WorldWideScience

Sample records for cell type-specific investigation

  1. Cell Type-Specific Contributions to the TSC Neuropathology

    2017-08-01

    AWARD NUMBER: W81XWH-16-1-0415 TITLE: Cell Type-Specific Contributions to the TSC Neuropathology PRINCIPAL INVESTIGATOR: Gabriella D’Arcangelo...AND SUBTITLE Cell Type-Specific Contributions to the TSC Neuropathology 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-16-1-0415 5c. PROGRAM...how heterozygous and homozygous Tsc2 mutations affect the development of mutant excitatory neurons as well as other surrounding brain cells , in vivo

  2. Cardiac Glycoside Glucoevatromonoside Induces Cancer Type-Specific Cell Death

    Naira F. Z. Schneider

    2018-03-01

    Full Text Available Cardiac glycosides (CGs are natural compounds used traditionally to treat congestive heart diseases. Recent investigations repositioned CGs as potential anticancer agents. To discover novel cytotoxic CG scaffolds, we selected the cardenolide glucoevatromonoside (GEV out of 46 CGs for its low nanomolar anti-lung cancer activity. GEV presented reduced toxicity toward non-cancerous cell types (lung MRC-5 and PBMC and high-affinity binding to the Na+/K+-ATPase α subunit, assessed by computational docking. GEV-induced cell death was caspase-independent, as investigated by a multiparametric approach, and culminates in severe morphological alterations in A549 cells, monitored by transmission electron microscopy, live cell imaging and flow cytometry. This non-canonical cell death was not preceded or accompanied by exacerbation of autophagy. In the presence of GEV, markers of autophagic flux (e.g. LC3I-II conversion were impacted, even in presence of bafilomycin A1. Cell death induction remained unaffected by calpain, cathepsin, parthanatos, or necroptosis inhibitors. Interestingly, GEV triggered caspase-dependent apoptosis in U937 acute myeloid leukemia cells, witnessing cancer-type specific cell death induction. Differential cell cycle modulation by this CG led to a G2/M arrest, cyclin B1 and p53 downregulation in A549, but not in U937 cells. We further extended the anti-cancer potential of GEV to 3D cell culture using clonogenic and spheroid formation assays and validated our findings in vivo by zebrafish xenografts. Altogether, GEV shows an interesting anticancer profile with the ability to exert cytotoxic effects via induction of different cell death modalities.

  3. Cell-Type-Specific Splicing of Piezo2 Regulates Mechanotransduction

    Marcin Szczot

    2017-12-01

    Full Text Available Summary: Piezo2 is a mechanically activated ion channel required for touch discrimination, vibration detection, and proprioception. Here, we discovered that Piezo2 is extensively spliced, producing different Piezo2 isoforms with distinct properties. Sensory neurons from both mice and humans express a large repertoire of Piezo2 variants, whereas non-neuronal tissues express predominantly a single isoform. Notably, even within sensory ganglia, we demonstrate the splicing of Piezo2 to be cell type specific. Biophysical characterization revealed substantial differences in ion permeability, sensitivity to calcium modulation, and inactivation kinetics among Piezo2 splice variants. Together, our results describe, at the molecular level, a potential mechanism by which transduction is tuned, permitting the detection of a variety of mechanosensory stimuli. : Szczot et al. find that the mechanoreceptor Piezo2 is extensively alternatively spliced, generating multiple distinct isoforms. Their findings indicate that these splice products have specific tissue and cell type expression patterns and exhibit differences in receptor properties. Keywords: Piezo, touch, sensation, ion-channel, splicing

  4. Freedom of expression: cell-type-specific gene profiling.

    Otsuki, Leo; Cheetham, Seth W; Brand, Andrea H

    2014-01-01

    Cell fate and behavior are results of differential gene regulation, making techniques to profile gene expression in specific cell types highly desirable. Many methods now enable investigation at the DNA, RNA and protein level. This review introduces the most recent and popular techniques, and discusses key issues influencing the choice between these such as ease, cost and applicability of information gained. Interdisciplinary collaborations will no doubt contribute further advances, including not just in single cell type but single-cell expression profiling. © 2014 Wiley Periodicals, Inc.

  5. Cell type-specific neuroprotective activity of untranslocated prion protein.

    Elena Restelli

    2010-10-01

    Full Text Available A key pathogenic role in prion diseases was proposed for a cytosolic form of the prion protein (PrP. However, it is not clear how cytosolic PrP localization influences neuronal viability, with either cytotoxic or anti-apoptotic effects reported in different studies. The cellular mechanism by which PrP is delivered to the cytosol of neurons is also debated, and either retrograde transport from the endoplasmic reticulum or inefficient translocation during biosynthesis has been proposed. We investigated cytosolic PrP biogenesis and effect on cell viability in primary neuronal cultures from different mouse brain regions.Mild proteasome inhibition induced accumulation of an untranslocated form of cytosolic PrP in cortical and hippocampal cells, but not in cerebellar granules. A cyclopeptolide that interferes with the correct insertion of the PrP signal sequence into the translocon increased the amount of untranslocated PrP in cortical and hippocampal cells, and induced its synthesis in cerebellar neurons. Untranslocated PrP boosted the resistance of cortical and hippocampal neurons to apoptotic insults but had no effect on cerebellar cells.These results indicate cell type-dependent differences in the efficiency of PrP translocation, and argue that cytosolic PrP targeting might serve a physiological neuroprotective function.

  6. ERE environment- and cell type-specific transcriptional effects of estrogen in normal endometrial cells.

    Lascombe, I; Sallot, M; Vuillermoz, C; Weisz, A; Adessi, G L; Jouvenot, M

    1998-04-30

    Our previous results have suggested a repression of E2 (17beta-estradiol) effect on the c-fos gene of cultured guinea-pig endometrial cells. To investigate this repression, the expression of three human c-fos gene recombinants, pFC1-BL (-2250/+41), pFC2-BL (-1400/+41) and pFC2E (-1300/-1050 and -230/+41), known to be E2-responsive in Hela cells, was studied in stromal (SC) and glandular epithelial cells (GEC). In both cellular types, pFC1-BL was not induced by E2, even in the presence of growth factors or co-transfected estrogen receptor. The pattern of pFC2-BL and pFC2E expression was strikingly different and depended on the cellular type: pFC2-BL and pFC2E induction was restricted to the glandular epithelial cells and did not occur in the SCs. We argue for a repression of E2 action which is dependent on the estrogen-responsive cis-acting element (ERE) environment and also cell type-specific involving DNA/protein and/or protein/protein interactions with cellular type-specific factors.

  7. Cell type-specific responses to salinity - the epidermal bladder cell transcriptome of Mesembryanthemum crystallinum.

    Oh, Dong-Ha; Barkla, Bronwyn J; Vera-Estrella, Rosario; Pantoja, Omar; Lee, Sang-Yeol; Bohnert, Hans J; Dassanayake, Maheshi

    2015-08-01

    Mesembryanthemum crystallinum (ice plant) exhibits extreme tolerance to salt. Epidermal bladder cells (EBCs), developing on the surface of aerial tissues and specialized in sodium sequestration and other protective functions, are critical for the plant's stress adaptation. We present the first transcriptome analysis of EBCs isolated from intact plants, to investigate cell type-specific responses during plant salt adaptation. We developed a de novo assembled, nonredundant EBC reference transcriptome. Using RNAseq, we compared the expression patterns of the EBC-specific transcriptome between control and salt-treated plants. The EBC reference transcriptome consists of 37 341 transcript-contigs, of which 7% showed significantly different expression between salt-treated and control samples. We identified significant changes in ion transport, metabolism related to energy generation and osmolyte accumulation, stress signalling, and organelle functions, as well as a number of lineage-specific genes of unknown function, in response to salt treatment. The salinity-induced EBC transcriptome includes active transcript clusters, refuting the view of EBCs as passive storage compartments in the whole-plant stress response. EBC transcriptomes, differing from those of whole plants or leaf tissue, exemplify the importance of cell type-specific resolution in understanding stress adaptive mechanisms. No claim to original US government works. New Phytologist © 2015 New Phytologist Trust.

  8. Cell-type specific four-component hydrogel.

    Timo Aberle

    Full Text Available In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin to generate a blend (technical term: quattroGel, an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appropriate for general cell adhesion, and restricted diffusion. Cell proliferation of endothelial cells, chondrocytes and fibroblasts was essentially unaffected. In contrast, on quattroGels neither endothelial cells formed vascular tubes nor did primary neurons extend neurites in significant amounts. Only chondrocytes differentiated properly as judged by collagen isoform expression. The biophysical quattroGel characteristics appeared to leave distinct cell processes such as mitosis unaffected and favored differentiation of sessile cells, but hampered differentiation of migratory cells. This cell-type selectivity is of interest e.g. during articular cartilage or invertebral disc repair, where pathological innervation and angiogenesis represent adverse events in tissue engineering.

  9. Cell type-specific suppression of mechanosensitive genes by audible sound stimulation.

    Kumeta, Masahiro; Takahashi, Daiji; Takeyasu, Kunio; Yoshimura, Shige H

    2018-01-01

    Audible sound is a ubiquitous environmental factor in nature that transmits oscillatory compressional pressure through the substances. To investigate the property of the sound as a mechanical stimulus for cells, an experimental system was set up using 94.0 dB sound which transmits approximately 10 mPa pressure to the cultured cells. Based on research on mechanotransduction and ultrasound effects on cells, gene responses to the audible sound stimulation were analyzed by varying several sound parameters: frequency, wave form, composition, and exposure time. Real-time quantitative PCR analyses revealed a distinct suppressive effect for several mechanosensitive and ultrasound-sensitive genes that were triggered by sounds. The effect was clearly observed in a wave form- and pressure level-specific manner, rather than the frequency, and persisted for several hours. At least two mechanisms are likely to be involved in this sound response: transcriptional control and RNA degradation. ST2 stromal cells and C2C12 myoblasts exhibited a robust response, whereas NIH3T3 cells were partially and NB2a neuroblastoma cells were completely insensitive, suggesting a cell type-specific response to sound. These findings reveal a cell-level systematic response to audible sound and uncover novel relationships between life and sound.

  10. Cell-type-specific gene delivery into neuronal cells in vitro and in vivo

    Parveen, Zahida; Mukhtar, Muhammad; Rafi, Mohammed; Wenger, David A.; Siddiqui, Khwaja M.; Siler, Catherine A.; Dietzschold, Bernhard; Pomerantz, Roger J.; Schnell, Matthias J.; Dornburg, Ralph

    2003-01-01

    The avian retroviruses reticuloendotheliosis virus strain A (REV-A) and spleen necrosis virus (SNV) are not naturally infectious in human cells. However, REV-A-derived viral vectors efficiently infect human cells when they are pseudotyped with envelope proteins displaying targeting ligands specific for human cell-surface receptors. Here we report that vectors containing the gag region of REV-A and pol of SNV can be pseudotyped with the envelope protein of vesicular stomatitis virus (VSV) and the glycoproteins of different rabies virus (RV) strains. Vectors pseudotyped with the envelope protein of the highly neurotropic RV strain CVS-N2c facilitated cell type-specific gene delivery into mouse and human neurons, but did not infect other human cell types. Moreover, when such vector particles were injected into the brain of newborn mice, only neuronal cells were infected in vivo. Cell-type-specific gene delivery into neurons may present quite specific gene therapy approaches for many degenerative diseases of the brain

  11. Modification of surface/neuron interfaces for neural cell-type specific responses: a review

    Chen, Cen; Kong, Xiangdong; Lee, In-Seop

    2016-01-01

    Surface/neuron interfaces have played an important role in neural repair including neural prostheses and tissue engineered scaffolds. This comprehensive literature review covers recent studies on the modification of surface/neuron interfaces. These interfaces are identified in cases both where the surfaces of substrates or scaffolds were in direct contact with cells and where the surfaces were modified to facilitate cell adhesion and controlling cell-type specific responses. Different sources of cells for neural repair are described, such as pheochromocytoma neuronal-like cell, neural stem cell (NSC), embryonic stem cell (ESC), mesenchymal stem cell (MSC) and induced pluripotent stem cell (iPS). Commonly modified methods are discussed including patterned surfaces at micro- or nano-scale, surface modification with conducting coatings, and functionalized surfaces with immobilized bioactive molecules. These approaches to control cell-type specific responses have enormous potential implications in neural repair. (paper)

  12. Long-term in vitro, cell-type-specific genome-wide reprogramming of gene expression

    Hakelien, Anne-Mari; Gaustad, Kristine G.; Taranger, Christel K.; Skalhegg, Bjorn S.; Kuentziger, Thomas; Collas, Philippe

    2005-01-01

    We demonstrate a cell extract-based, genome-wide and heritable reprogramming of gene expression in vitro. Kidney epithelial 293T cells have previously been shown to take on T cell properties following a brief treatment with an extract of Jurkat T cells. We show here that 293T cells exposed for 1 h to a Jurkat cell extract undergo genome-wide, target cell-type-specific and long-lasting transcriptional changes. Microarray analyses indicate that on any given week after extract treatment, ∼2500 genes are upregulated >3-fold, of which ∼900 are also expressed in Jurkat cells. Concomitantly, ∼1500 genes are downregulated or repressed, of which ∼500 are also downregulated in Jurkat cells. Gene expression changes persist for over 30 passages (∼80 population doublings) in culture. Target cell-type specificity of these changes is shown by the lack of activation or repression of Jurkat-specific genes by extracts of 293T cells or carcinoma cells. Quantitative RT-PCR analysis confirms the long-term transcriptional activation of genes involved in key T cell functions. Additionally, growth of cells in suspended aggregates, expression of CD3 and CD28 T cell surface markers, and interleukin-2 secretion by 293T cells treated with extract of adult peripheral blood T cells illustrate a functional nuclear reprogramming. Therefore, target cell-type-specific and heritable changes in gene expression, and alterations in cell function, can be promoted by extracts derived from transformed cells as well as from adult primary cells

  13. Cell-type specific roles for PTEN in establishing a functional retinal architecture.

    Robert Cantrup

    Full Text Available The retina has a unique three-dimensional architecture, the precise organization of which allows for complete sampling of the visual field. Along the radial or apicobasal axis, retinal neurons and their dendritic and axonal arbors are segregated into layers, while perpendicular to this axis, in the tangential plane, four of the six neuronal types form patterned cellular arrays, or mosaics. Currently, the molecular cues that control retinal cell positioning are not well-understood, especially those that operate in the tangential plane. Here we investigated the role of the PTEN phosphatase in establishing a functional retinal architecture.In the developing retina, PTEN was localized preferentially to ganglion, amacrine and horizontal cells, whose somata are distributed in mosaic patterns in the tangential plane. Generation of a retina-specific Pten knock-out resulted in retinal ganglion, amacrine and horizontal cell hypertrophy, and expansion of the inner plexiform layer. The spacing of Pten mutant mosaic populations was also aberrant, as were the arborization and fasciculation patterns of their processes, displaying cell type-specific defects in the radial and tangential dimensions. Irregular oscillatory potentials were also observed in Pten mutant electroretinograms, indicative of asynchronous amacrine cell firing. Furthermore, while Pten mutant RGC axons targeted appropriate brain regions, optokinetic spatial acuity was reduced in Pten mutant animals. Finally, while some features of the Pten mutant retina appeared similar to those reported in Dscam-mutant mice, PTEN expression and activity were normal in the absence of Dscam.We conclude that Pten regulates somal positioning and neurite arborization patterns of a subset of retinal cells that form mosaics, likely functioning independently of Dscam, at least during the embryonic period. Our findings thus reveal an unexpected level of cellular specificity for the multi-purpose phosphatase, and

  14. Bridging the Gap: Towards a Cell-Type Specific Understanding of Neural Circuits Underlying Fear Behaviors

    McCullough, KM; Morrison, FG; Ressler, KJ

    2016-01-01

    Fear and anxiety-related disorders are remarkably common and debilitating, and are often characterized by dysregulated fear responses. Rodent models of fear learning and memory have taken great strides towards elucidating the specific neuronal circuitries underlying the learning of fear responses. The present review addresses recent research utilizing optogenetic approaches to parse circuitries underlying fear behaviors. It also highlights the powerful advances made when optogenetic techniques are utilized in a genetically defined, cell-type specific, manner. The application of next-generation genetic and sequencing approaches in a cell-type specific context will be essential for a mechanistic understanding of the neural circuitry underlying fear behavior and for the rational design of targeted, circuit specific, pharmacologic interventions for the treatment and prevention of fear-related disorders. PMID:27470092

  15. Common themes and cell type specific variations of higher order chromatin arrangements in the mouse

    Cremer Thomas

    2005-12-01

    Full Text Available Abstract Background Similarities as well as differences in higher order chromatin arrangements of human cell types were previously reported. For an evolutionary comparison, we now studied the arrangements of chromosome territories and centromere regions in six mouse cell types (lymphocytes, embryonic stem cells, macrophages, fibroblasts, myoblasts and myotubes with fluorescence in situ hybridization and confocal laser scanning microscopy. Both species evolved pronounced differences in karyotypes after their last common ancestors lived about 87 million years ago and thus seem particularly suited to elucidate common and cell type specific themes of higher order chromatin arrangements in mammals. Results All mouse cell types showed non-random correlations of radial chromosome territory positions with gene density as well as with chromosome size. The distribution of chromosome territories and pericentromeric heterochromatin changed during differentiation, leading to distinct cell type specific distribution patterns. We exclude a strict dependence of these differences on nuclear shape. Positional differences in mouse cell nuclei were less pronounced compared to human cell nuclei in agreement with smaller differences in chromosome size and gene density. Notably, the position of chromosome territories relative to each other was very variable. Conclusion Chromosome territory arrangements according to chromosome size and gene density provide common, evolutionary conserved themes in both, human and mouse cell types. Our findings are incompatible with a previously reported model of parental genome separation.

  16. Cell-type-specific expression of NFIX in the developing and adult cerebellum.

    Fraser, James; Essebier, Alexandra; Gronostajski, Richard M; Boden, Mikael; Wainwright, Brandon J; Harvey, Tracey J; Piper, Michael

    2017-07-01

    Transcription factors from the nuclear factor one (NFI) family have been shown to play a central role in regulating neural progenitor cell differentiation within the embryonic and post-natal brain. NFIA and NFIB, for instance, promote the differentiation and functional maturation of granule neurons within the cerebellum. Mice lacking Nfix exhibit delays in the development of neuronal and glial lineages within the cerebellum, but the cell-type-specific expression of this transcription factor remains undefined. Here, we examined the expression of NFIX, together with various cell-type-specific markers, within the developing and adult cerebellum using both chromogenic immunohistochemistry and co-immunofluorescence labelling and confocal microscopy. In embryos, NFIX was expressed by progenitor cells within the rhombic lip and ventricular zone. After birth, progenitor cells within the external granule layer, as well as migrating and mature granule neurons, expressed NFIX. Within the adult cerebellum, NFIX displayed a broad expression profile, and was evident within granule cells, Bergmann glia, and interneurons, but not within Purkinje neurons. Furthermore, transcriptomic profiling of cerebellar granule neuron progenitor cells showed that multiple splice variants of Nfix are expressed within this germinal zone of the post-natal brain. Collectively, these data suggest that NFIX plays a role in regulating progenitor cell biology within the embryonic and post-natal cerebellum, as well as an ongoing role within multiple neuronal and glial populations within the adult cerebellum.

  17. Cell-type-specific role for nucleus accumbens neuroligin-2 in depression and stress susceptibility.

    Heshmati, Mitra; Aleyasin, Hossein; Menard, Caroline; Christoffel, Daniel J; Flanigan, Meghan E; Pfau, Madeline L; Hodes, Georgia E; Lepack, Ashley E; Bicks, Lucy K; Takahashi, Aki; Chandra, Ramesh; Turecki, Gustavo; Lobo, Mary Kay; Maze, Ian; Golden, Sam A; Russo, Scott J

    2018-01-30

    Behavioral coping strategies are critical for active resilience to stress and depression; here we describe a role for neuroligin-2 (NLGN-2) in the nucleus accumbens (NAc). Neuroligins (NLGN) are a family of neuronal postsynaptic cell adhesion proteins that are constituents of the excitatory and inhibitory synapse. Importantly, NLGN-3 and NLGN-4 mutations are strongly implicated as candidates underlying the development of neuropsychiatric disorders with social disturbances such as autism, but the role of NLGN-2 in neuropsychiatric disease states is unclear. Here we show a reduction in NLGN-2 gene expression in the NAc of patients with major depressive disorder. Chronic social defeat stress in mice also decreases NLGN-2 selectively in dopamine D1-positive cells, but not dopamine D2-positive cells, within the NAc of stress-susceptible mice. Functional NLGN-2 knockdown produces bidirectional, cell-type-specific effects: knockdown in dopamine D1-positive cells promotes subordination and stress susceptibility, whereas knockdown in dopamine D2-positive cells mediates active defensive behavior. These findings establish a behavioral role for NAc NLGN-2 in stress and depression; provide a basis for targeted, cell-type specific therapy; and highlight the role of active behavioral coping mechanisms in stress susceptibility.

  18. Layer- and Cell Type-Specific Modulation of Excitatory Neuronal Activity in the Neocortex

    Gabriele Radnikow

    2018-01-01

    Full Text Available From an anatomical point of view the neocortex is subdivided into up to six layers depending on the cortical area. This subdivision has been described already by Meynert and Brodmann in the late 19/early 20. century and is mainly based on cytoarchitectonic features such as the size and location of the pyramidal cell bodies. Hence, cortical lamination is originally an anatomical concept based on the distribution of excitatory neuron. However, it has become apparent in recent years that apart from the layer-specific differences in morphological features, many functional properties of neurons are also dependent on cortical layer or cell type. Such functional differences include changes in neuronal excitability and synaptic activity by neuromodulatory transmitters. Many of these neuromodulators are released from axonal afferents from subcortical brain regions while others are released intrinsically. In this review we aim to describe layer- and cell-type specific differences in the effects of neuromodulator receptors in excitatory neurons in layers 2–6 of different cortical areas. We will focus on the neuromodulator systems using adenosine, acetylcholine, dopamine, and orexin/hypocretin as examples because these neuromodulator systems show important differences in receptor type and distribution, mode of release and functional mechanisms and effects. We try to summarize how layer- and cell type-specific neuromodulation may affect synaptic signaling in cortical microcircuits.

  19. Systematic and Cell Type-Specific Telomere Length Changes in Subsets of Lymphocytes

    Jue Lin

    2016-01-01

    Full Text Available Telomeres, the protective DNA-protein complexes at the ends of linear chromosomes, are important for genome stability. Leukocyte or peripheral blood mononuclear cell (PBMC telomere length is a potential biomarker for human aging that integrates genetic, environmental, and lifestyle factors and is associated with mortality and risks for major diseases. However, only a limited number of studies have examined longitudinal changes of telomere length and few have reported data on sorted circulating immune cells. We examined the average telomere length (TL in CD4+, CD8+CD28+, and CD8+CD28− T cells, B cells, and PBMCs, cross-sectionally and longitudinally, in a cohort of premenopausal women. We report that TL changes over 18 months were correlated among these three T cell types within the same participant. Additionally, PBMC TL change was also correlated with those of all three T cell types, and B cells. The rate of shortening for B cells was significantly greater than for the three T cell types. CD8+CD28− cells, despite having the shortest TL, showed significantly more rapid attrition when compared to CD8+CD28+ T cells. These results suggest systematically coordinated, yet cell type-specific responses to factors and pathways contribute to telomere length regulation.

  20. Digital sorting of complex tissues for cell type-specific gene expression profiles.

    Zhong, Yi; Wan, Ying-Wooi; Pang, Kaifang; Chow, Lionel M L; Liu, Zhandong

    2013-03-07

    Cellular heterogeneity is present in almost all gene expression profiles. However, transcriptome analysis of tissue specimens often ignores the cellular heterogeneity present in these samples. Standard deconvolution algorithms require prior knowledge of the cell type frequencies within a tissue or their in vitro expression profiles. Furthermore, these algorithms tend to report biased estimations. Here, we describe a Digital Sorting Algorithm (DSA) for extracting cell-type specific gene expression profiles from mixed tissue samples that is unbiased and does not require prior knowledge of cell type frequencies. The results suggest that DSA is a specific and sensitivity algorithm in gene expression profile deconvolution and will be useful in studying individual cell types of complex tissues.

  1. Cell type-specific characterization of nuclear DNA contents within complex tissues and organs

    Lambert Georgina M

    2005-10-01

    Full Text Available Abstract Background Eukaryotic organisms are defined by the presence of a nucleus, which encloses the chromosomal DNA, and is characterized by its DNA content (C-value. Complex eukaryotic organisms contain organs and tissues that comprise interspersions of different cell types, within which polysomaty, endoreduplication, and cell cycle arrest is frequently observed. Little is known about the distribution of C-values across different cell types within these organs and tissues. Results We have developed, and describe here, a method to precisely define the C-value status within any specific cell type within complex organs and tissues of plants. We illustrate the application of this method to Arabidopsis thaliana, specifically focusing on the different cell types found within the root. Conclusion The method accurately and conveniently charts C-value within specific cell types, and provides novel insight into developmental processes. The method is, in principle, applicable to any transformable organism, including mammals, within which cell type specificity of regulation of endoreduplication, of polysomaty, and of cell cycle arrest is suspected.

  2. Cell type-specific translational repression of Cyclin B during meiosis in males.

    Baker, Catherine Craig; Gim, Byung Soo; Fuller, Margaret T

    2015-10-01

    The unique cell cycle dynamics of meiosis are controlled by layers of regulation imposed on core mitotic cell cycle machinery components by the program of germ cell development. Although the mechanisms that regulate Cdk1/Cyclin B activity in meiosis in oocytes have been well studied, little is known about the trans-acting factors responsible for developmental control of these factors in male gametogenesis. During meiotic prophase in Drosophila males, transcript for the core cell cycle protein Cyclin B1 (CycB) is expressed in spermatocytes, but the protein does not accumulate in spermatocytes until just before the meiotic divisions. Here, we show that two interacting proteins, Rbp4 and Fest, expressed at the onset of spermatocyte differentiation under control of the developmental program of male gametogenesis, function to direct cell type- and stage-specific repression of translation of the core G2/M cell cycle component cycB during the specialized cell cycle of male meiosis. Binding of Fest to Rbp4 requires a 31-amino acid region within Rbp4. Rbp4 and Fest are required for translational repression of cycB in immature spermatocytes, with Rbp4 binding sequences in a cell type-specific shortened form of the cycB 3' UTR. Finally, we show that Fest is required for proper execution of meiosis I. © 2015. Published by The Company of Biologists Ltd.

  3. The female gametophyte: an emerging model for cell type-specific systems biology in plant development

    Marc William Schmid

    2015-11-01

    Full Text Available Systems biology, a holistic approach describing a system emerging from the interactions of its molecular components, critically depends on accurate qualitative determination and quantitative measurements of these components. Development and improvement of large-scale profiling methods (omics now facilitates comprehensive measurements of many relevant molecules. For multicellular organisms, such as animals, fungi, algae, and plants, the complexity of the system is augmented by the presence of specialized cell types and organs, and a complex interplay within and between them. Cell type-specific analyses are therefore crucial for the understanding of developmental processes and environmental responses. This review first gives an overview of current methods used for large-scale profiling of specific cell types exemplified by recent advances in plant biology. The focus then lies on suitable model systems to study plant development and cell type specification. We introduce the female gametophyte of flowering plants as an ideal model to study fundamental developmental processes. Moreover, the female reproductive lineage is of importance for the emergence of evolutionary novelties such as an unequal parental contribution to the tissue nurturing the embryo or the clonal production of seeds by asexual reproduction (apomixis. Understanding these processes is not only interesting from a developmental or evolutionary perspective, but bears great potential for further crop improvement and the simplification of breeding efforts. We finally highlight novel methods, which are already available or which will likely soon facilitate large-scale profiling of the specific cell types of the female gametophyte in both model and non-model species. We conclude that it may take only few years until an evolutionary systems biology approach toward female gametogenesis may decipher some of its biologically most interesting and economically most valuable processes.

  4. Cell Type-Specific Chromatin Signatures Underline Regulatory DNA Elements in Human Induced Pluripotent Stem Cells and Somatic Cells.

    Zhao, Ming-Tao; Shao, Ning-Yi; Hu, Shijun; Ma, Ning; Srinivasan, Rajini; Jahanbani, Fereshteh; Lee, Jaecheol; Zhang, Sophia L; Snyder, Michael P; Wu, Joseph C

    2017-11-10

    Regulatory DNA elements in the human genome play important roles in determining the transcriptional abundance and spatiotemporal gene expression during embryonic heart development and somatic cell reprogramming. It is not well known how chromatin marks in regulatory DNA elements are modulated to establish cell type-specific gene expression in the human heart. We aimed to decipher the cell type-specific epigenetic signatures in regulatory DNA elements and how they modulate heart-specific gene expression. We profiled genome-wide transcriptional activity and a variety of epigenetic marks in the regulatory DNA elements using massive RNA-seq (n=12) and ChIP-seq (chromatin immunoprecipitation combined with high-throughput sequencing; n=84) in human endothelial cells (CD31 + CD144 + ), cardiac progenitor cells (Sca-1 + ), fibroblasts (DDR2 + ), and their respective induced pluripotent stem cells. We uncovered 2 classes of regulatory DNA elements: class I was identified with ubiquitous enhancer (H3K4me1) and promoter (H3K4me3) marks in all cell types, whereas class II was enriched with H3K4me1 and H3K4me3 in a cell type-specific manner. Both class I and class II regulatory elements exhibited stimulatory roles in nearby gene expression in a given cell type. However, class I promoters displayed more dominant regulatory effects on transcriptional abundance regardless of distal enhancers. Transcription factor network analysis indicated that human induced pluripotent stem cells and somatic cells from the heart selected their preferential regulatory elements to maintain cell type-specific gene expression. In addition, we validated the function of these enhancer elements in transgenic mouse embryos and human cells and identified a few enhancers that could possibly regulate the cardiac-specific gene expression. Given that a large number of genetic variants associated with human diseases are located in regulatory DNA elements, our study provides valuable resources for deciphering

  5. Cell-type-specific roles for COX-2 in UVB-induced skin cancer

    Herschman, Harvey

    2014-01-01

    In human tumors, and in mouse models, cyclooxygenase-2 (COX-2) levels are frequently correlated with tumor development/burden. In addition to intrinsic tumor cell expression, COX-2 is often present in fibroblasts, myofibroblasts and endothelial cells of the tumor microenvironment, and in infiltrating immune cells. Intrinsic cancer cell COX-2 expression is postulated as only one of many sources for prostanoids required for tumor promotion/progression. Although both COX-2 inhibition and global Cox-2 gene deletion ameliorate ultraviolet B (UVB)-induced SKH-1 mouse skin tumorigenesis, neither manipulation can elucidate the cell type(s) in which COX-2 expression is required for tumorigenesis; both eliminate COX-2 activity in all cells. To address this question, we created Cox-2 flox/flox mice, in which the Cox-2 gene can be eliminated in a cell-type-specific fashion by targeted Cre recombinase expression. Cox-2 deletion in skin epithelial cells of SKH-1 Cox-2 flox/flox;K14Cre + mice resulted, following UVB irradiation, in reduced skin hyperplasia and increased apoptosis. Targeted epithelial cell Cox-2 deletion also resulted in reduced tumor incidence, frequency, size and proliferation rate, altered tumor cell differentiation and reduced tumor vascularization. Moreover, Cox-2 flox/flox;K14Cre + papillomas did not progress to squamous cell carcinomas. In contrast, Cox-2 deletion in SKH-1 Cox-2 flox/flox; LysMCre + myeloid cells had no effect on UVB tumor induction. We conclude that (i) intrinsic epithelial COX-2 activity plays a major role in UVB-induced skin cancer, (ii) macrophage/myeloid COX-2 plays no role in UVB-induced skin cancer and (iii) either there may be another COX-2-dependent prostanoid source(s) that drives UVB skin tumor induction or there may exist a COX-2-independent pathway(s) to UVB-induced skin cancer. PMID:24469308

  6. PINK1 is required for timely cell-type specific mitochondrial clearance during Drosophila midgut metamorphosis.

    Liu, Yan; Lin, Jingjing; Zhang, Minjie; Chen, Kai; Yang, Shengxi; Wang, Qun; Yang, Hongqin; Xie, Shusen; Zhou, Yongjian; Zhang, Xi; Chen, Fei; Yang, Yufeng

    2016-11-15

    Mitophagy is the selective degradation of mitochondria by autophagy, which is an important mitochondrial quality and quantity control process. During Drosophila metamorphosis, the degradation of midgut involves a large change in length and organization, which is mediated by autophagy. Here we noticed a cell-type specific mitochondrial clearance process that occurs in enterocytes (ECs), while most mitochondria remain in intestinal stem cells (ISCs) during metamorphosis. Although PINK1/PARKIN represent the canonical pathway for the elimination of impaired mitochondria in varied pathological conditions, their roles in developmental processes or normal physiological conditions have been less studied. To examine the potential contribution of PINK1 in developmental processes, we monitored the dynamic expression pattern of PINK1 in the midgut development by taking advantage of a newly CRISPR/Cas9 generated knock-in fly strain expressing PINK1-mCherry fusion protein that presumably recapitulates the endogenous expression pattern of PINK1. We disclosed a spatiotemporal correlation between the expression pattern of PINK1 and the mitochondrial clearance or persistence in ECs or ISCs respectively. By mosaic genetic analysis, we then demonstrated that PINK1 and PARKIN function epistatically to mediate the specific timely removal of mitochondria, and are involved in global autophagy in ECs during Drosophila midgut metamorphosis, with kinase-dead PINK1 exerting dominant negative effects. Taken together, our studies concluded that the PINK1/PARKIN is crucial for timely cell-type specific mitophagy under physiological conditions and demonstrated again that Drosophila midgut metamorphosis might serve as an elegant in vivo model to study autophagy. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Cell type-specific roles of Jak3 in IL-2-induced proliferative signal transduction

    Fujii, Hodaka

    2007-01-01

    Binding of interleukin-2 (IL-2) to its specific receptor induces activation of two members of Jak family protein tyrosine kinases, Jak1 and Jak3. An IL-2 receptor (IL-2R)-reconstituted NIH 3T3 fibroblast cell line proliferates in response to IL-2 only when hematopoietic lineage-specific Jak3 is ectopically expressed. However, the mechanism of Jak3-dependent proliferation in the fibroblast cell line is not known. Here, I showed that Jak3 expression is dispensable for IL-2-induced activation of Jak1 and Stat proteins and expression of nuclear proto-oncogenes in the IL-2R-reconstituted fibroblast cell line. Jak3 expression markedly enhanced these IL-2-induced signaling events. In contrast, Jak3 expression was essential for induction of cyclin genes involved in the G1-S transition. These data suggest a critical role of Jak3 in IL-2 signaling in the fibroblast cell line and may provide further insight into the cell type-specific mechanism of cytokine signaling

  8. Cell type-specific roles of Jak3 in IL-2-induced proliferative signal transduction

    Fujii, Hodaka

    2007-01-01

    Binding of IL-2 to its specific receptor induces activation of two members of Jak family protein tyrosine kinases, Jak1 and Jak3. An IL-2R-reconstituted NIH 3T3 fibroblast cell line proliferates in response to IL-2 only when hematopoietic lineage-specific Jak3 is ectopically expressed. However, the mechanism of Jak3-dependent proliferation in the fibroblast cell line is not known. Here, I showed that Jak3 expression is dispensable for IL-2-induced activation of Jak1 and Stat proteins and expression of nuclear proto-oncogenes in the IL-2R-reconstituted fibroblast cell line. However, Jak3 expression markedly enhanced these IL-2-induced signaling events. In contrast, Jak3 expression was essential for induction of cyclin genes involved in the G1-S transition. These data suggest a critical role of Jak3 in IL-2 signaling in the fibroblast cell line and may provide further insight into the cell type-specific mechanism of cytokine signaling. PMID:17266928

  9. Protein conservation and variation suggest mechanisms of cell type-specific modulation of signaling pathways.

    Martin H Schaefer

    2014-06-01

    Full Text Available Many proteins and signaling pathways are present in most cell types and tissues and yet perform specialized functions. To elucidate mechanisms by which these ubiquitous pathways are modulated, we overlaid information about cross-cell line protein abundance and variability, and evolutionary conservation onto functional pathway components and topological layers in the pathway hierarchy. We found that the input (receptors and the output (transcription factors layers evolve more rapidly than proteins in the intermediary transmission layer. In contrast, protein expression variability decreases from the input to the output layer. We observed that the differences in protein variability between the input and transmission layer can be attributed to both the network position and the tendency of variable proteins to physically interact with constitutively expressed proteins. Differences in protein expression variability and conservation are also accompanied by the tendency of conserved and constitutively expressed proteins to acquire somatic mutations, while germline mutations tend to occur in cell type-specific proteins. Thus, conserved core proteins in the transmission layer could perform a fundamental role in most cell types and are therefore less tolerant to germline mutations. In summary, we propose that the core signal transmission machinery is largely modulated by a variable input layer through physical protein interactions. We hypothesize that the bow-tie organization of cellular signaling on the level of protein abundance variability contributes to the specificity of the signal response in different cell types.

  10. Epigenetic regulation of normal human mammary cell type-specific miRNAs

    Vrba, Lukas [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center; Inst. of Plant Molecular Biology, Ceske Budejovice (Czech Republic). Biology Centre ASCR; Garbe, James C. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Center; Stampfer, Martha R. [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Center; Futscher, Bernard W. [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center and Dept. of Pharmacology & Toxicology

    2011-08-26

    Epigenetic mechanisms are important regulators of cell type–specific genes, including miRNAs. In order to identify cell type-specific miRNAs regulated by epigenetic mechanisms, we undertook a global analysis of miRNA expression and epigenetic states in three isogenic pairs of human mammary epithelial cells (HMEC) and human mammary fibroblasts (HMF), which represent two differentiated cell types typically present within a given organ, each with a distinct phenotype and a distinct epigenotype. While miRNA expression and epigenetic states showed strong interindividual concordance within a given cell type, almost 10% of the expressed miRNA showed a cell type–specific pattern of expression that was linked to the epigenetic state of their promoter. The tissue-specific miRNA genes were epigenetically repressed in nonexpressing cells by DNA methylation (38%) and H3K27me3 (58%), with only a small set of miRNAs (21%) showing a dual epigenetic repression where both DNA methylation and H3K27me3 were present at their promoters, such as MIR10A and MIR10B. Individual miRNA clusters of closely related miRNA gene families can each display cell type–specific repression by the same or complementary epigenetic mechanisms, such as the MIR200 family, and MIR205, where fibroblasts repress MIR200C/141 by DNA methylation, MIR200A/200B/429 by H3K27me3, and MIR205 by both DNA methylation and H3K27me3. Since deregulation of many of the epigenetically regulated miRNAs that we identified have been linked to disease processes such as cancer, it is predicted that compromise of the epigenetic control mechanisms is important for this process. Overall, these results highlight the importance of epigenetic regulation in the control of normal cell type–specific miRNA expression.

  11. Cell type-specific termination of transcription by transposable element sequences.

    Conley, Andrew B; Jordan, I King

    2012-09-30

    Transposable elements (TEs) encode sequences necessary for their own transposition, including signals required for the termination of transcription. TE sequences within the introns of human genes show an antisense orientation bias, which has been proposed to reflect selection against TE sequences in the sense orientation owing to their ability to terminate the transcription of host gene transcripts. While there is evidence in support of this model for some elements, the extent to which TE sequences actually terminate transcription of human gene across the genome remains an open question. Using high-throughput sequencing data, we have characterized over 9,000 distinct TE-derived sequences that provide transcription termination sites for 5,747 human genes across eight different cell types. Rarefaction curve analysis suggests that there may be twice as many TE-derived termination sites (TE-TTS) genome-wide among all human cell types. The local chromatin environment for these TE-TTS is similar to that seen for 3' UTR canonical TTS and distinct from the chromatin environment of other intragenic TE sequences. However, those TE-TTS located within the introns of human genes were found to be far more cell type-specific than the canonical TTS. TE-TTS were much more likely to be found in the sense orientation than other intragenic TE sequences of the same TE family and TE-TTS in the sense orientation terminate transcription more efficiently than those found in the antisense orientation. Alu sequences were found to provide a large number of relatively weak TTS, whereas LTR elements provided a smaller number of much stronger TTS. TE sequences provide numerous termination sites to human genes, and TE-derived TTS are particularly cell type-specific. Thus, TE sequences provide a powerful mechanism for the diversification of transcriptional profiles between cell types and among evolutionary lineages, since most TE-TTS are evolutionarily young. The extent of transcription

  12. Cell type-specific termination of transcription by transposable element sequences

    Conley Andrew B

    2012-09-01

    Full Text Available Abstract Background Transposable elements (TEs encode sequences necessary for their own transposition, including signals required for the termination of transcription. TE sequences within the introns of human genes show an antisense orientation bias, which has been proposed to reflect selection against TE sequences in the sense orientation owing to their ability to terminate the transcription of host gene transcripts. While there is evidence in support of this model for some elements, the extent to which TE sequences actually terminate transcription of human gene across the genome remains an open question. Results Using high-throughput sequencing data, we have characterized over 9,000 distinct TE-derived sequences that provide transcription termination sites for 5,747 human genes across eight different cell types. Rarefaction curve analysis suggests that there may be twice as many TE-derived termination sites (TE-TTS genome-wide among all human cell types. The local chromatin environment for these TE-TTS is similar to that seen for 3′ UTR canonical TTS and distinct from the chromatin environment of other intragenic TE sequences. However, those TE-TTS located within the introns of human genes were found to be far more cell type-specific than the canonical TTS. TE-TTS were much more likely to be found in the sense orientation than other intragenic TE sequences of the same TE family and TE-TTS in the sense orientation terminate transcription more efficiently than those found in the antisense orientation. Alu sequences were found to provide a large number of relatively weak TTS, whereas LTR elements provided a smaller number of much stronger TTS. Conclusions TE sequences provide numerous termination sites to human genes, and TE-derived TTS are particularly cell type-specific. Thus, TE sequences provide a powerful mechanism for the diversification of transcriptional profiles between cell types and among evolutionary lineages, since most TE-TTS are

  13. Focusing on neuronal cell-type specific mechanisms for brain circuit organization, function and dysfunction

    Lu Li

    2017-01-01

    Mammalian brain circuits consist of dynamically interconnected neurons with characteristic morphology, physiology, connectivity and genetics which are often called neuronal cell types. Neuronal cell types have been considered as building blocks of brain circuits, but knowledge of how neuron types or subtypes connect to and interact with each other to perform neural computation is still lacking. Such mechanistic insights are critical not only to our understanding of normal brain functions, such as perception, motion and cognition, but also to brain disorders including Alzheimer's disease, Schizophrenia and epilepsy, to name a few. Thus it is necessary to carry out systematic and standardized studies on neuronal cell-type specific mechanisms for brain circuit organization and function, which will provide good opportunities to bridge basic and clinical research. Here based on recent technology advancements, we discuss the strategy to target and manipulate specific populations of neuronsin vivo to provide unique insights on how neuron types or subtypes behave, interact, and generate emergent properties in a fully connected brain network. Our approach is highlighted by combining transgenic animal models, targeted electrophysiology and imaging with robotics, thus complete and standardized mapping ofin vivo properties of genetically defined neuron populations can be achieved in transgenic mouse models, which will facilitate the development of novel therapeutic strategies for brain disorders.

  14. Cell type-specific genetic and optogenetic tools reveal hippocampal CA2 circuits.

    Kohara, Keigo; Pignatelli, Michele; Rivest, Alexander J; Jung, Hae-Yoon; Kitamura, Takashi; Suh, Junghyup; Frank, Dominic; Kajikawa, Koichiro; Mise, Nathan; Obata, Yuichi; Wickersham, Ian R; Tonegawa, Susumu

    2014-02-01

    The formation and recall of episodic memory requires precise information processing by the entorhinal-hippocampal network. For several decades, the trisynaptic circuit entorhinal cortex layer II (ECII)→dentate gyrus→CA3→CA1 and the monosynaptic circuit ECIII→CA1 have been considered the primary substrates of the network responsible for learning and memory. Circuits linked to another hippocampal region, CA2, have only recently come to light. Using highly cell type-specific transgenic mouse lines, optogenetics and patch-clamp recordings, we found that dentate gyrus cells, long believed to not project to CA2, send functional monosynaptic inputs to CA2 pyramidal cells through abundant longitudinal projections. CA2 innervated CA1 to complete an alternate trisynaptic circuit, but, unlike CA3, projected preferentially to the deep, rather than to the superficial, sublayer of CA1. Furthermore, contrary to existing knowledge, ECIII did not project to CA2. Our results allow a deeper understanding of the biology of learning and memory.

  15. Cell-type specificity of ChIP-predicted transcription factor binding sites

    Håndstad Tony

    2012-08-01

    Full Text Available Abstract Background Context-dependent transcription factor (TF binding is one reason for differences in gene expression patterns between different cellular states. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq identifies genome-wide TF binding sites for one particular context—the cells used in the experiment. But can such ChIP-seq data predict TF binding in other cellular contexts and is it possible to distinguish context-dependent from ubiquitous TF binding? Results We compared ChIP-seq data on TF binding for multiple TFs in two different cell types and found that on average only a third of ChIP-seq peak regions are common to both cell types. Expectedly, common peaks occur more frequently in certain genomic contexts, such as CpG-rich promoters, whereas chromatin differences characterize cell-type specific TF binding. We also find, however, that genotype differences between the cell types can explain differences in binding. Moreover, ChIP-seq signal intensity and peak clustering are the strongest predictors of common peaks. Compared with strong peaks located in regions containing peaks for multiple transcription factors, weak and isolated peaks are less common between the cell types and are less associated with data that indicate regulatory activity. Conclusions Together, the results suggest that experimental noise is prevalent among weak peaks, whereas strong and clustered peaks represent high-confidence binding events that often occur in other cellular contexts. Nevertheless, 30-40% of the strongest and most clustered peaks show context-dependent regulation. We show that by combining signal intensity with additional data—ranging from context independent information such as binding site conservation and position weight matrix scores to context dependent chromatin structure—we can predict whether a ChIP-seq peak is likely to be present in other cellular contexts.

  16. Lim homeobox genes in the Ctenophore Mnemiopsis leidyi: the evolution of neural cell type specification

    Simmons David K

    2012-01-01

    Full Text Available Abstract Background Nervous systems are thought to be important to the evolutionary success and diversification of metazoans, yet little is known about the origin of simple nervous systems at the base of the animal tree. Recent data suggest that ctenophores, a group of macroscopic pelagic marine invertebrates, are the most ancient group of animals that possess a definitive nervous system consisting of a distributed nerve net and an apical statocyst. This study reports on details of the evolution of the neural cell type specifying transcription factor family of LIM homeobox containing genes (Lhx, which have highly conserved functions in neural specification in bilaterian animals. Results Using next generation sequencing, the first draft of the genome of the ctenophore Mnemiopsis leidyi has been generated. The Lhx genes in all animals are represented by seven subfamilies (Lhx1/5, Lhx3/4, Lmx, Islet, Lhx2/9, Lhx6/8, and LMO of which four were found to be represented in the ctenophore lineage (Lhx1/5, Lhx3/4, Lmx, and Islet. Interestingly, the ctenophore Lhx gene complement is more similar to the sponge complement (sponges do not possess neurons than to either the cnidarian-bilaterian or placozoan Lhx complements. Using whole mount in situ hybridization, the Lhx gene expression patterns were examined and found to be expressed around the blastopore and in cells that give rise to the apical organ and putative neural sensory cells. Conclusion This research gives us a first look at neural cell type specification in the ctenophore M. leidyi. Within M. leidyi, Lhx genes are expressed in overlapping domains within proposed neural cellular and sensory cell territories. These data suggest that Lhx genes likely played a conserved role in the patterning of sensory cells in the ancestor of sponges and ctenophores, and may provide a link to the expression of Lhx orthologs in sponge larval photoreceptive cells. Lhx genes were later co-opted into patterning more

  17. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells.

    Ruth Merkle

    2016-08-01

    Full Text Available Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC, is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO. However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR. The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in

  18. Cell-Type Specific Development of the Hyperpolarization-Activated Current, Ih, in Prefrontal Cortical Neurons

    Sha-Sha Yang

    2018-05-01

    Full Text Available H-current, also known as hyperpolarization-activated current (Ih, is an inward current generated by the hyperpolarization-activated cyclic nucleotide-gated (HCN cation channels. Ih plays an essential role in regulating neuronal properties, synaptic integration and plasticity, and synchronous activity in the brain. As these biological factors change across development, the brain undergoes varying levels of vulnerability to disorders like schizophrenia that disrupt prefrontal cortex (PFC-dependent function. However, developmental changes in Ih in PFC neurons remains untested. Here, we examine Ih in pyramidal neurons vs. gamma-aminobutyric acid (GABAergic parvalbumin-expressing (PV+ interneurons in developing mouse PFC. Our findings show that the amplitudes of Ih in these cell types are identical during the juvenile period but differ at later time points. In pyramidal neurons, Ih amplitude significantly increases from juvenile to adolescence and follows a similar trend into adulthood. In contrast, the amplitude of Ih in PV+ interneurons decreases from juvenile to adolescence, and does not change from adolescence to adulthood. Moreover, the kinetics of HCN channels in pyramidal neurons is significantly slower than in PV+ interneurons, with a gradual decrease in pyramidal neurons and a gradual increase in PV+ cells across development. Our study reveals distinct developmental trajectories of Ih in pyramidal neurons and PV+ interneurons. The cell-type specific alteration of Ih during the critical period from juvenile to adolescence reflects the contribution of Ih to the maturation of the PFC and PFC-dependent function. These findings are essential for a better understanding of normal PFC function, and for elucidating Ih’s crucial role in the pathophysiology of neurodevelopmental disorders.

  19. Discovery of cell-type specific DNA motif grammar in cis-regulatory elements using random Forest.

    Wang, Xin; Lin, Peijie; Ho, Joshua W K

    2018-01-19

    It has been observed that many transcription factors (TFs) can bind to different genomic loci depending on the cell type in which a TF is expressed in, even though the individual TF usually binds to the same core motif in different cell types. How a TF can bind to the genome in such a highly cell-type specific manner, is a critical research question. One hypothesis is that a TF requires co-binding of different TFs in different cell types. If this is the case, it may be possible to observe different combinations of TF motifs - a motif grammar - located at the TF binding sites in different cell types. In this study, we develop a bioinformatics method to systematically identify DNA motifs in TF binding sites across multiple cell types based on published ChIP-seq data, and address two questions: (1) can we build a machine learning classifier to predict cell-type specificity based on motif combinations alone, and (2) can we extract meaningful cell-type specific motif grammars from this classifier model. We present a Random Forest (RF) based approach to build a multi-class classifier to predict the cell-type specificity of a TF binding site given its motif content. We applied this RF classifier to two published ChIP-seq datasets of TF (TCF7L2 and MAX) across multiple cell types. Using cross-validation, we show that motif combinations alone are indeed predictive of cell types. Furthermore, we present a rule mining approach to extract the most discriminatory rules in the RF classifier, thus allowing us to discover the underlying cell-type specific motif grammar. Our bioinformatics analysis supports the hypothesis that combinatorial TF motif patterns are cell-type specific.

  20. TACO: a general-purpose tool for predicting cell-type-specific transcription factor dimers.

    Jankowski, Aleksander; Prabhakar, Shyam; Tiuryn, Jerzy

    2014-03-19

    Cooperative binding of transcription factor (TF) dimers to DNA is increasingly recognized as a major contributor to binding specificity. However, it is likely that the set of known TF dimers is highly incomplete, given that they were discovered using ad hoc approaches, or through computational analyses of limited datasets. Here, we present TACO (Transcription factor Association from Complex Overrepresentation), a general-purpose standalone software tool that takes as input any genome-wide set of regulatory elements and predicts cell-type-specific TF dimers based on enrichment of motif complexes. TACO is the first tool that can accommodate motif complexes composed of overlapping motifs, a characteristic feature of many known TF dimers. Our method comprehensively outperforms existing tools when benchmarked on a reference set of 29 known dimers. We demonstrate the utility and consistency of TACO by applying it to 152 DNase-seq datasets and 94 ChIP-seq datasets. Based on these results, we uncover a general principle governing the structure of TF-TF-DNA ternary complexes, namely that the flexibility of the complex is correlated with, and most likely a consequence of, inter-motif spacing.

  1. Cell-type-specific responses of RT4 neural cell lines to dibutyryl-cAMP: branch determination versus maturation

    Droms, K.; Sueoka, N.

    1987-01-01

    This report describes the induction of cell-type-specific maturation, by dibutyryl-cAMP and testololactone, of neuronal and glial properties in a family of cell lines derived from a rat peripheral neurotumor, RT4. This maturation allows further understanding of the process of determination because of the close lineage relationship between the cell types of the RT4 family. The RT4 family is characterized by the spontaneous conversion of one of the cell types, RT4-AC (stem-cell type), to any of three derivative cell types, RT4-B, RT4-D, or RT4-E, with a frequency of about 10(-5). The RT4-AC cells express some properties characteristic of both neuronal and glial cells. Of these neural properties expressed by RT4-AC cells, only the neuronal properties are expressed by the RT4-B and RT4-E cells, and only the glial properties are expressed by the RT4-D cells. This in vitro cell-type conversion of RT4-AC to three derivative cell types is a branch point for the coordinate regulation of several properties and seems to resemble determination in vivo. In our standard culture conditions, several other neuronal and glial properties are not expressed by these cell types. However, addition of dibutyryl-cAMP induces expression of additional properties, in a cell-type-specific manner: formation of long cellular processes in the RT4-B8 and RT4-E5 cell lines and expression of high-affinity uptake of gamma-aminobutyric acid, by a glial-cell-specific mechanism, in the RT4-D6-2 cell line. These new properties are maximally expressed 2-3 days after addition of dibutyryl-cAMP

  2. Genetically-directed, cell type-specific sparse labeling for the analysis of neuronal morphology.

    Thomas Rotolo

    Full Text Available In mammals, genetically-directed cell labeling technologies have not yet been applied to the morphologic analysis of neurons with very large and complex arbors, an application that requires extremely sparse labeling and that is only rendered practical by limiting the labeled population to one or a few predetermined neuronal subtypes.In the present study we have addressed this application by using CreER technology to non-invasively label very small numbers of neurons so that their morphologies can be fully visualized. Four lines of IRES-CreER knock-in mice were constructed to permit labeling selectively in cholinergic or catecholaminergic neurons [choline acetyltransferase (ChAT-IRES-CreER or tyrosine hydroxylase (TH-IRES-CreER], predominantly in projection neurons [neurofilament light chain (NFL-IRES-CreER], or broadly in neurons and some glia [vesicle-associated membrane protein2 (VAMP2-IRES-CreER]. When crossed to the Z/AP reporter and exposed to 4-hydroxytamoxifen in the early postnatal period, the number of neurons expressing the human placental alkaline phosphatase reporter can be reproducibly lowered to fewer than 50 per brain. Sparse Cre-mediated recombination in ChAT-IRES-CreER;Z/AP mice shows the full axonal and dendritic arbors of individual forebrain cholinergic neurons, the first time that the complete morphologies of these very large neurons have been revealed in any species.Sparse genetically-directed, cell type-specific neuronal labeling with IRES-creER lines should prove useful for studying a wide variety of questions in neuronal development and disease.

  3. Regulatory domain selectivity in the cell-type specific PKN-dependence of cell migration.

    Sylvie Lachmann

    Full Text Available The mammalian protein kinase N (PKN family of Serine/Threonine kinases comprises three isoforms, which are targets for Rho family GTPases. Small GTPases are major regulators of the cellular cytoskeleton, generating interest in the role(s of specific PKN isoforms in processes such as cell migration and invasion. It has been reported that PKN3 is required for prostate tumour cell invasion but not PKN1 or 2. Here we employ a cell model, the 5637 bladder tumour cell line where PKN2 is relatively highly expressed, to assess the potential redundancy of these isoforms in migratory responses. It is established that PKN2 has a critical role in the migration and invasion of these cells. Furthermore, using a PKN wild-type and chimera rescue strategy, it is shown that PKN isoforms are not simply redundant in supporting migration, but appear to be linked through isoform specific regulatory domain properties to selective upstream signals. It is concluded that intervention in PKNs may need to be directed at multiple isoforms to be effective in different cell types.

  4. Regulatory domain selectivity in the cell-type specific PKN-dependence of cell migration.

    Lachmann, Sylvie; Jevons, Amy; De Rycker, Manu; Casamassima, Adele; Radtke, Simone; Collazos, Alejandra; Parker, Peter J

    2011-01-01

    The mammalian protein kinase N (PKN) family of Serine/Threonine kinases comprises three isoforms, which are targets for Rho family GTPases. Small GTPases are major regulators of the cellular cytoskeleton, generating interest in the role(s) of specific PKN isoforms in processes such as cell migration and invasion. It has been reported that PKN3 is required for prostate tumour cell invasion but not PKN1 or 2. Here we employ a cell model, the 5637 bladder tumour cell line where PKN2 is relatively highly expressed, to assess the potential redundancy of these isoforms in migratory responses. It is established that PKN2 has a critical role in the migration and invasion of these cells. Furthermore, using a PKN wild-type and chimera rescue strategy, it is shown that PKN isoforms are not simply redundant in supporting migration, but appear to be linked through isoform specific regulatory domain properties to selective upstream signals. It is concluded that intervention in PKNs may need to be directed at multiple isoforms to be effective in different cell types.

  5. Mutant PIK3CA Induces EMT in a Cell Type Specific Manner.

    Divya Bhagirath

    Full Text Available Breast cancer is characterized into different molecular subtypes, and each subtype is characterized by differential gene expression that are associated with distinct survival outcomes in patients. PIK3CA mutations are commonly associated with most breast cancer subtypes. More recently PIK3CA mutations have been shown to induce tumor heterogeneity and are associated with activation of EGFR-signaling and reduced relapse free survival in basal subtype of breast cancer. Thus, understanding what determines PIK3CA induced heterogeneity and oncogenesis, is an important area of investigation. In this study, we assessed the effect of mutant PIK3CA together with mutant Ras plus mutant p53 on oncogenic behavior of two distinct stem/progenitor breast cell lines, designated as K5+/K19- and K5+/K19+. Constructs were ectopically overexpressed in K5+/K19- and K5+/K19+ stem/progenitor cells, followed by various in-vitro and in-vivo analyses. Oncogene combination m-Ras/m-p53/m-PIK3CA efficiently transformed both K5+/K19- and K5+/K19+ cell lines in-vitro, as assessed by anchorage-independent soft agar colony formation assay. Significantly, while this oncogene combination induced a complete epithelial-to-mesenchymal transition (EMT in K5+/K19- cell line, mostly epithelial phenotype with minor EMT component was seen in K5+/K19+ cell line. However, both K5+/K19- and K5+/K19+ transformed cells exhibited increased invasion and migration abilities. Analyses of CD44 and CD24 expression showed both cell lines had tumor-initiating CD44+/CD24low cell population, however transformed K5+/K19- cells had more proportion of these cells. Significantly, both cell types exhibited in-vivo tumorigenesis, and maintained their EMT and epithelial nature in-vivo in mice tumors. Notably, while both cell types exhibited increase in tumor-initiating cell population, differential EMT phenotype was observed in these cell lines. These results suggest that EMT is a cell type dependent

  6. Invariant TAD Boundaries Constrain Cell-Type-Specific Looping Interactions between Promoters and Distal Elements around the CFTR Locus.

    Smith, Emily M; Lajoie, Bryan R; Jain, Gaurav; Dekker, Job

    2016-01-07

    Three-dimensional genome structure plays an important role in gene regulation. Globally, chromosomes are organized into active and inactive compartments while, at the gene level, looping interactions connect promoters to regulatory elements. Topologically associating domains (TADs), typically several hundred kilobases in size, form an intermediate level of organization. Major questions include how TADs are formed and how they are related to looping interactions between genes and regulatory elements. Here we performed a focused 5C analysis of a 2.8 Mb chromosome 7 region surrounding CFTR in a panel of cell types. We find that the same TAD boundaries are present in all cell types, indicating that TADs represent a universal chromosome architecture. Furthermore, we find that these TAD boundaries are present irrespective of the expression and looping of genes located between them. In contrast, looping interactions between promoters and regulatory elements are cell-type specific and occur mostly within TADs. This is exemplified by the CFTR promoter that in different cell types interacts with distinct sets of distal cell-type-specific regulatory elements that are all located within the same TAD. Finally, we find that long-range associations between loci located in different TADs are also detected, but these display much lower interaction frequencies than looping interactions within TADs. Interestingly, interactions between TADs are also highly cell-type-specific and often involve loci clustered around TAD boundaries. These data point to key roles of invariant TAD boundaries in constraining as well as mediating cell-type-specific long-range interactions and gene regulation. Copyright © 2016 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  7. Cell type-specific response to high intracellular loading of polyacrylic acid-coated magnetic nanoparticles

    Lojk, Jasna; Bregar, Vladimir B; Rajh, Maruša; Miš, Katarina; Kreft, Mateja Erdani; Pirkmajer, Sergej; Veranič, Peter; Pavlin, Mojca

    2015-01-01

    Magnetic nanoparticles (NPs) are a special type of NP with a ferromagnetic, electron-dense core that enables several applications such as cell tracking, hyperthermia, and magnetic separation, as well as multimodality. So far, superparamagnetic iron oxide NPs (SPIONs) are the only clinically approved type of metal oxide NPs, but cobalt ferrite NPs have properties suitable for biomedical applications as well. In this study, we analyzed the cellular responses to magnetic cobalt ferrite NPs coated with polyacrylic acid (PAA) in three cell types: Chinese Hamster Ovary (CHO), mouse melanoma (B16) cell line, and primary human myoblasts (MYO). We compared the internalization pathway, intracellular trafficking, and intracellular fate of our NPs using fluorescence and transmission electron microscopy (TEM) as well as quantified NP uptake and analyzed uptake dynamics. We determined cell viability after 24 or 96 hours’ exposure to increasing concentrations of NPs, and quantified the generation of reactive oxygen species (ROS) upon 24 and 48 hours’ exposure. Our NPs have been shown to readily enter and accumulate in cells in high quantities using the same two endocytic pathways; mostly by macropinocytosis and partially by clathrin-mediated endocytosis. The cell types differed in their uptake rate, the dynamics of intracellular trafficking, and the uptake capacity, as well as in their response to higher concentrations of internalized NPs. The observed differences in cell responses stress the importance of evaluation of NP–cell interactions on several different cell types for better prediction of possible toxic effects on different cell and tissue types in vivo. PMID:25733835

  8. Cell-type-specific enrichment of risk-associated regulatory elements at ovarian cancer susceptibility loci.

    Coetzee, Simon G; Shen, Howard C; Hazelett, Dennis J; Lawrenson, Kate; Kuchenbaecker, Karoline; Tyrer, Jonathan; Rhie, Suhn K; Levanon, Keren; Karst, Alison; Drapkin, Ronny; Ramus, Susan J; Couch, Fergus J; Offit, Kenneth; Chenevix-Trench, Georgia; Monteiro, Alvaro N A; Antoniou, Antonis; Freedman, Matthew; Coetzee, Gerhard A; Pharoah, Paul D P; Noushmehr, Houtan; Gayther, Simon A

    2015-07-01

    Understanding the regulatory landscape of the human genome is a central question in complex trait genetics. Most single-nucleotide polymorphisms (SNPs) associated with cancer risk lie in non-protein-coding regions, implicating regulatory DNA elements as functional targets of susceptibility variants. Here, we describe genome-wide annotation of regions of open chromatin and histone modification in fallopian tube and ovarian surface epithelial cells (FTSECs, OSECs), the debated cellular origins of high-grade serous ovarian cancers (HGSOCs) and in endometriosis epithelial cells (EECs), the likely precursor of clear cell ovarian carcinomas (CCOCs). The regulatory architecture of these cell types was compared with normal human mammary epithelial cells and LNCaP prostate cancer cells. We observed similar positional patterns of global enhancer signatures across the three different ovarian cancer precursor cell types, and evidence of tissue-specific regulatory signatures compared to non-gynecological cell types. We found significant enrichment for risk-associated SNPs intersecting regulatory biofeatures at 17 known HGSOC susceptibility loci in FTSECs (P = 3.8 × 10(-30)), OSECs (P = 2.4 × 10(-23)) and HMECs (P = 6.7 × 10(-15)) but not for EECs (P = 0.45) or LNCaP cells (P = 0.88). Hierarchical clustering of risk SNPs conditioned on the six different cell types indicates FTSECs and OSECs are highly related (96% of samples using multi-scale bootstrapping) suggesting both cell types may be precursors of HGSOC. These data represent the first description of regulatory catalogues of normal precursor cells for different ovarian cancer subtypes, and provide unique insights into the tissue specific regulatory variation with respect to the likely functional targets of germline genetic susceptibility variants for ovarian cancer. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  9. Assessment of Membrane Fluidity Fluctuations during Cellular Development Reveals Time and Cell Type Specificity

    Noutsi, Bakiza Kamal; Gratton, Enrico; Chaieb, Saharoui

    2016-01-01

    Cell membrane is made up of a complex structure of lipids and proteins that diffuse laterally giving rise to what we call membrane fluidity. During cellular development, such as differentiation cell membranes undergo dramatic fluidity changes induced by proteins such as ARC and Cofilin among others. In this study we used the generalized polarization (GP) property of fluorescent probe Laurdan using two-photon microscopy to determine membrane fluidity as a function of time and for various cell lines. A low GP value corresponds to a higher fluidity and a higher GP value is associated with a more rigid membrane. Four different cell lines were monitored such as hN2, NIH3T3, HEK293 and L6 cells. Membrane fluidity was measured at 12h, 72h and 92 h. Our results show significant changes in membrane fluidity among all cell types at different time points. GP values tend to increase significantly within 92 h in hN2 cells and 72 h in NIH3T3 cells and only at 92 h in HEK293 cells. L6 showed a marked decrease in membrane fluidity at 72 h and starts to increase at 92 h. As expected, NIH3T3 cells have more rigid membrane at earlier time points. On the other hand, neurons tend to have the highest membrane fluidity at early time points emphasizing its correlation with plasticity and the need for this malleability during differentiation. This study sheds light on the involvement of membrane fluidity during neuronal differentiation and development of other cell lines.

  10. Assessment of Membrane Fluidity Fluctuations during Cellular Development Reveals Time and Cell Type Specificity

    Noutsi, Bakiza Kamal

    2016-06-30

    Cell membrane is made up of a complex structure of lipids and proteins that diffuse laterally giving rise to what we call membrane fluidity. During cellular development, such as differentiation cell membranes undergo dramatic fluidity changes induced by proteins such as ARC and Cofilin among others. In this study we used the generalized polarization (GP) property of fluorescent probe Laurdan using two-photon microscopy to determine membrane fluidity as a function of time and for various cell lines. A low GP value corresponds to a higher fluidity and a higher GP value is associated with a more rigid membrane. Four different cell lines were monitored such as hN2, NIH3T3, HEK293 and L6 cells. Membrane fluidity was measured at 12h, 72h and 92 h. Our results show significant changes in membrane fluidity among all cell types at different time points. GP values tend to increase significantly within 92 h in hN2 cells and 72 h in NIH3T3 cells and only at 92 h in HEK293 cells. L6 showed a marked decrease in membrane fluidity at 72 h and starts to increase at 92 h. As expected, NIH3T3 cells have more rigid membrane at earlier time points. On the other hand, neurons tend to have the highest membrane fluidity at early time points emphasizing its correlation with plasticity and the need for this malleability during differentiation. This study sheds light on the involvement of membrane fluidity during neuronal differentiation and development of other cell lines.

  11. Cell Type-specific Intrinsic Perithreshold Oscillations in Hippocampal GABAergic Interneurons.

    Kang, Young-Jin; Lewis, Hannah Elisabeth Smashey; Young, Mason William; Govindaiah, Gubbi; Greenfield, Lazar John; Garcia-Rill, Edgar; Lee, Sang-Hun

    2018-04-15

    The hippocampus plays a critical role in learning, memory, and spatial processing through coordinated network activity including theta and gamma oscillations. Recent evidence suggests that hippocampal subregions (e.g., CA1) can generate these oscillations at the network level, at least in part, through GABAergic interneurons. However, it is unclear whether specific GABAergic interneurons generate intrinsic theta and/or gamma oscillations at the single-cell level. Since major types of CA1 interneurons (i.e., parvalbumin-positive basket cells (PVBCs), cannabinoid type 1 receptor-positive basket cells (CB 1 BCs), Schaffer collateral-associated cells (SCAs), neurogliaform cells and ivy cells) are thought to play key roles in network theta and gamma oscillations in the hippocampus, we tested the hypothesis that these cells generate intrinsic perithreshold oscillations at the single-cell level. We performed whole-cell patch-clamp recordings from GABAergic interneurons in the CA1 region of the mouse hippocampus in the presence of synaptic blockers to identify intrinsic perithreshold membrane potential oscillations. The majority of PVBCs (83%), but not the other interneuron subtypes, produced intrinsic perithreshold gamma oscillations if the membrane potential remained above -45 mV. In contrast, CB 1 BCs, SCAs, neurogliaform cells, ivy cells, and the remaining PVBCs (17%) produced intrinsic theta, but not gamma, oscillations. These oscillations were prevented by blockers of persistent sodium current. These data demonstrate that the major types of hippocampal interneurons produce distinct frequency bands of intrinsic perithreshold membrane oscillations. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

  12. Cell-Type Specific DNA Methylation Patterns Define Human Breast Cellular Identity

    Novák, Petr; Stampfer, M.R.; Munoz-Rodriguez, J.L.; Garbe, J.C.; Ehrich, M.; Futscher, B. W.; Jensen, T.J.

    2012-01-01

    Roč. 7, č. 12 (2012), e52299 E-ISSN 1932-6203 Institutional research plan: CEZ:AV0Z50510513 Institutional support: RVO:60077344 Keywords : MAMMARY EPITHELIAL-CELLS * PLURIPOTENT STEM-CELLS * CPG ISLAND SHORES Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.730, year: 2012

  13. Gene Expression Programs in Response to Hypoxia: Cell Type Specificity and Prognostic Significance in Human Cancers.

    2006-01-01

    Full Text Available BACKGROUND: Inadequate oxygen (hypoxia triggers a multifaceted cellular response that has important roles in normal physiology and in many human diseases. A transcription factor, hypoxia-inducible factor (HIF, plays a central role in the hypoxia response; its activity is regulated by the oxygen-dependent degradation of the HIF-1alpha protein. Despite the ubiquity and importance of hypoxia responses, little is known about the variation in the global transcriptional response to hypoxia among different cell types or how this variation might relate to tissue- and cell-specific diseases. METHODS AND FINDINGS: We analyzed the temporal changes in global transcript levels in response to hypoxia in primary renal proximal tubule epithelial cells, breast epithelial cells, smooth muscle cells, and endothelial cells with DNA microarrays. The extent of the transcriptional response to hypoxia was greatest in the renal tubule cells. This heightened response was associated with a uniquely high level of HIF-1alpha RNA in renal cells, and it could be diminished by reducing HIF-1alpha expression via RNA interference. A gene-expression signature of the hypoxia response, derived from our studies of cultured mammary and renal tubular epithelial cells, showed coordinated variation in several human cancers, and was a strong predictor of clinical outcomes in breast and ovarian cancers. In an analysis of a large, published gene-expression dataset from breast cancers, we found that the prognostic information in the hypoxia signature was virtually independent of that provided by the previously reported wound signature and more predictive of outcomes than any of the clinical parameters in current use. CONCLUSIONS: The transcriptional response to hypoxia varies among human cells. Some of this variation is traceable to variation in expression of the HIF1A gene. A gene-expression signature of the cellular response to hypoxia is associated with a significantly poorer prognosis

  14. High intensity training may reverse the fiber type specific decline in myogenic stem cells in multiple sclerosis patients

    Jean eFarup

    2016-05-01

    Full Text Available Multiple sclerosis (MS is associated with loss of skeletal muscle mass and function. The myogenic stem cells (satellite cells – SCs are instrumental to accretion of myonuclei, but remain to be investigated in MS. The present study aimed to compare the SC and myonuclei content between MS patients (n=23 and age matched healthy controls (HC, n=18. Furthermore, the effects of 12 weeks of high intensity training on SC and myonuclei content were explored in MS. Muscle biopsies were obtained from m. Vastus Lateralis at baseline (MS+HC and following 12 weeks of training (MS only. Frozen biopsies were sectioned followed by immunohistochemical analysis for fiber type specific SCs (Pax7+, myonuclei (MN and central nuclei content and fiber cross-sectional area (fCSA using ATPase histochemistry. At baseline the SCs per fiber was lower in type II compared to type I fiber in both MS (119%, p<0.01 and HC (69%, p<0.05, whereas the SCs per fCSA was lower in type II fibers compared to type I only in MS (72%, p<0.05. No differences were observed in MN or central nuclei between MS and HC. Following training the type II fiber SCs per fiber and fCSA in MS patients increased by 165% (p<0.05 and 135% (p<0.05, respectively. Furthermore, the type II fiber MN content increased by 35% (p<0.05 following training. In conclusion, the SC content is lower in type II compared to type I fibers in both MS and HC. Furthermore, high intensity training was observed to selectively increase the SC and myonuclei content in type II fibers in MS patients.

  15. Cell-Type-Specific Gene Programs of the Normal Human Nephron Define Kidney Cancer Subtypes.

    Lindgren, David; Eriksson, Pontus; Krawczyk, Krzysztof; Nilsson, Helén; Hansson, Jennifer; Veerla, Srinivas; Sjölund, Jonas; Höglund, Mattias; Johansson, Martin E; Axelson, Håkan

    2017-08-08

    Comprehensive transcriptome studies of cancers often rely on corresponding normal tissue samples to serve as a transcriptional reference. In this study, we performed in-depth analyses of normal kidney tissue transcriptomes from the TCGA and demonstrate that the histological variability in cellularity, inherent in the kidney architecture, lead to considerable transcriptional differences between samples. This should be considered when comparing expression profiles of normal and cancerous kidney tissues. We exploited these differences to define renal-cell-specific gene signatures and used these as a framework to analyze renal cell carcinoma (RCC) ontogeny. Chromophobe RCCs express FOXI1-driven genes that define collecting duct intercalated cells, whereas HNF-regulated genes, specific for proximal tubule cells, are an integral part of clear cell and papillary RCC transcriptomes. These networks may be used as a framework for understanding the interplay between genomic changes in RCC subtypes and the lineage-defining regulatory machinery of their non-neoplastic counterparts. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  16. Unique cell type-specific junctional complexes in vascular endothelium of human and rat liver sinusoids.

    Cyrill Géraud

    Full Text Available Liver sinusoidal endothelium is strategically positioned to control access of fluids, macromolecules and cells to the liver parenchyma and to serve clearance functions upstream of the hepatocytes. While clearance of macromolecular debris from the peripheral blood is performed by liver sinusoidal endothelial cells (LSECs using a delicate endocytic receptor system featuring stabilin-1 and -2, the mannose receptor and CD32b, vascular permeability and cell trafficking are controlled by transcellular pores, i.e. the fenestrae, and by intercellular junctional complexes. In contrast to blood vascular and lymphatic endothelial cells in other organs, the junctional complexes of LSECs have not yet been consistently characterized in molecular terms. In a comprehensive analysis, we here show that LSECs express the typical proteins found in endothelial adherens junctions (AJ, i.e. VE-cadherin as well as α-, β-, p120-catenin and plakoglobin. Tight junction (TJ transmembrane proteins typical of endothelial cells, i.e. claudin-5 and occludin, were not expressed by rat LSECs while heterogenous immunreactivity for claudin-5 was detected in human LSECs. In contrast, junctional molecules preferentially associating with TJ such as JAM-A, B and C and zonula occludens proteins ZO-1 and ZO-2 were readily detected in LSECs. Remarkably, among the JAMs JAM-C was considerably over-expressed in LSECs as compared to lung microvascular endothelial cells. In conclusion, we show here that LSECs form a special kind of mixed-type intercellular junctions characterized by co-occurrence of endothelial AJ proteins, and of ZO-1 and -2, and JAMs. The distinct molecular architecture of the intercellular junctional complexes of LSECs corroborates previous ultrastructural findings and provides the molecular basis for further analyses of the endothelial barrier function of liver sinusoids under pathologic conditions ranging from hepatic inflammation to formation of liver metastasis.

  17. Analysis of cell-type-specific gene expression during mouse spermatogenesis

    Almstrup, Kristian; Nielsen, John E; Hansen, Martin Asser

    2004-01-01

    In rodents, changes in gene expression during spermatogenesis can be monitored by sampling testis from each day during postnatal development. However, changes in gene expression at the tissue level can reflect changes in the concentration of an mRNA in a specific cell type, changes in volume of s...

  18. Cell-Type Specific Determinants of NRAMP1 Expression in Professional Phagocytes

    Mathieu F. M. Cellier

    2013-01-01

    Full Text Available The Natural resistance-associated macrophage protein 1 (Nramp1 or Solute carrier 11 member 1, Slc11a1 transports divalent metals across the membrane of late endosomes and lysosomes in professional phagocytes. Nramp1 represents an ancient eukaryotic cell-autonomous defense whereas the gene duplication that yielded Nramp1 and Nramp2 predated the origin of Sarcopterygians (lobe-finned fishes and tetrapods. SLC11A1 genetic polymorphisms associated with human resistance to tuberculosis consist of potential regulatory variants. Herein, current knowledge of the regulation of SLC11A1 gene expression is reviewed and comprehensive analysis of ENCODE data available for hematopoietic cell-types suggests a hypothesis for the regulation of SLC11A1 expression during myeloid development and phagocyte functional polarization. SLC11A1 is part of a 34.6 kb CTCF-insulated locus scattered with predicted regulatory elements: a 3' enhancer, a large 5' enhancer domain and four elements spread around the transcription start site (TSS, including several C/EBP and PU.1 sites. SLC11A1 locus ends appear mobilized by ETS-related factors early during myelopoiesis; activation of both 5' and 3' enhancers in myelo-monocytic cells correlate with transcription factor binding at the TSS. Characterizing the corresponding cis/trans determinants functionally will establish the mechanisms involved and possibly reveal genetic variation that impacts susceptibility to infectious or immune diseases.

  19. Cell Type-Specific Manipulation with GFP-Dependent Cre Recombinase

    Tang, Jonathan C Y; Rudolph, Stephanie; Dhande, Onkar S; Abraira, Victoria E; Choi, Seungwon; Lapan, Sylvain; Drew, Iain R; Drokhlyansky, Eugene; Huberman, Andrew D; Regehr, Wade G; Cepko, Constance L

    2016-01-01

    Summary There are many transgenic GFP reporter lines that allow visualization of specific populations of cells. Using such lines for functional studies requires a method that transforms GFP into a molecule that enables genetic manipulation. Here we report the creation of a method that exploits GFP for gene manipulation, Cre Recombinase Dependent on GFP (CRE-DOG), a split component system that uses GFP and its derivatives to directly induce Cre/loxP recombination. Using plasmid electroporation and AAV viral vectors, we delivered CRE-DOG to multiple GFP mouse lines, leading to effective recombination selectively in GFP-labeled cells. Further, CRE-DOG enabled optogenetic control of these neurons. Beyond providing a new set of tools for manipulation of gene expression selectively in GFP+ cells, we demonstrate that GFP can be used to reconstitute the activity of a protein not known to have a modular structure, suggesting that this strategy might be applicable to a wide range of proteins. PMID:26258682

  20. Cell-type specific mechanisms of D-serine uptake and release in the brain

    Magalie eMartineau

    2014-05-01

    Full Text Available Accumulating evidence during the last decade established that D-serine is a key signaling molecule utilized by neurons and astroglia in the mammalian central nervous system. D-serine is increasingly appreciated as the main physiological endogenous coagonist for synaptic NMDA receptors at central excitatory synapses; it is mandatory for long-term changes in synaptic strength, memory, learning, and social interactions. Alterations in the extracellular levels of D-serine leading to disrupted cell-cell signaling are a trademark of many chronic or acute neurological (i.e. Alzheimer disease, epilepsy, stroke and psychiatric (i.e. schizophrenia disorders, and are associated with addictive behavior (i.e. cocaine addiction. Indeed, fine tuning of the extracellular levels of D-serine, achieved by various molecular machineries and signaling pathways, is necessary for maintenance of accurate NMDA receptor functions. Here, we review the experimental data supporting the notion that astroglia and neurons use different pathways to regulate levels of extracellular D-serine.

  1. ICK is essential for cell type-specific ciliogenesis and the regulation of ciliary transport.

    Chaya, Taro; Omori, Yoshihiro; Kuwahara, Ryusuke; Furukawa, Takahisa

    2014-06-02

    Cilia and flagella are formed and maintained by intraflagellar transport (IFT) and play important roles in sensing and moving across species. At the distal tip of the cilia/flagella, IFT complexes turn around to switch from anterograde to retrograde transport; however, the underlying regulatory mechanism is unclear. Here, we identified ICK localization at the tip of cilia as a regulator of ciliary transport. In ICK-deficient mice, we found ciliary defects in neuronal progenitor cells with Hedgehog signal defects. ICK-deficient cells formed cilia with mislocalized Hedgehog signaling components. Loss of ICK caused the accumulation of IFT-A, IFT-B, and BBSome components at the ciliary tips. In contrast, overexpression of ICK induced the strong accumulation of IFT-B, but not IFT-A or BBSome components at ciliary tips. In addition, ICK directly phosphorylated Kif3a, while inhibition of this Kif3a phosphorylation affected ciliary formation. Our results suggest that ICK is a Kif3a kinase and essential for proper ciliogenesis in development by regulating ciliary transport at the tip of cilia. © 2014 The Authors.

  2. Towards identifying host cell-type specific response patterns to bacterial endosymbiosis

    Gavrilovic, Srdjan

    The establishment of Symbiotic Nitrogen Fixation (SNF) is a complex process. It requires highly sophisticated signal exchanges between host plant and bacteria in order to fine-tune the molecular mechanisms necessary for optimal performance of the symbiosis, which ultimately determines the evoluti......The establishment of Symbiotic Nitrogen Fixation (SNF) is a complex process. It requires highly sophisticated signal exchanges between host plant and bacteria in order to fine-tune the molecular mechanisms necessary for optimal performance of the symbiosis, which ultimately determines......, and whole plant transformants were regenerated. These will form a basis for isolating transcriptionally active mRNA fractions associated with ribosomes and 21 nt long small RNAs from targeted cell populations....

  3. Partitioning Heritability of Regulatory and Cell-Type-Specific Variants across 11 Common Diseases

    Gusev, Alexander; Lee, S Hong; Trynka, Gosia

    2014-01-01

    Regulatory and coding variants are known to be enriched with associations identified by genome-wide association studies (GWASs) of complex disease, but their contributions to trait heritability are currently unknown. We applied variance-component methods to imputed genotype data for 11 common...... diseases to partition the heritability explained by genotyped SNPs (hg(2)) across functional categories (while accounting for shared variance due to linkage disequilibrium). Extensive simulations showed that in contrast to current estimates from GWAS summary statistics, the variance-component approach...... partitions heritability accurately under a wide range of complex-disease architectures. Across the 11 diseases DNaseI hypersensitivity sites (DHSs) from 217 cell types spanned 16% of imputed SNPs (and 24% of genotyped SNPs) but explained an average of 79% (SE = 8%) of hg(2) from imputed SNPs (5.1× enrichment...

  4. Homeostasis or channelopathy? Acquired cell type-specific ion channel changes in temporal lobe epilepsy and their antiepileptic potential

    Wolfart, Jakob; Laker, Debora

    2015-01-01

    Neurons continuously adapt the expression and functionality of their ion channels. For example, exposed to chronic excitotoxicity, neurons homeostatically downscale their intrinsic excitability. In contrast, the “acquired channelopathy” hypothesis suggests that proepileptic channel characteristics develop during epilepsy. We review cell type-specific channel alterations under different epileptic conditions and discuss the potential of channels that undergo homeostatic adaptations, as targets for antiepileptic drugs (AEDs). Most of the relevant studies have been performed on temporal lobe epilepsy (TLE), a widespread AED-refractory, focal epilepsy. The TLE patients, who undergo epilepsy surgery, frequently display hippocampal sclerosis (HS), which is associated with degeneration of cornu ammonis subfield 1 pyramidal cells (CA1 PCs). Although the resected human tissue offers insights, controlled data largely stem from animal models simulating different aspects of TLE and other epilepsies. Most of the cell type-specific information is available for CA1 PCs and dentate gyrus granule cells (DG GCs). Between these two cell types, a dichotomy can be observed: while DG GCs acquire properties decreasing the intrinsic excitability (in TLE models and patients with HS), CA1 PCs develop channel characteristics increasing intrinsic excitability (in TLE models without HS only). However, thorough examination of data on these and other cell types reveals the coexistence of protective and permissive intrinsic plasticity within neurons. These mechanisms appear differentially regulated, depending on the cell type and seizure condition. Interestingly, the same channel molecules that are upregulated in DG GCs during HS-related TLE, appear as promising targets for future AEDs and gene therapies. Hence, GCs provide an example of homeostatic ion channel adaptation which can serve as a primer when designing novel anti-epileptic strategies. PMID:26124723

  5. Cell-type-specific activation of mitogen-activated protein kinases in PAN-induced progressive renal disease in rats

    Park, Sang-Joon; Jeong, Kyu-Shik

    2004-01-01

    We examined the time-course activation and the cell-type specific role of MAP kinases in puromycin aminonucleoside (PAN)-induced renal disease. The maximal activation of c-Jun-NH 2 -terminal kinase (JNK), extracellular signal regulated kinase (ERK), and p38 MAP kinase was detected on Days 52, 38, and 38 after PAN-treatment, respectively. p-JNK was localized in mesangial and proximal tubular cells at the early renal injury. It was expressed, therefore, in the inflammatory cells of tubulointerstitial lesions. While, p-ERK was markedly increased in the glomerular regions and macrophages p-p38 was observed in glomerular endothelial cells, tubular cells, and some inflammatory cells. The results show that the activation of MAP kinases in the early renal injury by PAN-treatment involves cellular changes such as cell proliferation or apoptosis in renal native cells. The activation of MAP kinases in infiltrated inflammatory cells and fibrotic cells plays an important role in destructive events such as glomerulosclerosis and tubulointerstitial fibrosis

  6. Cell type-specific anti-cancer properties of valproic acid: independent effects on HDAC activity and Erk1/2 phosphorylation

    Gotfryd, Kamil; Skladchikova, Galina; Lepekhin, Eugene E

    2010-01-01

    lines (BT4C, BT4Cn, U87MG, N2a, PC12-E2, CSML0, CSML100, HeLa, L929, Swiss 3T3). Results: VPA induced significant histone deacetylase (HDAC) inhibition in most of the cell lines, but the degree of inhibition was highly cell type-specific. Moreover, cell growth, motility and the degree of Erk1......ABSTRACT: BACKGROUND: The anti-epileptic drug valproic acid (VPA) has attracted attention as an anti-cancer agent. Methods: The present study investigated effects of VPA exposure on histone deacetylase (HDAC) inhibition, cell growth, cell speed, and the degree of Erk1/2 phosphorylation in 10 cell....../2 phosphorylation were inhibited, activated, or unaffected by VPA in a cell type-specific manner. Importantly, no relationship was found between the effects of VPA on HDAC inhibition and changes in the degree of Erk1/2 phosphorylation, cell growth, or motility. In contrast, VPA-induced modulation of the MAPK...

  7. Samd7 is a cell type-specific PRC1 component essential for establishing retinal rod photoreceptor identity.

    Omori, Yoshihiro; Kubo, Shun; Kon, Tetsuo; Furuhashi, Mayu; Narita, Hirotaka; Kominami, Taro; Ueno, Akiko; Tsutsumi, Ryotaro; Chaya, Taro; Yamamoto, Haruka; Suetake, Isao; Ueno, Shinji; Koseki, Haruhiko; Nakagawa, Atsushi; Furukawa, Takahisa

    2017-09-26

    Precise transcriptional regulation controlled by a transcription factor network is known to be crucial for establishing correct neuronal cell identities and functions in the CNS. In the retina, the expression of various cone and rod photoreceptor cell genes is regulated by multiple transcription factors; however, the role of epigenetic regulation in photoreceptor cell gene expression has been poorly understood. Here, we found that Samd7, a rod-enriched sterile alpha domain (SAM) domain protein, is essential for silencing nonrod gene expression through H3K27me3 regulation in rod photoreceptor cells. Samd7- null mutant mice showed ectopic expression of nonrod genes including S-opsin in rod photoreceptor cells and rod photoreceptor cell dysfunction. Samd7 physically interacts with Polyhomeotic homologs (Phc proteins), components of the Polycomb repressive complex 1 (PRC1), and colocalizes with Phc2 and Ring1B in Polycomb bodies. ChIP assays showed a significant decrease of H3K27me3 in the genes up-regulated in the Samd7 -deficient retina, showing that Samd7 deficiency causes the derepression of nonrod gene expression in rod photoreceptor cells. The current study suggests that Samd7 is a cell type-specific PRC1 component epigenetically defining rod photoreceptor cell identity.

  8. ON Bipolar Cells in Macaque Retina: Type-Specific Synaptic Connectivity with Special Reference to OFF Counterparts

    Tsukamoto, Yoshihiko; Omi, Naoko

    2016-01-01

    To date, 12 macaque bipolar cell types have been described. This list includes all morphology types first outlined by Polyak (1941) using the Golgi method in the primate retina and subsequently identified by other researchers using electron microscopy (EM) combined with the Golgi method, serial section transmission EM (SSTEM), and immunohistochemical imaging. We used SSTEM for the rod-dense perifoveal area of macaque retina, reconfirmed ON (cone) bipolar cells to be classified as invaginating midget bipolar (IMB), diffuse bipolar (DB)4, DB5, DB6, giant bipolar (GB), and blue bipolar (BB) types, and clarified their type-specific connectivity. DB4 cells made reciprocal synapses with a kind of ON-OFF lateral amacrine cell, similar to OFF DB2 cells. GB cells contacted rods and cones, similar to OFF DB3b cells. Retinal circuits formed by GB and DB3b cells are thought to substantiate the psychophysical finding of fast rod signals in mesopic vision. DB6 cell output synapses were directed to ON midget ganglion (MG) cells at 70% of ribbon contacts, similar to OFF DB1 cells that directed 60% of ribbon contacts to OFF MG cells. IMB cells contacted medium- or long-wavelength sensitive (M/L-) cones but not short-wavelength sensitive (S-) cones, while BB cells contacted S-cones but not M/L-cones. However, IMB and BB dendrites had similar morphological architectures, and a BB cell contacting a single S-cone resembled an IMB cell. Thus, both IMB and BB may be the ON bipolar counterparts of the OFF flat midget bipolar (FMB) type, likewise DB4 of DB2, DB5 of DB3a, DB6 of DB1, and GB of DB3b OFF bipolar type. The ON DB plus GB, and OFF DB cells predominantly contacted M/L-cones and their outputs were directed mainly to parasol ganglion (PG) cells but also moderately to MG cells. BB cells directed S-cone-driven outputs almost exclusively to small bistratified ganglion (SBG) cells. Some FMB cells predominantly contacted S-cones and their outputs were directed to OFF MG cells. Thus, two

  9. Cell-type specific oxytocin gene expression from AAV delivered promoter deletion constructs into the rat supraoptic nucleus in vivo.

    Raymond L Fields

    Full Text Available The magnocellular neurons (MCNs in the hypothalamus selectively express either oxytocin (OXT or vasopressin (AVP neuropeptide genes, a property that defines their phenotypes. Here we examine the molecular basis of this selectivity in the OXT MCNs by stereotaxic microinjections of adeno-associated virus (AAV vectors that contain various OXT gene promoter deletion constructs using EGFP as the reporter into the rat supraoptic nucleus (SON. Two weeks following injection of the AAVs, immunohistochemical assays of EGFP expression from these constructs were done to determine whether the EGFP reporter co-localizes with either the OXT- or AVP-immunoreactivity in the MCNs. The results show that the key elements in the OT gene promoter that regulate the cell-type specific expression the SON are located -216 to -100 bp upstream of the transcription start site. We hypothesize that within this 116 bp domain a repressor exists that inhibits expression specifically in AVP MCNs, thereby leading to the cell-type specific expression of the OXT gene only in the OXT MCNs.

  10. Tissue- and Cell Type-Specific Expression of the Long Noncoding RNA Klhl14-AS in Mouse

    Sara Carmela Credendino

    2017-01-01

    Full Text Available lncRNAs are acquiring increasing relevance as regulators in a wide spectrum of biological processes. The extreme heterogeneity in the mechanisms of action of these molecules, however, makes them very difficult to study, especially regarding their molecular function. A novel lncRNA has been recently identified as the most enriched transcript in mouse developing thyroid. Due to its genomic localization antisense to the protein-encoding Klhl14 gene, we named it Klhl14-AS. In this paper, we highlight that mouse Klhl14-AS produces at least five splicing variants, some of which have not been previously described. Klhl14-AS is expressed with a peculiar pattern, characterized by diverse relative abundance of its isoforms in different mouse tissues. We examine the whole expression level of Klhl14-AS in a panel of adult mouse tissues, showing that it is expressed in the thyroid, lung, kidney, testis, ovary, brain, and spleen, although at different levels. In situ hybridization analysis reveals that, in the context of each organ, Klhl14-AS shows a cell type-specific expression. Interestingly, databases report a similar expression profile for human Klhl14-AS. Our observations suggest that this lncRNA could play cell type-specific roles in several organs and pave the way for functional characterization of this gene in appropriate biological contexts.

  11. Cell type-specific localization of Ephs pairing with ephrin-B2 in the rat postnatal pituitary gland.

    Yoshida, Saishu; Kato, Takako; Kanno, Naoko; Nishimura, Naoto; Nishihara, Hiroto; Horiguchi, Kotaro; Kato, Yukio

    2017-10-01

    Sox2-expressing stem/progenitor cells in the anterior lobe of the pituitary gland form two types of micro-environments (niches): the marginal cell layer and dense cell clusters in the parenchyma. In relation to the mechanism of regulation of niches, juxtacrine signaling via ephrin and its receptor Eph is known to play important roles in various niches. The ephrin and Eph families are divided into two subclasses to create ephrin/Eph signaling in co-operation with confined partners. Recently, we reported that ephrin-B2 localizes specifically to both pituitary niches. However, the Ephs interacting with ephrin-B2 in these pituitary niches have not yet been identified. Therefore, the present study aims to identify the Ephs interacting with ephrin-B2 and the cells that produce them in the rat pituitary gland. In situ hybridization and immunohistochemistry demonstrated cell type-specific localization of candidate interacting partners for ephrin-B2, including EphA4 in cells located in the posterior lobe, EphB1 in gonadotropes, EphB2 in corticotropes, EphB3 in stem/progenitor cells and EphB4 in endothelial cells in the adult pituitary gland. In particular, double-immunohistochemistry showed cis-interactions between EphB3 and ephrin-B2 in the apical cell membranes of stem/progenitor cell niches throughout life and trans-interactions between EphB2 produced by corticotropes and ephrin-B2 located in the basolateral cell membranes of stem/progenitor cells in the early postnatal pituitary gland. These data indicate that ephrin-B2 plays a role in pituitary stem/progenitor cell niches by selective interaction with EphB3 in cis and EphB2 in trans.

  12. Spatial separation of photosynthesis and ethanol production by cell type-specific metabolic engineering of filamentous cyanobacteria.

    Ehira, Shigeki; Takeuchi, Takuto; Higo, Akiyoshi

    2018-02-01

    Cyanobacteria, which perform oxygenic photosynthesis, have drawn attention as hosts for the direct production of biofuels and commodity chemicals from CO 2 and H 2 O using light energy. Although cyanobacteria capable of producing diverse chemicals have been generated by metabolic engineering, anaerobic non-photosynthetic culture conditions are often necessary for their production. In this study, we conducted cell type-specific metabolic engineering of the filamentous cyanobacterium Anabaena sp. PCC 7120, which forms a terminally differentiated cell called a heterocyst with a semi-regular spacing of 10-15 cells. Because heterocysts are specialized cells for nitrogen fixation, the intracellular oxygen level of heterocysts is maintained very low even when adjacent cells perform oxygenic photosynthesis. Pyruvate decarboxylase of Zymomonas mobilis and alcohol dehydrogenase of Synechocystis sp. PCC 6803 were exclusively expressed in heterocysts. Ethanol production was concomitant with nitrogen fixation in genetically engineered Anabaena sp. PCC 7120. Engineering of carbon metabolism in heterocysts improved ethanol production, and strain ET14, with an extra copy of the invB gene expressed from a heterocyst-specific promoter, produced 130.9 mg L -1 of ethanol after 9 days. ET14 produced 1681.9 mg L -1 of ethanol by increasing the CO 2 supply. Ethanol production per heterocyst cell was approximately threefold higher than that per cell of unicellular cyanobacterium. This study demonstrates the potential of heterocysts for anaerobic production of biofuels and commodity chemicals under oxygenic photosynthetic conditions.

  13. A quantitative comparison of cell-type-specific microarray gene expression profiling methods in the mouse brain.

    Benjamin W Okaty

    Full Text Available Expression profiling of restricted neural populations using microarrays can facilitate neuronal classification and provide insight into the molecular bases of cellular phenotypes. Due to the formidable heterogeneity of intermixed cell types that make up the brain, isolating cell types prior to microarray processing poses steep technical challenges that have been met in various ways. These methodological differences have the potential to distort cell-type-specific gene expression profiles insofar as they may insufficiently filter out contaminating mRNAs or induce aberrant cellular responses not normally present in vivo. Thus we have compared the repeatability, susceptibility to contamination from off-target cell-types, and evidence for stress-responsive gene expression of five different purification methods--Laser Capture Microdissection (LCM, Translating Ribosome Affinity Purification (TRAP, Immunopanning (PAN, Fluorescence Activated Cell Sorting (FACS, and manual sorting of fluorescently labeled cells (Manual. We found that all methods obtained comparably high levels of repeatability, however, data from LCM and TRAP showed significantly higher levels of contamination than the other methods. While PAN samples showed higher activation of apoptosis-related, stress-related and immediate early genes, samples from FACS and Manual studies, which also require dissociated cells, did not. Given that TRAP targets actively translated mRNAs, whereas other methods target all transcribed mRNAs, observed differences may also reflect translational regulation.

  14. Combinatorial Modulation of Signaling Pathways Reveals Cell-Type-Specific Requirements for Highly Efficient and Synchronous iPSC Reprogramming

    Simon E. Vidal

    2014-10-01

    Full Text Available The differentiated state of somatic cells provides barriers for the derivation of induced pluripotent stem cells (iPSCs. To address why some cell types reprogram more readily than others, we studied the effect of combined modulation of cellular signaling pathways. Surprisingly, inhibition of transforming growth factor β (TGF-β together with activation of Wnt signaling in the presence of ascorbic acid allows >80% of murine fibroblasts to acquire pluripotency after 1 week of reprogramming factor expression. In contrast, hepatic and blood progenitors predominantly required only TGF-β inhibition or canonical Wnt activation, respectively, to reprogram at efficiencies approaching 100%. Strikingly, blood progenitors reactivated endogenous pluripotency loci in a highly synchronous manner, and we demonstrate that expression of specific chromatin-modifying enzymes and reduced TGF-β/mitogen-activated protein (MAP kinase activity are intrinsic properties associated with the unique reprogramming response of these cells. Our observations define cell-type-specific requirements for the rapid and synchronous reprogramming of somatic cells.

  15. Dual-specificity anti-sigma factor reinforces control of cell-type specific gene expression in Bacillus subtilis.

    Mónica Serrano

    2015-04-01

    Full Text Available Gene expression during spore development in Bacillus subtilis is controlled by cell type-specific RNA polymerase sigma factors. σFand σE control early stages of development in the forespore and the mother cell, respectively. When, at an intermediate stage in development, the mother cell engulfs the forespore, σF is replaced by σG and σE is replaced by σK. The anti-sigma factor CsfB is produced under the control of σF and binds to and inhibits the auto-regulatory σG, but not σF. A position in region 2.1, occupied by an asparagine in σG and by a glutamate in οF, is sufficient for CsfB discrimination of the two sigmas, and allows it to delay the early to late switch in forespore gene expression. We now show that following engulfment completion, csfB is switched on in the mother cell under the control of σK and that CsfB binds to and inhibits σE but not σK, possibly to facilitate the switch from early to late gene expression. We show that a position in region 2.3 occupied by a conserved asparagine in σE and by a conserved glutamate in σK suffices for discrimination by CsfB. We also show that CsfB prevents activation of σG in the mother cell and the premature σG-dependent activation of σK. Thus, CsfB establishes negative feedback loops that curtail the activity of σE and prevent the ectopic activation of σG in the mother cell. The capacity of CsfB to directly block σE activity may also explain how CsfB plays a role as one of the several mechanisms that prevent σE activation in the forespore. Thus the capacity of CsfB to differentiate between the highly similar σF/σG and σE/σK pairs allows it to rinforce the cell-type specificity of these sigma factors and the transition from early to late development in B. subtilis, and possibly in all sporeformers that encode a CsfB orthologue.

  16. Cell-type-specific predictive network yields novel insights into mouse embryonic stem cell self-renewal and cell fate.

    Karen G Dowell

    Full Text Available Self-renewal, the ability of a stem cell to divide repeatedly while maintaining an undifferentiated state, is a defining characteristic of all stem cells. Here, we clarify the molecular foundations of mouse embryonic stem cell (mESC self-renewal by applying a proven Bayesian network machine learning approach to integrate high-throughput data for protein function discovery. By focusing on a single stem-cell system, at a specific developmental stage, within the context of well-defined biological processes known to be active in that cell type, we produce a consensus predictive network that reflects biological reality more closely than those made by prior efforts using more generalized, context-independent methods. In addition, we show how machine learning efforts may be misled if the tissue specific role of mammalian proteins is not defined in the training set and circumscribed in the evidential data. For this study, we assembled an extensive compendium of mESC data: ∼2.2 million data points, collected from 60 different studies, under 992 conditions. We then integrated these data into a consensus mESC functional relationship network focused on biological processes associated with embryonic stem cell self-renewal and cell fate determination. Computational evaluations, literature validation, and analyses of predicted functional linkages show that our results are highly accurate and biologically relevant. Our mESC network predicts many novel players involved in self-renewal and serves as the foundation for future pluripotent stem cell studies. This network can be used by stem cell researchers (at http://StemSight.org to explore hypotheses about gene function in the context of self-renewal and to prioritize genes of interest for experimental validation.

  17. Improving sensitivity of linear regression-based cell type-specific differential expression deconvolution with per-gene vs. global significance threshold.

    Glass, Edmund R; Dozmorov, Mikhail G

    2016-10-06

    The goal of many human disease-oriented studies is to detect molecular mechanisms different between healthy controls and patients. Yet, commonly used gene expression measurements from blood samples suffer from variability of cell composition. This variability hinders the detection of differentially expressed genes and is often ignored. Combined with cell counts, heterogeneous gene expression may provide deeper insights into the gene expression differences on the cell type-specific level. Published computational methods use linear regression to estimate cell type-specific differential expression, and a global cutoff to judge significance, such as False Discovery Rate (FDR). Yet, they do not consider many artifacts hidden in high-dimensional gene expression data that may negatively affect linear regression. In this paper we quantify the parameter space affecting the performance of linear regression (sensitivity of cell type-specific differential expression detection) on a per-gene basis. We evaluated the effect of sample sizes, cell type-specific proportion variability, and mean squared error on sensitivity of cell type-specific differential expression detection using linear regression. Each parameter affected variability of cell type-specific expression estimates and, subsequently, the sensitivity of differential expression detection. We provide the R package, LRCDE, which performs linear regression-based cell type-specific differential expression (deconvolution) detection on a gene-by-gene basis. Accounting for variability around cell type-specific gene expression estimates, it computes per-gene t-statistics of differential detection, p-values, t-statistic-based sensitivity, group-specific mean squared error, and several gene-specific diagnostic metrics. The sensitivity of linear regression-based cell type-specific differential expression detection differed for each gene as a function of mean squared error, per group sample sizes, and variability of the proportions

  18. Folic acid induces cell type-specific changes in the transcriptome of breast cancer cell lines: a proof-of-concept study.

    Price, R Jordan; Lillycrop, Karen A; Burdge, Graham C

    2016-01-01

    The effect of folic acid (FA) on breast cancer (BC) risk is uncertain. We hypothesised that this uncertainty may be due, in part, to differential effects of FA between BC cells with different phenotypes. To test this we investigated the effect of treatment with FA concentrations within the range of unmetabolised FA reported in humans on the expression of the transcriptome of non-transformed (MCF10A) and cancerous (MCF7 and Hs578T) BC cells. The total number of transcripts altered was: MCF10A, seventy-five (seventy up-regulated); MCF7, twenty-four (fourteen up-regulated); and Hs578T, 328 (156 up-regulated). Only the cancer-associated gene TAGLN was altered by FA in all three cell lines. In MCF10A and Hs578T cells, FA treatment decreased pathways associated with apoptosis, cell death and senescence, but increased those associated with cell proliferation. The folate transporters SLC19A1, SLC46A1 and FOLR1 were differentially expressed between cell lines tested. However, the level of expression was not altered by FA treatment. These findings suggest that physiological concentrations of FA can induce cell type-specific changes in gene regulation in a manner that is consistent with proliferative phenotype. This has implications for understanding the role of FA in BC risk. In addition, these findings support the suggestion that differences in gene expression induced by FA may involve differential activities of folate transporters. Together these findings indicate the need for further studies of the effect of FA on BC.

  19. In Vivo Senescence in the Sbds-Deficient Murine Pancreas: Cell-Type Specific Consequences of Translation Insufficiency.

    Marina E Tourlakis

    2015-06-01

    Full Text Available Genetic models of ribosome dysfunction show selective organ failure, highlighting a gap in our understanding of cell-type specific responses to translation insufficiency. Translation defects underlie a growing list of inherited and acquired cancer-predisposition syndromes referred to as ribosomopathies. We sought to identify molecular mechanisms underlying organ failure in a recessive ribosomopathy, with particular emphasis on the pancreas, an organ with a high and reiterative requirement for protein synthesis. Biallelic loss of function mutations in SBDS are associated with the ribosomopathy Shwachman-Diamond syndrome, which is typified by pancreatic dysfunction, bone marrow failure, skeletal abnormalities and neurological phenotypes. Targeted disruption of Sbds in the murine pancreas resulted in p53 stabilization early in the postnatal period, specifically in acinar cells. Decreased Myc expression was observed and atrophy of the adult SDS pancreas could be explained by the senescence of acinar cells, characterized by induction of Tgfβ, p15(Ink4b and components of the senescence-associated secretory program. This is the first report of senescence, a tumour suppression mechanism, in association with SDS or in response to a ribosomopathy. Genetic ablation of p53 largely resolved digestive enzyme synthesis and acinar compartment hypoplasia, but resulted in decreased cell size, a hallmark of decreased translation capacity. Moreover, p53 ablation resulted in expression of acinar dedifferentiation markers and extensive apoptosis. Our findings indicate a protective role for p53 and senescence in response to Sbds ablation in the pancreas. In contrast to the pancreas, the Tgfβ molecular signature was not detected in fetal bone marrow, liver or brain of mouse models with constitutive Sbds ablation. Nevertheless, as observed with the adult pancreas phenotype, disease phenotypes of embryonic tissues, including marked neuronal cell death due to apoptosis

  20. In Vivo Senescence in the Sbds-Deficient Murine Pancreas: Cell-Type Specific Consequences of Translation Insufficiency

    Tourlakis, Marina E.; Zhang, Siyi; Ball, Heather L.; Gandhi, Rikesh; Liu, Hongrui; Zhong, Jian; Yuan, Julie S.; Guidos, Cynthia J.; Durie, Peter R.; Rommens, Johanna M.

    2015-01-01

    Genetic models of ribosome dysfunction show selective organ failure, highlighting a gap in our understanding of cell-type specific responses to translation insufficiency. Translation defects underlie a growing list of inherited and acquired cancer-predisposition syndromes referred to as ribosomopathies. We sought to identify molecular mechanisms underlying organ failure in a recessive ribosomopathy, with particular emphasis on the pancreas, an organ with a high and reiterative requirement for protein synthesis. Biallelic loss of function mutations in SBDS are associated with the ribosomopathy Shwachman-Diamond syndrome, which is typified by pancreatic dysfunction, bone marrow failure, skeletal abnormalities and neurological phenotypes. Targeted disruption of Sbds in the murine pancreas resulted in p53 stabilization early in the postnatal period, specifically in acinar cells. Decreased Myc expression was observed and atrophy of the adult SDS pancreas could be explained by the senescence of acinar cells, characterized by induction of Tgfβ, p15Ink4b and components of the senescence-associated secretory program. This is the first report of senescence, a tumour suppression mechanism, in association with SDS or in response to a ribosomopathy. Genetic ablation of p53 largely resolved digestive enzyme synthesis and acinar compartment hypoplasia, but resulted in decreased cell size, a hallmark of decreased translation capacity. Moreover, p53 ablation resulted in expression of acinar dedifferentiation markers and extensive apoptosis. Our findings indicate a protective role for p53 and senescence in response to Sbds ablation in the pancreas. In contrast to the pancreas, the Tgfβ molecular signature was not detected in fetal bone marrow, liver or brain of mouse models with constitutive Sbds ablation. Nevertheless, as observed with the adult pancreas phenotype, disease phenotypes of embryonic tissues, including marked neuronal cell death due to apoptosis, were determined to

  1. Knockdown of the fat mass and obesity gene disrupts cellular energy balance in a cell-type specific manner.

    Ryan T Pitman

    Full Text Available Recent studies suggest that FTO variants strongly correlate with obesity and mainly influence energy intake with little effect on the basal metabolic rate. We suggest that FTO influences eating behavior by modulating intracellular energy levels and downstream signaling mechanisms which control energy intake and metabolism. Since FTO plays a particularly important role in adipocytes and in hypothalamic neurons, SH-SY5Y neuronal cells and 3T3-L1 adipocytes were used to understand how siRNA mediated knockdown of FTO expression alters cellular energy homeostasis. Cellular energy status was evaluated by measuring ATP levels using a luminescence assay and uptake of fluorescent glucose. FTO siRNA in SH-SY5Y cells mediated mRNA knockdown (-82%, increased ATP concentrations by up to 46% (P = 0.013 compared to controls, and decreased phosphorylation of AMPk and Akt in SH-SY5Y by -52% and -46% respectively as seen by immunoblotting. In contrast, FTO siRNA in 3T3-L1 cells decreased ATP concentration by -93% (p<0.0005, and increased AMPk and Akt phosphorylation by 204% and 70%, respectively suggesting that FTO mediates control of energy levels in a cell-type specific manner. Furthermore, glucose uptake was decreased in both SH-SY5Y (-51% p = 0.015 and 3T3-L1 cells (-30%, p = 0.0002. We also show that FTO knockdown decreases NPY mRNA expression in SH-SY5Y cells (-21% through upregulation of pSTAT3 (118%. These results provide important evidence that FTO-variant linked obesity may be associated with altered metabolic functions through activation of downstream metabolic mediators including AMPk.

  2. Synthesis of Heparan Sulfate with Cyclophilin B-binding Properties Is Determined by Cell Type-specific Expression of Sulfotransferases*

    Deligny, Audrey; Denys, Agnès; Marcant, Adeline; Melchior, Aurélie; Mazurier, Joël; van Kuppevelt, Toin H.; Allain, Fabrice

    2010-01-01

    Cyclophilin B (CyPB) induces migration and adhesion of T lymphocytes via a mechanism that requires interaction with 3-O-sulfated heparan sulfate (HS). HS biosynthesis is a complex process with many sulfotransferases involved. N-Deacetylases/N-sulfotransferases are responsible for N-sulfation, which is essential for subsequent modification steps, whereas 3-O-sulfotransferases (3-OSTs) catalyze the least abundant modification. These enzymes are represented by several isoforms, which differ in term of distribution pattern, suggesting their involvement in making tissue-specific HS. To elucidate how the specificity of CyPB binding is determined, we explored the relationships between the expression of these sulfotransferases and the generation of HS motifs with CyPB-binding properties. We demonstrated that high N-sulfate density and the presence of 2-O- and 3-O-sulfates determine binding of CyPB, as evidenced by competitive experiments with heparin derivatives, soluble HS, and anti-HS antibodies. We then showed that target cells, i.e. CD4+ lymphocyte subsets, monocytes/macrophages, and related cell lines, specifically expressed high levels of NDST2 and 3-OST3 isoforms. Silencing the expression of NDST1, NDST2, 2-OST, and 3-OST3 by RNA interference efficiently decreased binding and activity of CyPB, thus confirming their involvement in the biosynthesis of binding sequences for CyPB. Moreover, we demonstrated that NDST1 was able to partially sulfate exogenous substrate in the absence of NDST2 but not vice versa, suggesting that both isoenzymes do not have redundant activities but do have rather complementary activities in making N-sulfated sequences with CyPB-binding properties. Altogether, these results suggest a regulatory mechanism in which cell type-specific expression of certain HS sulfotransferases determines the specific binding of CyPB to target cells. PMID:19940140

  3. Synthesis of heparan sulfate with cyclophilin B-binding properties is determined by cell type-specific expression of sulfotransferases.

    Deligny, Audrey; Denys, Agnès; Marcant, Adeline; Melchior, Aurélie; Mazurier, Joël; van Kuppevelt, Toin H; Allain, Fabrice

    2010-01-15

    Cyclophilin B (CyPB) induces migration and adhesion of T lymphocytes via a mechanism that requires interaction with 3-O-sulfated heparan sulfate (HS). HS biosynthesis is a complex process with many sulfotransferases involved. N-Deacetylases/N-sulfotransferases are responsible for N-sulfation, which is essential for subsequent modification steps, whereas 3-O-sulfotransferases (3-OSTs) catalyze the least abundant modification. These enzymes are represented by several isoforms, which differ in term of distribution pattern, suggesting their involvement in making tissue-specific HS. To elucidate how the specificity of CyPB binding is determined, we explored the relationships between the expression of these sulfotransferases and the generation of HS motifs with CyPB-binding properties. We demonstrated that high N-sulfate density and the presence of 2-O- and 3-O-sulfates determine binding of CyPB, as evidenced by competitive experiments with heparin derivatives, soluble HS, and anti-HS antibodies. We then showed that target cells, i.e. CD4+ lymphocyte subsets, monocytes/macrophages, and related cell lines, specifically expressed high levels of NDST2 and 3-OST3 isoforms. Silencing the expression of NDST1, NDST2, 2-OST, and 3-OST3 by RNA interference efficiently decreased binding and activity of CyPB, thus confirming their involvement in the biosynthesis of binding sequences for CyPB. Moreover, we demonstrated that NDST1 was able to partially sulfate exogenous substrate in the absence of NDST2 but not vice versa, suggesting that both isoenzymes do not have redundant activities but do have rather complementary activities in making N-sulfated sequences with CyPB-binding properties. Altogether, these results suggest a regulatory mechanism in which cell type-specific expression of certain HS sulfotransferases determines the specific binding of CyPB to target cells.

  4. Layer- and cell-type-specific subthreshold and suprathreshold effects of long-term monocular deprivation in rat visual cortex.

    Medini, Paolo

    2011-11-23

    Connectivity and dendritic properties are determinants of plasticity that are layer and cell-type specific in the neocortex. However, the impact of experience-dependent plasticity at the level of synaptic inputs and spike outputs remains unclear along vertical cortical microcircuits. Here I compared subthreshold and suprathreshold sensitivity to prolonged monocular deprivation (MD) in rat binocular visual cortex in layer 4 and layer 2/3 pyramids (4Ps and 2/3Ps) and in thick-tufted and nontufted layer 5 pyramids (5TPs and 5NPs), which innervate different extracortical targets. In normal rats, 5TPs and 2/3Ps are the most binocular in terms of synaptic inputs, and 5NPs are the least. Spike responses of all 5TPs were highly binocular, whereas those of 2/3Ps were dominated by either the contralateral or ipsilateral eye. MD dramatically shifted the ocular preference of 2/3Ps and 4Ps, mostly by depressing deprived-eye inputs. Plasticity was profoundly different in layer 5. The subthreshold ocular preference shift was sevenfold smaller in 5TPs because of smaller depression of deprived inputs combined with a generalized loss of responsiveness, and was undetectable in 5NPs. Despite their modest ocular dominance change, spike responses of 5TPs consistently lost their typically high binocularity during MD. The comparison of MD effects on 2/3Ps and 5TPs, the main affected output cells of vertical microcircuits, indicated that subthreshold plasticity is not uniquely determined by the initial degree of input binocularity. The data raise the question of whether 5TPs are driven solely by 2/3Ps during MD. The different suprathreshold plasticity of the two cell populations could underlie distinct functional deficits in amblyopia.

  5. Cell type-specific variations in the induction of hsp70 in human leukocytes by feverlike whole body hyperthermia.

    Oehler, R; Pusch, E; Zellner, M; Dungel, P; Hergovics, N; Homoncik, M; Eliasen, M M; Brabec, M; Roth, E

    2001-10-01

    Fever has been associated with shortened duration and improved survival in infectious disease. The mechanism of this beneficial response is still poorly understood. The heat-inducible 70-kDa heat shock protein (Hsp70) has been associated with protection of leukocytes against the cytotoxicity of inflammatory mediators and with improved survival of severe infections. This study characterizes the induction of Hsp70 by feverlike temperatures in human leukocytes in vitro and in vivo. Using flow cytometry, Hsp70 expression was determined in whole blood samples. This approach eliminated cell isolation procedures that would greatly affect the results. Heat treatment of whole blood in vitro for 2 hours at different temperatures revealed that Hsp70 expression depends on temperature and cell type; up to 41 degrees C, Hsp70 increased only slightly in lymphocytes and polymorphonuclear leukocytes. However, in monocytes a strong induction was already seen at 39 degrees C, and Hsp70 levels at 41 degrees C were 10-fold higher than in the 37 degrees C control. To be as close as possible to the physiological situation during fever, we immersed healthy volunteers in a hot water bath, inducing whole body hyperthermia (39 degrees C), and measured leukocyte Hsp70 expression. Hsp70 was induced in all leukocytes with comparable but less pronounced cell type-specific variations as observed in vitro. Thus, a systemic increase of body temperature as triggered by fever stimulates Hsp70 expression in peripheral leukocytes, especially in monocytes. This fever-induced Hsp70 expression may protect monocytes when confronted with cytotoxic inflammatory mediators, thereby improving the course of the disease.

  6. Adolescent maturation of inhibitory inputs onto cingulate cortex neurons is cell-type specific and TrkB dependent

    Angela eVandenberg

    2015-02-01

    Full Text Available The maturation of inhibitory circuits during adolescence may be tied to the onset of mental health disorders such as schizophrenia. Neurotrophin signaling likely plays a critical role in supporting inhibitory circuit development and is also implicated in psychiatric disease. Within the neocortex, subcircuits may mature at different times and show differential sensitivity to neurotrophin signaling. We measured miniature inhibitory and excitatory postsynaptic currents (mIPSC and mEPSCs in Layer 5 cell-types in the mouse anterior cingulate across the periadolescent period. We differentiated cell-types mainly by Thy1 YFP transgene expression and also retrobead injection labeling in the contralateral cingulate and ipsilateral pons. We found that YFP- neurons and commissural projecting neurons had lower frequency of mIPSCs than neighboring YFP+ neurons or pons projecting neurons in juvenile mice (P21-25. YFP- neurons and to a lesser extent commissural projecting neurons also showed a significant increase in mIPSC amplitude during the periadolescent period (P21-25 vs. P40-50, which was not seen in YFP+ neurons or pons projecting neurons. Systemic disruption of tyrosine kinase receptor B (TrkB signaling during P23-50 in TrkBF616A mice blocked developmental changes in mIPSC amplitude, without affecting miniature excitatory post synaptic currents (mEPSCs. Our data suggest that the maturation of inhibitory inputs onto layer 5 pyramidal neurons is cell-type specific. These data may inform our understanding of adolescent brain development across species and aid in identifying candidate subcircuits that may show greater vulnerability in mental illness.

  7. Npas4 regulates excitatory-inhibitory balance within neural circuits through cell-type-specific gene programs.

    Spiegel, Ivo; Mardinly, Alan R; Gabel, Harrison W; Bazinet, Jeremy E; Couch, Cameron H; Tzeng, Christopher P; Harmin, David A; Greenberg, Michael E

    2014-05-22

    The nervous system adapts to experience by inducing a transcriptional program that controls important aspects of synaptic plasticity. Although the molecular mechanisms of experience-dependent plasticity are well characterized in excitatory neurons, the mechanisms that regulate this process in inhibitory neurons are only poorly understood. Here, we describe a transcriptional program that is induced by neuronal activity in inhibitory neurons. We find that, while neuronal activity induces expression of early-response transcription factors such as Npas4 in both excitatory and inhibitory neurons, Npas4 activates distinct programs of late-response genes in inhibitory and excitatory neurons. These late-response genes differentially regulate synaptic input to these two types of neurons, promoting inhibition onto excitatory neurons while inducing excitation onto inhibitory neurons. These findings suggest that the functional outcomes of activity-induced transcriptional responses are adapted in a cell-type-specific manner to achieve a circuit-wide homeostatic response. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Drug and cell type-specific regulation of genes with different classes of estrogen receptor beta-selective agonists.

    Sreenivasan Paruthiyil

    2009-07-01

    Full Text Available Estrogens produce biological effects by interacting with two estrogen receptors, ERalpha and ERbeta. Drugs that selectively target ERalpha or ERbeta might be safer for conditions that have been traditionally treated with non-selective estrogens. Several synthetic and natural ERbeta-selective compounds have been identified. One class of ERbeta-selective agonists is represented by ERB-041 (WAY-202041 which binds to ERbeta much greater than ERalpha. A second class of ERbeta-selective agonists derived from plants include MF101, nyasol and liquiritigenin that bind similarly to both ERs, but only activate transcription with ERbeta. Diarylpropionitrile represents a third class of ERbeta-selective compounds because its selectivity is due to a combination of greater binding to ERbeta and transcriptional activity. However, it is unclear if these three classes of ERbeta-selective compounds produce similar biological activities. The goals of these studies were to determine the relative ERbeta selectivity and pattern of gene expression of these three classes of ERbeta-selective compounds compared to estradiol (E(2, which is a non-selective ER agonist. U2OS cells stably transfected with ERalpha or ERbeta were treated with E(2 or the ERbeta-selective compounds for 6 h. Microarray data demonstrated that ERB-041, MF101 and liquiritigenin were the most ERbeta-selective agonists compared to estradiol, followed by nyasol and then diarylpropionitrile. FRET analysis showed that all compounds induced a similar conformation of ERbeta, which is consistent with the finding that most genes regulated by the ERbeta-selective compounds were similar to each other and E(2. However, there were some classes of genes differentially regulated by the ERbeta agonists and E(2. Two ERbeta-selective compounds, MF101 and liquiritigenin had cell type-specific effects as they regulated different genes in HeLa, Caco-2 and Ishikawa cell lines expressing ERbeta. Our gene profiling studies

  9. Microarray Gene Expression Analysis to Evaluate Cell Type Specific Expression of Targets Relevant for Immunotherapy of Hematological Malignancies.

    M J Pont

    Full Text Available Cellular immunotherapy has proven to be effective in the treatment of hematological cancers by donor lymphocyte infusion after allogeneic hematopoietic stem cell transplantation and more recently by targeted therapy with chimeric antigen or T-cell receptor-engineered T cells. However, dependent on the tissue distribution of the antigens that are targeted, anti-tumor responses can be accompanied by undesired side effects. Therefore, detailed tissue distribution analysis is essential to estimate potential efficacy and toxicity of candidate targets for immunotherapy of hematological malignancies. We performed microarray gene expression analysis of hematological malignancies of different origins, healthy hematopoietic cells and various non-hematopoietic cell types from organs that are often targeted in detrimental immune responses after allogeneic stem cell transplantation leading to graft-versus-host disease. Non-hematopoietic cells were also cultured in the presence of IFN-γ to analyze gene expression under inflammatory circumstances. Gene expression was investigated by Illumina HT12.0 microarrays and quality control analysis was performed to confirm the cell-type origin and exclude contamination of non-hematopoietic cell samples with peripheral blood cells. Microarray data were validated by quantitative RT-PCR showing strong correlations between both platforms. Detailed gene expression profiles were generated for various minor histocompatibility antigens and B-cell surface antigens to illustrate the value of the microarray dataset to estimate efficacy and toxicity of candidate targets for immunotherapy. In conclusion, our microarray database provides a relevant platform to analyze and select candidate antigens with hematopoietic (lineage-restricted expression as potential targets for immunotherapy of hematological cancers.

  10. Delta-9-tetrahydrocannabinol accumulation, metabolism and cell-type-specific adverse effects in aggregating brain cell cultures

    Monnet-Tschudi, Florianne [Department of Physiology, University of Lausanne, 7, rue du Bugnon CH-1005 Lausanne (Switzerland); Hazekamp, Arno [Department of Plant Metabolomics, University of Leiden (Netherlands); Perret, Nicolas; Zurich, Marie-Gabrielle [Department of Physiology, University of Lausanne, 7, rue du Bugnon CH-1005 Lausanne (Switzerland); Mangin, Patrice; Giroud, Christian [Laboratory of Forensic Toxicology and Chemistry, Institute of Legal Medicine, University Hospital Center and University of Lausanne (Switzerland); Honegger, Paul [Department of Physiology, University of Lausanne, 7, rue du Bugnon CH-1005 Lausanne (Switzerland)

    2008-04-01

    Despite the widespread use of Cannabis as recreational drug or as medicine, little is known about its toxicity. The accumulation, metabolism and toxicity of THC were analyzed 10 days after a single treatment, and after repeated exposures during 10 days. Mixed-cell aggregate cultures of fetal rat telencephalon were used as in vitro model, as well as aggregates enriched either in neurons or in glial cells. It was found that THC accumulated preferentially in neurons, and that glia-neuron interactions decreased THC accumulation. The quantification of 11-OH-THC and of THC-COOH showed that brain aggregates were capable of THC metabolism. No cell-type difference was found for the metabolite 11-OH-THC, whereas the THC-COOH content was higher in mixed-cell cultures. No cell death was found at THC concentrations of 2 {mu}M in single treatment and of 1 {mu}M and 2 {mu}M in repeated treatments. Neurons, and particularly GABAergic neurons, were most sensitive to THC. Only the GABAergic marker was affected after the single treatment, whereas the GABAergic, cholinergic and astrocytic markers were decreased after the repeated treatments. JWH 015, a CB2 receptor agonist, showed effects similar to THC, whereas ACEA, a CB1 receptor agonist, had no effect. The expression of the cytokine IL-6 was upregulated 48 h after the single treatment with 5 {mu}M of THC or JWH 015, whereas the expression of TNF-{alpha} remained unchanged. These results suggest that the adverse effects of THC were related either to THC accumulation or to cannabinoid receptor activation and associated with IL-6 upregulation.

  11. Delta-9-tetrahydrocannabinol accumulation, metabolism and cell-type-specific adverse effects in aggregating brain cell cultures

    Monnet-Tschudi, Florianne; Hazekamp, Arno; Perret, Nicolas; Zurich, Marie-Gabrielle; Mangin, Patrice; Giroud, Christian; Honegger, Paul

    2008-01-01

    Despite the widespread use of Cannabis as recreational drug or as medicine, little is known about its toxicity. The accumulation, metabolism and toxicity of THC were analyzed 10 days after a single treatment, and after repeated exposures during 10 days. Mixed-cell aggregate cultures of fetal rat telencephalon were used as in vitro model, as well as aggregates enriched either in neurons or in glial cells. It was found that THC accumulated preferentially in neurons, and that glia-neuron interactions decreased THC accumulation. The quantification of 11-OH-THC and of THC-COOH showed that brain aggregates were capable of THC metabolism. No cell-type difference was found for the metabolite 11-OH-THC, whereas the THC-COOH content was higher in mixed-cell cultures. No cell death was found at THC concentrations of 2 μM in single treatment and of 1 μM and 2 μM in repeated treatments. Neurons, and particularly GABAergic neurons, were most sensitive to THC. Only the GABAergic marker was affected after the single treatment, whereas the GABAergic, cholinergic and astrocytic markers were decreased after the repeated treatments. JWH 015, a CB2 receptor agonist, showed effects similar to THC, whereas ACEA, a CB1 receptor agonist, had no effect. The expression of the cytokine IL-6 was upregulated 48 h after the single treatment with 5 μM of THC or JWH 015, whereas the expression of TNF-α remained unchanged. These results suggest that the adverse effects of THC were related either to THC accumulation or to cannabinoid receptor activation and associated with IL-6 upregulation

  12. Differential microRNA expression signatures and cell type-specific association with Taxol resistance in ovarian cancer cells

    Kim YW

    2014-02-01

    Full Text Available Yong-Wan Kim,1 Eun Young Kim,1 Doin Jeon,1 Juinn-Lin Liu,2 Helena Suhyun Kim,3 Jin Woo Choi,4 Woong Shick Ahn5 1Cancer Research Institute of Medical Science, The Catholic University of Korea, Seoul, Republic of Korea; 2Brain Tumor Center, Department of Neuro-Oncology, The University of Texas MD Anderson Cancer Center, TX, USA; 3Cancer Rehab Laboratory, RH Healthcare Systems Inc, TX, USA; 4Harvard Medical School and Wellman Center for Photomedicine, Cambridge, MA, USA; 5Department of Obstetrics and Gynecology, The Catholic University of Korea, Seoul, Republic of Korea Abstract: Paclitaxel (Taxol resistance remains a major obstacle for the successful treatment of ovarian cancer. MicroRNAs (miRNAs have oncogenic and tumor suppressor activity and are associated with poor prognosis phenotypes. miRNA screenings for this drug resistance are needed to estimate the prognosis of the disease and find better drug targets. miRNAs that were differentially expressed in Taxol-resistant ovarian cancer cells, compared with Taxol-sensitive cells, were screened by Illumina Human MicroRNA Expression BeadChips. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR was used to identify target genes of selected miRNAs. Kaplan–Meier survival analysis was applied to identify dysregulated miRNAs in ovarian cancer patients using data from The Cancer Genome Atlas. A total of 82 miRNAs were identified in ovarian carcinoma cells compared to normal ovarian cells. miR-141, miR-106a, miR-200c, miR-96, and miR-378 were overexpressed, and miR-411, miR-432, miR-494, miR-409-3p, and miR-655 were underexpressed in ovarian cancer cells. Seventeen miRNAs were overexpressed in Taxol-resistant cells, including miR-663, miR-622, and HS_188. Underexpressed miRNAs in Taxol-sensitive cells included miR-497, miR-187, miR-195, and miR-107. We further showed miR-663 and miR-622 as significant prognosis markers of the chemo-resistant patient group. In particular, the

  13. Coordinated cell type-specific epigenetic remodeling in prefrontal cortex begins before birth and continues into early adulthood.

    Hennady P Shulha

    2013-04-01

    Full Text Available Development of prefrontal and other higher-order association cortices is associated with widespread changes in the cortical transcriptome, particularly during the transitions from prenatal to postnatal development, and from early infancy to later stages of childhood and early adulthood. However, the timing and longitudinal trajectories of neuronal gene expression programs during these periods remain unclear in part because of confounding effects of concomitantly occurring shifts in neuron-to-glia ratios. Here, we used cell type-specific chromatin sorting techniques for genome-wide profiling of a histone mark associated with transcriptional regulation--H3 with trimethylated lysine 4 (H3K4me3--in neuronal chromatin from 31 subjects from the late gestational period to 80 years of age. H3K4me3 landscapes of prefrontal neurons were developmentally regulated at 1,157 loci, including 768 loci that were proximal to transcription start sites. Multiple algorithms consistently revealed that the overwhelming majority and perhaps all of developmentally regulated H3K4me3 peaks were on a unidirectional trajectory defined by either rapid gain or loss of histone methylation during the late prenatal period and the first year after birth, followed by similar changes but with progressively slower kinetics during early and later childhood and only minimal changes later in life. Developmentally downregulated H3K4me3 peaks in prefrontal neurons were enriched for Paired box (Pax and multiple Signal Transducer and Activator of Transcription (STAT motifs, which are known to promote glial differentiation. In contrast, H3K4me3 peaks subject to a progressive increase in maturing prefrontal neurons were enriched for activating protein-1 (AP-1 recognition elements that are commonly associated with activity-dependent regulation of neuronal gene expression. We uncovered a developmental program governing the remodeling of neuronal histone methylation landscapes in the prefrontal

  14. Fiber type specific response of skeletal muscle satellite cells to high-intensity resistance training in dialysis patients

    Molsted, Stig; Andersen, Jesper Løvind; Harrison, Adrian Paul

    2015-01-01

    Introduction. The aim was to investigate the effect of high-intensity resistance training on satellite cell (SC) and myonuclear number in the muscle of patients undergoing dialysis. Methods. Patients (n=21) underwent a 16-week control period, followed by 16 weeks of resistance training thrice...

  15. Characterisation of CD4 T cells in healthy and diseased koalas (Phascolarctos cinereus) using cell-type-specific monoclonal antibodies.

    Mangar, Chandan; Armitage, Charles W; Timms, Peter; Corcoran, Lynn M; Beagley, Kenneth W

    2016-07-01

    The koala (Phascolarctos cinereus) is an arboreal herbivorous marsupial that is an Australian icon. Koalas in many parts of Australia are under multiple threats including habitat destruction, dog attacks, vehicular accidents, and infectious diseases such as Chlamydia spp. and the koala retrovirus (KoRV), which may contribute to the incidence of lymphoma and leukaemia in this species. Due to a lack of koala-specific immune reagents and assays there is currently no way to adequately analyse the immune response in healthy, diseased or vaccinated animals. This paper reports the production and characterisation of the first anti-koala CD4 monoclonal antibody (mAb). The koala CD4 gene was identified and used to develop recombinant proteins for mAb production. Fluorochrome-conjugated anti-CD4 mAb was used to measure the levels of CD4(+) lymphocytes collected from koala spleens (41.1%, range 20-45.1%) lymph nodes (36.3%, range 19-55.9%) and peripheral blood (23.8%, range 17.3-35%) by flow cytometry. Biotin-conjugated anti-CD4 mAb was used for western blot to determine an approximate size of 52 kDa for the koala CD4 molecule and used in immunohistochemistry to identify CD4(+) cells in the paracortical region and germinal centres of spleen and lymph nodes. Using the anti-CD4 mab we showed that CD4 cells from vaccinated, but not control, koalas proliferated following in vitro stimulation with UV-inactivated Chlamydia pecorum and recombinant chlamydial antigens. Since CD4(+) T cells have been shown to play a pivotal role in clearing chlamydial infection in both human and mouse infections, using this novel antibody will help determine the role CD4(+) T cells play in protection against chlamydial infection in koalas and also enhance our knowledge of how KoRV affects the koala immune system. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. CTCF-mediated transcriptional regulation through cell type-specific chromosome organization in the β-globin locus

    Junier, Ivan; Dale, Ryan K.; Hou, Chunhui; Képès, François; Dean, Ann

    2012-01-01

    International audience; The principles underlying the architectural landscape of chromatin beyond the nucleosome level in living cells remains largely unknown despite its potential to play a role in mammalian gene regulation. We investigated the three-dimensional folding of a 1 Mbp region of human chromosome 11 containing the β-globin genes by integrating looping interactions of the CCCTC-binding insulator protein CTCF determined comprehensively by chromosome conformation capture (3C) into a ...

  17. Cell-type-specific H+-ATPase activity in root tissues enables K+ retention and mediates acclimation of barley (Hordeum vulgare) to salinity stress

    Shabala, Lana; Zhang, Jingyi; Pottosin, Igor

    2016-01-01

    While the importance of cell type specificity in plant adaptive responses is widely accepted, only a limited number of studies have addressed this issue at the functional level. We have combined electrophysiological, imaging, and biochemical techniques to reveal the physiological mechanisms confe...

  18. In Vivo Zonal Variation and Liver Cell-Type Specific NF-κB Localization after Chronic Adaptation to Ethanol and following Partial Hepatectomy.

    Harshavardhan Nilakantan

    Full Text Available NF-κB is a major inflammatory response mediator in the liver, playing a key role in the pathogenesis of alcoholic liver injury. We investigated zonal as well as liver cell type-specific distribution of NF-κB activation across the liver acinus following adaptation to chronic ethanol intake and 70% partial hepatectomy (PHx. We employed immunofluorescence staining, digital image analysis and statistical distributional analysis to quantify subcellular localization of NF-κB in hepatocytes and hepatic stellate cells (HSCs. We detected significant spatial heterogeneity of NF-κB expression and cellular localization between cytoplasm and nucleus across liver tissue. Our main aims involved investigating the zonal bias in NF-κB localization and determining to what extent chronic ethanol intake affects this zonal bias with in hepatocytes at baseline and post-PHx. Hepatocytes in the periportal area showed higher NF-κB expression than in the pericentral region in the carbohydrate-fed controls, but not in the ethanol group. However, the distribution of NF-κB nuclear localization in hepatocytes was shifted towards higher levels in pericentral region than in periportal area, across all treatment conditions. Chronic ethanol intake shifted the NF-κB distribution towards higher nuclear fraction in hepatocytes as compared to the pair-fed control group. Ethanol also stimulated higher NF-κB expression in a subpopulation of HSCs. In the control group, PHx elicited a shift towards higher NF-κB nuclear fraction in hepatocytes. However, this distribution remained unchanged in the ethanol group post-PHx. HSCs showed a lower NF-κB expression following PHx in both ethanol and control groups. We conclude that adaptation to chronic ethanol intake attenuates the liver zonal variation in NF-κB expression and limits the PHx-induced NF-κB activation in hepatocytes, but does not alter the NF-κB expression changes in HSCs in response to PHx. Our findings provide new

  19. Integrative modeling of eQTLs and cis-regulatory elements suggests mechanisms underlying cell type specificity of eQTLs.

    Christopher D Brown

    Full Text Available Genetic variants in cis-regulatory elements or trans-acting regulators frequently influence the quantity and spatiotemporal distribution of gene transcription. Recent interest in expression quantitative trait locus (eQTL mapping has paralleled the adoption of genome-wide association studies (GWAS for the analysis of complex traits and disease in humans. Under the hypothesis that many GWAS associations tag non-coding SNPs with small effects, and that these SNPs exert phenotypic control by modifying gene expression, it has become common to interpret GWAS associations using eQTL data. To fully exploit the mechanistic interpretability of eQTL-GWAS comparisons, an improved understanding of the genetic architecture and causal mechanisms of cell type specificity of eQTLs is required. We address this need by performing an eQTL analysis in three parts: first we identified eQTLs from eleven studies on seven cell types; then we integrated eQTL data with cis-regulatory element (CRE data from the ENCODE project; finally we built a set of classifiers to predict the cell type specificity of eQTLs. The cell type specificity of eQTLs is associated with eQTL SNP overlap with hundreds of cell type specific CRE classes, including enhancer, promoter, and repressive chromatin marks, regions of open chromatin, and many classes of DNA binding proteins. These associations provide insight into the molecular mechanisms generating the cell type specificity of eQTLs and the mode of regulation of corresponding eQTLs. Using a random forest classifier with cell specific CRE-SNP overlap as features, we demonstrate the feasibility of predicting the cell type specificity of eQTLs. We then demonstrate that CREs from a trait-associated cell type can be used to annotate GWAS associations in the absence of eQTL data for that cell type. We anticipate that such integrative, predictive modeling of cell specificity will improve our ability to understand the mechanistic basis of human

  20. Metabolic characterization of isocitrate dehydrogenase (IDH) mutant and IDH wildtype gliomaspheres uncovers cell type-specific vulnerabilities.

    Garrett, Matthew; Sperry, Jantzen; Braas, Daniel; Yan, Weihong; Le, Thuc M; Mottahedeh, Jack; Ludwig, Kirsten; Eskin, Ascia; Qin, Yue; Levy, Rachelle; Breunig, Joshua J; Pajonk, Frank; Graeber, Thomas G; Radu, Caius G; Christofk, Heather; Prins, Robert M; Lai, Albert; Liau, Linda M; Coppola, Giovanni; Kornblum, Harley I

    2018-01-01

    There is considerable interest in defining the metabolic abnormalities of IDH mutant tumors to exploit for therapy. While most studies have attempted to discern function by using cell lines transduced with exogenous IDH mutant enzyme, in this study, we perform unbiased metabolomics to discover metabolic differences between a cohort of patient-derived IDH1 mutant and IDH wildtype gliomaspheres. Using both our own microarray and the TCGA datasets, we performed KEGG analysis to define pathways differentially enriched in IDH1 mutant and IDH wildtype cells and tumors. Liquid chromatography coupled to mass spectrometry analysis with labeled glucose and deoxycytidine tracers was used to determine differences in overall cellular metabolism and nucleotide synthesis. Radiation-induced DNA damage and repair capacity was assessed using a comet assay. Differences between endogenous IDH1 mutant metabolism and that of IDH wildtype cells transduced with the IDH1 (R132H) mutation were also investigated. Our KEGG analysis revealed that IDH wildtype cells were enriched for pathways involved in de novo nucleotide synthesis, while IDH1 mutant cells were enriched for pathways involved in DNA repair. LC-MS analysis with fully labeled 13 C-glucose revealed distinct labeling patterns between IDH1 mutant and wildtype cells. Additional LC-MS tracing experiments confirmed increased de novo nucleotide synthesis in IDH wildtype cells relative to IDH1 mutant cells. Endogenous IDH1 mutant cultures incurred less DNA damage than IDH wildtype cultures and sustained better overall growth following X-ray radiation. Overexpression of mutant IDH1 in a wildtype line did not reproduce the range of metabolic differences observed in lines expressing endogenous mutations, but resulted in depletion of glutamine and TCA cycle intermediates, an increase in DNA damage following radiation, and a rise in intracellular ROS. These results demonstrate that IDH1 mutant and IDH wildtype cells are easily distinguishable

  1. Mosaic Expression of Thyroid Hormone Regulatory Genes Defines Cell Type-Specific Dependency in the Developing Chicken Cerebellum.

    Delbaere, Joke; Van Herck, Stijn L J; Bourgeois, Nele M A; Vancamp, Pieter; Yang, Shuo; Wingate, Richard J T; Darras, Veerle M

    2016-12-01

    The cerebellum is a morphologically unique brain structure that requires thyroid hormones (THs) for the correct coordination of key cellular events driving its development. Unravelling the interplay between the multiple factors that can regulate intracellular TH levels is a key step to understanding their role in the regulation of these cellular processes. We therefore investigated the regional/cell-specific expression pattern of TH transporters and deiodinases in the cerebellum using the chicken embryo as a model. In situ hybridisation revealed expression of the TH transporters monocarboxylate transporter 8 (MCT8) and 10 (MCT10), L-type amino acid transporter 1 (LAT1) and organic anion transporting polypeptide 1C1 (OATP1C1) as well as the inactivating type 3 deiodinase (D3) in the fourth ventricle choroid plexus, suggesting a possible contribution of the resulting proteins to TH exchange and subsequent inactivation of excess hormone at the blood-cerebrospinal fluid barrier. Exclusive expression of LAT1 and the activating type 2 deiodinase (D2) mRNA was found at the level of the blood-brain barrier, suggesting a concerted function for LAT1 and D2 in the direct access of active T 3 to the developing cerebellum via the capillary endothelial cells. The presence of MCT8 mRNA in Purkinje cells and cerebellar nuclei during the first 2 weeks of embryonic development points to a potential role of this transporter in the uptake of T 3 in central neurons. At later stages, together with MCT10, detection of MCT8 signal in close association with the Purkinje cell dendritic tree suggests a role of both transporters in TH signalling during Purkinje cell synaptogenesis. MCT10 was also expressed in late-born cells in the rhombic lip lineage with a clear hybridisation signal in the outer external granular layer, indicating a potential role for MCT10 in the proliferation of granule cell precursors. By contrast, expression of D3 in the first-born rhombic lip-derived population may

  2. Mammalian transcriptional hotspots are enriched for tissue specific enhancers near cell type specific highly expressed genes and are predicted to act as transcriptional activator hubs.

    Joshi, Anagha

    2014-12-30

    Transcriptional hotspots are defined as genomic regions bound by multiple factors. They have been identified recently as cell type specific enhancers regulating developmentally essential genes in many species such as worm, fly and humans. The in-depth analysis of hotspots across multiple cell types in same species still remains to be explored and can bring new biological insights. We therefore collected 108 transcription-related factor (TF) ChIP sequencing data sets in ten murine cell types and classified the peaks in each cell type in three groups according to binding occupancy as singletons (low-occupancy), combinatorials (mid-occupancy) and hotspots (high-occupancy). The peaks in the three groups clustered largely according to the occupancy, suggesting priming of genomic loci for mid occupancy irrespective of cell type. We then characterized hotspots for diverse structural functional properties. The genes neighbouring hotspots had a small overlap with hotspot genes in other cell types and were highly enriched for cell type specific function. Hotspots were enriched for sequence motifs of key TFs in that cell type and more than 90% of hotspots were occupied by pioneering factors. Though we did not find any sequence signature in the three groups, the H3K4me1 binding profile had bimodal peaks at hotspots, distinguishing hotspots from mono-modal H3K4me1 singletons. In ES cells, differentially expressed genes after perturbation of activators were enriched for hotspot genes suggesting hotspots primarily act as transcriptional activator hubs. Finally, we proposed that ES hotspots might be under control of SetDB1 and not DNMT for silencing. Transcriptional hotspots are enriched for tissue specific enhancers near cell type specific highly expressed genes. In ES cells, they are predicted to act as transcriptional activator hubs and might be under SetDB1 control for silencing.

  3. A functional SNP associated with atopic dermatitis controls cell type-specific methylation of the VSTM1 gene locus

    Dilip Kumar

    2017-02-01

    Full Text Available Abstract Background Expression quantitative trait loci (eQTL databases represent a valuable resource to link disease-associated SNPs to specific candidate genes whose gene expression is significantly modulated by the SNP under investigation. We previously identified signal inhibitory receptor on leukocytes-1 (SIRL-1 as a powerful regulator of human innate immune cell function. While it is constitutively high expressed on neutrophils, on monocytes the SIRL-1 surface expression varies strongly between individuals. The underlying mechanism of regulation, its genetic control as well as potential clinical implications had not been explored yet. Methods Whole blood eQTL data of a Chinese cohort was used to identify SNPs regulating the expression of VSTM1, the gene encoding SIRL-1. The genotype effect was validated by flow cytometry (cell surface expression, correlated with electrophoretic mobility shift assay (EMSA, chromatin immunoprecipitation (ChIP and bisulfite sequencing (C-methylation and its functional impact studied the inhibition of reactive oxygen species (ROS. Results We found a significant association of a single CpG-SNP, rs612529T/C, located in the promoter of VSTM1. Through flow cytometry analysis we confirmed that primarily in the monocytes the protein level of SIRL-1 is strongly associated with genotype of this SNP. In monocytes, the T allele of this SNP facilitates binding of the transcription factors YY1 and PU.1, of which the latter has been recently shown to act as docking site for modifiers of DNA methylation. In line with this notion rs612529T associates with a complete demethylation of the VSTM1 promoter correlating with the allele-specific upregulation of SIRL-1 expression. In monocytes, this upregulation strongly impacts the IgA-induced production of ROS by these cells. Through targeted association analysis we found a significant Meta P value of 1.14 × 10–6 for rs612529 for association to atopic dermatitis (AD

  4. Opening up the blackbox: an interpretable deep neural network-based classifier for cell-type specific enhancer predictions.

    Kim, Seong Gon; Theera-Ampornpunt, Nawanol; Fang, Chih-Hao; Harwani, Mrudul; Grama, Ananth; Chaterji, Somali

    2016-08-01

    Gene expression is mediated by specialized cis-regulatory modules (CRMs), the most prominent of which are called enhancers. Early experiments indicated that enhancers located far from the gene promoters are often responsible for mediating gene transcription. Knowing their properties, regulatory activity, and genomic targets is crucial to the functional understanding of cellular events, ranging from cellular homeostasis to differentiation. Recent genome-wide investigation of epigenomic marks has indicated that enhancer elements could be enriched for certain epigenomic marks, such as, combinatorial patterns of histone modifications. Our efforts in this paper are motivated by these recent advances in epigenomic profiling methods, which have uncovered enhancer-associated chromatin features in different cell types and organisms. Specifically, in this paper, we use recent state-of-the-art Deep Learning methods and develop a deep neural network (DNN)-based architecture, called EP-DNN, to predict the presence and types of enhancers in the human genome. It uses as features, the expression levels of the histone modifications at the peaks of the functional sites as well as in its adjacent regions. We apply EP-DNN to four different cell types: H1, IMR90, HepG2, and HeLa S3. We train EP-DNN using p300 binding sites as enhancers, and TSS and random non-DHS sites as non-enhancers. We perform EP-DNN predictions to quantify the validation rate for different levels of confidence in the predictions and also perform comparisons against two state-of-the-art computational models for enhancer predictions, DEEP-ENCODE and RFECS. We find that EP-DNN has superior accuracy and takes less time to make predictions. Next, we develop methods to make EP-DNN interpretable by computing the importance of each input feature in the classification task. This analysis indicates that the important histone modifications were distinct for different cell types, with some overlaps, e.g., H3K27ac was

  5. Histone modification profiles are predictive for tissue/cell-type specific expression of both protein-coding and microRNA genes

    Zhang Michael Q

    2011-05-01

    Full Text Available Abstract Background Gene expression is regulated at both the DNA sequence level and through modification of chromatin. However, the effect of chromatin on tissue/cell-type specific gene regulation (TCSR is largely unknown. In this paper, we present a method to elucidate the relationship between histone modification/variation (HMV and TCSR. Results A classifier for differentiating CD4+ T cell-specific genes from housekeeping genes using HMV data was built. We found HMV in both promoter and gene body regions to be predictive of genes which are targets of TCSR. For example, the histone modification types H3K4me3 and H3K27ac were identified as the most predictive for CpG-related promoters, whereas H3K4me3 and H3K79me3 were the most predictive for nonCpG-related promoters. However, genes targeted by TCSR can be predicted using other type of HMVs as well. Such redundancy implies that multiple type of underlying regulatory elements, such as enhancers or intragenic alternative promoters, which can regulate gene expression in a tissue/cell-type specific fashion, may be marked by the HMVs. Finally, we show that the predictive power of HMV for TCSR is not limited to protein-coding genes in CD4+ T cells, as we successfully predicted TCSR targeted genes in muscle cells, as well as microRNA genes with expression specific to CD4+ T cells, by the same classifier which was trained on HMV data of protein-coding genes in CD4+ T cells. Conclusion We have begun to understand the HMV patterns that guide gene expression in both tissue/cell-type specific and ubiquitous manner.

  6. Intricate and Cell Type-Specific Populations of Endogenous Circular DNA (eccDNA) in Caenorhabditis elegans and Homo sapiens.

    Shoura, Massa J; Gabdank, Idan; Hansen, Loren; Merker, Jason; Gotlib, Jason; Levene, Stephen D; Fire, Andrew Z

    2017-10-05

    Investigations aimed at defining the 3D configuration of eukaryotic chromosomes have consistently encountered an endogenous population of chromosome-derived circular genomic DNA, referred to as extrachromosomal circular DNA (eccDNA). While the production, distribution, and activities of eccDNAs remain understudied, eccDNA formation from specific regions of the linear genome has profound consequences on the regulatory and coding capabilities for these regions. Here, we define eccDNA distributions in Caenorhabditis elegans and in three human cell types, utilizing a set of DNA topology-dependent approaches for enrichment and characterization. The use of parallel biophysical, enzymatic, and informatic approaches provides a comprehensive profiling of eccDNA robust to isolation and analysis methodology. Results in human and nematode systems provide quantitative analysis of the eccDNA loci at both unique and repetitive regions. Our studies converge on and support a consistent picture, in which endogenous genomic DNA circles are present in normal physiological states, and in which the circles come from both coding and noncoding genomic regions. Prominent among the coding regions generating DNA circles are several genes known to produce a diversity of protein isoforms, with mucin proteins and titin as specific examples. Copyright © 2017 Shoura et al.

  7. Cell type-specific response to high intracellular loading of polyacrylic acid-coated magnetic nanoparticles

    Lojk J

    2015-02-01

    Full Text Available Jasna Lojk,1 Vladimir B Bregar,1 Maruša Rajh,1 Katarina Miš,2 Mateja Erdani Kreft,3 Sergej Pirkmajer,2 Peter Veranič,3 Mojca Pavlin1 1Group for Nano and Biotechnological Applications, Faculty of Electrical Engineering, 2Institute of Pathophysiology, Faculty of Medicine, 3Institute of Cell Biology, Faculty of Medicine, University of Ljubljana, Ljubljana, Slovenia Abstract: Magnetic nanoparticles (NPs are a special type of NP with a ferromagnetic, electron-dense core that enables several applications such as cell tracking, hyperthermia, and magnetic separation, as well as multimodality. So far, superparamagnetic iron oxide NPs (SPIONs are the only clinically approved type of metal oxide NPs, but cobalt ferrite NPs have properties suitable for biomedical applications as well. In this study, we analyzed the cellular responses to magnetic cobalt ferrite NPs coated with polyacrylic acid (PAA in three cell types: Chinese Hamster Ovary (CHO, mouse melanoma (B16 cell line, and primary human myoblasts (MYO. We compared the internalization pathway, intracellular trafficking, and intracellular fate of our NPs using fluorescence and transmission electron microscopy (TEM as well as quantified NP uptake and analyzed uptake dynamics. We determined cell viability after 24 or 96 hours’ exposure to increasing concentrations of NPs, and quantified the generation of reactive oxygen species (ROS upon 24 and 48 hours’ exposure. Our NPs have been shown to readily enter and accumulate in cells in high quantities using the same two endocytic pathways; mostly by macropinocytosis and partially by clathrin-mediated endocytosis. The cell types differed in their uptake rate, the dynamics of intracellular trafficking, and the uptake capacity, as well as in their response to higher concentrations of internalized NPs. The observed differences in cell responses stress the importance of evaluation of NP–cell interactions on several different cell types for better

  8. Cell type-specific deficiency of c-kit gene expression in mutant mice of mi/mi genotype.

    Isozaki, K.; Tsujimura, T.; Nomura, S.; Morii, E.; Koshimizu, U.; Nishimune, Y.; Kitamura, Y.

    1994-01-01

    The mi locus of mice encodes a novel member of the basic-helix-loop-helix-leucine zipper protein family of transcription factors (hereafter called mi factor). In addition to microphthalmus, osteopetrosis, and lack of melanocytes, mice of mi/mi genotype are deficient in mast cells. Since the c-kit receptor tyrosine kinase plays an important role in the development of mast cells, and since the c-kit expression by cultured mast cells from mi/mi mice is deficient in both mRNA and protein levels, the mast cell deficiency of mi/mi mice has been attributed at least in part to the deficient expression of c-kit. However, it remained to be examined whether the c-kit expression was also deficient in tissues of mi/mi mice. In the present study, we examined the c-kit expression by mi/mi skin mast cells using in situ hybridization and immunohistochemistry. Moreover, we examined the c-kit expression by various cells other than mast cells in tissues of mi/mi mice. We found that the c-kit expression was deficient in mast cells but not in erythroid precursors, testicular germ cells, and neurons of mi/mi mice. This suggested that the regulation of the c-kit transcription by the mi factor was dependent on cell types. Mice of mi/mi genotype appeared to be a useful model to analyze the function of transcription factors in the whole-animal level. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7524330

  9. Cell-type-specific genome editing with a microRNA-responsive CRISPR-Cas9 switch.

    Hirosawa, Moe; Fujita, Yoshihiko; Parr, Callum J C; Hayashi, Karin; Kashida, Shunnichi; Hotta, Akitsu; Woltjen, Knut; Saito, Hirohide

    2017-07-27

    The CRISPR-Cas9 system is a powerful genome-editing tool useful in a variety of biotechnology and biomedical applications. Here we developed a synthetic RNA-based, microRNA (miRNA)-responsive CRISPR-Cas9 system (miR-Cas9 switch) in which the genome editing activity of Cas9 can be modulated through endogenous miRNA signatures in mammalian cells. We created miR-Cas9 switches by using a miRNA-complementary sequence in the 5΄-UTR of mRNA encoding Streptococcus pyogenes Cas9. The miR-21-Cas9 or miR-302-Cas9 switches selectively and efficiently responded to miR-21-5p in HeLa cells or miR-302a-5p in human induced pluripotent stem cells, and post-transcriptionally attenuated the Cas9 activity only in the target cells. Moreover, the miR-Cas9 switches could differentially control the genome editing by sensing endogenous miRNA activities within a heterogeneous cell population. Our miR-Cas9 switch system provides a promising framework for cell-type selective genome editing and cell engineering based on intracellular miRNA information. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. Cell-type-specific genome editing with a microRNA-responsive CRISPR–Cas9 switch

    Hirosawa, Moe; Fujita, Yoshihiko; Parr, Callum J. C.; Hayashi, Karin; Kashida, Shunnichi; Hotta, Akitsu; Woltjen, Knut

    2017-01-01

    Abstract The CRISPR–Cas9 system is a powerful genome-editing tool useful in a variety of biotechnology and biomedical applications. Here we developed a synthetic RNA-based, microRNA (miRNA)-responsive CRISPR–Cas9 system (miR-Cas9 switch) in which the genome editing activity of Cas9 can be modulated through endogenous miRNA signatures in mammalian cells. We created miR-Cas9 switches by using a miRNA-complementary sequence in the 5΄-UTR of mRNA encoding Streptococcus pyogenes Cas9. The miR-21-Cas9 or miR-302-Cas9 switches selectively and efficiently responded to miR-21-5p in HeLa cells or miR-302a-5p in human induced pluripotent stem cells, and post-transcriptionally attenuated the Cas9 activity only in the target cells. Moreover, the miR-Cas9 switches could differentially control the genome editing by sensing endogenous miRNA activities within a heterogeneous cell population. Our miR-Cas9 switch system provides a promising framework for cell-type selective genome editing and cell engineering based on intracellular miRNA information. PMID:28525578

  11. Cell-Type-Specific Transcriptome Analysis in the Drosophila Mushroom Body Reveals Memory-Related Changes in Gene Expression.

    Crocker, Amanda; Guan, Xiao-Juan; Murphy, Coleen T; Murthy, Mala

    2016-05-17

    Learning and memory formation in Drosophila rely on a network of neurons in the mushroom bodies (MBs). Whereas numerous studies have delineated roles for individual cell types within this network in aspects of learning or memory, whether or not these cells can also be distinguished by the genes they express remains unresolved. In addition, the changes in gene expression that accompany long-term memory formation within the MBs have not yet been studied by neuron type. Here, we address both issues by performing RNA sequencing on single cell types (harvested via patch pipets) within the MB. We discover that the expression of genes that encode cell surface receptors is sufficient to identify cell types and that a subset of these genes, required for sensory transduction in peripheral sensory neurons, is not only expressed within individual neurons of the MB in the central brain, but is also critical for memory formation. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  12. Cell type-specific gene expression of midbrain dopaminergic neurons reveals molecules involved in their vulnerability and protection.

    Chung, Chee Yeun; Seo, Hyemyung; Sonntag, Kai Christian; Brooks, Andrew; Lin, Ling; Isacson, Ole

    2005-07-01

    Molecular differences between dopamine (DA) neurons may explain why the mesostriatal DA neurons in the A9 region preferentially degenerate in Parkinson's disease (PD) and toxic models, whereas the adjacent A10 region mesolimbic and mesocortical DA neurons are relatively spared. To characterize innate physiological differences between A9 and A10 DA neurons, we determined gene expression profiles in these neurons in the adult mouse by laser capture microdissection, microarray analysis and real-time PCR. We found 42 genes relatively elevated in A9 DA neurons, whereas 61 genes were elevated in A10 DA neurons [> 2-fold; false discovery rate (FDR) neurotoxic or protective biochemical pathways. Three A9-elevated molecules [G-protein coupled inwardly rectifying K channel 2 (GIRK2), adenine nucleotide translocator 2 (ANT-2) and the growth factor IGF-1] and three A10-elevated peptides (GRP, CGRP and PACAP) were further examined in both alpha-synuclein overexpressing PC12 (PC12-alphaSyn) cells and rat primary ventral mesencephalic (VM) cultures exposed to MPP+ neurotoxicity. GIRK2-positive DA neurons were more vulnerable to MPP+ toxicity and overexpression of GIRK2 increased the vulnerability of PC12-alphaSyn cells to the toxin. Blocking of ANT decreased vulnerability to MPP+ in both cell culture systems. Exposing cells to IGF-1, GRP and PACAP decreased vulnerability of both cell types to MPP+, whereas CGRP protected PC12-alphaSyn cells but not primary VM DA neurons. These results indicate that certain differentially expressed molecules in A9 and A10 DA neurons may play key roles in their relative vulnerability to toxins and PD.

  13. Cell-Type-Specific Regulation of the Retinoic Acid Receptor Mediated by the Orphan Nuclear Receptor TLX†

    Kobayashi, Mime; Yu, Ruth T.; Yasuda, Kunio; Umesono, Kazuhiko

    2000-01-01

    Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor β (RARβ). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARβ2 promoter. The region surrounding the transcription start site of the avian RARβ2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARβ2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARβ gene in the eye. PMID:11073974

  14. Cell-type-specific regulation of the retinoic acid receptor mediated by the orphan nuclear receptor TLX.

    Kobayashi, M; Yu, R T; Yasuda, K; Umesono, K

    2000-12-01

    Malformations in the eye can be caused by either an excess or deficiency of retinoids. An early target gene of the retinoid metabolite, retinoic acid (RA), is that encoding one of its own receptors, the retinoic acid receptor beta (RARbeta). To better understand the mechanisms underlying this autologous regulation, we characterized the chick RARbeta2 promoter. The region surrounding the transcription start site of the avian RARbeta2 promoter is over 90% conserved with the corresponding region in mammals and confers strong RA-dependent transactivation in primary cultured embryonic retina cells. This response is selective for RAR but not retinoid X receptor-specific agonists, demonstrating a principal role for RAR(s) in retina cells. Retina cells exhibit a far higher sensitivity to RA than do fibroblasts or osteoblasts, a property we found likely due to expression of the orphan nuclear receptor TLX. Ectopic expression of TLX in fibroblasts resulted in increased sensitivity to RA induction, an effect that is conserved between chick and mammals. We have identified a cis element, the silencing element relieved by TLX (SET), within the RARbeta2 promoter region which confers TLX- and RA-dependent transactivation. These results indicate an important role for TLX in autologous regulation of the RARbeta gene in the eye.

  15. Cell-Type-Specific Circuit Connectivity of Hippocampal CA1 Revealed through Cre-Dependent Rabies Tracing

    Yanjun Sun

    2014-04-01

    Full Text Available We developed and applied a Cre-dependent, genetically modified rabies-based tracing system to map direct synaptic connections to specific CA1 neuron types in the mouse hippocampus. We found common inputs to excitatory and inhibitory CA1 neurons from CA3, CA2, the entorhinal cortex (EC, the medial septum (MS, and, unexpectedly, the subiculum. Excitatory CA1 neurons receive inputs from both cholinergic and GABAergic MS neurons, whereas inhibitory neurons receive a great majority of inputs from GABAergic MS neurons. Both cell types also receive weaker input from glutamatergic MS neurons. Comparisons of inputs to CA1 PV+ interneurons versus SOM+ interneurons showed similar strengths of input from the subiculum, but PV+ interneurons received much stronger input than SOM+ neurons from CA3, the EC, and the MS. Thus, rabies tracing identifies hippocampal circuit connections and maps how the different input sources to CA1 are distributed with different strengths on each of its constituent cell types.

  16. Cell-Type Specific Changes in Glial Morphology and Glucocorticoid Expression During Stress and Aging in the Medial Prefrontal Cortex

    Thomas E. Chan

    2018-05-01

    Full Text Available Repeated exposure to stressors is known to produce large-scale remodeling of neurons within the prefrontal cortex (PFC. Recent work suggests stress-related forms of structural plasticity can interact with aging to drive distinct patterns of pyramidal cell morphological changes. However, little is known about how other cellular components within PFC might be affected by these challenges. Here, we examined the effects of stress exposure and aging on medial prefrontal cortical glial subpopulations. Interestingly, we found no changes in glial morphology with stress exposure but a profound morphological change with aging. Furthermore, we found an upregulation of non-nuclear glucocorticoid receptors (GR with aging, while nuclear levels remained largely unaffected. Both changes are selective for microglia, with no stress or aging effect found in astrocytes. Lastly, we show that the changes found within microglia inversely correlated with the density of dendritic spines on layer III pyramidal cells. These findings suggest microglia play a selective role in synaptic health within the aging brain.

  17. An Atlas for Schistosoma mansoni Organs and Life-Cycle Stages Using Cell Type-Specific Markers and Confocal Microscopy

    Cogswell, Alexis; Williams, David L.; Newmark, Phillip A.

    2011-01-01

    Schistosomiasis (bilharzia) is a tropical disease caused by trematode parasites (Schistosoma) that affects hundreds of millions of people in the developing world. Currently only a single drug (praziquantel) is available to treat this disease, highlighting the importance of developing new techniques to study Schistosoma. While molecular advances, including RNA interference and the availability of complete genome sequences for two Schistosoma species, will help to revolutionize studies of these animals, an array of tools for visualizing the consequences of experimental perturbations on tissue integrity and development needs to be made widely available. To this end, we screened a battery of commercially available stains, antibodies and fluorescently labeled lectins, many of which have not been described previously for analyzing schistosomes, for their ability to label various cell and tissue types in the cercarial stage of S. mansoni. This analysis uncovered more than 20 new markers that label most cercarial tissues, including the tegument, the musculature, the protonephridia, the secretory system and the nervous system. Using these markers we present a high-resolution visual depiction of cercarial anatomy. Examining the effectiveness of a subset of these markers in S. mansoni adults and miracidia, we demonstrate the value of these tools for labeling tissues in a variety of life-cycle stages. The methodologies described here will facilitate functional analyses aimed at understanding fundamental biological processes in these parasites. PMID:21408085

  18. Cell and receptor type-specific alterations in markers of GABA neurotransmission in the prefrontal cortex of subjects with schizophrenia.

    Lewis, David A; Hashimoto, Takanori; Morris, Harvey M

    2008-10-01

    Impairments in cognitive control, such as those involved in working memory, are associated with dysfunction of the dorsolateral prefrontal cortex (DLPFC) in individuals with schizophrenia. This dysfunction appears to result, at least in part, from abnormalities in GABA-mediated neurotransmission. In this paper, we review recent findings indicating that the altered DLPFC circuitry in subjects with schizophrenia reflects changes in the expression of genes that encode selective presynaptic and postsynaptic components of GABA neurotransmission. Specifically, using a combination of methods, we found that subjects with schizophrenia exhibited expression deficits in GABA-related transcripts encoding presynaptic regulators of GABA neurotransmission, neuropeptide markers of specific subpopulations of GABA neurons, and certain subunits of the GABA(A) receptor. In particular, alterations in the expression of the neuropeptide somatostatin suggested that GABA neurotransmission is impaired in the Martinotti subset of GABA neurons that target the dendrites of pyramidal cells. In contrast, none of the GABA-related transcripts assessed to date were altered in the DLPFC of monkeys chronically exposed to antipsychotic medications, suggesting that the effects observed in the human studies reflect the disease process and not its treatment. In concert with previous findings, these data suggest that working memory dysfunction in schizophrenia may be attributable to altered GABA neurotransmission in specific DLPFC microcircuits.

  19. Triiodothyronine regulates angiogenic growth factor and cytokine secretion by isolated human decidual cells in a cell-type specific and gestational age-dependent manner.

    Vasilopoulou, E; Loubière, L S; Lash, G E; Ohizua, O; McCabe, C J; Franklyn, J A; Kilby, M D; Chan, S Y

    2014-06-01

    cell isolates were unaffected by T3 so changes in cell numbers could not account for any observed effects. In the first trimester, T3 decreased VEGF-A secretion by total decidual cells (P < 0.05) and increased angiopoietin-2 secretion by stromal-depleted cells (P < 0.05) but in the second trimester total decidual cells showed only increased angiogenin secretion (P < 0.05). In the first trimester, T3 reduced IL-10 secretion by total decidual cells (P < 0.05), and reduced granulocyte macrophage colony stimulating factor (P < 0.01), IL-8 (P < 0.05), IL-10 (P < 0.01), IL-1β (P < 0.05) and monocyte chemotactic protein -1 (P < 0.001) secretion by macrophages, but increased tumour necrosis factor-α secretion by stromal-depleted cells (P < 0.05) and increased IL-6 by uNK cells (P < 0.05). In contrast, in the second trimester T3 increased IL-10 secretion by total decidual cells (P < 0.01) but did not affect cytokine secretion by uNK cells and macrophages. Conditioned media from first trimester T3-treated total decidual cells and macrophages did not alter EVT invasion compared with untreated controls. Thus, treatment of decidual cells with T3 resulted in changes in both angiogenic growth factor and cytokine secretion in a cell type-specific and gestational age-dependent manner, with first trimester decidual macrophages being the most responsive to T3 treatment, but these changes in decidual cell secretome did not affect EVT invasion in vitro. Our results are based on in vitro findings and we cannot be certain if a similar response occurs in human pregnancy in vivo. Optimal maternal thyroid hormone concentrations could play a critical role in maintaining a balanced inflammatory response in early pregnancy to prevent fetal immune rejection and promote normal placental development through the regulation of the secretion of critical cytokines and angiogenic growth factors by human decidual cells. Our data suggest that there is an ontogenically determined regulatory 'switch' in T3

  20. Mapping Mammalian Cell-type-specific Transcriptional Regulatory Networks Using KD-CAGE and ChIP-seq Data in the TC-YIK Cell Line

    Lizio, Marina; Ishizu, Yuri; Itoh, Masayoshi; Lassmann, Timo; Hasegawa, Akira; Kubosaki, Atsutaka; Severin, Jessica; Kawaji, Hideya; Nakamura, Yukio; Suzuki, Harukazu; Hayashizaki, Yoshihide; Carninci, Piero; Forrest, Alistair R. R.

    2015-01-01

    Mammals are composed of hundreds of different cell types with specialized functions. Each of these cellular phenotypes are controlled by different combinations of transcription factors. Using a human non islet cell insulinoma cell line (TC-YIK) which expresses insulin and the majority of known pancreatic beta cell specific genes as an example, we describe a general approach to identify key cell-type-specific transcription factors (TFs) and their direct and indirect targets. By ranking all human TFs by their level of enriched expression in TC-YIK relative to a broad collection of samples (FANTOM5), we confirmed known key regulators of pancreatic function and development. Systematic siRNA mediated perturbation of these TFs followed by qRT-PCR revealed their interconnections with NEUROD1 at the top of the regulation hierarchy and its depletion drastically reducing insulin levels. For 15 of the TF knock-downs (KD), we then used Cap Analysis of Gene Expression (CAGE) to identify thousands of their targets genome-wide (KD-CAGE). The data confirm NEUROD1 as a key positive regulator in the transcriptional regulatory network (TRN), and ISL1, and PROX1 as antagonists. As a complimentary approach we used ChIP-seq on four of these factors to identify NEUROD1, LMX1A, PAX6, and RFX6 binding sites in the human genome. Examining the overlap between genes perturbed in the KD-CAGE experiments and genes with a ChIP-seq peak within 50 kb of their promoter, we identified direct transcriptional targets of these TFs. Integration of KD-CAGE and ChIP-seq data shows that both NEUROD1 and LMX1A work as the main transcriptional activators. In the core TRN (i.e., TF-TF only), NEUROD1 directly transcriptionally activates the pancreatic TFs HSF4, INSM1, MLXIPL, MYT1, NKX6-3, ONECUT2, PAX4, PROX1, RFX6, ST18, DACH1, and SHOX2, while LMX1A directly transcriptionally activates DACH1, SHOX2, PAX6, and PDX1. Analysis of these complementary datasets suggests the need for caution in interpreting Ch

  1. Correlation of mRNA and protein levels: Cell type-specific gene expression of cluster designation antigens in the prostate

    Deutsch Eric W

    2008-05-01

    Full Text Available Abstract Background: Expression levels of mRNA and protein by cell types exhibit a range of correlations for different genes. In this study, we compared levels of mRNA abundance for several cluster designation (CD genes determined by gene arrays using magnetic sorted and laser-capture microdissected human prostate cells with levels of expression of the respective CD proteins determined by immunohistochemical staining in the major cell types of the prostate – basal epithelial, luminal epithelial, stromal fibromuscular, and endothelial – and for prostate precursor/stem cells and prostate carcinoma cells. Immunohistochemical stains of prostate tissues from more than 50 patients were scored for informative CD antigen expression and compared with cell-type specific transcriptomes. Results: Concordance between gene and protein expression findings based on 'present' vs. 'absent' calls ranged from 46 to 68%. Correlation of expression levels was poor to moderate (Pearson correlations ranged from 0 to 0.63. Divergence between the two data types was most frequently seen for genes whose array signals exceeded background (> 50 but lacked immunoreactivity by immunostaining. This could be due to multiple factors, e.g. low levels of protein expression, technological sensitivities, sample processing, probe set definition or anatomical origin of tissue and actual biological differences between transcript and protein abundance. Conclusion: Agreement between these two very different methodologies has great implications for their respective use in both molecular studies and clinical trials employing molecular biomarkers.

  2. Staphylococcus aureus alpha-toxin mediates general and cell type-specific changes in metabolite concentrations of immortalized human airway epithelial cells.

    Philipp Gierok

    Full Text Available Staphylococcus aureus alpha-toxin (Hla is a potent pore-forming cytotoxin that plays an important role in the pathogenesis of S. aureus infections, including pneumonia. The impact of Hla on the dynamics of the metabolome in eukaryotic host cells has not been investigated comprehensively. Using 1H-NMR, GC-MS and HPLC-MS, we quantified the concentrations of 51 intracellular metabolites and assessed alterations in the amount of 25 extracellular metabolites in the two human bronchial epithelial cell lines S9 and 16HBE14o- under standard culture conditions and after treatment with sub-lethal amounts (2 µg/ml of recombinant Hla (rHla in a time-dependent manner. Treatment of cells with rHla caused substantial decreases in the concentrations of intracellular metabolites from different metabolic pathways in both cell lines, including ATP and amino acids. Concomitant increases in the extracellular concentrations were detected for various intracellular compounds, including nucleotides, glutathione disulfide and NAD+. Our results indicate that rHla has a major impact on the metabolome of eukaryotic cells as a consequence of direct rHla-mediated alterations in plasma membrane permeability or indirect effects mediated by cellular signalling. However, cell-specific changes also were observed. Glucose consumption and lactate production rates suggest that the glycolytic activity of S9 cells, but not of 16HBE14o- cells, is increased in response to rHla. This could contribute to the observed higher level of resistance of S9 cells against rHla-induced membrane damage.

  3. Gene expression relationship between prostate cancer cells of Gleason 3, 4 and normal epithelial cells as revealed by cell type-specific transcriptomes

    Pascal, Laura E; Liu, Alvin Y; Vêncio, Ricardo ZN; Page, Laura S; Liebeskind, Emily S; Shadle, Christina P; Troisch, Pamela; Marzolf, Bruz; True, Lawrence D; Hood, Leroy E

    2009-01-01

    Prostate cancer cells in primary tumors have been typed CD10 - /CD13 - /CD24 hi /CD26 + /CD38 lo /CD44 - /CD104 - . This CD phenotype suggests a lineage relationship between cancer cells and luminal cells. The Gleason grade of tumors is a descriptive of tumor glandular differentiation. Higher Gleason scores are associated with treatment failure. CD26 + cancer cells were isolated from Gleason 3+3 (G3) and Gleason 4+4 (G4) tumors by cell sorting, and their gene expression or transcriptome was determined by Affymetrix DNA array analysis. Dataset analysis was used to determine gene expression similarities and differences between G3 and G4 as well as to prostate cancer cell lines and histologically normal prostate luminal cells. The G3 and G4 transcriptomes were compared to those of prostatic cell types of non-cancer, which included luminal, basal, stromal fibromuscular, and endothelial. A principal components analysis of the various transcriptome datasets indicated a closer relationship between luminal and G3 than luminal and G4. Dataset comparison also showed that the cancer transcriptomes differed substantially from those of prostate cancer cell lines. Genes differentially expressed in cancer are potential biomarkers for cancer detection, and those differentially expressed between G3 and G4 are potential biomarkers for disease stratification given that G4 cancer is associated with poor outcomes. Differentially expressed genes likely contribute to the prostate cancer phenotype and constitute the signatures of these particular cancer cell types

  4. Cell type specific DNA methylation in cord blood: A 450K-reference data set and cell count-based validation of estimated cell type composition.

    Gervin, Kristina; Page, Christian Magnus; Aass, Hans Christian D; Jansen, Michelle A; Fjeldstad, Heidi Elisabeth; Andreassen, Bettina Kulle; Duijts, Liesbeth; van Meurs, Joyce B; van Zelm, Menno C; Jaddoe, Vincent W; Nordeng, Hedvig; Knudsen, Gunn Peggy; Magnus, Per; Nystad, Wenche; Staff, Anne Cathrine; Felix, Janine F; Lyle, Robert

    2016-09-01

    Epigenome-wide association studies of prenatal exposure to different environmental factors are becoming increasingly common. These studies are usually performed in umbilical cord blood. Since blood comprises multiple cell types with specific DNA methylation patterns, confounding caused by cellular heterogeneity is a major concern. This can be adjusted for using reference data consisting of DNA methylation signatures in cell types isolated from blood. However, the most commonly used reference data set is based on blood samples from adult males and is not representative of the cell type composition in neonatal cord blood. The aim of this study was to generate a reference data set from cord blood to enable correct adjustment of the cell type composition in samples collected at birth. The purity of the isolated cell types was very high for all samples (>97.1%), and clustering analyses showed distinct grouping of the cell types according to hematopoietic lineage. We explored whether this cord blood and the adult peripheral blood reference data sets impact the estimation of cell type composition in cord blood samples from an independent birth cohort (MoBa, n = 1092). This revealed significant differences for all cell types. Importantly, comparison of the cell type estimates against matched cell counts both in the cord blood reference samples (n = 11) and in another independent birth cohort (Generation R, n = 195), demonstrated moderate to high correlation of the data. This is the first cord blood reference data set with a comprehensive examination of the downstream application of the data through validation of estimated cell types against matched cell counts.

  5. Genome-Wide Progesterone Receptor Binding: Cell Type-Specific and Shared Mechanisms in T47D Breast Cancer Cells and Primary Leiomyoma Cells

    Huang, Lei; Owen, Jonas K.; Xie, Anna; Navarro, Antonia; Monsivais, Diana; Coon V, John S.; Kim, J. Julie; Dai, Yang; Bulun, Serdar E.

    2012-01-01

    Background Progesterone, via its nuclear receptor (PR), exerts an overall tumorigenic effect on both uterine fibroid (leiomyoma) and breast cancer tissues, whereas the antiprogestin RU486 inhibits growth of these tissues through an unknown mechanism. Here, we determined the interaction between common or cell-specific genome-wide binding sites of PR and mRNA expression in RU486-treated uterine leiomyoma and breast cancer cells. Principal Findings ChIP-sequencing revealed 31,457 and 7,034 PR-binding sites in breast cancer and uterine leiomyoma cells, respectively; 1,035 sites overlapped in both cell types. Based on the chromatin-PR interaction in both cell types, we statistically refined the consensus progesterone response element to G•ACA• • •TGT•C. We identified two striking differences between uterine leiomyoma and breast cancer cells. First, the cis-regulatory elements for HSF, TEF-1, and C/EBPα and β were statistically enriched at genomic RU486/PR-targets in uterine leiomyoma, whereas E2F, FOXO1, FOXA1, and FOXF sites were preferentially enriched in breast cancer cells. Second, 51.5% of RU486-regulated genes in breast cancer cells but only 6.6% of RU486-regulated genes in uterine leiomyoma cells contained a PR-binding site within 5 kb from their transcription start sites (TSSs), whereas 75.4% of RU486-regulated genes contained a PR-binding site farther than 50 kb from their TSSs in uterine leiomyoma cells. RU486 regulated only seven mRNAs in both cell types. Among these, adipophilin (PLIN2), a pro-differentiation gene, was induced via RU486 and PR via the same regulatory region in both cell types. Conclusions Our studies have identified molecular components in a RU486/PR-controlled gene network involved in the regulation of cell growth, cell migration, and extracellular matrix function. Tissue-specific and common patterns of genome-wide PR binding and gene regulation may determine the therapeutic effects of antiprogestins in uterine fibroids and

  6. Global mapping of cell type-specific open chromatin by FAIRE-seq reveals the regulatory role of the NFI family in adipocyte differentiation.

    Hironori Waki

    2011-10-01

    Full Text Available Identification of regulatory elements within the genome is crucial for understanding the mechanisms that govern cell type-specific gene expression. We generated genome-wide maps of open chromatin sites in 3T3-L1 adipocytes (on day 0 and day 8 of differentiation and NIH-3T3 fibroblasts using formaldehyde-assisted isolation of regulatory elements coupled with high-throughput sequencing (FAIRE-seq. FAIRE peaks at the promoter were associated with active transcription and histone modifications of H3K4me3 and H3K27ac. Non-promoter FAIRE peaks were characterized by H3K4me1+/me3-, the signature of enhancers, and were largely located in distal regions. The non-promoter FAIRE peaks showed dynamic change during differentiation, while the promoter FAIRE peaks were relatively constant. Functionally, the adipocyte- and preadipocyte-specific non-promoter FAIRE peaks were, respectively, associated with genes up-regulated and down-regulated by differentiation. Genes highly up-regulated during differentiation were associated with multiple clustered adipocyte-specific FAIRE peaks. Among the adipocyte-specific FAIRE peaks, 45.3% and 11.7% overlapped binding sites for, respectively, PPARγ and C/EBPα, the master regulators of adipocyte differentiation. Computational motif analyses of the adipocyte-specific FAIRE peaks revealed enrichment of a binding motif for nuclear family I (NFI transcription factors. Indeed, ChIP assay showed that NFI occupy the adipocyte-specific FAIRE peaks and/or the PPARγ binding sites near PPARγ, C/EBPα, and aP2 genes. Overexpression of NFIA in 3T3-L1 cells resulted in robust induction of these genes and lipid droplet formation without differentiation stimulus. Overexpression of dominant-negative NFIA or siRNA-mediated knockdown of NFIA or NFIB significantly suppressed both induction of genes and lipid accumulation during differentiation, suggesting a physiological function of these factors in the adipogenic program. Together, our

  7. Cell-type-specific and differentiation-status-dependent variations in cytotoxicity of tributyltin in cultured rat cerebral neurons and astrocytes.

    Oyanagi, Koshi; Tashiro, Tomoko; Negishi, Takayuki

    2015-08-01

    Tributyltin (TBT) is an organotin used as an anti-fouling agent for fishing nets and ships and it is a widespread environmental contaminant at present. There is an increasing concern about imperceptible but serious adverse effect(s) of exposure to chemicals existing in the environment on various organs and their physiological functions, e.g. brain and mental function. Here, so as to contribute to improvement of and/or advances in in vitro cell-based assay systems for evaluating brain-targeted adverse effect of chemicals, we tried to evaluate cell-type-specific and differentiation-status-dependent variations in the cytotoxicity of TBT towards neurons and astrocytes using the four culture systems differing in the relative abundance of these two types of cells; primary neuron culture (> 95% neurons), primary neuron-astrocyte (2 : 1) mix culture, primary astrocyte culture (> 95% astrocytes), and passaged astrocyte culture (100% proliferative astrocytes). Cell viability was measured at 48 hr after exposure to TBT in serum-free medium. IC50's of TBT were 198 nM in primary neuron culture, 288 nM in primary neuron-astrocyte mix culture, 2001 nM in primary astrocyte culture, and 1989 nM in passaged astrocyte culture. Furthermore, in primary neuron-astrocyte mix culture, vulnerability of neurons cultured along with astrocytes to TBT toxicity was lower than that of neurons cultured purely in primary neuron culture. On the other hand, astrocytes in primary neuron-astrocyte mix culture were considered to be more vulnerable to TBT than those in primary or passaged astrocyte culture. The present study demonstrated variable cytotoxicity of TBT in neural cells depending on the culture condition.

  8. Comparative Analysis of WUSCHEL-Related Homeobox Genes Revealed Their Parent-of-Origin and Cell Type-Specific Expression Pattern During Early Embryogenesis in Tobacco

    Xuemei Zhou

    2018-03-01

    Full Text Available WUSCHEL-related homeobox (WOX gene is a plant-specific clade of homeobox transcription factors. Increasing evidences reveal that WOXs play critical roles in early embryogenesis, which involves zygote development, initiation of zygote division, and apical or basal cell lineage establishment. However, how WOXs regulate these developmental events remains largely unknown, and even detailed expression pattern in gametes and early proembryos is not yet available. Here, 13 WOX family genes were identified in Nicotiana tabacum genome. Comparative analysis of 13 WOX family genes with their homologs in Arabidopsis thaliana reveals relatively conserved expression pattern of WUS and WOX5 in shoot/root apical meristem. Whereas variations were also found, e.g., lacking homolog of WOX8 (a marker for suspensor cell in tobacco genome and the expression of WOX2/WOX9 in both apical cell and basal cell. Transient transcriptional activity analysis revealed that WOXs in WUS clade have repressive activities for their target's transcription, whereas WOXs in ancient and intermediate clade have activation activities, giving a molecular basis for the phylogenetic classification of tobacco WOXs into three major clades. Expression pattern analysis revealed that some WOXs (e.g., WOX 13a expressed in both male and female gametes and some WOXs (e.g., WOX 11 and WOX 13b displayed the characteristics of parent-of-origin genes. Interestingly, some WOXs (e.g., WOX2 and WOX9, which are essential for early embryo patterning, were de novo transcribed in zygote, indicating relevant mechanism for embryo pattern formation is only established in zygote right after fertilization and not carried in by gametes. We also found that most WOXs displayed a stage-specific and cell type-specific expression pattern. Taken together, this work provides a detailed landscape of WOXs in tobacco during fertilization and early embryogenesis, which will facilitate the understanding of their specific roles

  9. Cell type specific DNA methylation in cord blood: A 450K-reference data set and cell count-based validation of estimated cell type composition

    Gervin, K. (Kristina); Page, C.M. (Christian Magnus); H.C.D. Aass (Hans Christian Dalsbotten); M.A.E. Jansen (Michelle); Fjeldstad, H.E. (Heidi Elisabeth); B.K. Andreassen (Bettina Kulle); L. Duijts (Liesbeth); J.B.J. van Meurs (Joyce); M.C. van Zelm (Menno); V.W.V. Jaddoe (Vincent); Nordeng, H. (Hedvig); Knudsen, G.P. (Gunn Peggy); P. Magnus (Per); W. Nystad (Wenche); Staff, A.C. (Anne Cathrine); J.F. Felix (Janine); R. Lyle (Robert)

    2016-01-01

    textabstractEpigenome-wide association studies of prenatal exposure to different environmental factors are becoming increasingly common. These studies are usually performed in umbilical cord blood. Since blood comprises multiple cell types with specific DNA methylation patterns, confounding caused

  10. Cell type-specific recruitment of Drosophila Lin-7 to distinct MAGUK-based protein complexes defines novel roles for Sdt and Dlg-S97.

    Bachmann, André; Timmer, Marco; Sierralta, Jimena; Pietrini, Grazia; Gundelfinger, Eckart D; Knust, Elisabeth; Thomas, Ulrich

    2004-04-15

    Stardust (Sdt) and Discs-Large (Dlg) are membrane-associated guanylate kinases (MAGUKs) involved in the organization of supramolecular protein complexes at distinct epithelial membrane compartments in Drosophila. Loss of either Sdt or Dlg affects epithelial development with severe effects on apico-basal polarity. Moreover, Dlg is required for the structural and functional integrity of synaptic junctions. Recent biochemical and cell culture studies have revealed that various mammalian MAGUKs can interact with mLin-7/Veli/MALS, a small PDZ-domain protein. To substantiate these findings for their in vivo significance with regard to Sdt- and Dlg-based protein complexes, we analyzed the subcellular distribution of Drosophila Lin-7 (DLin-7) and performed genetic and biochemical assays to characterize its interaction with either of the two MAGUKs. In epithelia, Sdt mediates the recruitment of DLin-7 to the subapical region, while at larval neuromuscular junctions, a particular isoform of Dlg, Dlg-S97, is required for postsynaptic localization of DLin-7. Ectopic expression of Dlg-S97 in epithelia, however, was not sufficient to induce a redistribution of DLin-7. These results imply that the recruitment of DLin-7 to MAGUK-based protein complexes is defined by cell-type specific mechanisms and that DLin-7 acts downstream of Sdt in epithelia and downstream of Dlg at synapses.

  11. p53 shapes genome-wide and cell type-specific changes in microRNA expression during the human DNA damage response.

    Hattori, Hiroyoshi; Janky, Rekin's; Nietfeld, Wilfried; Aerts, Stein; Madan Babu, M; Venkitaraman, Ashok R

    2014-01-01

    The human DNA damage response (DDR) triggers profound changes in gene expression, whose nature and regulation remain uncertain. Although certain micro-(mi)RNA species including miR34, miR-18, miR-16 and miR-143 have been implicated in the DDR, there is as yet no comprehensive description of genome-wide changes in the expression of miRNAs triggered by DNA breakage in human cells. We have used next-generation sequencing (NGS), combined with rigorous integrative computational analyses, to describe genome-wide changes in the expression of miRNAs during the human DDR. The changes affect 150 of 1523 miRNAs known in miRBase v18 from 4-24 h after the induction of DNA breakage, in cell-type dependent patterns. The regulatory regions of the most-highly regulated miRNA species are enriched in conserved binding sites for p53. Indeed, genome-wide changes in miRNA expression during the DDR are markedly altered in TP53-/- cells compared to otherwise isogenic controls. The expression levels of certain damage-induced, p53-regulated miRNAs in cancer samples correlate with patient survival. Our work reveals genome-wide and cell type-specific alterations in miRNA expression during the human DDR, which are regulated by the tumor suppressor protein p53. These findings provide a genomic resource to identify new molecules and mechanisms involved in the DDR, and to examine their role in tumor suppression and the clinical outcome of cancer patients.

  12. Morphological changes in different populations of bladder afferent neurons detected by herpes simplex virus (HSV) vectors with cell-type-specific promoters in mice with spinal cord injury.

    Shimizu, Nobutaka; Doyal, Mark F; Goins, William F; Kadekawa, Katsumi; Wada, Naoki; Kanai, Anthony J; de Groat, William C; Hirayama, Akihide; Uemura, Hirotsugu; Glorioso, Joseph C; Yoshimura, Naoki

    2017-11-19

    Functional and morphological changes in C-fiber bladder afferent pathways are reportedly involved in neurogenic detrusor overactivity (NDO) after spinal cord injury (SCI). This study examined the morphological changes in different populations of bladder afferent neurons after SCI using replication-defective herpes simplex virus (HSV) vectors encoding the mCherry reporter driven by neuronal cell-type-specific promoters. Spinal intact (SI) and SCI mice were injected into the bladder wall with HSV mCherry vectors driven by the cytomegalovirus (CMV) promoter, CGRP promoter, TRPV1 promoter or neurofilament 200 (NF200) promoter. Two weeks after vector inoculation into the bladder wall, L1 and L6 dorsal root ganglia (DRG) were removed bilaterally for immunofluorescent staining using anti-mCherry antibody. The number of CMV promoter vector-labeled neurons was not altered after SCI. The number of CGRP and TRPV1 promoter vector-labeled neurons was significantly increased whereas the number of NF200 vector-labeled neurons was decreased in L6 DRG after SCI. The median size of CGRP promoter-labeled C-fiber neurons was increased from 247.0 in SI mice to 271.3μm 2 in SCI mice whereas the median cell size of TRPV1 promoter vector-labeled neurons was decreased from 245.2 in SI mice to 216.5μm 2 in SCI mice. CGRP and TRPV1 mRNA levels of laser-captured bladder afferent neurons labeled with Fast Blue were significantly increased in SCI mice compared to SI mice. Thus, using a novel HSV vector-mediated neuronal labeling technique, we found that SCI induces expansion of the CGRP- and TRPV1-expressing C-fiber cell population, which could contribute to C-fiber afferent hyperexcitability and NDO after SCI. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

  13. Genome-Wide Maps of m6A circRNAs Identify Widespread and Cell-Type-Specific Methylation Patterns that Are Distinct from mRNAs

    Chan Zhou

    2017-08-01

    Full Text Available N6-methyladenosine (m6A is the most abundant internal modification of mRNAs and is implicated in all aspects of post-transcriptional RNA metabolism. However, little is known about m6A modifications to circular (circ RNAs. We developed a computational pipeline (AutoCirc that, together with depletion of ribosomal RNA and m6A immunoprecipitation, defined thousands of m6A circRNAs with cell-type-specific expression. The presence of m6A circRNAs is corroborated by interaction between circRNAs and YTHDF1/YTHDF2, proteins that read m6A sites in mRNAs, and by reduced m6A levels upon depletion of METTL3, the m6A writer. Despite sharing m6A readers and writers, m6A circRNAs are frequently derived from exons that are not methylated in mRNAs, whereas mRNAs that are methylated on the same exons that compose m6A circRNAs exhibit less stability in a process regulated by YTHDF2. These results expand our understanding of the breadth of m6A modifications and uncover regulation of circRNAs through m6A modification.

  14. Complexity of type-specific 56 kDa antigen CD4 T-cell epitopes of Orientia tsutsugamushi strains causing scrub typhus in India.

    Arunachalam Ramaiah

    Full Text Available Orientia tsutsugamushi (Ots is an obligate, intracellular, mite-transmitted human pathogen which causes scrub typhus. Understanding the diversity of Ots antigens is essential for designing specific diagnostic assays and efficient vaccines. The protective immunodominant type-specific 56 kDa antigen (TSA of Ots varies locally and across its geographic distribution. TSA contains four hypervariable domains. We bioinformatically analyzed 345 partial sequences of TSA available from India, most of which contain only the three variable domains (VDI-III and three spacer conserved domains (SVDI, SVDII/III, SVDIII. The total number (152 of antigenic types (amino acid variants varied from 14-36 in the six domains of TSA that we studied. Notably, 55% (787/1435 of the predicted CD4 T-cell epitopes (TCEs from all the six domains had high binding affinities (HBA to at least one of the prevalent Indian human leukocyte antigen (HLA alleles. A surprisingly high proportion (61% of such TCEs were from spacer domains; indeed 100% of the CD4 TCEs in the SVDI were HBA. TSA sequences from India had more antigenic types (AT than TSA from Korea. Overall, >90% of predicted CD4 TCEs from spacer domains were predicted to have HBA against one or more prevalent HLA types from Indian, Korean, Asia-Pacific region or global population data sets, while only <50% of CD4 TCEs in variable domains exhibited such HBA. The phylogenetically and immunologically important amino acids in the conserved spacer domains were identified. Our results suggest that the conserved spacer domains are predicted to be functionally more important than previously appreciated in immune responses to Ots infections. Changes occurring at the TCE level of TSA may contribute to the wide range of pathogenicity of Ots in humans and mouse models. CD4 T-cell functional experiments are needed to assess the immunological significance of these HBA spacer domains and their role in clearance of Ots from Indian patients.

  15. Nuclear and cellular expression data from the whole 16-cell stage Arabidopsis thaliana embryo and a cell type-specific expression atlas of the early Arabidopsis embryo

    Palovaara, J.P.J.

    2017-01-01

    SuperSeries contain expression data from the nuclei of cell types involved in patterning events, with focus on root apical stem cell formation, at 16-cell stage, early globular stage and late globular stage in the early Arabidopsis embryo (atlas). Expression data comparing nuclear and cellular RNA

  16. Biological adhesion of the flatworm Macrostomum lignano relies on a duo-gland system and is mediated by a cell type-specific intermediate filament protein.

    Lengerer, Birgit; Pjeta, Robert; Wunderer, Julia; Rodrigues, Marcelo; Arbore, Roberto; Schärer, Lukas; Berezikov, Eugene; Hess, Michael W; Pfaller, Kristian; Egger, Bernhard; Obwegeser, Sabrina; Salvenmoser, Willi; Ladurner, Peter

    2014-02-12

    mediated changes of the anchor cell morphology are comparable to situations observed in human gut epithelia. Therefore, our current findings and future investigations using this powerful flatworm model system might contribute to a better understanding of the function of intermediate filaments and their associated human diseases.

  17. Enterovirus strain and type-specific differences in growth kinetics and virus-induced cell destruction in human pancreatic duct epithelial HPDE cells.

    Smura, Teemu; Natri, Olli; Ylipaasto, Petri; Hellman, Marika; Al-Hello, Haider; Piemonti, Lorenzo; Roivainen, Merja

    2015-12-02

    Enterovirus infections have been suspected to be involved in the development of type 1 diabetes. However, the pathogenetic mechanism of enterovirus-induced type 1 diabetes is not known. Pancreatic ductal cells are closely associated with pancreatic islets. Therefore, enterovirus infections in ductal cells may also affect beta-cells and be involved in the induction of type 1 diabetes. The aim of this study was to assess the ability of different enterovirus strains to infect, replicate and produce cytopathic effect in human pancreatic ductal cells. Furthermore, the viral factors that affect these capabilities were studied. The pancreatic ductal cells were highly susceptible to enterovirus infections. Both viral growth and cytolysis were detected for several enterovirus serotypes. However, the viral growth and capability to induce cytopathic effect (cpe) did not correlate completely. Some of the virus strains replicated in ductal cells without apparent cpe. Furthermore, there were strain-specific differences in the growth kinetics and the ability to cause cpe within some serotypes. Viral adaptation experiments were carried out to study the potential genetic determinants behind these phenotypic differences. The blind-passage of non-lytic CV-B6-Schmitt strain in HPDE-cells resulted in lytic phenotype and increased progeny production. This was associated with the substitution of a single amino acid (K257E) in the virus capsid protein VP1 and the viral ability to use decay accelerating factor (DAF) as a receptor. This study demonstrates considerable plasticity in the cell tropism, receptor usage and cytolytic properties of enteroviruses and underlines the strong effect of single or few amino acid substitutions in cell tropism and lytic capabilities of a given enterovirus. Since ductal cells are anatomically close to pancreatic islets, the capability of enteroviruses to infect and destroy pancreatic ductal cells may also implicate in respect to enterovirus induced type 1

  18. Cell-type specific short-term plasticity at auditory nerve synapses controls feed-forward inhibition in the dorsal cochlear nucleus

    Miloslav eSedlacek

    2014-07-01

    Full Text Available Feedforward inhibition represents a powerful mechanism by which control of the timing and fidelity of action potentials in local synaptic circuits of various brain regions is achieved. In the cochlear nucleus, the auditory nerve provides excitation to both principal neurons and inhibitory interneurons. Here, we investigated the synaptic circuit associated with fusiform cells (FCs, principal neurons of the dorsal cochlear nucleus (DCN that receive excitation from auditory nerve fibers and inhibition from tuberculoventral cells (TVCs on their basal dendrites in the deep layer of DCN. Despite the importance of these inputs in regulating fusiform cell firing behavior, the mechanisms determining the balance of excitation and feed-forward inhibition in this circuit are not well understood. Therefore, we examined the timing and plasticity of auditory nerve driven feed-forward inhibition (FFI onto FCs. We find that in some FCs, excitatory and inhibitory components of feed-forward inhibition had the same stimulation thresholds indicating they could be triggered by activation of the same fibers. In other FCs, excitation and inhibition exhibit different stimulus thresholds, suggesting FCs and TVCs might be activated by different sets of fibers. In addition we find that during repetitive activation, synapses formed by the auditory nerve onto TVCs and FCs exhibit distinct modes of short-term plasticity. Feed-forward inhibitory post-synaptic currents (IPSCs in FCs exhibit short-term depression because of prominent synaptic depression at the auditory nerve-TVC synapse. Depression of this feedforward inhibitory input causes a shift in the balance of fusiform cell synaptic input towards greater excitation and suggests that fusiform cell spike output will be enhanced by physiological patterns of auditory nerve activity.

  19. Cell-type specific short-term plasticity at auditory nerve synapses controls feed-forward inhibition in the dorsal cochlear nucleus.

    Sedlacek, Miloslav; Brenowitz, Stephan D

    2014-01-01

    Feed-forward inhibition (FFI) represents a powerful mechanism by which control of the timing and fidelity of action potentials in local synaptic circuits of various brain regions is achieved. In the cochlear nucleus, the auditory nerve provides excitation to both principal neurons and inhibitory interneurons. Here, we investigated the synaptic circuit associated with fusiform cells (FCs), principal neurons of the dorsal cochlear nucleus (DCN) that receive excitation from auditory nerve fibers and inhibition from tuberculoventral cells (TVCs) on their basal dendrites in the deep layer of DCN. Despite the importance of these inputs in regulating fusiform cell firing behavior, the mechanisms determining the balance of excitation and FFI in this circuit are not well understood. Therefore, we examined the timing and plasticity of auditory nerve driven FFI onto FCs. We find that in some FCs, excitatory and inhibitory components of FFI had the same stimulation thresholds indicating they could be triggered by activation of the same fibers. In other FCs, excitation and inhibition exhibit different stimulus thresholds, suggesting FCs and TVCs might be activated by different sets of fibers. In addition, we find that during repetitive activation, synapses formed by the auditory nerve onto TVCs and FCs exhibit distinct modes of short-term plasticity. Feed-forward inhibitory post-synaptic currents (IPSCs) in FCs exhibit short-term depression because of prominent synaptic depression at the auditory nerve-TVC synapse. Depression of this feedforward inhibitory input causes a shift in the balance of fusiform cell synaptic input towards greater excitation and suggests that fusiform cell spike output will be enhanced by physiological patterns of auditory nerve activity.

  20. The M-current contributes to high threshold membrane potential oscillations in a cell type-specific way in the pedunculopontine nucleus of mice

    Csilla eBordas

    2015-04-01

    Full Text Available The pedunculopontine nucleus is known as a cholinergic nucleus of the reticular activating system, participating in regulation of sleep and wakefulness. Besides cholinergic neurons, it consists of GABAergic and glutamatergic neurons as well. According to classical and recent studies, more subgroups of neurons were defined. Groups based on the neurotransmitter released by a neuron are not homogenous, but can be further subdivided.The PPN neurons do not only provide cholinergic and non-cholinergic inputs to several subcortical brain areas but they are also targets of cholinergic and other different neuromodulatory actions. Although cholinergic neuromodulation has been already investigated in the nucleus, one of its characteristic targets, the M-type potassium current has not been described yet.Using slice electrophysiology, we provide evidence in the present work that cholinergic neurons possess M-current, whereas GABAergic neurons lack it. The M-current contributes to certain functional differences of cholinergic and GABAergic neurons, as spike frequency adaptation, action potential firing frequency or the amplitude difference of medium afterhyperpolarizations. Furthermore, we showed that high threshold membrane potential oscillation with high power, around 20 Hz frequency is a functional property of almost all cholinergic cells, whereas GABAergic neurons have only low amplitude oscillations. Blockade of the M-current abolished the oscillatory activity at 20 Hz, and largely diminished it at other frequencies.Taken together, the M-current seems to be characteristic for PPN cholinergic neurons. It provides a possibility for modulating gamma band activity of these cells, thus contributing to neuromodulatory regulation of the reticular activating system.

  1. Biological adhesion of the flatworm Macrostomum lignano relies on a duo-gland system and is mediated by a cell type-specific intermediate filament protein

    Lengerer, Birgit; Pjeta, Robert; Wunderer, Julia; Rodrigues, Marcelo; Arbore, Roberto; Schaerer, Lukas; Berezikov, Eugene; Hess, Michael W.; Pfaller, Kristian; Egger, Bernhard; Obwegeser, Sabrina; Salvenmoser, Willi; Ladurner, Peter

    2014-01-01

    Background: Free-living flatworms, in both marine and freshwater environments, are able to adhere to and release from a substrate several times within a second. This reversible adhesion relies on adhesive organs comprised of three cell types: an adhesive gland cell, a releasing gland cell, and an

  2. Cell-type specific role of the RNA-binding protein, NONO, in the DNA double-strand break response in the mouse testes.

    Li, Shuyi; Shu, Feng-Jue; Li, Zhentian; Jaafar, Lahcen; Zhao, Shourong; Dynan, William S

    2017-03-01

    The tandem RNA recognition motif protein, NONO, was previously identified as a candidate DNA double-strand break (DSB) repair factor in a biochemical screen for proteins with end-joining stimulatory activity. Subsequent work showed that NONO and its binding partner, SFPQ, have many of the properties expected for bona fide repair factors in cell-based assays. Their contribution to the DNA damage response in intact tissue in vivo has not, however, been demonstrated. Here we compare DNA damage sensitivity in the testes of wild-type mice versus mice bearing a null allele of the NONO homologue (Nono gt ). In wild-type mice, NONO protein was present in Sertoli, peritubular myoid, and interstitial cells, with an increase in expression following induction of DNA damage. As expected for the product of an X-linked gene, NONO was not detected in germ cells. The Nono gt/0 mice had at most a mild testis developmental phenotype in the absence of genotoxic stress. However, following irradiation at sublethal, 2-4 Gy doses, Nono gt/0 mice displayed a number of indicators of radiosensitivity as compared to their wild-type counterparts. These included higher levels of persistent DSB repair foci, increased numbers of apoptotic cells in the seminiferous tubules, and partial degeneration of the blood-testis barrier. There was also an almost complete loss of germ cells at later times following irradiation, evidently arising as an indirect effect reflecting loss of stromal support. Results demonstrate a role for NONO protein in protection against direct and indirect biological effects of ionizing radiation in the whole animal. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Construction of an Aptamer-SiRNA Chimera-Modified Tissue-Engineered Blood Vessel for Cell-Type-Specific Capture and Delivery.

    Chen, Wen; Zeng, Wen; Sun, Jiansen; Yang, Mingcan; Li, Li; Zhou, Jingting; Wu, Yangxiao; Sun, Jun; Liu, Ge; Tang, Rui; Tan, Ju; Zhu, Chuhong

    2015-06-23

    The application of tissue-engineered blood vessels (TEBVs) is the main developmental direction of vascular replacement therapy. Due to few and/or dysfunctional endothelial progenitor cells (EPCs), it is difficult to successfully construct EPC capture TEBVs in diabetes. RNA has a potential application in cell protection and diabetes treatment, but poor specificity and low efficiency of RNA transfection in vivo limit the application of RNA. On the basis of an acellular vascular matrix, we propose an aptamer-siRNA chimera-modified TEBV that can maintain a satisfactory patency in diabetes. This TEBV consists of two parts, CD133-adenosine kinase (ADK) chimeras and a TEBV scaffold. Our results showed that CD133-ADK chimeras could selectively capture the CD133-positive cells in vivo, and then captured cells can internalize the bound chimeras to achieve RNA self-transfection. Subsequently, CD133-ADK chimeras were cut into ADK siRNA by a dicer, resulting in depletion of ADK. An ADK-deficient cell may act as a bioreactor that sustainably releases adenosine. To reduce nonspecific RNA transfection, we increased the proportion of HAuCl4 during the material preparation, through which the transfection capacity of polyethylenimine (PEI)/polyethylene glycol (PEG)-capped gold nanoparticles (PEI/PEG-AuNPs) was significantly decreased and the ability of TEBV to resist tensile and liquid shear stress was greatly enhanced. PEG and 2'-O-methyl modification was used to enhance the in vivo stability of RNA chimeras. At day 30 postgrafting, the patency rate of CD133-ADK chimera-modified TEBVs reached 90% in diabetic rats and good endothelialization was observed.

  4. Expression of the Ly-6 family proteins Lynx1 and Ly6H in the rat brain is compartmentalized, cell-type specific, and developmentally regulated

    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...

  5. Type-specific detection of high-risk human papillomavirus (HPV) in self-sampled cervicovaginal cells applied to FTA elute cartridge.

    Gustavsson, Inger; Sanner, Karin; Lindell, Monica; Strand, Anders; Olovsson, Matts; Wikström, Ingrid; Wilander, Erik; Gyllensten, Ulf

    2011-08-01

    Most procedures for self-sampling of cervical cells are based on liquid-based media for transportation and storage. An alternative is to use a solid support, such as dry filter paper media. To evaluate if self-sampling of cervicovaginal fluid using a cytobrush (Viba-brush; Rovers Medical Devices B.V., Oss, The Netherlands) and a solid support such as the Whatman Indicating FTA Elute cartridge (GE Healthcare, United Kingdom) can be used for reliable typing of human papillomavirus (HPV), as compared to cervical samples obtained by a physician using a cytobrush and the indicating FTA Elute Micro card and biopsy analysis. A total of 50 women with a previous high-risk (HR) HPV positive test were invited to perform self-sampling using the Viba-brush and the FTA cartridge and thereafter a physician obtained a cervical sample using the cytobrush and a FTA card, together with a cervical biopsy for histology and HPV typing. Detection of HR-HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58 and 59 was performed using three multiplex real-time polymerase chain reaction (PCR) assays. All samples contained sufficient amounts of genomic DNA and the self-samples yielded on average 3.5 times more DNA than those obtained by the physician. All women that were positive for HR-HPV in the biopsy sample also typed positive both by self-sampling and physician-obtained sampling. For women with a histological diagnosis of cervical intraepithelial neoplasia grades 2-3 (CIN 2-3) all three HPV samples showed 100% concordance. A higher number of women were HPV positive by self-sampling than by physician-obtained sampling or by biopsy analysis. The Viba-brush and the FTA cartridge are suitable for self-sampling of vaginal cells and subsequent HR-HPV typing. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Time- and cell-type specific changes in iron, ferritin, and transferrin in the gerbil hippocampal CA1 region after transient forebrain ischemia

    Dae Young Yoo

    2016-01-01

    Full Text Available In the present study, we used immunohistochemistry and western blot analysis to examine changes in the levels and cellular localization of iron, heavy chain ferritin (ferritin-H, and transferrin in the gerbil hippocampal CA1 region from 30 minutes to 7 days following transient forebrain ischemia. Relative to sham controls, iron reactivity increased significantly in the stratum pyramidale and stratum oriens at 12 hours following ischemic insult, transiently decreased at 1-2 days and then increased once again within the CA1 region at 4-7 days after ischemia. One day after ischemia, ferritin-H immunoreactivity increased significantly in the stratum pyramidale and decreased at 2 days. At 4-7 days after ischemia, ferritin-H immunoreactivity in the glial components in the CA1 region was significantly increased. Transferrin immunoreactivity was increased significantly in the stratum pyramidale at 12 hours, peaked at 1 day, and then decreased significantly at 2 days after ischemia. Seven days after ischemia, Transferrin immunoreactivity in the glial cells of the stratum oriens and radiatum was significantly increased. Western blot analyses supported these results, demonstrating that compared to sham controls, ferritin H and transferrin protein levels in hippocampal homogenates significantly increased at 1 day after ischemia, peaked at 4 days and then decreased. These results suggest that iron overload-induced oxidative stress is most prominent at 12 hours after ischemia in the stratum pyramidale, suggesting that this time window may be the optimal period for therapeutic intervention to protect neurons from ischemia-induced death.

  7. The complex becomes more complex: protein-protein interactions of SnRK1 with DUF581 family proteins provide a framework for cell- and stimulus type-specific SnRK1 signaling in plants

    Madlen eNietzsche

    2014-02-01

    Full Text Available In plants, SNF1-related kinase (SnRK1 responds to the availability of carbohydrates as well as to environmental stresses by down-regulating ATP consuming biosynthetic processes, while stimulating energy-generating catabolic reactions through gene expression and post-transcriptional regulation. The functional SnRK1 complex is a heterotrimer where the catalytic alpha subunit associates with a regulatory beta subunit and an activating gamma subunit. Several different metabolites as well as the hormone abscisic acid (ABA have been shown to modulate SnRK1 activity in a cell- and stimulus-type specific manner. It has been proposed that tissue- or stimulus-specific expression of adapter proteins mediating SnRK1 regulation can at least partly explain the differences observed in SnRK1 signaling. By using yeast two-hybrid and in planta bi-molecular fluorescence complementation assays we were able to demonstrate that proteins containing the domain of unknown function (DUF 581 could interact with both isoforms of the SnRK1 alpha subunit (AKIN10/11 of Arabidopsis. A structure/function analysis suggests that the DUF581 is a generic SnRK1 interaction module and co-expression with DUF581 proteins in plant cells leads to reallocation of the kinase to specific regions within the nucleus. Yeast two-hybrid analyses suggest that SnRK1 and DUF581 proteins can share common interaction partners inside the nucleus. The analysis of available microarray data implies that expression of the 19 members of the DUF581 encoding gene family in Arabidopsis is differentially regulated by hormones and environmental cues, indicating specialized functions of individual family members. We hypothesize that DUF581 proteins could act as mediators conferring tissue- and stimulus-type specific differences in SnRK1 regulation.

  8. Androgen and taxol cause cell type-specific alterations of centrosome and DNA organization in androgen-responsive LNCaP and androgen-independent DU145 prostate cancer cells

    Schatten, H.; Ripple, M.; Balczon, R.; Weindruch, R.; Chakrabarti, A.; Taylor, M.; Hueser, C. N.

    2000-01-01

    We investigated the effects of androgen and taxol on the androgen-responsive LNCaP and androgen-independent DU145 prostate cancer cell lines. Cells were treated for 48 and 72 h with 0.05-1 nM of the synthetic androgen R1881 and with 100 nM taxol. Treatment of LNCaP cells with 0.05 nM R1881 led to increased cell proliferation, whereas treatment with 1 nM R1881 resulted in inhibited cell division, DNA cycle arrest, and altered centrosome organization. After treatment with 1 nM R1881, chromatin became clustered, nuclear envelopes convoluted, and mitochondria accumulated around the nucleus. Immunofluorescence microscopy with antibodies to centrosomes showed altered centrosome structure. Although centrosomes were closely associated with the nucleus in untreated cells, they dispersed into the cytoplasm after treatment with 1 nM R1881. Microtubules were only faintly detected in 1 nM R1881-treated LNCaP cells. The effects of taxol included microtubule bundling and altered mitochondria morphology, but not DNA organization. As expected, the androgen-independent prostate cancer cell line DU145 was not affected by R1881. Treatment with taxol resulted in bundling of microtubules in both cell lines. Additional taxol effects were seen in DU145 cells with micronucleation of DNA, an indication of apoptosis. Simultaneous treatment with R1881 and taxol had no additional effects on LNCaP or DU145 cells. These results suggest that LNCaP and DU145 prostate cancer cells show differences not only in androgen responsiveness but in sensitivity to taxol as well. Copyright 2000 Wiley-Liss, Inc.

  9. Cell-type specific DNA-protein interactions at the tissue-type plasminogen activator promoter in human endothelial and HeLa cells in vivo and in vitro

    Arts, J.; Herr, I.; Lansink, M.; Angel, P.; Kooistra, T.

    1997-01-01

    Tissue-type plasminogen activator (t-PA) gene expression in human endothelial cells and HeLa cells is stimulated by the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) at the level of transcription. To study the mechanism of transcriptional regulation, we have characterized a

  10. Type I interferon production during herpes simplex virus infection is controlled by cell-type-specific viral recognition through Toll-like receptor 9, the mitochondrial antiviral signaling protein pathway, and novel recognition systems

    Rasmussen, Simon Brandtoft; Sørensen, Louise Nørgaard; Malmgaard, Lene

    2007-01-01

    Recognition of viruses by germ line-encoded pattern recognition receptors of the innate immune system is essential for rapid production of type I interferon (IFN) and early antiviral defense. We investigated the mechanisms of viral recognition governing production of type I IFN during herpes...... simplex virus (HSV) infection. We show that early production of IFN in vivo is mediated through Toll-like receptor 9 (TLR9) and plasmacytoid dendritic cells, whereas the subsequent alpha/beta IFN (IFN-alpha/beta) response is derived from several cell types and induced independently of TLR9...

  11. The cell-type specific uptake of polymer-coated or micelle-embedded QDs and SPIOs does not provoke an acute pro-inflammatory response in the liver

    Markus Heine

    2014-09-01

    Full Text Available Semiconductor quantum dots (QD and superparamagnetic iron oxide nanocrystals (SPIO have exceptional physical properties that are well suited for biomedical applications in vitro and in vivo. For future applications, the direct injection of nanocrystals for imaging and therapy represents an important entry route into the human body. Therefore, it is crucial to investigate biological responses of the body to nanocrystals to avoid harmful side effects. In recent years, we established a system to embed nanocrystals with a hydrophobic oleic acid shell either by lipid micelles or by the amphiphilic polymer poly(maleic anhydride-alt-1-octadecene (PMAOD. The goal of the current study is to investigate the uptake processes as well as pro-inflammatory responses in the liver after the injection of these encapsulated nanocrystals. By immunofluorescence and electron microscopy studies using wild type mice, we show that 30 min after injection polymer-coated nanocrystals are primarily taken up by liver sinusoidal endothelial cells. In contrast, by using wild type, Ldlr-/- as well as Apoe-/- mice we show that nanocrystals embedded within lipid micelles are internalized by Kupffer cells and, in a process that is dependent on the LDL receptor and apolipoprotein E, by hepatocytes. Gene expression analysis of pro-inflammatory markers such as tumor necrosis factor alpha (TNFα or chemokine (C-X-C motif ligand 10 (Cxcl10 indicated that 48 h after injection internalized nanocrystals did not provoke pro-inflammatory pathways. In conclusion, internalized nanocrystals at least in mouse liver cells, namely endothelial cells, Kupffer cells and hepatocytes are at least not acutely associated with potential adverse side effects, underlining their potential for biomedical applications.

  12. Neuronal survival in the brain: neuron type-specific mechanisms

    Pfisterer, Ulrich Gottfried; Khodosevich, Konstantin

    2017-01-01

    Neurogenic regions of mammalian brain produce many more neurons that will eventually survive and reach a mature stage. Developmental cell death affects both embryonically produced immature neurons and those immature neurons that are generated in regions of adult neurogenesis. Removal of substantial...... numbers of neurons that are not yet completely integrated into the local circuits helps to ensure that maturation and homeostatic function of neuronal networks in the brain proceed correctly. External signals from brain microenvironment together with intrinsic signaling pathways determine whether...... for survival in a certain brain region. This review focuses on how immature neurons survive during normal and impaired brain development, both in the embryonic/neonatal brain and in brain regions associated with adult neurogenesis, and emphasizes neuron type-specific mechanisms that help to survive for various...

  13. OpenDMAP: An open source, ontology-driven concept analysis engine, with applications to capturing knowledge regarding protein transport, protein interactions and cell-type-specific gene expression

    Johnson Helen L

    2008-01-01

    Full Text Available Abstract Background Information extraction (IE efforts are widely acknowledged to be important in harnessing the rapid advance of biomedical knowledge, particularly in areas where important factual information is published in a diverse literature. Here we report on the design, implementation and several evaluations of OpenDMAP, an ontology-driven, integrated concept analysis system. It significantly advances the state of the art in information extraction by leveraging knowledge in ontological resources, integrating diverse text processing applications, and using an expanded pattern language that allows the mixing of syntactic and semantic elements and variable ordering. Results OpenDMAP information extraction systems were produced for extracting protein transport assertions (transport, protein-protein interaction assertions (interaction and assertions that a gene is expressed in a cell type (expression. Evaluations were performed on each system, resulting in F-scores ranging from .26 – .72 (precision .39 – .85, recall .16 – .85. Additionally, each of these systems was run over all abstracts in MEDLINE, producing a total of 72,460 transport instances, 265,795 interaction instances and 176,153 expression instances. Conclusion OpenDMAP advances the performance standards for extracting protein-protein interaction predications from the full texts of biomedical research articles. Furthermore, this level of performance appears to generalize to other information extraction tasks, including extracting information about predicates of more than two arguments. The output of the information extraction system is always constructed from elements of an ontology, ensuring that the knowledge representation is grounded with respect to a carefully constructed model of reality. The results of these efforts can be used to increase the efficiency of manual curation efforts and to provide additional features in systems that integrate multiple sources for

  14. Bonafide, type-specific human papillomavirus persistence among HIV-positive pregnant women: predictive value for cytological abnormalities, a longitudinal cohort study

    Angela RI Meyrelles

    2016-02-01

    Full Text Available This study investigated the rate of human papillomavirus (HPV persistence, associated risk factors, and predictors of cytological alteration outcomes in a cohort of human immunodeficiency virus-infected pregnant women over an 18-month period. HPV was typed through L1 gene sequencing in cervical smears collected during gestation and at 12 months after delivery. Outcomes were defined as nonpersistence (clearance of the HPV in the 2nd sample, re-infection (detection of different types of HPV in the 2 samples, and type-specific HPV persistence (the same HPV type found in both samples. An unfavourable cytological outcome was considered when the second exam showed progression to squamous intraepithelial lesion or high squamous intraepithelial lesion. Ninety patients were studied. HPV DNA persistence occurred in 50% of the cases composed of type-specific persistence (30% or re-infection (20%. A low CD4+T-cell count at entry was a risk factor for type-specific, re-infection, or HPV DNA persistence. The odds ratio (OR was almost three times higher in the type-specific group when compared with the re-infection group (OR = 2.8; 95% confidence interval: 0.43-22.79. Our findings show that bonafide (type-specific HPV persistence is a stronger predictor for the development of cytological abnormalities, highlighting the need for HPV typing as opposed to HPV DNA testing in the clinical setting.

  15. Investigation of solar cell radiation damage

    Bernard, J.; Reulet, R.; Arndt, R.A.

    1974-01-01

    Development of communications satellites has led to the requirement for a greater and longer lived solar cell power source. Accordingly, studies have been undertaken with the aim of determining which solar cell array provides the greatest power at end of life and the amount of degradation. Investigation of the damage done to thin silicon and thin film CdS solar cells is being carried out in two steps. First, irradiations were performed singly with 0.15, 1.0 and 2.0MeV electrons and 0.7, 2.5 and 22MeV proton. Solar cells and their cover materials were irradiated separately in order to locate the sites of the damage. Diffusion length and I.V. characteristics of the cells and transmission properties of the cover materials were measured. All neasurements were made in vacuum immediately after irradiation. In the second part it is intended to study the effect of various combinations of proton, electron and photon irradiation both with and without an electrical load. The results of this part show whether synergism is involved in solar cell damage and the relative importance of each of three radiation sources if synergism is found [fr

  16. Well-Controlled Cell-Trapping Systems for Investigating Heterogeneous Cell-Cell Interactions.

    Kamiya, Koki; Abe, Yuta; Inoue, Kosuke; Osaki, Toshihisa; Kawano, Ryuji; Miki, Norihisa; Takeuchi, Shoji

    2018-03-01

    Microfluidic systems have been developed for patterning single cells to study cell-cell interactions. However, patterning multiple types of cells to understand heterogeneous cell-cell interactions remains difficult. Here, it is aimed to develop a cell-trapping device to assemble multiple types of cells in the well-controlled order and morphology. This device mainly comprises a parylene sheet for assembling cells and a microcomb for controlling the cell-trapping area. The cell-trapping area is controlled by moving the parylene sheet on an SU-8 microcomb using tweezers. Gentle downward flow is used as a driving force for the cell-trapping. The assembly of cells on a parylene sheet with round and line-shaped apertures is demonstrated. The cell-cell contacts of the trapped cells are then investigated by direct cell-cell transfer of calcein via connexin nanopores. Finally, using the device with a system for controlling the cell-trapping area, three different types of cells in the well-controlled order are assembled. The correct cell order rate obtained using the device is 27.9%, which is higher than that obtained without the sliding parylene system (0.74%). Furthermore, the occurrence of cell-cell contact between the three cell types assembled is verified. This cell-patterning device will be a useful tool for investigating heterogeneous cell-cell interactions. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. XAS Investigations of PEM Fuel Cells

    Roth, Christina; Ramaker, David E.

    Polymer-electrolyte membrane (PEM) fuel cells are still far from an area-wide market launch due in part to long-term stability, reliability and cost issues. A more detailed knowledge of the underlying reaction mechanisms is expected to further their application, as it would allow for the design of tailor-made catalysts. However, this will only be possible by complementing traditional in situ studies on single-crystals in electrochemical cells with more sophisticated metal/electrolyte interfacial studies by novel spectroscopic methodologies, which can provide complementary insights into the behaviour of commercial catalysts under real fuel cell operating conditions. This review will focus on the advances of Xray absorption spectroscopy (XAS) in applied fuel cell research utilizing several examples. XAS enables both the nanoparticle morphology and the adsorbate coverage and binding site to be investigated with just one technique. The latter is possible when complementing the conventional extended X-ray absorption fine structure (EXAFS) analysis with the more novel Δμ XANES approach.

  18. Microfabricated physical spatial gradients for investigating cell migration and invasion dynamics.

    Michael Mak

    Full Text Available We devise a novel assay that introduces micro-architectures into highly confining microchannels to probe the decision making processes of migrating cells. The conditions are meant to mimic the tight spaces in the physiological environment that cancer cells encounter during metastasis within the matrix dense stroma and during intravasation and extravasation through the vascular wall. In this study we use the assay to investigate the relative probabilities of a cell 1 permeating and 2 repolarizing (turning around when it migrates into a spatially confining region. We observe the existence of both states even within a single cell line, indicating phenotypic heterogeneity in cell migration invasiveness and persistence. We also show that varying the spatial gradient of the taper can induce behavioral changes in cells, and different cell types respond differently to spatial changes. Particularly, for bovine aortic endothelial cells (BAECs, higher spatial gradients induce more cells to permeate (60% than lower gradients (12%. Furthermore, highly metastatic breast cancer cells (MDA-MB-231 demonstrate a more invasive and permeative nature (87% than non-metastatic breast epithelial cells (MCF-10A (25%. We examine the migration dynamics of cells in the tapered region and derive characteristic constants that quantify this transition process. Our data indicate that cell response to physical spatial gradients is both cell-type specific and heterogeneous within a cell population, analogous to the behaviors reported to occur during tumor progression. Incorporation of micro-architectures in confined channels enables the probing of migration behaviors specific to defined geometries that mimic in vivo microenvironments.

  19. Ctip2-, Satb2-, Prox1-, and GAD65-Expressing Neurons in Rat Cultures: Preponderance of Single- and Double-Positive Cells, and Cell Type-Specific Expression of Neuron-Specific Gene Family Members, Nsg-1 (NEEP21) and Nsg-2 (P19).

    Digilio, Laura; Yap, Chan Choo; Winckler, Bettina

    2015-01-01

    The brain consists of many distinct neuronal cell types, but which cell types are present in widely used primary cultures of embryonic rodent brain is often not known. We characterized how abundantly four cell type markers (Ctip2, Satb2, Prox1, GAD65) were represented in cultured rat neurons, how easily neurons expressing different markers can be transfected with commonly used plasmids, and whether neuronal-enriched endosomal proteins Nsg-1 (NEEP21) and Nsg-2 (P19) are ubiquitously expressed in all types of cultured neurons. We found that cultured neurons stably maintain cell type identities that are reflective of cell types in vivo. This includes neurons maintaining simultaneous expression of two transcription factors, such as Ctip2+/Satb2+ or Prox1+/Ctip2+ double-positive cells, which have also been described in vivo. Secondly, we established the superior efficiency of CAG promoters for both Lipofectamine-mediated transfection as well as for electroporation. Thirdly, we discovered that Nsg-1 and Nsg-2 were not expressed equally in all neurons: whereas high levels of both Nsg-1 and Nsg-2 were found in Satb2-, Ctip2-, and GAD65-positive neurons, Prox1-positive neurons in hippocampal cultures expressed low levels of both. Our findings thus highlight the importance of identifying neuronal cell types for doing cell biology in cultured neurons: Keeping track of neuronal cell type might uncover effects in assays that might otherwise be masked by the mixture of responsive and non-responsive neurons in the dish.

  20. Theoretical investigation on heterojunction solar cell

    Prema, K.; Geetha, K.

    1986-11-01

    The study of thin film solar cells has proved that the surface is rough. A two-dimensional method based on the integral equation technique to analyse thin film solar cells has been developed by DeMey et al. In this paper we present our analysis of a thin film solar cell using the above techniques. Variation of the minority carrier concentration, the saturation current and the junction current of the solar cell with surface roughness is presented. (author). 8 refs, 4 figs

  1. Neuronal type-specific gene expression profiling and laser-capture microdissection.

    Pietersen, Charmaine Y; Lim, Maribel P; Macey, Laurel; Woo, Tsung-Ung W; Sonntag, Kai C

    2011-01-01

    The human brain is an exceptionally heterogeneous structure. In order to gain insight into the neurobiological basis of neural circuit disturbances in various neurologic or psychiatric diseases, it is often important to define the molecular cascades that are associated with these disturbances in a neuronal type-specific manner. This can be achieved by the use of laser microdissection, in combination with molecular techniques such as gene expression profiling. To identify neurons in human postmortem brain tissue, one can use the inherent properties of the neuron, such as pigmentation and morphology or its structural composition through immunohistochemistry (IHC). Here, we describe the isolation of homogeneous neuronal cells and high-quality RNA from human postmortem brain material using a combination of rapid IHC, Nissl staining, or simple morphology with Laser-Capture Microdissection (LCM) or Laser Microdissection (LMD).

  2. Feasibility investigation of allogeneic endometrial regenerative cells

    Reid Michael

    2009-02-01

    Full Text Available Abstract Endometrial Regenerative Cells (ERC are a population of mesenchymal-like stem cells having pluripotent differentiation activity and ability to induce neoangiogenesis. In vitro and animal studies suggest ERC are immune privileged and in certain situations actively suppress ongoing immune responses. In this paper we describe the production of clinical grade ERC and initial safety experiences in 4 patients with multiple sclerosis treated intravenously and intrathecally. The case with the longest follow up, of more than one year, revealed no immunological reactions or treatment associated adverse effects. These preliminary data suggest feasibility of clinical ERC administration and support further studies with this novel stem cell type.

  3. Cybernetical investigations on the radiation effects on tumour cells

    Iliakis, G.

    1978-01-01

    The goal of this work was to find suitable conditions for the in-vitro investigations with these E.A.T. cells so that basic experiments carried out with yeast cells will also be possible. Systematic investigations on cell multiplication led to reliable and definite test conditions for these cells. For the evaluation of the data which had been measured with several sorts of radiation the cybernetic model was applied. (orig./MG) [de

  4. Optical Investigations of Endothelial Cell Motility

    Rossen, Ninna Struck

    A monolayer of endothelial cells lines the entire circulatory system and create a barrier between the circulatory system and the tissues. To create and maintain an intact barrier, the individual cells have to connect tightly with their neighbors, which causes a highly correlated motion between...... are fascinating from a biophysical point of view. The vasculature also plays a signi cant role in many pathologies. In diabetic blindness or ischemic diseases the ow of blood is insucient to sustain certain tissues or whole limbs. The creation of new blood vessels can relieve or treat such diseases. In other...... pathologies, such as the growth of cancerous tumors and metastasis, the creation of new blood vessels to these tumors worsen the condition and an inhibition of blood vessel creation will relieve the pathology. The thesis is divided into three parts; Part 1 provides some general background knowledge...

  5. Investigation of the selenium metabolism in cancer cell lines

    Lunøe, Kristoffer; Gabel-Jensen, Charlotte; Stürup, Stefan

    2011-01-01

    The aim of this work was to compare different selenium species for their ability to induce cell death in different cancer cell lines, while investigating the underlying chemistry by speciation analysis. A prostate cancer cell line (PC-3), a colon cancer cell line (HT-29) and a leukaemia cell line...... (Jurkat E6-1) were incubated with five selenium compounds representing inorganic as well as organic Se compounds in different oxidation states. Selenomethionine (SeMet), Se-methylselenocysteine (MeSeCys), methylseleninic acid (MeSeA), selenite and selenate in the concentration range 5-100 mu M were...... incubated with cells for 24 h and the induction of cell death was measured using flow cytometry. The amounts of total selenium in cell medium, cell lysate and the insoluble fractions was determined by ICP-MS. Speciation analysis of cellular fractions was performed by reversed phase, anion exchange and size...

  6. Investigating reliability attributes of silicon photovoltaic cells - An overview

    Royal, E. L.

    1982-01-01

    Reliability attributes are being developed on a wide variety of advanced single-crystal silicon solar cells. Two separate investigations: cell-contact integrity (metal-to-silicon adherence), and cracked cells identified with fracture-strength-reducing flaws are discussed. In the cell-contact-integrity investigation, analysis of contact pull-strength data shows that cell types made with different metallization technologies, i.e., vacuum, plated, screen-printed and soldered, have appreciably different reliability attributes. In the second investigation, fracture strength was measured using Czochralski wafers and cells taken at various stages of processing and differences were noted. Fracture strength, which is believed to be governed by flaws introduced during wafer sawing, was observed to improve (increase) after chemical polishing and other process steps that tend to remove surface and edge flaws.

  7. Evolution of sexes from an ancestral mating-type specification pathway.

    Sa Geng

    2014-07-01

    Full Text Available Male and female sexes have evolved repeatedly in eukaryotes but the origins of dimorphic sexes and their relationship to mating types in unicellular species are not understood. Volvocine algae include isogamous species such as Chlamydomonas reinhardtii, with two equal-sized mating types, and oogamous multicellular species such as Volvox carteri with sperm-producing males and egg-producing females. Theoretical work predicts genetic linkage of a gamete cell-size regulatory gene(s to an ancestral mating-type locus as a possible step in the evolution of dimorphic gametes, but this idea has not been tested. Here we show that, contrary to predictions, a single conserved mating locus (MT gene in volvocine algae-MID, which encodes a RWP-RK domain transcription factor-evolved from its ancestral role in C. reinhardtii as a mating-type specifier, to become a determinant of sperm and egg development in V. carteri. Transgenic female V. carteri expressing male MID produced functional sperm packets during sexual development. Transgenic male V. carteri with RNA interference (RNAi-mediated knockdowns of VcMID produced functional eggs, or self-fertile hermaphrodites. Post-transcriptional controls were found to regulate cell-type-limited expression and nuclear localization of VcMid protein that restricted its activity to nuclei of developing male germ cells and sperm. Crosses with sex-reversed strains uncoupled sex determination from sex chromosome identity and revealed gender-specific roles for male and female mating locus genes in sexual development, gamete fitness and reproductive success. Our data show genetic continuity between the mating-type specification and sex determination pathways of volvocine algae, and reveal evidence for gender-specific adaptations in the male and female mating locus haplotypes of Volvox. These findings will enable a deeper understanding of how a master regulator of mating-type determination in an ancestral unicellular species was

  8. Muscle type-specific responses to NAD+ salvage biosynthesis promote muscle function in Caenorhabditis elegans.

    Vrablik, Tracy L; Wang, Wenqing; Upadhyay, Awani; Hanna-Rose, Wendy

    2011-01-15

    Salvage biosynthesis of nicotinamide adenine dinucleotide (NAD(+)) from nicotinamide (NAM) lowers NAM levels and replenishes the critical molecule NAD(+) after it is hydrolyzed. This pathway is emerging as a regulator of multiple biological processes. Here we probe the contribution of the NAM-NAD(+) salvage pathway to muscle development and function using Caenorhabditis elegans. C. elegans males with mutations in the nicotinamidase pnc-1, which catalyzes the first step of this NAD(+) salvage pathway, cannot mate due to a spicule muscle defect. Multiple muscle types are impaired in the hermaphrodites, including body wall muscles, pharyngeal muscles and vulval muscles. An active NAD(+) salvage pathway is required for optimal function of each muscle cell type. However, we found surprising muscle-cell-type specificity in terms of both the timing and relative sensitivity to perturbation of NAD(+) production or NAM levels. Active NAD(+) biosynthesis during development is critical for function of the male spicule protractor muscles during adulthood, but these muscles can surprisingly do without salvage biosynthesis in adulthood under the conditions examined. The body wall muscles require ongoing NAD(+) salvage biosynthesis both during development and adulthood for maximum function. The vulval muscles do not function in the presence of elevated NAM concentrations, but NAM supplementation is only slightly deleterious to body wall muscles during development or upon acute application in adults. Thus, the pathway plays distinct roles in different tissues. As NAM-NAD(+) biosynthesis also impacts muscle differentiation in vertebrates, we propose that similar complexities may be found among vertebrate muscle cell types. Copyright © 2010 Elsevier Inc. All rights reserved.

  9. Tissue-type-specific transcriptome analysis identifies developing xylem-specific promoters in poplar.

    Ko, Jae-Heung; Kim, Hyun-Tae; Hwang, Ildoo; Han, Kyung-Hwan

    2012-06-01

    Plant biotechnology offers a means to create novel phenotypes. However, commercial application of biotechnology in crop improvement programmes is severely hindered by the lack of utility promoters (or freedom to operate the existing ones) that can drive gene expression in a tissue-specific or temporally controlled manner. Woody biomass is gaining popularity as a source of fermentable sugars for liquid fuel production. To improve the quantity and quality of woody biomass, developing xylem (DX)-specific modification of the feedstock is highly desirable. To develop utility promoters that can drive transgene expression in a DX-specific manner, we used the Affymetrix Poplar Genome Arrays to obtain tissue-type-specific transcriptomes from poplar stems. Subsequent bioinformatics analysis identified 37 transcripts that are specifically or strongly expressed in DX cells of poplar. After further confirmation of their DX-specific expression using semi-quantitative PCR, we selected four genes (DX5, DX8, DX11 and DX15) for in vivo confirmation of their tissue-specific expression in transgenic poplars. The promoter regions of the selected DX genes were isolated and fused to a β-glucuronidase (GUS)-reported gene in a binary vector. This construct was used to produce transgenic poplars via Agrobacterium-mediated transformation. The GUS expression patterns of the resulting transgenic plants showed that these promoters were active in the xylem cells at early seedling growth and had strongest expression in the developing xylem cells at later growth stages of poplar. We conclude that these DX promoters can be used as a utility promoter for DX-specific biomass engineering. © 2012 The Authors. Plant Biotechnology Journal © 2012 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  10. INVESTIGATION OF PEM FUEL CELL FOR AUTOMOTIVE USE

    A. K. M. Mohiuddin

    2015-11-01

    Full Text Available This paper provides a brief investigation on suitability of Proton-exchange  membrane fuel cells (PEMFCs as the source of power for transportation purposes. Hydrogen is an attractive alternative transportation fuel. It is the least polluting fuel that can be used in an internal combustion engine (ICE and it is widely available. If hydrogen is used in a fuel cell which converts the chemical energy of hydrogen into electricity, (NOx emissions are eliminated. The investigation was carried out on a  fuel cell car model by implementing polymer electrolyte membrane (PEM types of fuel cell as the source of power to propel the prototype car. This PEMFC has capability to propel the electric motor by converting chemical energy stored in hydrogen gas into useful electrical energy. PEM fuel cell alone is used as the power source for the electric motor without the aid of any other power source such as battery associated with it. Experimental investigations were carried out to investigate the characteristics of fuel cell used and the performance of the fuel cell car. Investigated papameters are the power it develops, voltage, current and speed it produces under different load conditions. KEYWORDS: fuel cell; automotive; proton exchange membrane; polymer electrolyte membrane; internal combustion engine

  11. Magnetic resonance investigation of magnetic-labeled baker's yeast cells

    Godoy Morais, J.P.M.; Azevedo, R.B.; Silva, L.P.; Lacava, Z.G.M.; Bao, S.N.; Silva, O.; Pelegrini, F.; Gansau, C.; Buske, N.; Safarik, I.; Safarikova, M.; Morais, P.C.

    2004-01-01

    In this study, the interaction of DMSA-coated magnetite nanoparticles (5 and 10 nm core-size) with Saccharomyces cerevisae was investigated using magnetic resonance (MR) and transmission electron microscopy (TEM). The TEM micrographs revealed magnetite nanoparticles attached externally to the cell wall. The MR data support the strong interaction among the nanoparticles supported by the cells. A remarkable shift in the resonance field was used as signature of particle attachment to the cell wall

  12. Investigation of Pockels Cells Crystal Contrast Ratio Distribution

    Giedrius Sinkevičius

    2017-07-01

    Full Text Available The BBO Pockel’s cell has been investigated. The investigation results of optimal operating area on the surface of the crystal dependent of intrinsic contrast ratio (ICR and voltage contrast ratio (VCR for Pockel’s cell are presented. The block diagram of Pockel’s cells contrast measurement stand and measurement methodology are introduced and discussed. The graphs of intrinsic contrast ratio distribution on crystal surface, contrast ratio with voltage dependency and voltage contrast ratio distribution on crystal surface with half-wave voltage are presented.

  13. Porosimetry as an effective method of fuel cell investigation

    Kazarinov, V.E.

    1996-04-01

    A porosimetric method is described for the investigation of all kinds of porous materials including soft or frail materials and powders. The method is well suited for the investigation of electrodes in fuel cells and batteries. The method is nondestructive and allows for repeated measurements on the same sample.

  14. Investigation of autofocus algorithms for brightfield microscopy of unstained cells

    Wu, Shu Yu; Dugan, Nazim; Hennelly, Bryan M.

    2014-05-01

    In the past decade there has been significant interest in image processing for brightfield cell microscopy. Much of the previous research on image processing for microscopy has focused on fluorescence microscopy, including cell counting, cell tracking, cell segmentation and autofocusing. Fluorescence microscopy provides functional image information that involves the use of labels in the form of chemical stains or dyes. For some applications, where the biochemical integrity of the cell is required to remain unchanged so that sensitive chemical testing can later be applied, it is necessary to avoid staining. For this reason the challenge of processing images of unstained cells has become a topic of increasing attention. These cells are often effectively transparent and appear to have a homogenous intensity profile when they are in focus. Bright field microscopy is the most universally available and most widely used form of optical microscopy and for this reason we are interested in investigating image processing of unstained cells recorded using a standard bright field microscope. In this paper we investigate the application of a range of different autofocus metrics applied to unstained bladder cancer cell lines using a standard inverted bright field microscope with microscope objectives that have high magnification and numerical aperture. We present a number of conclusions on the optimum metrics and the manner in which they should be applied for this application.

  15. Investigating the role of retinal Müller cells with approaches in genetics and cell biology.

    Fu, Suhua; Zhu, Meili; Ash, John D; Wang, Yunchang; Le, Yun-Zheng

    2014-01-01

    Müller cells are major macroglia and play many essential roles as a supporting cell in the retina. As Müller cells only constitute a small portion of retinal cells, investigating the role of Müller glia in retinal biology and diseases is particularly challenging. To overcome this problem, we first generated a Cre/lox-based conditional gene targeting system that permits the genetic manipulation and functional dissection of gene of interests in Müller cells. To investigate diabetes-induced alteration of Müller cells, we recently adopted methods to analyze Müller cells survival/death in vitro and in vivo. We also used normal and genetically altered primary cell cultures to reveal the mechanistic insights for Müller cells in biological and disease processes. In this article, we will discuss the applications and limitations of these methodologies, which may be useful for research in retinal Müller cell biology and pathophysiology.

  16. Real-time PCR for type-specific identification of herpes simplex in clinical samples: evaluation of type-specific results in the context of CNS diseases.

    Meylan, Sylvain; Robert, Daniel; Estrade, Christine; Grimbuehler, Valérie; Péter, Olivier; Meylan, Pascal R; Sahli, Roland

    2008-02-01

    HSV-1 and HSV-2 cause CNS infections of dissimilar clinico-pathological characteristics with prognostic and therapeutic implications. To validate a type-specific real-time PCR that uses MGB/LNA Taqman probes and to review the virologico-clinical data of 25 eligible patients with non-neonatal CNS infections. This real-time PCR was evaluated against conventional PCR (26 CSF and 20 quality controls), and LightCycler assay (51 mucocutaneous, 8 CSF and 32 quality controls) and culture/immunofluorescence (75 mucocutaneous) to assess typing with independent methods. Taqman real-time PCR detected 240 HSV genomes per ml CSF, a level appropriate for the management of patients, and provided unambiguous typing for the 104 positive (62 HSV-1 and 42 HSV-2) out the 160 independent clinical samples tested. HSV type diagnosed by Taqman real-time PCR predicted final diagnosis (meningitis versus encephalitis/meningoencephalitis, p<0.001) in 24/25 patients at time of presentation, in contrast to clinical evaluation. Our real-time PCR, as a sensitive and specific means for type-specific HSV diagnosis, provided rapid prognostic information for patient management.

  17. Investigating microenvironmental regulation of human chordoma cell behaviour.

    Priya Patel

    Full Text Available The tumour microenvironment is complex and composed of many different constituents, including matricellular proteins such as connective tissue growth factor (CCN2, and is characterized by gradients in oxygen levels. In various cancers, hypoxia and CCN2 promote stem and progenitor cell properties, and regulate the proliferation, migration and phenotype of cancer cells. Our study was aimed at investigating the effects of hypoxia and CCN2 on chordoma cells, using the human U-CH1 cell line. We demonstrate that under basal conditions, U-CH1 cells express multiple CCN family members including CCN1, CCN2, CCN3 and CCN5. Culture of U-CH1 cells in either hypoxia or in the presence of recombinant CCN2 peptide promoted progenitor cell-like characteristics specific to the notochordal tissue of origin. Specifically, hypoxia induced the most robust increase in progenitor-like characteristics in U-CH1 cells, including increased expression of the notochord-associated markers T, CD24, FOXA1, ACAN and CA12, increased cell growth and tumour-sphere formation, and a decrease in the percentage of vacuolated cells present in the heterogeneous population. Interestingly, the effects of recombinant CCN2 peptide on U-CH1 cells were more pronounced under normoxia than hypoxia, promoting increased expression of CCN1, CCN2, CCN3 and CCN5, the notochord-associated markers SOX5, SOX6, T, CD24, and FOXA1 as well as increased tumour-sphere formation. Overall, this study highlights the importance of multiple factors within the tumour microenvironment and how hypoxia and CCN2 may regulate human chordoma cell behaviour.

  18. Fundamental investigations on periodic nano- and microstructured organic solar cells

    Niggemann, M.

    2005-03-15

    Using organic semiconducting materials in solar cells is a new approach with promising possibilities. The great potential of low cost production combined with mechanical flexibility gives rise to new applications. Due to the relatively simple fabrication process from solution and the mechanical flexibility, the production of organic solar cells by the cost effective roll-to-roll process appears promising. However, the preconditions for commercialization are not fulfilled as yet. The demands on organic solar cells strongly depend on the type of application. The highest demands on solar cell technologies are set by the energy market. Organic solar cells are only expected to be competitive on the energy market when the requirements on efficiency, lifetime and costs are fulfilled at the same time. Regarding this as a long term goal, a less demanding but still challenging medium term goal would be the application of relatively small organic solar cell modules for i.e. portable electronic devices. The integration of Organic Field Effect Transistors (OFET) and Organic Light Emitting Diodes (OLED) to all-polymer electronic devices is still under development. Nevertheless, the integration of organic solar cells as one functional component appears promising as the production technologies are expected to be compatible. The innovative contribution of this thesis to the development of organic solar cells is as follows: Motivated by the desire to fabricate efficient and cost effective organic solar cells, the approach of developing novel solar cell architectures based on periodic nano- and microstructures is followed. At present, planar organic solar cells with indium tin oxide (ITO) as a transparent electrode are intensively studied. One decisive cost factor would, however, be the indium price, which is the key component of the ITO electrode. The planar cell architecture can be conceived as a one-dimensional photonic device, however the presented work widens the investigations

  19. Investigating the effect of cell substrate on cancer cell stiffness by optical tweezers.

    Yousafzai, Muhammad Sulaiman; Coceano, Giovanna; Bonin, Serena; Niemela, Joseph; Scoles, Giacinto; Cojoc, Dan

    2017-07-26

    The mechanical properties of cells are influenced by their microenvironment. Here we report cell stiffness alteration by changing the cell substrate stiffness for isolated cells and cells in contact with other cells. Polydimethylsiloxane (PDMS) is used to prepare soft substrates with three different stiffness values (173, 88 and 17kPa respectively). Breast cancer cells lines, namely HBL-100, MCF-7 and MDA-MB-231 with different level of aggressiveness are cultured on these substrates and their local elasticity is investigated by vertical indentation of the cell membrane. Our preliminary results show an unforeseen behavior of the MDA-MB-231 cells. When cultured on glass substrate as isolated cells, they are less stiff than the other two types of cells, in agreement with the general statement that more aggressive and metastatic cells are softer. However, when connected to other cells the stiffness of MDA-MB-231 cells becomes similar to the other two cell lines. Moreover, the stiffness of MDA-MB-231 cells cultured on soft PDMS substrates is significantly higher than the stiffness of the other cell types, demonstrating thus the strong influence of the environmental conditions on the mechanical properties of the cells. Copyright © 2017. Published by Elsevier Ltd.

  20. The role of myostatin and activin receptor IIB in the regulation of unloading-induced myofiber type-specific skeletal muscle atrophy.

    Babcock, Lyle W; Knoblauch, Mark; Clarke, Mark S F

    2015-09-15

    Chronic unloading induces decrements in muscle size and strength. This adaptation is governed by a number of molecular factors including myostatin, a potent negative regulator of muscle mass. Myostatin must first be secreted into the circulation and then bind to the membrane-bound activin receptor IIB (actRIIB) to exert its atrophic action. Therefore, we hypothesized that myofiber type-specific atrophy observed after hindlimb suspension (HLS) would be related to myofiber type-specific expression of myostatin and/or actRIIB. Wistar rats underwent HLS for 10 days, after which the tibialis anterior was harvested for frozen cross sectioning. Simultaneous multichannel immunofluorescent staining combined with differential interference contrast imaging was employed to analyze myofiber type-specific expression of myostatin and actRIIB and myofiber type cross-sectional area (CSA) across fiber types, myonuclei, and satellite cells. Hindlimb suspension (HLS) induced significant myofiber type-specific atrophy in myosin heavy chain (MHC) IIx (P Myostatin staining associated with myonuclei was less in HLS rats compared with controls, while satellite cell staining for myostatin remained unchanged. In contrast, the total number myonuclei and satellite cells per myofiber was reduced in HLS compared with ambulatory control rats (P myostatin-induced myofiber type-selective atrophy observed during chronic unloading. Copyright © 2015 the American Physiological Society.

  1. Bending spring rate investigation of nanopipette for cell injection

    Shen, Yajing; Zhang, Zhenhai; Fukuda, Toshio

    2015-04-01

    Bending of nanopipette tips during cell penetration is a major cause of cell injection failure. However, the flexural rigidity of nanopipettes is little known due to their irregular structure. In this paper, we report a quantitative method to estimate the flexural rigidity of a nanopipette by investigating its bending spring rate. First nanopipettes with a tip size of 300 nm are fabricated from various glass tubes by laser pulling followed by focused ion beam (FIB) milling. Then the bending spring rate of the nanopipettes is investigated inside a scanning electron microscope (SEM). Finally, a yeast cell penetration test is performed on these nanopipettes, which have different bending spring rates. The results show that nanopipettes with a higher bending spring rate have better cell penetration capability, which confirms that the bending spring rate may well reflect the flexural rigidity of a nanopipette. This method provides a quantitative parameter for characterizing the mechanical property of a nanopipette that can be potentially taken as a standard specification in the future. This general method can also be used to estimate other one-dimensional structures for cell injection, which will greatly benefit basic cell biology research and clinical applications.

  2. Bending spring rate investigation of nanopipette for cell injection

    Shen, Yajing; Zhang, Zhenhai; Fukuda, Toshio

    2015-01-01

    Bending of nanopipette tips during cell penetration is a major cause of cell injection failure. However, the flexural rigidity of nanopipettes is little known due to their irregular structure. In this paper, we report a quantitative method to estimate the flexural rigidity of a nanopipette by investigating its bending spring rate. First nanopipettes with a tip size of 300 nm are fabricated from various glass tubes by laser pulling followed by focused ion beam (FIB) milling. Then the bending spring rate of the nanopipettes is investigated inside a scanning electron microscope (SEM). Finally, a yeast cell penetration test is performed on these nanopipettes, which have different bending spring rates. The results show that nanopipettes with a higher bending spring rate have better cell penetration capability, which confirms that the bending spring rate may well reflect the flexural rigidity of a nanopipette. This method provides a quantitative parameter for characterizing the mechanical property of a nanopipette that can be potentially taken as a standard specification in the future. This general method can also be used to estimate other one-dimensional structures for cell injection, which will greatly benefit basic cell biology research and clinical applications. (paper)

  3. Cell-type specific increases in female hamster nucleus accumbens spine density following female sexual experience.

    Staffend, Nancy A; Hedges, Valerie L; Chemel, Benjamin R; Watts, Val J; Meisel, Robert L

    2014-11-01

    Female sexual behavior is an established model of a naturally motivated behavior which is regulated by activity within the mesolimbic dopamine system. Repeated activation of the mesolimbic circuit by female sexual behavior elevates dopamine release and produces persistent postsynaptic alterations to dopamine D1 receptor signaling within the nucleus accumbens. Here we demonstrate that sexual experience in female Syrian hamsters significantly increases spine density and alters morphology selectively in D1 receptor-expressing medium spiny neurons within the nucleus accumbens core, with no corresponding change in dopamine receptor binding or protein expression. Our findings demonstrate that previous life experience with a naturally motivated behavior has the capacity to induce persistent structural alterations to the mesolimbic circuit that can increase reproductive success and are analogous to the persistent structural changes following repeated exposure to many drugs of abuse.

  4. eFORGE : A Tool for Identifying Cell Type-Specific Signal in Epigenomic Data

    Breeze, Charles E.; Paul, Dirk S.; van Dongen, Jenny; Butcher, Lee M.; Ambrose, John C.; Barrett, James E.; Lowe, Robert; Rakyan, Vardhman K.; Iotchkova, Valentina; Frontini, Mattia; Downes, Kate; Ouwehand, Willem H.; Laperle, Jonathan; Jacques, Pierre-ETienne; Bourque, Guillaume; Bergmann, Anke K.; Siebert, Reiner; Vellenga, Edo; Saeed, Sadia; Matarese, Filomena; Martens, Joost H. A.; Stunnenberg, Hendrik G.; Teschendorff, Andrew E.; Herrero, Javier; Birney, Ewan; Dunham, Ian; Beck, Stephan

    2016-01-01

    Epigenome-wide association studies (EWAS) provide an alternative approach for studying human disease through consideration of non-genetic variants such as altered DNA methylation. To advance the complex interpretation of EWAS, we developed eFORGE (http://eforge.cs.ucl.ac.uk/), a new stand-alone and

  5. Cell-Type-Specific Cytokinin Distribution within the Arabidopsis Primary Root Apex

    Antoniadi, I.; Plačková, Lenka; Simonovik, B.; Doležal, Karel; Turnbull, C.; Ljung, K.; Novák, Ondřej

    2015-01-01

    Roč. 27, č. 7 (2015), s. 1955-1967 ISSN 1040-4651 R&D Projects: GA ČR GA14-34792S; GA MŠk(CZ) LO1204; GA MŠk LK21306 Institutional support: RVO:61389030 Keywords : SOLID-PHASE EXTRACTION * ENDOPLASMIC-RETICULUM * MERISTEM ACTIVITY Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 8.538, year: 2015

  6. Mapping functional connectivity in barrel-related columns reveals layer- and cell type-specific microcircuits.

    Schubert, D.; Kotter, R.; Staiger, J.F.

    2007-01-01

    Synaptic circuits bind together functional modules of the neocortex. We aim to clarify in a rodent model how intra- and transcolumnar microcircuits in the barrel cortex are laid out to segregate and also integrate sensory information. The primary somatosensory (barrel) cortex of rodents is the ideal

  7. A cell-type-specific role for murine Commd1 in liver inflammation

    Bartuzi, Paulina; Wijshake, Tobias; Dekker, Daphne C.; Fedoseienko, Alina; Kloosterhuis, Niels J.; Youssef, Sameh A.; Li, Haiying; Shiri-Sverdlov, Ronit; Kuivenhoven, Jan-Albert; de Bruin, Alain; Burstein, Ezra; Hofker, Marten H.; van de Sluis, Bait

    2014-01-01

    The transcription factor NF-kappa B plays a critical role in the inflammatory response and it has been implicated in various diseases, including non-alcoholic fatty liver disease (NAFLD). Although transient NF-kappa B activation may protect tissues from stress, a prolonged NF-kappa B activation can

  8. Frequency of nuclear mutant huntingtin inclusion formation in neurons and glia is cell-type-specific

    Jansen, Anne H. P.; van Hal, Maurik; Op den Kelder, Ilse C.; Meier, Romy T.; de Ruiter, Anna-Aster; Schut, Menno H.; Smith, Donna L.; Grit, Corien; Brouwer, Nieske; Kamphuis, Willem; Boddeke, H. W. G. M.; den Dunnen, Wilfred F. A.; van Roon, Willeke M. C.; Bates, Gillian P.; Hol, Elly M.; Reits, Eric A.

    2017-01-01

    Huntington's disease (HD) is an autosomal dominant inherited neurodegenerative disorder that is caused by a CAG expansion in the Huntingtin (HTT) gene, leading to HTT inclusion formation in the brain. The mutant huntingtin protein (mHTT) is ubiquitously expressed and therefore nuclear inclusions

  9. Frequency of Nuclear Mutant Huntingtin Inclusion Formation in Neurons and Glia is Cell-Type-Specific

    Jansen, Anne H P; van Hal, Maurik; op den Kelder, Ilse C.; Meier, Romy T.; de Ruiter, Anna-Aster; Schut, Menno H.; Smith, Donna L.; Grit, Corien; Brouwer, Nieske; Kamphuis, Willem; Boddeke, H. W. G. M.; den Dunnen, Wilfred F. A.; van Roon, Willeke M. C.; Bates, Gillian P.; Hol, Elly M.; Reits, Eric A.

    Huntington's disease (HD) is an autosomal dominant inherited neurodegenerative disorder that is caused by a CAG expansion in the Huntingtin (HTT) gene, leading to HTT inclusion formation in the brain. The mutant huntingtin protein (mHTT) is ubiquitously expressed and therefore nuclear inclusions

  10. Cell-Type-Specific Translation Profiling Reveals a Novel Strategy for Treating Fragile X Syndrome.

    Thomson, Sophie R; Seo, Sang S; Barnes, Stephanie A; Louros, Susana R; Muscas, Melania; Dando, Owen; Kirby, Caoimhe; Wyllie, David J A; Hardingham, Giles E; Kind, Peter C; Osterweil, Emily K

    2017-08-02

    Excessive mRNA translation downstream of group I metabotropic glutamate receptors (mGlu 1/5 ) is a core pathophysiology of fragile X syndrome (FX); however, the differentially translating mRNAs that contribute to altered neural function are not known. We used translating ribosome affinity purification (TRAP) and RNA-seq to identify mistranslating mRNAs in CA1 pyramidal neurons of the FX mouse model (Fmr1 -/y ) hippocampus, which exhibit exaggerated mGlu 1/5 -induced long-term synaptic depression (LTD). In these neurons, we find that the Chrm4 transcript encoding muscarinic acetylcholine receptor 4 (M 4 ) is excessively translated, and synthesis of M 4 downstream of mGlu 5 activation is mimicked and occluded. Surprisingly, enhancement rather than inhibition of M 4 activity normalizes core phenotypes in the Fmr1 -/y , including excessive protein synthesis, exaggerated mGluR-LTD, and audiogenic seizures. These results suggest that not all excessively translated mRNAs in the Fmr1 -/y brain are detrimental, and some may be candidates for enhancement to correct pathological changes in the FX brain. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  11. Investigation of an Aberrant Cell Voltage During the Filling of a Large Lithium Thionyl Chloride Cell

    Thaller, Lawrence H.; Quinzio, Michael V.

    1997-01-01

    The investigation of an aberrant cell voltage during the filling of a large lithium thionyl chloride cell summary is at: an aberrant voltage trace was noted during the review of cell filling data; incident was traced to an interruption during filling; experimentation suggested oxidizable sites within the carbon electrode were responsible for the drop in voltage; the voltage anomaly could be reproduced by interrupting the filling of similar cells; and anomalous voltage dip was not due to a short.

  12. Investigation of cancer cell behavior on nanofibrous scaffolds

    Szot, Christopher S.; Buchanan, Cara F. [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Gatenholm, Paul [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Department of Chemical and Biological Engineering, Chalmers University of Technology, SE-412 96 Goeteborg (Sweden); Rylander, Marissa Nichole [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States); Freeman, Joseph W., E-mail: jwfreeman@vt.edu [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061 (United States)

    2011-01-01

    Tissue engineering and the use of nanofibrous biomaterial scaffolds offer a unique perspective for studying cancer development in vitro. Current in vitro models of tumorigenesis are limited by the use of static, two-dimensional (2D) cell culture monolayers that lack the structural architecture necessary for cell-cell interaction and three-dimensional (3D) scaffolds that are too simplistic for studying basic pathological mechanisms. In this study, two nanofibrous biomaterials that mimic the structure of the extracellular matrix, bacterial cellulose and electrospun polycaprolactone (PCL)/collagen I, were investigated as potential 3D scaffolds for an in vitro cancer model. Multiple cancer cell lines were cultured on each scaffold material and monitored for cell viability, proliferation, adhesion, infiltration, and morphology. Both bacterial cellulose and electrospun PCL/collagen I, which have nano-scale structures on the order of 100-500 nm, have been used in many diverse tissue engineering applications. Cancer cell adhesion and growth were limited on bacterial cellulose, while all cellular processes were enhanced on the electrospun scaffolds. This initial analysis has demonstrated the potential of electrospun PCL/collagen I scaffolds toward the development of an improved 3D in vitro cancer model.

  13. Prevalence of type-specific HPV among female university students from northern Brazil

    Vieira, Rodrigo Covre; Monteiro, Jeniffer do Socorro Valente; Manso, Est?fane Primo; dos Santos, Maria Renata Mendon?a; Tsutsumi, Mihoko Yamamoto; Ishikawa, Edna Aoba Yassui; Ferrari, Stephen Francis; Lima, Karla Val?ria Batista; de Sousa, Ma?sa Silva

    2015-01-01

    Background Human papillomavirus (HPV) infection is associated with cervical cancer, the most frequent cancer in women from northern Brazil. Assessment of the short-term impact of HPV vaccination depends on the availability of data on the prevalence of type-specific HPV in young women in the pre-immunization period, although these data are currently unavailable for the study region. The aim of this study was to estimate the distribution of all mucosal HPV genotypes, including low- and high-ris...

  14. Micro solid-phase radioimmunoassay for detection of herpesvirus type-specific antibody: specificity and sensitivity

    Adler-Storthz, K.; Matson, D.O.; Adam, E.; Dreesman, G.R. (Baylor Univ., Houston, TX (USA). Coll. of Medicine)

    1983-02-01

    The specificity and sensitivity of a micro solid-phase radioimmunoassay (micro-SPRIA) that detects type-specific IgG antibody to herpes simplex virus types 1 and 2 (HSV1 and HSV2) were evaluated. Glycoproteins VP123 (molecular weight, 123,000) of HSV1 and VP119 (molecular weight, 119,000) of HSV2 were found to display the greatest degree of antigenic type-specificity of several HSV antigens tested with the micro-SPRIA technique. When testing a group of sera, negative for anti-HSV antibodies by microneutralization, in the micro-SPRIA, a range of negative reactivities was noted, suggesting that cut-points should be determined for each antigen preparation. The micro-SPRIA detected appropriate antibody activity in patients with recurrent infection and a marked agreement was noted in comparison to detection of anti-HSV antibodies measured with the microneutralization test. The type-specificity of the micro-SPRIA was substantiated by the independence of test results using VP119 and VP123 antigens for a random group of positive sera. The assay is rapid, specific, and sensitive and allows the testing of multiple serum samples with a standardized set of reagents.

  15. Human muscle fibre type-specific regulation of AMPK and downstream targets by exercise

    Kristensen, Dorte Enggaard; Albers, Peter Hjorth; Prats, Clara

    2015-01-01

    are expressed in a fibre type-dependent manner and that fibre type-specific activation of AMPK and downstream targets is dependent on exercise intensity. Pools of type I and II fibres were prepared from biopsies of m. vastus lateralis from healthy men before and after two exercise trials; A) continuous cycling......AMP-activated protein kinase (AMPK) is a regulator of energy homeostasis during exercise. Studies suggest muscle fibre type-specific AMPK expression. However, fibre type-specific regulation of AMPK and downstream targets during exercise has not been proven. We hypothesized that AMPK subunits...... (CON) 30 min at 69 ± 1% VO2peak or B) interval cycling (INT) 30 min with 6 × 1.5 min high-intense bouts peaking at 95 ± 2% VO2peak . In type I vs. II fibres a higher β1 AMPK (+215%) and lower γ3 AMPK expression (-71%) was found. α1 , α2 , β2 and γ1 AMPK expression was similar between fibre types...

  16. Fuel cell bus operation, system investigation H2 bus

    Anon.

    1992-01-01

    The WP covers two tasks: - Prepartion of Technical Catalogue: In cooperation with ICIL, AR have compiled a technical catalogue, providing the impartial descriptions, both of existing technology and regulations, and the likely future developments of these, as to remedy the first problem faced by a potential hydrogen bus fleet operator viz the absence of an impartial description of the available vehicle and fuels systems together with the absence of a description of regulatory and safety factors which need consideration. - Fuel Cell Bus Operation - System Investigation H 2 Bus: The application of fuel cell electric generation systems to hybrid electrical buses or electrical busses without any storage system on board is considered. The task will cover safety and environmental aspects, a cost estimate and a market evaluation. (orig.)

  17. Fnip1 regulates skeletal muscle fiber type specification, fatigue resistance, and susceptibility to muscular dystrophy

    Reyes, Nicholas L.; Banks, Glen B.; Tsang, Mark; Margineantu, Daciana; Gu, Haiwei; Djukovic, Danijel; Chan, Jacky; Torres, Michelle; Liggitt, H. Denny; Hirenallur-S, Dinesh K.; Hockenbery, David M.; Raftery, Daniel; Iritani, Brian M.

    2015-01-01

    Mammalian skeletal muscle is broadly characterized by the presence of two distinct categories of muscle fibers called type I “red” slow twitch and type II “white” fast twitch, which display marked differences in contraction strength, metabolic strategies, and susceptibility to fatigue. The relative representation of each fiber type can have major influences on susceptibility to obesity, diabetes, and muscular dystrophies. However, the molecular factors controlling fiber type specification remain incompletely defined. In this study, we describe the control of fiber type specification and susceptibility to metabolic disease by folliculin interacting protein-1 (Fnip1). Using Fnip1 null mice, we found that loss of Fnip1 increased the representation of type I fibers characterized by increased myoglobin, slow twitch markers [myosin heavy chain 7 (MyH7), succinate dehydrogenase, troponin I 1, troponin C1, troponin T1], capillary density, and mitochondria number. Cultured Fnip1-null muscle fibers had higher oxidative capacity, and isolated Fnip1-null skeletal muscles were more resistant to postcontraction fatigue relative to WT skeletal muscles. Biochemical analyses revealed increased activation of the metabolic sensor AMP kinase (AMPK), and increased expression of the AMPK-target and transcriptional coactivator PGC1α in Fnip1 null skeletal muscle. Genetic disruption of PGC1α rescued normal levels of type I fiber markers MyH7 and myoglobin in Fnip1-null mice. Remarkably, loss of Fnip1 profoundly mitigated muscle damage in a murine model of Duchenne muscular dystrophy. These results indicate that Fnip1 controls skeletal muscle fiber type specification and warrant further study to determine whether inhibition of Fnip1 has therapeutic potential in muscular dystrophy diseases. PMID:25548157

  18. Human Papilloma Virus prevalence and type-specific relative contribution in invasive cervical cancer specimens from Italy

    Lloveras Belén

    2010-06-01

    Full Text Available Abstract Background Cervical cancer represents an important global public health problem. It is the 2nd most common cancer among women worldwide. Human Papillomavirus (HPV infection is now well-established as a necessary cause of invasive cervical cancer (ICC development. Only a few studies on HPV prevalence and type-specific distribution in ICC have been conducted in Italy. Aim To describe the prevalence of HPV and the HPV type-specific distribution in ICC cases identified in Rome, Italy. Methods 140 paraffin embedded tissue blocks of primary ICC diagnosed between 2001 and 2006 were identified at the Regina Elena Cancer Institute in Rome (Italy. HPV was detected through amplification of HPV DNA using SPF-10 HPV broad-spectrum primers followed by DEIA and then genotyping by LiPA25 (version 1. Results 134 cases were considered suitable for HPV DNA detection after histological evaluation; and overall, 90.3% (121/134 HPV prevalence was detected. 111 cases had a single HPV type, 4 cases had an uncharacterized type (HPVX and 6 cases had multiple HPV infections. The five most common single HPV types among positive cases were: HPV16 (71/121; 58.7%, HPV18 (12/121; 9.9%, HPV31, HPV45 and HPV58 (5/121; 4.1% each. 2 (1.5% of the single infections and 2 (1.5% of the multiple infections contained low risk types. Statistically significant differences in the relative contribution of HPV18 were found when comparing squamous cell carcinomas with adenocarcinomas. Conclusions HPV16 and HPV18 accounted for almost 70% of all the HPV positive ICC cases. The study provides baseline information for further evaluation on the impact of recently introduced HPV vaccines in Italy.

  19. Investigating energy deposition within cell populations using Monte Carlo simulations.

    Oliver, Patricia A K; Thomson, Rowan M

    2018-06-27

    In this work, we develop multicellular models of healthy and cancerous human soft tissues, which are used to investigate energy deposition in subcellular targets, quantify the microdosimetric spread in a population of cells, and determine how these results depend on model details. Monte Carlo (MC) tissue models combining varying levels of detail on different length scales are developed: microscopically-detailed regions of interest (>1500 explicitly-modelled cells) are embedded in bulk tissue phantoms irradiated by photons (20 keV to 1.25 MeV). Specific energy (z; energy imparted per unit mass) is scored in nuclei and cytoplasm compartments using the EGSnrc user-code egs_chamber; specific energy mean, <z>, standard deviation, σz, and distribution, f(z,D), are calculated for a variety of macroscopic doses, D. MC-calculated f(z,D) are compared with normal distributions having the same mean and standard deviation. For mGy doses, there is considerable variation in energy deposition (microdosimetric spread) throughout a cell population: e.g., for 30 keV photons irradiating melanoma with 7.5 μm cell radius and 3 μm nuclear radius, σz/<z> for nuclear targets is 170%, and the fraction of nuclei receiving no energy deposition, fz=0, is 0.31 for a dose of 10 mGy. If cobalt-60 photons are considered instead, then σz/<z> decreases to 84%, and fz=0 decreases to 0.036. These results correspond to randomly arranged cells with cell/nucleus sizes randomly sampled from a normal distribution with a standard deviation of 1 μm. If cells are arranged in a hexagonal lattice and cell/nucleus sizes are uniform throughout the population, then σz/<z> decreases to 106% and 68% for 30 keV and cobalt-60,respectively; fz=0

  20. Analysis of proteins of mouse sarcoma pseudotype viruses: type-specific radioimmunoassays for ecotropic virus p30's

    Kennel, S.J.; Tennant, R.W.

    1979-01-01

    Murine sarcoma virus pseudotypes were prepared by infection of nonproducer cells (A1-2), which were transformed by the Gazdar strain of mouse sarcoma virus, with Gross (N-tropic), WN1802B (B-tropic), or Moloney (NB-tropic) viruses. The respective host range pseudotype sarcoma viruses were defined by the tritration characteristics on cells with the appropriate Fv-1 genotype. Proteins from virus progeny were analyzed by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Bands present in both the 65,000- and the 10,000- to 20,000-molecular-weight regions of the gel distinguished the pseudotype viruses from their respective helpers. Furthermore, two protein bands were noted in the p30 region of murine sarcoma virus (Gross), one corresponding to Gross virus p30, and another of slightly slower mobility. However, since the mobility of the putative sarcoma p30 is nearly indentical to that of WN1802B, its presence could not be established by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Type-specific radioimmunossays for Gross virus p30 and for WN1802B p30 were applied for analysis of pseudotype preparations, and among several ecotropic viruses tested, only the homologous virus scored in the respective assay. By use of these assays, pseudotype viruses were found to contain only 8 to 48% helper-specific p30's; the remainder is presumably derived from the sarcoma virus

  1. Human papillomavirus type-specific prevalence in the cervical cancer screening population of Czech women.

    Ruth Tachezy

    Full Text Available BACKGROUND: Infection with high-risk human papillomavirus (HPVtypes has been recognized as a causal factor for the development of cervical cancer and a number of other malignancies. Today, vaccines against HPV, highly effective in the prevention of persistent infection and precancerous lesions, are available for the routine clinical practice. OBJECTIVES: The data on the prevalence and type-specific HPV distribution in the population of each country are crucial for the surveillance of HPV type-specific prevalence at the onset of vaccination against HPV. METHODS: Women attending a preventive gynecological examination who had no history of abnormal cytological finding and/or surgery for cervical lesions were enrolled. All samples were tested for the presence of HPV by High-Risk Hybrid Capture 2 (HR HC2 and by a modified PCR-reverse line blot assay with broad spectrum primers (BS-RLB. RESULTS: Cervical smears of 1393 women were analyzed. In 6.5% of women, atypical cytological findings were detected. Altogether, 28.3% (394/1393 of women were positive for any HPV type by BS-RLB, 18.2% (254/1393 by HR HC2, and 22.3% (310/1393 by BS-RLB for HR HPV types. In women with atypical findings the prevalence for HR and any HPV types were significantly higher than in women with normal cytological findings. Overall, 36 different HPV types were detected, with HPV 16 being the most prevalent (4.8%. HPV positivity decreased with age; the highest prevalence was 31.5% in the age group 21-25 years. CONCLUSIONS: Our study subjects represent the real screening population. HPV prevalence in this population in the Czech Republic is higher than in other countries of Eastern Europe. Also the spectrum of the most prevalent HPV types differs from those reported by others but HPV 16 is, concordantly, the most prevalent type. Country-specific HPV type-specific prevalences provide baseline information which will enable to measure the impact of HPV vaccination in the future.

  2. Indirect micro-immunofluorescence test for detecting type-specific antibodies to herpes simplex virus.

    Forsey, T; Darougar, S

    1980-02-01

    A rapid indirect micro-immunofluorescence test capable of detecting and differentiating type-specific antibodies to herpes simplex virus is described. The test proved highly sensitive and, in 80 patients with active herpes ocular infection, antibody was detected in 94%. No anti-herpes antibody was detected in a control group of 20 patients with adenovirus infections. Testing of animal sera prepared against herpes simplex virus types 1 and 2 and of human sera from cases of ocular and genital herpes infections showed that the test can differentiate antibodies to the infecting serotypes. Specimens of whole blood, taken by fingerprick, and eye secretions, both collected on cellulose sponges, could be tested by indirect micro-immunofluorescence. Anti-herpes IgG, IgM, and IgA can also be detected.

  3. Type-specific proactive interference in patients with semantic and phonological STM deficits.

    Harris, Lara; Olson, Andrew; Humphreys, Glyn

    2014-01-01

    Prior neuropsychological evidence suggests that semantic and phonological components of short-term memory (STM) are functionally and neurologically distinct. The current paper examines proactive interference (PI) from semantic and phonological information in two STM-impaired patients, DS (semantic STM deficit) and AK (phonological STM deficit). In Experiment 1 probe recognition tasks with open and closed sets of stimuli were used. Phonological PI was assessed using nonword items, and semantic and phonological PI was assessed using words. In Experiment 2 phonological and semantic PI was elicited by an item recognition probe test with stimuli that bore phonological and semantic relations to the probes. The data suggested heightened phonological PI for the semantic STM patient, and exaggerated effects of semantic PI in the phonological STM case. The findings are consistent with an account of extremely rapid decay of activated type-specific representations in cases of severely impaired phonological and semantic STM.

  4. Experimental investigation and mathematical modeling of triode PEM fuel cells

    Martino, E.; Koilias, G.; Athanasiou, M.; Katsaounis, A.; Dimakopoulos, Y.; Tsamopoulos, J.; Vayenas, C.G.

    2017-01-01

    Highlights: •The triode fuel cell operation was tested using novel comb-type electrode designs. •Triode operation enhances the PEMFC power output by up to 500%. •Power output enhancement exceeds auxiliary power by up to 20%. •Good agreement with mathematical model based on the laws of Kirchhoff. •Proton fluxes in the membrane found via solution of the Nernst Planck equation -- Abstract: The triode operation of humidified PEM fuel cells has been investigated both with pure H 2 and with CO poisoned H 2 feed over commercial Vulcan supported Pt(30%)-Ru(15%) anodes. It was found that triode operation, which involves the use of a third, auxiliary, electrode, leads to up to 400% power output increase with the same CO poisoned H 2 gas feed. At low current densities, the power increase is accompanied by an increase in overall thermodynamic efficiency. A mathematical model, based on Kirchhoff’s laws, has been developed which is in reasonably good agreement with the experimental results. In order to gain some additional insight into the mechanism of triode operation, the model has been also extended to describe the potential distribution inside the Nafion membrane via the numerical solution of the Nernst-Planck equation. Both model and experiment have shown the critical role of minimizing the auxiliary-anode or auxiliary-cathode resistance, and this has led to improved comb-shaped anode or cathode electrode geometries.

  5. Inference of type-specific HPV transmissibility, progression and clearance rates: a mathematical modelling approach.

    Helen C Johnson

    Full Text Available Quantifying rates governing the clearance of Human Papillomavirus (HPV and its progression to clinical disease, together with viral transmissibility and the duration of naturally-acquired immunity, is essential in estimating the impact of vaccination programmes and screening or testing regimes. However, the complex natural history of HPV makes this difficult. We infer the viral transmissibility, rate of waning natural immunity and rates of progression and clearance of infection of 13 high-risk and 2 non-oncogenic HPV types, making use of a number of rich datasets from Sweden. Estimates of viral transmissibility, clearance of initial infection and waning immunity were derived in a Bayesian framework by fitting a susceptible-infectious-recovered-susceptible (SIRS transmission model to age- and type-specific HPV prevalence data from both a cross-sectional study and a randomised controlled trial (RCT of primary HPV screening. The models fitted well, but over-estimated the prevalence of four high-risk types with respect to the data. Three of these types (HPV-33, -35 and -58 are among the most closely related phylogenetically to the most prevalent HPV-16. The fourth (HPV-45 is the most closely related to HPV-18; the second most prevalent type. We suggest that this may be an indicator of cross-immunity. Rates of progression and clearance of clinical lesions were additionally estimated from longitudinal data gathered as part of the same RCT. Our estimates of progression and clearance rates are consistent with the findings of survival analysis studies and we extend the literature by estimating progression and clearance rates for non-16 and non-18 high-risk types. We anticipate that such type-specific estimates will be useful in the parameterisation of further models and in developing our understanding of HPV natural history.

  6. Rapid, sensitive, type specific PCR detection of the E7 region of human papillomavirus type 16 and 18 from paraffin embedded sections of cervical carcinoma

    Lesnikova, Iana; Lidang, Marianne; Hamilton-Dutoit, Steven

    2010-01-01

    ABSTRACT: Human papillomavirus (HPV) infection, and in particularly infection with HPVs 16 and 18, is a central carcinogenic factor in the uterine cervix. We established and optimized a PCR assay for the detection and discrimination of HPV types 16 and 18 in archival formaldehyde fixed and paraffin...... embedded (FFPE) sections of cervical cancer.Tissue blocks from 35 cases of in situ or invasive cervical squamous cell carcinoma and surrogate FFPE sections containing the cell lines HeLa and SiHa were tested for HPV 16 and HPV18 by conventional PCR using type specific primers, and for the housekeeping gene...

  7. Investigation of microstructural and mechanical properties of cell walls of closed-cell aluminium alloy foams

    Islam, M.A.; Kader, M.A.; Hazell, P.J.; Brown, A.D. [School of Engineering and Information Technology, UNSW Canberra, ACT 2610 (Australia); Saadatfar, M. [Department of Applied Mathematics, Australian National University, Canberra ACT 0200 (Australia); Quadir, M.Z [Electron Microscope Unit, Mark Wainwright Analytical Centre (MWAC), The University of New South Wales, Sydney, NSW 2052 (Australia); Microscopy and Microanalysis Facility (MMF), John de Laeter Centre (JdLC), Curtin University, WA 6102 (Australia); Escobedo, J.P., E-mail: J.Escobedo-Diaz@adfa.edu.au [School of Engineering and Information Technology, UNSW Canberra, ACT 2610 (Australia)

    2016-06-01

    This study investigates the influence of microstructure on the strength properties of individual cell walls of closed-cell stabilized aluminium foams (SAFs). Optical microscopy (OM), micro-computed X-ray tomography (µ-CT), electron backscattering diffraction (EBSD), and energy dispersive X-ray spectroscopy (EDS) analyses were conducted to examine the microstructural properties of SAF cell walls. Novel micro-tensile tests were performed to investigate the strength properties of individual cell walls. Microstructural analysis of the SAF cell walls revealed that the material consists of eutectic Al-Si and dendritic a-Al with an inhomogeneous distribution of intermetallic particles and micro-pores (void defects). These microstructural features affected the micro-mechanism fracture behaviour and tensile strength of the specimens. Laser-based extensometer and digital image correlation (DIC) analyses were employed to observe the strain fields of individual tensile specimens. The tensile failure mode of these materials has been evaluated using microstructural analysis of post-mortem specimens, revealing a brittle cleavage fracture of the cell wall materials. The micro-porosities and intermetallic particles reduced the strength under tensile loading, limiting the elongation to fracture on average to ~3.2% and an average ultimate tensile strength to ~192 MPa. Finally, interactions between crack propagation and obstructing intermetallic compounds during the tensile deformation have been elucidated.

  8. Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

    Leonardo D'Aiuto

    Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

  9. Investigation progress of imaging techniques monitoring stem cell therapy

    Wu Jun; An Rui

    2006-01-01

    Recently stem cell therapy has showed potential clinical application in diabetes mellitus, cardiovascular diseases, malignant tumor and trauma. Efficient techniques of non-invasively monitoring stem cell transplants will accelerate the development of stem cell therapies. This paper briefly reviews the clinical practice of stem cell, in addition, makes a review of monitoring methods including magnetic resonance and radionuclide imaging which have been used in stem cell therapy. (authors)

  10. The role of Sox6 in zebrafish muscle fiber type specification.

    Jackson, Harriet E; Ono, Yosuke; Wang, Xingang; Elworthy, Stone; Cunliffe, Vincent T; Ingham, Philip W

    2015-01-01

    The transcription factor Sox6 has been implicated in regulating muscle fiber type-specific gene expression in mammals. In zebrafish, loss of function of the transcription factor Prdm1a results in a slow to fast-twitch fiber type transformation presaged by ectopic expression of sox6 in slow-twitch progenitors. Morpholino-mediated Sox6 knockdown can suppress this transformation but causes ectopic expression of only one of three slow-twitch specific genes assayed. Here, we use gain and loss of function analysis to analyse further the role of Sox6 in zebrafish muscle fiber type specification. The GAL4 binary misexpression system was used to express Sox6 ectopically in zebrafish embryos. Cis-regulatory elements were characterized using transgenic fish. Zinc finger nuclease mediated targeted mutagenesis was used to analyse the effects of loss of Sox6 function in embryonic, larval and adult zebrafish. Zebrafish transgenic for the GCaMP3 Calcium reporter were used to assay Ca2+ transients in wild-type and mutant muscle fibres. Ectopic Sox6 expression is sufficient to downregulate slow-twitch specific gene expression in zebrafish embryos. Cis-regulatory elements upstream of the slow myosin heavy chain 1 (smyhc1) and slow troponin c (tnnc1b) genes contain putative Sox6 binding sites required for repression of the former but not the latter. Embryos homozygous for sox6 null alleles expressed tnnc1b throughout the fast-twitch muscle whereas other slow-specific muscle genes, including smyhc1, were expressed ectopically in only a subset of fast-twitch fibers. Ca2+ transients in sox6 mutant fast-twitch fibers were intermediate in their speed and amplitude between those of wild-type slow- and fast-twitch fibers. sox6 homozygotes survived to adulthood and exhibited continued misexpression of tnnc1b as well as smaller slow-twitch fibers. They also exhibited a striking curvature of the spine. The Sox6 transcription factor is a key regulator of fast-twitch muscle fiber differentiation

  11. Development and Evaluation of a PCR and Mass Spectroscopy-based (PCR-MS) Method for Quantitative, Type-specific Detection of Human Papillomavirus

    Patel, Divya A.; Shih, Yang-Jen; Newton, Duane W.; Michael, Claire W.; Oeth, Paul A.; Kane, Michael D.; Opipari, Anthony W.; Ruffin, Mack T.; Kalikin, Linda M.; Kurnit, David M.

    2010-01-01

    Knowledge of the central role of high-risk human papillomavirus (HPV) in cervical carcinogenesis, coupled with an emerging need to monitor the efficacy of newly introduced HPV vaccines, warrant development and evaluation of type-specific, quantitative HPV detection methods. In the present study, a prototype PCR and mass spectroscopy (PCR-MS)-based method to detect and quantitate 13 high-risk HPV types is compared to the Hybrid Capture 2 High Risk HPV DNA test (HC2; Digene Corp., Gaithersburg, MD) in 199 cervical scraping samples and to DNA sequencing in 77 cervical tumor samples. High-risk HPV types were detected in 76/77 (98.7%) cervical tumor samples by PCR-MS. Degenerate and type-specific sequencing confirmed the types detected by PCR-MS. In 199 cervical scraping samples, all 13 HPV types were detected by PCR-MS. Eighteen (14.5%) of 124 cervical scraping samples that were positive for high-risk HPV by HC2 were negative by PCR-MS. In all these cases, degenerate DNA sequencing failed to detect any of the 13 high-risk HPV types. Nearly half (46.7%) of the 75 cervical scraping samples that were negative for high-risk HPV by the HC2 assay were positive by PCR-MS. Type-specific sequencing in a subset of these samples confirmed the HPV type detected by PCR-MS. Quantitative PCR-MS results demonstrated that 11/75 (14.7%) samples contained as much HPV copies/cell as HC2-positive samples. These findings suggest that this prototype PCR-MS assay performs at least as well as HC2 for HPV detection, while offering the additional, unique advantages of type-specific identification and quantitation. Further validation work is underway to define clinically meaningful HPV detection thresholds and to evaluate the potential clinical application of future generations of the PCR-MS assay. PMID:19410602

  12. Development and evaluation of a PCR and mass spectroscopy (PCR-MS)-based method for quantitative, type-specific detection of human papillomavirus.

    Patel, Divya A; Shih, Yang-Jen; Newton, Duane W; Michael, Claire W; Oeth, Paul A; Kane, Michael D; Opipari, Anthony W; Ruffin, Mack T; Kalikin, Linda M; Kurnit, David M

    2009-09-01

    Knowledge of the central role of high-risk human papillomavirus (HPV) in cervical carcinogenesis, coupled with an emerging need to monitor the efficacy of newly introduced HPV vaccines, warrant development and evaluation of type-specific, quantitative HPV detection methods. In the present study, a prototype PCR and mass spectroscopy (PCR-MS)-based method to detect and quantitate 13 high-risk HPV types is compared to the Hybrid Capture 2 High-Risk HPV DNA test (HC2; Digene Corp., Gaithersburg, MD) in 199 cervical scraping samples and to DNA sequencing in 77 cervical tumor samples. High-risk HPV types were detected in 76/77 (98.7%) cervical tumor samples by PCR-MS. Degenerate and type-specific sequencing confirmed the types detected by PCR-MS. In 199 cervical scraping samples, all 13 HPV types were detected by PCR-MS. Eighteen (14.5%) of 124 cervical scraping samples that were positive for high-risk HPV by HC2 were negative by PCR-MS. In all these cases, degenerate DNA sequencing failed to detect any of the 13 high-risk HPV types. Nearly half (46.7%) of the 75 cervical scraping samples that were negative for high-risk HPV by the HC2 assay were positive by PCR-MS. Type-specific sequencing in a subset of these samples confirmed the HPV type detected by PCR-MS. Quantitative PCR-MS results demonstrated that 11/75 (14.7%) samples contained as much HPV copies/cell as HC2-positive samples. These findings suggest that this prototype PCR-MS assay performs at least as well as HC2 for HPV detection, while offering the additional, unique advantages of type-specific identification and quantitation. Further validation work is underway to define clinically meaningful HPV detection thresholds and to evaluate the potential clinical application of future generations of the PCR-MS assay.

  13. Investigation of aluminosilicate refractory for solid oxide fuel cell applications

    Gentile, Paul Steven

    Stationary solid oxide fuel cells (SOFCs) have been demonstrated to provide clean and reliable electricity through electro-chemical conversion of various fuel sources (CH4 and other light hydrocarbons). To become a competitive conversion technology the costs of SOFCs must be reduced to less than $400/kW. Aluminosilicate represents a potential low cost alternative to high purity alumina for SOFC refractory applications. The objectives of this investigation are to: (1) study changes of aluminosilicate chemistry and morphology under SOFC conditions, (2) identify volatile silicon species released by aluminosilicates, (3) identify the mechanisms of aluminosilicate vapor deposition on SOFC materials, and (4) determine the effects of aluminosilicate vapors on SOFC electrochemical performance. It is shown thermodynamically and empirically that low cost aluminosilicate refractory remains chemically and thermally unstable under SOFC operating conditions between 800°C and 1000°C. Energy dispersive spectroscopy (EDS) and X-ray photoelectron spectroscopy (XPS) of the aluminosilicate bulk and surface identified increased concentrations of silicon at the surface after exposure to SOFC gases at 1000°C for 100 hours. The presence of water vapor accelerated surface diffusion of silicon, creating a more uniform distribution. Thermodynamic equilibrium modeling showed aluminosilicate remains stable in dry air, but the introduction of water vapor indicative of actual SOFC gas streams creates low temperature (active anode interface.

  14. Investigation of materials for inert electrodes in aluminum electrodeposition cells

    Haggerty, J. S.; Sadoway, D. R.

    1987-09-14

    Work was divided into major efforts. The first was the growth and characterization of specimens; the second was Hall cell performance testing. Cathode and anode materials were the subject of investigation. Preparation of specimens included growth of single crystals and synthesis of ultra high purity powders. Special attention was paid to ferrites as they were considered to be the most promising anode materials. Ferrite anode corrosion rates were studied and the electrical conductivities of a set of copper-manganese ferrites were measured. Float Zone, Pendant Drop Cryolite Experiments were undertaken because unsatisfactory choices of candidate materials were being made on the basis of a flawed set of selection criteria applied to an incomplete and sometimes inaccurate data base. This experiment was then constructed to determine whether the apparatus used for float zone crystal growth could be adapted to make a variety of important based melts and their interactions with candidate inert anode materials. The third major topic was Non Consumable Anode (Data Base, Candidate Compositions), driven by our perception that the basis for prior selection of candidate materials was inadequate. Results are presented. 162 refs., 39 figs., 18 tabs.

  15. Human muscle fiber type-specific insulin signaling: Impact of obesity and type 2 diabetes

    Albers, Peter Hjorth; Pedersen, Andreas J T; Birk, Jesper Bratz

    2015-01-01

    Skeletal muscle is a heterogeneous tissue composed of different fiber types. Studies suggest that insulin-mediated glucose metabolism is different between muscle fiber types. We hypothesized that differences are due to fiber-type specific expression/regulation of insulin signaling elements and....../or metabolic enzymes. Pools of type I and II fibers were prepared from biopsies of the vastus lateralis muscles from lean, obese and type 2 diabetic subjects before and after a hyperinsulinemic-euglycemic clamp. Type I fibers compared to type II fibers have higher protein levels of the insulin receptor, GLUT4......, hexokinase II, glycogen synthase (GS), pyruvate dehydrogenase (PDH-E1α) and a lower protein content of Akt2, TBC1D4 and TBC1D1. In type I fibers compared to type II fibers, the phosphorylation-response to insulin was similar (TBC1D4, TBC1D1 and GS) or decreased (Akt and PDH-E1α). Phosphorylation...

  16. Systematic review of type-specific pathophysiological symptoms of Sasang typology

    Yoo Ri Han

    2016-06-01

    Full Text Available Previous studies on the Sasang typology have focused on the differential diagnosis of each Sasang type with type-specific pathophysiological symptoms (TSPS. The purpose of this study was to elucidate the latent physiological mechanism related to these clinical indicators. We searched six electronic databases for articles published from 1990 to 2015 using the Sasang typology-related keywords, and found and analyzed 35 such articles. The results were summarized into six TSPS categories: perspiration, temperature preference, sleep, defecation, urination, and susceptibility to stress. The Tae-Eum and So-Eum types showed contrasting features with TSPS, and the So-Yang type was in the middle. The Tae-Eum type has good digestive function, regular bowel movement and defecation, high sleep quality, and low susceptibility to stress and cold. The Tae-Eum type has relatively large volumes of sweat and feels fresh after sweating; however, the urine is highly concentrated. These clinical features might be related to the biopsychological traits of the Tae-Eum type, including a low trait anxiety level and high ponderal and body mass indices. This study used the autonomic reactivity hypothesis for explaining the pathophysiological predispositions in the Sasang typology. The Tae-Eum and So-Eum Sasang types have a low threshold in parasympathetic and sympathetic activation, respectively. This study provides a foundation for integrating traditional Korean personalized medicine and Western biomedicine.

  17. Investigation of back surface fields effect on bifacial solar cells

    Sepeai, Suhaila; Sulaiman, M. Y.; Sopian, Kamaruzzaman; Zaidi, Saleem H.

    2012-11-01

    A bifacial solar cell, in contrast with a conventional monofacial solar cell, produces photo-generated current from both front and back sides. Bifacial solar cell is an attractive candidate for enhancing photovoltaic (PV) market competitiveness as well as supporting the current efforts to increase efficiency and lower material costs. This paper reports on the fabrication of bifacial solar cells using phosphorus-oxytrichloride (POCl3) emitter formation on p-type, nanotextured silicon (Si) wafer. Backside surface field was formed through Al-diffusion using conventional screen-printing process. Bifacial solar cells with a structure of n+pp+ with and without back surface field (BSF) were fabricated in which silicon nitride (SiN) anti reflection and passivation films were coated on both sides, followed by screen printing of Argentum (Ag) and Argentum/Aluminum (Ag/Al) on front and back contacts, respectively. Bifacial solar cells without BSF exhibited open circuit voltage (VOC) of 535 mV for front and 480 mV for back surface. With Al-alloyed BSF bifacial solar cells, the VOC improved to 580 mV for the front surface and 560 mV for the back surface. Simulation of bifacial solar cells using PC1D and AFORS software demonstrated good agreement with experimental results. Simulations showed that best bifacial solar cells are achieved through a combination of high lifetime wafer, low recombination back surface field, reduced contact resistance, and superior surface passivation.

  18. Live-cell imaging: new avenues to investigate retinal regeneration

    Manuela Lahne

    2017-01-01

    Full Text Available Sensing and responding to our environment requires functional neurons that act in concert. Neuronal cell loss resulting from degenerative diseases cannot be replaced in humans, causing a functional impairment to integrate and/or respond to sensory cues. In contrast, zebrafish (Danio rerio possess an endogenous capacity to regenerate lost neurons. Here, we will focus on the processes that lead to neuronal regeneration in the zebrafish retina. Dying retinal neurons release a damage signal, tumor necrosis factor α, which induces the resident radial glia, the Müller glia, to reprogram and re-enter the cell cycle. The Müller glia divide asymmetrically to produce a Müller glia that exits the cell cycle and a neuronal progenitor cell. The arising neuronal progenitor cells undergo several rounds of cell divisions before they migrate to the site of damage to differentiate into the neuronal cell types that were lost. Molecular and immunohistochemical studies have predominantly provided insight into the mechanisms that regulate retinal regeneration. However, many processes during retinal regeneration are dynamic and require live-cell imaging to fully discern the underlying mechanisms. Recently, a multiphoton imaging approach of adult zebrafish retinal cultures was developed. We will discuss the use of live-cell imaging, the currently available tools and those that need to be developed to advance our knowledge on major open questions in the field of retinal regeneration.

  19. Muscle-Type Specific Autophosphorylation of CaMKII Isoforms after Paced Contractions

    Wouter Eilers

    2014-01-01

    Full Text Available We explored to what extent isoforms of the regulator of excitation-contraction and excitation-transcription coupling, calcium/calmodulin protein kinase II (CaMKII contribute to the specificity of myocellular calcium sensing between muscle types and whether concentration transients in its autophosphorylation can be simulated. CaMKII autophosphorylation at Thr287 was assessed in three muscle compartments of the rat after slow or fast motor unit-type stimulation and was compared against a computational model (CaMuZclE coupling myocellular calcium dynamics with CaMKII Thr287 phosphorylation. Qualitative differences existed between fast- (gastrocnemius medialis and slow-type muscle (soleus for the expression pattern of CaMKII isoforms. Phospho-Thr287 content of δA CaMKII, associated with nuclear functions, demonstrated a transient and compartment-specific increase after excitation, which contrasted to the delayed autophosphorylation of the sarcoplasmic reticulum-associated βM CaMKII. In soleus muscle, excitation-induced δA CaMKII autophosphorylation demonstrated frequency dependence (P = 0.02. In the glycolytic compartment of gastrocnemius medialis, CaMKII autophosphorylation after excitation was blunted. In silico assessment emphasized the importance of mitochondrial calcium buffer capacity for excitation-induced CaMKII autophosphorylation but did not predict its isoform specificity. The findings expose that CaMKII autophosphorylation with paced contractions is regulated in an isoform and muscle type-specific fashion and highlight properties emerging for phenotype-specific regulation of CaMKII.

  20. World Health Organization Guidelines for Containment of Poliovirus Following Type-Specific Polio Eradication - Worldwide, 2015.

    Previsani, Nicoletta; Tangermann, Rudolph H; Tallis, Graham; Jafari, Hamid S

    2015-08-28

    In 1988, the World Health Assembly of the World Health Organization (WHO) resolved to eradicate polio worldwide. Among the three wild poliovirus (WPV) types (type 1, type 2, and type 3), WPV type 2 (WPV2) has been eliminated in the wild since 1999, and WPV type 3 (WPV3) has not been reported since 2012. In 2015, only Afghanistan and Pakistan have reported WPV transmission. On May 25, 2015, all WHO Member States endorsed World Health Assembly resolution 68.3 on full implementation of the Polio Eradication and Endgame Strategic Plan 2013-2018 (the Endgame Plan), and with it, the third Global Action Plan to minimize poliovirus facility-associated risk (GAPIII). All WHO Member States have committed to implementing appropriate containment of WPV2 in essential laboratory and vaccine production facilities* by the end of 2015 and of type 2 oral poliovirus vaccine (OPV2) within 3 months of global withdrawal of OPV2, which is planned for April 2016. This report summarizes critical steps for essential laboratory and vaccine production facilities that intend to retain materials confirmed to contain or potentially containing type-specific WPV, vaccine-derived poliovirus (VDPV), or OPV/Sabin viruses, and steps for nonessential facilities† that process specimens that contain or might contain polioviruses. National authorities will need to certify that the essential facilities they host meet the containment requirements described in GAPIII. After certification of WPV eradication, the use of all OPV will cease; final containment of all polioviruses after polio eradication and OPV cessation will minimize the risk for reintroduction of poliovirus into a polio-free world.

  1. Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer

    Tai Phillip WL

    2011-07-01

    Full Text Available Abstract Background Hundreds of genes, including muscle creatine kinase (MCK, are differentially expressed in fast- and slow-twitch muscle fibers, but the fiber type-specific regulatory mechanisms are not well understood. Results Modulatory region 1 (MR1 is a 1-kb regulatory region within MCK intron 1 that is highly active in terminally differentiating skeletal myocytes in vitro. A MCK small intronic enhancer (MCK-SIE containing a paired E-box/myocyte enhancer factor 2 (MEF2 regulatory motif resides within MR1. The SIE's transcriptional activity equals that of the extensively characterized 206-bp MCK 5'-enhancer, but the MCK-SIE is flanked by regions that can repress its activity via the individual and combined effects of about 15 different but highly conserved 9- to 24-bp sequences. ChIP and ChIP-Seq analyses indicate that the SIE and the MCK 5'-enhancer are occupied by MyoD, myogenin and MEF2. Many other E-boxes located within or immediately adjacent to intron 1 are not occupied by MyoD or myogenin. Transgenic analysis of a 6.5-kb MCK genomic fragment containing the 5'-enhancer and proximal promoter plus the 3.2-kb intron 1, with and without MR1, indicates that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers (types I and IIa, respectively, but is not required for expression in fast-twitch muscle fibers (types IIb and IId. Conclusions In this study, we discovered that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers and that MR1's positive transcriptional activity depends on a paired E-box MEF2 site motif within a SIE. This is the first study to delineate the DNA controls for MCK expression in different skeletal muscle fiber types.

  2. Investigating evolutionary conservation of dendritic cell subset identity and functions

    Thien-Phong eVu Manh

    2015-06-01

    Full Text Available Dendritic cells (DC were initially defined as mononuclear phagocytes with a dendritic morphology and an exquisite efficiency for naïve T cell activation. DC encompass several subsets initially identified by their expression of specific cell surface molecules and later shown to excel in distinct functions and to develop under the instruction of different transcription factors or cytokines. Very few cell surface molecules are expressed in a specific manner on any immune cell type. Hence, to identify cell types, the sole use of a small number of cell surface markers in classical flow cytometry can be deceiving. Moreover, the markers currently used to define mononuclear phagocyte subsets vary depending on the tissue and animal species studied and even between laboratories. This has led to confusion in the definition of DC subset identity and in their attribution of specific functions. There is a strong need to identify a rigorous and consensus way to define mononuclear phagocyte subsets, with precise guidelines potentially applicable throughout tissues and species. We will discuss the advantages, drawbacks and complementarities of different methodologies: cell surface phenotyping, ontogeny, functional characterization and molecular profiling. We will advocate that gene expression profiling is a very rigorous, largely unbiased and accessible method to define the identity of mononuclear phagocyte subsets, which strengthens and refines surface phenotyping. It is uniquely powerful to yield new, experimentally testable, hypotheses on the ontogeny or functions of mononuclear phagocyte subsets, their molecular regulation and their evolutionary conservation. We propose defining cell populations based on a combination of cell surface phenotyping, expression analysis of hallmark genes and robust functional assays, in order to reach a consensus and integrate faster the huge but scattered knowledge accumulated by different laboratories on different cell types

  3. Muscle fiber type specific induction of slow myosin heavy chain 2 gene expression by electrical stimulation

    Crew, Jennifer R.; Falzari, Kanakeshwari; DiMario, Joseph X.

    2010-01-01

    Vertebrate skeletal muscle fiber types are defined by a broad array of differentially expressed contractile and metabolic protein genes. The mechanisms that establish and maintain these different fiber types vary throughout development and with changing functional demand. Chicken skeletal muscle fibers can be generally categorized as fast and fast/slow based on expression of the slow myosin heavy chain 2 (MyHC2) gene in fast/slow muscle fibers. To investigate the cellular and molecular mechanisms that control fiber type formation in secondary or fetal muscle fibers, myoblasts from the fast pectoralis major (PM) and fast/slow medial adductor (MA) muscles were isolated, allowed to differentiate in vitro, and electrically stimulated. MA muscle fibers were induced to express the slow MyHC2 gene by electrical stimulation, whereas PM muscle fibers did not express the slow MyHC2 gene under identical stimulation conditions. However, PM muscle fibers did express the slow MyHC2 gene when electrical stimulation was combined with inhibition of inositol triphosphate receptor (IP3R) activity. Electrical stimulation was sufficient to increase nuclear localization of expressed nuclear-factor-of-activated-T-cells (NFAT), NFAT-mediated transcription, and slow MyHC2 promoter activity in MA muscle fibers. In contrast, both electrical stimulation and inhibitors of IP3R activity were required for these effects in PM muscle fibers. Electrical stimulation also increased levels of peroxisome-proliferator-activated receptor-γ co-activator-1 (PGC-1α) protein in PM and MA muscle fibers. These results indicate that MA muscle fibers can be induced by electrical stimulation to express the slow MyHC2 gene and that fast PM muscle fibers are refractory to stimulation-induced slow MyHC2 gene expression due to fast PM muscle fiber specific cellular mechanisms involving IP3R activity.

  4. Investigation of high-rate lithium-thionyl chloride cells

    Hayes, Catherine A.; Gust, Steven; Farrington, Michael D.; Lockwood, Judith A.; Donaldson, George J.

    Chemical analysis of a commercially produced high-rate D-size lithium-thionyl cell was carried out, as a function of rate of discharge (1 ohm and 5 ohms), depth of discharge, and temperature (25 C and -40 C), using specially developed methods for identifying suspected minor cell products or impurities which may effect cell performance. These methods include a product-retrieval system which involves solvent extraction to enhance the recovery of suspected semivolatile minor chemicals, and methods of quantitative GC analysis of volatile and semivolatile products. The nonvolatile products were analyzed by wet chemical methods. The results of the analyses indicate that the predominant discharge reaction in this cell is 4Li + 2SOCl2 going to 4LiCl + S + SO2, with SO2 formation decreasing towards the end of cell life (7 to 12 Ah). The rate of discharge had no effect on the product distribution. Upon discharge of the high-rate cell at -40 C, one cell exploded, and all others exhibited overheating and rapid internal pressure rise when allowed to warm up to room temperature.

  5. Diffusion cell investigations into the acidic degradation of organic coatings

    Møller, Victor Buhl; Wang, Ting; Dam-Johansen, Kim

    2018-01-01

    Protective organic coatings work by preventing contact between an aggressive environment and a vulnerable substrate. However, the long required lifetime of a barrier coating provides a challenge when attempting to evaluate coating performance. Diffusion cells can be used as a tool to estimate...... coating barrier properties and lifetime. In this work, a diffusion cell array was designed, constructed, and compared to previous designs, with simplicity being the most important design parameter. Sulfuric acid diffusion through five different coatings was monitored using a battery of cells...

  6. Investigations of Laser Pumped Gas Cell Atomic Frequency Standard

    Volk, C. H; Camparo, J. C; Frueholz, R. P

    1981-01-01

    Recently it has been suggested that the performance characteristics of a rubidium gas cell atomic frequency standard might be improved by replacing the standard rubidium discharge lamp with a single mode laser diode...

  7. Autoradiographic investigations of cells from the cerebrospinal fluid

    Thamm, W.

    1983-01-01

    A total of 155 samples of cerebrospinal fluid obtained from 61 patients were subjected to cytological examination and incubated together with 3 1 H-thymidine. Radioactive labelling was thus achieved for lymphocytes, monocytes and tumour cells. The highest contents of radioactivity were seen in tumour cells. To a lesser degree had the isotope become attached to the lymphocytes, while the tendency to take up radioactivity was lowest in cells from autochthonous tumours of the brain. In cerebral and meningeal metastases the labelling index rose proportionately to the progress of the disease. Autoradiography can be used to monitor cytostatic treatment carried out to control the spread of meningeal carcinoma cells. In non-inflammatory disorders, lymphocytes and monocytes of the cerebrospinal fluid only rarely show radioactivity. A suprisignly high labelling index was determined for lymphocytes and monocytes of cerebrospinal fluid from young children, which was considered to be attributable to the high proliferation rates to be expected here. (orig./MG) [de

  8. Quantitative image analysis for investigating cell-matrix interactions

    Burkel, Brian; Notbohm, Jacob

    2017-07-01

    The extracellular matrix provides both chemical and physical cues that control cellular processes such as migration, division, differentiation, and cancer progression. Cells can mechanically alter the matrix by applying forces that result in matrix displacements, which in turn may localize to form dense bands along which cells may migrate. To quantify the displacements, we use confocal microscopy and fluorescent labeling to acquire high-contrast images of the fibrous material. Using a technique for quantitative image analysis called digital volume correlation, we then compute the matrix displacements. Our experimental technology offers a means to quantify matrix mechanics and cell-matrix interactions. We are now using these experimental tools to modulate mechanical properties of the matrix to study cell contraction and migration.

  9. Investigations on quinquethiophenes as donor materials in organic solar cells

    Schulze, Kerstin

    2008-01-01

    Organic photovoltaics could in the future represent a possibility for energy production from renewable energy sources. The advance consists here first of all in the potential of a very reasonable fabrication, for instance a production in the role-to-role procedurre, which can be prusued so on flexible substrates like for instance foils. Although the material costs are low, until the commercialization of organic solar cells among others an increasement of their power efficiency is necessary. Preferably in organic solar cells donor and acceptor materials should be applied, the absorption spectra and energy levels of which are ideally matched, because so can high zero-current voltages be reached. Additionally high absorption coefficents of the materials over a large spectral range can lead to high current densities in these photovoltaic components. In this thesis novel quinquethiophenes as donors in organic solar cells are studied, which consist as basic unit of five thiophene rings as well as dicyanovinyl end groups and alkyl side chains. The studied materials possess a high absorption coefficient and reach because of the high ionization potential high zero-current voltages in organic solar cells under application of the fullerenet C 60 as acceptor. Simultaneously a efficient separation of the excitons on the acceptor-donor interface occurs. However the high ionization potential of the quinquethiophenes puts special requirements to the further solar-cell structure. Within this thesis it is shown that adifference between internal voltage and zero-current voltage influences decidingly the shape of the solar-cell characteristic and can generate a S-shape in the neighbourhood of the zero-current voltage. The internal voltage is hereby determined by the contacting of the photoactive layers. An increasement of the internal voltage of the solar cell can be reached by a corresponding material choice. So in this thesis it is shown that organic solar cells based on these

  10. Spin-cast bulk heterojunction solar cells: A dynamical investigation

    Chou, Kang Wei

    2013-02-22

    Spin-coating is extensively used in the lab-based manufacture of organic solar cells, including most of the record-setting solution-processed cells. We report the first direct observation of photoactive layer formation as it occurs during spin-coating. The study provides new insight into mechanisms and kinetics of bulk heterojunction formation, which may be crucial for its successful transfer to scalable printing processes. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Electrocatalysis of fuel cell reactions: Investigation of alternate electrolytes

    Chin, D. T.; Hsueh, K. L.; Chang, H. H.

    1984-01-01

    Oxygen reduction and transport properties of the electrolyte in the phosphoric acid fuel cell are studied. The areas covered were: (1) development of a theoretical expression for the rotating ring disk electrode technique; (2) determination of the intermediate reaction rate constants for oxygen reduction on platinum in phosphoric acid electrolyte; (3) determination of oxygen reduction mechanism in trifluoreomethanesulfonic acid (TFMSA) which was considered as an alternate electrolyte for the acid fuel cells; and (4) the measurement of transport properties of the phosphoric acid electrolyte at high concentrations and temperatures.

  12. Spin-cast bulk heterojunction solar cells: A dynamical investigation

    Chou, Kang Wei; Yan, Buyi; Li, Ruipeng; Li, Erqiang; Zhao, Kui; Anjum, Dalaver H.; Alvarez, Steven; Gassaway, Robert; Biocca, Alan K.; Thoroddsen, Sigurdur T; Hexemer, Alexander; Amassian, Aram

    2013-01-01

    Spin-coating is extensively used in the lab-based manufacture of organic solar cells, including most of the record-setting solution-processed cells. We report the first direct observation of photoactive layer formation as it occurs during spin-coating. The study provides new insight into mechanisms and kinetics of bulk heterojunction formation, which may be crucial for its successful transfer to scalable printing processes. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Persistence of type-specific human papillomavirus infection and increased long-term risk of cervical cancer.

    Chen, Hui-Chi; Schiffman, Mark; Lin, Ching-Yu; Pan, Mei-Hung; You, San-Lin; Chuang, Li-Chung; Hsieh, Chang-Yao; Liaw, Kai-Li; Hsing, Ann W; Chen, Chien-Jen

    2011-09-21

    Human papillomavirus (HPV) persistence is the pivotal event in cervical carcinogenesis. We followed a large-scale community-based cohort for 16 years to investigate the role of genotype-specific HPV persistence in predicting cervical cancer including invasive and in situ carcinoma. At the baseline examination in 1991-1992, 11,923 participants (aged 30-65 years) consented to HPV testing and cytology; 6923 participants were reexamined in 1993-1995. For HPV testing, we used a polymerase chain reaction-based assay that detected 39 HPV types. Women who developed cervical cancer were identified from cancer and death registries. Cumulative risks for developing cervical cancer among infected and persistently infected women were calculated by the Kaplan-Meier method. Of 10,123 women who were initially cytologically normal, 68 developed cervical cancer. The 16-year cumulative risks of subsequent cervical cancer for women with HPV16, HPV58 (without HPV16), or other carcinogenic HPV types (without HPV16 or HPV58) were 13.5%, 10.3%, and 4.0%, respectively, compared with 0.26% for HPV-negative women. Women with type-specific persistence of any carcinogenic HPV had greatly increased risk compared with women who were HPV-negative at both visits (hazard ratio = 75.4, 95% confidence interval = 31.8 to 178.9). The cumulative cervical cancer risks following persistent carcinogenic HPV infections increased with age: The risks were 5.5%, 14.4%, and 18.1% for women aged 30-44 years, 45-54 years, and 55 years and older, respectively. However, newly acquired infections were associated with a low risk of cervical cancer regardless of age. HPV negativity was associated with a very low long-term risk of cervical cancer. Persistent detection of HPV among cytologically normal women greatly increased risk. Thus, it is useful to perform repeated HPV testing following an initial positive test.

  14. Investigating the gene expression profiles of metastasising sarcoma cells

    Cavanna, T.; Pokorná, Eva; Veselý, Pavel; Zicha, D.

    2005-01-01

    Roč. 40, č. 1 (2005), s. 37 ISSN 0035-9017. [Cytokinematics 2004. International Symposium on Microscopy of Live Cells in the Post Genomics Era /8./. 05.09.2004-07.09.2004, Hradec Králové] Institutional research plan: CEZ:AV0Z5052915 Keywords : sarcoma * metastasis Subject RIV: EB - Genetics ; Molecular Biology

  15. Investigation of adhesion and mechanical properties of human glioma cells by single cell force spectroscopy and atomic force microscopy.

    Andolfi, Laura; Bourkoula, Eugenia; Migliorini, Elisa; Palma, Anita; Pucer, Anja; Skrap, Miran; Scoles, Giacinto; Beltrami, Antonio Paolo; Cesselli, Daniela; Lazzarino, Marco

    2014-01-01

    Active cell migration and invasion is a peculiar feature of glioma that makes this tumor able to rapidly infiltrate into the surrounding brain tissue. In our recent work, we identified a novel class of glioma-associated-stem cells (defined as GASC for high-grade glioma--HG--and Gasc for low-grade glioma--LG) that, although not tumorigenic, act supporting the biological aggressiveness of glioma-initiating stem cells (defined as GSC for HG and Gsc for LG) favoring also their motility. Migrating cancer cells undergo considerable molecular and cellular changes by remodeling their cytoskeleton and cell interactions with surrounding environment. To get a better understanding about the role of the glioma-associated-stem cells in tumor progression, cell deformability and interactions between glioma-initiating stem cells and glioma-associated-stem cells were investigated. Adhesion of HG/LG-cancer cells on HG/LG-glioma-associated stem cells was studied by time-lapse microscopy, while cell deformability and cell-cell adhesion strengths were quantified by indentation measurements by atomic force microscopy and single cell force spectroscopy. Our results demonstrate that for both HG and LG glioma, cancer-initiating-stem cells are softer than glioma-associated-stem cells, in agreement with their neoplastic features. The adhesion strength of GSC on GASC appears to be significantly lower than that observed for Gsc on Gasc. Whereas, GSC spread and firmly adhere on Gasc with an adhesion strength increased as compared to that obtained on GASC. These findings highlight that the grade of glioma-associated-stem cells plays an important role in modulating cancer cell adhesion, which could affect glioma cell migration, invasion and thus cancer aggressiveness. Moreover this work provides evidence about the importance of investigating cell adhesion and elasticity for new developments in disease diagnostics and therapeutics.

  16. Investigations on the insulin receptor of isolated fat cells

    Eichler, W.

    1980-01-01

    Fat cells, isolated from the epididymal adipose tissue of rats, were incubed with iodine 125 insulin after previous incubation with various antagonists. By varying the antagonist concentration, it was possible to determine the effect these substances have on the insulin receptor, i.e. the insulin similarity. By varying the preincubation time, toxicity of the test substances could be detected, which pretended repression effects; and by finally verying the incubation time the effects on the receptor via the membrane could be distinguished from direct receptor bindings of the antagonist. (orig./MG) [de

  17. Investigating Aspergillus nidulans secretome during colonisation of cork cell walls.

    Martins, Isabel; Garcia, Helga; Varela, Adélia; Núñez, Oscar; Planchon, Sébastien; Galceran, Maria Teresa; Renaut, Jenny; Rebelo, Luís P N; Silva Pereira, Cristina

    2014-02-26

    Cork, the outer bark of Quercus suber, shows a unique compositional structure, a set of remarkable properties, including high recalcitrance. Cork colonisation by Ascomycota remains largely overlooked. Herein, Aspergillus nidulans secretome on cork was analysed (2DE). Proteomic data were further complemented by microscopic (SEM) and spectroscopic (ATR-FTIR) evaluation of the colonised substrate and by targeted analysis of lignin degradation compounds (UPLC-HRMS). Data showed that the fungus formed an intricate network of hyphae around the cork cell walls, which enabled polysaccharides and lignin superficial degradation, but probably not of suberin. The degradation of polysaccharides was suggested by the identification of few polysaccharide degrading enzymes (β-glucosidases and endo-1,5-α-l-arabinosidase). Lignin degradation, which likely evolved throughout a Fenton-like mechanism relying on the activity of alcohol oxidases, was supported by the identification of small aromatic compounds (e.g. cinnamic acid and veratrylaldehyde) and of several putative high molecular weight lignin degradation products. In addition, cork recalcitrance was corroborated by the identification of several protein species which are associated with autolysis. Finally, stringent comparative proteomics revealed that A. nidulans colonisation of cork and wood share a common set of enzymatic mechanisms. However the higher polysaccharide accessibility in cork might explain the increase of β-glucosidase in cork secretome. Cork degradation by fungi remains largely overlook. Herein we aimed at understanding how A. nidulans colonise cork cell walls and how this relates to wood colonisation. To address this, the protein species consistently present in the secretome were analysed, as well as major alterations occurring in the substrate, including lignin degradation compounds being released. The obtained data demonstrate that this fungus has superficially attacked the cork cell walls apparently by

  18. Numerical cell model investigating cellular carbon fluxes in Emiliania huxleyi.

    Holtz, Lena-Maria; Wolf-Gladrow, Dieter; Thoms, Silke

    2015-01-07

    Coccolithophores play a crucial role in the marine carbon cycle and thus it is interesting to know how they will respond to climate change. After several decades of research the interplay between intracellular processes and the marine carbonate system is still not well understood. On the basis of experimental findings given in literature, a numerical cell model is developed that describes inorganic carbon fluxes between seawater and the intracellular sites of calcite precipitation and photosynthetic carbon fixation. The implemented cell model consists of four compartments, for each of which the carbonate system is resolved individually. The four compartments are connected to each other via H(+), CO2, and HCO3(-) fluxes across the compartment-confining membranes. For CO2 accumulation around RubisCO, an energy-efficient carbon concentrating mechanism is proposed that relies on diffusive CO2 uptake. At low external CO2 concentrations and high light intensities, CO2 diffusion does not suffice to cover the carbon demand of photosynthesis and an additional uptake of external HCO3(-) becomes essential. The model is constrained by data of Emiliania huxleyi, the numerically most abundant coccolithophore species in the present-day ocean. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  19. Experimental investigation two phase flow in direct methanol fuel cells

    Mat, M. D.; Kaplan, Y.; Celik, S.; Oeztural, A.

    2007-01-01

    Direct methanol fuel cells (DMFC) have received many attentions specifically for portable electronic applications since it utilize methanol which is in liquid form in atmospheric condition and high energy density of the methanol. Thus it eliminates the storage problem of hydrogen. It also eliminates humidification requirement of polymeric membrane which is a problem in PEM fuel cells. Some electronic companies introduced DMFC prototypes for portable electronic applications. Presence of carbon dioxide gases due to electrochemical reactions in anode makes the problem a two phase problem. A two phase flow may occur at cathode specifically at high current densities due to the excess water. Presence of gas phase in anode region and liquid phase in cathode region prevents diffusion of fuel and oxygen to the reaction sites thus reduces the performance of the system. Uncontrolled pressure buildup in anode region increases methanol crossover through membrane and adversely effect the performance. Two phase flow in both anode and cathode region is very effective in the performance of DMYC system and a detailed understanding of two phase flow for high performance DMFC systems. Although there are many theoretical and experimental studies available on the DMFC systems in the literature, only few studies consider problem as a two-phase flow problem. In this study, an experimental set up is developed and species distributions on system are measured with a gas chromatograph. System performance characteristics (V-I curves) is measured depending on the process parameters (temperature, fuel ad oxidant flow rates, methanol concentration etc)

  20. Investigation of Ruthenium Dissolution in Advanced Membrane Electrode Assemblies for Direct Methanol Based Fuel Cells Stacks

    Valdez, T. I.; Firdosy, S.; Koel, B. E.; Narayanan, S. R.

    2005-01-01

    This viewgraph presentation gives a detailed review of the Direct Methanol Based Fuel Cell (DMFC) stack and investigates the Ruthenium that was found at the exit of the stack. The topics include: 1) Motivation; 2) Pathways for Cell Degradation; 3) Cell Duration Testing; 4) Duration Testing, MEA Analysis; and 5) Stack Degradation Analysis.

  1. Serial type-specific human papillomavirus (HPV) load measurement allows differentiation between regressing cervical lesions and serial virion productive transient infections

    Depuydt, Christophe E; Jonckheere, Jef; Berth, Mario; Salembier, Geert M; Vereecken, Annie J; Bogers, Johannes J

    2015-01-01

    Persistent high-risk human papillomavirus (HPV) infection is strongly associated with the development of high-grade cervical intraepithelial neoplasia (CIN) or cancer. Not all persistent infections lead to cancer. Viral load measured at a single time-point is a poor predictor of the natural history of HPV infections. However the profile of viral load evolution over time could distinguish nonprogressive from progressive (carcinogenic) infections. A retrospective natural history study was set up using a Belgian laboratory database including more than 800,000 liquid cytology specimens. All samples were submitted to qPCR identifying E6/E7 genes of 18 HPV types. Viral load changes over time were assessed by the linear regression slope. Database search identified 261 untreated women with persistent type-specific HPV DNA detected (270 infections) in at least three of the last smears for a average period of 3.2 years. Using the coefficient of determination (R²) infections could be subdivided in a latency group (n = 143; R² < 0.85) and a regressing group (n = 127; R² ≥ 0.85). In (≥3) serial viral load measurements, serial transient infections with latency is characterized by a nonlinear limited difference in decrease or increase of type-specific viral load (R² < 0.85 and slopes between 2 measurements 0.0010 and −0.0010 HPV copies/cell per day) over a longer period of time (1553 days), whereas regression of a clonal cell population is characterized by a linear (R² ≥ 0.85) decrease (−0.0033 HPV copies/cell per day) over a shorter period of time (708 days; P < 0.001). Using serial HPV type-specific viral load measurements we could for the first time identify regressing CIN2 and CIN3 lesions. Evolution of the viral load is an objective measurable indicator of the natural history of HPV infections and could be used for future triage in HPV-based cervical screening programs

  2. Microfluidics for investigating vaso-occlusions in sickle cell disease.

    Horton, Renita E

    2017-07-01

    SCD stems from amutation in the beta globin gene. Upon deoxygenation, hemoglobin polymerizes and triggers RBC remodeling. This phenomenon is central to SCD pathogenesis as individuals suffering from the disease are plagued by painful vaso-occlusive crises episodes. These episodes are the result of a combination of processes including inflammation, thrombosis, and blood cell adhesion to the vascular wall which leads to blockages within the vasculature termed vaso-occlusions. Vaso-occlusive episodes deprive tissues of oxygen and are a major contributor to SCD-related complications; unfortunately, the complex mechanisms that contribute to vaso-occlusions are not well understood. Vaso-occlusions can occur in post-capillary venules; hence, the microvasculature is a prime target for SCD therapies. Traditional in vitro systems poorly recapitulate architectural and dynamic flow properties of in vivo systems. However, microfluidic devices can capture features of the native vasculature such as cellular composition, flow, geometry, and ECM presentation. This review, although not comprehensive, highlights microfluidic approaches that aim to improve our current understanding of the pathophysiological mechanisms surrounding SCD. Microfluidic platforms can aid in identifying factors that may contribute to disease severity and can serve as suitable test beds for novel treatment strategies which may improve patient outcomes. © 2017 John Wiley & Sons Ltd.

  3. Experimental Investigation on an Absorption Refrigerator Driven by Solar Cells

    Zi-Jie Chien

    2013-01-01

    Full Text Available This experiment is to study an absorption refrigerator driven by solar cells. Hand-held or carried in vehicle can be powered by solar energy in places without power. In the evenings or rainy days, it is powered by storage battery, and it can be directly powered by alternating current (AC power supply if available, and the storage battery can be charged full as a backup supply. The proposed system was tested by the alternation of solar irradiance 550 to 700 W/m2 as solar energy and 500ml ambient temperature water as cooling load. After 160 minutes, the proposal refrigerator can maintain the temperature at 5–8°C, and the coefficient of performance (COP of NH3-H2O absorption refrigeration system is about 0.25. Therefore, this system can be expected to be used in remote areas for refrigeration of food and beverages in outdoor activities in remote and desert areas or long-distance road transportation of food or low temperature refrigeration of vaccine to avoid the deterioration of the food or the vaccines.

  4. Assessment of centrifugation using for accelerated immunological microarray analysis for blood cells investigation

    A. V. Shishkin

    2011-01-01

    Full Text Available Phase of incubation microarray with cell suspension is prolonged when cells are investigated. It takes from 20 to 60 min if cell sedimentation on the surface of microarray is the result of gravity . Decrease of this stage duration is possible due to centrifugation. In th is article influence of centrifugation on results of analysis is considered. Changes of morphological description of cells are estimated when they a re precipitatedwith different acceleration. Also availability of centrifugation using when it is necessary to obtain the high density of cell binding in test regions of microarray if cells concentration in sample is small is demonstrated.

  5. Micro solid-phase radioimmunoassay for detection of herpesvirus type-specific antibody: parameters involved in standardization

    Matson, D.O.; Adler-Storthz, K.; Adam, E.; Dreesman, G.R. (Baylor Univ., Houston, TX (USA). Coll. of Medicine)

    1983-02-01

    A micro solid-phase radioimmunoassay (micro-SPRIA) was developed to demonstrate type-specific antibodies to herpes simplex virus types 1 and 2 (HSV1 and HSV2). Glycoproteins from the 123,000 dalton region of HSV1 (VP123) and the 119,000 dalton region of HSV2 (VP119) were isolated on preparative polyacrylamide gels for use as antigens in the micro-SPRIA. Human sera selected from clinical samples by virological history and appropriate microneutralization data were used to standardize the micro-SPRIA. Optimization of the assay required the use of siliconized microtiter wells for adsorption of antigen. Maximized results were highly dependent on the concentrations of antigen, primary antibody, and secondary antibody as well as the diluents used for these principal test reagents. Incorporation of HSV glycoproteins of each respective type with the optimal condition established in this study facilitates the direct detection of type-specific antibody in human sera.

  6. Herpes simplex virus latency-associated transcript sequence downstream of the promoter influences type-specific reactivation and viral neurotropism.

    Bertke, Andrea S; Patel, Amita; Krause, Philip R

    2007-06-01

    Herpes simplex virus (HSV) establishes latency in sensory nerve ganglia during acute infection and may later periodically reactivate to cause recurrent disease. HSV type 1 (HSV-1) reactivates more efficiently than HSV-2 from trigeminal ganglia while HSV-2 reactivates more efficiently than HSV-1 from lumbosacral dorsal root ganglia (DRG) to cause recurrent orofacial and genital herpes, respectively. In a previous study, a chimeric HSV-2 that expressed the latency-associated transcript (LAT) from HSV-1 reactivated similarly to wild-type HSV-1, suggesting that the LAT influences the type-specific reactivation phenotype of HSV-2. To further define the LAT region essential for type-specific reactivation, we constructed additional chimeric HSV-2 viruses by replacing the HSV-2 LAT promoter (HSV2-LAT-P1) or 2.5 kb of the HSV-2 LAT sequence (HSV2-LAT-S1) with the corresponding regions from HSV-1. HSV2-LAT-S1 was impaired for reactivation in the guinea pig genital model, while its rescuant and HSV2-LAT-P1 reactivated with a wild-type HSV-2 phenotype. Moreover, recurrences of HSV-2-LAT-S1 were frequently fatal, in contrast to the relatively mild recurrences of the other viruses. During recurrences, HSV2-LAT-S1 DNA increased more in the sacral cord compared to its rescuant or HSV-2. Thus, the LAT sequence region, not the LAT promoter region, provides essential elements for type-specific reactivation of HSV-2 and also plays a role in viral neurotropism. HSV-1 DNA, as quantified by real-time PCR, was more abundant in the lumbar spinal cord, while HSV-2 DNA was more abundant in the sacral spinal cord, which may provide insights into the mechanism for type-specific reactivation and different patterns of central nervous system infection of HSV-1 and HSV-2.

  7. Comparison between 125IUdR and 51Cr as cell labels in investigations of tumor cell migration

    Basse, P; Hokland, P; Hokland, M

    1991-01-01

    YAC-1 tumor cells double-labeled with Na2[51Cr]O4 [51Cr] and [125I]iododeoxyuridine [125IUdR] were injected intravenously into Balb/c mice in order to investigate their migration and fate 0-4 h after the injection. Whereas the clearance of tumor cells from the lung tissue was similar as judged...

  8. Cell-type specific expression of constitutively-active Rheb promotes regeneration of bulbospinal respiratory axons following cervical SCI.

    Urban, Mark W; Ghosh, Biswarup; Strojny, Laura R; Block, Cole G; Blazejewski, Sara M; Wright, Megan C; Smith, George M; Lepore, Angelo C

    2018-05-01

    Damage to respiratory neural circuitry and consequent loss of diaphragm function is a major cause of morbidity and mortality in individuals suffering from traumatic cervical spinal cord injury (SCI). Repair of CNS axons after SCI remains a therapeutic challenge, despite current efforts. SCI disrupts inspiratory signals originating in the rostral ventral respiratory group (rVRG) of the medulla from their phrenic motor neuron (PhMN) targets, resulting in loss of diaphragm function. Using a rat model of cervical hemisection SCI, we aimed to restore rVRG-PhMN-diaphragm circuitry by stimulating regeneration of injured rVRG axons via targeted induction of Rheb (ras homolog enriched in brain), a signaling molecule that regulates neuronal-intrinsic axon growth potential. Following C2 hemisection, we performed intra-rVRG injection of an adeno-associated virus serotype-2 (AAV2) vector that drives expression of a constitutively-active form of Rheb (cRheb). rVRG neuron-specific cRheb expression robustly increased mTOR pathway activity within the transduced rVRG neuron population ipsilateral to the hemisection, as assessed by levels of phosphorylated ribosomal S6 kinase. By co-injecting our novel AAV2-mCherry/WGA anterograde/trans-synaptic axonal tracer into rVRG, we found that cRheb expression promoted regeneration of injured rVRG axons into the lesion site, while we observed no rVRG axon regrowth with AAV2-GFP control. AAV2-cRheb also significantly reduced rVRG axonal dieback within the intact spinal cord rostral to the lesion. However, cRheb expression did not promote any recovery of ipsilateral hemi-diaphragm function, as assessed by inspiratory electromyography (EMG) burst amplitudes. This lack of functional recovery was likely because regrowing rVRG fibers did not extend back into the caudal spinal cord to synaptically reinnervate PhMNs that we retrogradely-labeled with cholera toxin B from the ipsilateral hemi-diaphragm. Our findings demonstrate that enhancing neuronal-intrinsic axon growth capacity can promote regeneration of injured bulbospinal respiratory axons after SCI, but this strategy may need to be combined with other manipulations to achieve reconnection of damaged neural circuitry and ultimately recovery of diaphragm function. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. Cell type-specific modulation of lipid mediator's formation in murine adipose tissue by omega-3 fatty acids

    Kuda, Ondřej; Rombaldová, Martina; Janovská, Petra; Flachs, Pavel; Kopecký, Jan

    2016-01-01

    Roč. 469, č. 3 (2016), s. 731-736 ISSN 0006-291X R&D Projects: GA ČR(CZ) GP13-04449P; GA ČR(CZ) GA13-00871S; GA MŠk(CZ) LH14040 Institutional support: RVO:67985823 Keywords : adipose tissue macrophages * omega-3 PUFA * protectin D1 * lipid mediators * lipidomics Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.466, year: 2016

  10. CTCF Mediates the Cell-Type Specific Spatial Organization of the Kcnq5 Locus and the Local Gene Regulation

    Ren, Licheng; Wang, Yang; Shi, Minglei; Wang, Xiaoning; Yang, Zhong; Zhao, Zhihu

    2012-01-01

    Chromatin loops play important roles in the dynamic spatial organization of genes in the nucleus. Growing evidence has revealed that the multivalent functional zinc finger protein CCCTC-binding factor (CTCF) is a master regulator of genome spatial organization, and mediates the ubiquitous chromatin loops within the genome. Using circular chromosome conformation capture (4C) methodology, we discovered that CTCF may be a master organizer in mediating the spatial organization of the kcnq5 gene l...

  11. Cell type-specific sorting of neuropeptides : a mechanism to modulate peptide composition of large dense core vesicles

    Klumperman, J.; Spijker, S.; Minnen, J. van; Sharp-Baker, H.; Smit, A.B.; Geraerts, W.P.M.

    1996-01-01

    The CNS of Lymnaea stagnalis contains two populations of egg-laying hormone (ELH)-producing neurons that differ in size and topology. In type I neurons, all peptides located C-terminally from the cleavage site Arg-Ser-Arg-Arg180-183 are sorted into secretory large dense-core vesicles (LDCV), whereas

  12. Synthesis of heparan sulfate with cyclophilin B-binding properties is determined by cell type-specific expression of sulfotransferases.

    Deligny, A.; Denys, A.; Marcant, A.; Melchior, A.; Mazurier, J.; Kuppevelt, A.H.M.S.M. van; Allain, F.

    2010-01-01

    Cyclophilin B (CyPB) induces migration and adhesion of T lymphocytes via a mechanism that requires interaction with 3-O-sulfated heparan sulfate (HS). HS biosynthesis is a complex process with many sulfotransferases involved. N-Deacetylases/N-sulfotransferases are responsible for N-sulfation, which

  13. Cortical deficits of glutamic acid decarboxylase 67 expression in schizophrenia: clinical, protein, and cell type-specific features.

    Curley, Allison A; Arion, Dominique; Volk, David W; Asafu-Adjei, Josephine K; Sampson, Allan R; Fish, Kenneth N; Lewis, David A

    2011-09-01

    Cognitive deficits in schizophrenia are associated with altered activity of the dorsolateral prefrontal cortex, which has been attributed to lower expression of the 67 kDa isoform of glutamic acid decarboxylase (GAD67), the major γ-aminobutyric acid (GABA)-synthesizing enzyme. However, little is known about the relationship of prefrontal GAD67 mRNA levels and illness severity, translation of the transcript into protein, and protein levels in axon terminals, the key site of GABA production and function. Quantitative polymerase chain reaction was used to measure GAD67 mRNA levels in postmortem specimens of dorsolateral prefrontal cortex from subjects with schizophrenia and matched comparison subjects with no known history of psychiatric or neurological disorders (N=42 pairs). In a subset of this cohort in which potential confounds of protein measures were controlled (N=19 pairs), Western blotting was used to quantify tissue levels of GAD67 protein in tissue. In five of these pairs, multilabel confocal immunofluorescence was used to quantify GAD67 protein levels in the axon terminals of parvalbumin-containing GABA neurons, which are known to have low levels of GAD67 mRNA in schizophrenia. GAD67 mRNA levels were significantly lower in schizophrenia subjects (by 15%), but transcript levels were not associated with predictors or measures of illness severity or chronicity. In schizophrenia subjects, GAD67 protein levels were significantly lower in total gray matter (by 10%) and in parvalbumin axon terminals (by 49%). The findings that lower GAD67 mRNA expression is common in schizophrenia, that it is not a consequence of having the illness, and that it leads to less translation of the protein, especially in the axon terminals of parvalbumin-containing neurons, support the hypothesis that lower GABA synthesis in parvalbumin neurons contributes to dorsolateral prefrontal cortex dysfunction and impaired cognition in schizophrenia.

  14. Cell and Receptor Type-Specific Alterations in Markers of GABA Neurotransmission in the Prefrontal Cortex of Subjects with Schizophrenia

    Lewis, David A.; Hashimoto, Takanori; Morris, Harvey M.

    2008-01-01

    Impairments in cognitive control, such as those involved in working memory, are associated with dysfunction of the dorsolateral prefrontal cortex (DLPFC) in individuals with schizophrenia. This dysfunction appears to result, at least in part, from abnormalities in GABA-mediated neurotransmission. In this paper, we review recent findings indicating that the altered DLPFC circuitry in subjects with schizophrenia reflects changes in the expression of genes that encode selective presynaptic and p...

  15. Spiking in primary somatosensory cortex during natural whisking in awake head-restrained rats is cell-type specific

    de Kock, C.P.J.; Sakmann, B.

    2009-01-01

    Sensation involves active movement of sensory organs, but it remains unknown how position or movement of sensory organs is encoded in cortex. In the rat whisker system, each whisker is represented by an individual cortical (barrel) column. Here, we quantified in awake, head-fixed rats the impact of

  16. Investigation of the bystander effect in MRC5 cells after acute and fractionated irradiation in vitro

    Shokouhozaman Soleymanifard

    2014-01-01

    Full Text Available Radiation-induced bystander effect (RIBE has been defined as radiation responses observed in nonirradiated cells. It has been the focus of investigators worldwide due to the deleterious effects it induces in nonirradiated cells. The present study was performed to investigate whether acute or fractionated irradiation will evoke a differential bystander response in MRC5 cells. A normal human cell line (MRC5, and a human lung tumor cell line (QU-DB were exposed to 0, 1, 2, and 4Gy of single acute or fractionated irradiation of equal fractions with a gap of 6 h. The MRC5 cells were supplemented with the media of irradiated cells and their micronucleus frequency was determined. The micronucleus frequency after single and fractionated irradiation did not vary significantly in the MRC5 cells conditioned with autologous or QU-DB cell-irradiated media, except for 4Gy where the frequency of micronucleated cells was lower in those MRC5 cells cultured in the media of QU-DB-exposed with a single dose of 4Gy. Our study demonstrates that the radiation-induced bystander effect was almost similar after single acute and fractionated exposure in MRC5 cells.

  17. Investigation of the bystander effect in MRC5 cells after acute and fractionated irradiation in vitro

    Soleymanifard, Shokouhozaman; Toossi, Mohammad Taghi Bahreyni; Samani, Roghayeh Kamran; Mohebbi, Shokoufeh

    2014-01-01

    Radiation-induced bystander effect (RIBE) has been defined as radiation responses observed in nonirradiated cells. It has been the focus of investigators worldwide due to the deleterious effects it induces in nonirradiated cells. The present study was performed to investigate whether acute or fractionated irradiation will evoke a differential bystander response in MRC5 cells. A normal human cell line (MRC5), and a human lung tumor cell line (QU-DB) were exposed to 0, 1, 2, and 4Gy of single acute or fractionated irradiation of equal fractions with a gap of 6 h. The MRC5 cells were supplemented with the media of irradiated cells and their micronucleus frequency was determined. The micronucleus frequency after single and fractionated irradiation did not vary significantly in the MRC5 cells conditioned with autologous or QU-DB cell-irradiated media, except for 4Gy where the frequency of micronucleated cells was lower in those MRC5 cells cultured in the media of QU-DB-exposed with a single dose of 4Gy. Our study demonstrates that the radiation-induced bystander effect was almost similar after single acute and fractionated exposure in MRC5 cells. (author)

  18. Analytical investigation on cell temperature control method of planar solid oxide fuel cell

    Inui, Y.; Ito, N.; Nakajima, T.; Urata, A. [Department of Electrical and Electronic Engineering, Toyohashi University of Technology, Tempaku-cho, Toyohashi (Japan)

    2006-09-15

    The solid oxide fuel cell (SOFC) has a problem in durability of the ceramics used as its cell materials because its operating temperature is very high and the cell temperature fluctuation induces thermal stress in the ceramics. The cell temperature distribution in the SOFC, therefore, should be kept as constant as possible during variable load operation through control of the average current density in the cell. Considering this fact, the authors numerically optimize the operating parameters of air utilization and the inlet gas temperature of the planar SOFC by minimizing the cell temperature shift from its nominal value and propose a new cell temperature control method that adopts these optimum operating parameters for each average current density. The effectiveness of the proposed method is very high and the temperature variation is suppressed to a very low level without lowering the single cell voltage for both the co-flow and counter-flow type cells, indicating that the proposed cell temperature control method makes variable load operation of the planar SOFC possible. (author)

  19. Self-organising aggregates of zebrafish retinal cells for investigating mechanisms of neural lamination.

    Eldred, Megan K; Charlton-Perkins, Mark; Muresan, Leila; Harris, William A

    2017-03-15

    To investigate the cell-cell interactions necessary for the formation of retinal layers, we cultured dissociated zebrafish retinal progenitors in agarose microwells. Within these wells, the cells re-aggregated within hours, forming tight retinal organoids. Using a Spectrum of Fates zebrafish line, in which all different types of retinal neurons show distinct fluorescent spectra, we found that by 48 h in culture, the retinal organoids acquire a distinct spatial organisation, i.e. they became coarsely but clearly laminated. Retinal pigment epithelium cells were in the centre, photoreceptors and bipolar cells were next most central and amacrine cells and retinal ganglion cells were on the outside. Image analysis allowed us to derive quantitative measures of lamination, which we then used to find that Müller glia, but not RPE cells, are essential for this process. © 2017. Published by The Company of Biologists Ltd.

  20. Identification and Investigation of Native Chromosomal Fragile Sites in the Avian Cell Line DT40

    Pentzold, Constanze

    cell systems. With the identification and investigation of CFSs in avian DT40 cells, this study reveals the genome-­‐wide evolutionary conservation of CFSs beyond the mammalian lineage for the first time. It opens the way for speculations on the beneficial existence of CFSs throughout the animal...... kingdom....

  1. Investigation of InGaN/Si double junction tandem solar cells | Bouzid ...

    In this work, the solar power conversion efficiency of InGaN/Si double junction tandem solar cells was investigated under 1-sun AM1.5 illumination, using realistic material parameters. With this intention, the current-voltage curves are calculated for different front recombination velocities and the influence of the bottom cell ...

  2. Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

    Karimpoor, Mahroo; Yebra-Fernandez, Eva; Parhizkar, Maryam; Orlu, Mine; Craig, Duncan; Khorashad, Jamshid S; Edirisinghe, Mohan

    2018-04-01

    The development of assays for evaluating the sensitivity of leukaemia cells to anti-cancer agents is becoming an important aspect of personalized medicine. Conventional cell cultures lack the three-dimensional (3D) structure of the bone marrow (BM), the extracellular matrix and stromal components which are crucial for the growth and survival of leukaemia stem cells. To accurately predict the sensitivity of the leukaemia cells in an in vitro assay a culturing system containing the essential components of BM is required. In this study, we developed a porous calcium alginate foam-based scaffold to be used for 3D culture. The new 3D culture was shown to be cell compatible as it supported the proliferation of both normal haematopoietic and leukaemia cells. Our cell differential assay for myeloid markers showed that the porous foam-based 3D culture enhanced myeloid differentiation in both leukaemia and normal haematopoietic cells compared to two-dimensional culture. The foam-based scaffold reduced the sensitivity of the leukaemia cells to the tested antileukaemia agents in K562 and HL60 leukaemia cell line model and also primary myeloid leukaemia cells. This observation supports the application of calcium alginate foams as scaffold components of the 3D cultures for investigation of sensitivity to antileukaemia agents in primary myeloid cells. © 2018 The Author(s).

  3. Investigation of reliability attributes and accelerated stress factors on terrestrial solar cells

    Lathrop, J. W.; Prince, J. L.

    1980-04-01

    Three tasks were undertaken to investigate reliability attributes of terrestrial solar cells: (1) a study of the electrical behavior of cells in the second (reverse) quadrant; (2) the accelerated stress testing of three new state-of-the-art cells; and (3) the continued bias-temperature testing of four block 2 type silicon cells at 78 C and 135 C. Electrical characteristics measured in the second quadrant were determined to be a function of the cell's thermal behavior with breakdown depending on the initiation of localized heating. This implied that high breakdown cells may be more fault tolerant when forced to operate in the second quadrant, a result contrary to conventional thinking. The accelerated stress tests used in the first (power) quadrant were bias-temperature, bias-temperature-humidity, temperature-humidity, thermal shock, and thermal cycle. The new type cells measured included an EFG cell, a polycrystalline cell, and a Czochralski cell. Significant differences in the response to the various tests were observed between cell types. A microprocessed controlled, short interval solar cell tester was designed and construction initiated on a prototype.

  4. Murine transgenic embryonic stem cell lines for the investigation of sinoatrial node-related molecular pathways

    Stefanie Schmitteckert

    2017-12-01

    Full Text Available The elucidation of molecular mechanisms that restrict the potential of pluripotent stem cells and promote cardiac lineage differentiation is of crucial relevance, since embryonic stem cells (ESCs hold great potential for cell based heart therapies. The homeodomain transcription factor Shox2 is essential for the development and proper function of the native cardiac pacemaker, the sinoatrial node. This prompted us to develop a cardiac differentiation model using ESC lines isolated from blastocysts of Shox2-deficient mice. The established cell model provides a fundamental basis for the investigation of molecular pathways under physiological and pathophysiological conditions for evaluating novel therapeutic approaches.

  5. Prevalence of type-specific HPV infection by age and grade of cervical cytology: data from the ARTISTIC trial

    Sargent, A; Bailey, A; Almonte, M; Turner, A; Thomson, C; Peto, J; Desai, M; Mather, J; Moss, S; Roberts, C; Kitchener, H C

    2008-01-01

    Human papillomavirus (HPV) infection causes cervical cancer and premalignant dysplasia. Type-specific HPV prevalence data provide a basis for assessing the impact of HPV vaccination programmes on cervical cytology. We report high-risk HPV (HR-HPV) type-specific prevalence data in relation to cervical cytology for 24 510 women (age range: 20–64; mean age 40.2 years) recruited into the ARTISTIC trial, which is being conducted within the routine NHS Cervical Screening Programme in Greater Manchester. The most common HR-HPV types were HPV16, 18, 31, 51 and 52, which accounted for 60% of all HR-HPV types detected. There was a marked decline in the prevalence of HR-HPV infection with age, but the proportion due to each HPV type did not vary greatly with age. Multiple infections were common below the age of 30 years but less so between age 30 and 64 years. Catch-up vaccination of this sexually active cohort would be expected to reduce the number of women with moderate or worse cytology by 45%, but the number with borderline or mild cytology would fall by only 7%, giving an overall reduction of 12% in the number of women with abnormal cytology and 27% in the number with any HR-HPV infection. In the absence of broader cross-protection, the large majority of low-grade and many high-grade abnormalities may still occur in sexually active vaccinated women. PMID:18392052

  6. Atomic force microscopy as a tool for the investigation of living cells.

    Morkvėnaitė-Vilkončienė, Inga; Ramanavičienė, Almira; Ramanavičius, Arūnas

    2013-01-01

    Atomic force microscopy is a valuable and useful tool for the imaging and investigation of living cells in their natural environment at high resolution. Procedures applied to living cell preparation before measurements should be adapted individually for different kinds of cells and for the desired measurement technique. Different ways of cell immobilization, such as chemical fixation on the surface, entrapment in the pores of a membrane, or growing them directly on glass cover slips or on plastic substrates, result in the distortion or appearance of artifacts in atomic force microscopy images. Cell fixation allows the multiple use of samples and storage for a prolonged period; it also increases the resolution of imaging. Different atomic force microscopy modes are used for the imaging and analysis of living cells. The contact mode is the best for cell imaging because of high resolution, but it is usually based on the following: (i) image formation at low interaction force, (ii) low scanning speed, and (iii) usage of "soft," low resolution cantilevers. The tapping mode allows a cell to behave like a very solid material, and destructive shear forces are minimized, but imaging in liquid is difficult. The force spectroscopy mode is used for measuring the mechanical properties of cells; however, obtained results strongly depend on the cell fixation method. In this paper, the application of 3 atomic force microscopy modes including (i) contact, (ii) tapping, and (iii) force spectroscopy for the investigation of cells is described. The possibilities of cell preparation for the measurements, imaging, and determination of mechanical properties of cells are provided. The applicability of atomic force microscopy to diagnostics and other biomedical purposes is discussed.

  7. To investigate the necessity of STRA6 upregulation in T cells during T cell immune responses.

    Rafik Terra

    Full Text Available Our earlier study revealed that STRA6 (stimulated by retinoic acid gene 6 was up-regulated within 3 h of TCR stimulation. STRA6 is the high-affinity receptor for plasma retinol-binding protein (RBP and mediates cellular vitamin A uptake. We generated STRA6 knockout (KO mice to assess whether such up-regulation was critical for T-cell activation, differentiation and function. STRA6 KO mice under vitamin A sufficient conditions were fertile without apparent anomalies upon visual inspection. The size, cellularity and lymphocyte subpopulations of STRA6 KO thymus and spleen were comparable to those of their wild type (WT controls. KO and WT T cells were similar in terms of TCR-stimulated proliferation in vitro and homeostatic expansion in vivo. Naive KO CD4 cells differentiated in vitro into Th1, Th2, Th17 as well as regulatory T cells in an analogous manner as their WT counterparts. In vivo experiments revealed that anti-viral immune responses to lymphocytic choriomeningitis virus in KO mice were comparable to those of WT controls. We also demonstrated that STRA6 KO and WT mice had similar glucose tolerance. Total vitamin A levels are dramatically lower in the eyes of KO mice as compared to those of WT mice, but the levels in other organs were not significantly affected after STRA6 deletion under vitamin A sufficient conditions, indicating that the eye is the mouse organ most sensitive to the loss of STRA6. Our results demonstrate that 1 in vitamin A sufficiency, the deletion of STRA6 in T cells does no affect the T-cell immune responses so-far tested, including those depend on STAT5 signaling; 2 STRA6-independent vitamin A uptake compensated the lack of STRA6 in lymphoid organs under vitamin A sufficient conditions in mice; 3 STRA6 is critical for vitamin A uptake in the eyes even in vitamin A sufficiency.

  8. Investigation of Plant Cell Wall Properties: A Study of Contributions from the Nanoscale to the Macroscale Impacting Cell Wall Recalcitrance

    Crowe, Jacob Dillon

    Biochemical conversion of lignocellulosic biomass to fuel ethanol is one of a few challenging, yet opportune technologies that can reduce the consumption of petroleum-derived transportation fuels, while providing parallel reductions in greenhouse gas emissions. Biomass recalcitrance, or resistance to deconstruction, is a major technical challenge that limits effective conversion of biomass to fermentable sugars, often requiring a costly thermochemical pretreatment step to improve biomass deconstruction. Biomass recalcitrance is imparted largely by the secondary cell wall, a complex polymeric matrix of cell wall polysaccharides and aromatic heteropolymers, that provides structural stability to cells and enables plant upright growth. Polymers within the cell wall can vary both compositionally and structurally depending upon plant species and anatomical fraction, and have varied responses to thermochemical pretreatments. Cell wall properties impacting recalcitrance are still not well understood, and as a result, the goal of this dissertation is to investigate structural features of the cell wall contributing to recalcitrance (1) in diverse anatomical fractions of a single species, (2) in response to diverse pretreatments, and (3) resulting from genetic modification. In the first study, feedstock cell wall heterogeneity was investigated in anatomical (stem, leaf sheaths, and leaf blades) and internode fractions of switchgrass at varying tissue maturities. Lignin content was observed as the key contributor to recalcitrance in maturing stem tissues only, with non-cellulosic substituted glucuronoarabinoxylans and pectic polysaccharides contributing to cell wall recalcitrance in leaf sheath and leaf blades. Hydroxycinnamate (i.e., saponifiable p-coumarate and ferulate) content along with xylan and pectin extractability decreased with tissue maturity, suggesting lignification is only one component imparting maturity specific cell wall recalcitrance. In the second study

  9. [Investigation of neural stem cell-derived donor contribution in the inner ear following blastocyst injection].

    Volkenstein, S; Brors, D; Hansen, S; Mlynski, R; Dinger, T C; Müller, A M; Dazert, S

    2008-03-01

    Utilising the enormous proliferation and multi-lineage differentiation potentials of somatic stem cells represents a possible therapeutical strategy for diseases of non-regenerative tissues like the inner ear. In the current study, the possibility of murine neural stem cells to contribute to the developing inner ear following blastocyst injection was investigated. Fetal brain-derived neural stem cells from the embryonic day 14 cortex of male mice were isolated and expanded for four weeks in neurobasal media supplemented with bFGF and EGF. Neural stem cells of male animals were harvested, injected into blastocysts and the blastocysts were transferred into pseudo-pregnant foster animals. Each blastocyst was injected with 5-15 microspheres growing from single cell suspension from neurospheres dissociated the day before. The resulting mice were investigated six months POST PARTUM for the presence of donor cells. Brainstem evoked response audiometry (BERA) was performed in six animals. To visualize donor cells Lac-Z staining was performed on sliced cochleas of two animals. In addition, the cochleas of four female animals were isolated and genomic DNA of the entire cochlea was analyzed for donor contribution by Y-chromosome-specific PCR. All animals had normal thresholds in brainstem evoked response audiometry. The male-specific PCR product indicating the presence of male donor cells were detected in the cochleas of three of the four female animals investigated. In two animals, male donor cells were detected unilateral, in one animal bilateral. The results suggest that descendants of neural stem cells are detectable in the inner ear after injection into blastocysts and possess the ability to integrate into the developing inner ear without obvious loss in hearing function.

  10. Extensions of the Rosner-Colditz breast cancer prediction model to include older women and type-specific predicted risk.

    Glynn, Robert J; Colditz, Graham A; Tamimi, Rulla M; Chen, Wendy Y; Hankinson, Susan E; Willett, Walter W; Rosner, Bernard

    2017-08-01

    A breast cancer risk prediction rule previously developed by Rosner and Colditz has reasonable predictive ability. We developed a re-fitted version of this model, based on more than twice as many cases now including women up to age 85, and further extended it to a model that distinguished risk factor prediction of tumors with different estrogen/progesterone receptor status. We compared the calibration and discriminatory ability of the original, the re-fitted, and the type-specific models. Evaluation used data from the Nurses' Health Study during the period 1980-2008, when 4384 incident invasive breast cancers occurred over 1.5 million person-years. Model development used two-thirds of study subjects and validation used one-third. Predicted risks in the validation sample from the original and re-fitted models were highly correlated (ρ = 0.93), but several parameters, notably those related to use of menopausal hormone therapy and age, had different estimates. The re-fitted model was well-calibrated and had an overall C-statistic of 0.65. The extended, type-specific model identified several risk factors with varying associations with occurrence of tumors of different receptor status. However, this extended model relative to the prediction of any breast cancer did not meaningfully reclassify women who developed breast cancer to higher risk categories, nor women remaining cancer free to lower risk categories. The re-fitted Rosner-Colditz model has applicability to risk prediction in women up to age 85, and its discrimination is not improved by consideration of varying associations across tumor subtypes.

  11. Investigation of water distribution in proton exchange membrane fuel cells via Terahertz imaging

    Thamboon, P.; Buaphad, P.; Thongbai, C.; Saisud, J.; Kusoljariyakul, K.; Rhodes, M.W.; Vilaithong, T.

    2011-01-01

    Coherent transition radiation in a THz regime generated from a femtosecond electron bunch is explored for its potential use in imaging applications. Due to water sensitivity, the THz imaging experiment is performed on a proton exchange membrane fuel cell (PEMFC) to assess the ability to quantify water in the flow field of the cell. In this investigation, the PEMFC design and the experimental setup for the THz imaging is described. The results of the THz images in the flow field are also discussed.

  12. Biomaterials and computation: a strategic alliance to investigate emergent responses of neural cells.

    Sergi, Pier Nicola; Cavalcanti-Adam, Elisabetta Ada

    2017-03-28

    Topographical and chemical cues drive migration, outgrowth and regeneration of neurons in different and crucial biological conditions. In the natural extracellular matrix, their influences are so closely coupled that they result in complex cellular responses. As a consequence, engineered biomaterials are widely used to simplify in vitro conditions, disentangling intricate in vivo behaviours, and narrowing the investigation on particular emergent responses. Nevertheless, how topographical and chemical cues affect the emergent response of neural cells is still unclear, thus in silico models are used as additional tools to reproduce and investigate the interactions between cells and engineered biomaterials. This work aims at presenting the synergistic use of biomaterials-based experiments and computation as a strategic way to promote the discovering of complex neural responses as well as to allow the interactions between cells and biomaterials to be quantitatively investigated, fostering a rational design of experiments.

  13. Investigating the role of caveolin-2 in prostate cancer cell line

    Jin-Yih Low

    2017-02-01

    Full Text Available Prostate cancer is a worldwide problem. While the role of caveolin-1 has been extensively studied, little is known about the role of caveolin-2 (CAV2 in prostate cancer. Up-regulation of CAV2 in androgen independent PC3 cells compared to normal prostate cell line and androgen dependent prostate cancer cell lines has been observed. Recent studies suggest that up-regulation of CAV2 plays an important role in androgen independent prostate cancer. This study investigates whether CAV2 is important in mediating the aggressive phenotypes seen in androgen independent prostate cancer cells. The androgen independent prostate cancer cell line, PC3 was used that has been shown to express CAV2, and CAV2 knock down was performed using siRNA system. Changes to cell number, migration and invasion were assessed after knocking down CAV2. Our results showed that down-regulating CAV2 resulted in reduced cell numbers, migration and invasion in PC3 cells. This preliminary study suggests that CAV2 may act to promote malignant behavior in an androgen independent prostate cancer cell line. Further studies are required to fully elucidate the role of CAV2 in androgen independent prostate cancer.

  14. Opto-electronic analysis of silicon solar cells by LBIC investigations and current-voltage characterization

    Thantsha, N.M.; Macabebe, E.Q.B.; Vorster, F.J.; Dyk, E.E. van

    2009-01-01

    A different laser beam induced current (LBIC) mapping technique has been used for the measurements of spatial variation of light generated current of a solar cell. These variations are caused by parasitic resistances and defects at grain boundaries (GBs) in multicrystalline silicon solar cells (mc-Si). This study investigates and identifies the regions within mc-Si solar cells where dominating recombination and lifetime limiting processes occur. A description of the LBIC technique is presented and the results show how multicrystalline GBs and other defects affect the light generated current of a spot illuminated mc-Si solar cell. The results of the internal quantum efficiency (IQE) at wavelength of 660 nm revealed that some regions in mc-Si solar cell give rise to paths that lead current away from the intended load.

  15. Opto-electronic analysis of silicon solar cells by LBIC investigations and current-voltage characterization

    Thantsha, N.M.; Macabebe, E.Q.B.; Vorster, F.J. [Department of Physics, PO Box 77000, Nelson Mandela Metropolitan University, Port Elizabeth 6031 (South Africa); Dyk, E.E. van, E-mail: ernest.vandyk@nmmu.ac.z [Department of Physics, PO Box 77000, Nelson Mandela Metropolitan University, Port Elizabeth 6031 (South Africa)

    2009-12-01

    A different laser beam induced current (LBIC) mapping technique has been used for the measurements of spatial variation of light generated current of a solar cell. These variations are caused by parasitic resistances and defects at grain boundaries (GBs) in multicrystalline silicon solar cells (mc-Si). This study investigates and identifies the regions within mc-Si solar cells where dominating recombination and lifetime limiting processes occur. A description of the LBIC technique is presented and the results show how multicrystalline GBs and other defects affect the light generated current of a spot illuminated mc-Si solar cell. The results of the internal quantum efficiency (IQE) at wavelength of 660 nm revealed that some regions in mc-Si solar cell give rise to paths that lead current away from the intended load.

  16. An investigation of cell growth and detachment from thermoresponsive physically crosslinked networks.

    Healy, Deirdre; Nash, Maria; Gorleov, Alexander; Thompson, Kerry; Dockery, Peter; Rochev, Yury

    2017-11-01

    The primary aim of this investigation was to determine the biocompatibility and cell culture potential of a newly designed class of thermoresponsive polymers. The attractiveness of these polymers lies in the fact that they swell rather than dissolve when the temperature is reduced below their respective lower critical solution temperature, due to the incorporation of octadecyl methacrylate (ODMA). The ODMA monomer acts as a physical crosslinker, preventing polymer dissolution upon temperature reduction. Two polymers were studied in this investigation poly(N isorpoylacrylamide (NIPAm)(99.25%)-co-ODMA(0.75%)) and poly(NIPAm(65%)-co-N-tert-butylacrylamide (NtBAm)(34.25%)-co-ODMA(0.75%)). Thin thermoresponsive films of the polymers were prepared via spin coating. 3T3 cells were then seeded on the prepared films and cell viability was assessed quantitatively through cell viability and activity assays and qualitatively by light microscopy. Cells were successfully seeded and grown on the poly(NIPAm-co-ODMA) and poly(NIPAm-co-NtBAm-co-ODMA) copolymer films after film modification with cell adhesion promoters (CAPs). Cell sheets successfully detached from the CAP coated poly(NIPAm-co-ODMA) platforms upon temperature reduction. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Dye-sensitized solar cells: Atomic scale investigation of interface structure and dynamics

    Ma Wei; Zhang Fan; Meng Sheng

    2014-01-01

    Recent progress in dye-sensitized solar cells (DSC) research is reviewed, focusing on atomic-scale investigations of the interface electronic structures and dynamical processes, including the structure of dye adsorption onto TiO 2 , ultrafast electron injection, hot-electron injection, multiple-exciton generation, and electron—hole recombination. Advanced experimental techniques and theoretical approaches are briefly summarized, and then progressive achievements in photovoltaic device optimization based on insights from atomic scale investigations are introduced. Finally, some challenges and opportunities for further improvement of dye solar cells are presented. (invited review — international conference on nanoscience and technology, china 2013)

  18. Investigate The Role Of Cells To Cell Adhesion During Wound Healing

    Montes, D. R.; Rodriguez, A. J.; Uyoh, E.; Antwi, P.; Olusegun, S.; Murray, P.

    2017-12-01

    Improvements in wound healing can save living organism and cut misery in health care centers and on the military battlefield. Microscopic examination of how Madin-Darby Canine Kidney (MDCK) epithelial cells respond to wounds under various treatments that block general calcium absorption and e-cadherin mechanics may yield novel wound treatment insights. We used MDCK cells as our tissue model for this experiment.Each tissue was grown on mattek dishes at high, middle, and low densities. Next, we incubated the tissue at 37 degrees celsius overnight. The next day, we wounded the MDCK tissue by scratching them with a 21G1 ½(38.1 mm) syringe needle. Before the scratch we took an image of the tissues. With the tissue wound we made a time-lapse movie of about 3 hours long with a interval of 1 min. Afterward, we compared each MDCK tissue by adding a calcium chelator named EGTA. As so, this will sequester calcium use as well to block e-cadherin mechanics. We then looked at the relationship between a wounded tissue and a healthy tissue. As such, wound dynamics can be observed. We asked ourselves, will EGTA prevent wound closure?

  19. A physiologic three-dimensional cell culture system to investigate the role of decorin in matrix organisation and cell survival

    Seidler, Daniela G.; Schaefer, Liliana; Robenek, Horst; Iozzo, Renato V.; Kresse, Hans; Schoenherr, Elke

    2005-01-01

    In vivo cells exist in a three-dimensional environment generated and maintained by multiple cell-cell and cell-matrix interactions. Proteoglycans, like decorin, affect these complex interactions. Thus, we sought to investigate the role of decorin in a three-dimensional environment where the matrix was generated over time by decorin-deficient fibroblasts in the presence of L-ascorbic acid 2-phosphate. The cells were viable and proliferated in response to FGF2. Decorin was incorporated in the matrix and caused a ∼2 nm shift in the average diameter of the collagen fibrils, and the range and distribution of the fibrils became narrower and more uniform. Although there were no appreciable changes in collagen composition, we found that exogenous decorin induced the de novo synthesis of collagen I and V and cross-linked β (I). In the early phases of the three-dimensional culture, decorin reduced apoptosis. However, following the establishment of a three-dimensional matrix, the cells did not require decorin for their survival

  20. INVESTIGATION OF HYPOLIPIDEMIC EFFECT OF SESQUITERPENE Γ-LACTONE AHILLIN IN HEPATOMA TISSUE CULTURE (HTC CELLS

    V. V. Ivanov

    2014-01-01

    Full Text Available Objective. Investigation of hypolipidemic effect of sesquiterpene γ-lactone ahillin in hepatoma tissue culture (HTC cells.Material and methods. In this study we’ve evaluated the effect of γ-lactone sesquiterpene aсhillin and gemfibrozil (comparator drug on the lipid content in the hepatoma tissue culture (HTC cell which were incubated with a fat emulsion lipofundin by fluorescent method with vital dye Nile Redand staining the cells with the dye Oil Red O. The cell viability was investigated using the MTT-test and staining with Trypan blue.Results. Cultivation cells HTC with aсhillin and gemfibrozilat concentrations ranging from 0.5 to1.5 mM and from0.25 mM to0.5 mM, respectively, resulted in dose-dependent decrease of the fluorescence’s intensity Nile Red. It reflects a decrease in lipid content in the cells. At these concentrations the drugs didn’t have cytotoxic effect and the cell viability didn’t change compared to the control culture.An experimental hyperlipidemia in the hepatoma culture cells was induced by adding to the incubation medium a fat emulsion lipofundin at a final concentration 0.05%. The intensity of fluorescence Nile Red in the cells was increased 4 fold (p < 0.05. This result suggests the significant accumulation of lipids in the cell’s cytosol and confirmed by microscopy after staining neutral lipids with the dye Oil Red O. Under these conditions aсhillin and gemfibrozil reduced lipid content in cells and hadthe effect at concentrations of0.5 mM and0.25 mM respectively.Conclusion. In the lipofundin-mediated model of hyperlipidemia the sesquiterpene lactone aсhillin prevents the lipid accumulation in cells. It confirms by decrease of fluorescence Nile Red and reduction lipid drops which were stained with Oil Red O in cytosol. To establish the molecular targets of aсhillin’saction on lipid metabolism in cell culture HTC we need to investigate a gene expression of key enzymes of lipid metabolism.

  1. Investigation into local cell mechanics by atomic force microscopy mapping and optical tweezer vertical indentation.

    Coceano, G; Yousafzai, M S; Ma, W; Ndoye, F; Venturelli, L; Hussain, I; Bonin, S; Niemela, J; Scoles, G; Cojoc, D; Ferrari, E

    2016-02-12

    Investigating the mechanical properties of cells could reveal a potential source of label-free markers of cancer progression, based on measurable viscoelastic parameters. The Young's modulus has proved to be the most thoroughly studied so far, however, even for the same cell type, the elastic modulus reported in different studies spans a wide range of values, mainly due to the application of different experimental conditions. This complicates the reliable use of elasticity for the mechanical phenotyping of cells. Here we combine two complementary techniques, atomic force microscopy (AFM) and optical tweezer microscopy (OTM), providing a comprehensive mechanical comparison of three human breast cell lines: normal myoepithelial (HBL-100), luminal breast cancer (MCF-7) and basal breast cancer (MDA-MB-231) cells. The elastic modulus was measured locally by AFM and OTM on single cells, using similar indentation approaches but different measurement parameters. Peak force tapping AFM was employed at nanonewton forces and high loading rates to draw a viscoelastic map of each cell and the results indicated that the region on top of the nucleus provided the most meaningful results. OTM was employed at those locations at piconewton forces and low loading rates, to measure the elastic modulus in a real elastic regime and rule out the contribution of viscous forces typical of AFM. When measured by either AFM or OTM, the cell lines' elasticity trend was similar for the aggressive MDA-MB-231 cells, which were found to be significantly softer than the other two cell types in both measurements. However, when comparing HBL-100 and MCF-7 cells, we found significant differences only when using OTM.

  2. Investigation of biomaterials by human epithelial gingiva cells: an in vitro study

    Neunzehn Jörg

    2012-12-01

    Full Text Available Abstract Introduction In modern medicine and dentistry the use of biomaterials is a fast developing field of increasing interest. Especially in dentistry the interaction between biomaterials like implant materials and the soft tissue in the oral cavity is in the focus of daily research. In this context the high importance of testing materials and their surfaces concerning their biocompatibility towards corresponding cells is very likely. For this purpose this study investigates cells derived from human gingival biopsies on different materials and surfaces. Methods Cells in this study were cultivated out of human biopsies by a grow out explant technique and were sub cultivated on titanium, zirconium dioxide and collagen membrane specimens. To characterise the cells on the material surfaces used in this study immunohistochemical and histological staining techniques as well as different methods of microscopy (light microscopy and SEM were applied. Results With the aid of the explant technique and the chosen cell cultivation method it was possible to investigate the human gingiva derived cells on different materials. The data of the present study show that the human gingival cells attach and proliferate on all three tested materials by exhibiting characteristic gingival keratinocyte protein expression even after long periods of culture e.g. up to 70 days. Conclusions It could be shown that the three tested materials titanium, zirconium dioxide and collagen membrane (and their special surfaces are good candidates for the application as materials in the dental gingival environment or, in the case of the collagen membrane as scaffold/cell-carrier for human gingival cells in tissue engineering.

  3. The hot cell laboratories for material investigations of the Institute for Safety Research

    Viehrig, H W

    1998-10-01

    Special facilities for handling and testing of irradiated specimens are necessary, to perform the investigation of activated material. The Institute for Safety Research has two hot cell laboratories: - the preparation laboratory and - the materials testing laboratory. This report is intended to give an overview of the available facilities and developed techniques in the laboratories. (orig.)

  4. An Investigation on Changing Behaviours of University Students Switching from Using Classical Cell Phones to Smartphones

    Arslan, Yusuf

    2016-01-01

    In this study, it was tried to comprehend whether there occur any changes in behaviours of university students switching from classical cell phones to smartphones. The investigation was carried out according to quantitative research method. Questionnaire was employed as data collection tool. The datum of the study was limited with the information…

  5. Using a Cell Phone to Investigate the Skin Depth Effect in Salt Water

    Rayner, John

    2017-01-01

    This paper describes an experimental investigation of the skin depth effect for electromagnetic waves in salt water using a cell phone that is immersed to a critical depth where it no longer responds when called. We show that this critical depth is directly proportional to the theoretical skin depth for a range of salt concentrations.

  6. Investigation of Novel Electrocatalysts for Metal Supported Solid Oxide Fuel Cells - Ru:GDC

    Sudireddy, Bhaskar Reddy; Nielsen, Jimmy; Thydén, Karl Tor Sune

    2015-01-01

    The electrochemical performance and stability of the planar metal supported solid oxide fuel cells (MS-SOFC) with two different electrocatalytically active materials, namely, Ni:GDC and Ru:GDC were investigated. Ru:GDC with an ASR of 0.322 Ωcm2 performed better than Ni:GDC with an ASR of 0.453 Ωc...

  7. Wide-range stiffness gradient PVA/HA hydrogel to investigate stem cell differentiation behavior.

    Oh, Se Heang; An, Dan Bi; Kim, Tae Ho; Lee, Jin Ho

    2016-04-15

    Although stiffness-controllable substrates have been developed to investigate the effect of stiffness on cell behavior and function, the use of separate substrates with different degrees of stiffness, substrates with a narrow range stiffness gradient, toxicity of residues, different surface composition, complex fabrication procedures/devices, and low cell adhesion are still considered as hurdles of conventional techniques. In this study, a cylindrical polyvinyl alcohol (PVA)/hyaluronic acid (HA) hydrogel with a wide-range stiffness gradient (between ∼20kPa and ∼200kPa) and cell adhesiveness was prepared by a liquid nitrogen (LN2)-contacting gradual freezing-thawing method that does not use any additives or specific devices to produce the stiffness gradient hydrogel. From an in vitro cell culture using the stiffness gradient PVA/HA hydrogel, it was observed that human bone marrow mesenchymal stem cells have favorable stiffness ranges for induction of differentiation into specific cell types (∼20kPa for nerve cell, ∼40kPa for muscle cell, ∼80kPa for chondrocyte, and ∼190kPa for osteoblast). The PVA/HA hydrogel with a wide range of stiffness spectrum can be a useful tool for basic studies related with the stem cell differentiation, cell reprogramming, cell migration, and tissue regeneration in terms of substrate stiffness. It is postulated that the stiffness of the extracellular matrix influences cell behavior. To prove this concept, various techniques to prepare substrates with a stiffness gradient have been developed. However, the narrow ranges of stiffness gradient and complex fabrication procedures/devices are still remained as limitations. Herein, we develop a substrate (hydrogel) with a wide-range stiffness gradient using a gradual freezing-thawing method which does not need specific devices to produce a stiffness gradient hydrogel. From cell culture experiments using the hydrogel, it is observed that human bone marrow mesenchymal stem cells have

  8. The jet impingement cell: A valuable device for investigating CO{sub 2} corrosion

    Kinsella, Brian; John, Douglas; Bailey, Stuart; De Marco, Roland [Western Australian Corrosion Research Group, School of Applied Chemistry, Curtin University of Technology, GPO Box U1987, Perth WA, 6845 (Australia)

    2004-07-01

    The jet impingement cell (JIC) is a valuable technique for the investigation of carbon dioxide corrosion of steel and its inhibition under high flow conditions or high wall shear stress. Despite the use of the JIC in corrosion testing laboratories not a great deal has been published on the design and calibration of these cells. In the evaluation of corrosion inhibitors, the type of corrosion and relative performance of the inhibitors depends on the metallurgy of the steel used to manufacture electrodes and measure the corrosion rate. This paper covers aspects of cell design and the determination of mass transfer and wall shear stress at electrodes used in the cell. The performance of different generic type corrosion inhibitors and their affect on the type of corrosion (i.e, uniform, pitting and crevice corrosion) is shown and discussed. (authors)

  9. Dynamic modeling and experimental investigation of a high temperature PEM fuel cell stack

    Nguyen, Gia; Sahlin, Simon Lennart; Andreasen, Søren Juhl

    2016-01-01

    High temperature polymer fuel cells operating at 100 to 200◦C require simple fuel processing and produce high quality heat that can integrate well with domestic heating systems. Because the transportation of hydrogen is challenging, an alternative option is to reform natural gas on site....... This article presents the development of a dynamic model and the comparison with experimental data from a high temperature proton exchange membrane fuel cell stack operating on hydrogen with carbon monoxide concentrations up to 0.8%, and temperatures from 155 to 175◦C. The dynamic response of the fuel cell...... is investigated with simulated reformate gas. The dynamic response of the fuel cell stack was compared with a step change in current from 0.09 to 0.18 and back to 0.09 A/cm2 . This article shows that the dynamic model calculates the voltage at steady state well. The dynamic response for a change in current shows...

  10. Investigating the dynamics of a direct parallel combination of supercapacitors and polymer electrolyte fuel cells

    Papra, M.; Buechi, F.N.; Koetz, R. [Electrochemistry Laboratory, Paul Scherrer Institute, CH-5232 Villigen PSI (Switzerland)

    2010-10-15

    Hydrogen fuelled vehicles with a fuel cell based powertrain are considered to contribute to sustainable mobility by reducing CO{sub 2} emissions from road transport. In such vehicles the fuel cell system is typically hybridised with an energy storage device such as a battery or a supercapacitor (SC) to allow for recovering braking energy and assist the fuel cell system for peak power. The direct parallel combination of a polymer electrolyte fuel cell (PEFC) and a SC without any control electronics is investigated in the present study. It is demonstrated that the combination enhances the dynamics of the PEFC significantly during load changes. However, due to the lack of a power electronic interface the SC cannot be utilised to its optimum capacity. (Abstract Copyright [2010], Wiley Periodicals, Inc.)

  11. Numerical investigation of flow field configuration and contact resistance for PEM fuel cell performance

    Akbari, Mohammad Hadi; Rismanchi, Behzad [Department of Mechanical Engineering, Shiraz University, Shiraz 71348-51154 (Iran)

    2008-08-15

    A steady-state three-dimensional non-isothermal computational fluid dynamics (CFD) model of a proton exchange membrane fuel cell is presented. Conservation of mass, momentum, species, energy, and charge, as well as electrochemical kinetics are considered. In this model, the effect of interfacial contact resistance is also included. The numerical solution is based on a finite-volume method. In this study the effects of flow channel dimensions on the cell performance are investigated. Simulation results indicate that increasing the channel width will improve the limiting current density. However, it is observed that an optimum shoulder size of the flow channels exists for which the cell performance is the highest. Polarization curves are obtained for different operating conditions which, in general, compare favorably with the corresponding experimental data. Such a CFD model can be used as a tool in the development and optimization of PEM fuel cells. (author)

  12. Polarized Ends of Human Macula Densa Cells: Ultrastructural Investigation and Morphofunctional Correlations.

    Cangiotti, Angela Maria; Lorenzi, Teresa; Zingaretti, Maria Cristina; Fabri, Mara; Morroni, Manrico

    2018-05-01

    The morphology of the kidney macula densa (MD) has extensively been investigated in animals, whereas human studies are scanty. We studied the fine structure of human MD cells focusing on their apical and basal ends and correlating structure and function. The MD region was examined by transmission electron microscopy in six renal biopsies from patients with kidney disease. Ultrastructural analysis of MD cells was performed on serial sections. MD cells show two polarized ends. The apical portion is characterized by a single, immotile cilium associated with microvilli; apically, cells are joined by adhering junctions. In the basal portion, the cytoplasm contains small, dense granules and numerous, irregular cytoplasmic projections extending to the adjacent extraglomerular mesangium. The projections often contain small, dense granules. A reticulated basement membrane around MD cells separates them from the extraglomerular mesangium. Although the fact that tissue specimens came from patients with kidney disease mandates extreme caution, ultrastructural examination confirmed that MD cells have sensory features due to the presence of the primary cilium, that they are connected by apical adhering junctions forming a barrier that separates the tubular flow from the interstitium, and that they present numerous basal interdigitations surrounded by a reticulated basement membrane. Conceivably, the latter two features are related to the functional activity of the MD. The small, dense granules in the basal cytoplasm and in cytoplasmic projections are likely related to the paracrine function of MD cells. Anat Rec, 301:922-931, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  13. Numerical investigation of the effect of operating parameters on a planar solid oxide fuel cell

    Raj, Abhishek; Sasmito, Agus P.; Shamim, Tariq

    2015-01-01

    Highlights: • Effects of operating parameters on a planar type of SOFC are investigated. • The studies carried out by developing a three dimensional mathematical model. • The cell performance is enhanced at high temperatures and cathode stoichiometry. • Cathode stoichiometry has a high influence on the cell performance. • The effect of anode stoichiometry on the cell performance is low. - Abstract: The three operating parameters – temperature, stoichiometry and the degree of humidification – constitute key factors required to ensure high performance of the solid oxide fuel cell (SOFC). A careful trade-off between performance and parasitic loads is required in order to optimize the output. The present study numerically analyzes the influence of the key operating parameters on the performance of planar type of SOFC and parasitic loads utilizing a validated three dimensional mathematical model which takes into account of the conservation of mass, momentum, species and charge. The numerical results indicate that the cell performance is enhanced at high temperatures and cathode stoichiometry and it declines with increasing cathode relative humidity. Furthermore, cathode stoichiometry is found to have higher influence on the cell performance as compared to the anode stoichiometry. The gain in cell performance however, has to be balanced with the changing parasitic load requirement from pumping, humidification and heating. The results presented herein can assist in the selection of optimum or near-to-optimum operating parameters for high performance planar type SOFC

  14. Modeling T-cell proliferation: an investigation of the consequences of the Hayflick limit.

    Pilyugin, S; Mittler, J; Antia, R

    1997-05-07

    Somatic cells, including immune cells such as T-cells have a limited capacity for proliferation and can only replicate for a finite number of generations (known as the Hayflick limit) before dying. In this paper we use mathematical models to investigate the consequences of introducing a Hayflick limit on the dynamics of T-cells stimulated with specific antigen. We show that while the Hayflick limit does not alter the dynamics of T-cell response to antigen over the short term, it may have a profound effect on the long-term immune response. In particular we show that over the long term the Hayflick limit may be important in determining whether an immune response can be maintained to a persistent antigen (or parasite). The eventual outcome is determined by the magnitude of the Hayflick limit, the extent to which antigen reduces the input of T-cells from the thymus, and the rate of antigen-induced proliferation of T-cells. Counter to what might be expected we show that the persistence of an immune response (immune memory) requires the density of persistent antigen to be less than a defined threshold value. If the amount of persistent antigen (or parasite) is greater than this threshold value then immune memory will be relatively short lived. The consequences of this threshold for persistent mycobacterial and HIV infections and for the generation of vaccines are discussed.

  15. Label-free investigation of the effects of lithium niobate polarization on cell adhesion

    Mandracchia, B.; Gennari, O.; Paturzo, M.; Grilli, S.; Ferraro, P.

    2017-06-01

    The determination of contact area is pivotal to understand how biomaterials properties influence cell adhesion. In particular, the influence of surface charges is well-known but still controversial, especially when new functional materials and methods are introduced. Here, we use for the first time Holographic Total Internal Reflection Microscopy (HoloTIRM) to study the influence of the spontaneous polarization of ferroelectric lithium niobate (LN) on the adhesion properties of fibroblast cells. The selective illumination of a very thin region directly above the substrate, achieved by Total Internal Reflection, provides high-contrast images of the contact regions. Holographic recording, on the other hand, allows for label-free quantitative phase imaging of the contact areas between cells and LN. Phase signal is more sensitive in the first 100nm and, thus more reliable in order to locate focal contacts. This work shows that cells adhering on negatively polarized LN present a significant increase of the contact area in comparison with cells adhering on the positively polarized LN substrate, as well as an intensification of contact vicinity. This confirms the potential of LN as a platform for investigating the role of charges on cellular processes. The similarity of cell adhesion behavior on negatively polarized LN and glass control also confirms the possibility to use LN as an active substrate without impairing cell behavior.

  16. Investigation of Halohydrins Degradation by Whole Cells and Cell-free Extract of Pseudomonas putida DSM 437: A Kinetic Approach

    A. Konti

    2017-10-01

    Full Text Available The biodegradation of two halohydrins (1,3-dichloro-2-propanol and 3-chloro-1,2-propanediol by P. putida DSM 437 was investigated. Intact cells of previously acclimatized P. putida DSM 437 as well as cell-free extracts were used in order to study the degradation kinetics. When whole cells were used, a maximum biodegradation rate of 3-CPD (vmax = 1.28.10–5 mmol mg–1 DCW h–1 was determined, which was more than 4 times higher than that of 1,3-DCP. However, the affinity towards both halohydrins (Km was practically the same. When using cell-free extract, the apparent vmax and Km values for 1,3-DCP were estimated at 9.61.10–6 mmol mg–1 protein h–1 and 8.00 mM, respectively, while for 3-CPD the corresponding values were 2.42.10–5 mmol mg–1 protein h–1 and 9.07 mM. GC-MS analysis of cell-free extracts samples spiked with 1,3-DCP revealed the presence of 3-CPD and glycerol, intermediates of 1,3-DCP degradation pathway. 3-CPD degradation was strongly inhibited by the presence of epichlorohydrin and to a lesser extent by glycidol, intermediates of dehalogenation pathway.

  17. Human airway epithelial cells investigated by atomic force microscopy: A hint to cystic fibrosis epithelial pathology

    Lasalvia, Maria [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Castellani, Stefano [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); D’Antonio, Palma [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Perna, Giuseppe [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Carbone, Annalucia [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); Colia, Anna Laura; Maffione, Angela Bruna [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Capozzi, Vito [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Conese, Massimo, E-mail: massimo.conese@unifg.it [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy)

    2016-10-15

    The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in

  18. Human airway epithelial cells investigated by atomic force microscopy: A hint to cystic fibrosis epithelial pathology

    Lasalvia, Maria; Castellani, Stefano; D’Antonio, Palma; Perna, Giuseppe; Carbone, Annalucia; Colia, Anna Laura; Maffione, Angela Bruna; Capozzi, Vito; Conese, Massimo

    2016-01-01

    The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in the

  19. Clustered Regularly Interspaced Short Palindromic Repeats Are emm Type-Specific in Highly Prevalent Group A Streptococci.

    Zheng, Po-Xing; Chan, Yuen-Chi; Chiou, Chien-Shun; Chiang-Ni, Chuan; Wang, Shu-Ying; Tsai, Pei-Jane; Chuang, Woei-Jer; Lin, Yee-Shin; Liu, Ching-Chuan; Wu, Jiunn-Jong

    2015-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) are the bacterial adaptive immune system against foreign nucleic acids. Given the variable nature of CRISPR, it could be a good marker for molecular epidemiology. Group A streptococcus is one of the major human pathogens. It has two CRISPR loci, including CRISPR01 and CRISPR02. The aim of this study was to analyze the distribution of CRISPR-associated gene cassettes (cas) and CRISPR arrays in highly prevalent emm types. The cas cassette and CRISPR array in two CRISPR loci were analyzed in a total of 332 strains, including emm1, emm3, emm4, emm12, and emm28 strains. The CRISPR type was defined by the spacer content of each CRISPR array. All strains had at least one cas cassette or CRISPR array. More than 90% of the spacers were found in one emm type, specifically. Comparing the consistency between emm and CRISPR types by Simpson's index of diversity and the adjusted Wallace coefficient, CRISPR01 type was concordant to emm type, and CRISPR02 showed unidirectional congruence to emm type, suggesting that at least for the majority of isolates causing infection in high income countries, the emm type can be inferred from CRISPR analysis, which can further discriminate isolates sharing the same emm type.

  20. Direct methods and residue type specific isotope labeling in NMR structure determination and model-driven sequential assignment

    Schedlbauer, Andreas; Auer, Renate; Ledolter, Karin; Tollinger, Martin; Kloiber, Karin; Lichtenecker, Roman; Ruedisser, Simon; Hommel, Ulrich; Schmid, Walther; Konrat, Robert; Kontaxis, Georg

    2008-01-01

    Direct methods in NMR based structure determination start from an unassigned ensemble of unconnected gaseous hydrogen atoms. Under favorable conditions they can produce low resolution structures of proteins. Usually a prohibitively large number of NOEs is required, to solve a protein structure ab-initio, but even with a much smaller set of distance restraints low resolution models can be obtained which resemble a protein fold. One problem is that at such low resolution and in the absence of a force field it is impossible to distinguish the correct protein fold from its mirror image. In a hybrid approach these ambiguous models have the potential to aid in the process of sequential backbone chemical shift assignment when 13 C β and 13 C' shifts are not available for sensitivity reasons. Regardless of the overall fold they enhance the information content of the NOE spectra. These, combined with residue specific labeling and minimal triple-resonance data using 13 C α connectivity can provide almost complete sequential assignment. Strategies for residue type specific labeling with customized isotope labeling patterns are of great advantage in this context. Furthermore, this approach is to some extent error-tolerant with respect to data incompleteness, limited precision of the peak picking, and structural errors caused by misassignment of NOEs

  1. Rhizoctonia solani and Bacterial Inoculants Stimulate Root Exudation of Antifungal Compounds in Lettuce in a Soil-Type Specific Manner

    Saskia Windisch

    2017-06-01

    Full Text Available Previous studies conducted on a unique field site comprising three contrasting soils (diluvial sand DS, alluvial loam AL, loess loam LL under identical cropping history, demonstrated soil type-dependent differences in biocontrol efficiency against Rhizoctonia solani-induced bottom rot disease in lettuce by two bacterial inoculants (Pseudomonas jessenii RU47 and Serratia plymuthica 3Re-4-18. Disease severity declined in the order DS > AL > LL. These differences were confirmed under controlled conditions, using the same soils in minirhizotron experiments. Gas chromatography-mass spectrometry (GC-MS profiling of rhizosphere soil solutions revealed benzoic and lauric acids as antifungal compounds; previously identified in root exudates of lettuce. Pathogen inoculation and pre-inoculation with bacterial inoculants significantly increased the release of antifungal root exudates in a soil type-specific manner; with the highest absolute levels detected on the least-affected LL soil. Soil type-dependent differences were also recorded for the biocontrol effects of the two bacterial inoculants; showing the highest efficiency after double-inoculation on the AL soil. However, this was associated with a reduction of shoot growth and root hair development and a limited micronutrient status of the host plants. Obviously, disease severity and the expression of biocontrol effects are influenced by soil properties with potential impact on reproducibility of practical applications.

  2. Exercise in the fasted state facilitates fibre type-specific intramyocellular lipid breakdown and stimulates glycogen resynthesis in humans

    De Bock, K.; Richter, Erik; Russell, A.P.

    2005-01-01

    g (kg bw)-1 h-1) exercise. In both conditions, subjects ingested 5 g carbohydrates per kg body weight during recovery. Fibre type-specific relative IMTG content was determined by Oil red O staining in needle biopsies from m. vastus lateralis before, immediately after and 4 h after exercise. During F...... sessions with an interval of 3 weeks. In each session subjects performed 2 h of constant-load bicycle exercise (~75% VO2,max), followed by 4 h of controlled recovery. On one occasion they exercised after an overnight fast (F), and on the other (CHO) they received carbohydrates before (~150 g) and during (1...... but not during CHO, the exercise bout decreased IMTG content in type I fibres from 18 ± 2% to 6 ± 2% (P = 0.007) area lipid staining. Conversely, during recovery, IMTG in type I fibres decreased from 15 ± 2% to 10 ± 2% in CHO, but did not change in F. Neither exercise nor recovery changed IMTG in type IIa fibres...

  3. Dynamics of bacterial communities in two unpolluted soils after spiking with phenanthrene: soil type specific and common responders

    Guo-Chun eDing

    2012-08-01

    Full Text Available Considering their key role for ecosystem processes, it is important to understand the response of microbial communities in unpolluted soils to pollution with polycyclic aromatic hydrocarbons (PAH. Phenanthrene, a model compound for PAH, was spiked to a Cambisol and a Luvisol soil. Total community DNA from phenanthrene-spiked and control soils collected on days 0, 21 and 63 were analyzed based on PCR-amplified 16S rRNA genefragments. Denaturing gradient gel electrophoresis (DGGE fingerprints of bacterial communities increasingly deviated with time between spiked and control soils. In taxon specific DGGE, significant responses of Alphaproteobacteria and Actinobacteria became only detectable after 63 days, while significant effects on Betaproteobacteria were detectable in both soils after 21 days. Comparison of the taxonomic distribution of bacteria in spiked and control soils on day 63 as revealed by pyrosequencing indicated soil type specific negative effects of phenanthrene on several taxa, many of them belonging to the Gamma-, Beta- or Deltaproteobacteria. Bacterial richness and evenness decreased in spiked soils. Despite the significant differences in the bacterial community structure between both soils on day 0, similar genera increased in relative abundance after PAH spiking, especially Sphingomonas and Polaromonas. However, this did not result in an increased overall similarity of the bacterial communities in both soils.

  4. Comprehensive analysis of ultrasonic vocalizations in a mouse model of fragile X syndrome reveals limited, call type specific deficits.

    Snigdha Roy

    Full Text Available Fragile X syndrome (FXS is a well-recognized form of inherited mental retardation, caused by a mutation in the fragile X mental retardation 1 (Fmr1 gene. The gene is located on the long arm of the X chromosome and encodes fragile X mental retardation protein (FMRP. Absence of FMRP in fragile X patients as well as in Fmr1 knockout (KO mice results, among other changes, in abnormal dendritic spine formation and altered synaptic plasticity in the neocortex and hippocampus. Clinical features of FXS include cognitive impairment, anxiety, abnormal social interaction, mental retardation, motor coordination and speech articulation deficits. Mouse pups generate ultrasonic vocalizations (USVs when isolated from their mothers. Whether those social ultrasonic vocalizations are deficient in mouse models of FXS is unknown. Here we compared isolation-induced USVs generated by pups of Fmr1-KO mice with those of their wild type (WT littermates. Though the total number of calls was not significantly different between genotypes, a detailed analysis of 10 different categories of calls revealed that loss of Fmr1 expression in mice causes limited and call-type specific deficits in ultrasonic vocalization: the carrier frequency of flat calls was higher, the percentage of downward calls was lower and that the frequency range of complex calls was wider in Fmr1-KO mice compared to their WT littermates.

  5. Numerical investigation of the coupled water and thermal management in PEM fuel cell

    Cao, Tao-Feng; Lin, Hong; Chen, Li; He, Ya-Ling; Tao, Wen-Quan

    2013-01-01

    Highlights: ► A fully coupled, non-equilibrium, anisotropic PEM fuel cell computational model is developed. ► The coupled water and heat transport processes are numerically investigated. ► Anisotropic property of gas diffusion layer has an effect on local cell performance. ► The boundary temperature greatly affects the cell local temperature and indirectly influences the saturation profile. ► The cathode gas inlet humidity slightly affects the local temperature distribution. - Abstract: Water and thermal managements are the most important issue in the operation and optimization of proton exchange membrane fuel cell (PEMFC). A three-dimensional, two-phase, non-isothermal model of PEMFC is presented in this paper. The model is used to investigate the interaction between water and thermal transport processes, the effects of anisotropic characters of gas diffusion layer, different boundary temperature of flow plate and the effect of gas inlet humidity. By comparing the numerical results of different cases, it is found that maximum cell temperature is higher in the isotropic gas diffusion layer; in contrast, the liquid saturation is lower than other case. Moreover, the boundary temperature greatly affects the temperature distribution in PEMFC, and indirectly influences the water saturation distribution. This indicates that the coupled relationship between water and thermal managements cannot be ignored, and these two processes must be considered simultaneously in the optimization of PEMFC

  6. Investigating the Thermodynamic Performances of TO-Based Metamaterial Tunable Cells with an Entropy Generation Approach

    Guoqiang Xu

    2017-10-01

    Full Text Available Active control of heat flux can be realized with transformation optics (TO thermal metamaterials. Recently, a new class of metamaterial tunable cells has been proposed, aiming to significantly reduce the difficulty of fabrication and to flexibly switch functions by employing several cells assembled on related positions following the TO design. However, owing to the integration and rotation of materials in tunable cells, they might lead to extra thermal losses as compared with the previous continuum design. This paper focuses on investigating the thermodynamic properties of tunable cells under related design parameters. The universal expression for the local entropy generation rate in such metamaterial systems is obtained considering the influence of rotation. A series of contrast schemes are established to describe the thermodynamic process and thermal energy distributions from the viewpoint of entropy analysis. Moreover, effects of design parameters on thermal dissipations and system irreversibility are investigated. In conclusion, more thermal dissipations and stronger thermodynamic processes occur in a system with larger conductivity ratios and rotation angles. This paper presents a detailed description of the thermodynamic properties of metamaterial tunable cells and provides reference for selecting appropriate design parameters on related positions to fabricate more efficient and energy-economical switchable TO devices.

  7. Investigation of an alkaline direct ethanol fuel cell with non Pt-catalyst

    Beyer, M.; Guelzow, E.; Uhm, S. [DLR Deutsches Zentrum fuer Luft- und Raumfahrt e.V., Stuttgart (Germany). Inst. fuer Technische Thermodynamik

    2010-07-01

    This paper focuses on the characterisation of an alkaline direct ethanol fuel cell (ADEFC). Ethanol and for comparison also methanol was fed to the anode in a caustic potash solution at different concentrations and temperatures. An anion exchange membrane (Tokuyama) sandwiched between two Hypermec electrodes (Acta SpA.) was investigated in a single cell. Current-voltage-measurements (U(I) characteristics), short term operation under load, electrochemical impedance spectroscopy (EIS) and pH recording were carried out to characterize fuel cell performance. The long term objective is to investigate the mechanism of ethanol electro oxidation reaction (EOR). 18 mW/cm{sup 2} was reached at room temperature with a technically oriented 50 cm{sup 2} cell with ethanol. However, poor long term stability under load of the fuel cell is observed. Furthermore in the U(I) characteristics a negative hysteresis is present in the forward and backward scan at room temperature which indicates poisoning intermediates of electrode reactions. A pH decline appears during operation indicating a development of either acetic acid or acetates or acetaldehyde as main products of the ethanol oxidation, which may be responsible for rate decrease of ethanol oxidation with time. EIS measurement shows an increased membrane resistance. (orig.)

  8. Microfluidic biofunctionalisation protocols to form multi-valent interactions for cell rolling and phenotype modification investigations

    Perozziello, Gerardo

    2013-07-01

    In this study, we propose a fast, simple method to biofunctionalise microfluidic systems for cellomic investigations based on micro-fluidic protocols. Many available processes either require expensive and time-consuming protocols or are incompatible with the fabrication of microfluidic systems. Our method differs from the existing since it is applicable to an assembled system, uses few microlitres of reagents and it is based on the use of microbeads. The microbeads have specific surface moieties to link the biomolecules and couple cell receptors. Furthermore, the microbeads serve as arm spacer and offer the benefit of the multi-valent interaction. Microfluidics was adapted together with topology and biochemistry surface modifications to offer the microenvironment for cellomic studies. Based on this principle, we exploit the streptavidin-biotin interaction to couple antibodies to the biofunctionalised microfluidic environment within 5 h using 200 μL of reagents and biomolecules. We selected the antibodies able to form complexes with the MHC class I (MHC-I) molecules present on the cell membrane and involved in the immune surveillance. To test the microfluidic system, tumour cell lines (RMA) were rolled across the coupled antibodies to recognise and strip MHC-I molecules. As result, we show that cell rolling performed inside a microfluidic chamber functionalised with beads and the opportune antibody facilitate the removal of MHC class I molecules. We showed that the level of median fluorescent intensity of the MHC-I molecules is 300 for cells treated in a not biofunctionalised surface. It decreased to 275 for cells treated in a flat biofunctionalised surface and to 250 for cells treated on a surface where biofunctionalised microbeads were immobilised. The cells with reduced expression of MHC-I molecules showed, after cytotoxicity tests, susceptibility 3.5 times higher than normal cells. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. [Investigation the Inhibitory Effects of Kaempferol on Rat Renalmesangial Cells Proliferation under High Glucose Condition].

    Chen, Ni; Han, Peng-Ding; Chen, Wen; Deng, Yan

    2017-07-01

    To investigate the protective effects of kaempferol on rat renal mesangial cells under high glucose condition and explore its mechanism. The HBZY-1 cells were divided into normal glucose group (5.5 mmol/L), high glucose group (25 mmol/L), 10 μmol/L kaempferol+high glucose group, and 30 μmol/L kaempferol+high glucose group. Cell proliferative ability was measured by MTT; cell cycle was analyzed by flow cytometry; mRNA and protein levels were determined by Real-time PCR and Western blot, respectively. Kaempferol had no effect on the proliferative ability of rat renal mesangial cells under normal glucose (5.5 mmol/L) condition. High glucose (25 mmol/L) enhanced the cell proliferative ability, and this effect was antagonized by kaempferol (10-30 μmol/L) treatment. High glucose reduced the cell population at G 0 /G 1 phase with an associated increase in S phase, and had no effect on G₂/M phase; and kaempferol treatment restored high glucose-induced changes in cell cycle. Kaempferol also prevented high glucose-induced increase in fibronectin and connective tissue growth factor mRNA and protein expression levels. Kaempferol also prevented high glucose-induced increase in fibronectin and connective tissue growth factor mRNA and protein expression levels. Further, high glucose caused an increase in protein level of phosphorylated p38 mitogen-activated protein kinases (p38 MAPK), which was antagonized by kaempferol treatment. Our results suggest that kaempferol exerts its protective effect on rat renal mesangial cells under high glucose condition via p38 MAPK signaling pathway.

  10. Investigating Ceria Nanocrystals Uptake by Glioblastoma Multiforme Cells and its Related Effects: An Electron Microscopy Study

    Aloufi, Bader

    2017-01-22

    Cerium oxide nanoparticles have been utilized widely nowadays in cancer research. It has been suggested by many studies that these nanoparticles are capable of having dual antioxidant behavior in healthy and cancer microenvironment; where in physiological condition, they act as antioxidant and do not affect the healthy cells, while in tumor-like condition; they act as an oxidase, and result in a selective killing for the cancer cells. In this experiment, the interaction of nanoceria with glioblastoma and healthy astrocyte cells was examined, and further correlated with the in vitro cytotoxic effects of various nanoceria concentrations (100 and 300 µg/ml) and exposure times (12, 24, and 48 hours). Electron microscopes were used to investigate the cellular-NPs interactions, and to examine the related cytotoxic effects in combination with trypan blue and propidium iodide viability assays. Our data suggest the following results. First, the two cell lines demonstrated capability of taken up the ceria through endocytosis pathway, where the NPs were recognized engulfed by double membrane vesicles at various regions over the cellular cytoplasm. Secondly, cerium oxide nanoparticles were found to affect the glioblastoma cells, but not so severely the corresponding healthy astrocytes at the various concentrations and incubation times, as revealed by the viability assays and the electron microscopy analysis. Thirdly, the viability of the glioblastoma cells after the treatment displayed a declined trend when increasing the ceria concentrations, but did not show such dependency with regard to the different time points. In all cases, the healthy astrocyte cells showed slight alterations in mitochondrial shape which did not influence their viability. Among the various nanoceria concentrations and exposure times, the most efficient dose of treatment was found to be with a concentration of 300 µg/ml at a time point of 24-hour, where higher reduction on the viability of

  11. Dual Role of miR-21 in CD4+T-Cells : Activation-Induced miR-21 Supports Survival of Memory T-Cells and Regulates CCR7 Expression in Naive T-Cells

    Smigielska-Czepiel, Katarzyna; van den Berg, Anke; Jellema, Pytrick; Slezak-Prochazka, Izabella; Maat, Henny; van den Bos, Hilda; van der Lei, Roelof Jan; Kluiver, Joost; Brouwer, Elisabeth; Boots, Anne Mieke H.; Kroesen, Bart-Jan

    2013-01-01

    Immune cell-type specific miRNA expression patterns have been described but the detailed role of single miRNAs in the function of T-cells remains largely unknown. We investigated the role of miR-21 in the function of primary human CD4+ T-cells. MiR-21 is substantially expressed in T-cells with a

  12. Investigation of integrin expression on the surface of osteoblast-like cells by atomic force microscopy

    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.

  13. Investigation of integrin expression on the surface of osteoblast-like cells by atomic force microscopy

    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.

  14. Investigation of Cytocidal Activity of Bacillus Thuringiensis Parasporal Toxin on CCRF-CEM Cell Line

    Elham Moazamian

    2013-03-01

    Full Text Available Background & Objective: Parasporin is a parasporal protein of Bacillus thuringiensis and exhibits special cytocidal activity against human cancer cells. Similar to other insecticidal Bacillus thuringiensis crystal toxins, parasporin shows target specificity and damages the cellular membrane. In this study, different strains of Bacillus thuringiensis isolated from various regions of Iran and their cytocidal activity against CCRF-CEM cell line and human erythrocyte were investigated.   Materials & Methods: Fifty soil samples were collected from different Iranian provinces, and characterization was performed based on protein crystal morphology by phase-contrast microscope and variations of Cry protein toxin using SDS-PAGE. After parasporin was processed with proteinase K, the active form was produced and protein activity on the cell line was evaluated. Results: Parasporal inclusion proteins showed different cytotoxicity against acute lymphoblastic leukemia cells (ALL, but not against normal lymphocyte. Isolated parasporin demonstrated no hemolytic activity against human erythrocyte. It appears that these proteins have the ability to differentiate between normal lymphocytes and leukemia cells and have specific receptors on specific cancer cell lines. Conclusion: Our results provide evidence that the parasporin-producing organism is a common member in Bacillus thuringiensis populations occurring in the natural environments of Iran.

  15. Ceratopteris richardii (C-fern: A model for investigating adaptive modification of vascular plant cell walls

    Olivier eLeroux

    2013-09-01

    Full Text Available Plant cell walls are essential for most aspects of plant growth, development, and survival, including cell division, expansive cell growth, cell-cell communication, biomechanical properties, and stress responses. Therefore, characterising cell wall diversity contributes to our overall understanding of plant evolution and development. Recent biochemical analyses, concomitantly with whole genome sequencing of plants located at pivotal points in plant phylogeny, have helped distinguish between homologous characters and those which might be more derived. Most plant lineages now have at least one fully sequenced representative and although genome sequences for fern species are in progress they not yet available this group. Ferns offer key advantages for the study of developmental processes leading to vascularisation and complex organs as well as the specific differences between diploid sporophyte tissues and haploid gametophyte tissues and the interplay between them. Ceratopteris richardii has been well investigated building a body of knowledge which combined with the genomic and biochemical information available for other plants will progress our understanding of wall diversity and its impact on evolution and development.

  16. Detection of FMD virus type specific IgG1, IgG2 and IgA antibodies in milk and serum of buffaloes vaccinated with oil adjuvanted polyvalent FMD vaccine

    R. Sharma

    2010-02-01

    Full Text Available The present investigation was carried out on 15 randomly selected milch buffaloes divided into three groups on the basis of lactation at an organized farm, to study the foot and mouth disease virus type specific antibodies in milk and serum following FMD vaccination. Milk and serum samples collected before vaccination i.e. 0 day and on 7, 14, 28, 42 and 56 days post vaccination, were analyzed for the detection of FMD virus specific IgG1, IgG2 and IgA antibody response by indirect double antibody sandwich ELISA. Significant FMD virus type specific antibody titres (IgG1, IgG2 and IgA were detected in milk and serum of buffaloes on different days post vaccination, though the levels of antibodies were lower in milk as compared to serum. FMD virus type specific IgG1 was found to be the predominant subclass as compared to IgG2 and IgA both in milk and serum of vaccinated buffaloes. Milk and serum IgG1, IgG2 and IgA antibody titres were positively correlated with values of regression coefficient (R as 0.506, 0.434 and 0.396, respectively.

  17. Systematic investigation of the impact of operation conditions on the degradation behaviour of perovskite solar cells

    Domanski, Konrad; Alharbi, Essa A.; Hagfeldt, Anders; Grätzel, Michael; Tress, Wolfgang

    2018-01-01

    Perovskite solar cells have achieved power-conversion efficiency values approaching those of established photovoltaic technologies, making the reliable assessment of their operational stability the next essential step towards commercialization. Although studies increasingly often involve a form of stability characterization, they are conducted in non-standardized ways, which yields data that are effectively incomparable. Furthermore, stability assessment of a novel material system with its own peculiarities might require an adjustment of common standards. Here, we investigate the effects of different environmental factors and electrical load on the ageing behaviour of perovskite solar cells. On this basis, we comment on our perceived relevance of the different ways these are currently aged. We also demonstrate how the results of the experiments can be distorted and how to avoid the common pitfalls. We hope this work will initiate discussion on how to age perovskite solar cells and facilitate the development of consensus stability measurement protocols.

  18. Metallofullerenes as fuel cell electrocatalysts: a theoretical investigation of adsorbates on C59Pt.

    Gabriel, Margaret A; Genovese, Luigi; Krosnicki, Guillaume; Lemaire, Olivier; Deutsch, Thierry; Franco, Alejandro A

    2010-08-28

    Nano-structured electrode degradation in state-of-the-art polymer electrolyte membrane fuel cells (PEMFCs) is one of the main shortcomings that limit the large-scale development and commercialization of this technology. During normal operating conditions of the fuel cell, the PEMFC lifetime tends to be limited by coarsening of the cathode's Pt-based catalyst and by corrosion of the cathode's carbon black support. Because of their chemical properties, metallofullerenes such as C(59)Pt may be more electrochemically stable than the Pt/C mixture. In this paper we investigate, by theoretical methods, the stability of oxygen reduction reaction (ORR) adsorbates on the metallofullerene C(59)Pt and evaluate its potential as a PEMFC fuel cell catalyst.

  19. Investigation of ITO layers for application as transparent contacts in flexible photovoltaic cell structures

    Znajdek, Katarzyna; Sibiński, Maciej

    2013-07-01

    In this paper authors present the mechanical, optical and electrical parameters of Indium Tin Oxide (ITO) Transparent Conductive Layers (TCL) deposited on flexible substrate. Layers' properties are analyzed and verified. Investigated Transparent Conductive Oxide (TCO) was deposited, using magnetron sputtering method. Flexible polymer PET (polyethylene terephthalate) foil was used as a substrate, in order to photovoltaic (PV) cell's emitter contact application of investigated material. ITO-coated PET foils have been dynamically bent on numerous cylinders of various diameters according to the standard requirements. Resistance changes for each measured sample were measured and recorded during bending cycle. Thermal durability, as well as temperature influence on resistance and optical transmission are verified. Presented results were conducted to verify practical suitability and to evaluate possible applications of Indium Tin Oxide as a front contact in flexible photovoltaic cell structures.

  20. Investigation of Micro- and Macro-Scale Transport Processes for Improved Fuel Cell Performance

    Gu, Wenbin [General Motors LLC, Pontiac, MI (United States)

    2014-08-29

    This report documents the work performed by General Motors (GM) under the Cooperative agreement No. DE-EE0000470, “Investigation of Micro- and Macro-Scale Transport Processes for Improved Fuel Cell Performance,” in collaboration with the Penn State University (PSU), University of Tennessee Knoxville (UTK), Rochester Institute of Technology (RIT), and University of Rochester (UR) via subcontracts. The overall objectives of the project are to investigate and synthesize fundamental understanding of transport phenomena at both the macro- and micro-scales for the development of a down-the-channel model that accounts for all transport domains in a broad operating space. GM as a prime contractor focused on cell level experiments and modeling, and the Universities as subcontractors worked toward fundamental understanding of each component and associated interface.

  1. Cellular Analysis of Adult Neural Stem Cells for Investigating Prion Biology.

    Haigh, Cathryn L

    2017-01-01

    Traditional primary and secondary cell cultures have been used for the investigation of prion biology and disease for many years. While both types of cultures produce highly valid and immensely valuable results, they also have their limitations; traditional cell lines are often derived from cancers, therefore subject to numerous DNA changes, and primary cultures are labor-intensive and expensive to produce requiring sacrifice of many animals. Neural stem cell (NSC) cultures are a relatively new technology to be used for the study of prion biology and disease. While NSCs are subject to their own limitations-they are generally cultured ex vivo in environments that artificially force their growth-they also have their own unique advantages. NSCs retain the ability for self-renewal and can therefore be propagated in culture similarly to secondary cultures without genetic manipulation. In addition, NSCs are multipotent; they can be induced to differentiate into mature cells of central nervous system (CNS) linage. The combination of self-renewal and multipotency allows NSCs to be used as a primary cell line over multiple generations saving time, costs, and animal harvests, thus providing a valuable addition to the existing cell culture repertoire used for investigation of prion biology and disease. Furthermore, NSC cultures can be generated from mice of any genotype, either by embryonic harvest or harvest from adult brain, allowing gene expression to be studied without further genetic manipulation. This chapter describes a standard method of culturing adult NSCs and assays for monitoring NSC growth, migration, and differentiation and revisits basic reactive oxygen species detection in the context of NSC cultures.

  2. A preliminary investigation into the prevalence and prediction of problematic cell phone use.

    Smetaniuk, Peter

    2014-03-01

    Likening mobile phone use dependency to the classification of excessive behaviors may be necessarily equivalent in seriousness to previously established addictions such as problematic computing or excessive gambling. The aim of the study explores into the behavior of excessive use of mobile phones as a pathological behavior. Two studies investigated criteria for problematic mobile phone usage by examining student (Study 1, N = 301) and nonstudent (Study 2, N = 362) responses to a set of adapted mobile phone addiction inventories. Study 1 investigated cell phone addiction inventories as constructs designed to measure problematic cell phone use. Additionally, Study 2 sought to predict age, depression, extraversion, emotional stability, impulse control, and self-esteem as independent variables that augment respondents' perceptions of problematic use. The results from Study 1 and Study 2 indicate that 10 to 25% of the participants tested exhibited problematic cell phone usage. Additionally, age, depression, extraversion, and low impulse control are the most suitable predictors for problematic use. The results of the two studies indicate that problematic mobile phone use does occur and ought to be taken seriously by the psychological community. Presently, there is limited data providing conclusive evidence for a comprehensible categorization of cell phone addiction, as well as a unified explanatory model specific to problematic mobile phone use. Studies such as this one may contribute substantial findings, adding scientific significance, and offering a valuable submission for the ongoing progress of creating intervention frameworks relative to "virtual addictions".

  3. A preliminary investigation into the prevalence and prediction of problematic cell phone use

    Smetaniuk, Peter

    2014-01-01

    Background and aims: Likening mobile phone use dependency to the classification of excessive behaviors may be necessarily equivalent in seriousness to previously established addictions such as problematic computing or excessive gambling. The aim of the study explores into the behavior of excessive use of mobile phones as a pathological behavior. Methods: Two studies investigated criteria for problematic mobile phone usage by examining student (Study 1, N = 301) and nonstudent (Study 2, N = 362) responses to a set of adapted mobile phone addiction inventories. Study 1 investigated cell phone addiction inventories as constructs designed to measure problematic cell phone use. Additionally, Study 2 sought to predict age, depression, extraversion, emotional stability, impulse control, and self-esteem as independent variables that augment respondents’ perceptions of problematic use. Results: The results from Study 1 and Study 2 indicate that 10 to 25% of the participants tested exhibited problematic cell phone usage. Additionally, age, depression, extraversion, and low impulse control are the most suitable predictors for problematic use. Conclusions: The results of the two studies indicate that problematic mobile phone use does occur and ought to be taken seriously by the psychological community. Presently, there is limited data providing conclusive evidence for a comprehensible categorization of cell phone addiction, as well as a unified explanatory model specific to problematic mobile phone use. Studies such as this one may contribute substantial findings, adding scientific significance, and offering a valuable submission for the ongoing progress of creating intervention frameworks relative to “virtual addictions”. PMID:25215213

  4. Cell Systems to Investigate the Impact of Polyphenols on Cardiovascular Health

    Charlotte Grootaert

    2015-11-01

    Full Text Available Polyphenols are a diverse group of micronutrients from plant origin that may serve as antioxidants and that contribute to human health in general. More specifically, many research groups have investigated their protective effect against cardiovascular diseases in several animal studies and human trials. Yet, because of the excessive processing of the polyphenol structure by human cells and the residing intestinal microbial community, which results in a large variability between the test subjects, the exact mechanisms of their protective effects are still under investigation. To this end, simplified cell culture systems have been used to decrease the inter-individual variability in mechanistic studies. In this review, we will discuss the different cell culture models that have been used so far for polyphenol research in the context of cardiovascular diseases. We will also review the current trends in cell culture research, including co-culture methodologies. Finally, we will discuss the potential of these advanced models to screen for cardiovascular effects of the large pool of bioactive polyphenols present in foods and their metabolites.

  5. Quantum Dot Nanotoxicity Investigations Using Human Lung Cells and TOXOR Electrochemical Enzyme Assay Methodology.

    O'Hara, Tony; Seddon, Brian; O'Connor, Andrew; McClean, Siobhán; Singh, Baljit; Iwuoha, Emmanuel; Fuku, Xolile; Dempsey, Eithne

    2017-01-27

    Recent studies have suggested that certain nanomaterials can interfere with optically based cytotoxicity assays resulting in underestimations of nanomaterial toxicity. As a result there has been growing interest in the use of whole cell electrochemical biosensors for nanotoxicity applications. Herein we report application of an electrochemical cytotoxicity assay developed in house (TOXOR) in the evaluation of toxic effects of mercaptosuccinic acid capped cadmium telluride quantum dots (MSA capped CdTe QDs), toward mammalian cells. MSA capped CdTe QDs were synthesized, characterized, and their cytotoxicity toward A549 human lung epithelial cells investigated. The internalization of QDs within cells was scrutinized via confocal microscopy. The cytotoxicity assay is based on the measurement of changes in cellular enzyme acid phosphatase upon 24 h exposure to QDs. Acid phosphatase catalyzes dephosphorylation of 2-naphthyl phosphate to 2-naphthol (determined by chronocoulometry) and is indicative of metabolic activity in cells. The 24 h IC50 (concentration resulting in 50% reduction in acid phosphatase activity) value for MSA capped CdTe QDs was found to be 118 ± 49 μg/mL using the TOXOR assay and was in agreement with the MTT assay (157 ± 31 μg/mL). Potential uses of this electrochemical assay include the screening of nanomaterials, environmental toxins, in addition to applications in the pharmaceutical, food, and health sectors.

  6. Integration of statistical modeling and high-content microscopy to systematically investigate cell-substrate interactions.

    Chen, Wen Li Kelly; Likhitpanichkul, Morakot; Ho, Anthony; Simmons, Craig A

    2010-03-01

    Cell-substrate interactions are multifaceted, involving the integration of various physical and biochemical signals. The interactions among these microenvironmental factors cannot be facilely elucidated and quantified by conventional experimentation, and necessitate multifactorial strategies. Here we describe an approach that integrates statistical design and analysis of experiments with automated microscopy to systematically investigate the combinatorial effects of substrate-derived stimuli (substrate stiffness and matrix protein concentration) on mesenchymal stem cell (MSC) spreading, proliferation and osteogenic differentiation. C3H10T1/2 cells were grown on type I collagen- or fibronectin-coated polyacrylamide hydrogels with tunable mechanical properties. Experimental conditions, which were defined according to central composite design, consisted of specific permutations of substrate stiffness (3-144 kPa) and adhesion protein concentration (7-520 microg/mL). Spreading area, BrdU incorporation and Runx2 nuclear translocation were quantified using high-content microscopy and modeled as mathematical functions of substrate stiffness and protein concentration. The resulting response surfaces revealed distinct patterns of protein-specific, substrate stiffness-dependent modulation of MSC proliferation and differentiation, demonstrating the advantage of statistical modeling in the detection and description of higher-order cellular responses. In a broader context, this approach can be adapted to study other types of cell-material interactions and can facilitate the efficient screening and optimization of substrate properties for applications involving cell-material interfaces. Copyright 2009 Elsevier Ltd. All rights reserved.

  7. Disruption of steroidogenesis: Cell models for mechanistic investigations and as screening tools.

    Odermatt, Alex; Strajhar, Petra; Engeli, Roger T

    2016-04-01

    In the modern world, humans are exposed during their whole life to a large number of synthetic chemicals. Some of these chemicals have the potential to disrupt endocrine functions and contribute to the development and/or progression of major diseases. Every year approximately 1000 novel chemicals, used in industrial production, agriculture, consumer products or as pharmaceuticals, are reaching the market, often with limited safety assessment regarding potential endocrine activities. Steroids are essential endocrine hormones, and the importance of the steroidogenesis pathway as a target for endocrine disrupting chemicals (EDCs) has been recognized by leading scientists and authorities. Cell lines have a prominent role in the initial stages of toxicity assessment, i.e. for mechanistic investigations and for the medium to high throughput analysis of chemicals for potential steroidogenesis disrupting activities. Nevertheless, the users have to be aware of the limitations of the existing cell models in order to apply them properly, and there is a great demand for improved cell-based testing systems and protocols. This review intends to provide an overview of the available cell lines for studying effects of chemicals on gonadal and adrenal steroidogenesis, their use and limitations, as well as the need for future improvements of cell-based testing systems and protocols. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Investigating the use of in situ liquid cell scanning transmission electron microscopy

    Nguy, Amanda [Iowa State Univ., Ames, IA (United States)

    2016-02-19

    Engineering nanoparticles with desired shape-dependent properties is the key to many applications in nanotechnology. Although many synthetic procedures exist to produce anisotropic gold nanoparticles, the dynamics of growth are typically unknown or hypothetical. In the case of seed-mediated growth in the presence of DNA into anisotropic nanoparticles, it is not known exactly how DNA directs growth into specific morphologies. A series of preliminary experiments were carried out to contribute to the investigation of the possible mechanism of DNA-mediated growth of gold nanoprisms into gold nanostars using liquid cell scanning transmission electron microscopy (STEM). Imaging in the liquid phase was achieved through the use of a liquid cell platform and liquid cell holder that allow the sample to be contained within a “chip sandwich” between two electron transparent windows. Ex situ growth experiments were performed using Au-T30 NPrisms (30-base thymine oligonucleotide-coated gold nanoprisms) that are expected to grow into gold nanostars. Growth to form these nanostars were imaged using TEM (transmission electron microscopy) and liquid cell STEM (scanning transmission electron microscopy). An attempt to perform in situ growth experiments with the same Au-T30 nanoprisms revealed challenges in obtaining desired morphology results due to the environmental differences within the liquid cell compared to the ex situ environment. Different parameters in the experimental method were explored including fluid line set up, simultaneous and alternating reagent addition, and the effect of different liquid cell volumes to ensure adequate flow of reagents into the liquid cell. Lastly, the binding affinities were compared for T30 and A30 DNA incubated with gold nanoparticles using zeta potential measurements, absorption spectroscopy, and isothermal titration calorimetry (ITC). It was previously reported thymine bases have a lower binding affinity to gold surfaces than adenine

  9. Proteomic investigation into betulinic acid-induced apoptosis of human cervical cancer HeLa cells.

    Xu, Tao; Pang, Qiuying; Zhou, Dong; Zhang, Aiqin; Luo, Shaman; Wang, Yang; Yan, Xiufeng

    2014-01-01

    Betulinic acid is a pentacyclic triterpenoid that exhibits anticancer functions in human cancer cells. This study provides evidence that betulinic acid is highly effective against the human cervical cancer cell line HeLa by inducing dose- and time-dependent apoptosis. The apoptotic process was further investigated using a proteomics approach to reveal protein expression changes in HeLa cells following betulinic acid treatment. Proteomic analysis revealed that there were six up- and thirty down-regulated proteins in betulinic acid-induced HeLa cells, and these proteins were then subjected to functional pathway analysis using multiple analysis software. UDP-glucose 6-dehydrogenase, 6-phosphogluconate dehydrogenase decarboxylating, chain A Horf6-a novel human peroxidase enzyme that involved in redox process, was found to be down-regulated during the apoptosis process of the oxidative stress response pathway. Consistent with our results at the protein level, an increase in intracellular reactive oxygen species was observed in betulinic acid-treated cells. The proteins glucose-regulated protein and cargo-selection protein TIP47, which are involved in the endoplasmic reticulum pathway, were up-regulated by betulinic acid treatment. Meanwhile, 14-3-3 family proteins, including 14-3-3β and 14-3-3ε, were down-regulated in response to betulinic acid treatment, which is consistent with the decrease in expression of the target genes 14-3-3β and 14-3-3ε. Furthermore, it was found that the antiapoptotic bcl-2 gene was down-regulated while the proapoptotic bax gene was up-regulated after betulinic acid treatment in HeLa cells. These results suggest that betulinic acid induces apoptosis of HeLa cells by triggering both the endoplasmic reticulum pathway and the ROS-mediated mitochondrial pathway.

  10. Investigation of triamterene as an inhibitor of the TGR5 receptor: identification in cells and animals.

    Li, Yingxiao; Cheng, Kai Chun; Niu, Chiang-Shan; Lo, Shih-Hsiang; Cheng, Juei-Tang; Niu, Ho-Shan

    2017-01-01

    G-protein-coupled bile acid receptor 1 (GPBAR1, also known as TGR5) has been shown to participate in glucose homeostasis. In animal models, a TGR5 agonist increases incretin secretion to reduce hyperglycemia. Many agonists have been developed for clinical use. However, the effects of TGR5 blockade have not been studied extensively, with the exception of studies using TGR5 knockout mice. Therefore, we investigated the potential effect of triamterene on TGR5. We transfected the TGR5 gene into cultured Chinese hamster ovary cells (CHO-K1 cells) to express TGR5. Then, we applied a fluorescent indicator to examine the glucose uptake of these transfected cells. In addition, NCI-H716 cells that secrete incretin were also evaluated. Fura-2, a fluorescence indicator, was applied to determine the changes in calcium concentrations. The levels of cyclic adenosine monophosphate (cAMP) and glucagon-like peptide (GLP-1) were estimated using enzyme-linked immunosorbent assay kits. Moreover, rats with streptozotocin (STZ)-induced type 1-like diabetes were used to investigate the effects in vivo. Triamterene dose dependently inhibits the increase in glucose uptake induced by TGR5 agonists in CHO-K1 cells expressing the TGR5 gene. In cultured NCI-H716 cells, TGR5 activation also increases GLP-1 secretion by increasing calcium levels. Triamterene inhibits the increased calcium levels by TGR5 activation through competitive antagonism. Moreover, the GLP-1 secretion and increased cAMP levels induced by TGR5 activation are both dose dependently reduced by triamterene. However, treatment with KB-R7943 at a dose sufficient to block the Na + /Ca 2+ exchanger (NCX) failed to modify the responses to TGR5 activation in NCI-H716 cells or CHO-K1 cells expressing TGR5. Therefore, the inhibitory effects of triamterene on TGR5 activation do not appear to be related to NCX inhibition. Blockade of TGR5 activation by triamterene was further characterized in vivo using the STZ-induced diabetic rats

  11. Investigation of the response of low-dose irradiated cells. Pt. 2. Radio-adaptive response of human embryonic cells is related to cell-to-cell communication

    Ishii, Keiichiro; Watanabe, Masami.

    1994-01-01

    To clarify the radio-adaptive response of normal cells to low-dose radiation, we irradiated human embryonic cells and HeLa cells with low-dose X-ray and examined the changes in sensitivity to subsequent high-dose X-irradiation. The results obtained were as follows; (1) When HE cells were irradiated by a high-dose of 200 cGy, the growth ratio of the living cells five days after the irradiation decreased to 37% of that of the cells which received no X-irradiation. When the cells received a preliminary irradiation of 10 to 20 cGy four hours before the irradiation of 200 cGy, the relative growth ratios increased significantly to 45-53%. (2) This preliminary irradiation effect was not observed in HeLa cells, being cancer cells. (3) When the HE cells suspended in a Ca 2+ iron-free medium or TPA added medium while receiving the preliminary irradiation of 13 cGy, the effect of the preliminary irradiation in increasing the relative growth ratio of living cells was not observed. (4) This indicates that normal cells shows an adaptive response to low-dose radiation and become more radioresistant. This phenomenon is considered to involve cell-to-cell communication maintained in normal cells and intracellular signal transduction in which Ca 2+ ion plays a role. (author)

  12. An eight-year epidemiologic study based on baculovirus-expressed type-specific spike proteins for the differentiation of type I and II feline coronavirus infections

    2014-01-01

    Background Feline infectious peritonitis (FIP) is a fatal disease caused by feline coronavirus (FCoV). FCoVs are divided into two serotypes with markedly different infection rates among cat populations around the world. A baculovirus-expressed type-specific domain of the spike proteins of FCoV was used to survey the infection of the two viruses over the past eight years in Taiwan. Results An immunofluorescence assay based on cells infected with the recombinant viruses that was capable of distinguishing between the two types of viral infection was established. A total of 833 cases from a teaching hospital was surveyed for prevalence of different FCoV infections. Infection of the type I FCoV was dominant, with a seropositive rate of 70.4%, whereas 3.5% of cats were infected with the type II FCoV. In most cases, results derived from serotyping and genotyping were highly agreeable. However, 16.7% (4/24) FIP cats and 9.8% (6/61) clinically healthy cats were found to possess antibodies against both viruses. Moreover, most of the cats (84.6%, 22/26) infected with a genotypic untypable virus bearing a type I FCoV antibody. Conclusion A relatively simple serotyping method to distinguish between two types of FCoV infection was developed. Based on this method, two types of FCoV infection in Taiwan was first carried out. Type I FCoV was found to be predominant compared with type II virus. Results derived from serotyping and genotyping support our current understanding of evolution of disease-related FCoV and transmission of FIP. PMID:25123112

  13. Effects of temperature and cellular interactions on the mechanics and morphology of human cancer cells investigated by atomic force microscopy.

    Li, Mi; Liu, LianQing; Xi, Ning; Wang, YueChao; Xiao, XiuBin; Zhang, WeiJing

    2015-09-01

    Cell mechanics plays an important role in cellular physiological activities. Recent studies have shown that cellular mechanical properties are novel biomarkers for indicating the cell states. In this article, temperature-controllable atomic force microscopy (AFM) was applied to quantitatively investigate the effects of temperature and cellular interactions on the mechanics and morphology of human cancer cells. First, AFM indenting experiments were performed on six types of human cells to investigate the changes of cellular Young's modulus at different temperatures and the results showed that the mechanical responses to the changes of temperature were variable for different types of cancer cells. Second, AFM imaging experiments were performed to observe the morphological changes in living cells at different temperatures and the results showed the significant changes of cell morphology caused by the alterations of temperature. Finally, by co-culturing human cancer cells with human immune cells, the mechanical and morphological changes in cancer cells were investigated. The results showed that the co-culture of cancer cells and immune cells could cause the distinct mechanical changes in cancer cells, but no significant morphological differences were observed. The experimental results improved our understanding of the effects of temperature and cellular interactions on the mechanics and morphology of cancer cells.

  14. Analysis of cell flow and cell loss following X-irradiation using sequential investigation of the total number of cells in the various parts of the cell cycle

    Skog, S.; Tribukait, B.

    1985-01-01

    The cell flow and cell loss of an in vivo growing Ehrlich ascites tumour were calculated by sequential estimation of changes in total number of cells in the cell cycle compartments. Normal growth was compared with the grossly disturbed cell flow evident after a 5 Gy X-irradiation. The doubling time of normal, exponentially growing cells was 24 hr. The generation time was 21 hr and the potential doubling time was 21 hr. Thus, the growth fraction was 1.0 and the cell loss rate about 0.5%/hr. Following irradiation, a transiently increased relative outflow rate from all cell cycle compartments was found at about 3 and 40 hr, and from S phase at 24 hr after irradiation. Increase in cell loss as well as non-viable cells was observed at 24 hr after irradiation at the time of release of the irradiation-induced G 2 blockage. The experiments show the applicability and limitations of cell flow and cell loss calculations by sequential analysis of the total number of cells in the various parts of the cell cycle. (author)

  15. In-depth investigation of spin-on doped solar cells with thermally grown oxide passivation

    Samir Mahmmod Ahmad

    Full Text Available Solar cell industrial manufacturing, based largely on proven semiconductor processing technologies supported by significant advancements in automation, has reached a plateau in terms of cost and efficiency. However, solar cell manufacturing cost (dollar/watt is still substantially higher than fossil fuels. The route to lowering cost may not lie with continuing automation and economies of scale. Alternate fabrication processes with lower cost and environmental-sustainability coupled with self-reliance, simplicity, and affordability may lead to price compatibility with carbon-based fuels. In this paper, a custom-designed formulation of phosphoric acid has been investigated, for n-type doping in p-type substrates, as a function of concentration and drive-in temperature. For post-diffusion surface passivation and anti-reflection, thermally-grown oxide films in 50–150-nm thickness were grown. These fabrication methods facilitate process simplicity, reduced costs, and environmental sustainability by elimination of poisonous chemicals and toxic gases (POCl3, SiH4, NH3. Simultaneous fire-through contact formation process based on screen-printed front surface Ag and back surface through thermally grown oxide films was optimized as a function of the peak temperature in conveyor belt furnace. Highest efficiency solar cells fabricated exhibited efficiency of ∼13%. Analysis of results based on internal quantum efficiency and minority carried measurements reveals three contributing factors: high front surface recombination, low minority carrier lifetime, and higher reflection. Solar cell simulations based on PC1D showed that, with improved passivation, lower reflection, and high lifetimes, efficiency can be enhanced to match with commercially-produced PECVD SiN-coated solar cells. Keywords: Crystalline Si solar cells, Phosphoric acid spin-on doping, Screen printing, Thermal oxide passivation

  16. Numerical investigation of optimized CZTSSe based solar cell in Wx-Amps environment

    Mohanty, Soumya Priyadarshini; Padhy, Srinibasa; Chowdhury, Joy; Sing, Udai P.

    2018-05-01

    The CZTSSe is the modified version of CZTS with selenium infusion. It shows maximum efficiency in the band gap from 1 to 1.4 eV. In our present work CZTSSe based solar cell is investigated using Wx-Amps tool. The Mo layer, absorber layer, CdS layer, i-ZnO [4]and Al-ZnO layers with their electrical, optical and material parameters are fitted in the tool. The vital parameters such as carrier density, thickness of the CZTSSe absorber layer, operating temperature, CdS buffer layer thickness and its carrier density on the cell interpretation are calculated. From[4] the simulation results it is apparent that the optimal absorber layer varies from 2.9 µm to 3.7 µm. The temperature variation has a strong influence on the efficiency of the cell. An optimal efficiency of 22% (With Jsc=33 mA/cm2, Voc=0.98 V, and fill factor= 68%) are attained. These results will give some insight for makeing higher efficiency CZTSSe based solar cell.

  17. An investigation of changes in element distribution and chemical states during differentiation of embryonic stem cells

    Sugimoto, T.; Ide-Ektessabi, A.; Ishihara, R.; Tanigaki, M.

    2004-01-01

    Metallic elements and their organic compounds have dynamic regulatory functions in cells. In this study, we implemented a new approach to investigate the mechanism of differentiation of embryonic stem cells, by measuring and analyzing the change in distribution and chemical states of intracellular trace elements. We anticipate that trace metal elements and metalloproteins play important roles in the direction of differentiation, both as active centers, and as factors in the death of neural cells in neurodegenerative disorders. The aim of this study is to analyze the distribution and chemical states of trace elements during the process of differentiation of mouse embryonic stem cells, and to understand how these factors relate to the differentiation process. Using the experimental results, some previously unexplained points are considered, namely (1) how the intracellular elements change during the process of neuronal differentiation, and (2) what the optimal conditions of such elements are for neuronal differentiation. The information obtained during this study is relevant to nervous system development and evolution

  18. An investigation of changes in element distribution and chemical states during differentiation of embryonic stem cells

    Sugimoto, T.; Ide-Ektessabi, A. E-mail: h51167@sakura.kudpc.kyoto-u.ac.jp; Ishihara, R.; Tanigaki, M

    2004-07-01

    Metallic elements and their organic compounds have dynamic regulatory functions in cells. In this study, we implemented a new approach to investigate the mechanism of differentiation of embryonic stem cells, by measuring and analyzing the change in distribution and chemical states of intracellular trace elements. We anticipate that trace metal elements and metalloproteins play important roles in the direction of differentiation, both as active centers, and as factors in the death of neural cells in neurodegenerative disorders. The aim of this study is to analyze the distribution and chemical states of trace elements during the process of differentiation of mouse embryonic stem cells, and to understand how these factors relate to the differentiation process. Using the experimental results, some previously unexplained points are considered, namely (1) how the intracellular elements change during the process of neuronal differentiation, and (2) what the optimal conditions of such elements are for neuronal differentiation. The information obtained during this study is relevant to nervous system development and evolution.

  19. Investigation of Novel Electrolytes for Use in Lithium-Ion Batteries and Direct Methanol Fuel Cells

    Pilar, Kartik

    Energy storage and conversion plays a critical role in the efficient use of available energy and is crucial for the utilization of renewable energy sources. To achieve maximum efficiency of renewable energy sources, improvements to energy storage materials must be developed. In this work, novel electrolytes for secondary batteries and fuel cells have been studied using nuclear magnetic resonance and high pressure x-ray scattering techniques to form a better understanding of dynamic and structural properties of these materials. Ionic liquids have been studied due to their potential as a safer alternative to organic solvent-based electrolytes in lithium-ion batteries and composite sulfonated polyetheretherketone (sPEEK) membranes have been investigated for their potential use as a proton exchange membrane electrolyte in direct methanol fuel cells. The characterization of these novel electrolytes is a step towards the development of the next generation of improved energy storage and energy conversion devices.

  20. The nanotoxicity investigation of optical nanoparticles to cultured cells in vitro.

    Zeng, Qinghui; Shao, Dan; Ji, Wenyu; Li, Jing; Chen, Li; Song, Jie

    2014-01-01

    Optical nanoparticles (NPs) have the potential to provide new tools for diagnosis and treatment of human diseases, however, their nanotoxicity and biological characteristics are still unclear. Here, we prepared a series of typical NPs (including gold nanospheres, gold nanorods, silver nanopheres, silver triangular nanoplates and quantum dots) with different material and surface chemical modification for nanotoxicity test. Cell proliferation was investigated by SRB assay where the NPs were co-cultured with cancer cells. It was found that NPs' toxicity was highly correlated to different factors-material selection, physical size/surface area, shape, and surface chemical property, etc. This work has the potential to provide a uniform and systematic information when they are applied as probes in biological and medical fields.

  1. Investigations of the temperature distribution in proton exchange membrane fuel cells

    Jung, Chi-Young; Shim, Hyo-Sub; Koo, Sang-Man; Lee, Sang-Hwan; Yi, Sung-Chul

    2012-01-01

    A two-dimensional, non-isothermal model of a proton exchange membrane fuel cell was implemented to elucidate heat balance through the membrane electrode assembly (MEA). To take local utilization of platinum catalyst into account, the model was presented by considering the formation of agglomerated catalyst structure in the electrodes. To estimate energy balance through the MEA, various modes of heat generation and depletion by reversible/irreversible heat release, ohmic heating and phase change of water were included in the present model. In addition, dual-pathway kinetics, that is a combination of Heyrovsky–Volmer and Tafel–Volmer kinetics, were employed to precisely describe the hydrogen oxidation reaction. The proposed model was validated with experimental cell polarization, resulting in excellent fit. The temperature distribution inside the MEA was analyzed by the model. Consequently, a thorough investigation was made of the relation between membrane thickness and the temperature distribution inside the MEA.

  2. Investigation of edge plasmas in the anchor cell region of GAMMA 10

    Islam, Khairul; Nakashima, Yousuke; Yatsu, Kiyoshi

    2000-01-01

    The first results of Langmuir probe measurements at the outer transition region of the anchor cell of GAMMA 10 are given. A probe current asymmetry in vertical direction is found in this region. It is also found that the asymmetry of probe current increases in outward direction and the direction of the asymmetry is independent on movable limiter position. A relation of the plasma asymmetry with the main magnetic field configuration is investigated. Plasma flow through the non-asymmetric magnetic field configuration region is thought to be the source of plasma asymmetry in this region, i.e., ∇B and curvature drifts are responsible for the asymmetry. Possibility of cold plasma formation in the anchor cell region is obtained during plug electron cyclotron resonance heating (ECRH) and can be explained with the desorption of particles due to the collision of the drifted out particles with the wall. (author)

  3. Cold-water immersion after training sessions: Effects on fiber type-specific adaptations in muscle K+ transport proteins to sprint-interval training in men.

    Christiansen, Danny; Bishop, David John; Broatch, James R; Bangsbo, Jens; McKenna, Michael John; Murphy, Robyn M

    2018-05-10

    Effects of regular use of cold-water immersion (CWI) on fiber type-specific adaptations in muscle K + transport proteins to intense training, along with their relationship to changes in mRNA levels after the first training session, were investigated in humans. Nineteen recreationally-active men (24{plus minus}6 y, 79.5{plus minus}10.8 kg, 44.6{plus minus}5.8 mL∙kg -1 ∙min -1 ) completed six weeks of sprint-interval cycling either without (passive rest; CON) or with training sessions followed by CWI (15 min at 10{degree sign}C; COLD). Muscle biopsies were obtained before and after training to determine abundance of Na + ,K + -ATPase isoforms (α 1-3 , β 1-3 ) and FXYD1, and after recovery treatments (+0h and +3h) on the first day of training to measure mRNA content. Training increased (ptraining (p>0.05). CWI after each session did not influence responses to training (p>0.05). However, α 2 mRNA increased after the first session in COLD (+0h, p0.05). In both conditions, α 1 and β 3 mRNA increased (+3h; p 0.05) after the first session. In summary, Na + ,K + -ATPase isoforms are differently regulated in type I and II muscle fibers by sprint-interval training in humans, which for most isoforms do not associate with changes in mRNA levels after the first training session. CWI neither impairs nor improves protein adaptations to intense training of importance for muscle K + regulation.

  4. In vitro investigations of Cynara scolymus L. extract on cell physiology of HepG2 liver cells

    Gesine Löhr

    2009-06-01

    Full Text Available The objective of this study was the investigation of a potential influence of artichoke leaf extract (ALE on the cell physiology and gene expression of phase I/II enzymes of human liver cells HepG2 and investigation on potential cell protective effects against ethanol-induced cell toxicity against HepG2 cells. Cell biological assays under in vitro conditions using HepG2 liver cells and investigation of mitochondrial activity (MTT test, proliferation assay (BrdU incorporation ELISA, LDH as toxicity marker, gene expression analysis by RT-PCR and enzyme activity of glutationtransferase. Artichocke extract, containing 27% caffeoylquinic acids and 7% flavonoids induced mitochondrial activity, proliferation and total protein content under in vitro conditions in human liver cells HepG2. These effects could not be correlated to the well-known artichoke secondary compounds cynarin, caffeic acid, chlorogenic acid, luteolin and luteolin-7-O-glucoside. The flavones luteolin and luteolin-7-O-glucoside had inhibitory effects at 100 µg/mL level on HepG2 cells, with luteolin being a significant stronger inhibitor compared to the respective glucoside. Artichoke leaf extract had minor stimulating effect on gene expression of CYP1A2, while CYP3A4, GGT, GPX2, GSR and GST were slightly inhibited. GST inhibition under in vitro conditions was also shown by quantification of GST enzyme activity. Induction of gene expression of CYP1A2 was shown to be supraadditive after simultaneous application of ethanol plus artichoke extract. Artichoke leaf extract exhibited cell protective effects against ethanol-induced toxicity within cotreatment under in vitro conditions. Also H2O2 damage was significantly inhibited by simultaneous artichoke incubation. Pre- and posttreatments did not exert protective effects. DMSO-induced toxicity was significantly reduced by pre-, post- and cotreatment with artichoke extract and especially with luteolin-7-O-glucoside, indicating a direct

  5. In-depth investigation of spin-on doped solar cells with thermally grown oxide passivation

    Ahmad, Samir Mahmmod; Cheow, Siu Leong; Ludin, Norasikin A.; Sopian, K.; Zaidi, Saleem H.

    Solar cell industrial manufacturing, based largely on proven semiconductor processing technologies supported by significant advancements in automation, has reached a plateau in terms of cost and efficiency. However, solar cell manufacturing cost (dollar/watt) is still substantially higher than fossil fuels. The route to lowering cost may not lie with continuing automation and economies of scale. Alternate fabrication processes with lower cost and environmental-sustainability coupled with self-reliance, simplicity, and affordability may lead to price compatibility with carbon-based fuels. In this paper, a custom-designed formulation of phosphoric acid has been investigated, for n-type doping in p-type substrates, as a function of concentration and drive-in temperature. For post-diffusion surface passivation and anti-reflection, thermally-grown oxide films in 50-150-nm thickness were grown. These fabrication methods facilitate process simplicity, reduced costs, and environmental sustainability by elimination of poisonous chemicals and toxic gases (POCl3, SiH4, NH3). Simultaneous fire-through contact formation process based on screen-printed front surface Ag and back surface through thermally grown oxide films was optimized as a function of the peak temperature in conveyor belt furnace. Highest efficiency solar cells fabricated exhibited efficiency of ∼13%. Analysis of results based on internal quantum efficiency and minority carried measurements reveals three contributing factors: high front surface recombination, low minority carrier lifetime, and higher reflection. Solar cell simulations based on PC1D showed that, with improved passivation, lower reflection, and high lifetimes, efficiency can be enhanced to match with commercially-produced PECVD SiN-coated solar cells.

  6. Investigation of retinal ganglion cells and axons of normal rats using fluorogold retrograde labeling

    Yin Xiaolei; Ye Jian; Chen Chunlin

    2006-01-01

    To investigate the retinal ganglion cells (RGCs) by means of fluorogold retrograde labeling, RGCs were labeled by injecting the fluorogold bilaterally into the superficial superior colliculus and lateral genicutate nucleus in six adult SD rats. One and two weeks (3 rats in each group) after injecting the fluorogold, RGCs FG-labeled were observed and the number of them were counted. The results showed that after a week mean density of fluorogold-labeled RGCs was 2210 ± 128/mm 2 , and it was 2164 ± 117/mm 2 after two weeks. Our conclusion is fluorogold retrograde labeling could be very useful in the research of RGCs. (authors)

  7. Thermodynamic Investigation of an Integrated Gasification Plant with Solid Oxide Fuel Cell and Steam Cycles

    Rokni, Masoud

    2012-01-01

    A gasification plant is integrated on the top of a solid oxide fuel cell (SOFC) cycle, while a steam turbine (ST) cycle is used as a bottoming cycle for the SOFC plant. The gasification plant was fueled by woodchips to produce biogas and the SOFC stacks were fired with biogas. The produced gas...... generator (HRSG). The steam cycle was modeled with a simple single pressure level. In addition, a hybrid recuperator was used to recover more energy from the HRSG and send it back to the SOFC cycle. Thus two different configurations were investigated to study the plants characteristic. Such system...

  8. Establishment of fruit bat cells (Rousettus aegyptiacus as a model system for the investigation of filoviral infection.

    Verena Krähling

    Full Text Available BACKGROUND: The fruit bat species Rousettus aegyptiacus was identified as a potential reservoir for the highly pathogenic filovirus Marburg virus. To establish a basis for a molecular understanding of the biology of filoviruses in the reservoir host, we have adapted a set of molecular tools for investigation of filovirus replication in a recently developed cell line, R06E, derived from the species Rousettus aegyptiacus. METHODOLOGY/PRINCIPAL FINDINGS: Upon infection with Ebola or Marburg viruses, R06E cells produced viral titers comparable to VeroE6 cells, as shown by TCID(50 analysis. Electron microscopic analysis of infected cells revealed morphological signs of filovirus infection as described for human- and monkey-derived cell lines. Using R06E cells, we detected an unusually high amount of intracellular viral proteins, which correlated with the accumulation of high numbers of filoviral nucleocapsids in the cytoplasm. We established protocols to produce Marburg infectious virus-like particles from R06E cells, which were then used to infect naïve target cells to investigate primary transcription. This was not possible with other cell lines previously tested. Moreover, we established protocols to reliably rescue recombinant Marburg viruses from R06E cells. CONCLUSION/SIGNIFICANCE: These data indicated that R06E cells are highly suitable to investigate the biology of filoviruses in cells derived from their presumed reservoir.

  9. Experimentally and numerically investigating cell performance and localized characteristics for a high-temperature proton exchange membrane fuel cell

    Su Ay; Ferng, Yuh Ming; Shih, Jah Ching

    2009-01-01

    This paper is to experimentally and numerically investigate the cell performance and the localized characteristics associated with a high-temperature proton exchange membrane fuel cell (PEMFC). Three experiments are carried out in order to study the performance of the PEMFC with different operating conditions and to validate the numerical simulation model. The model proposed herein is a three-dimensional (3-D) computational fluid dynamics (CFD) non-isothermal model that essentially consists of thermal-hydraulic equations and electrochemical model. The performance curves of the PEMFC predicted by the present model agree with the experimental measured data. In addition, both the experiments and the predictions precisely demonstrate the enhanced effects of inlet gas temperature and system pressure on the PEMFC performance. Based on the simulation results, the localized characteristics within a PEMFC can be reasonably captured. These parameters include the fuel gas distribution, liquid water saturation distribution, membrane conductivity distribution, temperature variation, and current density distribution etc. As the PEMFC is operated at the higher current density, the fuel gas would be insufficiently supplied to the catalyst layer, consequently causing the decline in the generation of power density. This phenomenon is so called mass transfer limitation, which can be precisely simulated by the present CFD model.

  10. Optical properties of photoreceptor and retinal pigment epithelium cells investigated with adaptive optics optical coherence tomography

    Liu, Zhuolin

    Human vision starts when photoreceptors collect and respond to light. Photoreceptors do not function in isolation though, but share close interdependence with neighboring photoreceptors and underlying retinal pigment epithelium (RPE) cells. These cellular interactions are essential for normal function of the photoreceptor-RPE complex, but methods to assess these in the living human eye are limited. One approach that has gained increased promise is high-resolution retinal imaging that has undergone tremendous technological advances over the last two decades to probe the living retina at the cellular level. Pivotal in these advances has been adaptive optics (AO) and optical coherence tomography (OCT) that together allow unprecedented spatial resolution of retinal structures in all three dimensions. Using these high-resolution systems, cone photoreceptor are now routinely imaged in healthy and diseased retina enabling fundamental structural properties of cones to be studied such as cell spacing, packing arrangement, and alignment. Other important cell properties, however, have remained elusive to investigation as even better imaging performance is required and thus has resulted in an incomplete understanding of how cells in the photoreceptor-RPE complex interact with light. To address this technical bottleneck, we expanded the imaging capability of AO-OCT to detect and quantify more accurately and completely the optical properties of cone photoreceptor and RPE cells at the cellular level in the living human retina. The first objective of this thesis was development of a new AO-OCT method that is more precise and sensitive, thus enabling a more detailed view of the 3D optical signature of the photoreceptor-RPE complex than was previously possible (Chapter 2). Using this new system, the second objective was quantifying the waveguide properties of individual cone photoreceptor inner and outer segments across the macula (Chapter 3). The third objective extended the AO

  11. Investigation of Some Transparent Metal Oxides as Damp Heat Protective Coating for CIGS Solar Cells: Preprint

    Pern, F. J.; Yan, F.; Zaaunbrecher, B.; To, B.; Perkins, J.; Noufi, R.

    2012-10-01

    We investigated the protective effectiveness of some transparent metal oxides (TMO) on CIGS solar cell coupons against damp heat (DH) exposure at 85oC and 85% relative humidity (RH). Sputter-deposited bilayer ZnO (BZO) with up to 0.5-um Al-doped ZnO (AZO) layer and 0.2-um bilayer InZnO were used as 'inherent' part of device structure on CdS/CIGS/Mo/SLG. Sputter-deposited 0.2-um ZnSnO and atomic layer deposited (ALD) 0.1-um Al2O3 were used as overcoat on typical BZO/CdS/CIGS/Mo/SLG solar cells. The results were all negative -- all TMO-coated CIGS cells exhibited substantial degradation in DH. Combining the optical photographs, PL and EL imaging, SEM surface micro-morphology, coupled with XRD, I-V and QE measurements, the causes of the device degradations are attributed to hydrolytic corrosion, flaking, micro-cracking, and delamination induced by the DH moisture. Mechanical stress and decrease in crystallinity (grain size effect) could be additional degrading factors for thicker AZO grown on CdS/CIGS.

  12. Investigations at INRIM on a Pd-C cell manufactured by NPL

    Battuello, M.; Florio, M.; Machin, G.

    2011-10-01

    One of a set of metal-carbon eutectic cells (a Pd-C cell, 1765 K) manufactured by NPL and used for a previous comparison of temperature scales with NIST has been investigated at INRIM. There it was implemented in two different furnaces, namely a single- and a three-zone, and measured with a standard radiation thermometer operating at 900 nm and 950 nm. Both ITS-90 and thermodynamic melting temperatures of the cell were determined by means of an extrapolation approach. The thermodynamic temperature differs by only -0.31 K from the NIST value whereas the ITS-90 temperature differs by only -0.46 K from the NPL value. The agreements, within the combined expanded uncertainties, are particularly significant, because of the different approach followed at INRIM, namely the extrapolation of multi-fixed-point scales (n = 3 and n = 4), as compared with a direct radiometric method at NIST and an ITS-90 realization traceable to the gold point at NPL.

  13. A numerical investigation of the effects of membrane swelling in polymer electrolyte fuel cells

    Tiss, Faysal; Chouikh, Ridha; Guizani, Amenallah

    2013-01-01

    Highlights: ► Membrane water content is controlled by the operating conditions in the cathode. ► When the membrane is in contact with water, only pore size varies. ► Membrane water content increase by increasing the functioning temperature. ► Good agreement between computational results and previous reported experimental data. - Abstract: A two-dimensional computational fluid dynamics model of PEM fuel cell is developed by taking into account the electrochemical, mass and heat transfer process occurring in the cathode compartment. Additionally, this model includes the effect of water content in the membrane swelling phenomenon. Several parameters such as gases temperature, inlet velocity and membrane characteristics are too investigated to establish their effect on the PEM fuel cell performance. The membrane water content and the air fraction variation in the gas channel are examined for diverse values of Reynolds number. In particular, the desirable inlet flow for enhancing the performance of the PEM fuel cell is determined by examining membrane water content patterns. The methodology in this study is useful to the control of water management and gas diffusion layer design

  14. Transient photoluminescence decay investigations of LPE GaAs heteroface solar cells

    Wettling, W.; Ehrhardt, A.; Brett, A.; Lutz, F.

    1990-01-01

    The transient photoluminescence decay (PLD) is investigated as a technique for the quality control of GaAs solar cells. An analytic expression for the PL intensity is derived from the time dependent continuity equation for minority carrier concentration in the emitter by the Fourier transform method. On both sides of the emitter, i.e. at the interface to the window layer and to the space charge region, surface recombination velocities that can vary between 0 and ∞ are allowed as boundary conditions. Experiments were performed using a mode-locked and cavity dumped laser as excitation source and an optical sampling oscilloscope as detector for the transient PL. PLD from GaAs wafers and solar cells was measured with time resolution of down to 20 ps for various intensities of laser excitation and (for the cells) under open-circuit and short-circuit condition. The results are discussed in respect to the theory together with a model of local internal boundary conditions at the junction near the exciting laser beam

  15. Investigation of venous blood cells parameters among 1180 healthy people in Tianjin area

    Jiang Liping; Hao Jianxiu; Li Jin; Xing Zhiwei; Zhao Xinran; Jiang Bo; Wang Xiaoguang; Jiang Enhai

    2013-01-01

    Objective: To investigate the reference values of 18 parameters of venous blood cells among healthy adults in Tianjin area. Methods: The values of 18 parameters of venous blood from 1180 healthy adults in Tianjin area were measured by Sysmex KX-21 hematology analyzer and the results were analyzed. Results: The statistical analysis of the test reveals that significant differences exist in most parameters of venous blood cells according to the gender and age of people. Except the parameters of mean corpuscular volume, lymphocytes percentage, mean platelet volume, platelet distribution width,there were significant differences in the remaining parameters between the males group and the females group. Except the parameters of white blood count,platelet count, mean corpuscular hemoglobin, neutrophil percentage,absolute neutrophil count, mean platelet volume, platelet distribution width, there were significant differences in the remaining parameters between the old male group and the adult male group. Except the parameters of white blood count, mean corpuscular volume, mean corpuscular hemoglobin, there was no significant difference in the remaining parameters between the old female group and the adult female group. Conclusions: There are some differences between the findings and the reference range provided by the National Guide to Clinical laboratory Procedure. Therefore, it is necessary for laboratory to establish the reference values of venous blood cells according to concrete conditions. (authors)

  16. Comparative proteomic investigation of metastatic and non-metastatic osteosarcoma cells of human and canine origin.

    Jahnabi Roy

    Full Text Available Osteosarcoma is the most common bone cancer in dogs and people. In order to improve clinical outcomes, it is necessary to identify proteins that are differentially expressed by metastatic cells. Membrane bound proteins are responsible for multiple pro-metastatic functions. Therefore characterizing the differential expression of membranous proteins between metastatic and non-metastatic clonal variants will allow the discovery of druggable targets and consequently improve treatment methodology. The objective of this investigation was to systemically identify the membrane-associated proteomics of metastatic and non-metastatic variants of human and canine origin. Two clonal variants of divergent in vivo metastatic potential from human and canine origins were used. The plasma membranes were isolated and peptide fingerprinting was used to identify differentially expressed proteins. Selected proteins were further validated using western blotting, flow cytometry, confocal microscopy and immunohistochemistry. Over 500 proteins were identified for each cell line with nearly 40% of the proteins differentially regulated. Conserved between both species, metastatic variants demonstrated significant differences in expression of membrane proteins that are responsible for pro-metastatic functions. Additionally, CD147, CD44 and vimentin were validated using various biochemical techniques. Taken together, through a comparative proteomic approach we have identified several differentially expressed cell membrane proteins that will help in the development of future therapeutics.

  17. Comparative proteomic investigation of metastatic and non-metastatic osteosarcoma cells of human and canine origin.

    Roy, Jahnabi; Wycislo, Kathryn L; Pondenis, Holly; Fan, Timothy M; Das, Aditi

    2017-01-01

    Osteosarcoma is the most common bone cancer in dogs and people. In order to improve clinical outcomes, it is necessary to identify proteins that are differentially expressed by metastatic cells. Membrane bound proteins are responsible for multiple pro-metastatic functions. Therefore characterizing the differential expression of membranous proteins between metastatic and non-metastatic clonal variants will allow the discovery of druggable targets and consequently improve treatment methodology. The objective of this investigation was to systemically identify the membrane-associated proteomics of metastatic and non-metastatic variants of human and canine origin. Two clonal variants of divergent in vivo metastatic potential from human and canine origins were used. The plasma membranes were isolated and peptide fingerprinting was used to identify differentially expressed proteins. Selected proteins were further validated using western blotting, flow cytometry, confocal microscopy and immunohistochemistry. Over 500 proteins were identified for each cell line with nearly 40% of the proteins differentially regulated. Conserved between both species, metastatic variants demonstrated significant differences in expression of membrane proteins that are responsible for pro-metastatic functions. Additionally, CD147, CD44 and vimentin were validated using various biochemical techniques. Taken together, through a comparative proteomic approach we have identified several differentially expressed cell membrane proteins that will help in the development of future therapeutics.

  18. Investigation of freeze/thaw durability in polymer electrolyte fuel cells

    Lim, Soo-Jin; Park, Gu-Gon; Sohn, Young-Jun; Yim, Sung-Dae; Yang, Tae-Hyun; Kim, Chang-Soo [Fuel Cell Research Center, Korea Institute of Energy Research, 102, Gajeong-ro, Yuseong-gu, Daejeon 305-343 (Korea, Republic of); Park, Jin-Soo [Department of Environmental Engineering, College of Engineering, Sanmyung University, 300 Anseo-dong, Dongnam-gu, Cheonam, Chungnam Province 330-720 (Korea, Republic of); Hong, Bo Ki [Fuel Cell Vehicle Team 1, Ecotechnology Center, Hyundai-Kia Motors Company, 104, Mabuk-dong, Giheung-gu, Yongin-si, Gyeonggi-do 446-912 (Korea, Republic of)

    2010-12-15

    This study aims to investigate the effect of different gas diffusion layers (GDLs) on freeze/thaw condition durability in polymer electrolyte fuel cells (PEFCs). Three kinds of GDLs-cloth, felt and paper type - with similar basic properties except thickness and bending stiffness were used. The changes in the properties and cell performance were investigated from the -30 to 70 C range of freeze/thaw cycles. The I-V performance degradation was observed to be negligible for the felt GDL whereas the cloth and paper GDLs showed a marked I-V performance loss. No distinctive correlation between the changes in electrochemical properties, such as active metal surface area, hydrogen crossover rates and decreased I-V performance, was observed except an increase in ohmic resistance revealed by ac-impedance spectroscopy. The physical destruction of electrodes was also shown by scanning electron microscope (SEM) analysis. The present study found that sufficient mechanical supporting force between the interfaces of materials enhances PEFC durability in sub-zero temperature conditions. (author)

  19. Investigation of flurbiprofen genotoxicity and cytotoxicity in rat bone marrow cells.

    Timocin, Taygun; Ila, Hasan B

    2015-01-01

    This study was performed to investigate cytogenetic effects of NSAID flurbiprofen which was used as active ingredient in some analgesic, antipyretic and anti-inflammatory drugs. Genotoxic effect of flurbiprofen was investigated using in vivo chromosome aberration (CA) test and random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) test. Also, oxidative stress potential of flurbiprofen was determined by measuring total oxidant and antioxidant level which occurred with flurbiprofen treatment in rat peripheral blood. For these purposes, rats were treated with three concentrations of flurbiprofen (29.25, 58.50 and 117 mg/kg, body weight) in single dose at two different treatment periods (12 and 24 h). According to the results, flurbiprofen did not affect chromosome aberrations in rat bone marrow cells with CA test. In RAPD-PCR test, polymorphic bands were unaffected. Also, test substance did not change total oxidant and antioxidant status (except for 58.50 and 117 mg/kg, 12 h) and therefore it did not lead to significant increase on oxidative stress (again except 58.50 and 117 mg/kg, 12 h). However, flurbiprofen reduced to mitotic indexes and these reductions were dose-dependent for 12 h treatment. In summary, flurbiprofen did not show significant genotoxic effect. But it caused cytotoxicity in rat bone marrow cells.

  20. Large prospective investigation of meat intake, related mutagens, and risk of renal cell carcinoma.

    Daniel, Carrie R; Cross, Amanda J; Graubard, Barry I; Park, Yikyung; Ward, Mary H; Rothman, Nathaniel; Hollenbeck, Albert R; Chow, Wong-Ho; Sinha, Rashmi

    2012-01-01

    The evidence for meat intake and renal cell carcinoma (RCC) risk is inconsistent. Mutagens related to meat cooking and processing, and variation by RCC subtype may be important to consider. In a large US cohort, we prospectively investigated intake of meat and meat-related compounds in relation to risk of RCC, as well as clear cell and papillary RCC histologic subtypes. Study participants (492,186) completed a detailed dietary assessment linked to a database of heme iron, heterocyclic amines (HCA), polycyclic aromatic hydrocarbons (PAHs), nitrate, and nitrite concentrations in cooked and processed meats. Over 9 (mean) y of follow-up, we identified 1814 cases of RCC (498 clear cell and 115 papillary adenocarcinomas). HRs and 95% CIs were estimated within quintiles by using multivariable Cox proportional hazards regression. Red meat intake [62.7 g (quintile 5) compared with 9.8 g (quintile 1) per 1000 kcal (median)] was associated with a tendency toward an increased risk of RCC [HR: 1.19; 95% CI: 1.01, 1.40; P-trend = 0.06] and a 2-fold increased risk of papillary RCC [P-trend = 0.002]. Intakes of benzo(a)pyrene (BaP), a marker of PAHs, and 2-amino-1-methyl-6-phenyl-imidazo[4,5-b]pyridine (PhIP), an HCA, were associated with a significant 20-30% elevated risk of RCC and a 2-fold increased risk of papillary RCC. No associations were observed for the clear cell subtype. Red meat intake may increase the risk of RCC through mechanisms related to the cooking compounds BaP and PhIP. Our findings for RCC appeared to be driven by strong associations with the rarer papillary histologic variant. This study is registered at clinicaltrials.gov as NCT00340015.

  1. Investigation of test methods, material properties, and processes for solar cell encapsulants. Annual report

    Willis, P. B.; Baum, B.

    1979-06-01

    The goal of this program is to identify, evaluate, and recommend encapsulant materials and processes for the production of cost-effective, long-life solar cell modules. During the past year, the technical activities emphasized the reformulation of a commercial grade of ethylene/vinyl acetate copolymer for use as a pottant in solar cell module manufacture. After experimenting with a variety of techniques, a vacuum-bag process was developed and found to be an excellent encapsulation method. Adhesive strengths and primers for the bonding of ethylene/vinyl acetate to superstrate and substrate materials was assessed with encouraging results. The weathering effects on ten other polymers exposed to twelve months of weathering in Arizona, Florida, and under EMMAQUA were evaluated by determination of tensile strengths, elongations, optical transmission, etc. As may be expected, the best overall retention of mechanical properties is found for the fluorocarbon polymers, especially FEP. Hard coatings containing ultraviolet absorbers were investigated for the purpose of providing a soil resistant surface and additional weathering stability to the soft EVA pottant. Corrosion studies using a standard salt spray test were used to determine the degree of protection offered to a variety of metals by encapsulation in EVA pottant. A survey of scrim materials was also conducted. These open hole weaves are intended for use as spacers between the cell and substrate to provide a mechanical barrier, improve insulation resistance and prevent migration of the pigmented pottant over the cell surface. A mechanical engineering analysis of composite structural materials for use as substrates was performed. Results are presented in detail. (WHK)

  2. Water transport in gas diffusion media for PEM fuel cells. Experimental and numerical investigation

    Roth, Joerg

    2010-08-20

    The water flux in partially saturated hydrophobic carbon fibre paper for polymer electrolyte membrane fuel cell applications is investigated and compared with the frequently used constitutive two-phase flow model based on Darcy's law. Further, the first steps towards a math-based material design for gas diffusion media are explored in this thesis. Two self-developed ex-situ experiments to investigate the liquid water transport are introduced. The first is a newly developed buoyancy-based measurement of the pressuresaturation relationship on thin porous material with an accuracy of 0.5 kPa for the pressure and {+-} 5% for the saturation. The second experiment measures the pressure drop in dependence of flow rates down to magnitudes of {mu}L/s across the partially saturated thin porous material. This flow rate is relevant for the fuel cell application. The liquid water transport through Toray 060 carbon fibre paper, impregnated with 7% and 10% PTFE is investigated at wet and dry boundary conditions. The experiments are also accompanied by analytical and numerical free surface modelling with the consideration of the material morphology and liquid-solid interaction. The imbibing and draining cases of an arrangement of six fibres at varying solid-liquid interaction and boundary conditions are studied with 'Surface Evolver'. In order to evaluate the findings of ex-situ and modelling work for applicability to water transport in fuel cell operation, the technique of nuclear magnetic resonance (NMR) imaging is assessed. The focus is on the visualisation of 2D and 3D water distribution in the operating fuel cell. The compatibility of the NMR experiment with fuel cell operation in relation to material selection, operating temperature, and current density is addressed. NMR imaging is employed for different current densities, stoichiometries, and fuel cell arrangements. The fuel cell arrangements differ by the cathode diffusion medium. Plain, hydrophobic, and

  3. Investigations into the molecular mechanism of chromatid breakage in the G2-phase of mammalian cells

    Bryant, P.E.; Armstrong, G.N.; Gray, L.; Frankenberg, D.; Mozdarani, H.

    2003-01-01

    Chromatid breakage following irradiation of cells in the G2-phase of the cell cycle results from the induction of DNA double-strand breaks (dsb). The conversion of dsb into chromatid breaks (cb) has a genetic basis, seemingly different from that of dsb rejoining. The variation in extent of this conversion is exemplified by the stiking variation in frequency of cb in irradiated cycling T-lymphocytes between different normal individuals. Elevated cb frequency in lymphocytes of around 40% of breast cancer patients and their first-degree relatives suggests the presence of mutations in low penetrance cancer predisposing genes that also affect conversion of dsb to cb. Investigation of the mechanism of chromatid radiosensitivity using genetically engineered rodent cell lines containing unique dsb break sites indicate that a single isolated dsb is sufficient to cause a cb. The single-event nature of chromatid breakage is confirmed by the fact that cb are induced as a linear function of radiation dose. Moreover, we have recently shown that ultrasoft carbon-K X-rays also induce chromatid breakage. In this case the energy of the secondary electrons produced by carbon-K X-rays is too low to span more than one DNA double helix, thus further supporting our conclusion that a single dsb is responsible for the formation of a cb. Chromatid breakage is thought to involve a rearrangement between DNA strands at the crossover points of chromatin loop(s) triggered by the presence of a dsb within the loop structure. The occasional observation of 'looped-out' sections of chromatin at cb sites supports this hypothesis. The occurrence of 'colour-switches' between FPG stained chromatids at a proportion of break sites (e.g. about 16% in CHO cells) shows that a significant proportion of cb definitely result from chromatin rearrangements. Measurements of altered colour-switch ratio (csr) in mutant rodent and human cells (irs1 and AT cells respectively) also indicate a genetic basis for the

  4. A microPIXE investigation of the interaction of cells of Schizosaccharomyces pombe with the culture medium

    Rombouts, P.M.M. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Gomez-Morilla, I. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Grime, G.W. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Webb, R.P. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Cuenca, L. [Fluids and Systems Research Centre, School of Engineering, University of Surrey, Guildford GU2 7XH (United Kingdom); Rodriguez, R. [Fluids and Systems Research Centre, School of Engineering, University of Surrey, Guildford GU2 7XH (United Kingdom); Browton, M. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Wardell, N. [School of Biomedical and Molecular Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom); Underwood, B. [Fluids and Systems Research Centre, School of Engineering, University of Surrey, Guildford GU2 7XH (United Kingdom); Kirkby, N.F. [Fluids and Systems Research Centre, School of Engineering, University of Surrey, Guildford GU2 7XH (United Kingdom); Kirkby, K.J. [Ion Beam Centre, Advanced Technology Institute, School of Electronics and Physical Sciences, University of Surrey, Guildford GU2 7XH (United Kingdom)]. E-mail: k.kirkby@surrey.ac.uk

    2007-07-15

    Schizosaccharomyces pombe (S. pombe) is a eucaryotic cell type similar to mammalian cells but much more simple. As it also executes its cell cycle rapidly it is very useful for investigating basic processes in cells. In this paper we report a feasibility study of the applicability of microPIXE to investigate the interaction between S. pombe cells and the surrounding culture medium. Cells were cultured in various growth medium prior to preparation for analysis. 1 {mu}l drops of medium and cells were spotted onto polypropylene foils held in contact with a polished copper block previously cooled in liquid nitrogen. The samples were dehydrated by freeze-drying. Micro PIXE analysis was carried out with the IBC microbeam facility using a beam of 2.5 MeV protons focused to 1-2 {mu}m diameter. Initially no elemental contrast was observed between the cells and the medium, but by modifying the dilution of the cell suspension, the cells could be distinguished from the surrounding medium through an increased concentration of P and reduced concentration of Cl. The distribution of Na in the medium around the cells showed evidence of the action of the Na pump. Sporulation appears to be induced in the cells by adding Cu to the growth medium and the uptake of Cu by the cells could be clearly observed. This study shows that it is possible to analyse the mass transport of elements in and out of cells In the future this will enable concentration gradients to be analysed and allow the rate of production or consumption of individual cells to be calculated. By observing these patterns for individual cells (not populations) at various known points in the cell cycle, fundamental data can be derived.

  5. A microPIXE investigation of the interaction of cells of Schizosaccharomyces pombe with the culture medium

    Rombouts, P.M.M.; Gomez-Morilla, I.; Grime, G.W.; Webb, R.P.; Cuenca, L.; Rodriguez, R.; Browton, M.; Wardell, N.; Underwood, B.; Kirkby, N.F.; Kirkby, K.J.

    2007-01-01

    Schizosaccharomyces pombe (S. pombe) is a eucaryotic cell type similar to mammalian cells but much more simple. As it also executes its cell cycle rapidly it is very useful for investigating basic processes in cells. In this paper we report a feasibility study of the applicability of microPIXE to investigate the interaction between S. pombe cells and the surrounding culture medium. Cells were cultured in various growth medium prior to preparation for analysis. 1 μl drops of medium and cells were spotted onto polypropylene foils held in contact with a polished copper block previously cooled in liquid nitrogen. The samples were dehydrated by freeze-drying. Micro PIXE analysis was carried out with the IBC microbeam facility using a beam of 2.5 MeV protons focused to 1-2 μm diameter. Initially no elemental contrast was observed between the cells and the medium, but by modifying the dilution of the cell suspension, the cells could be distinguished from the surrounding medium through an increased concentration of P and reduced concentration of Cl. The distribution of Na in the medium around the cells showed evidence of the action of the Na pump. Sporulation appears to be induced in the cells by adding Cu to the growth medium and the uptake of Cu by the cells could be clearly observed. This study shows that it is possible to analyse the mass transport of elements in and out of cells In the future this will enable concentration gradients to be analysed and allow the rate of production or consumption of individual cells to be calculated. By observing these patterns for individual cells (not populations) at various known points in the cell cycle, fundamental data can be derived

  6. Long-term HPV type-specific risks for ASCUS and LSIL: a 14-year follow-up of a randomized primary HPV screening trial.

    Elfström, K Miriam; Smelov, Vitaly; Johansson, Anna L V; Eklund, Carina; Naucler, Pontus; Arnheim-Dahlström, Lisen; Dillner, Joakim

    2015-01-15

    Human papillomavirus (HPV) infections result in a significant burden of low-grade cervical lesions. Between 1997 and 2000, our randomized trial of primary HPV screening enrolled 12,527 women participating in population-based screening. Women between 32 and 38 years of age (median: 34, interquartile range: 33-37) were randomized to HPV and cytology double testing (intervention arm, n = 6,257 enrolled, n = 5,888 followed-up) or to cytology, with samples frozen for future HPV testing (control arm, n = 6,270 enrolled, n = 5,795 followed-up). We estimated the HPV type-specific, long-term absolute risks (AR), and population attributable proportions (PAR) for cytological diagnoses of atypical squamous cells of undetermined significance (ASCUS) or low-grade squamous intraepithelial lesion (LSIL) and for histopathologically diagnosed cervical intraepithelial neoplasia grade 1 (CIN1). The women were followed using comprehensive, nationwide register-based follow-up. During a mean follow-up time of 11.07 years, 886 ASCUS and LSIL lesions were detected, 448 in the intervention arm and 438 in the control arm. Poisson regression estimated the incidence rate ratios (IRRs) of low-grade lesions by HPV type. The IRRs were strongly dependent on follow-up time. The IRRs for ASCUS/LSIL associated with high-risk HPV positivity were 18.6 (95% CI: 14.9-23.4) during the first screening round, 4.1 (95% CI: 2.8-6.2) during the second, 2.6 (95% CI: 1.7-4.1) during the third, and 1.1 (95% CI: 0.7-1.8) for >9 years of follow-up, with similar declines seen for the individual types. Type 16 contributed consistently to the greatest proportion of ASCUS, LSIL, and CIN1 risk in the population (first screening round PAR: ASCUS: 15.5% (95% CI: 9.7-21.9), LSIL: 14.7% (95% CI: 8.0-20.9), and CIN1: 13.4% (95% CI: 3.2-22.5)), followed by type 31 [8.4% (95% CI: 4.2-12.5) for ASCUS to 17.3% (95% CI: 6.8-26.6) for CIN1]. In summary, most ASCUS/LSIL lesions associated with HPV infection are caused by new HPV

  7. Experimental investigation of solid oxide fuel cells using biomass gasification producer gases

    Norheim, Arnstein

    2005-07-01

    The main objective of this thesis is theoretical and experimental investigations related to utilisation of biomass gasification producer gases as fuel for Solid Oxide Fuel Cells (SOFC). Initial fundamental steps towards a future system of combined heat and power production based on biomass gasification and SOFC are performed and include: 1) Theoretical modeling of the composition of biomass gasification producer gases. 2) Experimental investigation of SOFC performance using biomass gasification producer gas as fuel. 3) Experimental investigation of SOFC performance using biomass gasification producer gas containing high sulphur concentration. The modeling of the composition of gasifier producer gas was performed using the program FactSage. The main objective was to investigate the amount and speciation of trace species in the producer gases as several parameters were varied. Thus, the composition at thermodynamic equilibrium of sulphur, chlorine, potassium, sodium and compounds of these were established. This was done for varying content of the trace species in the biomass material at different temperatures and fuel utilisation i.e. varying oxygen content in the producer gas. The temperature interval investigated was in the range of normal SOFC operation. It was found that sulphur is expected to be found as H2S irrespective of temperature and amount of sulphur. Only at very high fuel utilisation some S02 is formed. Important potassium containing compounds in the gas are gaseous KOH and K. When chlorine is present, the amount of KOH and K will decrease due to the formation of KCI. The level of sodium investigated here was low, but some Na, NaOH and NaCl is expected to be formed. Below a certain temperature, condensation of alkali rich carbonates may occur. The temperature at which condensation begins is mainly depending on the amount of potassium present; the condensation temperature increases with increasing potassium content. In the first experimental work

  8. [Investigation of emergency capacities for occupational hazard accidents in silicon solar cell producing enterprises].

    Yang, D D; Xu, J N; Zhu, B L

    2016-11-20

    Objective: To investigate and analyze the influential factors of occupational hazard acci-dents, emergency facilities and emergency management in Silicon solar cell producing enterprises, then to pro-vide scientific strategies. Methods: The methods of occupationally healthy field investigating, inspecting of ven-tilation effectiveness, setup of emergency program and wearing chemical suit were used. Results: The mainly occupational hazard accidents factors in the process of Silicon solar cell producing included poisoning chemi-cals, high temperature, onizing radiation and some workplaces. The poisoning chemicals included nitric acid, hydrofluoric acid, sulfuric acid, hydrochloric acid, sodium hydroxide, potassium hydroxide, chlorine, phos-phorus oxychloride, phosphorus pentoxide, nitrogen dioxide, ammonia, silane, and so on; the workplaces in-cluded the area of producing battery slides and auxiliary producing area. Among the nine enterprises, gas detec-tors were installed in special gas supplying stations and sites, but the height, location and alarmvalues of gas detectors in six enterprises were not according with standard criteria; emergency shower and eyewash equip-ment were installed in workplaces with strong corrosive chemicals, but the issues of waste water were not solved; ventilation systems were set in the workplaces with ammonia and silane, but not qualified with part lo-cations and parameters in two enterprises; warehouses with materials of acid, alkali, chemical ammonia and phosphorus oxychloride were equipped with positive - pressure air respirator resuscitator and emergency cabi-nets, but with insufficient quantity in seven enterprises and expiration in part of products. The error rate of set-up emergency program and wearing chemical cloth were 30%~100% and 10%~30%, respectively. Among the nine enterprises, there were emergency rescue plans for dangerous chemical accidents, but without profession-al heatstroke and irradiation accident emergency plans

  9. Longitudinal investigation of natural killer cells and cytokines in chronic fatigue syndrome/myalgic encephalomyelitis

    Brenu Ekua W

    2012-05-01

    Full Text Available Abstract Background Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME is an etiologically unexplained disorder characterised by irregularities in various aspects of the immunological function. Presently, it is unknown whether these immunological changes remain consistent over time. This study investigates Natural Killer (NK cell cytotoxic activity, NK cell subsets (CD56brightCD16- and CD56dimCD16+ and cytokines, over the course of a12 month period in patients with CFS/ME. Methods The participants in the study comprised 65 (47.2 ± 11.5 years CFS/ME participants and 21 (45.2 ±9.3 years non-fatigued controls. Flow cytometry protocols were used to assess NK subsets and NK cytotoxic activity at various time points that included baseline (T1, 6 (T2 and 12 months (T3. Cytokine secretions were measured following mitogenic stimulation of peripheral blood mononuclear cells. Results NK cytotoxic activity was significantly decreased in the CFS/ME patients at T1, T2 and T3 compared to the non-fatigued group. Additionally, in comparison to the non-fatigued controls, the CFS/ME group had significantly lower numbers of CD56brightCD16- NK cells at both T1 and T2. Interestingly, following mitogenic stimulation, cytokine secretion revealed significant increases in IL-10, IFN-γ and TNF-α at T1 in the CFS/ME group. A significant decrease was observed at T2 in the CFS/ME group for IL-10 and IL-17A while at T3, IL-2 was increased in the CFS/ME group in comparison to the non-fatigued controls. Overall cytotoxic activity was significantly decreased at T3 compared to T1 and T2. CD56brightCD16- NK cells were much lower at T2 compared to T1 and T3. IL-10 and IL-17A secretion was elevated at T2 in comparison to T1 and T3. Conclusion These results confirm decreases in immune function in CFS/ME patients, suggesting an increased susceptibility to viral and other infections. Furthermore, NK cytotoxic activity may be a suitable biomarker for diagnosing CFS

  10. Hydrocortisone Increases the Vinblastine-Induced Chromosomal Damages in L929 Cells Investigated by the Micronucleus Assay on Cytokinesis-Blocked Binucleated Cells

    Tahere Ebrahimipour

    2017-03-01

    Full Text Available Background: Stress may cause damages to DNA or/and change the ability of the cells to overcome these damages. It may also cause irregularities in the cell cycle and induce abnormal cell divisions through glucocorticoid-dependent functions. The abnormal cell divisions, in turn, lead to chromosomal mal-segregation and aneuploidy. In this study, the effects of the stress hormone, hydrocortisone (HYD, were investigated on the induced chromosomal abnormalities by vinblastine (VIN during cell cycle in L929 cells. Methods: This work was performed in winter 2013 at Department of Biology, University of Ferdowsi, Mashhad, Iran. Cultured cells were divided into different groups including control, VIN-treated, HYD treated and VIN+HYD co-treated cells. The induced chromosomal damages were investigated by micronucleus assay in cytokinesis-blocked binucleated cells. Results: Although HYD by itself did not increase the micronuclei (Mn frequency, co-treatment of cells with VIN and HYD led to significant increase (P<0.05 in the frequency of Mn in comparison to control and VIN treated groups. Conclusion: Cells treated with stress hormone are more sensitive to damages induced by VIN. Therefore, stress may not directly result in genetic instability, it can increase the harmful effects associated with other genotoxic agents.

  11. Progenitor cells of erythroblasts: an in vitro investigation of erythropoietin-responsive cells of guinea pig bone marrow

    Rosse, C.; Beaufait, D.W.

    1978-01-01

    The experiments were designed to therst whether erythroblast progenitor cell function could be demonstrated in a morphological cell type designated as transitional cells. Two cell fractions were obtained from the bone marrow of normal and polycythemic guinea pigs. One fraction (F1) was enriched in transitional cells and contained few other cell types which could be considered as candidates for erythropoietin responsive cells (ERC). The other fraction (F2) contained undifferentiated blast cells as well as transitional cells. The effect of human urinary erythropoiesis stimulating factors (ESF) on heme synthesis was compared in these two fractions by measuring 59 Fe incorporation into heme. ESF was more effective in stimulating heme synthesis in guinea pig bone marrow cells than homologous sera obtained from anemic or hypoxic animals. The majority of ERC sedimented in F2, but the stimulation index was comparable in the two fractions. It was confirmed by radioautography that the ESF response in F1 was due to the generation of proerythroblasts and basophilic erythroblasts that incorporated 55 Fe. The generation of these cells in F1 was dependent on the addition of ESF to the cultures, whereas 55 Fe-labeled erythroblasts were recovered from cultures of F2 not supplemented with ESF. ESF induced a proportion of transitional cells to incorporate 55 Fe in both F1 and F2. Transitional cells were the only cell type in which heme synthesis was dependent on ESF. Radioautography with 55 Fe identified a proportion of these cells as ERC in both F1 and F2 fractions of bone marrow obtained from normal and polycythemic guinea pigs. The present studies show that some transitional cells function as progenitors of erythroblasts because they respond to ESF by initiation of heme synthesis and by transformation into the earliest recognizable erythroid cells

  12. An investigation into carbon nanostructured materials as catalyst support in proton exchange membrane fuel cells

    Veltzé, Sune

    acid treatment on the Vapour Grown Carbon Fibers™ manufactured by Showa Denko K. K. From these fibres, twelve platinised samples were investigated, of which one was platinised by a platinum phtalocyanine impregnation method, two were platinised by the polyol method and the remaining by the Bönnemann......Polymer electrolyte fuel cells (PEFCs) are among the key research areas concerning clean cost-effective energy. Carbon nano fibres (CNF), single walled carbon nano tubes (SWCNT), multi walled carbon nano tubes (MWCNT) and other related materials are among the possible successors to standard carbon...... black support materials for low platinum containing electrocatalyst. This is partly due to their high electronic conductivity. Partly due to their high surface area needed for the dispersion of nanoparticulate metal-clusters. In addition carbon nano-structures (CNF, SWCNT, MWCNT etc.) are more durable...

  13. Results of investigations in a fast reactor configuration with a strong heterogeneous cell structure

    Lehmann, E.; Albert, D.; Dietze, K.; Faehrmann, K.; Hansen, W.; Huettel, G.; Wand, H.; Osmera, B.

    1984-11-01

    Investigations of the neutron flux spectrum, of the energy dependent importance function and of sample reactivities were performed in a fast reactor configuration characterized by a marked neutronic microstructure which is produced by the insertion of pellets of polyethylene and cadmium. Contrary to results of calculations with a homogenized composition, values obtained by means of the tree programs CARMEN, YARAB and P1X, developed in Rossendorf, agree well with measurements. Furthermore it could be shown that an adjoint weighting for the determination of cell-averaged values of the importance function is justified. In a configuration with larger heterogeneity induced by the aggregation of four uranium pellets significant spectral differences between uranium and material zones, respectively, show up in activation measurements as well as in sample reactivity determinations. (author)

  14. Investigation of altenative carbon materials for fuel-cell catalyst support

    Larsen, Mikkel Juul

    In order to ensure high utilization of the catalyst material in a polymer electrolyte membrane fuel cell (PEMFC) it is usually fixed in the form of nanoparticles on a supporting material. The catalyst is platinum or a platinum alloy, and the commonly used support is carbon black (CB). Although...... structured carbon forms such as graphitized CBs, carbon nanotubes (CNTs), and carbon nanofibres (CNFs). This thesis concerns the investigation of an array of different materials which may prospec-tively replace the conventional materials used in the catalyst. The study comprised 13 carbon samples which...... nanotubes (GMWCNTs), and graphitized carbon nanofibre (CNF), while the Pt/C samples were platinized samples of some of the CNTs and CNFs (Pt/FWCNT, Pt/GMWCNT, and Pt/CNF, respectively) as well as two commercial Pt/CB reference catalysts. Comparative analyses have been performed in order to be able to assess...

  15. Experimental Investigation of a Direct Methanol Fuel Cell with Hilbert Fractal Current Collectors

    Jing-Yi Chang

    2014-01-01

    Full Text Available The Hilbert curve is a continuous type of fractal space-filling curve. This fractal curve visits every point in a square grid with a size of 2×2, 4×4, or any other power of two. This paper presents Hilbert fractal curve application to direct methanol fuel cell (DMFC current collectors. The current collectors are carved following first, second, and third order Hilbert fractal curves. These curves give the current collectors different free open ratios and opening perimeters. We conducted an experimental investigation into DMFC performance as a function of the free open ratio and opening perimeter on the bipolar plates. Nyquist plots of the bipolar plates are made and compared using electrochemical impedance spectroscopy (EIS experiments to understand the phenomena in depth. The results obtained in this paper could be a good reference for future current collector design.

  16. Investigation of metal/carbon-related materials for fuel cell applications by electronic structure calculations

    Kong, Ki-jeong [Korea Research Institute of Chemical Technology, P.O.Box 107, Yuseong, Daejeon 305-600 (Korea, Republic of)]. E-mail: kong@krict.re.kr; Choi, Youngmin [Korea Research Institute of Chemical Technology, P.O.Box 107, Yuseong, Daejeon 305-600 (Korea, Republic of); Ryu, Beyong-Hwan [Korea Research Institute of Chemical Technology, P.O.Box 107, Yuseong, Daejeon 305-600 (Korea, Republic of); Lee, Jeong-O [Korea Research Institute of Chemical Technology, P.O.Box 107, Yuseong, Daejeon 305-600 (Korea, Republic of); Chang, Hyunju [Korea Research Institute of Chemical Technology, P.O.Box 107, Yuseong, Daejeon 305-600 (Korea, Republic of)

    2006-07-15

    The potential of carbon-related materials, such as carbon nanotubes (CNTs) and graphite nanofibers (GNFs), supported metal catalysts as an electrode for fuel cell application was investigated using the first-principle electronic structure calculations. The stable binding geometries and energies of metal catalysts are determined on the CNT surface and the GNF edge. The catalyst metal is more tightly bound to the GNF edge than to the CNT surface because of the existence of active dangling bonds of edge carbon atoms. The diffusion barrier of metal atoms on the surface and edge is also obtained. From our calculation results, we have found that high dispersity is achievable for GNF due to high barrier against the diffusion of metal atoms, while CNT appears less suitable. The GNF with a large edge-to-wall ratio is more suitable for the high-performance electrode than perfect crystalline graphite or CNT.

  17. Investigation of metal/carbon-related materials for fuel cell applications by electronic structure calculations

    Kong, Ki-jeong; Choi, Youngmin; Ryu, Beyong-Hwan; Lee, Jeong-O; Chang, Hyunju

    2006-01-01

    The potential of carbon-related materials, such as carbon nanotubes (CNTs) and graphite nanofibers (GNFs), supported metal catalysts as an electrode for fuel cell application was investigated using the first-principle electronic structure calculations. The stable binding geometries and energies of metal catalysts are determined on the CNT surface and the GNF edge. The catalyst metal is more tightly bound to the GNF edge than to the CNT surface because of the existence of active dangling bonds of edge carbon atoms. The diffusion barrier of metal atoms on the surface and edge is also obtained. From our calculation results, we have found that high dispersity is achievable for GNF due to high barrier against the diffusion of metal atoms, while CNT appears less suitable. The GNF with a large edge-to-wall ratio is more suitable for the high-performance electrode than perfect crystalline graphite or CNT

  18. Solution-Processing of Organic Solar Cells: From In Situ Investigation to Scalable Manufacturing

    Abdelsamie, Maged

    2016-12-05

    implementation of organic solar cells with high efficiency and manufacturability. In this dissertation, we investigate the mechanism of the BHJ layer formation during solution processing from common lab-based processes, such as spin-coating, with the aim of understanding the roles of materials, formulations and processing conditions and subsequently using this insight to enable the scalable manufacturing of high efficiency organic solar cells by such methods as wire-bar coating and blade-coating. To do so, we have developed state-of-the-art in situ diagnostics techniques to provide us with insight into the thin film formation process. As a first step, we have developed a modified spin-coater which allows us to perform in situ UV-visible absorption measurements during spin coating and provides key insight into the formation and evolution of polymer aggregates in solution and during the transformation to the solid state. Using this method, we have investigated the formation of organic BHJs made of a blend of poly (3-hexylthiophene) (P3HT) and fullerene, reference materials in the organic solar cell field. We show that process kinetics directly influence the microstructure and morphology of the bulk heterojunction, highlighting the value of in situ measurements. We have investigated the influence of crystallization dynamics of a wide-range of small-molecule donors and their solidification pathways on the processing routes needed for attaining high-performance solar cells. The study revealed the reason behind the need of empirically-adopted processing strategies such as solvent additives or alternatively thermal or solvent vapor annealing for achieving optimal performance. The study has provided a new perspective to materials design linking the need for solvent additives or annealing to the ease of crystallization of small-molecule donors and the presence or absence of transient phases before crystallization. From there, we have extended our investigation to small-molecule (p

  19. Investigation of heating and cooling in a stand-alone high temperature PEM fuel cell system

    Zhang, Caizhi; Yu, Tao; Yi, Jun; Liu, Zhitao; Raj, Kamal Abdul Rasheedj; Xia, Lingchao; Tu, Zhengkai; Chan, Siew Hwa

    2016-01-01

    Highlights: • Heating-up and cooling-down processes of HT-PEMFC are the mainly interested topics. • Dynamic behaviours, power and energy demand of the heating and cooling was studied. • Hybrid system based on LiFeYPO_4 battery for heating and cooling is built and tested. • The concept of combining different heating sources together is recommended. - Abstract: One key issue pertaining to the cold-start of High temperature PEM fuel cell (HT-PEMFC) is the requirement of high amount of thermal energy for heating up the stack to a temperature of 120 °C or above before it can generate electricity. Furthermore, cooling down the stack to a certain temperature (e.g. 50 °C) is necessary before stopping. In this study, the dynamic behaviours, power and energy demand of a 6 kW liquid cooled HT-PEMFC stack during heating-up, operation and cooling-down were investigated experimentally. The dynamic behaviours of fuel cell under heating-up and cooling-down processes are the mainly interested topics. Then a hybridisation of HT-PEMFC with Li-ion battery to demonstrate the synergistic effect on dynamic behaviour was conducted and validated for its feasibility. At last, the concept of combining different heating sources together is analysed to reduce the heating time of the HT-PEMFC as well.

  20. Carrageenan activates monocytes via type-specific binding with interleukin-8: an implication for design of immuno-active biomaterials.

    Chan, Weng-I; Zhang, Guangpan; Li, Xin; Leung, Chung-Hang; Ma, Dik-Lung; Dong, Lei; Wang, Chunming

    2017-02-28

    Polymers that can activate the immune system may become useful biomaterials tools, given that the mechanisms underlying their actions are well understood. Herein, we report a novel type of interaction between polymers and immune cells - in studying the influence of the three major types of carrageenan (CGN) polysaccharides on monocyte behaviour in vitro, we found only the λ-type induced monocyte adhesion and this action requires the presence of an adequate amount of serum. Further analyses indicated λ-CGN bound interleukin-8 (IL-8) in the serum and activated the cultured monocytes through an IL-8-dependent pathway. This is the first demonstration that a polymer, with a renowned immunostimulatory effect, activates the immune system via binding and harnessing the function of a specific cytokine in the microenvironment. This is a new mechanism underlying polymer-immunity interactions that may shed light on future design and application of biomaterials tools targeting the immune system for a wide variety of therapeutic applications.

  1. Investigation of the functional role of CSLD proteins in plant cell wall deposition

    Nielsen, Erik Etlar [Univ. of Michigan, Ann Arbor, MI (United States)

    2017-11-21

    The overall goal of this research proposal was to characterize the molecular machinery responsible for polarized secretion of cell wall components in Arabidopsis thaliana. We have used the polarized expansion that occurs during root hair cell growth to identify membrane trafficking pathways involved in polarized secretion of cell wall components to the expanding tips of these cells, and we have recently shown that CSLD3 is preferentially targeted to the apical plasma membranes in root hair cells, where it plays essential roles during cell wall deposition in these cells. The specific aims of the project are designed to answer the following objective: Identification of the cell wall polysaccharide class that CSLD proteins synthesize.

  2. Investigating the role of the extracellular matrix on differentiation of human mesenchymal stem cells and MC3T3 cells

    Fernandes, H.A.M.; Dechering, Koen; Someren, Eugene; van Blitterswijk, Clemens; de Boer, Jan

    2008-01-01

    Human mesenchymal stem cells (hMSCs) are a promising cell source for bone tissue engineering, but due to their limited number and donor variation, other cell types are used to answer relevant questions in bone tissue engineering. Since the extracellular matrix (ECM) is a complex entity with

  3. Investigation of extracellular microRNAs in oral squamous cell carcinoma, rheumatoid arthritis and mesenchymal stem cell differentiation

    Yan, Yan

    serve as biomarkers for human diseases and can also act as mediators in cell-cell communication. In cancer, the abnormal expression of miRNAs in plasma has been observed. However, there is no report on the association of plasma miRNA expression with oral squamous cell carcinoma (OSCC) recurrence after...

  4. Non-Faradaic electrical impedimetric investigation of the interfacial effects of neuronal cell growth and differentiation on silicon nanowire transistors.

    Lin, Shu-Ping; Vinzons, Lester U; Kang, Yu-Shan; Lai, Tung-Yen

    2015-05-13

    Silicon nanowire field-effect transistor (SiNW FET) devices have been interfaced with cells; however, their application for noninvasive, real-time monitoring of interfacial effects during cell growth and differentiation on SiNW has not been fully explored. Here, we cultured rat adrenal pheochromocytoma (PC12) cells, a type of neural progenitor cell, directly on SiNW FET devices to monitor cell adhesion during growth and morphological changes during neuronal differentiation for a period of 5-7 d. Monitoring was performed by measuring the non-Faradaic electrical impedance of the cell-SiNW FET system using a precision LCR meter. Our SiNW FET devices exhibited changes in impedance parameters during cell growth and differentiation because of the negatively charged cell membrane, seal resistance, and membrane capacitance at the cell/SiNW interface. It was observed that during both PC12 cell growth and neuronal differentiation, the impedance magnitude increased and the phase shifted to more negative values. However, impedance changes during cell growth already plateaued 3 d after seeding, while impedance changes continued until the last observation day during differentiation. Our results also indicate that the frequency shift to above 40 kHz after growth factor induction resulted from a larger coverage of cell membrane on the SiNWs due to distinctive morphological changes according to vinculin staining. Encapsulation of PC12 cells in a hydrogel scaffold resulted in a lack of trend in impedance parameters and confirmed that impedance changes were due to the cells. Moreover, cytolysis of the differentiated PC12 cells led to significant changes in impedance parameters. Equivalent electrical circuits were used to analyze the changes in impedance values during cell growth and differentiation. The technique employed in this study can provide a platform for performing investigations of growth-factor-induced progenitor cell differentiation.

  5. Investigation of Near-Surface Defects Induced by Spike Rapid Thermal Annealing in c-SILICON Solar Cells

    Liu, Guodong; Ren, Pan; Zhang, Dayong; Wang, Weiping; Li, Jianfeng

    2016-01-01

    The defects induced by a spike rapid thermal annealing (RTA) process in crystalline silicon (c-Si) solar cells were investigated by the photoluminescence (PL) technique and the transmission electron microscopy (TEM), respectively. Dislocation defects were found to form in the near-surface junction region of the monocrystalline Si solar cell after a spike RTA process was performed at 1100∘C. Photo J-V characteristics were measured on the Si solar cell before and after the spike RTA treatments to reveal the effects of defects on the Si cell performances. In addition, the Silvaco device simulation program was used to study the effects of defects density on the cell performances by fitting the experimental data of RTA-treated cells. The results demonstrate that there was an obvious degradation in the Si solar cell performances when the defect density after the spike RTA treatment was above 1×1013cm-3.

  6. Life coaching following haematopoietic stem cell transplantation: a mixed-method investigation of feasibility and acceptability.

    Kenyon, M; Young, F; Mufti, G J; Pagliuca, A; Lim, Z; Ream, E

    2015-07-01

    Haematopoietic stem cell transplantation (HSCT) cures many haematological cancers. Recovery post-HSCT is physically and psychologically challenging, lasting several months. Beyond the first post-transplant year, a fifth report difficulties encompassing practical, social and emotional domains, including finance and employment. We investigated the feasibility, acceptability and impact of a life coaching intervention designed to address psychosocial 'survivor' concerns of HSCT recipients and facilitate transition to life post-treatment. A concurrent embedded experimental mixed-method design was employed. Pre- and post-intervention data collection comprised qualitative semi-structured telephone interviews and quantitative postal questionnaires. Seven purposively sampled HSCT recipients (life coaching delivered by a professional life coach fortnightly over 8 weeks. Participants reported less anxiety, depression and fewer survivor concerns post-intervention, with a trend for lower social difficulties and increased functional well-being. Perceived self-efficacy was unchanged. Life coaching was feasible to deliver and acceptable to the participants who indicated it was a positive experience, with benefits described in diverse areas including work, lifestyle and hobbies. Life coaching within cancer services potentially offers the means to address psychosocial concerns and support transition to life after treatment, enabling patients to reach their potential, e.g. returning to employment and financial independence. Further investigation of this intervention in cancer survivors is warranted. © 2015 John Wiley & Sons Ltd.

  7. Impedance Spectroscopic Investigation of the Degraded Dye-Sensitized Solar Cell due to Ageing

    Parth Bhatt

    2016-01-01

    Full Text Available This paper investigates the effect of ageing on the performance of dye-sensitized solar cells (DSCs. The electrical characterization of fresh and degraded DSCs is done under AM1.5G spectrum and the current density-voltage (J-V characteristics are analyzed. Short circuit current density (JSC decreases significantly whereas a noticeable increase in open circuit voltage is observed. These results have been further investigated electroanalytically using electrochemical impedance spectroscopy (EIS. An increase in net resistance results in a lower JSC for the degraded DSC. This decrease in current is mainly due to degradation of TiO2-dye interface, which is observed from light and dark J-V characteristics and is further confirmed by EIS measurements. A reduction in the chemical capacitance of the degraded DSC is observed, which is responsible for the shifting of Fermi level with respect to conduction band edge that further results in an increase of open circuit voltage for the degraded DSC. It is also confirmed from EIS that the degradation leads to a better contact formation between the electrolyte and Pt electrode, which improves the fill factor of the DSC. But the recombination throughout the DSC is found to increase along with degradation. This study suggests that the DSC should be used under low illumination conditions and around room temperature for a longer life.

  8. Investigation of test methods, material properties, and processes for solar cell encapsulants. Annual report

    Willis, P. B.; Baum, B.; Schnitzer, H. S.

    1980-07-01

    The goal of this program is to identify, evaluate, and recommend encapsulant materials and processes for the production of cost-effective, long-life solar cell modules. Technical activities during the past year have covered a number of topics and have emphasized the development of solar module encapsulation technology that employs ethylene/vinyl acetate, copolymer (EVA) as the pottant. These activities have included: (1) continued production of encapsulation grade EVA in sheet form to meet the needs of the photovoltaic industry; (2) investigations of three non-blocking techniques for EVA sheet; (3) performed an economic analysis of the high volume production of each pottant in order to estimate the large volume selling price (EVA, EPDM, aliphatic urethane, PVC plastisol, and butyl acrylate); (4) initiated an experimental corrosion protection program to determine if metal components could be successfully protected by encapsulation; (5) began an investigation to determine the maximum temperature which can be tolerated by the candidate pottant material in the event of hot spot heating or other temperature override; (6) continuation of surveys of potentially useful outer cover materials; and (7) continued with the accelerated artificial weathering of candidate encapsulation materials. Study results are presented. (WHK)

  9. Investigation of chemical and physical properties of carbon nanotubes and their effects on cell biomechanics

    Dong, Chenbo

    Carbon nanotubes (CNTs) are used for a variety of applications from nanocircuits, to hydrogen storage devices, and from designing optical fibers to forming conductive plastics. Recently, their functionalization with biomolecules led to exciting biological and biomedical applications in drug delivery or bioimaging. However, because of CNTs interactions with biological systems and their ability to translocate and persist into the circulatory and lymphatic systems and biological tissues, concerns about CNTs intrinsic toxicity have risen. It is thus necessary to develop and implement sensitive analysis technologies that allow investigation of CNTs toxicity upon uptake into a biological system. This thesis provides a comprehensive guide of experiments that have been performed during my Ph.D. tenure at West Virginia University in the Department of Chemical Engineering, in the group of Prof. Cerasela Zoica Dinu. Briefly: Chapter one presents a systematic study of the CNTs physical and chemical properties and how these properties are changed upon exposure to chemical agents normally used during their cleaning and purification processes. Also, this chapter shows how acid oxidation treatment leads to improved CNTs biocompatibility. Specifically, by incubating CNTs in a strong acid mixture we created a user-defined library of CNTs samples with different characteristics as recorded using Raman energy dispersive x-ray spectroscopy, atomic force microscopy, or solubility tests. Systematically characterized CNTs were subsequently tested for their biocompatibility in relation to human epithelial cells or enzymes. Such selected examples are building pertinent relationships between CNTs biocompatibility and their intrinsic properties by showing that acid oxidation treatment lowers CNTs toxicity making CNTs feasible platforms to be used for biomedical applications or the next generation of biosensors. (Publication: Chenbo Dong, Alan S Campell, Reem Eldawud, Gabriela Perhinschi, and

  10. Experimental investigation of the cytotoxicity of medium-borne signals in human prostate cancer cell line

    Sjostedt, Svetlana; Bezak, Eva

    2012-01-01

    Introduction. Evidence exists that exposure of non-irradiated cells to Irradiated Cell Conditioned Medium (ICCM) can cause effects similar to those resulting from direct radiation damage. This study attempts to validate the stochastic model, relating absorbed dose to the emission and processing of cell death signals by non-irradiated cells, in vitro in PC3 human prostate cancer cell line. Methods. The recipient cell survival was measured after exposure of cells to ICMM derived from donor cells: a) exposed to radiation doses from 2 Gy to 8 Gy and b) of concentrations varying from 2 x 10 2 to 6 x 10 6 irradiated with 2 Gy. Results. Exposure to ICCM, irradiated with doses between 2-8 Gy, resulted in a significant (p 2 cells was significantly higher (p < 0.5) compared to the rest of donor cell concentrations, indicating that the toxicity of ICCM depends on the cellular concentration of donor cells. Non-linear regression data fitting provided reasonable agreement with the microdosimetric model for the induction of cell killing through medium-borne signals. Conclusion. For the given cell line and given experimental conditions, significant decreases in cell survival were observed in non-irradiated cells exposed to ICCM derived from donor cells of various concentrations and irradiated with different doses

  11. AN INVESTIGATION TO RESOLVE THE INTERACTION BETWEEN FUEL CELL, POWER CONDITIONING SYSTEM AND APPLICATION LOADS

    Sudip K. Mazumder; Chuck McKintyre; Dan Herbison; Doug Nelson; Comas Haynes; Michael von Spakovsky; Joseph Hartvigsen; S. Elangovan

    2003-11-03

    Solid-Oxide Fuel Cell (SOFC) stacks respond quickly to changes in load and exhibit high part- and full-load efficiencies due to its rapid electrochemistry. However, this is not true for the thermal, mechanical, and chemical balance-of-plant subsystem (BOPS), where load-following time constants are, typically, several orders of magnitude higher. This dichotomy diminishes the reliability and performance of the electrode with increasing demand of load. Because these unwanted phenomena are not well understood, the manufacturers of SOFC use conservative schemes (such as, delayed load-following to compensate for slow BOPS response or expensive inductor filtering) to control stack responses to load variations. This limits the applicability of SOFC systems for load-varying stationary and transportation applications from a cost standpoint. Thus, a need exists for the synthesis of component- and system-level models of SOFC power-conditioning systems and the development of methodologies for investigating the system-interaction issues (which reduce the lifetime and efficiency of a SOFC) and optimizing the responses of each subsystem, leading to optimal designs of power-conditioning electronics and optimal control strategies, which mitigate the electrical-feedback effects. Equally important are ''multiresolution'' finite-element modeling and simulation studies, which can predict the impact of changes in system-level variables (e.g., current ripple and load-transients) on the local current densities, voltages, and temperature (these parameters are very difficult or cumbersome, if not impossible to obtain) within a SOFC cell. Towards that end, for phase I of this project, sponsored by the U.S. DOE (NETL), we investigate the interactions among fuel cell, power-conditioning system, and application loads and their effects on SOFC reliability (durability) and performance. A number of methodologies have been used in Phase I to develop the steady-state and transient

  12. Investigations of the functional states of dendritic cells under different conditioned microenvironments by Fourier transformed infrared spectroscopy.

    Dong, Rong; Long, Jinhua; Xu, Xiaoli; Zhang, Chunlin; Wen, Zongyao; Li, Long; Yao, Weijuan; Zeng, Zhu

    2014-01-10

    Dendritic cells are potent and specialized antigen presenting cells, which play a crucial role in initiating and amplifying both the innate and adaptive immune responses. The dendritic cell-based vaccination against cancer has been clinically achieved promising successes. But there are still many challenges in its clinical application, especially for how to identify the functional states. The CD14+ monocytes were isolated from human peripheral blood after plastic adherence and purified to approximately 98% with cocktail immunomagnetic beads. The immature dendritic cells and mature dendritic cells were induced by traditional protocols. The resulting dendritic cells were cocultured with normal cells and cancer cells. The functional state of dendritic cells including immature dendritic cells (imDCs) and mature dendritic cells (mDCs) under different conditioned microenvironments were investigated by Fourier transformed infrared spectroscopy (FTIR) and molecular biological methods. The results of Fourier transformed infrared spectroscopy showed that the gene transcription activity and energy states of dendritic cells were specifically suppressed by tumor cells (P Fourier transformed infrared spectroscopy at given wave numbers were closely correlated with the expression levels of NF-κB (R2:0.69 and R2:0.81, respectively). Our results confirmed that the ratios of absorption intensities of Fourier transformed infrared spectroscopy at given wave numbers were positively correlated with the expression levels of NF-κB, suggesting that Fourier transformed infrared spectroscopy technology could be clinically applied to identify the functional states of dendritic cell when performing dendritic cell-based vaccination. It's significant for the simplification and standardization of dendritic cell-based vaccination clinical preparation protocols.

  13. Investigations on the kinetics of the oxygen reduction in high temperature fuel cells

    Erning, J.W.

    1998-07-01

    Lanthan-Strontium-Manganite perowskites are the most widespread materials in use for solid oxide fuel cell cathodes. The electrode reaction taking place, i.e. the reduction of oxygen supplied by air, was investigated by electrochemical means to obtain further knowledge about the electrode processes. The high activation energy of this reaction (200 kJ/mol), preventing lower operation temperatures of the SOFC, was the starting point for the investigation. Quasi steady state current voltage measurements and impedance spectroscopy were performed in a three electrode configuration. The electrodes were of circular shape with a diameter of 10 mm. The preparation was made by screen printing as well as wet powder spraying onto plates made of Yttria-stabilized zirconia. Perowskite powders of varying chemical and stoichiometric composition were used. To obtain higher power densities and, more important, lower apparent activation energies, catalytic layers were added at the interface electrode/electrolyte. Additionally, a less complex system, a model electrode/electrolyte setup made from single-crystal YSZ as electrolyte and gold in liquid and solid state as electrode was developed to create a better defined system. This setup was used to investigate the behaviour of the electrode/electrolyte interface. Reliable, reproducible results could be obtained using either setup. The experimental conditions i.e. oxygen partial pressure, temperature and overpotential were varied in order to determine the kinetic properties of the electrodes. Apparent activation energies, pre-exponential factors, apparent charge-transfer coefficients and electrochemical orders of reaction were calculated from the current-voltage data in order to propose possible reaction steps. (orig.)

  14. Investigation of Membrane Receptors' Oligomers Using Fluorescence Resonance Energy Transfer and Multiphoton Microscopy in Living Cells

    Mishra, Ashish K.

    Investigating quaternary structure (oligomerization) of macromolecules (such as proteins and nucleic acids) in living systems (in vivo) has been a great challenge in biophysics, due to molecular diffusion, fluctuations in several biochemical parameters such as pH, quenching of fluorescence by oxygen (when fluorescence methods are used), etc. We studied oligomerization of membrane receptors in living cells by means of Fluorescence (Forster) Resonance Energy Transfer (FRET) using fluorescent markers and two photon excitation fluorescence micro-spectroscopy. Using suitable FRET models, we determined the stoichiometry and quaternary structure of various macromolecular complexes. The proteins of interest for this work are : (1) sigma-1 receptor and (2) rhodopsin, are described as below. (1) Sigma-1 receptors are molecular chaperone proteins, which also regulate ion channels. S1R seems to be involved in substance abuse, as well as several diseases such as Alzheimer's. We studied S1R in the presence and absence of its ligands haloperidol (an antagonist) and pentazocine +/- (an agonist), and found that at low concentration they reside as a mixture of monomers and dimers and that they may form higher order oligomers at higher concentrations. (2) Rhodopsin is a prototypical G protein coupled receptor (GPCR) and is directly involved in vision. GPCRs form a large family of receptors that participate in cell signaling by responding to external stimuli such as drugs, thus being a major drug target (more than 40% drugs target GPCRs). Their oligomerization has been largely controversial. Understanding this may help to understand the functional role of GPCRs oligomerization, and may lead to the discovery of more drugs targeting GPCR oligomers. It may also contribute toward finding a cure for Retinitis Pigmentosa, which is caused by a mutation (G188R) in rhodopsin, a disease which causes blindness and has no cure so far. Comparing healthy rhodopsin's oligomeric structure with that

  15. Experimental investigation of pulsating heat pipe performance with regard to fuel cell cooling application

    Clement, Jason; Wang Xia

    2013-01-01

    A pulsating heat pipe (PHP) is a closed loop, passive heat transfer device. Its operation depends on the phase change of a working fluid within the loop. Design and performance testing of a pulsating heat pipe was conducted under conditions to simulate heat dissipation requirements of a proton exchange membrane (PEM) fuel cell stack. Integration of pulsating heat pipes within bipolar plates of the stack would eliminate the need for ancillary cooling equipment, thus also reducing parasitic losses and increasing energy output. The PHP under investigation, having dimensions of 46.80 cm long and 14.70 cm wide, was constructed from 0.3175 cm copper tube. Heat pipes effectiveness was found to be dependent upon several factors such as energy input, types of working fluid and its filling ratio. Power inputs to the evaporator side of the pulsating heat pipe varied from 80 to 180 W. Working fluids tested included acetone, methanol, and deionized water. Filling ratios between 30 and 70 percent of the total working volume were also examined. Methanol outperformed other fluids tested; with a 45 percent fluid fill ratio and a 120 W power input, the apparatus took the shortest time to reach steady state and had one of the smallest steady state temperature differences. The various conditions studied were chosen to assess the heat pipe's potential as cooling media for PEM fuel cells. - Highlights: ► Methanol as a working fluid outperformed both acetone and water in a pulsating heat pipe. ► Performance for the PHP peaked with methanol and a fill ratio of 45 percent fluid to total volume. ► A smaller resistance was associated with a higher power input to the system.

  16. Mysterious inhibitory cell regulator investigated and found likely to be secretogranin II related

    John E. Hart

    2017-10-01

    Full Text Available In the context of a hunt for a postulated hormone that is tissue-mass inhibiting and reproductively associated, there is described probable relatedness to a granin protein. A 7–8 kDa polypeptide candidate (gels/MS appeared in a bioassay-guided fractionation campaign involving sheep plasma. An N-terminal sequence of 14 amino acids was obtained for the polypeptide by Edman degradation. Bioinformatics and molecular biology failed to illuminate any ovine or non-ovine protein which might relate to this sequence. The N-terminal sequence was synthesized as the 14mer EPL001 peptide and surprisingly found to be inhibitory in an assay in vivo of compensatory renal growth in the rat and modulatory of nematode fecundity, in line with the inhibitory hormone hypothesis. Antibodies were raised to EPL001 and their deployment upheld the hypothesis that the EPL001 amino acid sequence is meaningful and relevant, notwithstanding bioinformatic obscurity. Immunohistochemistry (IHC in sheep, rodents and humans yielded staining of seeming endocrine relevance (e.g. hypothalamus, gonads and neuroendocrine cells in diverse tissues, with apparent upregulation in certain human tumours (e.g. pheochromocytoma. Discrete IHC staining in Drosophila melanogaster embryo brain was seen in glia and in neuroendocrine cells, with staining likely in the corpus cardiacum. The search for the endogenous antigen involved immunoprecipitation (IP followed by liquid chromatography and mass spectrometry (LC–MS. Feedstocks were PC12 conditioned medium and aqueous extract of rat hypothalamus—both of which had anti-proliferative and pro-apoptotic effects in an assay in vitro involving rat bone marrow cells, which inhibition was subject to prior immunodepletion with an anti-EPL001 antibody—together with fruit fly embryo material. It is concluded that the mammalian antigen is likely secretogranin II (SgII related. The originally seen 7–8 kDa polypeptide is suggested to be a new proteoform

  17. Investigation of the optical properties of novel organic macromolecules for solar cell applications

    Adegoke, Oluwasegun Oluwasina

    The search for renewable energy sources to replace fossil fuel has been a major research focus in the energy sector. The sun, with its vast amount of energy, remains the most abundant and ubiquitous energy source that far exceeds the world energy demand. The ability to effectively capture and convert energy from the sun in the form of photons will be the key to its effective utilization. Organic macromolecules have tremendous potentials to replace and out-perform existing materials, due to their low-cost, ease of tunability, high absorption coefficient and "green" nature. In this dissertation, spectroscopic techniques of steady state absorption and time-resolved fluorescence spectroscopy were used to show the improved absorption of the oligothiophene-functionalized ZnPc through ultrafast energy transfer. ZnPc is known for its chemical and thermal stability. The power conversion efficiency (PCE) in ZnPc-based solar devices is however, very low because of the poor absorption of ZnPc in the 300 - 550 nm region of the solar spectrum. Oligothiophenes have good absorption in the spectral region where the absorption of ZnPc is poor. Other groups of organic compounds that have gained prominence in the study for the design of efficient active materials for photovoltaic cells are the polymers. In the dissertation, different factors which can affect the performance of organic polymers in photovoltaics systems were investigated and analyzed. The effects of the alteration of conjugation, donor-acceptor groups, heteroatoms and alkyl side chains on the photophysical properties and ultimately the performance of organic polymers in organic photovoltaics were investigated. The different effects were investigated using ultrafast spectroscopic techniques which are capable of providing insight of fluorescence decay dynamics at very short times in a time scale of femtosecond. The electronic structure calculations of the polymers were carried out to provide further evidence to the

  18. Solid electrolytes in thermodynamic investigations. Investigation of oxygen pressure effect in Ar + O2 type mixtures on cell potentials with CaF2 electrolyte and oxide electrodes

    Levitskij, V.A.; Narchuk, N.B.; Kashkarova, S.L.

    1982-01-01

    An experimental test of the P'sub(Osub(2))=P''sub(Osub(2)) condition (P'sub(Osub(2)) and P''sub(Osub(2)) - oxygen pressure above the first and the second electrodes) necessary for reversible work of the cells is carried out with the (-)O 2 , Pt (CaZrO 3 , 0.18CaOx0.82ZrO 2 )CaF 2 CaF 2 CaHfO 3 , HfO 2 , CaF 2 Pt, O 2 (+) cell as an example. The equilibrium potentials of the given cell are shown to be independent from Psub(Osub(2)) value above both electrodes up to the O 2 pressure equal to approximately 1 Pa at Psub(gen.)=Psub(Osub(2))+Psub(Ar)=10sup(5) Pa. Thermodynamic parameters of the CaO+HfO 2 =CaHfO 3 reaction obtained from the E=f(T) dependence in argon atmosphere under Psub(Osub(2))=1-10sup(2) Pa well agree with analogous values determined for the same cell under Psub(Osub(2)) = 10 5 Pa. Comparison of the results obtained by the e. m. f. method with F - - ion electrolyte both in pure oxygen and in argon atmosphere under low Psub(Osub(2)) with the present literature data testify to perspectives of using the cells of this type under controlled low Psub(Osub(2)) values for thermodynamic investigations

  19. Nanowire arrays as cell force sensors to investigate adhesin-enhanced holdfast of single cell bacteria and biofilm stability

    Sahoo, P.K.; Janissen, R.; Monteiro, M.P.; Cavalli, A.; Murillo, D.M.; Merfa, M.V.; Cesar, C.L.; Carvalho, H.F.; de Souza, A.A.; Bakkers, E.P.A.M.; Cotta, M.A.

    2016-01-01

    Surface attachment of a planktonic bacteria, mediated by adhesins and extracellular polymeric substances (EPS), is a crucial step for biofilm formation. Some pathogens can modulate cell adhesiveness, impacting host colonization and virulence. A framework able to quantify cell-surface interaction

  20. An Investigation to Resolve the Interaction Between Fuel Cell, Power Conditioning System and Application Loads

    Sudip K. Mazumder

    2005-12-31

    Development of high-performance and durable solidoxide fuel cells (SOFCs) and a SOFC power-generating system requires knowledge of the feedback effects from the power-conditioning electronics and from application-electrical-power circuits that may pass through or excite the power-electronics subsystem (PES). Therefore, it is important to develop analytical models and methodologies, which can be used to investigate and mitigate the effects of the electrical feedbacks from the PES and the application loads (ALs) on the reliability and performance of SOFC systems for stationary and non-stationary applications. However, any such attempt to resolve the electrical impacts of the PES on the SOFC would be incomplete unless one utilizes a comprehensive analysis, which takes into account the interactions of SOFC, PES, balance-of-plant system (BOPS), and ALs as a whole. SOFCs respond quickly to changes in load and exhibit high part- and full-load efficiencies due to its rapid electrochemistry, which is not true for the thermal and mechanical time constants of the BOPS, where load-following time constants are, typically, several orders of magnitude higher. This dichotomy can affect the lifetime and durability of the SOFCSs and limit the applicability of SOFC systems for load-varying stationary and transportation applications. Furthermore, without validated analytical models and investigative design and optimization methodologies, realizations of cost-effective, reliable, and optimal PESs (and power-management controls), in particular, and SOFC systems, in general, are difficult. On the whole, the research effort can lead to (a) cost-constrained optimal PES design for high-performance SOFCS and high energy efficiency and power density, (b) effective SOFC power-system design, analyses, and optimization, and (c) controllers and modulation schemes for mitigation of electrical impacts and wider-stability margin and enhanced system efficiency.

  1. [Fluorescence polarization used to investigate the cell membrane fluidity of Saccharomyces cerevisiae treated by pulsed electric field].

    Zhang, Ying; Zeng, Xin-An; Wen, Qi-Biao; Li, Lin

    2008-01-01

    To know the lethal mechanism of microorganisms under pulsed electric field treatment, the relationship between the inactivation of Saccharomyces cerevisiae (CICC1308) cell and the permeability and fluidity changes of its cell membrane treated by pulsed electric field (0-25 kV x cm(-1), 0-266 ms) was investigated. With 1,6-diphenyl-1,3,5-hexatriene (DPH) used as a probe, the cell membrane fluidity of Saccharomyces cerevisiae treated by pulsed electric field was expressed by fluorescence polarization. Results showed that the cell membrane fluidity decreases when the electric flied strength is up to 5 kV x cm(-1), and decreases with the increase in electric field strength and treatment time. The plate counting method and ultraviolet spectrophotometer were used to determine the cell viability and to investigate the cell membrane permeability, respectively, treated by pulsed electric field. Results showed that the lethal ratio and the content of protein and nucleic acid leaked from intracellular plasma increased with the increase in the electric field strength and the extension of treatment time. Even in a quite lower electric field of 5 kV x cm(-1) with a tiny microorganism lethal level, the increase in UV absorption value and the decrease in fluidity were significant. It was demonstrated that the cell membrane fluidity decreases with the increase in lethal ratio and cell membrane permeability. The viscosity of cell membrane increases with the decrease in fluidity. These phenomena indicated that cell membrane is one of the most key sites during the pulsed electric field treatment, and the increased membrane permeability and the decreased cell membrane fluidity contribute to the cell death.

  2. Program of scientific investigations and development of solid-oxide fuel cells (SOFC) in VIITF proposals on scientific and technical collaboration and SOFC commercialization

    Kleschev, Yu.N.; Chulharev, V.F.

    1996-04-01

    Investigations being performed at VNIITF covers the whole cycle of solid oxide fuel cell manufacturing. This report describes the main directions of investigations in materials, technologies, and commercialization.

  3. Investigation of dominant loss mechanisms in low-temperature polymer electrolyte membrane fuel cells

    Gerteisen, D.

    2010-01-01

    This thesis deals with the analysis of dominant loss mechanisms in direct methanol fuel cells (DMFC) and hydrogen fed polymer electrolyte membrane fuel cells (PEFC) by means of experimental characterization and modeling work.

  4. Investigation of HIV-1 infected and uninfected cells using the optical trapping technique

    Ombinda-Lemboumba, Saturnin

    2017-02-01

    Full Text Available Optical trapping has emerged as an essential tool for manipulating single biological material and performing sophisticated spectroscopy analysis on individual cell. The optical trapping technique has been used to grab and immobilize cells from a...

  5. Solution-Processed Organic Solar Cells from Dye Molecules: An Investigation of Diketopyrrolopyrrole:Vinazene Heterojunctions

    Walker, Bright; Han, Xu; Kim, Chunki; Sellinger, Alan; Nguyen, Thuc-Quyen

    2012-01-01

    Although one of the most attractive aspects of organic solar cells is their low cost and ease of fabrication, the active materials incorporated into the vast majority of reported bulk heterojunction (BHJ) solar cells include a semiconducting polymer

  6. An Investigation of the Effects of Raw Garlic on Radiation-induced Bystander Effects in MCF7 Cells

    Shokouhozaman Soleymanifard

    2014-11-01

    Full Text Available Introduction Radiation-induced bystander effect (RIBE is a phenomenon in which radiation signals are transmitted from irradiated cells to non-irradiated ones, inducing radiation effects in these cells. RIBE plays an effective role in radiation response at environmentally relevant low doses and in radiotherapy, given its impact on adjacent normal tissues or those far from the irradiated tumor. Reactive oxygen species contribute to RIBE induction. Therefore, the present study was conducted to investigate the possible inhibitory effects of garlic, as an antioxidant-containing plant, on RIBE. Materials and Methods MCF7 cells, treated with raw garlic extracts, were irradiated by 60Co gamma rays, and their culture medium was transferred to non-irradiated autologous bystander cells. Percentage cell viability and micronucleus formation in both irradiated and bystander cells were examined and compared with corresponding cell groups, not treated with garlic. Results Treatment with garlic extract reduced the number of micronucleus-containing cells in both irradiated and bystander cells. However, it only increased the percentage cell viability in bystander cells, not the irradiated ones. Conclusion RIBE was effectively suppressed by raw garlic extracts. Inhibitory effects of raw garlic may be of particular importance for exposure to environmentally relevant low doses, where RIBE dominates direct radiation effects. They are also partially important for addressing the limited therapeutic gain of radiotherapy, as they may only increase the percentage cell viability of bystander cells, not the directly irradiated tumor cells. However, more comprehensive in-vivo research regarding garlic treatment duration is required to support the obtained results.

  7. An Investigation of Short Circuits in All-solution Processed and All-organic Solar Cells

    Johansson, Jim

    2015-01-01

    Organic solar cells have shown great promise of becoming a cheaper alternative to inorganic solar cells. Additionally, they can also be made semitransparent. To avoid using expensive indium tin oxide electrodes in organic solar cells the electrodes can be made from conductive polymer, poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS). However, these so-called PEDOT-PEDOT solar cells are prone to short-circuiting. The work behind this thesis thus aimed to find the cause of the...

  8. Investigations of the toxic effects of glycans-based silver nanoparticles on different types of human cells

    Panzarini, E.; Mariano, S.; Dini, L.

    2017-08-01

    The effects of glycans-capped AgNPs (30±5 nm average diameter, spherical shape) on biocompatibility and uptake was studied in relation to the glycan capping (glucose AgNPs-G, glucose/sucrose AgNPs-GS, glucose/fructose AgNPs-GF), and to the cell types (HeLa cells, lymphocytes, and HepG2 cells). Glycan capping and type of cells drive morphological changes, viability loss and type and extent of cell death induction; in addition cells response is largely influenced by the AgNPs amount. The MTT photometric method to determine cell metabolism and the analysis of the membrane integrity by Annexin V-Propidium Iodide labelling were used to quantify cell viability and cell death with different concentrations of NPs. It turns out that i) AgNPs-GF are the most toxic, whereas ii) AgNPs-GS are the less toxic NPs, probably due to the stability of glucose/sucrose capping up to 5 days in culture medium; iii) HepG2 cells are the most sensitive to the presence of NPs. A deeper investigation is necessary to explain the interesting PBLs proliferation increase observed in the presence of AgNPs-GS.

  9. Biochemistry and biology: heart-to-heart to investigate cardiac progenitor cells.

    Chimenti, Isotta; Forte, Elvira; Angelini, Francesco; Messina, Elisa; Giacomello, Alessandro

    2013-02-01

    Cardiac regenerative medicine is a rapidly evolving field, with promising future developments for effective personalized treatments. Several stem/progenitor cells are candidates for cardiac cell therapy, and emerging evidence suggests how multiple metabolic and biochemical pathways strictly regulate their fate and renewal. In this review, we will explore a selection of areas of common interest for biology and biochemistry concerning stem/progenitor cells, and in particular cardiac progenitor cells. Numerous regulatory mechanisms have been identified that link stem cell signaling and functions to the modulation of metabolic pathways, and vice versa. Pharmacological treatments and culture requirements may be exploited to modulate stem cell pluripotency and self-renewal, possibly boosting their regenerative potential for cell therapy. Mitochondria and their many related metabolites and messengers, such as oxygen, ROS, calcium and glucose, have a crucial role in regulating stem cell fate and the balance of their functions, together with many metabolic enzymes. Furthermore, protein biochemistry and proteomics can provide precious clues on the definition of different progenitor cell populations, their physiology and their autocrine/paracrine regulatory/signaling networks. Interdisciplinary approaches between biology and biochemistry can provide productive insights on stem/progenitor cells, allowing the development of novel strategies and protocols for effective cardiac cell therapy clinical translation. This article is part of a Special Issue entitled Biochemistry of Stem Cells. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. 77 FR 5487 - Countervailing Duty Investigation of Crystalline Silicon Photovoltaic Cells, Whether or Not...

    2012-02-03

    ... modules or panels) and 8541.40.6030 (solar cells, not assembled into modules or made up into panels) for... crystalline silicon photovoltaic cells, whether or not assembled into modules (solar cells), from the People's Republic of China (PRC), filed in proper form by SolarWorld Industries America Inc. (Petitioner).\\1\\ The...

  11. Thermodynamic investigation of an integrated gasification plant with solid oxide fuel cell and steam cycles

    Rokni, Masoud [Technical Univ. of Denmark, Lyngby (Denmark). Dept. of Mechanical Engineering, Thermal Energy System

    2012-07-01

    A gasification plant is integrated on the top of a solid oxide fuel cell (SOFC) cycle, while a steam turbine (ST) cycle is used as a bottoming cycle for the SOFC plant. The gasification plant was fueled by woodchips to produce biogas and the SOFC stacks were fired with biogas. The produced gas was rather clean for feeding to the SOFC stacks after a simple cleaning step. Because all the fuel cannot be burned in the SOFC stacks, a burner was used to combust the remaining fuel. The off-gases from the burner were then used to produce steam for the bottoming steam cycle in a heat recovery steam generator (HRSG). The steam cycle was modeled with a simple single pressure level. In addition, a hybrid recuperator was used to recover more energy from the HRSG and send it back to the SOFC cycle. Thus two different configurations were investigated to study the plants characteristic. Such system integration configurations are completely novel and have not been studied elsewhere. Plant efficiencies of 56% were achieved under normal operation which was considerably higher than the IGCC (Integrated Gasification Combined Cycle) in which a gasification plant is integrated with a gas turbine and a steam turbine. Furthermore, it is shown that under certain operating conditions, plant efficiency of about 62 is also possible to achieve. (orig.)

  12. Social skills and executive function among youth with sickle cell disease: a preliminary investigation.

    Hensler, Molly; Wolfe, Kelly; Lebensburger, Jeffrey; Nieman, Jilian; Barnes, Margaux; Nolan, William; King, Allison; Madan-Swain, Avi

    2014-06-01

    To explore the relationship between executive function (EF) and social skills in youth with sickle cell disease (SCD).   20 youth with SCD completed objective tests of EF (Tasks of Executive Control; Animal Sorting subtest from the Developmental Neuropsychological Assessment-Second Edition), an IQ screener, and paper-and-pencil measures of social skills (Social Skills Improvement System [SSIS]). Primary caregivers completed paper-and-pencil measures of EF (Behavior Rating Inventory of Executive Function) and social skills (SSIS).   EF scores from the Behavior Rating Inventory of Executive Function related to parent- and child-reported social skills such that EF deficits correlated with poorer overall and domain-specific social skills. Similarly, EF scores from the Animal Sorting test related to child-reported social skills. Worse parent-reported EF predicted worse parent-reported social skills above the variance accounted for by IQ.   EF is related to social skills and may be necessary for successful social interaction among youth with SCD. These results provide rationale and guidance for future larger-scale investigations of EF and social skills among children with SCD. © The Author 2014. Published by Oxford University Press on behalf of the Society of Pediatric Psychology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. Investigation of membrane mechanics using spring networks: application to red-blood-cell modelling.

    Chen, Mingzhu; Boyle, Fergal J

    2014-10-01

    In recent years a number of red-blood-cell (RBC) models have been proposed using spring networks to represent the RBC membrane. Some results predicted by these models agree well with experimental measurements. However, the suitability of these membrane models has been questioned. The RBC membrane, like a continuum membrane, is mechanically isotropic throughout its surface, but the mechanical properties of a spring network vary on the network surface and change with deformation. In this work spring-network mechanics are investigated in large deformation for the first time via an assessment of the effect of network parameters, i.e. network mesh, spring type and surface constraint. It is found that a spring network is conditionally equivalent to a continuum membrane. In addition, spring networks are employed for RBC modelling to replicate the optical tweezers test. It is found that a spring network is sufficient for modelling the RBC membrane but strain-hardening springs are required. Moreover, the deformation profile of a spring network is presented for the first time via the degree of shear. It is found that spring-network deformation approaches continuous as the mesh density increases. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Investigation of gas flow characteristics in proton exchange membrane fuel cell

    Kwac, Lee Ku; Kim, Hong Gun

    2008-01-01

    An investigation of electrochemical behavior of PEMFC (proton exchange membrane fuel cell) is performed by using a single-phase two-dimensional finite element analysis. Equations of current balance, mass balance, and momentum balance are implemented to simulate the behavior of PEMFC. The analysis results for the co-flow and counterflow mode of gas flow direction are examined in detail in order to compare how the gas flow direction affects quantitatively. The characteristics of internal properties, such as gas velocity distribution, mass fraction of the reactants, fraction of water and current density distribution in PEMFC are illustrated in the electrode and GDL (gas diffusion layer). It is found that the dry reactant gases can be well internally humidified and maintain high performance in the case of the counter-flow mode without external humidification while it is not advantageous for highly humidified or saturated reactant gases. It is also found that the co-flow mode improves the current density distribution with humidified normal condition compared to the counter-flow mode

  15. Extension of Applicability of integral neutron transport theory in reactor cell and core investigation

    Pop-Jordanov, J.; Bosevski, T.; Kocic, A.; Altiparmakov, D.

    1980-01-01

    A Space-Point Energy-Group integral transport theory method (SPEG) is developed and applied to the local and global calculations of the Yugoslav RA reactor. Compared to other integral transport theory methods, the SPEG distinguishes by (1) the arbitrary order of the polynomial, (2) the effective determination of integral parameters through point flux values, (3) the use of neutron balance condition. as a posterior measure of the accuracy of the calculation and (4) the elimination of the subdivisions- into zones, in realistic cases. In addition, different direct (collision probability) and indirect (Monte Carlo) approaches to integral transport theory have been investigated and Some effective acceleration procedures introduced. The study was performed on three test problems in plane and cylindrical geometry, as well as on the nine-region cell of the RA reactor. In particular, the limitations of the integral transport theory including its non-applicability to optically large material regions and to global reactor calculations were examined. The proposed strictly multipoint approach, avoiding the subdivision into zones and groups, seems to provide a good starting point to overcome these limitations of the integral transport theory. (author)

  16. Investigation of methane steam reforming in planar porous support of solid oxide fuel cell

    Yang Yongping; Du Xiaoze; Yang Lijun; Huang Yuan; Xian Haizhen

    2009-01-01

    Adopting the porous support in integrated-planar solid oxide fuel cell (IP-SOFC) can reduce the operating temperature by reducing thickness of electrolyte layer, and also, provide internal reforming environment for hydrogen-rich fuel gas. The distributions of reactant and product components, and temperature of methane steam reforming for IP-SOFC were investigated by the developed physical and mathematical model with thermodynamic analysis, in which eleven possible reaction mechanisms were considered by the source terms and Arrhenius relationship. Numerical simulation of the model revealed that the progress of reforming reaction and the distribution of the product, H 2 , were influenced by the operating conditions, included that of temperature, ratio of H 2 O and CH 4 , as well as by the porosity of the supporting material. The simulating results indicate that the methane conversion rate can reach its maximum value under the operating temperature of 800 deg. C and porosity of ε = 0.4, which rather approximate to the practical operating conditions of IP-SOFC. In addition, characteristics of carbon deposition on surface of catalyst were discussed under various operating conditions and configuration parameters of the porous support. The present works provided some theoretical explanations to the numerous experimental observations and engineered practices

  17. Investigation of selective induction of breast cancer cells to death with treatment of plasma-activated medium

    Hashizume, Hiroshi; Tanaka, Hiromasa; Nakamura, Kae; Kano, Hiroyuki; Ishikawa, Kenji; Kikkawa, Fumitaka; Mizuno, Masaaki; Hori, Masaru

    2015-09-01

    The applications of plasma in medicine have much attention. We previously showed that plasma-activated medium (PAM) induced glioblastoma cells to apoptosis. However, it has not been elucidated the selectivity of PAM in detail. In this study, we investigated the selective effect of PAM on the death of human breast normal and cancer cells, MCF10A and MCF7, respectively, and observed the selective death with fluorescent microscopy. For the investigation of cell viability with PAM treatment, we prepared various PAMs according to the strengths, and treated each of cells with PAMs. Week PAM treatment only decreased the viability of MCF7 cells, while strong PAM treatment significantly affected both viabilities of MCF7 and MCF10A cells. For the fluorescent observation, we prepared the mixture of MCF7 and fluorescent-probed MCF10A cells, and seeded them. After the treatment of PAMs, the images showed that only MCF7 cells damaged in the mixture with week PAM treatment. These results suggested that a specific range existed with the selective effect in the strength of PAM. This work was partly supported by a Grant-in-Aid for Scientific Research on Innovative Areas ``Plasma Medical Innovation'' Grant No. 24108002 and 24108008 from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

  18. Investigating the feasibility of stem cell enrichment mediated by immobilized selectins.

    Charles, Nichola; Liesveld, Jane L; King, Michael R

    2007-01-01

    Hematopoietic stem cell therapy is used to treat both malignant and non-malignant diseases, and enrichment of the hematopoietic stem and progenitor cells (HSPCs) has the potential to reduce the likelihood of graft vs host disease or relapse, potentially fatal complications associated with the therapy. Current commercial HSPC isolation technologies rely solely on the CD34 surface marker, and while they have proven to be invaluable, they can be time-consuming with variable recoveries reported. We propose that selectin-mediated enrichment could prove to be a quick and effective method for recovering HSPCs from adult bone marrow (ABM) on the basis of differences in rolling velocities and independently of CD34 expression. Purified CD34+ ABM cells and the unselected CD34- ABM cells were perfused over immobilized P-, E-, and L-selectin-IgG at physiologic wall shear stresses, and rolling velocities and cell retention data were collected. CD34+ ABM cells generally exhibited lower rolling velocities and higher retention than the unselected CD34- ABM cells on all three selectins. For initial CD34+ ABM cell concentrations ranging from 1% to 5%, we predict an increase in purity ranging from 5.2% to 36.1%, depending on the selectin used. Additionally, selectin-mediated cell enrichment is not limited to subsets of cells with inherent differences in rolling velocities. CD34+ KG1a cells and CD34- HL60 cells exhibited nearly identical rolling velocities on immobilized P-selectin-IgG over the entire range of shear stresses studied. However, when anti-CD34 antibody was co-immobilized with the P-selectin-IgG, the rolling velocity of the CD34+ KG1a cells was significantly reduced, making selectin-mediated cell enrichment a feasible option. Optimal cell enrichment in immobilized selectin surfaces can be achieved within 10 min, much faster than most current commercially available systems.

  19. Cellular radiation effects and hyperthermia: Cytokinetic investigations with stationary phase yeast cells

    Fingerhut, R.; Otto, F.; Oldiges, H.; Kiefer, J.

    1980-01-01

    Wild type diploid yeast, Saccharomyces cerevisiae strain 211, was subjected to 250 kV X-rays or 50 0 C heat treatment for 30 min or to a combination of both. X-ray exposure took place either in air or in nitrogen. Cell number, percentage of budding cells and cell cycle progression was followed for up to 12 h post irradiation. The distribution of cell cycle stages was determined by flow cytofluorometry. All treatments cause a retardation of cell division rate. Hyperthermia leads mainly to a lengthening of G 1 , whereas X-rays arrest the cells reversibly in G 2 . The effect of the combined treatment appears to be merely additive. No selective action of hyperthermia on hypoxic cells was found. (orig.) [de

  20. Global investigation of interleukin-1β signaling in primary β-cells using quantitative phosphoproteomics

    Engholm-Keller, Kasper; Størling, Joachim; Pociot, Flemming

    in β-cells by which this cytokine can modulate cell-matrix interactions during inflammation, a regulation shown in other cell types. Further data analysis is currently ongoing, and the collective results of the experiments will hopefully facilitate additional insights into the effect of IL-1β......Novel Aspect: Global phosphoproteomic analysis of cytokine signaling in primary β-cells Introduction The insulin-producing β-cells of the pancreatic islets of Langerhans are targeted by aberrant immune system responses in diabetes mellitus involving cytokines, especially interleukin-1β (IL-1 β......), which initiate apoptosis of the β-cells. As only limited amounts of primary β-cells can be isolated from model organisms like mouse and rat, global phosphoproteomic analysis of these signaling events by mass spectrometry has generally been unfeasible. We have therefore developed a strategy...

  1. Investigation of functional activity human dental pulp stem cells at acute and chronic pulpitis.

    Ustiashvili, M; Kordzaia, D; Mamaladze, M; Jangavadze, M; Sanodze, L

    2014-09-01

    It is already recognized that together with the other connective tissues organ-specific progenic stem cells are also found in postnatal dental pulp. This group of undifferentiated cells is only 1% of total cell population of the pulp. The aim of the study was the identification of stem cells in human dental pulp, detection of their localization and assessment of functional activity during inflammation process and/or at norm. The obtained results showed that at acute pulpitis the pulp stroma is hypocellular in comparison with the norm but cells proliferative activity is low. CD 133 and NCAM (CD 56) positive stem cells were found in perivascularl space of the pulp stroma and in Hohle layer. At process prolongation and transition to the chronic phase pulp stroma is hypercellular, the cells with large, rounded or oval-shaped nuclei with clear chromatin appear together with fibroblasts. They are distributed as about entire thickness of the stroma as especially Hohle layer. In such cells higher proliferative activity (Ki67 expression) was observed. The cells in the mentioned proliferation phase are intensively marked by CD133, the rate of which is high in Hohle layer and along it. A large number of NCAM (CD 56) positive cells appear in pulp stroma. During pulpitis an involvement of stem cells into the process of reparative dentinogenesis should be conducted stepwise. In acute cases of the disease, stem cell perivascularl mobilization and proliferation and its migration to Hohle layer occur in response to irritation /stimulation. Chronification of the process leads not only to the migration of stem cells to the periphery of the pulp but also s their В«maturationВ» (increase of NCAM expression in the stem cells), which causes an increase the number of dentin producing active odontoblasts and initiation of reparative dentinogenesis.

  2. Investigations on Two Co-C Fixed-Point Cells Prepared at INRIM and LNE-Cnam

    Battuello, M.; Florio, M.; Sadli, M.; Bourson, F.

    2011-08-01

    INRIM and LNE-Cnam agreed to undertake a collaboration aimed to verify, through the use of metal-carbon eutectic fixed-point cells, methods and facilities used for defining the transition temperature of eutectic fixed points and manufacturing procedures of cells. To this purpose and as a first step of the cooperation, a Co-C cell manufactured at LNE-Cnam was measured at INRIM and compared with a local cell. The two cells were of different designs: the INRIM cell of 10 cm3 inner volume and the LNE-Cnam one of 3.9 cm3. The external dimensions of the two cells were noticeably different, namely, 40 mm in length and 24 mm in diameter for the LNE-Cnam cell 3Co4 and 110 mm in length and 42 mm in diameter for the INRIM cell. Consequently, the investigation of the effect of temperature distributions in the heating furnace was undertaken by implementing the cells inside single-zone and three-zone furnaces. The transition temperature of the cell was determined at the two institutes making use of different techniques: at INRIM radiation scales at 900 nm, 950 nm, and 1.6 μm were realized from In to Cu and then used to define T 90(Co-C) by extrapolation. At LNE-Cnam, a radiance comparator based on a grating monochromator was used for the extrapolation from the Cu fixed point. This paper presents a comparative description of the cells and the manufacturing methods and the results in terms of equivalence between the two cells and melting temperatures determined at INRIM and LNE-Cnam.

  3. Investigation of Biophysical Mechanisms in Gold Nanoparticle Mediated Laser Manipulation of Cells Using a Multimodal Holographic and Fluorescence Imaging Setup

    Rakoski, Mirko S.; Heinemann, Dag; Schomaker, Markus; Ripken, Tammo; Meyer, Heiko

    2015-01-01

    Laser based cell manipulation has proven to be a versatile tool in biomedical applications. In this context, combining weakly focused laser pulses and nanostructures, e.g. gold nanoparticles, promises to be useful for high throughput cell manipulation, such as transfection and photothermal therapy. Interactions between laser pulses and gold nanoparticles are well understood. However, it is still necessary to study cell behavior in gold nanoparticle mediated laser manipulation. While parameters like cell viability or perforation efficiency are commonly addressed, the influence of the manipulation process on other essential cell parameters is not sufficiently investigated yet. Thus, we set out to study four relevant cell properties: cell volume and area, ion exchange and cytoskeleton structure after gold nanoparticle based laser manipulation. For this, we designed a multimodal imaging and manipulation setup. 200 nm gold nanoparticles were attached unspecifically to canine cells and irradiated by weakly focused 850 ps laser pulses. Volume and area change in the first minute post laser manipulation was monitored using digital holography. Calcium imaging and cells expressing a marker for filamentous actin (F-actin) served to analyze the ion exchange and the cytoskeleton, respectively. High radiant exposures led to cells exhibiting a tendency to shrink in volume and area, possibly due to outflow of cytoplasm. An intracellular raise in calcium was observed and accompanied by an intercellular calcium wave. This multimodal approach enabled for the first time a comprehensive analysis of the cell behavior in gold nanoparticle mediated cell manipulation. Additionally, this work can pave the way for a better understanding and the evaluation of new applications in the context of cell transfection or photothermal therapy. PMID:25909631

  4. Investigation of biophysical mechanisms in gold nanoparticle mediated laser manipulation of cells using a multimodal holographic and fluorescence imaging setup.

    Stefan Kalies

    Full Text Available Laser based cell manipulation has proven to be a versatile tool in biomedical applications. In this context, combining weakly focused laser pulses and nanostructures, e.g. gold nanoparticles, promises to be useful for high throughput cell manipulation, such as transfection and photothermal therapy. Interactions between laser pulses and gold nanoparticles are well understood. However, it is still necessary to study cell behavior in gold nanoparticle mediated laser manipulation. While parameters like cell viability or perforation efficiency are commonly addressed, the influence of the manipulation process on other essential cell parameters is not sufficiently investigated yet. Thus, we set out to study four relevant cell properties: cell volume and area, ion exchange and cytoskeleton structure after gold nanoparticle based laser manipulation. For this, we designed a multimodal imaging and manipulation setup. 200 nm gold nanoparticles were attached unspecifically to canine cells and irradiated by weakly focused 850 ps laser pulses. Volume and area change in the first minute post laser manipulation was monitored using digital holography. Calcium imaging and cells expressing a marker for filamentous actin (F-actin served to analyze the ion exchange and the cytoskeleton, respectively. High radiant exposures led to cells exhibiting a tendency to shrink in volume and area, possibly due to outflow of cytoplasm. An intracellular raise in calcium was observed and accompanied by an intercellular calcium wave. This multimodal approach enabled for the first time a comprehensive analysis of the cell behavior in gold nanoparticle mediated cell manipulation. Additionally, this work can pave the way for a better understanding and the evaluation of new applications in the context of cell transfection or photothermal therapy.

  5. Detailed investigations of ZnO photoelectrodes preparation for dye sensitized solar cells.

    Marczak, Renata; Werner, Fabian; Ahmad, Rameez; Lobaz, Volodymyr; Guldi, Dirk M; Peukert, Wolfgang

    2011-04-05

    Wurtzite ZnO hexagonal nanopyramids were successfully synthesized in the liquid phase from homogeneous methanolic solutions of zinc acetate and tetramethylammonium hydroxide at an excess of zinc ions. The formation and properties of the nanocrystals were examined as a function of synthesis conditions. No significant influence of the [Zn(2+)]/[OH(-)] ratio was noticed on the final particle size, in spite of increased amounts of OH(-) ions, which tend to accelerate the particle nucleation and growth. Nevertheless, the reactant concentration ratio influences the surface properties of the ZnO nanocrystals. Mesoporous ZnO films were prepared by doctor blading ethanolic pastes containing ZnO nanoparticles and ethyl cellulose onto FTO conductive glass substrate followed by calcination. Additionally, the influence of a plasticizer (triacetin)-used during the paste preparation-on the film quality was investigated. A higher content of ZnO nanoparticles and plasticizer in the pastes improved the film quality. Four different temperatures (i.e., 400, 425, 450, and 475 °C) were used for the film calcination and their influence on the structural properties of the films was characterized. In principle, increasing the calcination temperature goes hand in hand with an increase of particle size, as well as the pore diameter and reduction of the surface area. Suitable mesoporous films were employed as photoanodes in dye sensitized solar cells (DSSCs). In order to assess the effect of the varied parameters on complete DSSC devices-using cis-diisothiocyanato-bis(2,2'-bipyridyl-4,4'-dicarboxylato) ruthenium(II)bis(tetrabutylammonium (N719) as a sensitizer-incident photon to current efficiency (IPCE) and current voltage measurements were carried out. The IPCE measurements confirmed photoinduced electron injection from the dye, reaching IPCE values up to 76%. Furthermore, current-voltage characteristics of complete cells emphasized the importance of the proper preparation methods and

  6. Cellular contractility and substrate elasticity: a numerical investigation of the actin cytoskeleton and cell adhesion.

    Ronan, William; Deshpande, Vikram S; McMeeking, Robert M; McGarry, J Patrick

    2014-04-01

    Numerous experimental studies have established that cells can sense the stiffness of underlying substrates and have quantified the effect of substrate stiffness on stress fibre formation, focal adhesion area, cell traction, and cell shape. In order to capture such behaviour, the current study couples a mixed mode thermodynamic and mechanical framework that predicts focal adhesion formation and growth with a material model that predicts stress fibre formation, contractility, and dissociation in a fully 3D implementation. Simulations reveal that SF contractility plays a critical role in the substrate-dependent response of cells. Compliant substrates do not provide sufficient tension for stress fibre persistence, causing dissociation of stress fibres and lower focal adhesion formation. In contrast, cells on stiffer substrates are predicted to contain large amounts of dominant stress fibres. Different levels of cellular contractility representative of different cell phenotypes are found to alter the range of substrate stiffness that cause the most significant changes in stress fibre and focal adhesion formation. Furthermore, stress fibre and focal adhesion formation evolve as a cell spreads on a substrate and leading to the formation of bands of fibres leading from the cell periphery over the nucleus. Inhibiting the formation of FAs during cell spreading is found to limit stress fibre formation. The predictions of this mutually dependent material-interface framework are strongly supported by experimental observations of cells adhered to elastic substrates and offer insight into the inter-dependent biomechanical processes regulating stress fibre and focal adhesion formation.

  7. Investigation of therapeutic efficiency of phenytoin (PHT) labeled with radioactive 131I in the cancer cell lines

    Cansu Uzaras; Ugur Avcibasi; Hasan Demiroglu; Emin Ilker Medine; Ayfer Yurt KiIcar; Fazilet Zuemruet Biber Mueftueler; Perihan Uenak

    2016-01-01

    The aim of this study is to determine the incorporations of PHT radiolabeled with 131 I ( 131 I-PHT) on U-87 MG, Daoy and A549 cancerous cell lines. For this, cold and radio-labeling studies were carried out. The radiolabeling yield of 131 I-PHT was obtained about 95 %. Subsequently, cell culture studies were carried out and radio-labeling yields of 131 I, 131 I-PHT on U-87 MG, Daoy and A549 cancerous cells were investigated. Cell culture studies demonstrated that the incorporation values of 131 IPHT on the three cell lines decreased with increasing radioactivity. Consequently, 131 I-PHT may be a good radiopharmaceutical for targeting radionuclide therapy of Central Nervous System Tumors. (author)

  8. Investigations on the change of texture of plant cells due to preservative treatments by digital holographic microscopy

    Vora, Priyanka; Anand, Arun

    2014-10-01

    Texture change is observed in preserved fruits and vegetables. Responsible factors for texture change during preservative treatments are cell morphology, cell wall structure, cell turger, water content and some biochemical components, and also the environmental conditions. Digital Holographic microscopy (DHM) is a quantitative phase contrast imaging technique, which provides three dimensional optical thickness profiles of transparent specimen. Using DHM the morphology of plant cells preserved by refrigeration or stored in vinegar or in sodium chloride can be obtained. This information about the spatio-temporal evolution of optical volume and thickness can be an important tool in area of food processing. Also from the three dimensional images, the texture of the cell can be retrieved and can be investigated under varying conditions.

  9. Opto-electro-modulated transient photovoltage and photocurrent system for investigation of charge transport and recombination in solar cells.

    Shi, Jiangjian; Li, Dongmei; Luo, Yanhong; Wu, Huijue; Meng, Qingbo

    2016-12-01

    An opto-electro-modulated transient photovoltage/photocurrent system has been developed to probe microscopic charge processes of a solar cell in its adjustable operating conditions. The reliability of this system is carefully determined by electric circuit simulations and experimental measurements. Using this system, the charge transport, recombination and storage properties of a conventional multicrystalline silicon solar cell under different steady-state bias voltages, and light illumination intensities are investigated. This system has also been applied to study the influence of the hole transport material layer on charge extraction and the microscopic charge processes behind the widely considered photoelectric hysteresis in perovskite solar cells.

  10. The investigation of influence of accelerated electrons on SiO2 used in silicon solar cells

    Abdullaev, G.B.; Bakirov, M.Ya; Akhmedov, G.M.; Safarov, N.A.; Safarova, F.D.

    1994-01-01

    The process of radiation defects production in enlightened SiO 2 layers coated on silicon solar cells was studied. During irradiation the silicon solar cells with enlightened layers radiation defects are formed both in silicon and SiO 2 thus making worse photo energetic parameters of cells. For investigation of radiation effects formed under irradiation by electrons with 5 MeV energy and cobalt-60 gamma-rays photoluminescence, absorption spectra and electron spin resonance methods were used. It is supposed that main radiation defects in silicon dioxide are E'-centers and oxygen vacancies. (A.D. Avezov). 10 refs.; 2 figs

  11. An investigation of hydrogen storage methods for fuel cell operation with man-portable equipment

    Browning, D [Defence Evaluation and Research Agency, Haslar (United Kingdom); Jones, P [Defence Evaluation and Research Agency, Haslar (United Kingdom); Packer, K [Defence Evaluation and Research Agency, Haslar (United Kingdom)

    1997-03-01

    Air breathing proton exchange membrane fuel cells (PEMFC) are being considered as a power source for man-portable equipment, such as army radios. In addition to the weight and volume of the fuel cell itself, the device producing hydrogen with which to fuel the cell is also of crucial importance. This paper describes a number of hydrogen storage methods and discusses their applicability to man-portable equipment. (orig.)

  12. Investigation of in vitro bone cell adhesion and proliferation on Ti using direct current stimulation

    Bodhak, Subhadip; Bose, Susmita; Kinsel, William C.; Bandyopadhyay, Amit

    2012-01-01

    Our objective was to establish an in vitro cell culture protocol to improve bone cell attachment and proliferation on Ti substrate using direct current stimulation. For this purpose, a custom made electrical stimulator was developed and a varying range of direct currents, from 5 to 25 μA, was used to study the current stimulation effect on bone cells cultured on conducting Ti samples in vitro. Cell–material interaction was studied for a maximum of 5 days by culturing with human fetal osteoblast cells (hFOB). The direct current was applied in every 8 h time interval and the duration of electrical stimulation was kept constant at 15 min for all cases. In vitro results showed that direct current stimulation significantly favored bone cell attachment and proliferation in comparison to nonstimulated Ti surface. Immunochemistry and confocal microscopy results confirmed that the cell adhesion was most pronounced on 25 μA direct current stimulated Ti surfaces as hFOB cells expressed higher vinculin protein with increasing amount of direct current. Furthermore, MTT assay results established that cells grew 30% higher in number under 25 μA electrical stimulation as compared to nonstimulated Ti surface after 5 days of culture period. In this work we have successfully established a simple and cost effective in vitro protocol offering easy and rapid analysis of bone cell–material interaction which can be used in promotion of bone cell attachment and growth on Ti substrate using direct current electrical stimulation in an in vitro model. - Highlights: ► D.C. stimulation was used to enhance in vitro bone cell adhesion and proliferation. ► Cells cultured on Ti were stimulated by using a custom made electrical stimulator. ► Optimization was performed by using a varying range of direct currents ∼ 5 to 25 μA. ► 25 μA stimulation was found most beneficial for promotion of cell adhesion/growth.

  13. Field and laboratory emission cell automation and control system for investigating surface chemistry reactions

    Flemmer, Michael M.; Ham, Jason E.; Wells, J. R.

    2007-01-01

    A novel system [field and laboratory emission cell (FLEC) automation and control system] has been developed to deliver ozone to a surface utilizing the FLEC to simulate indoor surface chemistry. Ozone, humidity, and air flow rate to the surface were continuously monitored using an ultraviolet ozone monitor, humidity, and flow sensors. Data from these sensors were used as feedback for system control to maintain predetermined experimental parameters. The system was used to investigate the chemistry of ozone with α-terpineol on a vinyl surface over 72h. Keeping all other experimental parameters the same, volatile organic compound emissions from the vinyl tile with α-terpineol were collected from both zero and 100ppb(partsper109) ozone exposures. System stability profiles collected from sensor data indicated experimental parameters were maintained to within a few percent of initial settings. Ozone data from eight experiments at 100ppb (over 339h) provided a pooled standard deviation of 1.65ppb and a 95% tolerance of 3.3ppb. Humidity data from 17 experiments at 50% relative humidity (over 664h) provided a pooled standard deviation of 1.38% and a 95% tolerance of 2.77%. Data of the flow rate of air flowing through the FLEC from 14 experiments at 300ml/min (over 548h) provided a pooled standard deviation of 3.02ml/min and a 95% tolerance range of 6.03ml/min. Initial experimental results yielded long term emissions of ozone/α-terpineol reaction products, suggesting that surface chemistry could play an important role in indoor environments.

  14. Investigation and Taguchi Optimization of Microbial Fuel Cell Salt Bridge Dimensional Parameters

    Sarma, Dhrupad; Barua, Parimal Bakul; Dey, Nabendu; Nath, Sumitro; Thakuria, Mrinmay; Mallick, Synthia

    2018-01-01

    One major problem of two chamber salt bridge microbial fuel cells (MFCs) is the high resistance offered by the salt bridge to anion flow. Many researchers who have studied and optimized various parameters related to salt bridge MFC, have not shed much light on the effect of salt bridge dimensional parameters on the MFC performance. Therefore, the main objective of this research is to investigate the effect of length and cross sectional area of salt bridge and the effect of solar radiation and atmospheric temperature on MFC current output. An experiment has been designed using Taguchi L9 orthogonal array, taking length and cross sectional area of salt bridge as factors having three levels. Nine MFCs were fabricated as per the nine trial conditions. Trials were conducted for 3 days and output current of each of the MFCs along with solar insolation and atmospheric temperature were recorded. Analysis of variance shows that salt bridge length has significant effect both on mean (with 53.90% contribution at 95% CL) and variance (with 56.46% contribution at 87% CL), whereas the effect of cross sectional area of the salt bridge and the interaction of these two factors is significant on mean only (with 95% CL). Optimum combination was found at 260 mm salt bridge length and 506.7 mm2 cross sectional area with 4.75 mA of mean output current. The temperature and solar insolation data when correlated with each of the MFCs average output current, revealed that both external factors have significant impact on MFC current output but the correlation coefficient varies from MFC to MFC depending on salt bridge dimensional parameters.

  15. An investigation into the activation and deactivation of chlorinated hydrocarbons to genotoxins in metabolically competent human cells.

    Doherty, A T; Ellard, S; Parry, E M; Parry, J M

    1996-05-01

    We have investigated the induction of micronuclei by 15 chlorinated hydrocarbons in the cytochalasin B-blocked micronucleus assay utilizing genetically engineered cell lines. The human lymphoblastoid cell line AHH-1, with native cytochrome CYP1A1 activity, the MCL-5 cell line, which stably expresses cDNAs encoding human CYP1A2, 2A6, 3A4, 2E1 and microsomal epoxide hydrolase, and the h2E1 cell line, containing a cDNA for CYP2E1, were used in this study. We have demonstrated the induction of kinetochore-positive micronuclei by two chlorinated solvents, 2,3-dichlorobutane and 1,1, 2-trichloroethane, in the metabolically competent cell lines MCL-5 and h2E1. The MCL-5 and h2E1 cell lines have in addition shown the capacity to produce metabolites in the presence of methylene chloride, carbon tetrachloride, 1,2,3-trichloropropane, tetrachloroethylene, toluene and n-hexane, wich yield elevated micronucleus frequencies compared with the parental cell line AHH-1. Hexachloroethane failed to induce micronuclei in any of the cell lines and 1,2-dichloroethane and 1-chlorohexane induced micronuclei without the requirement for metabolic activation in all three cell lines. The MCL-5 cell line exhibited reduced micronucleus frequencies compared with the AHH-1 and h2E1 cell lines following exposure to 1,2-dichloroethylene, 1,3-dichloropropane, 1,1, 1-trichloroethane and 1,2,3-trichloropropane. The methodology used has shown the ability of metabolically competent cell lines expressing cDNAs encoding the cytochrome P450 isoenzymes to metabolize halogenated hydrocarbons to genotoxic species, including both clastogens and aneugens. The biotransformation of chemicals to aneugenic species has not previously been demonstrated.

  16. Computational simulation of static/cyclic cell stimulations to investigate mechanical modulation of an individual mesenchymal stem cell using confocal microscopy

    Alihemmati, Zakieh; Vahidi, Bahman; Haghighipour, Nooshin; Salehi, Mohammad

    2017-01-01

    It has been found that cells react to mechanical stimuli, while the type and magnitude of these cells are different in various physiological and pathological conditions. These stimuli may affect cell behaviors via mechanotransduction mechanisms. The aim of this study is to evaluate mechanical responses of a mesenchymal stem cell (MSC) to a pressure loading using finite elements method (FEM) to clarify procedures of MSC mechanotransduction. The model is constructed based on an experimental set up in which statics and cyclic compressive loads are implemented on a model constructed from a confocal microscopy 3D image of a stem cell. Both of the applied compressive loads are considered in the physiological loading regimes. Moreover, a viscohyperelastic material model was assumed for the cell through which the finite elements simulation anticipates cell behavior based on strain and stress distributions in its components. As a result, high strain and stress values were captured from the viscohyperelastic model because of fluidic behavior of cytosol when compared with the obtained results through the hyperelastic models. It can be concluded that the generated strain produced by cyclic pressure is almost 8% higher than that caused by the static load and the von Mises stress distribution is significantly increased to about 150 kPa through the cyclic loading. In total, the results does not only trace the efficacy of an individual 3D model of MSC using biomechanical experiments of cell modulation, but these results provide knowledge in interpretations from cell geometry. The current study was performed to determine a realistic aspect of cell behavior. - Graphical abstract: Based on confocal microscopy images and through finite elements analysis, we simulate mechanical behavior of the stem cell components (the cell membrane, cytoplasm and nucleus) under a compressive load. A major novelty of this investigation is the usage of viscohyperelastic behavior for the realistic stem

  17. Investigation of microwave transitions and nonlinear magneto-optical rotation in anti-relaxation-coated cells

    Budker, D.; Hollberg, L.; Kimball, D.F.; Kitching, J.; Pustclny, S.; Robinson, H.G.; Yashchuk, V.V.

    2004-01-01

    Using laser optical pumping, widths and frequency shifts are determined for microwave transitions between the components of the ground-state hyperfine structure for 85 Rb and 87 Rb atoms contained in vapor cells with alkane anti-relaxation coatings. The results are compared with data on Zeeman relaxation obtained in nonlinear magneto-optical rotation (NMOR) experiments, a comparison important for quantitative understanding of spin-relaxation mechanisms in coated cells. By comparing cells manufactured over a forty-year period we demonstrate the long-term stability of coated cells, which may be useful for atomic clocks and magnetometers

  18. Investigation of microwave transitions and nonlinear magneto-optical rotation in anti-relaxation-coated cells

    Budker, Dmitry; Hollberg, Leo; Kimball, Derek F.; Kitching, J.; Pustelny, Szymon; Yashchuk, Valeriy V.

    2004-01-01

    Using laser optical pumping, widths and frequency shifts are determined for microwave transitions between ground-state hyperfine components of 85 Rb and 87 Rb atoms contained in vapor cells with alkane anti-relaxation coatings. The results are compared with data on Zeeman relaxation obtained in nonlinear magneto-optical rotation (NMOR) experiments, a comparison important for quantitative understanding of spin-relaxation mechanisms in coated cells. By comparing cells manufactured over a forty-year period we demonstrate the long-term stability of coated cells, an important property for atomic clocks and magnetometers

  19. Laser-capture micro dissection combined with next-generation sequencing analysis of cell type-specific deafness gene expression in the mouse cochlea.

    Nishio, Shin-Ya; Takumi, Yutaka; Usami, Shin-Ichi

    2017-05-01

    Cochlear implantation (CI), which directly stimulates the cochlear nerves, is the most effective and widely used medical intervention for patients with severe to profound sensorineural hearing loss. The etiology of the hearing loss is speculated to have a major influence of CI outcomes, particularly in cases resulting from mutations in genes preferentially expressed in the spiral ganglion region. To elucidate precise gene expression levels in each part of the cochlea, we performed laser-capture micro dissection in combination with next-generation sequencing analysis and determined the expression levels of all known deafness-associated genes in the organ of Corti, spiral ganglion, lateral wall, and spiral limbs. The results were generally consistent with previous reports based on immunocytochemistry or in situ hybridization. As a notable result, the genes associated with many kinds of syndromic hearing loss (such as Clpp, Hars2, Hsd17b4, Lars2 for Perrault syndrome, Polr1c and Polr1d for Treacher Collins syndrome, Ndp for Norrie Disease, Kal for Kallmann syndrome, Edn3 and Snai2 for Waardenburg Syndrome, Col4a3 for Alport syndrome, Sema3e for CHARGE syndrome, Col9a1 for Sticker syndrome, Cdh23, Cib2, Clrn1, Pcdh15, Ush1c, Ush2a, Whrn for Usher syndrome and Wfs1 for Wolfram syndrome) showed higher levels of expression in the spiral ganglion than in other parts of the cochlea. This dataset will provide a base for more detailed analysis in order to clarify gene functions in the cochlea as well as predict CI outcomes based on gene expression data. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  20. High throughput, cell type-specific analysis of key proteins in human endometrial biopsies of women from fertile and infertile couples

    Leach, Richard E.; Jessmon, Philip; Coutifaris, Christos; Kruger, Michael; Myers, Evan R.; Ali-Fehmi, Rouba; Carson, Sandra A.; Legro, Richard S.; Schlaff, William D.; Carr, Bruce R.; Steinkampf, Michael P.; Silva, Susan; Leppert, Phyllis C.; Giudice, Linda; Diamond, Michael P.; Armant, D. Randall

    2012-01-01

    BACKGROUND Although histological dating of endometrial biopsies provides little help for prediction or diagnosis of infertility, analysis of individual endometrial proteins, proteomic profiling and transcriptome analysis have suggested several biomarkers with altered expression arising from intrinsic abnormalities, inadequate stimulation by or in response to gonadal steroids or altered function due to systemic disorders. The objective of this study was to delineate the developmental dynamics of potentially important proteins in the secretory phase of the menstrual cycle, utilizing a collection of endometrial biopsies from women of fertile (n = 89) and infertile (n = 89) couples. METHODS AND RESULTS Progesterone receptor-B (PGR-B), leukemia inhibitory factor, glycodelin/progestagen-associated endometrial protein (PAEP), homeobox A10, heparin-binding EGF-like growth factor, calcitonin and chemokine ligand 14 (CXCL14) were measured using a high-throughput, quantitative immunohistochemical method. Significant cyclic and tissue-specific regulation was documented for each protein, as well as their dysregulation in women of infertile couples. Infertile patients demonstrated a delay early in the secretory phase in the decline of PGR-B (P localization provided important insights into the potential roles of these proteins in normal and pathological development of the endometrium that is not attainable from transcriptome analysis, establishing a basis for biomarker, diagnostic and targeted drug development for women with infertility. PMID:22215622

  1. Role of pH in determining the cell-type-specific residual activity of glucocerebrosidase in type 1 Gaucher disease

    van Weely, S.; van den Berg, M.; Barranger, J. A.; Sa Miranda, M. C.; Tager, J. M.; Aerts, J. M.

    1993-01-01

    The properties of control and 370Asn-->Ser glucocerebrosidase, the frequently encountered mutated form of the enzyme in type 1 Gaucher disease, were studied in vitro as well as in situ. The catalytic properties of purified 370Asn-->Ser glucocerebrosidase were highly dependent on the assay

  2. Thermal investigation of lithium-ion battery module with different cell arrangement structures and forced air-cooling strategies

    Wang, Tao; Tseng, K.J.; Zhao, Jiyun; Wei, Zhongbao

    2014-01-01

    Highlights: • Three-dimensional CFD model with forced air cooling are developed for battery modules. • Impact of different air cooling strategies on module thermal characteristics are investigated. • Impact of different model structures on module thermal responses are investigated. • Effect of inter-cell spacing on cell thermal characteristics are also studied. • The optimal battery module structure and air cooling strategy is recommended. - Abstract: Thermal management needs to be carefully considered in the lithium-ion battery module design to guarantee the temperature of batteries in operation within a narrow optimal range. This article firstly explores the thermal performance of battery module under different cell arrangement structures, which includes: 1 × 24, 3 × 8 and 5 × 5 arrays rectangular arrangement, 19 cells hexagonal arrangement and 28 cells circular arrangement. In addition, air-cooling strategies are also investigated by installing the fans in the different locations of the battery module to improve the temperature uniformity. Factors that influence the cooling capability of forced air cooling are discussed based on the simulations. The three-dimensional computational fluid dynamics (CFD) method and lumped model of single cell have been applied in the simulation. The temperature distributions of batteries are quantitatively described based on different module patterns, fan locations as well as inter-cell distance, and the conclusions are arrived as follows: when the fan locates on top of the module, the best cooling performance is achieved; the most desired structure with forced air cooling is cubic arrangement concerning the cooling effect and cost, while hexagonal structure is optimal when focus on the space utilization of battery module. Besides, the optimized inter-cell distance in battery module structure has been recommended

  3. HUMAN CAPSULE EPITHELIAL-CELL DEGENERATION A LM, SEM AND TEM INVESTIGATION

    JONGEBLOED, WL; KALICHARAN, D; WORST, JGF

    1993-01-01

    The degeneration of the capsule epithelium of cataractous lenses has been studied with LM, SEM on TEM with emphases on TEM. The observed degeneration of the epithelial cells can be described as follows: The cell nucleus becomes picnotic and desintegrates as result of change of the chromatin.

  4. Investigation of Ruthenium Dissolution in Advanced Membrane Electrode Assemblies for Direct Methanol Based Fuel Cell Stacks

    Valdez, Thomas I.; Firdosy, S.; Koel, B. E.; Narayanan, S. R.

    2005-01-01

    Dissolution of ruthenium was observed in the 80-cell stack. Duration testing was performed in single cell MEAs to determine the pathway of cell degradation. EDAX analysis on each of the single cell MEAs has shown that the Johnson Matthey commercial catalyst is stable in DMFC operation for 250 hours, no ruthenium dissolution was observed. Changes in the hydrophobicity of the cathode backing papers was minimum. Electrode polarization analysis revealed that the MEA performance loss is attributed to changes in the cathode catalyst layer. Ruthenium migration does not seem to occur during cell operation but can occur when methanol is absent from the anode compartment, the cathode compartment has access to air, and the cells in the stack are electrically connected to a load (Shunt Currents). The open-to-air cathode stack design allowed for: a) The MEAs to have continual access to oxygen; and b) The stack to sustain shunt currents. Ruthenium dissolution in a DMFC stack can be prevented by: a) Developing an internally manifolded stacks that seal reactant compartments when not in operation; b) Bringing the cell voltages to zero quickly when not in operation; and c) Limiting the total number of cells to 25 in an effort to limit shunt currents.

  5. Investigation on Cell Proliferation with a New Antibody against Thymidine Kinase 1

    Naining Wang

    2001-01-01

    Full Text Available The cytosolic thymidine kinase 1 (TK1 is one of the enzymes involved in DNA replication. Based on biochemical studies, TK1 is activated at late G1 of cell cycle, and its activity correlates with the cell proliferation. We have developed a polyclonal anti‐TK1 antibody against a synthetic peptide from the C‐terminus of human TK1. Using this antibody, here we demonstrate the exclusive location of TK1 in the cytoplasm of cells. Cell cycle dependent TK1 expression was studied by simultaneous fluorescence staining for TK1 and bromodeoxyuridine, by using elutriated cells, and by quantitation of the amount TK1 in relation to the cellular DNA content. TK1, which was strongly expressed in the cells in S+G2 period, raised at late G1 and decreased during mitosis. The amount of TK1 increased three folds from late G1 to G2. TK1 positive cells were demonstrated in areas of proliferation activity of various normal and malignant tissues. The new anti‐TK1 antibody works in archival specimens and is a specific marker of cell proliferation.

  6. Investigation of Anti-Relaxation Coatings for Alkali-Metal Vapor Cells using Surface Science Techniques

    2011-02-01

    addition to the inside surface of the cells. In order to avoid exposure to air, the cells were broken open inside a glovebag containing an argon ...unsaturated bonds increase the polar- izability of the surface, and effective coatings have long been assumed to require low polarizability to enable

  7. Development and Characterisation of a Novel NF-κB Reporter Cell Line for Investigation of Neuroinflammation

    Marie-Theres Zeuner

    2017-01-01

    Full Text Available Aberrant activation of the transcription factor NF-κB, as well as uncontrolled inflammation, has been linked to autoimmune diseases, development and progression of cancer, and neurological disorders like Alzheimer’s disease. Reporter cell lines are a valuable state-of-the art tool for comparative analysis of in vitro drug screening. However, a reporter cell line for the investigation of NF-κB-driven neuroinflammation has not been available. Thus, we developed a stable neural NF-κB-reporter cell line to assess the potency of proinflammatory molecules and peptides, as well as anti-inflammatory pharmaceuticals. We used lentivirus to transduce the glioma cell line U251-MG with a tandem NF-κB reporter construct containing GFP and firefly luciferase allowing an assessment of NF-κB activity via fluorescence microscopy, flow cytometry, and luminometry. We observed a robust activation of NF-κB after exposure of the reporter cell line to tumour necrosis factor alpha (TNFα and amyloid-β peptide [1-42] as well as to LPS derived from Salmonella minnesota and Escherichia coli. Finally, we demonstrate that the U251-NF-κB-GFP-Luc reporter cells can be used for assessing the anti-inflammatory potential of pharmaceutical compounds using Bay11-7082 and IMD0354. In summary, our newly generated cell line is a robust and cost-efficient tool to study pro- and anti-inflammatory potential of drugs and biologics in neural cells.

  8. Investigation of the metabolism of ergot alkaloids in cell culture by fourier transformation mass spectrometry.

    Mulac, Dennis; Grote, Anna-Karina; Kleigrewe, Karin; Humpf, Hans-Ulrich

    2011-07-27

    Ergot alkaloids are known toxic secondary metabolites of the fungus Claviceps purpurea occurring in various grains, especially rye products. The liver is responsible for converting the ergot alkaloids into metabolites; however, the toxic impact of these end products of metabolism is still unknown. The aim of this study was to analyze the metabolism of ergot alkaloids in colon and liver cell lines (HT-29, HepG2), as well as in human primary renal cells (RPTEC). It was shown that cells in vitro are able to metabolize ergot alkaloids, forming a variety of metabolic compounds. Significant differences between the used cell types could be identified, and a suitable model system was established using HT-29 cells, performing an intensive metabolism to hydroxylated metabolites. The formed substances were analyzed by coupling of high-performance liquid chromatography with fluorescence detection and Fourier transformation mass spectrometry (HPLC-FLD-FTMS) as a powerful tool to identify known and unknown metabolites.

  9. Dye-sensitized solar cells and solar module using polymer electrolytes: Stability and performance investigations

    Jilian Nei de Freitas

    2006-01-01

    Full Text Available We present recent results on solid-state dye-sensitized solar cell research using a polymer electrolyte based on a poly(ethylene oxide derivative. The stability and performance of the devices have been improved by a modification in the method of assembly of the cells and by the addition of plasticizers in the electrolyte. After 30 days of solar irradiation (100 mW cm-2 no changes in the cell's efficiency were observed using this new method. The effect of the active area size on cell performance and the first results obtained for the first solar module composed of 4.5 cm2 solid-state solar cells are also presented.

  10. Humidity-dependent bacterial cells functional morphometry investigations using atomic force microscope.

    Nikiyan, Hike; Vasilchenko, Alexey; Deryabin, Dmitry

    2010-01-01

    The effect of a relative humidity (RH) in a range of 93-65% on morphological and elastic properties of Bacillus cereus and Escherichia coli cells was evaluated using atomic force microscopy. It is shown that gradual dehumidification of bacteria environment has no significant effect on cell dimensional features and considerably decreases them only at 65% RH. The increasing of the bacteria cell wall roughness and elasticity occurs at the same time. Observed changes indicate that morphological properties of B. cereus are rather stable in wide range of relative humidity, whereas E. coli are more sensitive to drying, significantly increasing roughness and stiffness parameters at RH cell wall structure of gram-positive and gram-negative bacterial cells.

  11. Humidity-Dependent Bacterial Cells Functional Morphometry Investigations Using Atomic Force Microscope

    Hike Nikiyan

    2010-01-01

    Full Text Available The effect of a relative humidity (RH in a range of 93–65% on morphological and elastic properties of Bacillus cereus and Escherichia coli cells was evaluated using atomic force microscopy. It is shown that gradual dehumidification of bacteria environment has no significant effect on cell dimensional features and considerably decreases them only at 65% RH. The increasing of the bacteria cell wall roughness and elasticity occurs at the same time. Observed changes indicate that morphological properties of B. cereus are rather stable in wide range of relative humidity, whereas E. coli are more sensitive to drying, significantly increasing roughness and stiffness parameters at RH ≤ 84% RH. It is discussed the dependence of the response features on differences in cell wall structure of gram-positive and gram-negative bacterial cells.

  12. Comprehensive Investigation of Silver Nanoparticle/Aluminum Electrodes for Copper Indium Sulfide/Polymer Hybrid Solar Cells

    Arar, Mario; Pein, Andreas; Haas, Wernfried

    2012-01-01

    ,1,3-benzothiadiazole)] (PSiF-DBT) nanocomposite solar cells, which improves the fill factor compared to pure aluminum electrodes. A comprehensive structural investigation was performed by means of transmission electron microscopy and time-of-flight secondary ion mass spectrometry revealing the presence of silver...... nanoparticles in an aluminum oxide matrix between the absorber layer and the aluminum cathode. In combination with complementary optical investigations, the origin of the improvement is ascribed to a facilitated charge extraction....

  13. An in vitro investigation of immunomodulatory properties of Lactobacillus plantarum and L. delbrueckii cells and their extracellular polysaccharides.

    Kishimoto, Mana; Nomoto, Ryohei; Mizuno, Masashi; Osawa, Ro

    2017-01-01

    Many probiotic lactobacilli and their extracellular polysaccharides (EPS) have beneficial immunological properties. However, it is unclear how they elicit the host immune response. We thus investigated the immunological properties of UV-killed Lactobacillus delbrueckii TU-1 and L. plantarum KM-9 cells as well as their extracellular polysaccharides (EPSs). High-performance liquid chromatography and ion exchange chromatography analyses showed that their EPSs differ in sugar composition and sugar fractionation. The immunological properties were evaluated in a semi-intestinal model using a Transwell co-culture system that employed human intestinal epithelial (Caco-2) cells on the apical side and murine macrophage (RAW264.7) cells on the basolateral side. The UV-killed cells and EPSs were added to the apical side to allow direct contact with Caco-2 cells and incubated for 6 hr. After incubation, the amounts of tumor necrosis factor-α and several cytokines released by RAW264.7 or Caco-2 cells were quantified by cytotoxic activity on L929 cells (murine fibrosarcoma cell line) and quantitative reverse-transcriptase PCR. We found that the UV-killed cells and their EPSs had immunological effects on RAW264.7 cells via Caco-2 cells. The RAW264.7 cells showed different cytokine production profiles when treated with UV-killed cells and EPSs. The UV-killed cells and EPSs promoted a Th1-type cellular response. Furthermore, we found that the UV-killed cells sent positive signals through Toll-like receptor (TLR) 2. Meanwhile, neither EPS sent a positive signal through TLR4 and TLR2. This evidence suggests that both UV-killed cells of the lactobacillus strains and their EPSs trigger a Th1-type immune response in a human host, with the former triggering the response via the TLRs expressed on its epithelium and the latter employing a mechanism yet to be determined, possibly involving a novel receptor that is designed to recognize specific patterns of repeating sugar in the EPSs.

  14. Strategy for selecting disposable bags for cell culture media applications based on a root-cause investigation.

    Wood, Joseph; Mahajan, Ekta; Shiratori, Masaru

    2013-01-01

    The use of disposable bags for cell culture media storage has grown significantly in the past decade. Some of the key advantages of using disposable bags relative to non-disposable containers include increased product throughput, decreased cleaning validation costs, reduced risk of cross contamination and lower facility costs. As the scope of use of disposable bags for cell culture applications increases, problematic bags and scenarios should be identified and addressed to continue improving disposables technologies and meet the biotech industry's needs. In this article, we examine a cell culture application wherein media stored in disposable bags is warmed at 37°C before use for cell culture operations. A problematic bag film was identified through a prospective and retrospective cell culture investigation. The investigation provided information on the scope and variation of the issue with respect to different Chinese hamster ovary (CHO) cell lines, cell culture media, and application-specific parameters. It also led to the development of application-specific test methods and enabled a strategy for disposable bag film testing. The strategy was implemented for qualifying an alternative bag film for use in our processes. In this test strategy, multiple lots of 13 bag film types, encompassing eight vendors were evaluated using a three round, cell culture-based test strategy. The test strategy resulted in the determination of four viable bag film options based on the technical data. The results of this evaluation were used to conclude that a volatile or air-quenched compound, likely generated by gamma irradiation of the problematic bag film, negatively impacted cell culture performance. © 2013 American Institute of Chemical Engineers.

  15. Analysis of Individuals from a Dengue-Endemic Region Helps Define the Footprint and Repertoire of Antibodies Targeting Dengue Virus 3 Type-Specific Epitopes.

    Andrade, Daniela V; Katzelnick, Leah C; Widman, Doug G; Balmaseda, Angel; de Silva, Aravinda M; Baric, Ralph S; Harris, Eva

    2017-09-19

    The four dengue virus serotypes (DENV1 to 4) cause dengue, a major public health problem worldwide. Individuals exposed to primary DENV infections develop serotype-specific neutralizing antibodies, including strongly neutralizing antibodies targeting quaternary epitopes. To date, no studies have measured the levels and kinetics of serum antibodies directed to such epitopes among populations in regions where dengue is endemic. Here, we use a recombinant DENV4 (rDENV4/3-M14) displaying a major DENV3 type-specific quaternary epitope recognized by human monoclonal antibody 5J7 to measure the proportion, magnitude, and kinetics of DENV3 type-specific neutralizing antibody responses targeting this epitope. Primary DENV3 sera from 30 individuals in a dengue hospital-based study in Nicaragua were studied 3, 6, 12, and 18 months post-infection, alongside samples collected annually 1 to 4 years post-primary DENV3 infection from 10 individuals in a cohort study in Nicaragua. We found substantial individual variation in the proportion of DENV3 type-specific neutralizing antibody titers attributed to the 5J7 epitope (range, 0 to 100%), with the mean significantly increasing from 22.6% to 41.4% from 3 to 18 months. We extended the transplanted DENV3 5J7 epitope on the virion (rDENV4/3-M16), resulting in increased recognition in several individuals, helping define the footprint of the epitope. However, 37% and 13% of the subjects still showed little to no recognition of the 5J7 epitope at 3 and 18 months, respectively, indicating that one or more additional DENV3 type-specific epitopes exist. Overall, this study demonstrates how DENV-immune plasma from populations from areas of endemicity, when coupled with structurally guided recombinant viruses, can help characterize the epitope-specific neutralizing antibody response in natural DENV infections, with direct implications for design and evaluation of dengue vaccines. IMPORTANCE The four serotypes of dengue virus cause dengue

  16. A micro-Raman spectroscopic investigation of leukemic U-937 cells in aged cultures

    Fazio, Enza; Trusso, Sebastiano; Franco, Domenico; Nicolò, Marco Sebastiano; Allegra, Alessandro; Neri, Fortunato; Musolino, Caterina; Guglielmino, Salvatore P. P.

    2016-04-01

    Recently it has been shown that micro-Raman spectroscopy combined with multivariate analysis is able to discriminate among different types of tissues and tumoral cells by the detection of significant alterations and/or reorganizations of complex biological molecules, such as nucleic acids, lipids and proteins. Moreover, its use, being in principle a non-invasive technique, appears an interesting clinical tool for the evaluation of the therapeutical effects and of the disease progression. In this work we analyzed molecular changes in aged cultures of leukemia model U937 cells with respect to fresh cultures of the same cell line. In fact, structural variations of individual neoplastic cells on aging may lead to a heterogeneous data set, therefore falsifying confidence intervals, increasing error levels of analysis and consequently limiting the use of Raman spectroscopy analysis. We found that the observed morphological changes of U937 cells corresponded to well defined modifications of the Raman contributions in selected spectral regions, where markers of specific functional groups, useful to characterize the cell state, are present. A detailed subcellular analysis showed a change in cellular organization as a function of time, and correlated to a significant increase of apoptosis levels. Besides the aforementioned study, Raman spectra were used as input for principal component analysis (PCA) in order to detect and classify spectral changes among U937 cells.

  17. An investigation of the shedding of macromolecules from the Ehrlich mouse ascites tumor cell

    Edwards, E.H.

    1984-01-01

    The spontaneous release, or shedding, of cell surface components into the extracellular medium may be important in the determination of several features of the cancer cell phenotype. The release of macromolecules from the Erhlich mouse ascites tumor cell was studied under a variety of experimental conditions to elucidate the origin and the underlying mechanisms of release. The extrinsic macromolecules are a diverse group with apparent molecular weights ranging from 13,500 to 400,000 daltons. External labeling of the cell surface with tritiated 4,4'-diisothiocyano-1,2-diphenylethane-2,2-disulfonic acid ([ 3 H]H 2 DIDS) reveals a slow loss of labeled components at 4 degrees C, while at 21 degrees C and 37 degrees C an initial rapid loss is followed by a slower release. In vitro metabolic labeling with [1- 14 C]-D-glucosamine hydrochloride, D-[2- 3 H]-mannose and various [ 3 H]-L-amino acids results in the appearance of labeled macromolecules in the medium suggesting tumor, not mouse, origin. These data suggest that the extrinsic macromolecules originate from the cell surface. Macromolecules are shed by a temperature and pH sensitive process. These results suggest that a limited proteolytic digestion, or sublethal autolysis, of the cell surface may occur in this system. The macromolecules shed by the Ehrlich cell originate from the surface and are probably released by sublethal autolysis, direct secretion and a passive process

  18. Infrared microspectroscopy: a multiple-screening platform for investigating single-cell biochemical perturbations upon prion infection.

    Didonna, Alessandro; Vaccari, Lisa; Bek, Alpan; Legname, Giuseppe

    2011-03-16

    Prion diseases are a group of fatal neurodegenerative disorders characterized by the accumulation of prions in the central nervous system. The pathogenic prion (PrP(Sc)) possesses the capability to convert the host-encoded cellular isoform of the prion protein, PrP(C), into nascent PrP(Sc). The present work aims at providing novel insight into cellular response upon prion infection evidenced by synchrotron radiation infrared microspectroscopy (SR-IRMS). This non-invasive, label-free analytical technique was employed to investigate the biochemical perturbations undergone by prion infected mouse hypothalamic GT1-1 cells at the cellular and subcellular level. A decrement in total cellular protein content upon prion infection was identified by infrared (IR) whole-cell spectra and validated by bicinchoninic acid assay and single-cell volume analysis by atomic force microscopy (AFM). Hierarchical cluster analysis (HCA) of IR data discriminated between infected and uninfected cells and allowed to deduce an increment of lysosomal bodies within the cytoplasm of infected GT1-1 cells, a hypothesis further confirmed by SR-IRMS at subcellular spatial resolution and fluorescent microscopy. The purpose of this work, therefore, consists of proposing IRMS as a powerful multiscreening platform, drawing on the synergy with conventional biological assays and microscopy techniques in order to increase the accuracy of investigations performed at the single-cell level.

  19. Infrared Microspectroscopy: A Multiple-Screening Platform for Investigating Single-Cell Biochemical Perturbations upon Prion Infection

    2011-01-01

    Prion diseases are a group of fatal neurodegenerative disorders characterized by the accumulation of prions in the central nervous system. The pathogenic prion (PrPSc) possesses the capability to convert the host-encoded cellular isoform of the prion protein, PrPC, into nascent PrPSc. The present work aims at providing novel insight into cellular response upon prion infection evidenced by synchrotron radiation infrared microspectroscopy (SR-IRMS). This non-invasive, label-free analytical technique was employed to investigate the biochemical perturbations undergone by prion infected mouse hypothalamic GT1-1 cells at the cellular and subcellular level. A decrement in total cellular protein content upon prion infection was identified by infrared (IR) whole-cell spectra and validated by bicinchoninic acid assay and single-cell volume analysis by atomic force microscopy (AFM). Hierarchical cluster analysis (HCA) of IR data discriminated between infected and uninfected cells and allowed to deduce an increment of lysosomal bodies within the cytoplasm of infected GT1-1 cells, a hypothesis further confirmed by SR-IRMS at subcellular spatial resolution and fluorescent microscopy. The purpose of this work, therefore, consists of proposing IRMS as a powerful multiscreening platform, drawing on the synergy with conventional biological assays and microscopy techniques in order to increase the accuracy of investigations performed at the single-cell level. PMID:22778865

  20. The Investigation of ADAMTS16 in Insulin-Induced Human Chondrosarcoma Cells.

    Cakmak, Ozlem; Comertoglu, Ismail; Firat, Ridvan; Erdemli, Haci Kemal; Kursunlu, S Fatih; Akyol, Sumeyya; Ugurcu, Veli; Altuntas, Aynur; Adam, Bahattin; Demircan, Kadir

    2015-08-01

    A disintegrin-like metalloproteinase with thrombospondin motifs (ADAMTS) is a group of proteins that have enzymatic activity secreted by cells to the outside extracellular matrix. Insulin induces proteoglycan biosynthesis in chondrosarcoma chondrocytes. The purpose of the present in vitro study is to assess the time course effects of insulin on ADAMTS16 expression in OUMS-27 (human chondrosarcoma) cell line to examine whether insulin regulates ADAMTS16 expression as well as proteoglycan biosynthesis with multifaceted properties or not. Chondrosarcoma cells were cultured in Dulbecco's modified Eagle's medium having either 10 μg/mL insulin or not. While the experiment was going on, the medium containing insulin had been changed every other day. Cells were harvested at 1st, 3rd, 7th, and 11th days; subsequently, RNA and proteins were isolated in every experimental group according to their time interval. RNA expression of ADAMTS was estimated by quantitative real-time polymerase chain reaction (qRT-PCR) by using primers. Immunoreactive protein levels were encountered by the western blot protein detection technique by using proper anti-ADAMTS16 antibodies. ADAMTS16 mRNA expression level of chondrosarcoma cells was found to be insignificantly decreased in chondrosarcoma cells induced by insulin detected by the qRT-PCR instrument. On the other hand, there was a gradual decrease in immune-reactant ADAMTS16 protein amount by the time course in insulin-treated cell groups when compared with control cells. It has been suggested that insulin might possibly regulate ADAMTS16 levels/activities in OUMS-27 chondrosarcoma cells taking a role in extracellular matrix turnover.

  1. Investigation of the Biochemical Mechanism for Cell-Substrate Mechanical Sensing

    Ricotta, Vincent Anthony

    Advancements in stem cell biology and materials science have enabled the development of new treatments for tissue repair. Dental pulp stem cells (DPSCs), which are highly proliferative and can be induced to differentiate along several mesenchymal cell lineages, offer the possibility for pulpal regeneration and treatment of injured dentition. Polybutadiene (PB) may be used as a substrate for these cells. This elastomer can be spun casted into films of different thicknesses with different moduli. DPSCs grown on PB films, which are relatively hard (less than 1500 A thick), biomineralize depositing crystalline calcium phosphate without a requirement for the typical induction factor, dexamethasone (Dex). The moduli of cells track with the moduli of the surface suggesting that mechanics controls mineralization. The purpose of this study was to determine whether the major effect of Dex on biomineralization is the result of its ability to alter cell mechanics or its ability to induce osteogenesis/odontogenesis. DPSCs sense substrate mechanics through the focal adhesions, whose function is in part regulated by the Ras homolog gene (Rho) and its downstream effectors Rho associated kinases (ROCKs). ROCKs control actin filament polymerization and interactions with myosin light chain. Because cells sense substrate mechanics through focal adhesion proteins whose function is regulated by ROCKs, the impact of a ROCK inhibitor, Y-27632, was monitored. Blocking this pathway with Y-27632 suppressed the ability of DPSCs to sense the PB substrate. The cell modulus, plasma membrane stiffness, and cytosol stiffness were all lowered and biomineralization was suppressed in all cultures independent of substrate modulus or the presence of Dex. In other words, the inability of DPSCs to sense mechanical cues suppressed their ability to promote mineralization. On the other hand the expression of osteogenic/odontogenic markers (alkaline phosphatase and osteocalcin) was enhanced, perhaps due to Y

  2. Meat and fish consumption and the risk of renal cell carcinoma in the European prospective investigation into cancer and nutrition

    Rohrmann, Sabine; Linseisen, Jakob; Overvad, Kim; Wurtz, Anne Mette Lund; Roswall, Nina; Tjonneland, Anne; Boutron-Ruault, Marie-Christine; Racine, Antoine; Bastide, Nadia; Palli, Domenico; Agnoli, Claudia; Panico, Salvatore; Tumino, Rosario; Sacerdote, Carlotta; Weikert, Steffen; Steffen, Annika; Kuehn, Tilman; Li, Kuanrong; Khaw, Kay-Tee; Wareham, Nicholas J.; Bradbury, Kathryn E.; Peppa, Eleni; Trichopoulou, Antonia; Trichopoulos, Dimitrios; Bueno-de-Mesquita, H. Bas; Peeters, Petra H. M.; Hjartaker, Anette; Skeie, Guri; Weiderpass, Elisabete; Jakszyn, Paula; Dorronsoro, Miren; Barricarte, Aurelio; Santiuste de Pablos, Carmen; Molina-Montes, Esther; Alonso de la Torre, Ramon; Ericson, Ulrika; Sonestedt, Emily; Johansson, Mattias; Ljungberg, Borje; Freisling, Heinz; Romieu, Isabelle; Cross, Amanda J.; Vergnaud, Anne-Claire; Riboli, Elio; Boeing, Heiner

    2015-01-01

    Renal cell cancer (RCC) incidence varies worldwide with a higher incidence in developed countries and lifestyle is likely to contribute to the development of this disease. We examined whether meat and fish consumption were related to the risk of RCC in the European Prospective Investigation into

  3. Investigating a case of possible field cancerization in oral squamous cell carcinoma by the use of next-generation sequencing

    Tabatabaeifar, Siavosh; Larsen, Martin J.; Larsen, Stine R.

    2017-01-01

    Objectives Local recurrence and the development of second primary tumors (SPT) are important factors that can influence the survival rate of oral squamous cell carcinoma (OSCC) patients. We