separate Developmental Programs for HLA-A and -B Cell Surface Expression during Differentiation from Embryonic Stem Cells to Lymphocytes, Adipocytes and Osteoblasts

DEFF Research Database (Denmark)

Sabir, Hardee J; Nehlin, Jan O; Qanie, Diyako

2013-01-01

-A, but not -B) is seen on some multipotent stem cells, and this raises the question how this is in other stem cells and how it changes during differentiation. In this study, we have used flow cytometry to investigate the cell surface expression of HLA-A and -B on human embryonic stem cells (hESC), human...... hematopoietic stem cells (hHSC), human mesenchymal stem cells (hMSC) and their fully-differentiated progenies such as lymphocytes, adipocytes and osteoblasts. hESC showed extremely low levels of HLA-A and no -B. In contrast, multipotent hMSC and hHSC generally expressed higher levels of HLA-A and clearly HLA......A major problem of allogeneic stem cell therapy is immunologically mediated graft rejection. HLA class I A, B, and Cw antigens are crucial factors, but little is known of their respective expression on stem cells and their progenies. We have recently shown that locus-specific expression (HLA...
Spatiotemporal expression of chondroitin sulfate sulfotransferases in the postnatal developing mouse cerebellum.

Science.gov (United States)

Ishii, Maki; Maeda, Nobuaki

2008-08-01

Chondroitin sulfate (CS) proteoglycans are major components of the cell surface and the extracellular matrix in the developing brain and bind to various proteins via CS chains in a CS structure-dependent manner. This study demonstrated the expression pattern of three CS sulfotransferase genes, dermatan 4-O-sulfotransferase (D4ST), uronyl 2-O-sulfotransferase (UST), and N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S-6ST), in the mouse postnatal cerebellum. These sulfotransferases are responsible for the biosynthesis of oversulfated structures in CS chains such as B, D, and E units, which constitute the binding sites for various heparin-binding proteins. Real-time reverse transcription-polymerase chain reaction analysis indicated that the expression of UST increased remarkably during cerebellar development. The amounts of B and D units, which are generated by UST activity, in the cerebellar CS chains also increased during development. In contrast, the expression of GalNAc4S-6ST and its biosynthetic product, E unit, decreased during postnatal development. In situ hybridization experiments revealed the levels of UST and GalNAc4S-6ST mRNAs to correlate inversely in many cells including Purkinje cells, granule cells in the external granular layer, and inhibitory interneurons. In these neurons, the expression of UST increased and that of GalNAc4S-6ST decreased during development and/or maturation. D4ST was also expressed by many neurons, but its expression was not simply correlated with development, which might contribute to the diversification of CS structures expressed by distinct neurons. These results suggest that the CS structures of various cerebellar neurons change during development and such changes of CS are involved in the regulation of various signaling pathways.
Per a 10 protease activity modulates CD40 expression on dendritic cell surface by nuclear factor-kappaB pathway.

Science.gov (United States)

Goel, C; Kalra, N; Dwarakanath, B S; Gaur, S N; Arora, N

2015-05-01

Serine protease activity of Per a 10 from Periplaneta americana modulates dendritic cell (DC) functions by a mechanism(s) that remains unclear. In the present study, Per a 10 protease activity on CD40 expression and downstream signalling was evaluated in DCs. Monocyte-derived DCs from cockroach-allergic patients were treated with proteolytically active/heat-inactivated Per a 10. Stimulation with active Per a 10 demonstrated low CD40 expression on DCs surface (P Per a 10, suggesting cleavage of CD40. Per a 10 activity reduced the interleukin (IL)-12 and interferon (IFN)-γ secretion by DCs (P Per a 10, indicating that low CD40 expression is associated with low levels of IL-12 secretion. Active Per a 10 stimulation caused low nuclear factor-kappa B (NF-κB) activation in DCs compared to heat-inactivated Per a 10. Inhibition of the NF-κB pathway suppressed the CD40 expression and IL-12 secretion by DCs, further indicating that NF-κB is required for CD40 up-regulation. CD40 expression activated the tumour necrosis factor (TNF) receptor-associated factor 6 (TRAF6), thereby suggesting its involvement in NF-κB activation. Protease activity of Per a 10 induced p38 mitogen-activated protein kinase (MAPK) activation that showed no significant effect on CD40 expression by DCs. However, inhibiting p38 MAPK or NF-κB suppressed the secretion of IL-12, IFN-γ, IL-6 and TNF-α by DCs. Such DCs further reduced the secretion of IL-4, IL-6, IL-12 and TNF-α by CD4(+) T cells. In conclusion, protease activity of Per a 10 reduces CD40 expression on DCs. CD40 down-regulation leads to low NF-κB levels, thereby modulating DC-mediated immune responses. © 2014 British Society for Immunology.
Evaluation of Surface Fatigue Strength Based on Surface Temperature

Science.gov (United States)

Deng, Gang; Nakanishi, Tsutomu

Surface temperature is considered to be an integrated index that is dependent on not only the load and the dimensions at the contact point but also the sliding velocity, rolling velocity, surface roughness, and lubrication conditions. Therefore, the surface durability of rollers and gears can be evaluated more exactly and simply by the use of surface temperature rather than Hertzian stress. In this research, surface temperatures of rollers under different rolling and sliding conditions are measured using a thermocouple. The effects of load P, mean velocity Vm and sliding velocity Vs on surface temperature are clarified. An experimental formula, which expresses the linear relationship between surface temperature and the P0.86Vs1.31Vm-0.83 value, is used to determine surface temperature. By comparing calculated and measured temperature on the tooth surface of a gear, this formula is confirmed to be applicable for gear tooth surface temperature calculation.
Surface chemistry of gold nanoparticles determines the biocorona composition impacting cellular uptake, toxicity and gene expression profiles in human endothelial cells.

Science.gov (United States)

Chandran, Parwathy; Riviere, Jim E; Monteiro-Riviere, Nancy A

2017-05-01

This study investigated the role of nanoparticle size and surface chemistry on biocorona composition and its effect on uptake, toxicity and cellular responses in human umbilical vein endothelial cells (HUVEC), employing 40 and 80 nm gold nanoparticles (AuNP) with branched polyethyleneimine (BPEI), lipoic acid (LA) and polyethylene glycol (PEG) coatings. Proteomic analysis identified 59 hard corona proteins among the various AuNP, revealing largely surface chemistry-dependent signature adsorbomes exhibiting human serum albumin (HSA) abundance. Size distribution analysis revealed the relative instability and aggregation inducing potential of bare and corona-bound BPEI-AuNP, over LA- and PEG-AuNP. Circular dichroism analysis showed surface chemistry-dependent conformational changes of proteins binding to AuNP. Time-dependent uptake of bare, plasma corona (PC) and HSA corona-bound AuNP (HSA-AuNP) showed significant reduction in uptake with PC formation. Cell viability studies demonstrated dose-dependent toxicity of BPEI-AuNP. Transcriptional profiling studies revealed 126 genes, from 13 biological pathways, to be differentially regulated by 40 nm bare and PC-bound BPEI-AuNP (PC-BPEI-AuNP). Furthermore, PC formation relieved the toxicity of cationic BPEI-AuNP by modulating expression of genes involved in DNA damage and repair, heat shock response, mitochondrial energy metabolism, oxidative stress and antioxidant response, and ER stress and unfolded protein response cascades, which were aberrantly expressed in bare BPEI-AuNP-treated cells. NP surface chemistry is shown to play the dominant role over size in determining the biocorona composition, which in turn modulates cell uptake, and biological responses, consequently defining the potential safety and efficacy of nanoformulations.
Expression and Functional Pathway Analysis of Nuclear Receptor NR2F2 in Ovarian Cancer

Science.gov (United States)

Hawkins, Shannon M.; Loomans, Holli A.; Wan, Ying-Wooi; Ghosh-Choudhury, Triparna; Coffey, Donna; Xiao, Weimin; Liu, Zhandong; Sangi-Haghpeykar, Haleh

2013-01-01

Context: Recent evidence implicates the orphan nuclear receptor, nuclear receptor subfamily 2, group F, member 2 (NR2F2; chicken ovalbumin upstream promoter-transcription factor II) as both a master regulator of angiogenesis and an oncogene in prostate and other human cancers. Objective: The objective of the study was to determine whether NR2F2 plays a role in ovarian cancer and dissect its potential mechanisms of action. Design, Setting, and Patients: We examined NR2F2 expression in healthy ovary and ovarian cancers using quantitative PCR and immunohistochemistry. NR2F2 expression was targeted in established ovarian cancer cell lines to assess the impact of dysregulated NR2F2 expression in the epithelial compartment of ovarian cancers. Results: Our results indicate that NR2F2 is robustly expressed in the stroma of healthy ovary with little or no expression in epithelia lining the ovarian surface, clefts, or crypts. This pattern of NR2F2 expression was markedly disrupted in ovarian cancers, in which decreased levels of stromal expression and ectopic epithelial expression were frequently observed. Ovarian cancers with the most disrupted patterns of NR2F2 were associated with significantly shorter disease-free interval by Kaplan-Meier analysis. Targeting NR2F2 expression in established ovarian cancer cell lines enhanced apoptosis and increased proliferation. In addition, we found that NR2F2 regulates the expression of NEK2, RAI14, and multiple other genes involved in the cell cycle, suggesting potential pathways by which dysregulated expression of NR2F2 impacts ovarian cancer. Conclusions: These results uncover novel roles for NR2F2 in ovarian cancer and point to a unique scenario in which a single nuclear receptor plays potentially distinct roles in the stromal and epithelial compartments of the same tissue. PMID:23690307
Intraspinal serotonergic neurons consist of two, temporally distinct populations in developing zebrafish.

Science.gov (United States)

Montgomery, Jacob E; Wiggin, Timothy D; Rivera-Perez, Luis M; Lillesaar, Christina; Masino, Mark A

2016-06-01

Zebrafish intraspinal serotonergic neuron (ISN) morphology and distribution have been examined in detail at different ages; however, some aspects of the development of these cells remain unclear. Although antibodies to serotonin (5-HT) have detected ISNs in the ventral spinal cord of embryos, larvae, and adults, the only tryptophan hydroxylase (tph) transcript that has been described in the spinal cord is tph1a. Paradoxically, spinal tph1a is only expressed transiently in embryos, which brings the source of 5-HT in the ISNs of larvae and adults into question. Because the pet1 and tph2 promoters drive transgene expression in the spinal cord, we hypothesized that tph2 is expressed in spinal cords of zebrafish larvae. We confirmed this hypothesis through in situ hybridization. Next, we used 5-HT antibody labeling and transgenic markers of tph2-expressing neurons to identify a transient population of ISNs in embryos that was distinct from ISNs that appeared later in development. The existence of separate ISN populations may not have been recognized previously due to their shared location in the ventral spinal cord. Finally, we used transgenic markers and immunohistochemical labeling to identify the transient ISN population as GABAergic Kolmer-Agduhr double-prime (KA″) neurons. Altogether, this study revealed a novel developmental paradigm in which KA″ neurons are transiently serotonergic before the appearance of a stable population of tph2-expressing ISNs. © 2015 Wiley Periodicals, Inc.
Moving faces, looking places: validation of the Amsterdam Dynamic Facial Expression Set (ADFES)

NARCIS (Netherlands)

van der Schalk, J.; Hawk, S.T.; Fischer, A.H.; Doosje, B.

2011-01-01

We report two studies validating a new standardized set of filmed emotion expressions, the Amsterdam Dynamic Facial Expression Set (ADFES). The ADFES is distinct from existing datasets in that it includes a face-forward version and two different head-turning versions (faces turning toward and away
Decreased surface expression of the δ subunit of the GABAA receptor contributes to reduced tonic inhibition in dentate granule cells in a mouse model of fragile X syndrome.

Science.gov (United States)

Zhang, Nianhui; Peng, Zechun; Tong, Xiaoping; Lindemeyer, A Kerstin; Cetina, Yliana; Huang, Christine S; Olsen, Richard W; Otis, Thomas S; Houser, Carolyn R

2017-11-01

While numerous changes in the GABA system have been identified in models of Fragile X Syndrome (FXS), alterations in subunits of the GABA A receptors (GABA A Rs) that mediate tonic inhibition are particularly intriguing. Considering the key role of tonic inhibition in controlling neuronal excitability, reduced tonic inhibition could contribute to FXS-associated disorders such as hyperactivity, hypersensitivity, and increased seizure susceptibility. The current study has focused on the expression and function of the δ subunit of the GABA A R, a major subunit involved in tonic inhibition, in granule cells of the dentate gyrus in the Fmr1 knockout (KO) mouse model of FXS. Electrophysiological studies of dentate granule cells revealed a marked, nearly four-fold, decrease in tonic inhibition in the Fmr1 KO mice, as well as reduced effects of two δ subunit-preferring pharmacological agents, THIP and DS2, supporting the suggestion that δ subunit-containing GABA A Rs are compromised in the Fmr1 KO mice. Immunohistochemistry demonstrated a small but statistically significant decrease in δ subunit labeling in the molecular layer of the dentate gyrus in Fmr1 KO mice compared to wildtype (WT) littermates. The discrepancy between the large deficits in GABA-mediated tonic inhibition in granule cells in the Fmr1 KO mice and only modest reductions in immunolabeling of the δ subunit led to studies of surface expression of the δ subunit. Cross-linking experiments followed by Western blot analysis demonstrated a small, non-significant decrease in total δ subunit protein in the hippocampus of Fmr1 KO mice, but a four-fold decrease in surface expression of the δ subunit in these mice. No significant changes were observed in total or surface expression of the α4 subunit protein, a major partner of the δ subunit in the forebrain. Postembedding immunogold labeling for the δ subunit demonstrated a large, three-fold, decrease in the number of symmetric synapses with
Coral transcriptome and bacterial community profiles reveal distinct Yellow Band Disease states in Orbicella faveolata

KAUST Repository

Closek, Collin J.

2014-06-20

Coral diseases impact reefs globally. Although we continue to describe diseases, little is known about the etiology or progression of even the most common cases. To examine a spectrum of coral health and determine factors of disease progression we examined Orbicella faveolata exhibiting signs of Yellow Band Disease (YBD), a widespread condition in the Caribbean. We used a novel combined approach to assess three members of the coral holobiont: the coral-host, associated Symbiodinium algae, and bacteria. We profiled three conditions: (1) healthy-appearing colonies (HH), (2) healthy-appearing tissue on diseased colonies (HD), and (3) diseased lesion (DD). Restriction fragment length polymorphism analysis revealed health state-specific diversity in Symbiodinium clade associations. 16S ribosomal RNA gene microarrays (PhyloChips) and O. faveolata complimentary DNA microarrays revealed the bacterial community structure and host transcriptional response, respectively. A distinct bacterial community structure marked each health state. Diseased samples were associated with two to three times more bacterial diversity. HD samples had the highest bacterial richness, which included components associated with HH and DD, as well as additional unique families. The host transcriptome under YBD revealed a reduced cellular expression of defense- and metabolism-related processes, while the neighboring HD condition exhibited an intermediate expression profile. Although HD tissue appeared visibly healthy, the microbial communities and gene expression profiles were distinct. HD should be regarded as an additional (intermediate) state of disease, which is important for understanding the progression of YBD. © 2014 International Society for Microbial Ecology. All rights reserved.
Exposure of a distinct PDCA-1+ (CD317 B cell population to agonistic anti-4-1BB (CD137 inhibits T and B cell responses both in vitro and in vivo.

Directory of Open Access Journals (Sweden)

Dass S Vinay

Full Text Available 4-1BB (CD137 is an important T cell activating molecule. Here we report that it also promotes development of a distinct B cell subpopulation co-expressing PDCA-1. 4-1BB is expressed constitutively, and its expression is increased when PDCA-1(+ B cells are activated. We found that despite a high level of surface expression of 4-1BB on PDCA-1(+ B cells, treatment of these cells with agonistic anti-4-1BB mAb stimulated the expression of only a few activation markers (B7-2, MHC II, PD-L2, cytokines (IL-12p40/p70, and chemokines (MCP-1, RANTES, as well as sTNFR1, and the immunosuppressive enzyme, IDO. Although the PDCA-1(+ B cells stimulated by anti-4-1BB expressed MHC II at high levels and took up antigens efficiently, Ig class switching was inhibited when they were pulsed with T-independent (TI or T-dependent (TD Ags and adoptively transferred into syngeneic recipients. Furthermore, when anti-4-1BB-treated PDCA-1(+ B cells were pulsed with OVA peptide and combined with Vα2(+CD4(+ T cells, Ag-specific cell division was inhibited both in vitro and in vivo. Our findings suggest that the 4-1BB signal transforms PDCA-1(+ B cells into propagators of negative immune regulation, and establish an important role for 4-1BB in PDCA-1(+ B cell development and function.
Distinct epigenetic signatures elucidate enhancer-gene relationships that delineate CIMP and non-CIMP colorectal cancers.

Science.gov (United States)

Chong, Allen; Teo, Jing Xian; Ban, Kenneth H K

2016-05-10

Epigenetic changes, like DNA methylation, affect gene expression and in colorectal cancer (CRC), a distinct phenotype called the CpG island methylator phenotype ("CIMP") has significantly higher levels of DNA methylation at so-called "Type C loci" within the genome. We postulate that enhancer-gene pairs are coordinately controlled through DNA methylation in order to regulate the expression of key genes/biomarkers for a particular phenotype.Firstly, we found 24 experimentally-validated enhancers (VISTA enhancer browser) that contained statistically significant (FDR-adjusted q-value of CIMP versus non-CIMP CRCs. Of these, the methylation of 2 enhancers, 1702 and 1944, were found to be very well correlated with the methylation of the genes Wnt3A and IGDCC3, respectively, in two separate and independent datasets.We show for the first time that there are indeed distinct and dynamic changes in the methylation pattern of specific enhancer-gene pairs in CRCs. Such a coordinated epigenetic event could be indicative of an interaction between (1) enhancer 1702 and Wnt3A and (2) enhancer 1944 and IGDCC3. Moreover, our study shows that the methylation patterns of these 2 enhancer-gene pairs can potentially be used as biomarkers to delineate CIMP from non-CIMP CRCs.
Astrocyte-specific regulation of hMeCP2 expression in Drosophila

Directory of Open Access Journals (Sweden)

David L. Hess-Homeier

2014-10-01

Full Text Available Alterations in the expression of Methyl-CpG-binding protein 2 (MeCP2 either by mutations or gene duplication leads to a wide spectrum of neurodevelopmental disorders including Rett Syndrome and MeCP2 duplication disorder. Common features of Rett Syndrome (RTT, MeCP2 duplication disorder, and neuropsychiatric disorders indicate that even moderate changes in MeCP2 protein levels result in functional and structural cell abnormalities. In this study, we investigated two areas of MeCP2 pathophysiology using Drosophila as a model system: the effects of MeCP2 glial gain-of-function activity on circuits controlling sleep behavior, and the cell-type specific regulation of MeCP2 expression. In this study, we first examined the effects of elevated MeCP2 levels on microcircuits by expressing human MeCP2 (hMeCP2 in astrocytes and distinct subsets of amine neurons including dopamine and octopamine (OA neurons. Depending on the cell-type, hMeCP2 expression reduced sleep levels, altered daytime/nighttime sleep patterns, and generated sleep maintenance deficits. Second, we identified a 498 base pair region of the MeCP2e2 isoform that is targeted for regulation in distinct subsets of astrocytes. Levels of the full-length hMeCP2e2 and mutant RTT R106W protein decreased in astrocytes in a temporally and spatially regulated manner. In contrast, expression of the deletion Δ166 hMeCP2 protein was not altered in the entire astrocyte population. qPCR experiments revealed a reduction in full-length hMeCP2e2 transcript levels suggesting transgenic hMeCP2 expression is regulated at the transcriptional level. Given the phenotypic complexities that are caused by alterations in MeCP2 levels, our results provide insight into distinct cellular mechanisms that control MeCP2 expression and link microcircuit abnormalities with defined behavioral deficits.
Macrophage polarization alters the expression and sulfation pattern of glycosaminoglycans.

Science.gov (United States)

Martinez, Pierre; Denys, Agnès; Delos, Maxime; Sikora, Anne-Sophie; Carpentier, Mathieu; Julien, Sylvain; Pestel, Joël; Allain, Fabrice

2015-05-01

Macrophages are major cells of inflammatory process and take part in a large number of physiological and pathological processes. According to tissue environment, they can polarize into pro-inflammatory (M1) or alternative (M2) cells. Although many evidences have hinted to a potential role of cell-surface glycosaminoglycans (GAGs) in the functions of macrophages, the effect of M1 or M2 polarization on the biosynthesis of these polysaccharides has not been investigated so far. GAGs are composed of repeat sulfated disaccharide units. Heparan (HS) and chondroitin/dermatan sulfates (CS/DS) are the major GAGs expressed at the cell membrane. They are involved in numerous biological processes, which rely on their ability to selectively interact with a large panel of proteins. More than 20 genes encoding sulfotransferases have been implicated in HS and CS/DS biosynthesis, and the functional repertoire of HS and CS/DS has been related to the expression of these isoenzymes. In this study, we analyzed the expression of sulfotransferases as a response to macrophage polarization. We found that M1 and M2 activation drastically modified the profiles of expression of numerous HS and CS/DS sulfotransferases. This was accompanied by the expression of GAGs with distinct structural features. We then demonstrated that GAGs of M2 macrophages were efficient to present fibroblast growth factor-2 in an assay of tumor cell proliferation, thus indicating that changes in GAG structure may contribute to the functions of polarized macrophages. Altogether, our findings suggest a regulatory mechanism in which fine modifications in GAG biosynthesis may participate to the plasticity of macrophage functions. © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
Increased dysbindin-1B isoform expression in schizophrenia and its propensity in aggresome formation

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Xu, Yiliang; Sun, Yuhui; Ye, Haihong; Zhu, Li; Liu, Jianghong; Wu, Xiaofeng; Wang, Le; He, Tingting; Shen, Yan; Wu, Jane Y; Xu, Qi

2015-01-01

Genetic variations in the human dysbindin-1 gene (DTNBP1) have been associated with schizophrenia. As a result of alternative splicing, the human DTNBP1 gene generates at least three distinct protein isoforms, dysbindin-1A, -1B and -1C. Significant effort has focused on dysbindin-1A, an important player in multiple steps of neurodevelopment. However, the other isoforms, dysbindin-1B and dysbindin-1C have not been well characterized. Nor have been associated with human diseases. Here we report an increase in expression of DTNBP1b mRNA in patients with paranoid schizophrenia as compared with healthy controls. A single-nucleotide polymorphism located in intron 9, rs117610176, has been identified and associated with paranoid schizophrenia, and its C allele leads to an increase of DTNBP1b mRNA splicing. Our data show that different dysbindin splicing isoforms exhibit distinct subcellular distribution, suggesting their distinct functional activities. Dysbindin-1B forms aggresomes at the perinuclear region, whereas dysbindin-1A and -1C proteins exhibit diffused patterns in the cytoplasm. Dysbindin-1A interacts with dysbindin-1B, getting recruited to the aggresome structure when co-expressed with dysbindin-1B. Moreover, cortical neurons over-expressing dysbindin-1B show reduction in neurite outgrowth, suggesting that dysbindin-1B may interfere with dysbindin-1A function in a dominant-negative manner. Taken together, our study uncovers a previously unknown association of DTNBP1b expression with schizophrenia in addition to its distinct biochemical and functional properties. PMID:27462430
Imprinted Expression of SNRPN in Human Preimplantation Embryos

OpenAIRE

Huntriss, John; Daniels, Robert; Bolton, Virginia; Monk, Marilyn

1998-01-01

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are two clinically distinct neurogenetic disorders arising from a loss of expression of imprinted genes within the human chromosome region 15q11-q13. Recent evidence suggests that the SNRPN gene, which is defective in PWS, plays a central role in the imprinting-center regulation of the PWS/AS region. To increase our understanding of the regulation of expression of this imprinted gene, we have developed single-cell-sensitive procedures for...
Maximized Autotransporter-Mediated Expression (MATE for Surface Display and Secretion of Recombinant Proteins in Escherichia coli

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Shanna Sichwart

2015-01-01

Full Text Available A new optimized system for the surface display and secretion of recombinant proteins is described, termed MATE (maximized autotransporter-mediated expression. It is based on an artificial gene consisting of the coding region for the signal peptide of CtxB, a multiple cloning site for passenger gene insertion, flanked by coding sequences for linear epitopes for monoclonal antibodies and OmpT, and factor Xa protease cleavage sites followed by a codon-optimized DNA sequence of the linker and the β-barrel of the type V autotransporter EhaA from Escherichia coli under control of an IPTG-inducible T5 promoter. The MATE system enabled the continuous secretion of recombinant passenger mCherry via OmpT-mediated cleavage, using native OmpT protease activity in E. coli when grown at 37 °C. It is the first example to show that native OmpT activity is sufficient to facilitate the secretion of a correctly folded target protein in preparative amounts obtaining 240 μg of purified mCherry from 800 mL of crude culture supernatant. Because the release of mCherry was achieved by a simple transfer of the encoding plasmid from an OmpT-negative to an OmpT-positive strain, it bears the option to use surface display for screening purposes and secretion for production of the selected variant. A single plasmid could therefore be used for continuous secretion in OmpT-positive strains or surface display in OmpT-negative strains. In conclusion, the MATE system appears to be a versatile tool for the surface display and for the secretion of target proteins in E. coli.
Anxa4 Genes are Expressed in Distinct Organ Systems in Xenopus laevis and tropicalis But are Functionally Conserved

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Massé, Karine L; Collins, Robert J; Bhamra, Surinder; Seville, Rachel A

2007-01-01

Anxa4 belongs to the multigenic annexin family of proteins which are characterized by their ability to interact with membranes in a calcium-dependent manner. Defined as a marker for polarized epithelial cells, Anxa4 is believed to be involved in many cellular processes but its functions in vivo are still poorly understood. Previously, we cloned Xanx4 in Xenopus laevis (now referred to as anxa4a) and demonstrated its role during organogenesis of the pronephros, providing the first evidence of a specific function for this protein during the development of a vertebrate. Here, we describe the strict conservation of protein sequence and functional domains of anxa4 during vertebrate evolution. We also identify the paralog of anxa4a, anxa4b and show its specific temporal and spatial expression pattern is different from anxa4a. We show that anxa4 orthologs in X. laevis and tropicalis display expression domains in different organ systems. Whilst the anxa4a gene is mainly expressed in the kidney, Xt anxa4 is expressed in the liver. Finally, we demonstrate Xt anxa4 and anxa4a can display conserved function during kidney organogenesis, despite the fact that Xt anxa4 transcripts are not expressed in this domain. This study highlights the divergence of expression of homologous genes during Xenopus evolution and raises the potential problems of using X. tropicalis promoters in X. laevis. PMID:19279706
Albinism in phylogenetically and geographically distinct populations of Astyanax cavefish arises through the same loss-of-function Oca2 allele

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Gross, J B; Wilkens, H

2013-01-01

The Mexican tetra, Astyanax mexicanus, comprises 29 populations of cave-adapted fish distributed across a vast karst region in northeastern Mexico. These populations have a complex evolutionary history, having descended from ‘old' and ‘young' ancestral surface-dwelling stocks that invaded the region ∼6.7 and ∼2.8 MYa, respectively. This study investigates a set of captive, pigmented Astyanax cavefish collected from the Micos cave locality in 1970, in which albinism appeared over the past two decades. We combined novel coloration analyses, coding sequence comparisons and mRNA expression level studies to investigate the origin of albinism in captive-bred Micos cavefish. We discovered that albino Micos cavefish harbor two copies of a loss-of-function ocular and cutaneous albinism type II (Oca2) allele previously identified in the geographically distant Pachón cave population. This result suggests that phylogenetically young Micos cavefish and phylogenetically old Pachón cave fish inherited this Oca2 allele from the ancestral surface-dwelling taxon. This likely resulted from the presence of the loss-of-function Oca2 haplotype in the ‘young' ancestral surface-dwelling stock that colonized the Micos cave and also introgressed into the ancient Pachón cave population. The appearance of albinism in captive Micos cavefish, caused by the same loss-of-function allele present in Pachón cavefish, implies that geographically and phylogenetically distinct cave populations can evolve the same troglomorphic phenotype from standing genetic variation present in the ancestral taxon. PMID:23572122
Rat Models of Cardiovascular Disease Demonstrate Distinctive Pulmonary Gene Expressions for Vascular Response Genes: Impact of Ozone Exposure

Science.gov (United States)

Comparative gene expression profiling of multiple tissues from rat strains with genetic predisposition to diverse cardiovascular diseases (CVD) can help decode the transcriptional program that governs organ-specific functions. We examined expressions of CVD genes in the lungs of ...

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