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Sample records for cell receptor-t3 complex

  1. Characterization and expression of the human T cell receptor-T3 complex by monoclonal antibody F101.01

    DEFF Research Database (Denmark)

    Geisler, C; Plesner, T; Pallesen, G

    1988-01-01

    A murine monoclonal antibody (MoAb) F101.01 reacting with the T cell receptor (TCR)-T3 complex is presented. Immunohistological studies showed that F101.01 specifically stains T-zone lymphocytes in lymph nodes, tonsils, and splenic tissue. Two-colour immunofluorescence and flow cytometry...... demonstrated co-expression of the antigen defined by F101.01 and the pan-T cell antigens defined by CD2, CD3, CD5, and CD7 antibodies. Cells stained with CD4 and CD8 antibodies were both included in the F101.01-positive population, whereas CD16-positive natural killer cells (NK), B cells (CD19 and CD20......), and myeloid cells (CD13 and CD33) were excluded. The target antigen of F101.01 co-modulated with the CD3-defined antigen (T3) and the TCR recognized by the MoAb WT-31. CD3 antibody and WT-31 both blocked binding of F101.01. F101.01 precipitated the TCR-T3 complex from lysates of 125I-labelled peripheral blood...

  2. Characterization and expression of the human T cell receptor-T3 complex by monoclonal antibody F101.01

    DEFF Research Database (Denmark)

    Geisler, C; Plesner, T; Pallesen, G

    1988-01-01

    demonstrated co-expression of the antigen defined by F101.01 and the pan-T cell antigens defined by CD2, CD3, CD5, and CD7 antibodies. Cells stained with CD4 and CD8 antibodies were both included in the F101.01-positive population, whereas CD16-positive natural killer cells (NK), B cells (CD19 and CD20...

  3. Chansporter complexes in cell signaling.

    Science.gov (United States)

    Abbott, Geoffrey W

    2017-09-01

    Ion channels facilitate diffusion of ions across cell membranes for such diverse purposes as neuronal signaling, muscular contraction, and fluid homeostasis. Solute transporters often utilize ionic gradients to move aqueous solutes up their concentration gradient, also fulfilling a wide variety of tasks. Recently, an increasing number of ion channel-transporter ('chansporter') complexes have been discovered. Chansporter complex formation may overcome what could otherwise be considerable spatial barriers to rapid signal integration and feedback between channels and transporters, the ions and other substrates they transport, and environmental factors to which they must respond. Here, current knowledge in this field is summarized, covering both heterologous expression structure/function findings and potential mechanisms by which chansporter complexes fulfill contrasting roles in cell signaling in vivo. © 2017 Federation of European Biochemical Societies.

  4. Tensor Product of Polygonal Cell Complexes

    OpenAIRE

    Chien, Yu-Yen

    2017-01-01

    We introduce the tensor product of polygonal cell complexes, which interacts nicely with the tensor product of link graphs of complexes. We also develop the unique factorization property of polygonal cell complexes with respect to the tensor product, and study the symmetries of tensor products of polygonal cell complexes.

  5. General reformulation of hot cell complex

    International Nuclear Information System (INIS)

    Almeida, G.L. de; Souza, A.S.F. de; Souza, M.L.M. de; Rautenberg, F.A.

    1986-01-01

    The implantation of an operation philosophy without direct intervention of operator during isotope production process in hot cells of the CV-28 cyclotron is presented. The modifications carried out in equipments, systems and installations are described. (M.C.K.)

  6. Cell complexes of transition metals in biochemistry and medicine

    International Nuclear Information System (INIS)

    Voloshin, Ya.Z.; Varzatskij, O.A.; Bubnov, Yu.N.

    2007-01-01

    Basic directions and prospects of use of cell complexes of transition metals in medicine and biochemistry are considered: incapsulation of radioactive metal ions for radiotherapy and diagnostics; preparation of contrast compounds for magnetic resonance tomography, antidotes and pharmaceutical preparation of prolonged effect, preparations for boron-neutron-capture therapy of neoplasms, antioxidants; membrane transport of metal ions; study of interaction of cell metal complexes with nucleic acids; possibility of use of self-assembly of cell complexes for imitation of ligases and use of clathrochelates as linkers; design of inhibitors of viruses for AIDS therapy [ru

  7. Neurosecretory cells of the amygdaloid complex during estrous cycle.

    Science.gov (United States)

    Akhmadeev, A V; Kalimullina, L B

    2005-02-01

    Ultrastructure of neurosecretory cells of the dorsomedial nucleus of the cerebral amygdaloid complex (one of the main zones of sexual dimorphism) was studied in different phases of the estrous cycle. The characteristics of the "light" and "dark" cells change depending on the concentrations of sex steroids during estrus and metestrus.

  8. Characterising the cellulose synthase complexes of cell walls

    NARCIS (Netherlands)

    Mansoori Zangir, N.

    2012-01-01

    One of the characteristics of the plant kingdom is the presence of a structural cell wall. Cellulose is a major component in both the primary and secondary cell walls of plants. In higher plants cellulose is synthesized by so called rosette protein complexes with cellulose synthases (CESAs) as

  9. Antitumor Cell-Complex Vaccines Employing Genetically Modified Tumor Cells and Fibroblasts

    Directory of Open Access Journals (Sweden)

    Antonio Miguel

    2014-02-01

    Full Text Available The present study evaluates the immune response mediated by vaccination with cell complexes composed of irradiated B16 tumor cells and mouse fibroblasts genetically modified to produce GM-CSF. The animals were vaccinated with free B16 cells or cell complexes. We employed two gene plasmid constructions: one high producer (pMok and a low producer (p2F. Tumor transplant was performed by injection of B16 tumor cells. Plasma levels of total IgG and its subtypes were measured by ELISA. Tumor volumes were measured and survival curves were obtained. The study resulted in a cell complex vaccine able to stimulate the immune system to produce specific anti-tumor membrane proteins (TMP IgG. In the groups vaccinated with cells transfected with the low producer plasmid, IgG production was higher when we used free B16 cell rather than cell complexes. Nonspecific autoimmune response caused by cell complex was not greater than that induced by the tumor cells alone. Groups vaccinated with B16 transfected with low producer plasmid reached a tumor growth delay of 92% (p ≤ 0.01. When vaccinated with cell complex, the best group was that transfected with high producer plasmid, reaching a tumor growth inhibition of 56% (p ≤ 0.05. Significant survival (40% was only observed in the groups vaccinated with free transfected B16 cells.

  10. Trichomonas vaginalis perturbs the junctional complex in epithelial cells

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Trichomonas vaginalis, a protist parasite of the urogenital tract in humans, is the causative agent of trichomonosis,which in recent years have been associated with the cervical cancer development. In the present study we analyzed the modifications at the junctional complex level of Caco-2 cells after interaction with two isolates of T. vaginalis and the influence of the iron concentration present in the parasite's culture medium on the interaction effects. Our results show that T. vaginalis adheres to the epithelial cell causing alterations in the junctional complex, such as: (a) a decrease in transepithelial electrical resistance; (b) alteration in the pattern of junctional complex proteins distribution as obseryed for E-cadherin, occludin and ZO-1; and (c) enlargement of the spaces between epithelial cells. These effects were dependent on (a) the degree of the parasite virulence isolate, (b) the iron concentration in the culture medium, and (c) the expression of adhesin proteins on the parasite surface.

  11. Modeling of Complex Life Cycle Prediction Based on Cell Division

    Directory of Open Access Journals (Sweden)

    Fucheng Zhang

    2017-01-01

    Full Text Available Effective fault diagnosis and reasonable life expectancy are of great significance and practical engineering value for the safety, reliability, and maintenance cost of equipment and working environment. At present, the life prediction methods of the equipment are equipment life prediction based on condition monitoring, combined forecasting model, and driven data. Most of them need to be based on a large amount of data to achieve the problem. For this issue, we propose learning from the mechanism of cell division in the organism. We have established a moderate complexity of life prediction model across studying the complex multifactor correlation life model. In this paper, we model the life prediction of cell division. Experiments show that our model can effectively simulate the state of cell division. Through the model of reference, we will use it for the equipment of the complex life prediction.

  12. Tutorial: Electroporation of cells in complex materials and tissue

    Science.gov (United States)

    Rems, L.; Miklavčič, D.

    2016-05-01

    Electroporation is being successfully used in biology, medicine, food processing, and biotechnology, and in some environmental applications. Recent applications also include in addition to classical electroporation, where cells are exposed to micro- or milliseconds long pulses, exposures to extremely short nanosecond pulses, i.e., high-frequency electroporation. Electric pulses are applied to cells in different structural configurations ranging from suspended cells to cells in tissues. Understanding electroporation of cells in tissues and other complex environments is a key to its successful use and optimization in various applications. Thus, explanation will be provided theoretically/numerically with relation to experimental observations by scaling our understanding of electroporation from the molecular level of the cell membrane up to the tissue level.

  13. Nicotine affects protein complex rearrangement in Caenorhabditis elegans cells.

    Science.gov (United States)

    Sobkowiak, Robert; Zielezinski, Andrzej; Karlowski, Wojciech M; Lesicki, Andrzej

    2017-10-01

    Nicotine may affect cell function by rearranging protein complexes. We aimed to determine nicotine-induced alterations of protein complexes in Caenorhabditis elegans (C. elegans) cells, thereby revealing links between nicotine exposure and protein complex modulation. We compared the proteomic alterations induced by low and high nicotine concentrations (0.01 mM and 1 mM) with the control (no nicotine) in vivo by using mass spectrometry (MS)-based techniques, specifically the cetyltrimethylammonium bromide (CTAB) discontinuous gel electrophoresis coupled with liquid chromatography (LC)-MS/MS and spectral counting. As a result, we identified dozens of C. elegans proteins that are present exclusively or in higher abundance in either nicotine-treated or untreated worms. Based on these results, we report a possible network that captures the key protein components of nicotine-induced protein complexes and speculate how the different protein modules relate to their distinct physiological roles. Using functional annotation of detected proteins, we hypothesize that the identified complexes can modulate the energy metabolism and level of oxidative stress. These proteins can also be involved in modulation of gene expression and may be crucial in Alzheimer's disease. The findings reported in our study reveal putative intracellular interactions of many proteins with the cytoskeleton and may contribute to the understanding of the mechanisms of nicotinic acetylcholine receptor (nAChR) signaling and trafficking in cells.

  14. Internalisation of gonadotrophin-receptor complex in ovarian luteal cells

    International Nuclear Information System (INIS)

    Conn, P.M.; Conti, M.; Harwood, J.P.; Dufau, M.L.; Catt, K.J.

    1978-01-01

    Following evidence that certain protein hormones can enter target cells the present investigation was undertaken which shows that gonadotrophin-induced receptor loss may occur by a process of internalisation of the hormone-receptor complex following the initial interaction of gonadotrophin with the cell surface. Localisation studies were carried out in 33-d old female rats previously treated with pregnant mare serum gonadotrophin and human chorionic gonadotrophin (hCG) to induce ovarian luteinisation. Animals were injected with 125 I-hCG to label the ovarian receptors for luteinising hormone in vivo. Microscope autoradiographs demonstrating distribution of 125 I-hCG in ovaries at various times following injection are shown. The combined results from the autoradiographs and from solubilisation experiments were used to determine the location and nature of the hCG-receptor complex following occupancy and loss of receptors from the plasma membrane of luteinised ovarian cells. (U.K.)

  15. Enlightening intracellular complexity of living cells with quantitative phase microscopy

    Science.gov (United States)

    Martinez Torres, C.; Laperrousaz, B.; Berguiga, L.; Boyer Provera, E.; Elezgaray, J.; Nicolini, F. E.; Maguer-Satta, V.; Arneodo, A.; Argoul, F.

    2016-03-01

    The internal distribution of refractive indices (RIs) of a living cell is much more complex than usually admitted in multi-shell models. The reconstruction of RI maps from single phase images has rarely been achieved for several reasons: (i) we still have very little knowledge of the impact of internal macromolecular complexes on the local RI and (ii) phase changes produced by light propagation through the sample are mixed with diffraction effects by internal cell bodies. We propose the implementation a 2D wavelet-based contour chain detection method to distinguish internal boundaries thanks to their greatest optical path difference gradients. These contour chains correspond to the highest image phase contrast and follow the local RI inhomogeneities linked to the intracellular structural intricacy. Their statistics and spatial distribution are morphological indicators for distinguishing cells of different origins and to follow their transformation in pathologic situations. We use this method to compare non adherent blood cells from primary and laboratory culture origins, in healthy and pathological situations (chronic myelogenous leukaemia). In a second part of this presentation, we concentrate on the temporal dynamics of the phase contour chains and we discuss the spectral decomposition of their dynamics in both health and disease.

  16. Pea border cell maturation and release involve complex cell wall structural dynamics

    DEFF Research Database (Denmark)

    Mravec, Jozef; Guo, Xiaoyuan; Hansen, Aleksander Riise

    2017-01-01

    The adhesion of plant cells is vital for support and protection of the plant body and is maintained by a variety of molecular associations between cell wall components. In some specialized cases though, plant cells are programmed to detach and root cap-derived border cells are examples of this....... Border cells (in some species known as border-like cells) provide an expendable barrier between roots and the environment. Their maturation and release is an important but poorly characterized cell separation event. To gain a deeper insight into the complex cellular dynamics underlying this process, we...... undertook a systematic, detailed analysis of pea (Pisum sativum) root tip cell walls. Our study included immuno-carbohydrate microarray profiling, monosaccharide composition determination, Fourier-transformed infrared microspectroscopy (FT-IR), quantitative RT-PCR of cell wall biosynthetic genes, analysis...

  17. Mass transport and chloride ion complexes in occluded cell

    International Nuclear Information System (INIS)

    Tsuru, T.; Hashimoto, K.; Nishikata, A.; Haruyama, S.

    1989-01-01

    Changes in the transport and the concentration of ions in a model occluded cell are traced during galvanostatic anodic polarization of a mild steel and a stainless steel. Apparent transport numbers of anions and cations, which were estimated from chemical analysis of solution, were different from those calculated from known mobility data. At the initial stage of the polarization, the transport number of chloride ion was almost unity, and then decreased gradually. For the mild steel, the concentration of total chloride ion accumulated in the occluded compartment increased with the anodic charge passed, and the amount of chloride ion complexed with cations also increased. The chloride complex was estimated as FeCl + . For SUS304 stainless steel, the total chloride ion increased, however, the free chloride ion, which responded to an Ag/AgCl electrode remained approximately 2 mol/dm 3 . Therefore, most of the chloride ions transferred into the occluded cell formed complex ions, such as CrCl n 3-n . The number of chloride ion coordinated to ferrous and chromic ions was estimated from the data fo mass transport for the case of the mild steel and the stainless steel. (author) 9 refs., 14 figs

  18. Mechanics of Cellulose Synthase Complexes in Living Plant Cells

    Science.gov (United States)

    Zehfroosh, Nina; Liu, Derui; Ramos, Kieran P.; Yang, Xiaoli; Goldner, Lori S.; Baskin, Tobias I.

    The polymer cellulose is one of the major components of the world's biomass with unique and fascinating characteristics such as its high tensile strength, renewability, biodegradability, and biocompatibility. Because of these distinctive aspects, cellulose has been the subject of enormous scientific and industrial interest, yet there are still fundamental open questions about cellulose biosynthesis. Cellulose is synthesized by a complex of transmembrane proteins called ``Cellulose Synthase A'' (CESA) in the plasma membrane. Studying the dynamics and kinematics of the CESA complex will help reveal the mechanism of cellulose synthesis and permit the development and validation of models of CESA motility. To understand what drives these complexes through the cell membrane, we used total internal reflection fluorescence microscopy (TIRFM) and variable angle epi-fluorescence microscopy to track individual, fluorescently-labeled CESA complexes as they move in the hypocotyl and root of living plants. A mean square displacement analysis will be applied to distinguish ballistic, diffusional, and other forms of motion. We report on the results of these tracking experiments. This work was funded by NSF/PHY-1205989.

  19. Pea Border Cell Maturation and Release Involve Complex Cell Wall Structural Dynamics.

    Science.gov (United States)

    Mravec, Jozef; Guo, Xiaoyuan; Hansen, Aleksander Riise; Schückel, Julia; Kračun, Stjepan Krešimir; Mikkelsen, Maria Dalgaard; Mouille, Grégory; Johansen, Ida Elisabeth; Ulvskov, Peter; Domozych, David S; Willats, William George Tycho

    2017-06-01

    The adhesion of plant cells is vital for support and protection of the plant body and is maintained by a variety of molecular associations between cell wall components. In some specialized cases, though, plant cells are programmed to detach, and root cap-derived border cells are examples of this. Border cells (in some species known as border-like cells) provide an expendable barrier between roots and the environment. Their maturation and release is an important but poorly characterized cell separation event. To gain a deeper insight into the complex cellular dynamics underlying this process, we undertook a systematic, detailed analysis of pea ( Pisum sativum ) root tip cell walls. Our study included immunocarbohydrate microarray profiling, monosaccharide composition determination, Fourier-transformed infrared microspectroscopy, quantitative reverse transcription-PCR of cell wall biosynthetic genes, analysis of hydrolytic activities, transmission electron microscopy, and immunolocalization of cell wall components. Using this integrated glycobiology approach, we identified multiple novel modes of cell wall structural and compositional rearrangement during root cap growth and the release of border cells. Our findings provide a new level of detail about border cell maturation and enable us to develop a model of the separation process. We propose that loss of adhesion by the dissolution of homogalacturonan in the middle lamellae is augmented by an active biophysical process of cell curvature driven by the polarized distribution of xyloglucan and extensin epitopes. © 2017 American Society of Plant Biologists. All Rights Reserved.

  20. Pea Border Cell Maturation and Release Involve Complex Cell Wall Structural Dynamics1[OPEN

    Science.gov (United States)

    2017-01-01

    The adhesion of plant cells is vital for support and protection of the plant body and is maintained by a variety of molecular associations between cell wall components. In some specialized cases, though, plant cells are programmed to detach, and root cap-derived border cells are examples of this. Border cells (in some species known as border-like cells) provide an expendable barrier between roots and the environment. Their maturation and release is an important but poorly characterized cell separation event. To gain a deeper insight into the complex cellular dynamics underlying this process, we undertook a systematic, detailed analysis of pea (Pisum sativum) root tip cell walls. Our study included immunocarbohydrate microarray profiling, monosaccharide composition determination, Fourier-transformed infrared microspectroscopy, quantitative reverse transcription-PCR of cell wall biosynthetic genes, analysis of hydrolytic activities, transmission electron microscopy, and immunolocalization of cell wall components. Using this integrated glycobiology approach, we identified multiple novel modes of cell wall structural and compositional rearrangement during root cap growth and the release of border cells. Our findings provide a new level of detail about border cell maturation and enable us to develop a model of the separation process. We propose that loss of adhesion by the dissolution of homogalacturonan in the middle lamellae is augmented by an active biophysical process of cell curvature driven by the polarized distribution of xyloglucan and extensin epitopes. PMID:28400496

  1. Cell division control by the Chromosomal Passenger Complex

    Energy Technology Data Exchange (ETDEWEB)

    Waal, Maike S. van der; Hengeveld, Rutger C.C.; Horst, Armando van der; Lens, Susanne M.A., E-mail: s.m.a.lens@umcutrecht.nl

    2012-07-15

    The Chromosomal Passenger Complex (CPC) consisting of Aurora B kinase, INCENP, Survivin and Borealin, is essential for genomic stability by controlling multiple processes during both nuclear and cytoplasmic division. In mitosis it ensures accurate segregation of the duplicated chromosomes by regulating the mitotic checkpoint, destabilizing incorrectly attached spindle microtubules and by promoting the axial shortening of chromosomal arms in anaphase. During cytokinesis the CPC most likely prevents chromosome damage by imposing an abscission delay when a chromosome bridge connects the two daughter cells. Moreover, by controlling proper cytoplasmic division, the CPC averts tetraploidization. This review describes recent insights on how the CPC is capable of conducting its various functions in the dividing cell to ensure chromosomal stability.

  2. Role of Conserved Oligomeric Golgi Complex in the Abnormalities of Glycoprotein Processing in Breast Cancer Cells

    National Research Council Canada - National Science Library

    Zolov, Sergey N

    2006-01-01

    ...: protein glycosylation and its sorting. For analysis of COG complex function we utilized RNA interference assay to knockdown COG3p subunit of COG complex in normal and breast cancer cells and other tumor cell lines...

  3. Of macrophages and red blood cells; a complex love story.

    Directory of Open Access Journals (Sweden)

    Djuna Zoe de Back

    2014-01-01

    Full Text Available Macrophages tightly control the production and clearance of red blood cells (RBC. During steady state haematopoiesis, approximately 1010 red blood cells are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

  4. 89Zr-Oxine Complex PET Cell Imaging in Monitoring Cell-based Therapies

    Science.gov (United States)

    Wu, Haitao; Asiedu, Kingsley O.; Szajek, Lawrence P.; Griffiths, Gary L.; Choyke, Peter L.

    2015-01-01

    Purpose To develop a clinically translatable method of cell labeling with zirconium 89 (89Zr) and oxine to track cells with positron emission tomography (PET) in mouse models of cell-based therapy. Materials and Methods This study was approved by the institutional animal care committee. 89Zr-oxine complex was synthesized in an aqueous solution. Cell labeling conditions were optimized by using EL4 mouse lymphoma cells, and labeling efficiency was examined by using dendritic cells (DCs) (n = 4), naïve (n = 3) and activated (n = 3) cytotoxic T cells (CTLs), and natural killer (NK) (n = 4), bone marrow (n = 4), and EL4 (n = 4) cells. The effect of 89Zr labeling on cell survival, proliferation, and function were evaluated by using DCs (n = 3) and CTLs (n = 3). Labeled DCs (444–555 kBq/[5 × 106] cells, n = 5) and CTLs (185 kBq/[5 × 106] cells, n = 3) transferred to mice were tracked with microPET/CT. In a melanoma immunotherapy model, tumor targeting and cytotoxic function of labeled CTLs were evaluated with imaging (248.5 kBq/[7.7 × 106] cells, n = 4) and by measuring the tumor size (n = 6). Two-way analysis of variance was used to compare labeling conditions, the Wilcoxon test was used to assess cell survival and proliferation, and Holm-Sidak multiple tests were used to assess tumor growth and perform biodistribution analyses. Results 89Zr-oxine complex was synthesized at a mean yield of 97.3% ± 2.8 (standard deviation). It readily labeled cells at room temperature or 4°C in phosphate-buffered saline (labeling efficiency range, 13.0%–43.9%) and was stably retained (83.5% ± 1.8 retention on day 5 in DCs). Labeling did not affect the viability of DCs and CTLs when compared with nonlabeled control mice (P > .05), nor did it affect functionality. 89Zr-oxine complex enabled extended cell tracking for 7 days. Labeled tumor-specific CTLs accumulated in the tumor (4.6% on day 7) and induced tumor regression (P cell tracking technique for use with PET that is

  5. (89)Zr-Oxine Complex PET Cell Imaging in Monitoring Cell-based Therapies.

    Science.gov (United States)

    Sato, Noriko; Wu, Haitao; Asiedu, Kingsley O; Szajek, Lawrence P; Griffiths, Gary L; Choyke, Peter L

    2015-05-01

    To develop a clinically translatable method of cell labeling with zirconium 89 ((89)Zr) and oxine to track cells with positron emission tomography (PET) in mouse models of cell-based therapy. This study was approved by the institutional animal care committee. (89)Zr-oxine complex was synthesized in an aqueous solution. Cell labeling conditions were optimized by using EL4 mouse lymphoma cells, and labeling efficiency was examined by using dendritic cells (DCs) (n = 4), naïve (n = 3) and activated (n = 3) cytotoxic T cells (CTLs), and natural killer (NK) (n = 4), bone marrow (n = 4), and EL4 (n = 4) cells. The effect of (89)Zr labeling on cell survival, proliferation, and function were evaluated by using DCs (n = 3) and CTLs (n = 3). Labeled DCs (444-555 kBq/[5 × 10(6)] cells, n = 5) and CTLs (185 kBq/[5 × 10(6)] cells, n = 3) transferred to mice were tracked with microPET/CT. In a melanoma immunotherapy model, tumor targeting and cytotoxic function of labeled CTLs were evaluated with imaging (248.5 kBq/[7.7 × 10(6)] cells, n = 4) and by measuring the tumor size (n = 6). Two-way analysis of variance was used to compare labeling conditions, the Wilcoxon test was used to assess cell survival and proliferation, and Holm-Sidak multiple tests were used to assess tumor growth and perform biodistribution analyses. (89)Zr-oxine complex was synthesized at a mean yield of 97.3% ± 2.8 (standard deviation). It readily labeled cells at room temperature or 4°C in phosphate-buffered saline (labeling efficiency range, 13.0%-43.9%) and was stably retained (83.5% ± 1.8 retention on day 5 in DCs). Labeling did not affect the viability of DCs and CTLs when compared with nonlabeled control mice (P > .05), nor did it affect functionality. (89)Zr-oxine complex enabled extended cell tracking for 7 days. Labeled tumor-specific CTLs accumulated in the tumor (4.6% on day 7) and induced tumor regression (P cell tracking technique for use with PET that is applicable to a broad range of

  6. Of macrophages and red blood cells; a complex love story.

    Science.gov (United States)

    de Back, Djuna Z; Kostova, Elena B; van Kraaij, Marian; van den Berg, Timo K; van Bruggen, Robin

    2014-01-01

    Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

  7. Basal cell carcinoma of the nipple-areola complex.

    Science.gov (United States)

    Ferguson, Mark S; Nouraei, S A Reza; Davies, Ben J H; McLean, N R

    2009-11-01

    Basal cell carcinoma (BCC) of the nipple-areola complex is uncommon. It has been suggested that BCCs in this region behave more aggressively, with a higher potential for distant spread, than in other anatomical sites. To address questions about etiology, behavior, optimal treatment, and prognosis of this entity. A literature search identifying all cases of BCC of the nipple and nipple-areola complex in the English literature from 1893 to 2008. Thirty-four cases of BCC of the nipple, areola, or both were identified, mostly affecting middle-aged men. The majority of patients were treated with tissue-sparing surgery. There was a metastatic rate of 9.1%, and one patient died from the disease (3.0%). The optimal treatment of this condition should be local excision, but patients with this condition should be followed up for primary site recurrence and axillary metastasis, because there is greater incidence than with BCC at other anatomical sites. Furthermore, proven axillary metastasis should be surgically treated.

  8. A Single-Cell Biochemistry Approach Reveals PAR Complex Dynamics during Cell Polarization.

    Science.gov (United States)

    Dickinson, Daniel J; Schwager, Francoise; Pintard, Lionel; Gotta, Monica; Goldstein, Bob

    2017-08-21

    Regulated protein-protein interactions are critical for cell signaling, differentiation, and development. For the study of dynamic regulation of protein interactions in vivo, there is a need for techniques that can yield time-resolved information and probe multiple protein binding partners simultaneously, using small amounts of starting material. Here we describe a single-cell protein interaction assay. Single-cell lysates are generated at defined time points and analyzed using single-molecule pull-down, yielding information about dynamic protein complex regulation in vivo. We established the utility of this approach by studying PAR polarity proteins, which mediate polarization of many animal cell types. We uncovered striking regulation of PAR complex composition and stoichiometry during Caenorhabditis elegans zygote polarization, which takes place in less than 20 min. PAR complex dynamics are linked to the cell cycle by Polo-like kinase 1 and govern the movement of PAR proteins to establish polarity. Our results demonstrate an approach to study dynamic biochemical events in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Surface complexation of neptunium (V) onto whole cells and cell componets of Shewanella alga

    Energy Technology Data Exchange (ETDEWEB)

    Reed, Donald Timothy [Los Alamos National Laboratory; Deo, Randhir P [ASU; Rittmann, Bruce E [ASU; Songkasiri, Warinthorn [UNAFFILIATED

    2008-01-01

    We systematically quantified surface complexation of neptunium(V) onto whole cells of Shewanella alga strain BrY and onto cell wall and extracellular polymeric substances (EPS) of S. alga. We first performed acid and base titrations and used the mathematical model FITEQL with constant-capacitance surface-complexation to determine the concentrations and deprotonation constants of specific surface functional groups. Deprotonation constants most likely corresponded to a carboxyl site associated with amino acids (pK{sub a} {approx} 2.4), a carboxyl group not associated with amino acids (pK{sub a} {approx} 5), a phosphoryl site (pK{sub a} {approx} 7.2), and an amine site (pK{sub a} > 10). We then carried out batch sorption experiments with Np(V) and each of the S. alga components at different pHs. Results show that solution pH influenced the speciation of Np(V) and each of the surface functional groups. We used the speciation sub-model of the biogeochemical model CCBATCH to compute the stability constants for Np(V) complexation to each surface functional group. The stability constants were similar for each functional group on S. alga bacterial whole cells, cell walls, and EPS, and they explain the complicated sorption patterns when they are combined with the aqueous-phase speciation of Np(V). For pH < 8, NpO{sub 2}{sup +} was the dominant form of Np(V), and its log K values for the low-pK{sub a} carboxyl, other carboxyl, and phosphoryl groups were 1.75, 1.75, and 2.5 to 3.1, respectively. For pH greater than 8, the key surface ligand was amine >XNH3+, which complexed with NpO{sub 2}(CO{sub 3}){sub 3}{sup 5-}. The log K for NpO{sub 2}(CO{sub 3}){sub 3}{sup 5-} complexed onto the amine groups was 3.1 to 3.6. All of the log K values are similar to those of Np(V) complexes with aqueous carboxyl and N-containing carboxyl ligands. These results point towards the important role of surface complexation in defining key actinide-microbiological interactions in the subsurface.

  10. Relationships between Cargo, Cell Penetrating Peptides and Cell Type for Uptake of Non-Covalent Complexes into Live Cells

    Directory of Open Access Journals (Sweden)

    Andrea-Anneliese Keller

    2013-02-01

    Full Text Available Modulating signaling pathways for research and therapy requires either suppression or expression of selected genes or internalization of proteins such as enzymes, antibodies, nucleotide binding proteins or substrates including nucleoside phosphates and enzyme inhibitors. Peptides, proteins and nucleotides are transported by fusing or conjugating them to cell penetrating peptides or by formation of non-covalent complexes. The latter is often preferred because of easy handling, uptake efficiency and auto-release of cargo into the live cell. In our studies complexes are formed with labeled or readily detectable cargoes for qualitative and quantitative estimation of their internalization. Properties and behavior of adhesion and suspension vertebrate cells as well as the protozoa Leishmania tarentolae are investigated with respect to proteolytic activity, uptake efficiency, intracellular localization and cytotoxicity. Our results show that peptide stability to membrane-bound, secreted or intracellular proteases varies between different CPPs and that the suitability of individual CPPs for a particular cargo in complex formation by non-covalent interactions requires detailed studies. Cells vary in their sensitivity to increasing concentrations of CPPs. Thus, most cells can be efficiently transduced with peptides, proteins and nucleotides with intracellular concentrations in the low micromole range. For each cargo, cell type and CPP the optimal conditions must be determined separately.

  11. Slowness and sparseness have diverging effects on complex cell learning.

    Directory of Open Access Journals (Sweden)

    Jörn-Philipp Lies

    2014-03-01

    Full Text Available Following earlier studies which showed that a sparse coding principle may explain the receptive field properties of complex cells in primary visual cortex, it has been concluded that the same properties may be equally derived from a slowness principle. In contrast to this claim, we here show that slowness and sparsity drive the representations towards substantially different receptive field properties. To do so, we present complete sets of basis functions learned with slow subspace analysis (SSA in case of natural movies as well as translations, rotations, and scalings of natural images. SSA directly parallels independent subspace analysis (ISA with the only difference that SSA maximizes slowness instead of sparsity. We find a large discrepancy between the filter shapes learned with SSA and ISA. We argue that SSA can be understood as a generalization of the Fourier transform where the power spectrum corresponds to the maximally slow subspace energies in SSA. Finally, we investigate the trade-off between slowness and sparseness when combined in one objective function.

  12. Role of Conserved Oligomeric Golgi Complex in the Abnormalities of Glycoprotein Processing in Breast Cancer Cells

    Science.gov (United States)

    2006-05-01

    of COG complex function we utilized RNA interference assay to knockdown COG3p subunit of COG complex in normal and breast cancer cells and other tumor...protein trafficking, but the role of the COG complex in the abnormal glycosylation and secretion of tumor markers in breast cancer cells remains... COG complex in breast cancer cells MCF7 had been elevated 2-4 times in comparison to HB2 cells (Figure 5 A). The expression of HeLa COG3 CD44 ab

  13. Cationic Amphiphilic Tris-Cyclometalated Iridium(III) Complexes Induce Cancer Cell Death via Interaction with Ca2+-Calmodulin Complex.

    Science.gov (United States)

    Hisamatsu, Yosuke; Suzuki, Nozomi; Masum, Abdullah-Al; Shibuya, Ai; Abe, Ryo; Sato, Akira; Tanuma, Sei-Ichi; Aoki, Shin

    2017-02-15

    In our previous paper, we reported on the preparation of some cationic amphiphilic Ir complexes (2c, 2d) containing KKGG peptides that induce and detect cell death of Jurkat cells. Mechanistic studies suggest that 2c interacts with anionic molecules and/or membrane receptors on the cell surface to trigger an intracellular Ca 2+ response, resulting in the induction of cell death, accompanied by membrane disruption. We have continued the studies of cell death of Jurkat cells induced by 2c and found that xestospongin C, a selective inhibitor of an inositol 1,4,5-trisphosphate receptor located on the endoplasmic reticulum (ER), reduces the cytotoxicity of 2c, suggesting that 2c triggers the release of Ca 2+ from the ER, leading to an increase in the concentration of cytosolic Ca 2+ , thus inducing cell death. Moreover, we synthesized a series of new amphiphilic cationic Ir complexes 5a-c containing photoreactive 3-trifluoromethyl-3-phenyldiazirine (TFPD) groups, in an attempt to identify the target molecules of 2c. Interestingly, it was discovered that a TFPD group functions as a triplet quencher of Ir complexes. It was also found that 5b is useful as a turn-on phosphorescent probe of acidic proteins such as bovine serum albumin (BSA) (pI = 4.7) and their complexation was confirmed by luminescence titrations and SDS-PAGE of photochemical products between them. These successful results allowed us to carry out photoaffinity labeling of the target biomolecules of 5b (2c and analogues thereof) in Jurkat cells. A proteomic analysis of the products obtained by the photoirradiation of 5b with Jurkat cells suggests that the Ca 2+ -binding protein "calmodulin (CaM)" is one of target proteins of the Ir complexes. Indeed, 5b was found to interact with the Ca 2+ -CaM complex, as evidenced by luminescence titrations and the results of photochemical reactions of 5b with CaM in the presence of Ca 2+ (SDS-PAGE). A plausible mechanism for cell death induced by a cationic amphiphilic Ir

  14. Ganglion cell complex scan in the early prediction of glaucoma.

    Science.gov (United States)

    Ganekal, S

    2012-01-01

    To compare the macular ganglion cell complex (GCC) with peripapillary retinal fiber layer (RNFL) thickness map in glaucoma suspects and patients. Forty participants (20 glaucoma suspects and 20 glaucoma patients) were enrolled. Macular GCC and RNFL thickness maps were performed in both eyes of each participant in the same visit. The sensitivity and specificity of a color code less than 5% (red or yellow) for glaucoma diagnosis were calculated. Standard Automated Perimetry was performed with the Octopus 3.1.1 Dynamic 24-2 program. The statistical analysis was performed with the SPSS 10.1 (SPSS Inc. Chicago, IL, EUA). Results were expressed as mean +/- standard deviation and a p value of 0.05 or less was considered significant. Provide absolute numbers of these findings with their units of measurement. There was a statistically significant difference in average RNFL thickness (p=0.004), superior RNFL thickness (p=0.006), inferior RNFL thickness (p=0.0005) and average GCC (p=0.03) between the suspects and glaucoma patients. There was no difference in optic disc area (p=0.35) and vertical cup/disc ratio (p=0.234) in both groups. While 38% eyes had an abnormal GCC and 13% had an abnormal RNFL thickness in the glaucoma suspect group, 98% had an abnormal GCC and 90% had an abnormal RNFL thickness in the glaucoma group. The ability to diagnose glaucoma with macular GCC thickness is comparable to that with peripapillary RNFL thickness . Macular GCC thickness measurements may be a good alternative or a complementary measurement to RNFL thickness assessment in the clinical evaluation of glaucoma. © NEPjOPH.

  15. Synaptic Basis for Differential Orientation Selectivity between Complex and Simple Cells in Mouse Visual Cortex.

    Science.gov (United States)

    Li, Ya-tang; Liu, Bao-hua; Chou, Xiao-lin; Zhang, Li I; Tao, Huizhong W

    2015-08-05

    In the primary visual cortex (V1), orientation-selective neurons can be categorized into simple and complex cells primarily based on their receptive field (RF) structures. In mouse V1, although previous studies have examined the excitatory/inhibitory interplay underlying orientation selectivity (OS) of simple cells, the synaptic bases for that of complex cells have remained obscure. Here, by combining in vivo loose-patch and whole-cell recordings, we found that complex cells, identified by their overlapping on/off subfields, had significantly weaker OS than simple cells at both spiking and subthreshold membrane potential response levels. Voltage-clamp recordings further revealed that although excitatory inputs to complex and simple cells exhibited a similar degree of OS, inhibition in complex cells was more narrowly tuned than excitation, whereas in simple cells inhibition was more broadly tuned than excitation. The differential inhibitory tuning can primarily account for the difference in OS between complex and simple cells. Interestingly, the differential synaptic tuning correlated well with the spatial organization of synaptic input: the inhibitory visual RF in complex cells was more elongated in shape than its excitatory counterpart and also was more elongated than that in simple cells. Together, our results demonstrate that OS of complex and simple cells is differentially shaped by cortical inhibition based on its orientation tuning profile relative to excitation, which is contributed at least partially by the spatial organization of RFs of presynaptic inhibitory neurons. Simple and complex cells, two classes of principal neurons in the primary visual cortex (V1), are generally thought to be equally selective for orientation. In mouse V1, we report that complex cells, identified by their overlapping on/off subfields, has significantly weaker orientation selectivity (OS) than simple cells. This can be primarily attributed to the differential tuning selectivity

  16. Chromatin Repressive Complexes in Stem Cells, Development, and Cancer

    DEFF Research Database (Denmark)

    Laugesen, Anne; Helin, Kristian

    2014-01-01

    The chromatin environment is essential for the correct specification and preservation of cell identity through modulation and maintenance of transcription patterns. Many chromatin regulators are required for development, stem cell maintenance, and differentiation. Here, we review the roles...

  17. Variations in cell morphology in the canine cruciate ligament complex.

    Science.gov (United States)

    Smith, K D; Vaughan-Thomas, A; Spiller, D G; Clegg, P D; Innes, J F; Comerford, E J

    2012-08-01

    Cell morphology may reflect the mechanical environment of tissues and influence tissue physiology and response to injury. Normal cruciate ligaments (CLs) from disease-free stifle joints were harvested from dog breeds with a high (Labrador retriever) and low (Greyhound) risk of cranial cruciate ligament (CCL) rupture. Antibodies against the cytoskeletal components vimentin and alpha tubulin were used to analyse cell morphology; nuclei were stained with 4',6-diamidino-2-phenylindole, and images were collected using conventional and confocal microscopy. Both cranial and caudal CLs contained cells of heterogenous morphologies. Cells were arranged between collagen bundles and frequently had cytoplasmic processes. Some of these processes were long (type A cells), others were shorter, thicker and more branched (type B cells), and some had no processes (type C cells). Processes were frequently shown to contact other cells, extending longitudinally and transversely through the CLs. Cells with longer processes had fusiform nuclei, and those with no processes had rounded nuclei and were more frequent in the mid-substance of both CLs. Cells with long processes were more commonly noted in the CLs of the Greyhound. As contact between cells may facilitate direct communication, variances in cell morphology between breeds at a differing risk of CCL rupture may reflect differences in CL physiology. Copyright © 2012 Elsevier Ltd. All rights reserved.

  18. Trafficking of the IKs -Complex in MDCK Cells

    DEFF Research Database (Denmark)

    David, Jens-Peter; Andersen, Martin N; Olesen, Søren-Peter

    2013-01-01

    place in a post-endoplasmic reticulum compartment. We demonstrate that K 7.1 targets the I -complex to the basolateral membrane, but that KCNE1 can redirect the complex to the apical membrane upon mutation of critical K 7.1 basolateral targeting signals. Our data provide a possible explanation...

  19. Biotin-tagged platinum(iv) complexes as targeted cytostatic agents against breast cancer cells.

    Science.gov (United States)

    Muhammad, Nafees; Sadia, Nasreen; Zhu, Chengcheng; Luo, Cheng; Guo, Zijian; Wang, Xiaoyong

    2017-09-05

    A biotin-guided platinum IV complex is highly cytotoxic against breast cancer cells but hypotoxic against mammary epithelial cells. The mono-biotinylated Pt IV complex is superior to the di-biotinylated one and hence a promising drug candidate for the targeted therapy of breast cancer.

  20. Radio metal (169Yb) uptake in normal and tumour cells in vitro. Influence of metabolic cell activity and complex structure

    International Nuclear Information System (INIS)

    Franke, W.G.; Kampf, G.

    1996-01-01

    Trivalent radio metal tracers have been used for tumour imaging and metastatic pain palliation. For better understanding their tumour accumulation, basic model studies of uptake of different 169 Yb complexes into cultured normal and tumour cells were performed. Whereas the uptake of 169 Yb citrate is strongly dependent on the metabolic activity and is not tumour-cell pacific, the uptake of 169 Yb complexed with amino carbonic acid (NTA, EDTA, DTPA) does not correlate to the metabolic activities. These complexes are taken up to a greater amount by the tumour cells (by a factor of about 2). Uptake of both complex types leads to a stable association to cellular compounds, 169 Yb is not releasable by the strong complexing agent DTPA. Protein binding of the 169 Yb complexes shows great influence on their cellular uptake. The bound proportion is no more available,for cellular uptake. The results indicate that i 0 uptake of 169 Yb citrate is an active cellular transport process which i not tumor-specific, ii) the 169 Yb amino carbonic acid complexes show a weak favouring by the tumour cells, iii) different from earlier acceptions the Yb complexes studied are not taken up by the cells in protein-bound form. The structure of the Yb complex is decisive for its protein binding and cellular uptake. (author). 13 refs., 6 figs

  1. The cell shape proteins MreB and MreC control cell morphogenesis by positioning cell wall synthetic complexes.

    Science.gov (United States)

    Divakaruni, Arun V; Baida, Cyril; White, Courtney L; Gober, James W

    2007-10-01

    MreB, the bacterial actin homologue, is thought to function in spatially co-ordinating cell morphogenesis in conjunction with MreC, a protein that wraps around the outside of the cell within the periplasmic space. In Caulobacter crescentus, MreC physically associates with penicillin-binding proteins (PBPs) which catalyse the insertion of intracellularly synthesized precursors into the peptidoglycan cell wall. Here we show that MreC is required for the spatial organization of components of the peptidoglycan-synthesizing holoenzyme in the periplasm and MreB directs the localization of a peptidoglycan precursor synthesis protein in the cytosol. Additionally, fluorescent vancomycin (Van-FL) labelling revealed that the bacterial cytoskeletal proteins MreB and FtsZ, as well as MreC and RodA, were required for peptidoglycan synthetic activity. MreB and FtsZ were found to be required for morphogenesis of the polar stalk. FtsZ was required for a cell cycle-regulated burst of peptidoglycan synthesis early in the cell cycle resulting in the synthesis of cross-band structures, whereas MreB was required for lengthening of the stalk. Thus, the bacterial cytoskeleton and cell shape-determining proteins such as MreC, function in concert to orchestrate the localization of cell wall synthetic complexes resulting in spatially co-ordinated and efficient peptidoglycan synthetic activity.

  2. A Comparative Analysis of Ganglion Cell Complex Parameters in ...

    African Journals Online (AJOL)

    Dr femi Oderinlo

    in the eyes, the optic nerve head, nerve fibre layer and retinal ganglion cells. Retinal ganglion cells encompass three layers ... of the macula in eyes with mild diabetic retinopathy. 8. *Correspondence: O Oderinlo, Eye Foundation ... most sensitive detection of GCC thinning. FLV provides a. 10 quantitative measure of the ...

  3. Novel piplartine-containing ruthenium complexes: synthesis, cell growth inhibition, apoptosis induction and ROS production on HCT116 cells.

    Science.gov (United States)

    D'Sousa Costa, Cinara O; Araujo Neto, João H; Baliza, Ingrid R S; Dias, Rosane B; Valverde, Ludmila de F; Vidal, Manuela T A; Sales, Caroline B S; Rocha, Clarissa A G; Moreira, Diogo R M; Soares, Milena B P; Batista, Alzir A; Bezerra, Daniel P

    2017-11-28

    Piplartine (piperlongumine) is a plant-derived molecule that has been receiving intense interest due to its anticancer characteristics that target the oxidative stress. In the present paper, two novel piplartine-containing ruthenium complexes [Ru(piplartine)(dppf)(bipy)](PF 6 ) 2 (1) and [Ru(piplartine)(dppb)(bipy)](PF 6 ) 2 (2) were synthesized and investigated for their cellular and molecular responses on cancer cell lines. We found that both complexes are more potent than metal-free piplartine in a panel of cancer cell lines on monolayer cultures, as well in 3D model of cancer multicellular spheroids formed from human colon carcinoma HCT116 cells. Mechanistic studies uncovered that the complexes reduced the cell growth and caused phosphatidylserine externalization, internucleosomal DNA fragmentation, caspase-3 activation and loss of the mitochondrial transmembrane potential on HCT116 cells. Moreover, the pre-treatment with Z-VAD(OMe)-FMK, a pan-caspase inhibitor, reduced the complexes-induced apoptosis, indicating cell death by apoptosis through caspase-dependent and mitochondrial intrinsic pathways. Treatment with the complexes also caused a marked increase in the production of reactive oxygen species (ROS), including hydrogen peroxide, superoxide anion and nitric oxide, and decreased reduced glutathione levels. Application of N-acetyl-cysteine, an antioxidant, reduced the ROS levels and apoptosis induced by the complexes, indicating activation of ROS-mediated apoptosis pathway. RNA transcripts of several genes, including gene related to the cell cycle, apoptosis and oxidative stress, were regulated under treatment. However, the complexes failed to induce DNA intercalation. In conclusion, the complexes are more potent than piplartine against different cancer cell lines and are able to induce caspase-dependent and mitochondrial intrinsic apoptosis on HCT116 cells by ROS-mediated pathway.

  4. Induction of cell-cell fusion by ectromelia virus is not inhibited by its fusion inhibitory complex

    Directory of Open Access Journals (Sweden)

    Fuchs Pinhas

    2009-09-01

    Full Text Available Abstract Background Ectromelia virus, a member of the Orthopox genus, is the causative agent of the highly infectious mousepox disease. Previous studies have shown that different poxviruses induce cell-cell fusion which is manifested by the formation of multinucleated-giant cells (polykaryocytes. This phenomenon has been widely studied with vaccinia virus in conditions which require artificial acidification of the medium. Results We show that Ectromelia virus induces cell-cell fusion under neutral pH conditions and requires the presence of a sufficient amount of viral particles on the plasma membrane of infected cells. This could be achieved by infection with a replicating virus and its propagation in infected cells (fusion "from within" or by infection with a high amount of virus particles per cell (fusion "from without". Inhibition of virus maturation or inhibition of virus transport on microtubules towards the plasma membrane resulted in a complete inhibition of syncytia formation. We show that in contrast to vaccinia virus, Ectromelia virus induces cell-cell fusion irrespectively of its hemagglutination properties and cell-surface expression of the orthologs of the fusion inhibitory complex, A56 and K2. Additionally, cell-cell fusion was also detected in mice lungs following lethal respiratory infection. Conclusion Ectromelia virus induces spontaneous cell-cell fusion in-vitro and in-vivo although expressing an A56/K2 fusion inhibitory complex. This syncytia formation property cannot be attributed to the 37 amino acid deletion in ECTV A56.

  5. Metal Complex Dyes for Dye-Sensitized Solar Cells: Recent ...

    Indian Academy of Sciences (India)

    interests are in coding theory, error-correction in networks and wireless communication.“ Ruthenium ..... Calculate power conversion efficiency by using the formula: 100 ... Molar Extinction Coefficient Charge-Transfer Sensitizers for Solar Cell.

  6. Structural Studies of Complex Carbohydrates of Plant Cell Walls

    Energy Technology Data Exchange (ETDEWEB)

    Darvill, Alan [Univ. of Georgia, Athens, GA (United States); Hahn, Michael G. [Univ. of Georgia, Athens, GA (United States); O' Neill, Malcolm A. [Univ. of Georgia, Athens, GA (United States); York, William S. [Univ. of Georgia, Athens, GA (United States)

    2015-02-17

    Most of the solar energy captured by land plants is converted into the polysaccharides (cellulose, hemicellulose, and pectin) that are the predominant components of the cell wall. These walls, which account for the bulk of plant biomass, have numerous roles in the growth and development of plants. Moreover, these walls have a major impact on human life as they are a renewable source of biomass, a source of diverse commercially useful polymers, a major component of wood, and a source of nutrition for humans and livestock. Thus, understanding the molecular mechanisms that lead to wall assembly and how cell walls and their component polysaccharides contribute to plant growth and development is essential to improve and extend the productivity and value of plant materials. The proposed research will develop and apply advanced analytical and immunological techniques to study specific changes in the structures and interactions of the hemicellulosic and pectic polysaccharides that occur during differentiation and in response to genetic modification and chemical treatments that affect wall biosynthesis. These new techniques will make it possible to accurately characterize minute amounts of cell wall polysaccharides so that subtle changes in structure that occur in individual cell types can be identified and correlated to the physiological or developmental state of the plant. Successful implementation of this research will reveal fundamental relationships between polysaccharide structure, cell wall architecture, and cell wall functions.

  7. The human CTC1/STN1/TEN1 complex regulates telomere maintenance in ALT cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Chenhui; Jia, Pingping; Chastain, Megan; Shiva, Olga; Chai, Weihang, E-mail: wchai@wsu.edu

    2017-06-15

    Maintaining functional telomeres is important for long-term proliferation of cells. About 15% of cancer cells are telomerase-negative and activate the alternative-lengthening of telomeres (ALT) pathway to maintain their telomeres. Recent studies have shown that the human CTC1/STN1/TEN1 complex (CST) plays a multi-faceted role in telomere maintenance in telomerase-expressing cancer cells. However, the role of CST in telomere maintenance in ALT cells is unclear. Here, we report that human CST forms a functional complex localizing in the ALT-associated PML bodies (APBs) in ALT cells throughout the cell cycle. Suppression of CST induces telomere instabilities including telomere fragility and elevates telomeric DNA recombination, leading to telomere dysfunction. In addition, CST deficiency significantly diminishes the abundance of extrachromosomal circular telomere DNA known as C-circles and t-circles. Suppression of CST also results in multinucleation in ALT cells and impairs cell proliferation. Our findings imply that the CST complex plays an important role in regulating telomere maintenance in ALT cells. - Highlights: • CST localizes at telomeres and ALT-associated PML bodies in ALT cells throughout the cell cycle. • CST is important for promoting telomeric DNA replication in ALT cells. • CST deficiency decreases ECTR formation and increases T-SCE. • CST deficiency impairs ALT cell proliferation and results in multinucleation.

  8. The Cell Probe Complexity of Dynamic Range Counting

    DEFF Research Database (Denmark)

    Larsen, Kasper Green

    2012-01-01

    is the number of update operations, w the cell size, tq the query time and tu the update time. In the most natural setting of cell size w = (lg n), this gives a lower bound of tq = ((lg n/ lg lg n)2) for any polylogarithmic update time. This bound is almost a quadratic improvement over the highest previous...... is specified by a point q = (x, y), and the goal is to report the sum of the weights assigned to the points dominated by q, where a point (x0, y0) is dominated by q if x0 x and y0 y. In addition to being the highest cell probe lower bound to date, our lower bound is also tight for data struc- tures with update...

  9. Gold nanoparticle-mediated laser stimulation causes a complex stress signal in neuronal cells

    Science.gov (United States)

    Johannsmeier, Sonja; Heeger, Patrick; Terakawa, Mitsuhiro; Kalies, Stefan; Heisterkamp, Alexander; Ripken, Tammo; Heinemann, Dag

    2017-07-01

    Gold nanoparticle mediated laser stimulation of neuronal cells allows for cell activation on a single-cell level. It could therefore be considered an alternative to classical electric neurostimulation. The physiological impact of this new approach has not been intensively studied so far. Here, we investigate the targeted cell's reaction to a laser stimulus based on its calcium response. A complex cellular reaction involving multiple sources has been revealed.

  10. Telomere dysfunction and cell survival: Roles for distinct TIN2-containing complexes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sahn-ho; Davalos, Albert R.; Heo, Seok-Jin; Rodier, Francis; Zou, Ying; Beausejour, Christian; Kaminker, Patrick; Yannone, Steven M.; Campisi, Judith

    2007-10-02

    Telomeres are maintained by three DNA binding proteins (TRF1, TRF2 and POT1), and several associated factors. One factor, TIN2, binds TRF1 and TRF2 directly and POT1 indirectly. Along with two other proteins, TPP1 and hRap1, these form a soluble complex that may be the core telomere maintenance complex. It is not clear whether sub-complexes also exist in vivo. We provide evidence for two TIN2 sub-complexes with distinct functions in human cells. We isolated these two TIN2 sub-complexes from nuclear lysates of unperturbed cells and cells expressing TIN2 mutants TIN2-13, TIN2-15C, which cannot bind TRF2 or TRF1, respectively. In cells with wild-type p53 function, TIN2-15C was more potent than TIN2-13 in causing telomere uncapping and eventual growth arrest. In cells lacking p53 function, TIN2-15C was more potent than TIN2-13 in causing telomere dysfunction and cell death. Our findings suggest that distinct TIN2 complexes exist, and that TIN2-15C-sensitive subcomplexes are particularly important for cell survival in the absence of functional p53.

  11. Control board and utility system for cell complex

    International Nuclear Information System (INIS)

    Almeida, G.L. de; Silva, A.C.; Souza, A.S.F. de; Souza, M.L.M. de; Rautenberg, F.A.

    1986-01-01

    To attend necessities of hot cells operation and process control for isotope production in IEN cyclotron (Brazilian-CNEN) a utility system, such as, electricity, water, vacuum, air, and gas, and control board was constructed, which advantages are presented. (M.C.K.)

  12. Of macrophages and red blood cells; a complex love story

    NARCIS (Netherlands)

    de Back, Djuna Z.; Kostova, Elena B.; van Kraaij, Marian; van den Berg, Timo K.; van Bruggen, Robin

    2014-01-01

    Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with

  13. Ruthenium complexes with phenylterpyridine derivatives target cell membrane and trigger death receptors-mediated apoptosis in cancer cells.

    Science.gov (United States)

    Deng, Zhiqin; Gao, Pan; Yu, Lianling; Ma, Bin; You, Yuanyuan; Chan, Leung; Mei, Chaoming; Chen, Tianfeng

    2017-06-01

    Elucidation of the communication between metal complexes and cell membrane may provide useful information for rational design of metal-based anticancer drugs. Herein we synthesized a novel class of ruthenium (Ru) complexes containing phtpy derivatives (phtpy = phenylterpyridine), analyzed their structure-activity relationship and revealed their action mechanisms. The result showed that, the increase in the planarity of hydrophobic Ru complexes significantly enhanced their lipophilicity and cellular uptake. Meanwhile, the introduction of nitro group effectively improved their anticancer efficacy. Further mechanism studies revealed that, complex (2c), firstly accumulated on cell membrane and interacted with death receptors to activate extrinsic apoptosis signaling pathway. The complex was then transported into cell cytoplasm through transferrin receptor-mediated endocytosis. Most of the intracellular 2c accumulated in cell plasma, decreasing the level of cellular ROS, inducing the activation of caspase-9 and thus intensifying the apoptosis. At the same time, the residual 2c can translocate into cell nucleus to interact with DNA, induce DNA damage, activate p53 pathway and enhance apoptosis. Comparing with cisplatin, 2c possesses prolonged circulation time in blood, comparable antitumor ability and importantly, much lower toxicity in vivo. Taken together, this study uncovers the role of membrane receptors in the anticancer actions of Ru complexes, and provides fundamental information for rational design of membrane receptor targeting anticancer drugs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Cytotoxicity and anti-tumor effects of new ruthenium complexes on triple negative breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Cecília P Popolin

    Full Text Available Triple-negative breast cancer (TNBC is a highly aggressive breast cancer subtype. The high rate of metastasis associated to the fact that these cells frequently display multidrug resistance, make the treatment of metastatic disease difficult. Development of antitumor metal-based drugs was started with the discovery of cisplatin, however, the severe side effects represent a limitation for its clinical use. Ruthenium (Ru complexes with different ligands have been successfully studied as prospective antitumor drugs. In this work, we demonstrated the activity of a series of biphosphine bipyridine Ru complexes (1 [Ru(SO4(dppb(bipy], (2 [Ru(CO3(dppb(bipy], (3 [Ru(C2O4(dppb(bipy] and (4 [Ru(CH3CO2(dppb(bipy]PF6 [where dppb = 1,4-bis(diphenylphosphinobutane and bipy = 2,2'-bipyridine], on proliferation of TNBC (MDA-MB-231, estrogen-dependent breast tumor cells (MCF-7 and a non-tumor breast cell line (MCF-10A. Complex (4 was most effective among the complexes and was selected to be further investigated on effects on tumor cell adhesion, migration, invasion and in apoptosis. Moreover, DNA and HSA binding properties of this complex were also investigated. Results show that complex (4 was more efficient inhibiting proliferation of MDA-MB-231 cells over non-tumor cells. In addition, complex (4 was able to inhibit MDA-MB231 cells adhesion, migration and invasion and to induce apoptosis and inhibit MMP-9 secretion in TNBC cells. Complex (4 should be further investigated in vivo in order to stablish its potential to improve breast cancer treatment.

  15. The Cell Probe Complexity of Succinct Data Structures

    DEFF Research Database (Denmark)

    Gal, Anna; Miltersen, Peter Bro

    2003-01-01

    In the cell probe model with word size 1 (the bit probe model), a static data structure problem is given by a map , where is a set of possible data to be stored, is a set of possible queries (for natural problems, we have ) and is the answer to question about data . A solution is given by a repre......In the cell probe model with word size 1 (the bit probe model), a static data structure problem is given by a map , where is a set of possible data to be stored, is a set of possible queries (for natural problems, we have ) and is the answer to question about data . A solution is given...

  16. Chromosome Mis-segregation Generates Cell-Cycle-Arrested Cells with Complex Karyotypes that Are Eliminated by the Immune System.

    Science.gov (United States)

    Santaguida, Stefano; Richardson, Amelia; Iyer, Divya Ramalingam; M'Saad, Ons; Zasadil, Lauren; Knouse, Kristin A; Wong, Yao Liang; Rhind, Nicholas; Desai, Arshad; Amon, Angelika

    2017-06-19

    Aneuploidy, a state of karyotype imbalance, is a hallmark of cancer. Changes in chromosome copy number have been proposed to drive disease by modulating the dosage of cancer driver genes and by promoting cancer genome evolution. Given the potential of cells with abnormal karyotypes to become cancerous, do pathways that limit the prevalence of such cells exist? By investigating the immediate consequences of aneuploidy on cell physiology, we identified mechanisms that eliminate aneuploid cells. We find that chromosome mis-segregation leads to further genomic instability that ultimately causes cell-cycle arrest. We further show that cells with complex karyotypes exhibit features of senescence and produce pro-inflammatory signals that promote their clearance by the immune system. We propose that cells with abnormal karyotypes generate a signal for their own elimination that may serve as a means for cancer cell immunosurveillance. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Different types of nsP3-containing protein complexes in Sindbis virus-infected cells.

    Science.gov (United States)

    Gorchakov, Rodion; Garmashova, Natalia; Frolova, Elena; Frolov, Ilya

    2008-10-01

    Alphaviruses represent a serious public health threat and cause a wide variety of diseases, ranging from severe encephalitis, which can result in death or neurological sequelae, to mild infection, characterized by fever, skin rashes, and arthritis. In the infected cells, alphaviruses express only four nonstructural proteins, which function in the synthesis of virus-specific RNAs and in modification of the intracellular environment. The results of our study suggest that Sindbis virus (SINV) infection in BHK-21 cells leads to the formation of at least two types of nsP3-containing complexes, one of which was found in association with the plasma membrane and endosome-like vesicles, while the second was coisolated with cell nuclei. The latter complexes could be solubilized only with the cytoskeleton-destabilizing detergent. Besides viral nsPs, in the mammalian cells, both complexes contained G3BP1 and G3BP2 (which were found in different ratios), YBX1, and HSC70. Rasputin, an insect cell-specific homolog of G3BP1, was found in the nsP3-containing complexes isolated from mosquito cells, which was suggestive of a high conservation of the complexes in the cells of both vertebrate and invertebrate origin. The endosome- and plasma membrane-associated complexes contained a high concentration of double-stranded RNAs (dsRNAs), which is indicative of their function in viral-RNA synthesis. The dsRNA synthesis is likely to efficiently proceed on the plasma membrane, and at least some of the protein-RNA complexes would then be transported into the cytosol in association with the endosome-like vesicular organelles. These findings provide new insight into the mechanism of SINV replication and virus-host cell interactions.

  18. Mammalian aPKC/Par polarity complex mediated regulation of epithelial division orientation and cell fate

    Energy Technology Data Exchange (ETDEWEB)

    Vorhagen, Susanne; Niessen, Carien M., E-mail: carien.niessen@uni-koeln.de

    2014-11-01

    Oriented cell division is a key regulator of tissue architecture and crucial for morphogenesis and homeostasis. Balanced regulation of proliferation and differentiation is an essential property of tissues not only to drive morphogenesis but also to maintain and restore homeostasis. In many tissues orientation of cell division is coupled to the regulation of differentiation producing daughters with similar (symmetric cell division, SCD) or differential fate (asymmetric cell division, ACD). This allows the organism to generate cell lineage diversity from a small pool of stem and progenitor cells. Division orientation and/or the ratio of ACD/SCD need to be tightly controlled. Loss of orientation or an altered ratio can promote overgrowth, alter tissue architecture and induce aberrant differentiation, and have been linked to morphogenetic diseases, cancer and aging. A key requirement for oriented division is the presence of a polarity axis, which can be established through cell intrinsic and/or extrinsic signals. Polarity proteins translate such internal and external cues to drive polarization. In this review we will focus on the role of the polarity complex aPKC/Par3/Par6 in the regulation of division orientation and cell fate in different mammalian epithelia. We will compare the conserved function of this complex in mitotic spindle orientation and distribution of cell fate determinants and highlight common and differential mechanisms in which this complex is used by tissues to adapt division orientation and cell fate to the specific properties of the epithelium.

  19. CD1 and major histocompatibility complex II molecules follow a different course during dendritic cell maturation

    NARCIS (Netherlands)

    van der Wel, Nicole N.; Sugita, Masahiko; Fluitsma, Donna M.; Cao, Xaiochun; Schreibelt, Gerty; Brenner, Michael B.; Peters, Peter J.

    2003-01-01

    The maturation of dendritic cells is accompanied by the redistribution of major histocompatibility complex (MHC) class II molecules from the lysosomal MHC class IT compartment to the plasma membrane to mediate presentation of peptide antigens. Besides MHC molecules, dendritic cells also express CD1

  20. Unraveling the Complexities of Androgen Receptor Signaling in Prostate Cancer Cells

    OpenAIRE

    Heemers, Hannelore V.; Tindall, Donald J.

    2009-01-01

    Androgen signaling is critical for proliferation of prostate cancer cells but cannot be fully inhibited by current androgen deprivation therapies. A study by Xu et al. in this issue of Cancer Cell provides insights into the complexities of androgen signaling in prostate cancer and suggests avenues to target a subset of androgen-sensitive genes.

  1. Control of cell fate by the formation of an architecturally complex bacterial community.

    Science.gov (United States)

    Vlamakis, Hera; Aguilar, Claudio; Losick, Richard; Kolter, Roberto

    2008-04-01

    Bacteria form architecturally complex communities known as biofilms in which cells are held together by an extracellular matrix. Biofilms harbor multiple cell types, and it has been proposed that within biofilms individual cells follow different developmental pathways, resulting in heterogeneous populations. Here we demonstrate cellular differentiation within biofilms of the spore-forming bacterium Bacillus subtilis, and present evidence that formation of the biofilm governs differentiation. We show that motile, matrix-producing, and sporulating cells localize to distinct regions within the biofilm, and that the localization and percentage of each cell type is dynamic throughout development of the community. Importantly, mutants that do not produce extracellular matrix form unstructured biofilms that are deficient in sporulation. We propose that sporulation is a culminating feature of biofilm formation, and that spore formation is coupled to the formation of an architecturally complex community of cells.

  2. Cyclometalated Ruthenium(II) Anthraquinone Complexes Exhibit Strong Anticancer Activity in Hypoxic Tumor Cells.

    Science.gov (United States)

    Zeng, Leli; Chen, Yu; Huang, Huaiyi; Wang, Jinquan; Zhao, Donglei; Ji, Liangnian; Chao, Hui

    2015-10-19

    Hypoxia is the critical feature of the tumor microenvironment that is known to lead to resistance to many chemotherapeutic drugs. Six novel ruthenium(II) anthraquinone complexes were designed and synthesized; they exhibit similar or superior cytotoxicity compared to cisplatin in hypoxic HeLa, A549, and multidrug-resistant (A549R) tumor cell lines. Their anticancer activities are related to their lipophilicity and cellular uptake; therefore, these physicochemical properties of the complexes can be changed by modifying the ligands to obtain better anticancer candidates. Complex 1, the most potent member of the series, is highly active against hypoxic HeLa cancer cells (IC50 =0.53 μM). This complex likely has 46-fold better activity than cisplatin (IC50 =24.62 μM) in HeLa cells. This complex tends to accumulate in the mitochondria and the nucleus of hypoxic HeLa cells. Further mechanistic studies show that complex 1 induced cell apoptosis during hypoxia through multiple pathways, including those of DNA damage, mitochondrial dysfunction, and the inhibition of DNA replication and HIF-1α expression, making it an outstanding candidate for further in vivo studies. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Cytotoxicity of Titanate-Calcium Complexes to MC3T3 Osteoblast-Like Cells

    Science.gov (United States)

    Drury, Jeanie L.; Moussi, Joelle; Taylor-Pashow, Kathryn M. L.

    2016-01-01

    Monosodium titanates (MST) are a relatively novel form of particulate titanium dioxide that have been proposed for biological use as metal sorbents or delivery agents, most recently calcium (II). In these roles, the toxicity of the titanate or its metal complex is crucial to its biological utility. The aim of this study was to determine the cytotoxicity of MST and MST-calcium complexes with MC3T3 osteoblast-like cells; MST-Ca(II) complexes could be useful to promote bone formation in various hard tissue applications. MC3T3 cells were exposed to native MST or MST-Ca(II) complexes for 24–72 h. A CellTiter-Blue® assay was employed to assess the metabolic activity of the cells. The results showed that MST and MST-Ca(II) suppressed MC3T3 metabolic activity significantly in a dose-, time-, and cell-density-dependent fashion. MST-Ca(II) suppressed MC3T3 metabolism in a statistically identical manner as native MST at all concentrations. We concluded that MST and MST-Ca(II) are significantly cytotoxic to MC3T3 cells through a mechanism yet unknown; this is a potential problem to the biological utility of these complexes. PMID:28044136

  4. Cytotoxicity of Titanate-Calcium Complexes to MC3T3 Osteoblast-Like Cells

    Directory of Open Access Journals (Sweden)

    Yen-Wei Chen

    2016-01-01

    Full Text Available Monosodium titanates (MST are a relatively novel form of particulate titanium dioxide that have been proposed for biological use as metal sorbents or delivery agents, most recently calcium (II. In these roles, the toxicity of the titanate or its metal complex is crucial to its biological utility. The aim of this study was to determine the cytotoxicity of MST and MST-calcium complexes with MC3T3 osteoblast-like cells; MST-Ca(II complexes could be useful to promote bone formation in various hard tissue applications. MC3T3 cells were exposed to native MST or MST-Ca(II complexes for 24–72 h. A CellTiter-Blue® assay was employed to assess the metabolic activity of the cells. The results showed that MST and MST-Ca(II suppressed MC3T3 metabolic activity significantly in a dose-, time-, and cell-density-dependent fashion. MST-Ca(II suppressed MC3T3 metabolism in a statistically identical manner as native MST at all concentrations. We concluded that MST and MST-Ca(II are significantly cytotoxic to MC3T3 cells through a mechanism yet unknown; this is a potential problem to the biological utility of these complexes.

  5. Information maximization explains the emergence of complex cell-like neurons

    Directory of Open Access Journals (Sweden)

    Takuma eTanaka

    2013-11-01

    Full Text Available We propose models and a method to qualitatively explain the receptive field properties of complex cells in the primary visual cortex. We apply a learning method based on the information maximization principle in a feedforward network, which comprises an input layer of image patches, simple cell-like first-output-layer neurons, and second-output-layer neurons (Model 1. The information maximization results in the emergence of the complex cell-like receptive field properties in the second-output-layer neurons. After learning, second-output-layer neurons receive connection weights having the same size from two first-output-layer neurons with sign-inverted receptive fields. The second-output-layer neurons replicate the phase invariance and iso-orientation suppression. Furthermore, on the basis of these results, we examine a simplified model showing the emergence of complex cell-like receptive fields (Model 2. We show that after learning, the output neurons of this model exhibit iso-orientation suppression, cross-orientation facilitation, and end stopping, which are similar to those found in complex cells. These properties of model neurons suggest that complex cells in the primary visual cortex become selective to features composed of edges to increase the variability of the output.

  6. Structurally related hydrazone-based metal complexes with different antitumor activities variably induce apoptotic cell death.

    Science.gov (United States)

    Megger, Dominik A; Rosowski, Kristin; Radunsky, Christian; Kösters, Jutta; Sitek, Barbara; Müller, Jens

    2017-04-05

    Three new complexes bearing the tridentate hydrazone-based ligand 2-(2-(1-(pyridin-2-yl)ethylidene)hydrazinyl)pyridine (L) were synthesized and structurally characterized. Biological tests indicate that the Zn(ii) complex [ZnCl 2 (L)] is of low cytotoxicity against the hepatocellular carcinoma cell line HepG2. In contrast, the Cu(ii) and Mn(ii) complexes [CuCl 2 (L)] and [MnCl 2 (L)] are highly cytotoxic with EC 50 values of 1.25 ± 0.01 μM and 20 ± 1 μM, respectively. A quantitative proteome analysis reveals that treatment of the cells with the Cu(ii) complex leads to a significantly altered abundance of 102 apoptosis-related proteins, whereas 38 proteins were up- or down-regulated by the Mn(ii) complex. A closer inspection of those proteins regulated only by the Cu(ii) complex suggests that the superior cytotoxic activity of this complex is likely to be related to an initiation of the caspase-independent cell death (CICD). In addition, an increased generation of reactive oxygen species (ROS) and a strong up-regulation of proteins responsive to oxidative stress suggest that alterations of the cellular redox metabolism likely contribute to the cytotoxicity of the Cu(ii) complex.

  7. Taurine-modified Ru(ii)-complex targets cancerous brain cells for photodynamic therapy.

    Science.gov (United States)

    Du, Enming; Hu, Xunwu; Roy, Sona; Wang, Peng; Deasy, Kieran; Mochizuki, Toshiaki; Zhang, Ye

    2017-05-30

    The precision and efficacy of photodynamic therapy (PDT) is essential for the treatment of brain tumors because the cancer cells are within or adjacent to the delicate nervous system. Taurine is an abundant amino acid in the brain that serves the central nervous system (CNS). A taurine-modified polypyridyl Ru-complex was shown to have optimized intracellular affinity in cancer cells through accumulation in lysosomes. Symmetrical modification of this Ru-complex by multiple taurine molecules enhanced the efficiency of molecular emission with boosted generation of reactive oxygen species. These characteristic features make the taurine-modified Ru-complex a potentially effective photosensitizer for PDT of target cancer cells, with outstanding efficacy in cancerous brain cells.

  8. Complex heterogeneous tissue constructs containing multiple cell types prepared by inkjet printing technology.

    Science.gov (United States)

    Xu, Tao; Zhao, Weixin; Zhu, Jian-Ming; Albanna, Mohammad Z; Yoo, James J; Atala, Anthony

    2013-01-01

    This study was designed to develop a versatile method for fabricating complex and heterogeneous three-dimensional (3D) tissue constructs using simultaneous ink-jetting of multiple cell types. Human amniotic fluid-derived stem cells (hAFSCs), canine smooth muscle cells (dSMCs), and bovine aortic endothelial cells (bECs), were separately mixed with ionic cross-linker calcium chloride (CaCl(2)), loaded into separate ink cartridges and printed using a modified thermal inkjet printer. The three cell types were delivered layer-by-layer to pre-determined locations in a sodium alginate-collagen composite located in a chamber under the printer. The reaction between CaCl(2) and sodium alginate resulted in a rapid formation of a solid composite gel and the printed cells were anchored in designated areas within the gel. The printing process was repeated for several cycles leading to a complex 3D multi-cell hybrid construct. The biological functions of the 3D printed constructs were evaluated in vitro and in vivo. Each of the printed cell types maintained their viability and normal proliferation rates, phenotypic expression, and physiological functions within the heterogeneous constructs. The bioprinted constructs were able to survive and mature into functional tissues with adequate vascularization in vivo. These findings demonstrate the feasibility of fabricating complex heterogeneous tissue constructs containing multiple cell types using inkjet printing technology. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. The dynamic relationship between cerebellar Purkinje cell simple spikes and the spikelet number of complex spikes.

    Science.gov (United States)

    Burroughs, Amelia; Wise, Andrew K; Xiao, Jianqiang; Houghton, Conor; Tang, Tianyu; Suh, Colleen Y; Lang, Eric J; Apps, Richard; Cerminara, Nadia L

    2017-01-01

    Purkinje cells are the sole output of the cerebellar cortex and fire two distinct types of action potential: simple spikes and complex spikes. Previous studies have mainly considered complex spikes as unitary events, even though the waveform is composed of varying numbers of spikelets. The extent to which differences in spikelet number affect simple spike activity (and vice versa) remains unclear. We found that complex spikes with greater numbers of spikelets are preceded by higher simple spike firing rates but, following the complex spike, simple spikes are reduced in a manner that is graded with spikelet number. This dynamic interaction has important implications for cerebellar information processing, and suggests that complex spike spikelet number may maintain Purkinje cells within their operational range. Purkinje cells are central to cerebellar function because they form the sole output of the cerebellar cortex. They exhibit two distinct types of action potential: simple spikes and complex spikes. It is widely accepted that interaction between these two types of impulse is central to cerebellar cortical information processing. Previous investigations of the interactions between simple spikes and complex spikes have mainly considered complex spikes as unitary events. However, complex spikes are composed of an initial large spike followed by a number of secondary components, termed spikelets. The number of spikelets within individual complex spikes is highly variable and the extent to which differences in complex spike spikelet number affects simple spike activity (and vice versa) remains poorly understood. In anaesthetized adult rats, we have found that Purkinje cells recorded from the posterior lobe vermis and hemisphere have high simple spike firing frequencies that precede complex spikes with greater numbers of spikelets. This finding was also evident in a small sample of Purkinje cells recorded from the posterior lobe hemisphere in awake cats. In addition

  10. Werner complex deficiency in cells disrupts the Nuclear Pore Complex and the distribution of lamin B1.

    Science.gov (United States)

    Li, Zhi; Zhu, Yizhou; Zhai, Yujia; R Castroagudin, Michelle; Bao, Yifei; White, Tommy E; Glavy, Joseph S

    2013-12-01

    From the surrounding shell to the inner machinery, nuclear proteins provide the functional plasticity of the nucleus. This study highlights the nuclear association of Pore membrane (POM) protein NDC1 and Werner protein (WRN), a RecQ helicase responsible for the DNA instability progeria disorder, Werner Syndrome. In our previous publication, we connected the DNA damage sensor Werner's Helicase Interacting Protein (WHIP), a binding partner of WRN, to the NPC. Here, we confirm the association of the WRN/WHIP complex and NDC1. In established WRN/WHIP knockout cell lines, we further demonstrate the interdependence of WRN/WHIP and Nucleoporins (Nups). These changes do not completely abrogate the barrier of the Nuclear Envelope (NE) but do affect the distribution of FG Nups and the RAN gradient, which are necessary for nuclear transport. Evidence from WRN/WHIP knockout cell lines demonstrates changes in the processing and nucleolar localization of lamin B1. The appearance of "RAN holes" void of RAN corresponds to regions within the nucleolus filled with condensed pools of lamin B1. From WRN/WHIP knockout cell line extracts, we found three forms of lamin B1 that correspond to mature holoprotein and two potential post-translationally modified forms of the protein. Upon treatment with topoisomerase inhibitors lamin B1 cleavage occurs only in WRN/WHIP knockout cells. Our data suggest the link of the NDC1 and WRN as one facet of the network between the nuclear periphery and genome stability. Loss of WRN complex leads to multiple alterations at the NPC and the nucleolus. © 2013. Published by Elsevier B.V. All rights reserved.

  11. Thermal proximity coaggregation for system-wide profiling of protein complex dynamics in cells.

    Science.gov (United States)

    Tan, Chris Soon Heng; Go, Ka Diam; Bisteau, Xavier; Dai, Lingyun; Yong, Chern Han; Prabhu, Nayana; Ozturk, Mert Burak; Lim, Yan Ting; Sreekumar, Lekshmy; Lengqvist, Johan; Tergaonkar, Vinay; Kaldis, Philipp; Sobota, Radoslaw M; Nordlund, Pär

    2018-03-09

    Proteins differentially interact with each other across cellular states and conditions, but an efficient proteome-wide strategy to monitor them is lacking. We report the application of thermal proximity coaggregation (TPCA) for high-throughput intracellular monitoring of protein complex dynamics. Significant TPCA signatures observed among well-validated protein-protein interactions correlate positively with interaction stoichiometry and are statistically observable in more than 350 annotated human protein complexes. Using TPCA, we identified many complexes without detectable differential protein expression, including chromatin-associated complexes, modulated in S phase of the cell cycle. Comparison of six cell lines by TPCA revealed cell-specific interactions even in fundamental cellular processes. TPCA constitutes an approach for system-wide studies of protein complexes in nonengineered cells and tissues and might be used to identify protein complexes that are modulated in diseases. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  12. Granzyme A Cleaves a Mitochondrial Complex I Protein to Initiate Caspase-Independent Cell Death

    Science.gov (United States)

    Martinvalet, Denis; Dykxhoorn, Derek M.; Ferrini, Roger; Lieberman, Judy

    2010-01-01

    SUMMARY The killer lymphocyte protease granzyme A (GzmA) triggers caspase-independent target cell death with morphological features of apoptosis. We previously showed that GzmA acts directly on mitochondria to generate reactive oxygen species (ROS) and disrupt the transmembrane potential (ΔΨm) but does not permeabilize the mitochondrial outer membrane. Mitochondrial damage is critical to GzmA-induced cell death since cells treated with superoxide scavengers are resistant to GzmA. Here we find that GzmA accesses the mitochondrial matrix to cleave the complex I protein NDUFS3, an iron-sulfur subunit of the NADH:ubiquinone oxidoreductase complex I, after Lys56 to interfere with NADH oxidation and generate superoxide anions. Target cells expressing a cleavage site mutant of NDUFS3 are resistant to GzmA-mediated cell death but remain sensitive to GzmB. PMID:18485875

  13. Novel ruthenium methylcyclopentadienyl complex bearing a bipyridine perfluorinated ligand shows strong activity towards colorectal cancer cells.

    Science.gov (United States)

    Teixeira, Ricardo G; Brás, Ana Rita; Côrte-Real, Leonor; Tatikonda, Rajendhraprasad; Sanches, Anabela; Robalo, M Paula; Avecilla, Fernando; Moreira, Tiago; Garcia, M Helena; Haukka, Matti; Preto, Ana; Valente, Andreia

    2018-01-01

    Three new compounds have been synthesized and completely characterized by analytical and spectroscopic techniques. The new bipyridine-perfluorinated ligand L1 and the new organometallic complex [Ru(η 5 -MeCp)(PPh 3 ) 2 Cl] (Ru1) crystalize in the centrosymmetric triclinic space group P1¯. Analysis of the phenotypic effects induced by both organometallic complexes Ru1 and [Ru(η 5 -MeCp)(PPh 3 )(L1)][CF 3 SO 3 ] (Ru2), on human colorectal cancer cells (SW480 and RKO) survival, showed that Ru2 has a potent anti-proliferative activity, 4-6 times higher than cisplatin, and induce apoptosis in these cells. Data obtained in a noncancerous cell line derived from normal colon epithelial cells (NCM460) revealed an intrinsic selectivity of Ru2 for malignant cells at low concentrations, showing the high potential of this compound as a selective anticancer agent. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  14. Feeding cells induced by phytoparasitic nematodes require γ-tubulin ring complex for microtubule reorganization.

    Directory of Open Access Journals (Sweden)

    Mohamed Youssef Banora

    2011-12-01

    Full Text Available Reorganization of the microtubule network is important for the fast isodiametric expansion of giant-feeding cells induced by root-knot nematodes. The efficiency of microtubule reorganization depends on the nucleation of new microtubules, their elongation rate and activity of microtubule severing factors. New microtubules in plants are nucleated by cytoplasmic or microtubule-bound γ-tubulin ring complexes. Here we investigate the requirement of γ-tubulin complexes for giant feeding cells development using the interaction between Arabidopsis and Meloidogyne spp. as a model system. Immunocytochemical analyses demonstrate that γ-tubulin localizes to both cortical cytoplasm and mitotic microtubule arrays of the giant cells where it can associate with microtubules. The transcripts of two Arabidopsis γ-tubulin (TUBG1 and TUBG2 and two γ-tubulin complex proteins genes (GCP3 and GCP4 are upregulated in galls. Electron microscopy demonstrates association of GCP3 and γ-tubulin as part of a complex in the cytoplasm of giant cells. Knockout of either or both γ-tubulin genes results in the gene dose-dependent alteration of the morphology of feeding site and failure of nematode life cycle completion. We conclude that the γ-tubulin complex is essential for the control of microtubular network remodelling in the course of initiation and development of giant-feeding cells, and for the successful reproduction of nematodes in their plant hosts.

  15. Telomere dysfunction and cell survival: roles for distinctTIN2-containing complexes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sahn-Ho; Davalos, Albert R.; Heo, Seok-Jin; Rodier, Francis; Beausejour, Christian; Kaminker, Patrick; Campisi, Judith

    2006-11-07

    Telomeres are maintained by three DNA binding proteins, TRF1, TRF2 and POT1, and several associated factors. One factor, TIN2, binds TRF1 and TRF2 directly and POT1 indirectly. These and two other proteins form a soluble complex that may be the core telomere-maintenance complex. It is not clear whether subcomplexes exist or function in vivo. Here, we provide evidence for two TIN2 subcomplexes with distinct functions in human cells. TIN2 ablation by RNA interference caused telomere uncapping and p53-independent cell death in all cells tested. However, we isolated two TIN2 complexes from cell lysates, each selectively sensitive to a TIN2 mutant (TIN2-13, TIN2-15C). In cells with wild-type p53 function, TIN2-15C was more potent than TIN2-13 in causing telomere uncapping and eventual growth arrest. In cells lacking p53 function, TIN215C more than TIN2-13 caused genomic instability and cell death. Thus, TIN2 subcomplexes likely have distinct functions in telomere maintenance, and may provide selective targets for eliminating cells with mutant p53.

  16. Merkel cell polyomavirus recruits MYCL to the EP400 complex to promote oncogenesis.

    Directory of Open Access Journals (Sweden)

    Jingwei Cheng

    2017-10-01

    Full Text Available Merkel cell carcinoma (MCC frequently contains integrated copies of Merkel cell polyomavirus DNA that express a truncated form of Large T antigen (LT and an intact Small T antigen (ST. While LT binds RB and inactivates its tumor suppressor function, it is less clear how ST contributes to MCC tumorigenesis. Here we show that ST binds specifically to the MYC homolog MYCL (L-MYC and recruits it to the 15-component EP400 histone acetyltransferase and chromatin remodeling complex. We performed a large-scale immunoprecipitation for ST and identified co-precipitating proteins by mass spectrometry. In addition to protein phosphatase 2A (PP2A subunits, we identified MYCL and its heterodimeric partner MAX plus the EP400 complex. Immunoprecipitation for MAX and EP400 complex components confirmed their association with ST. We determined that the ST-MYCL-EP400 complex binds together to specific gene promoters and activates their expression by integrating chromatin immunoprecipitation with sequencing (ChIP-seq and RNA-seq. MYCL and EP400 were required for maintenance of cell viability and cooperated with ST to promote gene expression in MCC cell lines. A genome-wide CRISPR-Cas9 screen confirmed the requirement for MYCL and EP400 in MCPyV-positive MCC cell lines. We demonstrate that ST can activate gene expression in a EP400 and MYCL dependent manner and this activity contributes to cellular transformation and generation of induced pluripotent stem cells.

  17. Merkel cell polyomavirus recruits MYCL to the EP400 complex to promote oncogenesis.

    Science.gov (United States)

    Cheng, Jingwei; Park, Donglim Esther; Berrios, Christian; White, Elizabeth A; Arora, Reety; Yoon, Rosa; Branigan, Timothy; Xiao, Tengfei; Westerling, Thomas; Federation, Alexander; Zeid, Rhamy; Strober, Benjamin; Swanson, Selene K; Florens, Laurence; Bradner, James E; Brown, Myles; Howley, Peter M; Padi, Megha; Washburn, Michael P; DeCaprio, James A

    2017-10-01

    Merkel cell carcinoma (MCC) frequently contains integrated copies of Merkel cell polyomavirus DNA that express a truncated form of Large T antigen (LT) and an intact Small T antigen (ST). While LT binds RB and inactivates its tumor suppressor function, it is less clear how ST contributes to MCC tumorigenesis. Here we show that ST binds specifically to the MYC homolog MYCL (L-MYC) and recruits it to the 15-component EP400 histone acetyltransferase and chromatin remodeling complex. We performed a large-scale immunoprecipitation for ST and identified co-precipitating proteins by mass spectrometry. In addition to protein phosphatase 2A (PP2A) subunits, we identified MYCL and its heterodimeric partner MAX plus the EP400 complex. Immunoprecipitation for MAX and EP400 complex components confirmed their association with ST. We determined that the ST-MYCL-EP400 complex binds together to specific gene promoters and activates their expression by integrating chromatin immunoprecipitation with sequencing (ChIP-seq) and RNA-seq. MYCL and EP400 were required for maintenance of cell viability and cooperated with ST to promote gene expression in MCC cell lines. A genome-wide CRISPR-Cas9 screen confirmed the requirement for MYCL and EP400 in MCPyV-positive MCC cell lines. We demonstrate that ST can activate gene expression in a EP400 and MYCL dependent manner and this activity contributes to cellular transformation and generation of induced pluripotent stem cells.

  18. Biochemical characterization of native Usher protein complexes from a vesicular subfraction of tracheal epithelial cells.

    Science.gov (United States)

    Zallocchi, Marisa; Sisson, Joseph H; Cosgrove, Dominic

    2010-02-16

    Usher syndrome is the major cause of deaf/blindness in the world. It is a genetic heterogeneous disorder, with nine genes already identified as causative for the disease. We noted expression of all known Usher proteins in bovine tracheal epithelial cells and exploited this system for large-scale biochemical analysis of Usher protein complexes. The dissected epithelia were homogenized in nondetergent buffer and sedimented on sucrose gradients. At least two complexes were evident after the first gradient: one formed by specific isoforms of CDH23, PCDH15, and VLGR-1 and a different one at the top of the gradient that included all of the Usher proteins and rab5, a transport vesicle marker. TEM analysis of these top fractions found them enriched in 100-200 nm vesicles, confirming a vesicular association of the Usher complex(es). Immunoisolation of these vesicles confirmed some of the associations already predicted and identified novel interactions. When the vesicles are lysed in the presence of phenylbutyrate, most of the Usher proteins cosediment into the gradient at a sedimentation coefficient of approximately 50 S, correlating with a predicted molecular mass of 2 x 10(6) Da. Although it is still unclear whether there is only one complex or several independent complexes that are trafficked within distinct vesicular pools, this work shows for the first time that native Usher protein complexes occur in vivo. This complex(es) is present primarily in transport vesicles at the apical pole of tracheal epithelial cells, predicting that Usher proteins may be directionally transported as complexes in hair cells and photoreceptors.

  19. Gold nanoparticle-mediated laser stimulation induces a complex stress response in neuronal cells.

    Science.gov (United States)

    Johannsmeier, Sonja; Heeger, Patrick; Terakawa, Mitsuhiro; Kalies, Stefan; Heisterkamp, Alexander; Ripken, Tammo; Heinemann, Dag

    2018-04-25

    Stimulation of neuronal cells generally resorts to electric signals. Recent advances in laser-based stimulation methods could present an alternative with superior spatiotemporal resolution. The avoidance of electronic crosstalk makes these methods attractive for in vivo therapeutic application. In particular, nano-mediators, such as gold nanoparticles, can be used to transfer the energy from a laser pulse to the cell membrane and subsequently activate excitable cells. Although the underlying mechanisms of neuronal activation have been widely unraveled, the overall effect on the targeted cell is not understood. Little is known about the physiological and pathophysiological impact of a laser pulse targeted onto nanoabsorbers on the cell membrane. Here, we analyzed the reaction of the neuronal murine cell line Neuro-2A and murine primary cortical neurons to gold nanoparticle mediated laser stimulation. Our study reveals a severe, complex and cell-type independent stress response after laser irradiation, emphasizing the need for a thorough assessment of this approach's efficacy and safety.

  20. A new synthesis route for Os-complex modified redox polymers for potential biofuel cell applications.

    Science.gov (United States)

    Pöller, Sascha; Beyl, Yvonne; Vivekananthan, Jeevanthi; Guschin, Dmitrii A; Schuhmann, Wolfgang

    2012-10-01

    A new synthesis route for Os-complex modified redox polymers was developed. Instead of ligand exchange reactions for coordinative binding of suitable precursor Os-complexes at the polymer, Os-complexes already exhibiting the final ligand shell containing a suitable functional group were bound to the polymer via an epoxide opening reaction. By separation of the polymer synthesis from the ligand exchange reaction at the Os-complex, the modification of the same polymer backbone with different Os-complexes or the binding of the same Os-complex to a number of different polymer backbones becomes feasible. In addition, the Os-complex can be purified and characterized prior to its binding to the polymer. In order to further understand and optimize suitable enzyme/redox polymer systems concerning their potential application in biosensors or biofuel cells, a series of redox polymers was synthesized and used as immobilization matrix for Trametes hirsuta laccase. The properties of the obtained biofuel cell cathodes were compared with similar biocatalytic interfaces derived from redox polymers obtained via ligand exchange reaction of the parent Os-complex with a ligand integrated into the polymer backbone during the polymer synthesis. Copyright © 2011 Elsevier B.V. All rights reserved.

  1. Effects of reactive Mn(III)-oxalate complexes on structurally intact plant cell walls

    Science.gov (United States)

    Summering, J. A.; Keiluweit, M.; Goni, M. A.; Nico, P. S.; Kleber, M.

    2011-12-01

    Lignin components in the in plant litter are commonly assumed to have longer residence times in soil than many other compounds, which are supposedly, more easily degradable. The supposed resistance of lignin compounds to decomposition is generally attributed to the complex chain of biochemical steps required to create footholds in the non-porous structure of ligno-cellulose in cell walls. Interestingly, Mn(III) complexes have shown the ability to degrade ligno-cellulose. Mn(III) chelated by ligands such as oxalate are soluble oxidizers with a high affinity for lignin structures. Here we determined (i) the formation and decay kinetics of the Mn(III)-oxalate complexes in aqueous solution and (ii) the effects that these complexes have on intact ligno-cellulose. UV/vis spectroscopy and iodometric titrations confirmed the transient nature of Mn(III)-oxalate complexes with decay rates being in the order of hours. Zinnia elegans tracheary elements - a model ligno-cellulose substrate - were treated with Mn(III)-oxalate complexes in a newly developed flow-through reactor. Soluble decomposition products released during the treatment were analyzed by GC/MS and the degree of cell integrity was measured by cell counts, pre- and post-treatment counts indicate a decrease in intact Zinnia elegans as a result of Mn(III)-treatment. GC/MS results showed the release of a multitude of solubilized lignin breakdown products from plant cell walls. We conclude that Mn(III)-oxalate complexes have the ability to lyse intact plant cells and solubilize lignin. Lignin decomposition may thus be seen as resource dependent, with Mn(III) a powerful resource that should be abundant in terrestrial characterized by frequent redox fluctuations.

  2. A heteromeric molecular complex regulates the migration of lung alveolar epithelial cells during wound healing.

    Science.gov (United States)

    Ghosh, Manik C; Makena, Patrudu S; Kennedy, Joseph; Teng, Bin; Luellen, Charlean; Sinclair, Scott E; Waters, Christopher M

    2017-05-19

    Alveolar type II epithelial cells (ATII) are instrumental in early wound healing in response to lung injury, restoring epithelial integrity through spreading and migration. We previously reported in separate studies that focal adhesion kinase-1 (FAK) and the chemokine receptor CXCR4 promote epithelial repair mechanisms. However, potential interactions between these two pathways were not previously considered. In the present study, we found that wounding of rat ATII cells promoted increased association between FAK and CXCR4. In addition, protein phosphatase-5 (PP5) increased its association with this heteromeric complex, while apoptosis signal regulating kinase-1 (ASK1) dissociated from the complex. Cell migration following wounding was decreased when PP5 expression was decreased using shRNA, but migration was increased in ATII cells isolated from ASK1 knockout mice. Interactions between FAK and CXCR4 were increased upon depletion of ASK1 using shRNA in MLE-12 cells, but unaffected when PP5 was depleted. Furthermore, we found that wounded rat ATII cells exhibited decreased ASK1 phosphorylation at Serine-966, decreased serine phosphorylation of FAK, and decreased association of phosphorylated ASK1 with FAK. These changes in phosphorylation were dependent upon expression of PP5. These results demonstrate a unique molecular complex comprising CXCR4, FAK, ASK1, and PP5 in ATII cells during wound healing.

  3. Towards comprehensive cell lineage reconstructions in complex organisms using light-sheet microscopy.

    Science.gov (United States)

    Amat, Fernando; Keller, Philipp J

    2013-05-01

    Understanding the development of complex multicellular organisms as a function of the underlying cell behavior is one of the most fundamental goals of developmental biology. The ability to quantitatively follow cell dynamics in entire developing embryos is an indispensable step towards such a system-level understanding. In recent years, light-sheet fluorescence microscopy has emerged as a particularly promising strategy for recording the in vivo data required to realize this goal. Using light-sheet fluorescence microscopy, entire complex organisms can be rapidly imaged in three dimensions at sub-cellular resolution, achieving high temporal sampling and excellent signal-to-noise ratio without damaging the living specimen or bleaching fluorescent markers. The resulting datasets allow following individual cells in vertebrate and higher invertebrate embryos over up to several days of development. However, the complexity and size of these multi-terabyte recordings typically preclude comprehensive manual analyses. Thus, new computational approaches are required to automatically segment cell morphologies, accurately track cell identities and systematically analyze cell behavior throughout embryonic development. We review current efforts in light-sheet microscopy and bioimage informatics towards this goal, and argue that comprehensive cell lineage reconstructions are finally within reach for many key model organisms, including fruit fly, zebrafish and mouse. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

  4. Assembly and architecture of the EBV B cell entry triggering complex.

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    Karthik Sathiyamoorthy

    2014-08-01

    Full Text Available Epstein-Barr Virus (EBV is an enveloped double-stranded DNA virus of the gammaherpesvirinae sub-family that predominantly infects humans through epithelial cells and B cells. Three EBV glycoproteins, gH, gL and gp42, form a complex that targets EBV infection of B cells. Human leukocyte antigen (HLA class II molecules expressed on B cells serve as the receptor for gp42, triggering membrane fusion and virus entry. The mechanistic role of gHgL in herpesvirus entry has been largely unresolved, but it is thought to regulate the activation of the virally-encoded gB protein, which acts as the primary fusogen. Here we study the assembly and function of the reconstituted B cell entry complex comprised of gHgL, gp42 and HLA class II. The structure from negative-stain electron microscopy provides a detailed snapshot of an intermediate state in EBV entry and highlights the potential for the triggering complex to bring the two membrane bilayers into proximity. Furthermore, gHgL interacts with a previously identified, functionally important hydrophobic pocket on gp42, defining the overall architecture of the complex and playing a critical role in membrane fusion activation. We propose a macroscopic model of the initiating events in EBV B cell fusion centered on the formation of the triggering complex in the context of both viral and host membranes. This model suggests how the triggering complex may bridge the two membrane bilayers, orienting critical regions of the N- and C- terminal ends of gHgL to promote the activation of gB and efficient membrane fusion.

  5. A recombinant antibody with the antigen-specific, major histocompatibility complex-restricted specificity of T cells

    DEFF Research Database (Denmark)

    Andersen, P S; Stryhn, A; Hansen, B E

    1996-01-01

    Specific recognition of peptide/major histocompatibility complex (MHC) molecule complexes by the T-cell receptor is a key reaction in the specific immune response. Antibodies against peptide/MHC complexes would therefore be valuable tools in studying MHC function and T-cell recognition and might ...

  6. Cell division orientation is coupled to cell-cell adhesion by the E-cadherin/LGN complex

    NARCIS (Netherlands)

    Gloerich, Martijn; Bianchini, Julie M.; Siemers, Kathleen A.; Cohen, Daniel J.; Nelson, W. James

    2017-01-01

    Both cell-cell adhesion and oriented cell division play prominent roles in establishing tissue architecture, but it is unclear how they might be coordinated. Here, we demonstrate that the cell-cell adhesion protein E-cadherin functions as an instructive cue for cell division orientation. This is

  7. Cytotoxic property of surfactant-cobalt(III) complexes on a human breast cancer cell line.

    Science.gov (United States)

    Kumar, Rajendran Senthil; Riyasdeen, Anvarbatcha; Dinesh, Mohanakrishnan; Paul, Christo Preethy; Srinag, Suresh; Krishnamurthy, Hanumanthappa; Arunachalam, Sankaralingam; Akbarsha, Mohammad Abdulkadher

    2011-07-01

    The cancer chemotherapeutic potential of surfactant-cobalt(III) complexes, cis-[Co(bpy)(2)(C(14)H(29)NH(2))Cl](ClO(4))(2)·3 H(2)O (1) and cis-[Co(phen)(2)(C(14)H(29)NH(2))Cl](ClO(4))(2)·3 H(2)O (2) (bpy = 2,2'-bipyridine, phen = 1,10-phenanthroline) on MCF-7 breast cancer cell was determined adopting MTT assay and specific staining techniques. The complexes affected the viability of the cells significantly and the cells succumbed to apoptosis as seen in the changes in the nuclear morphology and cytoplasmic features. Since the complex 2 appeared to be more potent, further assays were carried out on the complex 2. Single-cell electrophoresis indicated DNA damage. The translocation of phosphatidyl serine and loss of mitochondrial potential was revealed by annexin V-Cy3 staining and JC-1 staining respectively. Western blot analysis revealed up-regulation of pro-apoptotic p53 and down-regulation of anti-apoptotic Bcl-2 protein. Taken together, the surfactant-cobalt(III) complex 2 would be a potential candidate for further investigation for application as a chemotherapeutic for cancers in general and estrogen receptor-positive breast cancer in particular. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Photodynamic killing of cancer cells by a Platinum(II) complex with cyclometallating ligand

    Science.gov (United States)

    Doherty, Rachel E.; Sazanovich, Igor V.; McKenzie, Luke K.; Stasheuski, Alexander S.; Coyle, Rachel; Baggaley, Elizabeth; Bottomley, Sarah; Weinstein, Julia A.; Bryant, Helen E.

    2016-03-01

    Photodynamic therapy that uses photosensitizers which only become toxic upon light-irradiation provides a strong alternative to conventional cancer treatment due to its ability to selectively target tumour material without affecting healthy tissue. Transition metal complexes are highly promising PDT agents due to intense visible light absorption, yet the majority are toxic even without light. This study introduces a small, photostable, charge-neutral platinum-based compound, Pt(II) 2,6-dipyrido-4-methyl-benzenechloride, complex 1, as a photosensitizer, which works under visible light. Activation of the new photosensitizer at low concentrations (0.1-1 μM) by comparatively low dose of 405 nm light (3.6 J cm-2) causes significant cell death of cervical, colorectal and bladder cancer cell lines, and, importantly, a cisplatin resistant cell line EJ-R. The photo-index of the complex is 8. We demonstrate that complex 1 induces irreversible DNA single strand breaks following irradiation, and that oxygen is essential for the photoinduced action. Neither light, nor compound alone led to cell death. The key advantages of the new drug include a remarkably fast accumulation time (diffusion-controlled, minutes), and photostability. This study demonstrates a highly promising new agent for photodynamic therapy, and attracts attention to photostable metal complexes as viable alternatives to conventional chemotherapeutics, such as cisplatin.

  9. B lymphocytes confer immune tolerance via cell surface GARP-TGF-β complex.

    Science.gov (United States)

    Wallace, Caroline H; Wu, Bill X; Salem, Mohammad; Ansa-Addo, Ephraim A; Metelli, Alessandra; Sun, Shaoli; Gilkeson, Gary; Shlomchik, Mark J; Liu, Bei; Li, Zihai

    2018-04-05

    GARP, a cell surface docking receptor for binding and activating latent TGF-β, is highly expressed by platelets and activated Tregs. While GARP is implicated in immune invasion in cancer, the roles of the GARP-TGF-β axis in systemic autoimmune diseases are unknown. Although B cells do not express GARP at baseline, we found that the GARP-TGF-β complex is induced on activated human and mouse B cells by ligands for multiple TLRs, including TLR4, TLR7, and TLR9. GARP overexpression on B cells inhibited their proliferation, induced IgA class-switching, and dampened T cell-independent antibody production. In contrast, B cell-specific deletion of GARP-encoding gene Lrrc32 in mice led to development of systemic autoimmune diseases spontaneously as well as worsening of pristane-induced lupus-like disease. Canonical TGF-β signaling more readily upregulates GARP in Peyer patch B cells than in splenic B cells. Furthermore, we demonstrated that B cells are required for the induction of oral tolerance of T cell-dependent antigens via GARP. Our studies reveal for the first time to our knowledge that cell surface GARP-TGF-β is an important checkpoint for regulating B cell peripheral tolerance, highlighting a mechanism of autoimmune disease pathogenesis.

  10. Stereological analysis of neuron, glial and endothelial cell numbers in the human amygdaloid complex.

    Directory of Open Access Journals (Sweden)

    María García-Amado

    Full Text Available Cell number alterations in the amygdaloid complex (AC might coincide with neurological and psychiatric pathologies with anxiety imbalances as well as with changes in brain functionality during aging. This stereological study focused on estimating, in samples from 7 control individuals aged 20 to 75 years old, the number and density of neurons, glia and endothelial cells in the entire AC and in its 5 nuclear groups (including the basolateral (BL, corticomedial and central groups, 5 nuclei and 13 nuclear subdivisions. The volume and total cell number in these territories were determined on Nissl-stained sections with the Cavalieri principle and the optical fractionator. The AC mean volume was 956 mm(3 and mean cell numbers (x10(6 were: 15.3 neurons, 60 glial cells and 16.8 endothelial cells. The numbers of endothelial cells and neurons were similar in each AC region and were one fourth the number of glial cells. Analysis of the influence of the individuals' age at death on volume, cell number and density in each of these 24 AC regions suggested that aging does not affect regional size or the amount of glial cells, but that neuron and endothelial cell numbers respectively tended to decrease and increase in territories such as AC or BL. These accurate stereological measures of volume and total cell numbers and densities in the AC of control individuals could serve as appropriate reference values to evaluate subtle alterations in this structure in pathological conditions.

  11. Treatment with at Homeopathic Complex Medication Modulates Mononuclear Bone Marrow Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Beatriz Cesar

    2011-01-01

    Full Text Available A homeopathic complex medication (HCM, with immunomodulatory properties, is recommended for patients with depressed immune systems. Previous studies demonstrated that the medication induces an increase in leukocyte number. The bone marrow microenvironment is composed of growth factors, stromal cells, an extracellular matrix and progenitor cells that differentiate into mature blood cells. Mice were our biological model used in this research. We now report in vivo immunophenotyping of total bone marrow cells and ex vivo effects of the medication on mononuclear cell differentiation at different times. Cells were examined by light microscopy and cytokine levels were measured in vitro. After in vivo treatment with HCM, a pool of cells from the new marrow microenvironment was analyzed by flow cytometry to detect any trend in cell alteration. The results showed decreases, mainly, in CD11b and TER-119 markers compared with controls. Mononuclear cells were used to analyze the effects of ex vivo HCM treatment and the number of cells showing ring nuclei, niche cells and activated macrophages increased in culture, even in the absence of macrophage colony-stimulating factor. Cytokines favoring stromal cell survival and differentiation in culture were induced in vitro. Thus, we observe that HCM is immunomodulatory, either alone or in association with other products.

  12. Stereological analysis of neuron, glial and endothelial cell numbers in the human amygdaloid complex.

    Science.gov (United States)

    García-Amado, María; Prensa, Lucía

    2012-01-01

    Cell number alterations in the amygdaloid complex (AC) might coincide with neurological and psychiatric pathologies with anxiety imbalances as well as with changes in brain functionality during aging. This stereological study focused on estimating, in samples from 7 control individuals aged 20 to 75 years old, the number and density of neurons, glia and endothelial cells in the entire AC and in its 5 nuclear groups (including the basolateral (BL), corticomedial and central groups), 5 nuclei and 13 nuclear subdivisions. The volume and total cell number in these territories were determined on Nissl-stained sections with the Cavalieri principle and the optical fractionator. The AC mean volume was 956 mm(3) and mean cell numbers (x10(6)) were: 15.3 neurons, 60 glial cells and 16.8 endothelial cells. The numbers of endothelial cells and neurons were similar in each AC region and were one fourth the number of glial cells. Analysis of the influence of the individuals' age at death on volume, cell number and density in each of these 24 AC regions suggested that aging does not affect regional size or the amount of glial cells, but that neuron and endothelial cell numbers respectively tended to decrease and increase in territories such as AC or BL. These accurate stereological measures of volume and total cell numbers and densities in the AC of control individuals could serve as appropriate reference values to evaluate subtle alterations in this structure in pathological conditions.

  13. Radiation-induced dissociation of stable DNA-protein complexes in Erlich ascites carcinoma cells

    International Nuclear Information System (INIS)

    Juhasz, P.P.; Sirota, N.P.; Gaziev, A.I.

    1982-01-01

    DNA of Ehrlich ascites carcinoma cells prepared under conditions that were highly denaturing for proteins but not for DNA, contained a group of nonhistone residual proteins. The amount of these proteins increased during DNA replication. The DNA-protein complex observed was sensitive to proteolytic enzymes and/or SH-reagents. γ-irradiation cells with moderate doses leads to a decrease in the amount of DNA-protein complexes. High-dose gamma-irradiation produces enhanced linking of chromosomal proteins with DNA. (author)

  14. Gamma-irradiation and neutron effect on DNA-membrane complexes of mammalian cells

    International Nuclear Information System (INIS)

    Lapidus, I.L.; Nazarov, V.M.; Ehrtsgreber, G.

    1984-01-01

    The first results of radiobiological investigations in the biophysical channel of the JINR reactor IBR-2 are presented. Sedimentation behaviour of DNA-membrane complexes has been studied at irradiation of the Chinese hamster cells (VT9-4) in a wide dose range of 137 Cs γ-irradiation and neutrons. An earlier assumption of the authors on the role of DNA double-strand breaks in changing the relative sedimentation velocity of complexes at irradiation of cells with doses over 50 Gy has been confirmed

  15. Complex forms of mitochondrial DNA in human B cells transformed by Epstein-Barr virus (EBV)

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Zeuthen, J

    1983-01-01

    Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed lymphoblast......Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed...

  16. Differential effects of a complex organochlorine mixture on the proliferation of breast cancer cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Aube, Michel, E-mail: 4aubem@videotron.ca [Axe de recherche en sante des populations et environnementale, Centre de recherche du Centre hospitalier universitaire de Quebec and Universite Laval, 2875 Boulevard Laurier, Edifice Delta 2, bureau 600, Quebec, QC, Canada G1V 2M2 (Canada); Larochelle, Christian, E-mail: christian.larochelle@inspq.qc.ca [Axe de recherche en sante des populations et environnementale, Centre de recherche du Centre hospitalier universitaire de Quebec and Universite Laval, 2875 Boulevard Laurier, Edifice Delta 2, bureau 600, Quebec, QC, Canada G1V 2M2 (Canada); Ayotte, Pierre, E-mail: pierre.ayotte@inspq.qc.ca [Axe de recherche en sante des populations et environnementale, Centre de recherche du Centre hospitalier universitaire de Quebec and Universite Laval, 2875 Boulevard Laurier, Edifice Delta 2, bureau 600, Quebec, QC, Canada G1V 2M2 (Canada); Laboratoire de Toxicologie, Institut national de sante publique du Quebec, 945 avenue Wolfe, Quebec, QC, Canada G1V 5B3 (Canada)

    2011-04-15

    Organochlorine compounds (OCs) are a group of persistent chemicals that accumulate in fatty tissues with age. Although OCs has been tested individually for their capacity to induce breast cancer cell proliferation, few studies examined the effect of complex mixtures that comprise compounds frequently detected in the serum of women. We constituted such an OC mixture containing 15 different components in environmentally relevant proportions and assessed its proliferative effects in four breast cancer cell lines (MCF-7, T47D, CAMA-1, MDAMB231) and in non-cancerous CV-1 cells. We also determined the capacity of the mixture to modulate cell cycle stage of breast cancer cells and to induce estrogenic and antiandrogenic effects using gene reporter assays. We observed that low concentrations of the mixture (100x10{sup 3} and 50x10{sup 3} dilutions) stimulated the proliferation of MCF-7 cells while higher concentrations (10x10{sup 3} and 5x10{sup 3} dilutions) had the opposite effect. In contrast, the mixture inhibited the proliferation of non-hormone-dependent cell lines. The mixture significantly increased the number of MCF-7 cells entering the S phase, an effect that was blocked by the antiestrogen ICI 182,780. Low concentrations of the mixture also caused an increase in CAMA-1 cell proliferation but only in the presence estradiol and dihydrotestosterone (p<0.05 at the 50x10{sup 3} dilution). DDT analogs and polychlorinated biphenyls all had the capacity to stimulate the proliferation of CAMA-1 cells in the presence of sex steroids. Reporter gene assays further revealed that the mixture and several of its constituents (DDT analogs, aldrin, dieldrin, {beta}-hexachlorocyclohexane, toxaphene) induced estrogenic effects, whereas the mixture and several components (DDT analogs, aldrin, dieldrin and PCBs) inhibited the androgen signaling pathway. Our results indicate that the complex OC mixture increases the proliferation of MCF-7 cells due to its estrogenic potential. The

  17. [Dinitrosyl iron complexes are endogenous signaling agents in animal and human cells and tissues (a hypothesis)].

    Science.gov (United States)

    Vanin, A F

    2004-01-01

    The hypothesis was advanced that dinitrosyl iron complexes generated in animal and human cells and tissues producing nitric oxide can function as endogenous universal regulators of biochemical and physiological processes. This function is realized by the ability of dinitrosyl iron complexes to act as donors of free nitric oxide molecules interacting with the heme groups of proteins, nitrosonium ions, or Fe+(NO+)2 interacting with the thiol groups of proteins. The effect of dinitrosyl iron complexes on the activity of some enzymes and the expression of the genome at the translation and transcription levels was considered.

  18. Proteomics-Based Analysis of Protein Complexes in Pluripotent Stem Cells and Cancer Biology.

    Science.gov (United States)

    Sudhir, Putty-Reddy; Chen, Chung-Hsuan

    2016-03-22

    A protein complex consists of two or more proteins that are linked together through protein-protein interactions. The proteins show stable/transient and direct/indirect interactions within the protein complex or between the protein complexes. Protein complexes are involved in regulation of most of the cellular processes and molecular functions. The delineation of protein complexes is important to expand our knowledge on proteins functional roles in physiological and pathological conditions. The genetic yeast-2-hybrid method has been extensively used to characterize protein-protein interactions. Alternatively, a biochemical-based affinity purification coupled with mass spectrometry (AP-MS) approach has been widely used to characterize the protein complexes. In the AP-MS method, a protein complex of a target protein of interest is purified using a specific antibody or an affinity tag (e.g., DYKDDDDK peptide (FLAG) and polyhistidine (His)) and is subsequently analyzed by means of MS. Tandem affinity purification, a two-step purification system, coupled with MS has been widely used mainly to reduce the contaminants. We review here a general principle for AP-MS-based characterization of protein complexes and we explore several protein complexes identified in pluripotent stem cell biology and cancer biology as examples.

  19. Contribution of Human Oral Cells to Astringency by Binding Salivary Protein/Tannin Complexes.

    Science.gov (United States)

    Soares, Susana; Ferrer-Galego, Raúl; Brandão, Elsa; Silva, Mafalda; Mateus, Nuno; Freitas, Victor de

    2016-10-10

    The most widely accepted mechanism to explain astringency is the interaction and precipitation of salivary proteins by food tannins, in particular proline-rich proteins. However, other mechanisms have been arising to explain astringency, such as binding of tannins to oral cells. In this work, an experimental method was adapted to study the possible contribution of both salivary proteins and oral cells to astringency induced by grape seed procyanidin fractions. Overall, in the absence of salivary proteins, the extent of procyanidin complexation with oral cells increased with increasing procyanidin degree of polymerization (mDP). Procyanidin fractions rich in monomers were the ones with the lowest ability to bind to oral cells. In the presence of salivary proteins and for procyanidins with mDP 2 the highest concentrations (1.5 and 2.0 mM) resulted in an increased binding of procyanidins to oral cells. This was even more evident for fractions III and IV at 1.0 mM and upper concentrations. Regarding the salivary proteins affected, it was possible to observe a decrease of P-B peptide and aPRP proteins for fractions II and III. This decrease is greater as the procyanidins' mDP increases. In fact, for fraction IV an almost total depletion of all salivary proteins was observed. This decrease is due to the formation of insoluble salivary protein/procyanidin complexes. Altogether, these data suggest that some procyanidins are able to bind to oral cells and that the salivary proteins interact with procyanidins forming salivary protein/procyanidin complexes that are also able to link to oral cells. The procyanidins that remain unbound to oral cells are able to bind to salivary proteins forming a large network of salivary protein/procyanidin complexes. Overall, the results presented herein provide one more step to understand food oral astringency onset.

  20. Processive movement of MreB-associated cell wall biosynthetic complexes in bacteria.

    Science.gov (United States)

    Domínguez-Escobar, Julia; Chastanet, Arnaud; Crevenna, Alvaro H; Fromion, Vincent; Wedlich-Söldner, Roland; Carballido-López, Rut

    2011-07-08

    The peptidoglycan cell wall and the actin-like MreB cytoskeleton are major determinants of cell shape in rod-shaped bacteria. The prevailing model postulates that helical, membrane-associated MreB filaments organize elongation-specific peptidoglycan-synthesizing complexes along sidewalls. We used total internal reflection fluorescence microscopy to visualize the dynamic relation between MreB isoforms and cell wall synthesis in live Bacillus subtilis cells. During exponential growth, MreB proteins did not form helical structures. Instead, together with other morphogenetic factors, they assembled into discrete patches that moved processively along peripheral tracks perpendicular to the cell axis. Patch motility was largely powered by cell wall synthesis, and MreB polymers restricted diffusion of patch components in the membrane and oriented patch motion.

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

    Directory of Open Access Journals (Sweden)

    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. Functional cooperation between FACT and MCM is coordinated with cell cycle and differential complex formation

    Directory of Open Access Journals (Sweden)

    Lin Chih-Li

    2010-02-01

    Full Text Available Abstract Background Functional cooperation between FACT and the MCM helicase complex constitutes an integral step during DNA replication initiation. However, mode of regulation that underlies the proper functional interaction of FACT and MCM is poorly understood. Methods & Results Here we present evidence indicating that such interaction is coordinated with cell cycle progression and differential complex formation. We first demonstrate the existence of two distinct FACT-MCM subassemblies, FACT-MCM2/4/6/7 and FACT-MCM2/3/4/5. Both complexes possess DNA unwinding activity and are subject to cell cycle-dependent enzymatic regulation. Interestingly, analysis of functional attributes further suggests that they act at distinct, and possibly sequential, steps during origin establishment and replication initiation. Moreover, we show that the phosphorylation profile of the FACT-associated MCM4 undergoes a cell cycle-dependent change, which is directly correlated with the catalytic activity of the FACT-MCM helicase complexes. Finally, at the quaternary structure level, physical interaction between FACT and MCM complexes is generally dependent on persistent cell cycle and further stabilized upon S phase entry. Cessation of mitotic cycle destabilizes the complex formation and likely leads to compromised coordination and activities. Conclusions Together, our results correlate FACT-MCM functionally and temporally with S phase and DNA replication. They further demonstrate that enzymatic activities intrinsically important for DNA replication are tightly controlled at various levels, thereby ensuring proper progression of, as well as exit from, the cell cycle and ultimately euploid gene balance.

  3. Arp2/3 complex activity in filopodia of spreading cells

    Directory of Open Access Journals (Sweden)

    Mendes Paula M

    2008-12-01

    Full Text Available Abstract Background Cells use filopodia to explore their environment and to form new adhesion contacts for motility and spreading. The Arp2/3 complex has been implicated in lamellipodial actin assembly as a major nucleator of new actin filaments in branched networks. The interplay between filopodial and lamellipodial protrusions is an area of much interest as it is thought to be a key determinant of how cells make motility choices. Results We find that Arp2/3 complex localises to dynamic puncta in filopodia as well as lamellipodia of spreading cells. Arp2/3 complex spots do not appear to depend on local adhesion or on microtubules for their localisation but their inclusion in filopodia or lamellipodia depends on the activity of the small GTPase Rac1. Arp2/3 complex spots in filopodia are capable of incorporating monomeric actin, suggesting the presence of available filament barbed ends for polymerisation. Arp2/3 complex in filopodia co-localises with lamellipodial proteins such as capping protein and cortactin. The dynamics of Arp2/3 complex puncta suggests that they are moving bi-directionally along the length of filopodia and that they may be regions of lamellipodial activity within the filopodia. Conclusion We suggest that filopodia of spreading cells have regions of lamellipodial activity and that this activity affects the morphology and movement of filopodia. Our work has implications for how we understand the interplay between lamellipodia and filopodia and for how actin networks are generated spatially in cells.

  4. X-ray-enhanced cancer cell migration requires the linker of nucleoskeleton and cytoskeleton complex.

    Science.gov (United States)

    Imaizumi, Hiromasa; Sato, Katsutoshi; Nishihara, Asuka; Minami, Kazumasa; Koizumi, Masahiko; Matsuura, Nariaki; Hieda, Miki

    2018-04-01

    The linker of nucleoskeleton and cytoskeleton (LINC) complex is a multifunctional protein complex that is involved in various processes at the nuclear envelope, including nuclear migration, mechanotransduction, chromatin tethering and DNA damage response. We recently showed that a nuclear envelope protein, Sad1 and UNC84 domain protein 1 (SUN1), a component of the LINC complex, has a critical function in cell migration. Although ionizing radiation activates cell migration and invasion in vivo and in vitro, the underlying molecular mechanism remains unknown. Here, we examined the involvement of the LINC complex in radiation-enhanced cell migration and invasion. A sublethal dose of X-ray radiation promoted human breast cancer MDA-MB-231 cell migration and invasion, whereas carbon ion beam radiation suppressed these processes in a dose-dependent manner. Depletion of SUN1 and SUN2 significantly suppressed X-ray-enhanced cell migration and invasion. Moreover, depletion or overexpression of each SUN1 splicing variant revealed that SUN1_888 containing 888 amino acids of SUN1 but not SUN1_916 was required for X-ray-enhanced migration and invasion. In addition, the results suggested that X-ray irradiation affected the expression level of SUN1 splicing variants and a SUN protein binding partner, nesprins. Taken together, our findings supported that the LINC complex contributed to photon-enhanced cell migration and invasion. © 2018 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  5. Detection of autoreactive CD4 T cells using major histocompatibility complex class II dextramers

    Directory of Open Access Journals (Sweden)

    Kuszynski Charles

    2011-07-01

    Full Text Available Abstract Background Tetramers are useful tools to enumerate the frequencies of antigen-specific T cells. However, unlike CD8 T cells, CD4 T cells - especially self-reactive cells - are challenging to detect with major histocompatibility complex (MHC class II tetramers because of low frequencies and low affinities of their T cell receptors to MHC-peptide complexes. Here, we report the use of fluorescent multimers, designated MHC dextramers that contain a large number of peptide-MHC complexes per reagent. Results The utility of MHC dextramers was evaluated in three autoimmune disease models: 1 proteolipid protein (PLP 139-151-induced experimental autoimmune encephalomyelitis in SJL/J (H-2s mice; 2 myelin oligodendrocyte glycoprotein (MOG 35-55-induced experimental autoimmune encephalomyelitis in C57Bl/6 (H-2b mice; and 3 cardiac myosin heavy chain (Myhc-α 334-352-induced experimental autoimmune myocarditis in A/J (H-2a mice. Flow cytometrically, we demonstrate that IAs/PLP 139-151, IAb/MOG 35-55 and IAk/Myhc-α 334-352 dextramers detect the antigen-sensitized cells with specificity, and with a detection sensitivity significantly higher than that achieved with conventional tetramers. Furthermore, we show that binding of dextramers, but not tetramers, is less dependent on the activation status of cells, permitting enumeration of antigen-specific cells ex vivo. Conclusions The data suggest that MHC dextramers are useful tools to track the generation and functionalities of self-reactive CD4 cells in various experimental systems.

  6. Rickettsia parkeri invasion of diverse host cells involves an Arp2/3 complex, WAVE complex and Rho-family GTPase-dependent pathway.

    Science.gov (United States)

    Reed, Shawna C O; Serio, Alisa W; Welch, Matthew D

    2012-04-01

    Rickettsiae are obligate intracellular pathogens that are transmitted to humans by arthropod vectors and cause diseases such as spotted fever and typhus. Although rickettsiae require the host cell actin cytoskeleton for invasion, the cytoskeletal proteins that mediate this process have not been completely described. To identify the host factors important during cell invasion by Rickettsia parkeri, a member of the spotted fever group (SFG), we performed an RNAi screen targeting 105 proteins in Drosophila melanogaster S2R+ cells. The screen identified 21 core proteins important for invasion, including the GTPases Rac1 and Rac2, the WAVE nucleation-promoting factor complex and the Arp2/3 complex. In mammalian cells, including endothelial cells, the natural targets of R. parkeri, the Arp2/3 complex was also crucial for invasion, while requirements for WAVE2 as well as Rho GTPases depended on the particular cell type. We propose that R. parkeri invades S2R+ arthropod cells through a primary pathway leading to actin nucleation, whereas invasion of mammalian endothelial cells occurs via redundant pathways that converge on the host Arp2/3 complex. Our results reveal a key role for the WAVE and Arp2/3 complexes, as well as a higher degree of variation than previously appreciated in actin nucleation pathways activated during Rickettsia invasion. © 2011 Blackwell Publishing Ltd.

  7. CD4/CD8/Dendritic cell complexes in the spleen: CD8+ T cells can directly bind CD4+ T cells and modulate their response

    Science.gov (United States)

    Barinov, Aleksandr; Galgano, Alessia; Krenn, Gerald; Tanchot, Corinne; Vasseur, Florence

    2017-01-01

    CD4+ T cell help to CD8+ T cell responses requires that CD4+ and CD8+ T cells interact with the same antigen presenting dendritic cell (Ag+DC), but it remains controversial whether helper signals are delivered indirectly through a licensed DC and/or involve direct CD4+/CD8+ T cell contacts and/or the formation of ternary complexes. We here describe the first in vivo imaging of the intact spleen, aiming to evaluate the first interactions between antigen-specific CD4+, CD8+ T cells and Ag+DCs. We show that in contrast to CD4+ T cells which form transient contacts with Ag+DC, CD8+ T cells form immediate stable contacts and activate the Ag+DC, acquire fragments of the DC membranes by trogocytosis, leading to their acquisition of some of the DC properties. They express MHC class II, and become able to present the specific Marilyn peptide to naïve Marilyn CD4+ T cells, inducing their extensive division. In vivo, these CD8+ T cells form direct stable contacts with motile naïve CD4+ T cells, recruiting them to Ag+DC binding and to the formation of ternary complexes, where CD4+ and CD8+ T cells interact with the DC and with one another. The presence of CD8+ T cells during in vivo immune responses leads to the early activation and up-regulation of multiple functions by CD4+ T lymphocytes. Thus, while CD4+ T cell help is important to CD8+ T cell responses, CD8+ T cells can interact directly with naïve CD4+ T cells impacting their recruitment and differentiation. PMID:28686740

  8. Cdt1 revisited: complex and tight regulation during the cell cycle and consequences of deregulation in mammalian cells

    Directory of Open Access Journals (Sweden)

    Fujita Masatoshi

    2006-10-01

    Full Text Available Abstract In eukaryotic cells, replication of genomic DNA initiates from multiple replication origins distributed on multiple chromosomes. To ensure that each origin is activated precisely only once during each S phase, a system has evolved which features periodic assembly and disassembly of essential pre-replication complexes (pre-RCs at replication origins. The pre-RC assembly reaction involves the loading of a presumptive replicative helicase, the MCM2-7 complexes, onto chromatin by the origin recognition complex (ORC and two essential factors, CDC6 and Cdt1. The eukaryotic cell cycle is driven by the periodic activation and inactivation of cyclin-dependent kinases (Cdks and assembly of pre-RCs can only occur during the low Cdk activity period from late mitosis through G1 phase, with inappropriate re-assembly suppressed during S, G2, and M phases. It was originally suggested that inhibition of Cdt1 function after S phase in vertebrate cells is due to geminin binding and that Cdt1 hyperfunction resulting from Cdt1-geminin imbalance induces re-replication. However, recent progress has revealed that Cdt1 activity is more strictly regulated by two other mechanisms in addition to geminin: (1 functional and SCFSkp2-mediated proteolytic regulation through phosphorylation by Cdks; and (2 replication-coupled proteolysis mediated by the Cullin4-DDB1Cdt2 ubiquitin ligase and PCNA, an eukaryotic sliding clamp stimulating replicative DNA polymerases. The tight regulation implies that Cdt1 control is especially critical for the regulation of DNA replication in mammalian cells. Indeed, Cdt1 overexpression evokes chromosomal damage even without re-replication. Furthermore, deregulated Cdt1 induces chromosomal instability in normal human cells. Since Cdt1 is overexpressed in cancer cells, this could be a new molecular mechanism leading to carcinogenesis. In this review, recent insights into Cdt1 function and regulation in mammalian cells are discussed.

  9. Complexities in human herpesvirus-6A and -6B binding to host cells

    International Nuclear Information System (INIS)

    Pedersen, Simon Metz; Hoellsberg, Per

    2006-01-01

    Human herpesvirus-6A and -6B uses the cellular receptor CD46 for fusion and infection of the host cell. The viral glycoprotein complex gH-gL from HHV-6A binds to the short consensus repeat 2 and 3 in CD46. Although all the major isoforms of CD46 bind the virus, certain isoforms may have higher affinity than others for the virus. Within recent years, elucidation of the viral complex has identified additional HHV-6A and -6B specific glycoproteins. Thus, gH-gL associates with a gQ1-gQ2 dimer to form a heterotetrameric complex. In addition, a novel complex consisting of gH-gL-gO has been described that does not bind CD46. Accumulating evidence suggests that an additional HHV-6A and -6B receptor exists. The previous simple picture of HHV-6A/B-host cell contact therefore includes more layers of complexities on both the viral and the host cell side of the interaction

  10. Toxicity study of complex CNT-PEG(-NH2)-DOX synthesis on neuroblastoma cells

    Science.gov (United States)

    Nurulhuda, I.; Mazatulikhma, M. Z.; Alrokayan, S.; Khan, H.; Rusop, M.

    2018-05-01

    The synthesized carbon nanotubes was functionalized with PEG and drug (doxorubicin) was tested on neuroblastoma cells. The treatment was done for 24 and 48 h. The concentration of CNT and doxorubicin were at 2.5, 5, 10 µg/ml and 0.5, 0.1, 0.05 µM, respectively. The result showed the longer time treatment do have effect on the cells viability and the complex functionalized CNT have high cells viability rather than the drug and CNT treatment alone.

  11. The number of cell types, information content, and the evolution of complex multicellularity

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    Karl J. Niklas

    2014-12-01

    Full Text Available The number of different cell types (NCT characterizing an organism is often used to quantify organismic complexity. This method results in the tautology that more complex organisms have a larger number of different kinds of cells, and that organisms with more different kinds of cells are more complex. This circular reasoning can be avoided (and simultaneously tested when NCT is plotted against different measures of organismic information content (e.g., genome or proteome size. This approach is illustrated by plotting the NCT of representative diatoms, green and brown algae, land plants, invertebrates, and vertebrates against data for genome size (number of base-pairs, proteome size (number of amino acids, and proteome functional versatility (number of intrinsically disordered protein domains or residues. Statistical analyses of these data indicate that increases in NCT fail to keep pace with increases in genome size, but exceed a one-to-one scaling relationship with increasing proteome size and with increasing numbers of intrinsically disordered protein residues. We interpret these trends to indicate that comparatively small increases in proteome (and not genome size are associated with disproportionate increases in NCT, and that proteins with intrinsically disordered domains enhance cell type diversity and thus contribute to the evolution of complex multicellularity.

  12. Experimental broadband absorption enhancement in silicon nanohole structures with optimized complex unit cells.

    Science.gov (United States)

    Lin, Chenxi; Martínez, Luis Javier; Povinelli, Michelle L

    2013-09-09

    We design silicon membranes with nanohole structures with optimized complex unit cells that maximize broadband absorption. We fabricate the optimized design and measure the optical absorption. We demonstrate an experimental broadband absorption about 3.5 times higher than an equally-thick thin film.

  13. Complexities in human herpesvirus-6A and -6B binding to host cells

    DEFF Research Database (Denmark)

    Pedersen, Simon Metz; Höllsberg, Per

    2006-01-01

    Human herpesvirus-6A and -6B uses the cellular receptor CD46 for fusion and infection of the host cell. The viral glycoprotein complex gH-gL from HHV-6A binds to the short consensus repeat 2 and 3 in CD46. Although all the major isoforms of CD46 bind the virus, certain isoforms may have higher...

  14. Paving the way towards complex blood-brain barrier models using pluripotent stem cells

    DEFF Research Database (Denmark)

    Lauschke, Karin; Frederiksen, Lise; Hall, Vanessa Jane

    2017-01-01

    , it is now possible to produce many cell types from the BBB and even partially recapitulate this complex tissue in vitro. In this review, we summarize the most recent developments in PSC differentiation and modelling of the BBB. We also suggest how patient-specific human induced PSCs could be used to model...

  15. Universal lab-on-a-chip platform for complex, perfused 3D cell cultures

    Science.gov (United States)

    Sonntag, F.; Schmieder, F.; Ströbel, J.; Grünzner, S.; Busek, M.; Günther, K.; Steege, T.; Polk, C.; Klotzbach, U.

    2016-03-01

    The miniaturization, rapid prototyping and automation of lab-on-a-chip technology play nowadays a very important role. Lab-on-a-chip technology is successfully implemented not only for environmental analysis and medical diagnostics, but also as replacement of animals used for the testing of substances in the pharmaceutical and cosmetics industries. For that purpose the Fraunhofer IWS and partners developed a lab-on-a-chip platform for perfused cell-based assays in the last years, which includes different micropumps, valves, channels, reservoirs and customized cell culture modules. This technology is already implemented for the characterization of different human cell cultures and organoids, like skin, liver, endothelium, hair follicle and nephron. The advanced universal lab-on-a-chip platform for complex, perfused 3D cell cultures is divided into a multilayer basic chip with integrated micropump and application-specific 3D printed cell culture modules. Moreover a technology for surface modification of the printed cell culture modules by laser micro structuring and a complex and flexibly programmable controlling device based on an embedded Linux system was developed. A universal lab-on-a-chip platform with an optional oxygenator and a cell culture module for cubic scaffolds as well as first cell culture experiments within the cell culture device will be presented. The module is designed for direct interaction with robotic dispenser systems. This offers the opportunity to combine direct organ printing of cells and scaffolds with the microfluidic cell culture module. The characterization of the developed system was done by means of Micro-Particle Image Velocimetry (μPIV) and an optical oxygen measuring system.

  16. Basal cell carcinomas of the areola-nipple complex: case reports and review of the literature.

    Science.gov (United States)

    Betti, Roberto; Martino, Patrizia; Moneghini, Laura; Vergani, Raffaelle; Tolomio, Elena; Crosti, Carlo

    2003-11-01

    Two white men 57 and 39 years old, and a 47-year-old white woman were seen with slowly developing papulo-nodular lesions of the areola-nipple complex. None of the patients presented with regional lymphadenopathy, history of trauma, or relevant sun-exposure. After excison of the mass, the histologic diagnosis of basal cell carcinoma was made. At two years of follow-up, no recurrence was evident. The low incidence of basal cell carcinoma in this particular site allows us to consider the areola-nipple complex location as unusual. Moreover, literature reports do not suggest that these BCCs have an increased potential for malignancy. The treatment options depend on the extension of the tumor and on the possible involvement of the areola-nipple complex and mammary tissue.

  17. FUNCTION OF MALATDEHYDROGENASE COMPLEX OF MAIZE MESOPHYLL AND BUNDLE SHEATH CELLS UNDER SALT STRESS CONDITION

    Directory of Open Access Journals (Sweden)

    Еprintsev А.Т.

    2006-12-01

    Full Text Available Salt-induced changes in malatdehydrogenase system activity make the essential contribution to cell adaptation to stress condition. The enzyme systems of C4-plants are most interesting due to their ability for adaptation to environment conditions. The role of separate components of malatdehydrogenase complex of mesophyll and bundle sheath cells of corn in formation of adaptive reaction in stressful conditions is investigated in presented work.The activation of all enzymes of malatdehydrogenase system and the subsequent decrease in their activity was observed in mesophyll durring the first stage of adaptation to salt influence. In bundle sheath cells such parameters are differed from control less essentially. Fast accumulation of piruvate in cells and malate in both investigated tissues was induced. The further salinity led to falling of concentration this intermediate. The concentration of piruvate was below control level, and it was raised by the end of an exposition.The results show that sodium chloride causes induction of Krebs-cycle in mesophyll and bundle sheath cells of corn and intensification of Hatch-Slack cycle. The described differences in function malatdehydrogenase systems of mesophyll and bundle sheath cells of leaves of corn under salinity mainly consist of the activity of enzymes of a studied complex in bundle sheath cells is subject to the minimal changes in comparison with mesophyll. Role of this enzymesystem in mechanisms of adaptive reaction of various tissues of corn to salt stress is discussed.

  18. A protocadherin-cadherin-FLRT3 complex controls cell adhesion and morphogenesis.

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    Xuejun Chen

    2009-12-01

    Full Text Available Paraxial protocadherin (PAPC and fibronectin leucine-rich domain transmembrane protein-3 (FLRT3 are induced by TGFbeta signaling in Xenopus embryos and both regulate morphogenesis by inhibiting C-cadherin mediated cell adhesion.We have investigated the functional and physical relationships between PAPC, FLRT3, and C-cadherin. Although neither PAPC nor FLRT3 are required for each other to regulate C-cadherin adhesion, they do interact functionally and physically, and they form a complex with cadherins. By itself PAPC reduces cell adhesion physiologically to induce cell sorting, while FLRT3 disrupts adhesion excessively to cause cell dissociation. However, when expressed together PAPC limits the cell dissociating and tissue disrupting activity of FLRT3 to make it effective in physiological cell sorting. PAPC counteracts FLRT3 function by inhibiting the recruitment of the GTPase RND1 to the FLRT3 cytoplasmic domain.PAPC and FLRT3 form a functional complex with cadherins and PAPC functions as a molecular "governor" to maintain FLRT3 activity at the optimal level for physiological regulation of C-cadherin adhesion, cell sorting, and morphogenesis.

  19. The Growing Complexity of Cancer Cell Response to DNA-Damaging Agents: Caspase 3 Mediates Cell Death or Survival?

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    Razmik Mirzayans

    2016-05-01

    Full Text Available It is widely stated that wild-type p53 either mediates the activation of cell cycle checkpoints to facilitate DNA repair and promote cell survival, or orchestrates apoptotic cell death following exposure to cancer therapeutic agents. This reigning paradigm has been challenged by numerous discoveries with different human cell types, including solid tumor-derived cell lines. Thus, activation of the p53 signaling pathway by ionizing radiation and other DNA-damaging agents hinders apoptosis and triggers growth arrest (e.g., through premature senescence in some genetic backgrounds; such growth arrested cells remain viable, secrete growth-promoting factors, and give rise to progeny with stem cell-like properties. In addition, caspase 3, which is best known for its role in the execution phase of apoptosis, has been recently reported to facilitate (rather than suppress DNA damage-induced genomic instability and carcinogenesis. This observation is consistent with an earlier report demonstrating that caspase 3 mediates secretion of the pro-survival factor prostaglandin E2, which in turn promotes enrichment of tumor repopulating cells. In this article, we review these and related discoveries and point out novel cancer therapeutic strategies. One of our objectives is to demonstrate the growing complexity of the DNA damage response beyond the conventional “repair and survive, or die” hypothesis.

  20. Toxic effect of C60 fullerene-doxorubicin complex towards tumor and normal cells in vitro

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    Prylutska S. V.

    2014-09-01

    Full Text Available Creation of new nanostructures possessing high antitumor activity is an important problem of modern biotechnology. Aim. To evaluate cytotoxicity of created complex of pristine C60 fullerene with the anthracycline antibiotic doxorubicin (Dox, as well as of free C60 fullerene and Dox, towards different cell types – tumor, normal immunocompetent and hepatocytes. Methods. Measurement of size distribution for particles in C60 + Dox mixture was performed by a dynamic light scattering (DLS technique. Toxic effect of C60 + Dox complex in vitro towards tumor and normal cells was studied using the MTT assay. Results. DLS experiment demonstrated that the main fraction of the particles in C60 + Dox mixture had a diameter in the range of about 132 nm. The toxic effect of C60 + Dox complex towards normal (lymphocytes, macrophages, hepatocytes and tumor (Ehrlich ascites carcinoma, leukemia L1210, Lewis lung carcinoma cells was decreased by ~10–16 % and ~7–9 %, accordingly, compared with the same effect of free Dox. Conclusions. The created C60 + Dox composite may be considered as a new pharmacological agent that kills effectively tumor cells in vitro and simultaneously prevents a toxic effect of the free form of Dox on normal cells.

  1. The Complex Interaction of Matrix Metalloproteinases in the Migration of Cancer Cells through Breast Tissue Stroma

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    Kerry J. Davies

    2014-01-01

    Full Text Available Breast cancer mortality is directly linked to metastatic spread. The metastatic cell must exhibit a complex phenotype that includes the capacity to escape from the primary tumour mass, invade the surrounding normal tissue, and penetrate into the circulation before proliferating in the parenchyma of distant organs to produce a metastasis. In the normal breast, cellular structures change cyclically in response to ovarian hormones leading to regulated cell proliferation and apoptosis. Matrix metalloproteinases (MMPs are a family of zinc dependent endopeptidases. Their primary function is degradation of proteins in the extracellular matrix to allow ductal progression through the basement membrane. A complex balance between matrix metalloproteinases and their inhibitors regulate these changes. These proteinases interact with cytokines, growth factors, and tumour necrosis factors to stimulate branching morphologies in normal breast tissues. In breast cancer this process is disrupted facilitating tumour progression and metastasis and inhibiting apoptosis increasing the life of the metastatic cells. This paper highlights the role of matrix metalloproteinases in cell progression through the breast stroma and reviews the complex relationships between the different proteinases and their inhibitors in relation to breast cancer cells as they metastasise.

  2. Ultrastructural Complexity of Nuclear Components During Early Apoptotic Phases in Breast Cancer Cells

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    Christian Castelli

    2001-01-01

    Full Text Available Fractal morphometry was used to investigate the ultrastructural features of the plasma membrane, perinuclear membrane and nuclear chromatin in SK‐BR‐3 human breast cancer cells undergoing apoptosis. Cells were incubated with 1 μM calcimycin (A23187 for 24 h. Cells in the early stage of apoptosis had fractal dimension (FD values indicating that their plasma membranes were less rough (lower FD than those of control cells, while their perinuclear membranes were unaffected. Changes of the chromatin texture within the entire nucleus and in selected nuclear domains were more pronounced in treated cells. This confirms that the morphological reorganization imputable to a loss of structural complexity (reduced FD occurs in the early stage of apoptosis, is accompanied by the inhibition of distinct enzymatic events and precedes the onset of conventional cellular markers, which can only be detected during the active phases of the apoptotic process.

  3. Nuclear DAMP complex-mediated RAGE-dependent macrophage cell death

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ruochan [Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213 (United States); Department of Infectious Diseases and State Key Lab of Viral Hepatitis, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Fu, Sha; Fan, Xue-Gong [Department of Infectious Diseases and State Key Lab of Viral Hepatitis, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Lotze, Michael T.; Zeh, Herbert J. [Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213 (United States); Tang, Daolin, E-mail: tangd2@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213 (United States); Kang, Rui, E-mail: kangr@upmc.edu [Department of Surgery, University of Pittsburgh, Pittsburgh, PA 15213 (United States)

    2015-03-13

    High mobility group box 1 (HMGB1), histone, and DNA are essential nuclear components involved in the regulation of chromosome structure and function. In addition to their nuclear function, these molecules act as damage-associated molecular patterns (DAMPs) alone or together when released extracellularly. The synergistic effect of these nuclear DNA-HMGB1-histone complexes as DAMP complexes (nDCs) on immune cells remains largely unexplored. Here, we demonstrate that nDCs limit survival of macrophages (e.g., RAW264.7 and peritoneal macrophages) but not cancer cells (e.g., HCT116, HepG2 and Hepa1-6). nDCs promote production of inflammatory tumor necrosis factor α (TNFα) release, triggering reactive oxygen species-dependent apoptosis and necrosis. Moreover, the receptor for advanced glycation end products (RAGE), but not toll-like receptor (TLR)-4 and TLR-2, was required for Akt-dependent TNFα release and subsequent cell death following treatment with nDCs. Genetic depletion of RAGE by RNAi, antioxidant N-Acetyl-L-cysteine, and TNFα neutralizing antibody significantly attenuated nDC-induced cell death. These findings provide evidence supporting novel signaling mechanisms linking nDCs and inflammation in macrophage cell death. - Highlights: • Nuclear DAMP complexes (nDCs) selectively induce cell death in macrophages, but not cancer cells. • TNFα-mediated oxidative stress is required for nDC-induced death. • RAGE-mediated Akt activation is required for nDC-induced TNFα release. • Blocking RAGE and TNFα inhibits nDC-induced macrophage cell death.

  4. Ultrastructural characteristics of nurse cell-larva complex of four species of Trichinella in several hosts

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    Sacchi L.

    2001-06-01

    Full Text Available The nurse cell-larva complex of nematodes of the genus Trichinella plays an Important role in the survival of the larva in decaying muscles, frequently favouring the transmission of the parasite in extreme environmental conditions. The ultrastructure of the nurse cell-larva complex in muscles from different hosts infected with T. nativa (a walrus and a polar bear, T. spiralis (horses and humans, T. pseudospiralis (a laboratory mouse and T. papuae (a laboratory mouse were examined. Analysis with transmission electron microscope showed that the typical nurse cell structure was present in all examined samples, irrespective of the species of larva, of the presence of a collagen capsule, of the age of infection and of the host species, suggesting that there exists a molecular mechanism that in the first stage of larva invasion is similar for encapsulated and non-encapsulated species.

  5. Swarming and complex pattern formation in Paenibacillus vortex studied by imaging and tracking cells

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    Jacob Eshel

    2008-02-01

    Full Text Available Abstract Background Swarming motility allows microorganisms to move rapidly over surfaces. The Gram-positive bacterium Paenibacillus vortex exhibits advanced cooperative motility on agar plates resulting in intricate colonial patterns with geometries that are highly sensitive to the environment. The cellular mechanisms that underpin the complex multicellular organization of such a simple organism are not well understood. Results Swarming by P. vortex was studied by real-time light microscopy, by in situ scanning electron microscopy and by tracking the spread of antibiotic-resistant cells within antibiotic-sensitive colonies. When swarming, P. vortex was found to be peritrichously flagellated. Swarming by the curved cells of P. vortex occurred on an extremely wide range of media and agar concentrations (0.3 to 2.2% w/v. At high agar concentrations (> 1% w/v rotating colonies formed that could be detached from the main mass of cells by withdrawal of cells into the latter. On lower percentage agars, cells moved in an extended network composed of interconnected "snakes" with short-term collision avoidance and sensitivity to extracts from swarming cells. P. vortex formed single Petri dish-wide "supercolonies" with a colony-wide exchange of motile cells. Swarming cells were coupled by rapidly forming, reversible and non-rigid connections to form a loose raft, apparently connected via flagella. Inhibitors of swarming (p-Nitrophenylglycerol and Congo Red were identified. Mitomycin C was used to trigger filamentation without inhibiting growth or swarming; this facilitated dissection of the detail of swarming. Mitomycin C treatment resulted in malcoordinated swarming and abortive side branch formation and a strong tendency by a subpopulation of the cells to form minimal rotating aggregates of only a few cells. Conclusion P. vortex creates complex macroscopic colonies within which there is considerable reflux and movement and interaction of cells. Cell

  6. T cell differentiation stages identified by molecular and immunologic analysis of the T cell receptor complex in childhood lymphoblastic leukemia.

    Science.gov (United States)

    Mirro, J; Kitchingman, G; Behm, F G; Murphy, S B; Goorha, R M

    1987-03-01

    T cell differentiation was investigated by determining the relationship of T cell receptor (Ti) gene rearrangement and transcription to the expression of surface and cytoplasmic T3 antigen using blast cells from five children with acute lymphoblastic leukemia of thymic origin. Patterns of monoclonal antibody (MoAb) reactivity indicated that these cases were representative of the three recognized stages (I, II, III) of human thymocyte development. The T3 antigen, which becomes linked to the Ti to form a functional T cell receptor complex on mature thymocytes, was expressed on the cell surface in two cases (stage III). However, in the remaining three cases that were surface T3 negative (stages I and II), large amounts of T3 were identified in the cytoplasm by immunoperoxidase staining and flow cytometry. Leukemic blasts from all five patients showed rearranged genes encoding the beta-chain portion of the Ti heterodimer. RNA transcripts of Ti beta-chain genes were also evident in lymphoblasts from all five cases, but transcripts coding for the alpha-chain portion of Ti were found only in cases that expressed T3 on the cell surface. Thus the absence of surface T3 (and presumably Ti) coincides with the absence of Ti alpha-chain RNA, suggesting that transcription of alpha-chain genes is a critical regulatory event in the surface expression of the Ti-T3 complex. Leukemic T cells that rearrange and express Ti beta-chain genes but lack Ti alpha-chain messenger RNA (mRNA) may represent a stage of differentiation analogous to pre-B cells, where heavy-chain immunoglobulin (Ig) genes are rearranged and expressed but light-chain Ig genes are not expressed.

  7. Sulforaphane induces cell cycle arrest by protecting RB-E2F-1 complex in epithelial ovarian cancer cells

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    Morris Robert

    2010-03-01

    Full Text Available Abstract Background Sulforaphane (SFN, an isothiocyanate phytochemical present predominantly in cruciferous vegetables such as brussels sprout and broccoli, is considered a promising chemo-preventive agent against cancer. In-vitro exposure to SFN appears to result in the induction of apoptosis and cell-cycle arrest in a variety of tumor types. However, the molecular mechanisms leading to the inhibition of cell cycle progression by SFN are poorly understood in epithelial ovarian cancer cells (EOC. The aim of this study is to understand the signaling mechanisms through which SFN influences the cell growth and proliferation in EOC. Results SFN at concentrations of 5 - 20 μM induced a dose-dependent suppression of growth in cell lines MDAH 2774 and SkOV-3 with an IC50 of ~8 μM after a 3 day exposure. Combination treatment with chemotherapeutic agent, paclitaxel, resulted in additive growth suppression. SFN at ~8 μM decreased growth by 40% and 20% on day 1 in MDAH 2774 and SkOV-3, respectively. Cells treated with cytotoxic concentrations of SFN have reduced cell migration and increased apoptotic cell death via an increase in Bak/Bcl-2 ratio and cleavage of procaspase-9 and poly (ADP-ribose-polymerase (PARP. Gene expression profile analysis of cell cycle regulated proteins demonstrated increased levels of tumor suppressor retinoblastoma protein (RB and decreased levels of E2F-1 transcription factor. SFN treatment resulted in G1 cell cycle arrest through down modulation of RB phosphorylation and by protecting the RB-E2F-1 complex. Conclusions SFN induces growth arrest and apoptosis in EOC cells. Inhibition of retinoblastoma (RB phosphorylation and reduction in levels of free E2F-1 appear to play an important role in EOC growth arrest.

  8. Cell surface clustering of Cadherin adhesion complex induced by antibody coated beads

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Cadherin receptors mediate cell-cell adhesion, signal transduction and assembly of cytoskeletons. How a single transmembrane molecule Cadherin can be involved in multiple functions through modulating its binding activities with many membrane adhesion molecules and cytoskeletal components is an unanswered question which can be elucidated by clues from bead experiments. Human lung cells expressing N-Cadherin were examined. After co-incubation with anti-N-Cadherin monoclonal antibody coated beads, cell surface clustering of N-Cadherin was induced. Immunofluorescent detection demonstrated that in addition to Cadherin, β-Catenin, α-Catenin, α-Actinin and Actin fluorescence also aggregated respectively at the membrane site of bead attachment. Myosin heavy chain (MHC), another major component of Actin cytoskeleton, did not aggregate at the membrane site of bead attachment. Adhesion unrelated protein Con A and polylysine conjugated beads did not induce the clustering of adhesion molecules. It is indicated that the Cadherin/Catenins/α-Actinin/Actin complex is formed at Cadherin mediated cell adherens junction; occupancy and cell surface clustering of Cadherin is crucial for the formation of Cadherin adhesion protein complexes.

  9. RYBP and Cbx7 Define Specific Biological Functions of Polycomb Complexes in Mouse Embryonic Stem Cells

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    Lluis Morey

    2013-01-01

    Full Text Available The Polycomb repressive complex 1 (PRC1 is required for decisions of stem cell fate. In mouse embryonic stem cells (ESCs, two major variations of PRC1 complex, defined by the mutually exclusive presence of Cbx7 or RYBP, have been identified. Here, we show that although the genomic localization of the Cbx7- and RYBP-containing PRC1 complexes overlaps in certain genes, it can also be mutually exclusive. At the molecular level, Cbx7 is necessary for recruitment of Ring1B to chromatin, whereas RYBP enhances the PRC1 enzymatic activity. Genes occupied by RYBP show lower levels of Ring1B and H2AK119ub and are consequently more highly transcribed than those bound by Cbx7. At the functional level, we show that genes occupied by RYBP are primarily involved in the regulation of metabolism and cell-cycle progression, whereas those bound by Cbx7 predominantly control early-lineage commitment of ESCs. Altogether, our results indicate that different PRC1 subtypes establish a complex pattern of gene regulation that regulates common and nonoverlapping aspects of ESC pluripotency and differentiation.

  10. A mononuclear zinc(II) complex with piroxicam: Crystal structure, DNA- and BSA-binding studies; in vitro cell cytotoxicity and molecular modeling of oxicam complexes

    Science.gov (United States)

    Jannesari, Zahra; Hadadzadeh, Hassan; Amirghofran, Zahra; Simpson, Jim; Khayamian, Taghi; Maleki, Batool

    2015-02-01

    A new mononuclear Zn(II) complex, trans-[Zn(Pir)2(DMSO)2], where Pir- is 4-hydroxy-2-methyl-N-2-pyridyl-2H-1,2-benzothiazine-3-carboxamide-1,1-dioxide (piroxicam), has been synthesized and characterized. The crystal structure of the complex was obtained by the single crystal X-ray diffraction technique. The interaction of the complex with DNA and BSA was investigated. The complex interacts with FS-DNA by two binding modes, viz., electrostatic and groove binding (major and minor). The microenvironment and the secondary structure of BSA are changed in the presence of the complex. The anticancer effects of the seven complexes of oxicam family were also determined on the human K562 cell lines and the results showed reasonable cytotoxicities. The interactions of the oxicam complexes with BSA and DNA were modeled by molecular docking and molecular dynamic simulation methods.

  11. Morphology and distribution of chandelier cell axon terminals in the mouse cerebral cortex and claustroamygdaloid complex.

    Science.gov (United States)

    Inda, M C; DeFelipe, J; Muñoz, A

    2009-01-01

    Chandelier cells represent a unique type of cortical gamma-aminobutityric acidergic interneuron whose axon terminals (Ch-terminals) only form synapses with the axon initial segments of some pyramidal cells. Here, we have used immunocytochemistry for the high-affinity plasma membrane transporter GAT-1 and the calcium-binding protein parvalbumin to analyze the morphology and distribution of Ch-terminals in the mouse cerebral cortex and claustroamygdaloid complex. In general, 2 types of Ch-terminals were distinguished on the basis of their size and the density of the axonal boutons that made up the terminal. Simple Ch-terminals were made up of 1 or 2 rows of labeled boutons, each row consisting of only 3-5 boutons. In contrast, complex Ch-terminals were tight cylinder-like structures made up of multiple rows of boutons. Simple Ch-terminals were detected throughout the cerebral cortex and claustroamygdaloid complex, the complex type was only occasionally found in certain regions, whereas in others they were very abundant. These results indicate that there are substantial differences in the morphology and distribution of Ch-terminals between different areas and layers of the mouse cerebral cortex. Furthermore, we suggest that the distribution of complex Ch-terminals may be related to the developmental origin of the different brain regions analyzed.

  12. Transforming growth factor beta 1 modulates extracellular matrix organization and cell-cell junctional complex formation during in vitro angiogenesis.

    Science.gov (United States)

    Merwin, J R; Anderson, J M; Kocher, O; Van Itallie, C M; Madri, J A

    1990-01-01

    Transforming growth factor-beta 1 (TGF-beta 1) is angiogenic in vivo. In two-dimensional (2-D) culture systems microvascular endothelial cell proliferation is inhibited up to 80% by TGF-beta 1; however, in three-dimensional (3-D) collagen gels TGF-beta 1 is found to have no effect on proliferation while eliciting the formation of calcium and magnesium dependent tube-like structures mimicking angiogenesis. DNA analyses performed on 3-D cell cultures reveal no significant difference in the amount of DNA or cell number in control versus TGF-beta 1 treated cultures. In 2-D cultures TGF-beta 1 is known to increase cellular fibronectin accumulation; however, in 3-D cultures no difference is seen between control and TGF-beta 1 treated cells as established by ELISA testing for type IV collagen, fibronectin, and laminin. In 3-D cultures there is increased synthesis and secretion of type V collagen in both control and TGF-beta 1 treated cultures over 2-D cultures. Even though an equal amount of type V collagen is seen in both 3-D conditions, there is a reorganization of the protein with concentration along an organizing basal lamina in TGF-beta 1 treated cultures. EM morphological analyses on 3-D cultures illustrate quiescent, control cells lacking cell contacts. In contrast, TGF-beta 1 treated cells show increased pseudopod formation, cell-cell contact, and organized basal lamina-like material closely apposed to the "abluminal" plasma membranes. TGF-beta 1 treated cells also appear to form junctional complexes between adjoining cells. Immunofluorescence using specific antibodies to the tight junction protein ZO-1 results in staining at apparent cell-cell junctions in the 3-D cultures. Northern blots of freshly isolated microvascular endothelium, 2-D and 3-D cultures, using cDNA and cRNA probes specific for the ZO-1 tight junction protein, reveal the presence of the 7.8 kb mRNA. Western blots of rat epididymal fat pad endothelial cells (RFC) monolayer lysates probed with

  13. Role for a Novel Usher Protein Complex in Hair Cell Synaptic Maturation

    Science.gov (United States)

    Zallocchi, Marisa; Meehan, Daniel T.; Delimont, Duane; Rutledge, Joseph; Gratton, Michael Anne; Flannery, John; Cosgrove, Dominic

    2012-01-01

    The molecular mechanisms underlying hair cell synaptic maturation are not well understood. Cadherin-23 (CDH23), protocadherin-15 (PCDH15) and the very large G-protein coupled receptor 1 (VLGR1) have been implicated in the development of cochlear hair cell stereocilia, while clarin-1 has been suggested to also play a role in synaptogenesis. Mutations in CDH23, PCDH15, VLGR1 and clarin-1 cause Usher syndrome, characterized by congenital deafness, vestibular dysfunction and retinitis pigmentosa. Here we show developmental expression of these Usher proteins in afferent spiral ganglion neurons and hair cell synapses. We identify a novel synaptic Usher complex comprised of clarin-1 and specific isoforms of CDH23, PCDH15 and VLGR1. To establish the in vivo relevance of this complex, we performed morphological and quantitative analysis of the neuronal fibers and their synapses in the Clrn1−/− mouse, which was generated by incomplete deletion of the gene. These mice showed a delay in neuronal/synaptic maturation by both immunostaining and electron microscopy. Analysis of the ribbon synapses in Ames waltzerav3J mice also suggests a delay in hair cell synaptogenesis. Collectively, these results show that, in addition to the well documented role for Usher proteins in stereocilia development, Usher protein complexes comprised of specific protein isoforms likely function in synaptic maturation as well. PMID:22363448

  14. Role for a novel Usher protein complex in hair cell synaptic maturation.

    Directory of Open Access Journals (Sweden)

    Marisa Zallocchi

    Full Text Available The molecular mechanisms underlying hair cell synaptic maturation are not well understood. Cadherin-23 (CDH23, protocadherin-15 (PCDH15 and the very large G-protein coupled receptor 1 (VLGR1 have been implicated in the development of cochlear hair cell stereocilia, while clarin-1 has been suggested to also play a role in synaptogenesis. Mutations in CDH23, PCDH15, VLGR1 and clarin-1 cause Usher syndrome, characterized by congenital deafness, vestibular dysfunction and retinitis pigmentosa. Here we show developmental expression of these Usher proteins in afferent spiral ganglion neurons and hair cell synapses. We identify a novel synaptic Usher complex comprised of clarin-1 and specific isoforms of CDH23, PCDH15 and VLGR1. To establish the in vivo relevance of this complex, we performed morphological and quantitative analysis of the neuronal fibers and their synapses in the Clrn1-/- mouse, which was generated by incomplete deletion of the gene. These mice showed a delay in neuronal/synaptic maturation by both immunostaining and electron microscopy. Analysis of the ribbon synapses in Ames waltzer(av3J mice also suggests a delay in hair cell synaptogenesis. Collectively, these results show that, in addition to the well documented role for Usher proteins in stereocilia development, Usher protein complexes comprised of specific protein isoforms likely function in synaptic maturation as well.

  15. Formation of wood secondary cell wall may involve two type cellulose synthase complexes in Populus.

    Science.gov (United States)

    Xi, Wang; Song, Dongliang; Sun, Jiayan; Shen, Junhui; Li, Laigeng

    2017-03-01

    Cellulose biosynthesis is mediated by cellulose synthases (CesAs), which constitute into rosette-like cellulose synthase complexe (CSC) on the plasma membrane. Two types of CSCs in Arabidopsis are believed to be involved in cellulose synthesis in the primary cell wall and secondary cell walls, respectively. In this work, we found that the two type CSCs participated cellulose biosynthesis in differentiating xylem cells undergoing secondary cell wall thickening in Populus. During the cell wall thickening process, expression of one type CSC genes increased while expression of the other type CSC genes decreased. Suppression of different type CSC genes both affected the wall-thickening and disrupted the multilaminar structure of the secondary cell walls. When CesA7A was suppressed, crystalline cellulose content was reduced, which, however, showed an increase when CesA3D was suppressed. The CesA suppression also affected cellulose digestibility of the wood cell walls. The results suggest that two type CSCs are involved in coordinating the cellulose biosynthesis in formation of the multilaminar structure in Populus wood secondary cell walls.

  16. Detection of circulating immune complexes by Raji cell assay: comparison of flow cytometric and radiometric methods

    International Nuclear Information System (INIS)

    Kingsmore, S.F.; Crockard, A.D.; Fay, A.C.; McNeill, T.A.; Roberts, S.D.; Thompson, J.M.

    1988-01-01

    Several flow cytometric methods for the measurement of circulating immune complexes (CIC) have recently become available. We report a Raji cell flow cytometric assay (FCMA) that uses aggregated human globulin (AHG) as primary calibrator. Technical advantages of the Raji cell flow cytometric assay are discussed, and its clinical usefulness is evaluated in a method comparison study with the widely used Raji cell immunoradiometric assay. FCMA is more precise and has greater analytic sensitivity for AHG. Diagnostic sensitivity by the flow cytometric method is superior in systemic lupus erythematosus (SLE), rheumatoid arthritis, and vasculitis patients: however, diagnostic specificity is similar for both assays, but the reference interval of FCMA is narrower. Significant correlations were found between CIC levels obtained with both methods in SLE, rheumatoid arthritis, and vasculitis patients and in longitudinal studies of two patients with cerebral SLE. The Raji cell FCMA is recommended for measurement of CIC levels to clinical laboratories with access to a flow cytometer

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

    Science.gov (United States)

    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.

  18. CRIM1 complexes with ß-catenin and cadherins, stabilizes cell-cell junctions and is critical for neural morphogenesis.

    Directory of Open Access Journals (Sweden)

    Virgilio G Ponferrada

    Full Text Available In multicellular organisms, morphogenesis is a highly coordinated process that requires dynamically regulated adhesion between cells. An excellent example of cellular morphogenesis is the formation of the neural tube from the flattened epithelium of the neural plate. Cysteine-rich motor neuron protein 1 (CRIM1 is a single-pass (type 1 transmembrane protein that is expressed in neural structures beginning at the neural plate stage. In the frog Xenopus laevis, loss of function studies using CRIM1 antisense morpholino oligonucleotides resulted in a failure of neural development. The CRIM1 knockdown phenotype was, in some cases, mild and resulted in perturbed neural fold morphogenesis. In severely affected embryos there was a dramatic failure of cell adhesion in the neural plate and complete absence of neural structures subsequently. Investigation of the mechanism of CRIM1 function revealed that it can form complexes with ß-catenin and cadherins, albeit indirectly, via the cytosolic domain. Consistent with this, CRIM1 knockdown resulted in diminished levels of cadherins and ß-catenin in junctional complexes in the neural plate. We conclude that CRIM1 is critical for cell-cell adhesion during neural development because it is required for the function of cadherin-dependent junctions.

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

    Directory of Open Access Journals (Sweden)

    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.

  20. Modification and uptake of a cisplatin carbonato complex by Jurkat cells.

    Science.gov (United States)

    Centerwall, Corey R; Tacka, Kirk A; Kerwood, Deborah J; Goodisman, Jerry; Toms, Bonnie B; Dubowy, Ronald L; Dabrowiak, James C

    2006-07-01

    The interactions of Jurkat cells with cisplatin, cis-[Pt(15NH3)2Cl2]1, are studied using 1H-15N heteronuclear single quantum coherence (HSQC) NMR and inductively coupled plasma mass spectrometry. We show that Jurkat cells in culture rapidly modify the monocarbonato complex cis-[Pt(15NH3)2(CO3)Cl]- (4), a cisplatin species that forms in culture media and probably also in blood. Analysis of the HSQC NMR peak intensity for 4 in the presence of different numbers of Jurkat cells reveals that each cell is capable of modifying 0.0028 pmol of 4 within approximately 0.6 h. The amounts of platinum taken up by the cell, weakly bound to the cell surface, remaining in the culture medium, and bound to genomic DNA were measured as functions of time of exposure to different concentrations of drug. The results show that most of the 4 that has been modified by the cells remains in the culture medium as a substance of molecular mass <3 kDa, which is HSQC NMR silent, and is not taken up by the cell. These results are consistent with a hitherto undocumented extracellular detoxification mechanism in which the cells rapidly modify 4, which is present in the culture medium, so it cannot bind to the cell. Because there is only a slow decrease in the amount of unmodified 4 remaining in the culture medium after 1 h, -1.1 +/- 0.4 microM h(-1), the cells subsequently lose their ability to modify 4. These observations have important implications for the mechanism of action of cisplatin.

  1. Polycomb repressive complex 2 (PRC2 restricts hematopoietic stem cell activity.

    Directory of Open Access Journals (Sweden)

    Ian J Majewski

    2008-04-01

    Full Text Available Polycomb group proteins are transcriptional repressors that play a central role in the establishment and maintenance of gene expression patterns during development. Using mice with an N-ethyl-N-nitrosourea (ENU-induced mutation in Suppressor of Zeste 12 (Suz12, a core component of Polycomb Repressive Complex 2 (PRC2, we show here that loss of Suz12 function enhances hematopoietic stem cell (HSC activity. In addition to these effects on a wild-type genetic background, mutations in Suz12 are sufficient to ameliorate the stem cell defect and thrombocytopenia present in mice that lack the thrombopoietin receptor (c-Mpl. To investigate the molecular targets of the PRC2 complex in the HSC compartment, we examined changes in global patterns of gene expression in cells deficient in Suz12. We identified a distinct set of genes that are regulated by Suz12 in hematopoietic cells, including eight genes that appear to be highly responsive to PRC2 function within this compartment. These data suggest that PRC2 is required to maintain a specific gene expression pattern in hematopoiesis that is indispensable to normal stem cell function.

  2. The Allelic Landscape of Human Blood Cell Trait Variation and Links to Common Complex Disease.

    Science.gov (United States)

    Astle, William J; Elding, Heather; Jiang, Tao; Allen, Dave; Ruklisa, Dace; Mann, Alice L; Mead, Daniel; Bouman, Heleen; Riveros-Mckay, Fernando; Kostadima, Myrto A; Lambourne, John J; Sivapalaratnam, Suthesh; Downes, Kate; Kundu, Kousik; Bomba, Lorenzo; Berentsen, Kim; Bradley, John R; Daugherty, Louise C; Delaneau, Olivier; Freson, Kathleen; Garner, Stephen F; Grassi, Luigi; Guerrero, Jose; Haimel, Matthias; Janssen-Megens, Eva M; Kaan, Anita; Kamat, Mihir; Kim, Bowon; Mandoli, Amit; Marchini, Jonathan; Martens, Joost H A; Meacham, Stuart; Megy, Karyn; O'Connell, Jared; Petersen, Romina; Sharifi, Nilofar; Sheard, Simon M; Staley, James R; Tuna, Salih; van der Ent, Martijn; Walter, Klaudia; Wang, Shuang-Yin; Wheeler, Eleanor; Wilder, Steven P; Iotchkova, Valentina; Moore, Carmel; Sambrook, Jennifer; Stunnenberg, Hendrik G; Di Angelantonio, Emanuele; Kaptoge, Stephen; Kuijpers, Taco W; Carrillo-de-Santa-Pau, Enrique; Juan, David; Rico, Daniel; Valencia, Alfonso; Chen, Lu; Ge, Bing; Vasquez, Louella; Kwan, Tony; Garrido-Martín, Diego; Watt, Stephen; Yang, Ying; Guigo, Roderic; Beck, Stephan; Paul, Dirk S; Pastinen, Tomi; Bujold, David; Bourque, Guillaume; Frontini, Mattia; Danesh, John; Roberts, David J; Ouwehand, Willem H; Butterworth, Adam S; Soranzo, Nicole

    2016-11-17

    Many common variants have been associated with hematological traits, but identification of causal genes and pathways has proven challenging. We performed a genome-wide association analysis in the UK Biobank and INTERVAL studies, testing 29.5 million genetic variants for association with 36 red cell, white cell, and platelet properties in 173,480 European-ancestry participants. This effort yielded hundreds of low frequency (<5%) and rare (<1%) variants with a strong impact on blood cell phenotypes. Our data highlight general properties of the allelic architecture of complex traits, including the proportion of the heritable component of each blood trait explained by the polygenic signal across different genome regulatory domains. Finally, through Mendelian randomization, we provide evidence of shared genetic pathways linking blood cell indices with complex pathologies, including autoimmune diseases, schizophrenia, and coronary heart disease and evidence suggesting previously reported population associations between blood cell indices and cardiovascular disease may be non-causal. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. The fabrication and cell culture of three-dimensional rolled scaffolds with complex micro-architectures

    International Nuclear Information System (INIS)

    Liu Yaxiong; Li Xiao; Qu Xiaoli; Zhu Lin; He Jiankang; Zhao Qian; Wu Wanquan; Li Dichen

    2012-01-01

    Cell cultures for tissue engineering are traditionally prepared on two-dimensional or three-dimensional scaffolds with simple pores; however, this limits mass transportation, which is necessary for cell viability and function. In this paper, an innovative method is proposed for fabricating porous scaffolds with designed complex micro-architectures. Channels devised by computer-aided design were used to simulate features of blood vessels in native rat liver. Rapid prototyping and microreplication were used to produce a negative polydimethylsiloxane mold, and then a planar porous scaffold with predefined microchannel parameters was obtained by freeze-drying a silk fibroin/gelatin solution of an optimized concentration. After seeding with rat primary hepatocytes, the planar scaffold was rolled up to build spatial channels. By reconstructing the three-dimensional channel model in the scaffold in the form of micro-computed topography data and observing the cross-sections of the scroll, we confirmed that the bent channels were still interconnected, with restricted deviations. A comparison of the primary hepatocyte culture in the scaffolds with and without the devised channels proved that our design influenced cell organization and improved cell survival and proliferation. This method can be used for the construction of complex tissues for implantation and for culturing cells in vitro for biological tests and observations.

  4. Ubiquitination regulates MHC class II-peptide complex retention and degradation in dendritic cells

    OpenAIRE

    Walseng, Even; Furuta, Kazuyuki; Bosch, Berta; Weih, Karis A.; Matsuki, Yohei; Bakke, Oddmund; Ishido, Satoshi; Roche, Paul A.

    2010-01-01

    The expression and turnover of MHC class II-peptide complexes (pMHC-II) on the surface of dendritic cells (DCs) is essential for their ability to activate CD4 T cells efficiently. The half-life of surface pMHC-II is significantly greater in activated (mature) DCs than in resting (immature) DCs, but the molecular mechanism leading to this difference remains unknown. We now show that ubiquitination of pMHC-II by the E3 ubiquitin ligase membrane-associated RING-CH 1 (March-I) regulates surface e...

  5. Engineering complex tissue-like microgel arrays for evaluating stem cell differentiation

    DEFF Research Database (Denmark)

    Guermani, Enrico; Shaki, Hossein; Mohanty, Soumyaranjan

    2016-01-01

    Development of tissue engineering scaffolds with native-like biology and microarchitectures is a prerequisite for stem cell mediated generation of off-the-shelf-tissues. So far, the field of tissue engineering has not full-filled its grand potential of engineering such combinatorial scaffolds...... for engineering functional tissues. This is primarily due to the many challenges associated with finding the right microarchitectures and ECM compositions for optimal tissue regeneration. Here, we have developed a new microgel array to address this grand challenge through robotic printing of complex stem cell...... platform will be used for high-throughput identification of combinatorial and native-like scaffolds for tissue engineering of functional organs....

  6. Electron Transfer Mediators for Photoelectrochemical Cells Based on Cu(I Metal Complexes

    Directory of Open Access Journals (Sweden)

    Michele Brugnati

    2007-01-01

    Full Text Available The preparation and the photoelectrochemical characterization of a series of bipyridine and pyridyl-quinoline Cu(I complexes, used as electron transfer mediators in regenerative photoelectrochemical cells, are reported. The best performing mediators produced maximum IPCEs of the order of 35–40%. The J-V curves recorded under monochromatic light showed that the selected Cu(I/(II couples generated higher Vocs and fill factors compared to an equivalent I-/I3- cell, due to a decreased dark current.

  7. A Recombinant Antibody with the Antigen-Specific, Major Histocompatibility Complex-Restricted Specificity of T Cells

    Science.gov (United States)

    Andersen, Peter S.; Stryhn, Anette; Hansen, Bjarke E.; Fugger, Lars; Engberg, Jan; Buus, Soren

    1996-03-01

    Specific recognition of peptide/major histocompatibility complex (MHC) molecule complexes by the T-cell receptor is a key reaction in the specific immune response. Antibodies against peptide/MHC complexes would therefore be valuable tools in studying MHC function and T-cell recognition and might lead to novel approaches in immunotherapy. However, it has proven difficult to generate antibodies with the specificity of T cells by conventional hybridoma techniques. Here we report that the phage display technology is a feasible alternative to generate antibodies recognizing specific, predetermined peptide/MHC complexes.

  8. Zika Virus Escapes NK Cell Detection by Upregulating Major Histocompatibility Complex Class I Molecules.

    Science.gov (United States)

    Glasner, Ariella; Oiknine-Djian, Esther; Weisblum, Yiska; Diab, Mohammad; Panet, Amos; Wolf, Dana G; Mandelboim, Ofer

    2017-11-15

    NK cells are innate lymphocytes that participate in many immune processes encompassing cancer, bacterial and fungal infection, autoimmunity, and even pregnancy and that specialize in antiviral defense. NK cells express inhibitory and activating receptors and kill their targets when activating signals overpower inhibitory signals. The NK cell inhibitory receptors include a uniquely diverse array of proteins named killer cell immunoglobulin-like receptors (KIRs), the CD94 family, and the leukocyte immunoglobulin-like receptor (LIR) family. The NK cell inhibitory receptors recognize mostly major histocompatibility complex (MHC) class I (MHC-I) proteins. Zika virus has recently emerged as a major threat due to its association with birth defects and its pandemic potential. How Zika virus interacts with the immune system, and especially with NK cells, is unclear. Here we show that Zika virus infection is barely sensed by NK cells, since little or no increase in the expression of activating NK cell ligands was observed following Zika infection. In contrast, we demonstrate that Zika virus infection leads to the upregulation of MHC class I proteins and consequently to the inhibition of NK cell killing. Mechanistically, we show that MHC class I proteins are upregulated via the RIGI-IRF3 pathway and that this upregulation is mediated via beta interferon (IFN-β). Potentially, countering MHC class I upregulation during Zika virus infection could be used as a prophylactic treatment against Zika virus. IMPORTANCE NK cells are innate lymphocytes that recognize and eliminate various pathogens and are known mostly for their role in controlling viral infections. NK cells express inhibitory and activating receptors, and they kill or spare their targets based on the integration of inhibitory and activating signals. Zika virus has recently emerged as a major threat to humans due to its pandemic potential and its association with birth defects. The role of NK cells in Zika virus

  9. Dye-sensitized solar cells and complexes between pyridines and iodines

    DEFF Research Database (Denmark)

    Hansen, Poul Erik; Phuong, Nguyen Tuyet; Krake, Jacob

    2012-01-01

    Interactions between triiodide (I3–) and 4-tert-butylpyridine (4TBP) as postulated in dye-sensitized solar cells (DSC) are investigated by means of 13C NMR and IR spectroscopy supported by DFT calculations. The charge transfer (CT) complex 4TBP∙I2 and potential salts such as (4TBP)2I+, I3– were...... synthesized and characterized by IR and 13C NMR spectroscopy. However, mixing (butyl)4N+, I3– and 4TBP at concentrations comparable to those of the DSC solar cell did not lead to any reaction. Neither CT complexes nor cationic species like (4TBP)2I+ were observed, judging from the 13C NMR spectroscopic...

  10. Odontoblasts: Specialized hard-tissue-forming cells in the dentin-pulp complex.

    Science.gov (United States)

    Kawashima, Nobuyuki; Okiji, Takashi

    2016-07-01

    Odontoblasts are specialized cells that produce dentin and exhibit unique morphological characteristics; i.e., they extend cytoplasmic processes into dentinal tubules. While osteoblasts, which are typical hard-tissue-forming cells, are generated from mesenchymal stem cells during normal and pathological bone metabolism, the induction of odontoblasts only occurs once during tooth development, and odontoblasts survive throughout the lives of healthy teeth. During the differentiation of odontoblasts, signaling molecules from the inner enamel epithelium are considered necessary for the differentiation of odontoblast precursors, i.e., peripheral dental papilla cells. If odontoblasts are destroyed by severe external stimuli, such as deep caries, the differentiation of dental pulp stem cells into odontoblast-like cells is induced. Various bioactive molecules, such as non-collagenous proteins, might be involved in this process, although the precise mechanisms responsible for odontoblast differentiation have not been fully elucidated. Recently, our knowledge about the other functional activities of odontoblasts (apart from dentin formation) has increased. For example, it has been suggested that odontoblasts might act as nociceptive receptors, and surveillance cells that detect the invasion of exogenous pathogens. The regeneration of the dentin-pulp complex has recently gained much attention as a promising future treatment modality that could increase the longevity of pulpless teeth. Finally, congenital dentin anomalies, which are concerned with the disturbance of odontoblast functions, are summarized. © 2016 Japanese Teratology Society.

  11. Cytotoxicity of Diimine Palladium (II) Complexes of Alkyldithiocarbamate Derivatives on Human Lung, Ovary and Liver Cells.

    Science.gov (United States)

    Aryanpour, Narges; Mansouri-Torshizi, Hassan; Nakhjavan, Maryam; H Shirazi, Farshad

    2012-01-01

    Three new Complexes of formula [pd(bpy)(R-NH-CSS)] Cl (where bpy is 2/2'- bipyridine, and R-NH-CSS is butylamine, hexylamine- and octyamine-dithiocabamate anion) have been synthesized by University of Sistan and Blachostan. These complexes have been characterized by spectroscopic methods such as ultraviolet-visible, infrared and (1)H-NMR as well as conductivity measurements and chemical analysis. In these complexes, each of the dithiocarbamate ligands coordinates to Pd (II) center as bidentate with two sulfur atoms. We have found a 1:1 electrolyte in water conductivity test for the above mentioned compounds. To measure the biologic activity and potential anticancer efficacy of these compounds, they have been compared with cisplatin and its palladium analogue of [Pd (NH3)2 Cl2] on three different cell lines of human hepatocarcinoma HepG2, human ovarian carcinoma OV2008, and human lung adenocarcinoma A549. Clonogenic assay has shown LD50s in the range of 0.131±0.025 to 0.934 ± 0.194 for these compounds on above cell lines. In comparison, cisplatin has shown LD50s of 0.838 ± 0.074, 2.196 ± 0.220, and 2.799 ± 0.733 on OV2008, HepG2 and A549 cell lines, respectively. As a conclusion, above three new complexes have shown higher cytotoxicities compared to cisplatin on three different human cell lines. Based on biological tests, these compounds may potentially be considered as good anticancer candidates for further pharmacological studies.

  12. An Exocyst Complex Functions in Plant Cell Growth in Arabidopsis and Tobacco

    Czech Academy of Sciences Publication Activity Database

    Hála, Michal; Cole, R.A.; Synek, Lukáš; Drdová, Edita; Pečenková, Tamara; Nordheim, A.; Lamkemeyer, T.; Madlung, J.; Hochholdinger, F.; Fowler, J.E.; Žárský, Viktor

    2008-01-01

    Roč. 20, č. 5 (2008), s. 1330-1345 ISSN 1040-4651 R&D Projects: GA MŠk ME 841; GA MŠk(CZ) LC06034; GA AV ČR IAA6038410 Institutional research plan: CEZ:AV0Z50380511 Keywords : EXOCYST * PROTEIN COMPLEX * CELL POLARITY * MORPHOGENESIS * GTPASES Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 9.296, year: 2008

  13. Basal cell carcinoma of the nipple-areola complex: a case report.

    Science.gov (United States)

    Huang, Ching-Wen; Pan, Ching-Kuen; Shih, Teng-Fu; Tsai, Cheng-Chien; Juan, Chung-Chou; Ker, Chen-Guo

    2005-10-01

    Basal cell carcinoma (BCC) of the nipple-areola complex is very rare. Only 24 cases were reported in the literature and 17 (70.8%) of these cases arose in men. Most of the cases were treated with simple excision. We report on a case of BCC of the nipple-areola complex in a 46-year-old woman, treated with partial mastectomy. Metastasis to the axillary lymph nodes had been noted in 3 (12.5%) of the 24 reported cases of BCC of the nipple-areola complex. Thus, we applied the concept of the sentinel lymph node to detect possible metastases of axillary lymph nodes, letting us avoid the unnecessary axillary lymph node dissection and possible future morbidity.

  14. Isolation and characterization of antigen-Ia complexes involved in T cell recognition

    DEFF Research Database (Denmark)

    Buus, S; Sette, A; Colon, S M

    1986-01-01

    Using equilibrium dialysis, it has been previously demonstrated that immunogenic peptides bind specifically to the Ia molecules serving as restriction elements in the immune response to these antigens. Using gel filtration to study the formation of ovalbumin (OVA) peptide-I-Ad complexes, it is he......Using equilibrium dialysis, it has been previously demonstrated that immunogenic peptides bind specifically to the Ia molecules serving as restriction elements in the immune response to these antigens. Using gel filtration to study the formation of ovalbumin (OVA) peptide-I-Ad complexes...... with glutaraldehyde revealed that the ovalbumin peptide was cross-linked solely to the alpha chain of I-Ad. Planar membranes containing I-Ad-OVA complexes stimulated a T cell response with 2 X 10(4) less antigen than required when uncomplexed antigen was used, thus demonstrating the biologic importance...

  15. Translational Capacity of a Cell Is Determined during Transcription Elongation via the Ccr4-Not Complex

    Directory of Open Access Journals (Sweden)

    Ishaan Gupta

    2016-05-01

    Full Text Available The current understanding of gene expression considers transcription and translation to be independent processes. Challenging this notion, we found that translation efficiency is determined during transcription elongation through the imprinting of mRNAs with Not1, the central scaffold of the Ccr4-Not complex. We determined that another subunit of the complex, Not5, defines Not1 binding to specific mRNAs, particularly those produced from ribosomal protein genes. This imprinting mechanism specifically regulates ribosomal protein gene expression, which in turn determines the translational capacity of cells. We validate our model by SILAC and polysome profiling experiments. As a proof of concept, we demonstrate that enhanced translation compensates for transcriptional elongation stress. Taken together, our data indicate that in addition to defining mRNA stability, components of the Ccr4-Not imprinting complex regulate RNA translatability, thus ensuring global gene expression homeostasis.

  16. Zebrafish model of tuberous sclerosis complex reveals cell-autonomous and non-cell-autonomous functions of mutant tuberin

    Directory of Open Access Journals (Sweden)

    Seok-Hyung Kim

    2011-03-01

    Tuberous sclerosis complex (TSC is an autosomal dominant disease caused by mutations in either the TSC1 (encodes hamartin or TSC2 (encodes tuberin genes. Patients with TSC have hamartomas in various organs throughout the whole body, most notably in the brain, skin, eye, heart, kidney and lung. To study the development of hamartomas, we generated a zebrafish model of TSC featuring a nonsense mutation (vu242 in the tsc2 gene. This tsc2vu242 allele encodes a truncated Tuberin protein lacking the GAP domain, which is required for inhibition of Rheb and of the TOR kinase within TORC1. We show that tsc2vu242 is a recessive larval-lethal mutation that causes increased cell size in the brain and liver. Greatly elevated TORC1 signaling is observed in tsc2vu242/vu242 homozygous zebrafish, and is moderately increased in tsc2vu242/+ heterozygotes. Forebrain neurons are poorly organized in tsc2vu242/vu242 homozygous mutants, which have extensive gray and white matter disorganization and ectopically positioned cells. Genetic mosaic analyses demonstrate that tsc2 limits TORC1 signaling in a cell-autonomous manner. However, in chimeric animals, tsc2vu242/vu242 mutant cells also mislocalize wild-type host cells in the forebrain in a non-cell-autonomous manner. These results demonstrate a highly conserved role of tsc2 in zebrafish and establish a new animal model for studies of TSC. The finding of a non-cell-autonomous function of mutant cells might help explain the formation of brain hamartomas and cortical malformations in human TSC.

  17. Lipofection indirectly increases expression of endogenous major histocompatibility complex class I molecules on tumor cells.

    Science.gov (United States)

    Fox, B A; Drury, M; Hu, H M; Cao, Z; Huntzicker, E G; Qie, W; Urba, W J

    1998-01-01

    Direct intratumoral injection of a lipid/DNA complex encoding an allogeneic major histocompatibility complex (MHC) class I molecule leads to regression of both an immunogenic murine tumor and also melanoma lesions in some patients. We have sought to understand the mechanism(s) for this augmentation of antitumor activity. While optimizing parameters for in vitro gene transfer into the D5 subclone of B16BL6, it was noted that lipofected tumors not only expressed the new alloantigen but also exhibited increased expression of endogenous MHC class I, both H-2 Kb and H-2 Db. This increase in expression was not restricted to the small percentage of cells that expressed the transfected gene, but appeared to affect the majority of cells in culture. Class I expression was not increased by lipopolysaccharide, DNA alone, lipid, or lipid/lipopolysaccharide mixtures. Enhanced class I expression required a DNA/lipid complex and was greatest when parameters optimized for gene transfer of the alloantigen were used. All DNA plasmids tested had this effect, including one plasmid whose DNA was not transcribed because it lacked an expression cassette. Because of the critical role that MHC class I antigens play in immune recognition, we propose that lipid complex-mediated gene transfer may provide immunological advantages beyond those that are attributable to expression of the specific gene transferred.

  18. Adenovirus type 5 DNA-protein complexes from formaldehyde cross-linked cells early after infection

    International Nuclear Information System (INIS)

    Spector, David J.; Johnson, Jeffrey S.; Baird, Nicholas L.; Engel, Daniel A.

    2003-01-01

    We report here the properties of viral DNA-protein complexes that purify with cellular chromatin following formaldehyde cross-linking of intact cells early after infection. The cross-linked viral DNA fractionated into shear-sensitive (S) and shear- resistant (R) components that were separable by sedimentation, which allowed independent characterization. The R component had the density and sedimentation properties expected for DNA-protein complexes and contained intact viral DNA. It accounted for about 50% of the viral DNA recovered at 1.5 h after infection but less than 20% by 4.5 h. The proportion of R component was independent of multiplicity of infection, even at less than one particle per cell. Viral hexon and protein VII, but not protein VI, were detected in the fractions containing the R component. These properties are consistent with those of partially uncoated virions associated with the nuclear envelope. A substantial proportion of the S component viral DNA had the same density as cellular chromatin. Protein VII was the most abundant viral protein present in gradient fractions that contained the S component. Complexes containing USF transcription factor cross-linked to the adenovirus major late promoter were detected by viral chromatin immunoprecipitation of the fractions containing S component. The S component probably contained uncoated nuclear viral DNA that assembles into early viral transcription complexes

  19. Monitoring ligand-dependent assembly of receptor ternary complexes in live cells by BRETFect.

    Science.gov (United States)

    Cotnoir-White, David; El Ezzy, Mohamed; Boulay, Pierre-Luc; Rozendaal, Marieke; Bouvier, Michel; Gagnon, Etienne; Mader, Sylvie

    2018-03-13

    There is currently an unmet need for versatile techniques to monitor the assembly and dynamics of ternary complexes in live cells. Here we describe bioluminescence resonance energy transfer with fluorescence enhancement by combined transfer (BRETFect), a high-throughput technique that enables robust spectrometric detection of ternary protein complexes based on increased energy transfer from a luciferase to a fluorescent acceptor in the presence of a fluorescent intermediate. Its unique donor-intermediate-acceptor relay system is designed so that the acceptor can receive energy either directly from the donor or indirectly via the intermediate in a combined transfer, taking advantage of the entire luciferase emission spectrum. BRETFect was used to study the ligand-dependent cofactor interaction properties of the estrogen receptors ERα and ERβ, which form homo- or heterodimers whose distinctive regulatory properties are difficult to dissect using traditional methods. BRETFect uncovered the relative capacities of hetero- vs. homodimers to recruit receptor-specific cofactors and regulatory proteins, and to interact with common cofactors in the presence of receptor-specific ligands. BRETFect was also used to follow the assembly of ternary complexes between the V2R vasopressin receptor and two different intracellular effectors, illustrating its use for dissection of ternary protein-protein interactions engaged by G protein-coupled receptors. Our results indicate that BRETFect represents a powerful and versatile technique to monitor the dynamics of ternary interactions within multimeric complexes in live cells.

  20. Tracking the Oxygen Status in the Cell Nucleus with a Hoechst-Tagged Phosphorescent Ruthenium Complex.

    Science.gov (United States)

    Hara, Daiki; Umehara, Yui; Son, Aoi; Asahi, Wataru; Misu, Sotaro; Kurihara, Ryohsuke; Kondo, Teruyuki; Tanabe, Kazuhito

    2018-05-04

    Molecular oxygen in living cells is distributed and consumed inhomogeneously, depending on the activity of each organelle. Therefore, tractable methods that can be used to monitor the oxygen status in each organelle are needed to understand cellular function. Here we report the design of a new oxygen-sensing probe for use in the cell nucleus. We prepared "Ru-Hoechsts", each consisting of a phosphorescent ruthenium complex linked to a Hoechst 33258 moiety, and characterized their properties as oxygen sensors. The Hoechst unit shows strong DNA-binding properties in the nucleus, and the ruthenium complex shows oxygen-dependent phosphorescence. Thus, Ru-Hoechsts accumulated in the cell nucleus and showed oxygen-dependent signals that could be monitored. Of the Ru-Hoechsts prepared in this study, Ru-Hoechst b, in which the ruthenium complex and the Hoechst unit were linked through a hexyl chain, showed the most suitable properties for monitoring the oxygen status. Ru-Hoechsts are probes with high potential for visualizing oxygen fluctuations in the nucleus. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Alkynyl gold(I) complex triggers necroptosis via ROS generation in colorectal carcinoma cells.

    Science.gov (United States)

    Mármol, Inés; Virumbrales-Muñoz, María; Quero, Javier; Sánchez-de-Diego, Cristina; Fernández, Luis; Ochoa, Ignacio; Cerrada, Elena; Yoldi, Mª Jesús Rodríguez

    2017-11-01

    Given the rise of apoptosis-resistant tumors, there exist a growing interest in developing new drugs capable of inducing different types of cell death to reduce colorectal cancer-related death rates. As apoptosis and necroptosis do not share cellular machinery, necroptosis induction may have a great therapeutic potential on those apoptosis-resistant cancers, despite the inflammatory effects associated with it. We have synthesized an alkynyl gold(I) complex [Au(CC-2-NC 5 H 4 )(PTA)] whose anticancer effect was tested on the colorectal adenocarcinoma Caco-2 cell line. With regard to its mechanism of action, this gold complex enters the mitochondria and disrupts its normal function, leading to an increase in ROS production, which triggers necroptosis. Necroptosis induction has been found dependent of TNF-α (Tumor necrosisfactor α) and TNFR1(Tumor necrosisfactor receptor 1) binding, RIP1(Receptor-Interacting Protein 1) activation and NF-κB (Nuclear Factor Kappa-Light-Chain-Enhancer of Activated B Cells) signaling. Moreover, the antitumor potential of [Au(CC-2-NC 5 H 4 )(PTA)] has also been confirmed on the 3D cancer model spheroid. Overall, the obtained data show firstly that gold complexes might have the ability of inducing necroptosis, and secondarily that our compound [Au(CC-2-NC 5 H 4 )(PTA)] is an interesting alternative to current chemotherapy drugs in cases of apoptosis resistance. Copyright © 2017. Published by Elsevier Inc.

  2. Repression of germline RNAi pathways in somatic cells by retinoblastoma pathway chromatin complexes.

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    Xiaoyun Wu

    Full Text Available The retinoblastoma (Rb tumor suppressor acts with a number of chromatin cofactors in a wide range of species to suppress cell proliferation. The Caenorhabditis elegans retinoblastoma gene and many of these cofactors, called synMuv B genes, were identified in genetic screens for cell lineage defects caused by growth factor misexpression. Mutations in many synMuv B genes, including lin-35/Rb, also cause somatic misexpression of the germline RNA processing P granules and enhanced RNAi. We show here that multiple small RNA components, including a set of germline-specific Argonaute genes, are misexpressed in the soma of many synMuv B mutant animals, revealing one node for enhanced RNAi. Distinct classes of synMuv B mutants differ in the subcellular architecture of their misexpressed P granules, their profile of misexpressed small RNA and P granule genes, as well as their enhancement of RNAi and the related silencing of transgenes. These differences define three classes of synMuv B genes, representing three chromatin complexes: a LIN-35/Rb-containing DRM core complex, a SUMO-recruited Mec complex, and a synMuv B heterochromatin complex, suggesting that intersecting chromatin pathways regulate the repression of small RNA and P granule genes in the soma and the potency of RNAi. Consistent with this, the DRM complex and the synMuv B heterochromatin complex were genetically additive and displayed distinct antagonistic interactions with the MES-4 histone methyltransferase and the MRG-1 chromodomain protein, two germline chromatin regulators required for the synMuv phenotype and the somatic misexpression of P granule components. Thus intersecting synMuv B chromatin pathways conspire with synMuv B suppressor chromatin factors to regulate the expression of small RNA pathway genes, which enables heightened RNAi response. Regulation of small RNA pathway genes by human retinoblastoma may also underlie its role as a tumor suppressor gene.

  3. Papillae formation on trichome cell walls requires the function of the mediator complex subunit Med25.

    Science.gov (United States)

    Fornero, Christy; Suo, Bangxia; Zahde, Mais; Juveland, Katelyn; Kirik, Viktor

    2017-11-01

    Glassy Hair 1 (GLH1) gene that promotes papillae formation on trichome cell walls was identified as a subunit of the transcriptional mediator complex MED25. The MED25 gene is shown to be expressed in trichomes. The expression of the trichome development marker genes GLABRA2 (GL2) and Ethylene Receptor2 (ETR2) is not affected in the glh1 mutant. Presented data suggest that Arabidopsis MED25 mediator component is likely involved in the transcription of genes promoting papillae deposition in trichomes. The plant cell wall plays an important role in communication, defense, organization and support. The importance of each of these functions varies by cell type. Specialized cells, such as Arabidopsis trichomes, exhibit distinct cell wall characteristics including papillae. To better understand the molecular processes important for papillae deposition on the cell wall surface, we identified the GLASSY HAIR 1 (GLH1) gene, which is necessary for papillae formation. We found that a splice-site mutation in the component of the transcriptional mediator complex MED25 gene is responsible for the near papillae-less phenotype of the glh1 mutant. The MED25 gene is expressed in trichomes. Reporters for trichome developmental marker genes GLABRA2 (GL2) and Ethylene Receptor2 (ETR2) were not affected in the glh1 mutant. Collectively, the presented results show that MED25 is necessary for papillae formation on the cell wall surface of leaf trichomes and suggest that the Arabidopsis MED25 mediator component is likely involved in the transcription of a subset of genes that promote papillae deposition in trichomes.

  4. A Cbx8-containing polycomb complex facilitates the transition to gene activation during ES cell differentiation.

    Directory of Open Access Journals (Sweden)

    Catherine Creppe

    2014-12-01

    Full Text Available Polycomb proteins play an essential role in maintaining the repression of developmental genes in self-renewing embryonic stem cells. The exact mechanism allowing the derepression of polycomb target genes during cell differentiation remains unclear. Our project aimed to identify Cbx8 binding sites in differentiating mouse embryonic stem cells. Therefore, we used a genome-wide chromatin immunoprecipitation of endogenous Cbx8 coupled to direct massive parallel sequencing (ChIP-Seq. Our analysis identified 171 high confidence peaks. By crossing our data with previously published microarray analysis, we show that several differentiation genes transiently recruit Cbx8 during their early activation. Depletion of Cbx8 partially impairs the transcriptional activation of these genes. Both interaction analysis, as well as chromatin immunoprecipitation experiments support the idea that activating Cbx8 acts in the context of an intact PRC1 complex. Prolonged gene activation results in eviction of PRC1 despite persisting H3K27me3 and H2A ubiquitination. The composition of PRC1 is highly modular and changes when embryonic stem cells commit to differentiation. We further demonstrate that the exchange of Cbx7 for Cbx8 is required for the effective activation of differentiation genes. Taken together, our results establish a function for a Cbx8-containing complex in facilitating the transition from a Polycomb-repressed chromatin state to an active state. As this affects several key regulatory differentiation genes this mechanism is likely to contribute to the robust execution of differentiation programs.

  5. Cell–cell signaling drives the evolution of complex traits: introduction—lung evo-devo

    Science.gov (United States)

    Torday, John S.; Rehan, V. K.

    2009-01-01

    Physiology integrates biology with the environment through cell–cell interactions at multiple levels. The evolution of the respiratory system has been “deconvoluted” (Torday and Rehan in Am J Respir Cell Mol Biol 31:8–12, 2004) through Gene Regulatory Networks (GRNs) applied to cell–cell communication for all aspects of lung biology development, homeostasis, regeneration, and aging. Using this approach, we have predicted the phenotypic consequences of failed signaling for lung development, homeostasis, and regeneration based on evolutionary principles. This cell–cell communication model predicts other aspects of vertebrate physiology as adaptational responses. For example, the oxygen-induced differentiation of alveolar myocytes into alveolar adipocytes was critical for the evolution of the lung in land dwelling animals adapting to fluctuating Phanarezoic oxygen levels over the past 500 million years. Adipocytes prevent lung injury due to oxygen radicals and facilitate the rise of endothermy. In addition, they produce the class I cytokine leptin, which augments pulmonary surfactant activity and alveolar surface area, increasing selection pressure for both respiratory oxygenation and metabolic demand initially constrained by high-systemic vascular pressure, but subsequently compensated by the evolution of the adrenomedullary beta-adrenergic receptor mechanism. Conserted positive selection for the lung and adrenals created further selection pressure for the heart, which becomes progressively more complex phylogenetically in tandem with the lung. Developmentally, increasing heart complexity and size impinges precociously on the gut mesoderm to induce the liver. That evolutionary-developmental interaction is significant because the liver provides regulated sources of glucose and glycogen to the evolving physiologic system, which is necessary for the evolution of the neocortex. Evolution of neocortical control furthers integration of physiologic systems. Such

  6. Enhancing the Photovoltaic Performance of Perovskite Solar Cells with a Down-Conversion Eu-Complex.

    Science.gov (United States)

    Jiang, Ling; Chen, Wangchao; Zheng, Jiawei; Zhu, Liangzheng; Mo, Li'e; Li, Zhaoqian; Hu, Linhua; Hayat, Tasawar; Alsaedi, Ahmed; Zhang, Changneng; Dai, Songyuan

    2017-08-16

    Organometal halide perovskite solar cells (PSCs) have shown high photovoltaic performance but poor utilization of ultraviolet (UV) irradiation. Lanthanide complexes have a wide absorption range in the UV region and they can down-convert the absorbed UV light into visible light, which provides a possibility for PSCs to utilize UV light for higher photocurrent, efficiency, and stability. In this study, we use a transparent luminescent down-converting layer (LDL) of Eu-4,7-diphenyl-1,10-phenanthroline (Eu-complex) to improve the light utilization efficiency of PSCs. Compared with the uncoated PSC, the PSC coated with Eu-complex LDL on the reverse of the fluorine-doped tin oxide glass displayed an enhancement of 11.8% in short-circuit current density (J sc ) and 15.3% in efficiency due to the Eu-complex LDL re-emitting UV light (300-380 nm) in the visible range. It is indicated that the Eu-complex LDL plays the role of enhancing the power conversion efficiency as well as reducing UV degradation for PSCs.

  7. A ghost-cell immersed boundary method for flow in complex geometry

    International Nuclear Information System (INIS)

    Tseng, Y.-H.; Ferziger, Joel H.

    2003-01-01

    An efficient ghost-cell immersed boundary method (GCIBM) for simulating turbulent flows in complex geometries is presented. A boundary condition is enforced through a ghost cell method. The reconstruction procedure allows systematic development of numerical schemes for treating the immersed boundary while preserving the overall second-order accuracy of the base solver. Both Dirichlet and Neumann boundary conditions can be treated. The current ghost cell treatment is both suitable for staggered and non-staggered Cartesian grids. The accuracy of the current method is validated using flow past a circular cylinder and large eddy simulation of turbulent flow over a wavy surface. Numerical results are compared with experimental data and boundary-fitted grid results. The method is further extended to an existing ocean model (MITGCM) to simulate geophysical flow over a three-dimensional bump. The method is easily implemented as evidenced by our use of several existing codes

  8. Strengthening of the DNA-protein complex during stationary phase aging of cell cultures

    International Nuclear Information System (INIS)

    Khokhlov, A.N.; Chirkova, E.Yu.; Gorin, A.I.

    1986-01-01

    The possibility of accumulation of cross-linkages in the DNA-protein complex was studied during stationary phase aging of cells in culture. Chinese hamster cells were used in the experiments, along with human fibroblasts. 3 H-thymidine, 14 C-valine, and 14 C-leucine were added to the medium. The quantity of protein firmly bound with DNA was judged from the value of the coefficient 14 C/ 3 H determined with allowance for penetration of counting from the 14 C-channel into the 3 H-channel. The authors maintain that the results presented in this paper provide further evidence of the value of stationary phase cell cultures for the study of the mechanisms of aging and also of some of the general principles underlying hereditary pathology

  9. Review of tritium confinement and atmosphere detritiation system in hot cells complex

    International Nuclear Information System (INIS)

    Rizzello, Claudio; Borgognoni, Fabio; Pinna, Tonio; Tosti, Silvano

    2010-01-01

    The tritium confinement strategy adopted during the past years in the ITER hot cell building is compared to the safety requirements given by the standard ISO-17873 'Nuclear facilities - criteria for the design and operation of ventilation systems for nuclear installations other than nuclear reactors'. In fact, this is the reference safety guideline recommended by French licensing authorities. Several features of the considered design of the hot cell building are not in agreement with these guidelines. Main discrepancies concern the zoning of the hot cell complex, the flow rates of ventilation, and the possibility to recycle the room atmosphere and to detritiate the effluent air. These aspects are discussed together with some proposed modifications of the design.

  10. A fluorescence anisotropy method for measuring protein concentration in complex cell culture media.

    Science.gov (United States)

    Groza, Radu Constantin; Calvet, Amandine; Ryder, Alan G

    2014-04-22

    The rapid, quantitative analysis of the complex cell culture media used in biopharmaceutical manufacturing is of critical importance. Requirements for cell culture media composition profiling, or changes in specific analyte concentrations (e.g. amino acids in the media or product protein in the bioprocess broth) often necessitate the use of complicated analytical methods and extensive sample handling. Rapid spectroscopic methods like multi-dimensional fluorescence (MDF) spectroscopy have been successfully applied for the routine determination of compositional changes in cell culture media and bioprocess broths. Quantifying macromolecules in cell culture media is a specific challenge as there is a need to implement measurements rapidly on the prepared media. However, the use of standard fluorescence spectroscopy is complicated by the emission overlap from many media components. Here, we demonstrate how combining anisotropy measurements with standard total synchronous fluorescence spectroscopy (TSFS) provides a rapid, accurate quantitation method for cell culture media. Anisotropy provides emission resolution between large and small fluorophores while TSFS provides a robust measurement space. Model cell culture media was prepared using yeastolate (2.5 mg mL(-1)) spiked with bovine serum albumin (0 to 5 mg mL(-1)). Using this method, protein emission is clearly discriminated from background yeastolate emission, allowing for accurate bovine serum albumin (BSA) quantification over a 0.1 to 4.0 mg mL(-1) range with a limit of detection (LOD) of 13.8 μg mL(-1). Copyright © 2014. Published by Elsevier B.V.

  11. Deciphering the internal complexity of living cells with quantitative phase microscopy: a multiscale approach

    Science.gov (United States)

    Martinez-Torres, Cristina; Laperrousaz, Bastien; Berguiga, Lotfi; Boyer-Provera, Elise; Elezgaray, Juan; Nicolini, Franck E.; Maguer-Satta, Veronique; Arneodo, Alain; Argoul, Françoise

    2015-09-01

    The distribution of refractive indices (RIs) of a living cell contributes in a nonintuitive manner to its optical phase image and quite rarely can be inverted to recover its internal structure. The interpretation of the quantitative phase images of living cells remains a difficult task because (1) we still have very little knowledge on the impact of its internal macromolecular complexes on the local RI and (2) phase changes produced by light propagation through the sample are mixed with diffraction effects by the internal cell bodies. We propose to implement a two-dimensional wavelet-based contour chain detection method to distinguish internal boundaries based on their greatest optical path difference gradients. These contour chains correspond to the highest image phase contrast and follow the local RI inhomogeneities linked to the intracellular structural intricacy. Their statistics and spatial distribution are the morphological indicators suited for comparing cells of different origins and/or to follow their transformation in pathologic situations. We use this method to compare nonadherent blood cells from primary and laboratory culture origins and to assess the internal transformation of hematopoietic stem cells by the transduction of the BCR-ABL oncogene responsible for the chronic myelogenous leukemia.

  12. Modeling the complex activity of sickle cell and thalassemia specialist nurses in England.

    Science.gov (United States)

    Leary, Alison; Anionwu, Elizabeth N

    2014-01-01

    Specialist advanced practice nursing in hemoglobinopathies has a rich historical and descriptive literature. Subsequent work has shown that the role is valued by patients and families and also by other professionals. However, there is little empirical research on the complexity of activity of these services in terms of interventions offered. In addition, the work of clinical nurse specialists in England has been devalued through a perception of oversimplification. The purpose of this study was to understand the complexity of expert nursing practice in sickle cell and thalassemia. The approach taken to modeling complexity was used from common methods in mathematical modeling and computational mathematics. Knowledge discovery through data was the underpinning framework used in this study using a priori mined data. This allowed categorization of activity and articulation of complexity. In total, 8966 nursing events were captured over 1639 hours from a total of 22.8 whole time equivalents, and several data sources were mined. The work of specialist nurses in this area is complex in terms of the physical and psychosocial care they provide. The nurses also undertook case management activity such as utilizing a very large network of professionals, and others participated in admission avoidance work and education of patients' families and other staff. The work of nurses specializing in hemoglobinopathy care is complex and multidimensional and is likely to contribute to the quality of care in a cost-effective way. An understanding of this complexity can be used as an underpinning to establishing key performance indicators, optimum caseload calculations, and economic evaluation.

  13. Joint use of developed collagen-containing complexes and cell cultures in creating new tissue equivalents

    Directory of Open Access Journals (Sweden)

    K. V. Kulakova

    2016-01-01

    Full Text Available The purpose of the study is to assess the possibility of applying the integrated module as the basis of a celltissue equivalent for treatment of wounds of skin and soft tissues. In the frame of the set task the following problems were being solved: research of the spatial structure and architectonics of the surface of the developed base collagen-containing materials and their biocompatibility with cell cultures.Materials and methods. The study of a material which is a two-layer complex film, consisting of collagen and polysaccharide components was carried out. The collagen was separated from the dermis and was then impregnated with particulate demineralized bone matrix (DCM according to the original methodology. For the purposes of the study the dehydrated material was created in the form of a film. Electron microscopic examination of surfaces was performed on scanning electron microscope JEOL JSM-IT300LV in high vacuum and at low values of probe current (< 0,1 nА. Studies to assess the viability of the cells cultivated on films of collagen material (tested for cytotoxicity and the adhesive capacity were performed in vitro using strains of diploid human fibroblasts 4–6 passage. The culture condition was visually assessed using an inverted Leica microscope DM IL (Carl Zeiss, Austria, equipped with a computerizes program of control of culture growth (Leica IM 1000.Results. The data obtained in the study of the surface structure of the developed complex module showed that it seems to be promising as a basic component of the cellular-tissue system with its large number of structural formations for fixation of the cells and a well-organized barrier layer capable of vapor - permeability. Experiments in vitro confirmed the absence of toxicity of the material being studied in relation to the culture of dermal human fibroblasts, suggesting the possibility of creation on its basis of cell-tissue complex and further experimental studies in vivo

  14. Arabidopsis thaliana plants lacking the ARP2/3 complex show defects in cell wall assembly and auxin distribution.

    Science.gov (United States)

    Pratap Sahi, Vaidurya; Cifrová, Petra; García-González, Judith; Kotannal Baby, Innu; Mouillé, Gregory; Gineau, Emilie; Müller, Karel; Baluška, František; Soukup, Aleš; Petrášek, Jan; Schwarzerová, Katerina

    2017-12-25

    The cytoskeleton plays an important role in the synthesis of plant cell walls. Both microtubules and actin cytoskeleton are known to be involved in the morphogenesis of plant cells through their role in cell wall building. The role of ARP2/3-nucleated actin cytoskeleton in the morphogenesis of cotyledon pavement cells has been described before. Seedlings of Arabidopsis mutants lacking a functional ARP2/3 complex display specific cell wall-associated defects. In three independent Arabidopsis mutant lines lacking subunits of the ARP2/3 complex, phenotypes associated with the loss of the complex were analysed throughout plant development. Organ size and anatomy, cell wall composition, and auxin distribution were investigated. ARP2/3-related phenotype is associated with changes in cell wall composition, and the phenotype is manifested especially in mature tissues. Cell walls of mature plants contain less cellulose and a higher amount of homogalacturonan, and display changes in cell wall lignification. Vascular bundles of mutant inflorescence stems show a changed pattern of AUX1-YFP expression. Plants lacking a functional ARP2/3 complex have decreased basipetal auxin transport. The results suggest that the ARP2/3 complex has a morphogenetic function related to cell wall synthesis and auxin transport. © The Author(s) 2017. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. The diabetes medication Canagliflozin reduces cancer cell proliferation by inhibiting mitochondrial complex-I supported respiration

    Directory of Open Access Journals (Sweden)

    Linda A. Villani

    2016-10-01

    Full Text Available Objective: The sodium-glucose transporter 2 (SGLT2 inhibitors Canagliflozin and Dapagliflozin are recently approved medications for type 2 diabetes. Recent studies indicate that SGLT2 inhibitors may inhibit the growth of some cancer cells but the mechanism(s remain unclear. Methods: Cellular proliferation and clonogenic survival were used to assess the sensitivity of prostate and lung cancer cell growth to the SGLT2 inhibitors. Oxygen consumption, extracellular acidification rate, cellular ATP, glucose uptake, lipogenesis, and phosphorylation of AMP-activated protein kinase (AMPK, acetyl-CoA carboxylase, and the p70S6 kinase were assessed. Overexpression of a protein that maintains complex-I supported mitochondrial respiration (NDI1 was used to establish the importance of this pathway for mediating the anti-proliferative effects of Canagliflozin. Results: Clinically achievable concentrations of Canagliflozin, but not Dapagliflozin, inhibit cellular proliferation and clonogenic survival of prostate and lung cancer cells alone and in combination with ionizing radiation and the chemotherapy Docetaxel. Canagliflozin reduced glucose uptake, mitochondrial complex-I supported respiration, ATP, and lipogenesis while increasing the activating phosphorylation of AMPK. The overexpression of NDI1 blocked the anti-proliferative effects of Canagliflozin indicating reductions in mitochondrial respiration are critical for anti-proliferative actions. Conclusion: These data indicate that like the biguanide metformin, Canagliflozin not only lowers blood glucose but also inhibits complex-I supported respiration and cellular proliferation in prostate and lung cancer cells. These observations support the initiation of studies evaluating the clinical efficacy of Canagliflozin on limiting tumorigenesis in pre-clinical animal models as well epidemiological studies on cancer incidence relative to other glucose lowering therapies in clinical populations. Keywords: AMP

  16. Composition and immunoreactivity of the A60 complex and other cell fractions from Mycobacterium bovis BCG.

    Science.gov (United States)

    Cocito, C; Vanlinden, F

    1995-02-01

    Surface static cultures of Mycobacterium bovis BCG contained cells embedded in an extracellular matrix, whose mechanical removal yielded free cells that were pressure disrupted and fractionated into cytoplasm and walls. Cell envelopes were either mechanically disrupted or extracted with detergents. Intracellular and extracellular fractions were analysed for proteins, polysaccharides, and antigen 6O (A60), a major complex immunodominant in tuberculosis. A60 was present in extracellular matrix, cytoplasm and walls: it represented a substantial portion of the proteins and polysaccharides of these fractions. While the protein/polysaccharide ratio varied according to the origin of A60 preparations, the electrophoretic patterns of A60 proteins (which accounted for the immunogenicity of the complex) remained unchanged. Western blots pointed to the proteins present within the 29-45 kDa range as the A60 components endowed with the highest immunogenicity level. Since the most heavily stained protein bands in SDS-PAGE patterns were located outside the region best recognized by antisera, a striking discordance was found between concentration and immunogenicity patterns of A60 proteins. The electrophoretic patterns of A60- and non-A60-proteins from cytoplasm were also different. A60 complexes in dot blots and some electrophoresed A60 proteins reacted with monoclonal antibodies directed against lipoarabinomannan (LAM), a highly immunogenic polymer of cell envelope. This contaminating compound was removed from A60 with organic solvents and detergents. SDS-PAGE and Western blot patterns of proteins from delipidated A60 were similar to those of native A60 proteins.

  17. Three-dimensional bioprinting of complex cell laden alginate hydrogel structures.

    Science.gov (United States)

    Tabriz, Atabak Ghanizadeh; Hermida, Miguel A; Leslie, Nicholas R; Shu, Wenmiao

    2015-12-21

    Different bioprinting techniques have been used to produce cell-laden alginate hydrogel structures, however these approaches have been limited to 2D or simple three-dimension (3D) structures. In this study, a new extrusion based bioprinting technique was developed to produce more complex alginate hydrogel structures. This was achieved by dividing the alginate hydrogel cross-linking process into three stages: primary calcium ion cross-linking for printability of the gel, secondary calcium cross-linking for rigidity of the alginate hydrogel immediately after printing and tertiary barium ion cross-linking for long-term stability of the alginate hydrogel in culture medium. Simple 3D structures including tubes were first printed to ensure the feasibility of the bioprinting technique and then complex 3D structures such as branched vascular structures were successfully printed. The static stiffness of the alginate hydrogel after printing was 20.18 ± 1.62 KPa which was rigid enough to sustain the integrity of the complex 3D alginate hydrogel structure during the printing. The addition of 60 mM barium chloride was found to significantly extend the stability of the cross-linked alginate hydrogel from 3 d to beyond 11 d without compromising the cellular viability. The results based on cell bioprinting suggested that viability of U87-MG cells was 93 ± 0.9% immediately after bioprinting and cell viability maintained above 88% ± 4.3% in the alginate hydrogel over the period of 11 d.

  18. Relationship between macular ganglion cell complex thickness and macular outer retinal thickness: a spectral-domain optical coherence tomography study.

    Science.gov (United States)

    Kita, Yoshiyuki; Kita, Ritsuko; Takeyama, Asuka; Anraku, Ayako; Tomita, Goji; Goldberg, Ivan

    2013-01-01

    To assess the relationship between macular ganglion cell complex and macular outer retinal thicknesses. Case-control study. Forty-two normal eyes and 91 eyes with primary open-angle glaucoma were studied. Spectral-domain optical coherence tomography (RTVue-100) was used to measure the macular ganglion cell complex and macular outer retinal thickness. Ganglion cell complex to outer retinal thickness ratio was also calculated. The relationships between the ganglion cell complex and outer retinal thicknesses and between the ganglion cell complex to outer retinal thickness ratio and outer retinal thickness were evaluated. There was a positive correlation between ganglion cell complex and outer retinal thicknesses in the normal group and the glaucoma group (r = 0.53, P variation in the outer retinal thickness. Therefore, when determining the ganglion cell complex, it seems necessary to consider the outer retinal thickness as well. We propose the ratio as a suitable parameter to account for individual variations in outer retinal thickness. © 2013 The Authors. Clinical and Experimental Ophthalmology © 2013 Royal Australian and New Zealand College of Ophthalmologists.

  19. Recycling of epidermal growth factor-receptor complexes in A431 cells: Identification of dual pathways

    International Nuclear Information System (INIS)

    Sorkin, A.; Krolenko, S.; Kudrjavtceva, N.; Lazebnik, J.; Teslenko, L.; Soderquist, A.M.; Nikolsky, N.

    1991-01-01

    The intracellular sorting of EGF-receptor complexes (EGF-RC) has been studied in human epidermoid carcinoma A431 cells. Recycling of EGF was found to occur rapidly after internalization at 37 degrees C. The initial rate of EGF recycling was reduced at 18 degrees C. A significant pool of internalized EGF was incapable of recycling at 18 degrees C but began to recycle when cells were warmed to 37 degrees C. The relative rate of EGF outflow at 37 degrees C from cells exposed to an 18 degrees C temperature block was slower (t1/2 approximately 20 min) than the rate from cells not exposed to a temperature block (t1/2 approximately 5-7 min). These data suggest that there might be both short- and long-time cycles of EGF recycling in A431 cells. Examination of the intracellular EGF-RC dissociation and dynamics of short- and long-time recycling indicated that EGF recycled as EGF-RC. Moreover, EGF receptors that were covalently labeled with a photoactivatable derivative of 125 I-EGF recycled via the long-time pathway at a rate similar to that of 125 I-EGF. Since EGF-RC degradation was also blocked at 18 degrees C, we propose that sorting to the lysosomal and long-time recycling pathway may occur after a highly temperature-sensitive step, presumably in the late endosomes

  20. Analysis of a fuel cell on-site integrated energy system for a residential complex

    Science.gov (United States)

    Simons, S. N.; Maag, W. L.

    1979-01-01

    The energy use and costs of the on-site integrated energy system (OS/IES) which provides electric power from an on-site power plant and recovers heat that would normally be rejected to the environment is compared to a conventional system purchasing electricity from a utility and a phosphoric acid fuel cell powered system. The analysis showed that for a 500-unit apartment complex a fuel OS/IES would be about 10% more energy conservative in terms of total coal consumption than a diesel OS/IES system or a conventional system. The fuel cell OS/IES capital costs could be 30 to 55% greater than the diesel OS/IES capital costs for the same life cycle costs. The life cycle cost of a fuel cell OS/IES would be lower than that for a conventional system as long as the cost of electricity is greater than $0.05 to $0.065/kWh. An analysis of several parametric combinations of fuel cell power plant and state-of-art energy recovery systems and annual fuel requirement calculations for four locations were made. It was shown that OS/IES component choices are a major factor in fuel consumption, with the least efficient system using 25% more fuel than the most efficient. Central air conditioning and heat pumps result in minimum fuel consumption while individual air conditioning units increase it, and in general the fuel cell of highest electrical efficiency has the lowest fuel consumption.

  1. A Complex Interaction Between Reduced Reelin Expression and Prenatal Organophosphate Exposure Alters Neuronal Cell Morphology

    Directory of Open Access Journals (Sweden)

    Brian R. Mullen

    2016-06-01

    Full Text Available Genetic and environmental factors are both likely to contribute to neurodevelopmental disorders including schizophrenia, autism spectrum disorders, and major depressive disorders. Prior studies from our laboratory and others have demonstrated that the combinatorial effect of two factors—reduced expression of reelin protein and prenatal exposure to the organophosphate pesticide chlorpyrifos oxon—gives rise to acute biochemical effects and to morphological and behavioral phenotypes in adolescent and young adult mice. In the current study, we examine the consequences of these factors on reelin protein expression and neuronal cell morphology in adult mice. While the cell populations that express reelin in the adult brain appear unchanged in location and distribution, the levels of full length and cleaved reelin protein show persistent reductions following prenatal exposure to chlorpyrifos oxon. Cell positioning and organization in the hippocampus and cerebellum are largely normal in animals with either reduced reelin expression or prenatal exposure to chlorpyrifos oxon, but cellular complexity and dendritic spine organization is altered, with a skewed distribution of immature dendritic spines in adult animals. Paradoxically, combinatorial exposure to both factors appears to generate a rescue of the dendritic spine phenotypes, similar to the mitigation of behavioral and morphological changes observed in our prior study. Together, our observations support an interaction between reelin expression and chlorpyrifos oxon exposure that is not simply additive, suggesting a complex interplay between genetic and environmental factors in regulating brain morphology.

  2. The subclonal complexity of STIL-TAL1+ T-cell acute lymphoblastic leukaemia.

    Science.gov (United States)

    Furness, Caroline L; Mansur, Marcela B; Weston, Victoria J; Ermini, Luca; van Delft, Frederik W; Jenkinson, Sarah; Gale, Rosemary; Harrison, Christine J; Pombo-de-Oliveira, Maria S; Sanchez-Martin, Marta; Ferrando, Adolfo A; Kearns, Pamela; Titley, Ian; Ford, Anthony M; Potter, Nicola E; Greaves, Mel

    2018-03-20

    Single-cell genetics were used to interrogate clonal complexity and the sequence of mutational events in STIL-TAL1+ T-ALL. Single-cell multicolour FISH was used to demonstrate that the earliest detectable leukaemia subclone contained the STIL-TAL1 fusion and copy number loss of 9p21.3 (CDKN2A/CDKN2B locus), with other copy number alterations including loss of PTEN occurring as secondary subclonal events. In three cases, multiplex qPCR and phylogenetic analysis were used to produce branching evolutionary trees recapitulating the snapshot history of T-ALL evolution in this leukaemia subtype, which confirmed that mutations in key T-ALL drivers, including NOTCH1 and PTEN, were subclonal and reiterative in distinct subclones. Xenografting confirmed that self-renewing or propagating cells were genetically diverse. These data suggest that the STIL-TAL1 fusion is a likely founder or truncal event. Therapies targeting the TAL1 auto-regulatory complex are worthy of further investigation in T-ALL.

  3. REST-mediated recruitment of polycomb repressor complexes in mammalian cells

    DEFF Research Database (Denmark)

    Dietrich, Nikolaj; Lerdrup, Mads; Landt, Eskild

    2012-01-01

    Polycomb Repressive Complex (PRC) 1 and PRC2 regulate genes involved in differentiation and development. However, the mechanism for how PRC1 and PRC2 are recruited to genes in mammalian cells is unclear. Here we present evidence for an interaction between the transcription factor REST, PRC1......, and increased gene expression. Genome-wide analysis of Polycomb binding in Rest¿/¿ and Eed¿/¿ mouse embryonic stem (mES) cells showed that Rest was required for PRC1 recruitment to a subset of Polycomb regulated neuronal genes. Furthermore, we found that PRC1 can be recruited to Rest binding sites independently...... of CpG islands and the H3K27Me3 mark. Surprisingly, PRC2 was frequently increased around Rest binding sites located in CpG-rich regions in the Rest¿/¿ mES cells, indicating a more complex interplay where Rest also can limit PRC2 recruitment. Therefore, we propose that Rest has context...

  4. The Yersinia pseudotuberculosis and Yersinia pestis toxin complex is active against cultured mammalian cells.

    Science.gov (United States)

    Hares, Michelle C; Hinchliffe, Stewart J; Strong, Philippa C R; Eleftherianos, Ioannis; Dowling, Andrea J; ffrench-Constant, Richard H; Waterfield, Nick

    2008-11-01

    The toxin complex (Tc) genes were first identified in the insect pathogen Photorhabdus luminescens and encode approximately 1 MDa protein complexes which are toxic to insect pests. Subsequent genome sequencing projects have revealed the presence of tc orthologues in a range of bacterial pathogens known to be associated with insects. Interestingly, members of the mammalian-pathogenic yersiniae have also been shown to encode Tc orthologues. Studies in Yersinia enterocolitica have shown that divergent tc loci either encode insect-active toxins or play a role in colonization of the gut in gastroenteritis models of rats. So far little is known about the activity of the Tc proteins in the other mammalian-pathogenic yersiniae. Here we present work to suggest that Tc proteins in Yersinia pseudotuberculosis and Yersinia pestis are not insecticidal toxins but have evolved for mammalian pathogenicity. We show that Tc is secreted by Y. pseudotuberculosis strain IP32953 during growth in media at 28 degrees C and 37 degrees C. We also demonstrate that oral toxicity of strain IP32953 to Manduca sexta larvae is not due to Tc expression and that lysates of Escherichia coli BL21 expressing the Yersinia Tc proteins are not toxic to Sf9 insect cells but are toxic to cultured mammalian cell lines. Cell lysates of E. coli BL21 expressing the Y. pseudotuberculosis Tc proteins caused actin ruffles, vacuoles and multi-nucleation in cultured human gut cells (Caco-2); similar morphology was observed after application of a lysate of E. coli BL21 expressing the Y. pestis Tc proteins to mouse fibroblast NIH3T3 cells, but not Caco-2 cells. Finally, transient expression of the individual Tc proteins in Caco-2 and NIH3T3 cell lines reproduced the actin and nuclear rearrangement observed with the topical applications. Together these results add weight to the growing hypothesis that the Tc proteins in Y. pseudotuberculosis and Y. pestis have been adapted for mammalian pathogenicity. We further

  5. Toroidal surface complexes of bacteriophage φ12 are responsible for host-cell attachment

    International Nuclear Information System (INIS)

    Leo-Macias, Alejandra; Katz, Garrett; Wei Hui; Alimova, Alexandra; Katz, A.; Rice, William J.; Diaz-Avalos, Ruben; Hu Guobin; Stokes, David L.; Gottlieb, Paul

    2011-01-01

    Cryo-electron tomography and subtomogram averaging are utilized to determine that the bacteriophage φ12, a member of the Cystoviridae family, contains surface complexes that are toroidal in shape, are composed of six globular domains with six-fold symmetry, and have a discrete density connecting them to the virus membrane-envelope surface. The lack of this kind of spike in a reassortant of φ12 demonstrates that the gene for the hexameric spike is located in φ12's medium length genome segment, likely to the P3 open reading frames which are the proteins involved in viral-host cell attachment. Based on this and on protein mass estimates derived from the obtained averaged structure, it is suggested that each of the globular domains is most likely composed of a total of four copies of P3a and/or P3c proteins. Our findings may have implications in the study of the evolution of the cystovirus species in regard to their host specificity. - Research Highlights: → Subtomogram averaging reveals enhanced detail of a φ12 cystovirus surface protein complex. → The surface protein complex has a toroidal shape and six-fold symmetry. → It is encoded by the medium-size genome segment. → The proteins of the surface complex most likely are one copy of P3a and three copies of P3c.

  6. Cellular responses induced by Cu(II quinolinonato complexes in human tumor and hepatic cells

    Directory of Open Access Journals (Sweden)

    Trávníček Zdeněk

    2012-12-01

    Full Text Available Abstract Background Inspired by the unprecedented historical success of cisplatin, one of the most important research directions in bioinorganic and medicinal chemistry is dedicated to the development of new anticancer compounds with the potential to surpass it in antitumor activity, while having lower unwanted side-effects. Therefore, a series of copper(II mixed-ligand complexes of the type [Cu(qui(L]Y · xH2O (1–6, where Hqui = 2-phenyl-3-hydroxy-4(1H-quinolinone, Y = NO3 (1, 3, 5 or BF4 (2, 4, 6, and L = 1,10-phenanthroline (phen (1, 2, 5-methyl-1,10-phenanthroline (mphen (3, 4 and bathophenanthroline (bphen (5, 6, was studied for their in vitro cytotoxicity against several human cancer cell lines (A549 lung carcinoma, HeLa cervix epitheloid carcinoma, G361 melanoma cells, A2780 ovarian carcinoma, A2780cis cisplatin-resistant ovarian carcinoma, LNCaP androgen-sensitive prostate adenocarcinoma and THP-1 monocytic leukemia. Results The tested complexes displayed a stronger cytotoxic effect against all the cancer cells as compared to cisplatin. The highest cytotoxicity was found for the complexes 4 (IC50 = 0.36 ± 0.05 μM and 0.56 ± 0.15 μM, 5 (IC50 = 0.66 ± 0.07 μM and 0.73 ± 0.08 μM and 6 (IC50 = 0.57 ± 0.11 μM and 0.70 ± 0.20 μM against A2780, and A2780cis respectively, as compared with the values of 12.0 ± 0.8 μM and 27.0 ± 4.6 μM determined for cisplatin. Moreover, the tested complexes were much less cytotoxic to primary human hepatocytes than to the cancer cells. The complexes 5 and 6 exhibited significantly high ability to modulate secretion of the pro-inflammatory cytokines TNF-α (2873 ± 238 pg/mL and 3284 ± 139 pg/mL for 5, and 6 respectively and IL-1β (1177 ± 128 pg/mL and 1087 ± 101 pg/mL for 5, and 6 respectively tested on the lipopolysaccharide (LPS-stimulated THP-1 cells as compared with the values of 1173

  7. Heptachlor induced mitochondria-mediated cell death via impairing electron transport chain complex III

    International Nuclear Information System (INIS)

    Hong, Seokheon; Kim, Joo Yeon; Hwang, Joohyun; Shin, Ki Soon; Kang, Shin Jung

    2013-01-01

    Highlights: •Heptachlor inhibited mitochondrial electron transport chain complex III activity. •Heptachlor promoted generation of reactive oxygen species. •Heptachlor induced Bax activation. •Heptachlor induced mitochondria-mediated and caspase-dependent apoptosis. -- Abstract: Environmental toxins like pesticides have been implicated in the pathogenesis of Parkinson’s disease (PD). Epidemiological studies suggested that exposures to organochlorine pesticides have an association with an increased PD risk. In the present study, we examined the mechanism of toxicity induced by an organochlorine pesticide heptachlor. In a human dopaminergic neuroblastoma SH-SY5Y cells, heptachlor induced both morphological and functional damages in mitochondria. Interestingly, the compound inhibited mitochondrial electron transport chain complex III activity. Rapid generation of reactive oxygen species and the activation of Bax were then detected. Subsequently, mitochondria-mediated, caspase-dependent apoptosis followed. Our results raise a possibility that an organochlorine pesticide heptachlor can act as a neurotoxicant associated with PD

  8. Structural measurements and cell line studies of the copper-PEG-Rifampicin complex against Mycobacterium tuberculosis.

    Science.gov (United States)

    Manning, Thomas; Mikula, Rachel; Wylie, Greg; Phillips, Dennis; Jarvis, Jackie; Zhang, Fengli

    2015-02-01

    The bacterium responsible for tuberculosis is increasing its resistance to antibiotics resulting in new multidrug-resistant Mycobacterium tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB). In this study, several analytical techniques including NMR, FT-ICR, MALDI-MS, LC-MS and UV/Vis are used to study the copper-Rifampicin-Polyethylene glycol (PEG-3350) complex. The copper (II) cation is a carrier for the antibiotic Rifampicin as well as nutrients for the bacterium. The NIH-NIAID cell line containing several Tb strains (including antibiotic resistant strains) is tested against seven copper-PEG-RIF complex variations. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. The Mediator complex: a master coordinator of transcription and cell lineage development.

    Science.gov (United States)

    Yin, Jing-wen; Wang, Gang

    2014-03-01

    Mediator is a multiprotein complex that is required for gene transcription by RNA polymerase II. Multiple subunits of the complex show specificity in relaying information from signals and transcription factors to the RNA polymerase II machinery, thus enabling control of the expression of specific genes. Recent studies have also provided novel mechanistic insights into the roles of Mediator in epigenetic regulation, transcriptional elongation, termination, mRNA processing, noncoding RNA activation and super enhancer formation. Based on these specific roles in gene regulation, Mediator has emerged as a master coordinator of development and cell lineage determination. Here, we describe the most recent advances in understanding the mechanisms of Mediator function, with an emphasis on its role during development and disease.

  10. Heptachlor induced mitochondria-mediated cell death via impairing electron transport chain complex III

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Seokheon; Kim, Joo Yeon; Hwang, Joohyun [Department of Molecular Biology, Sejong University, Seoul 143-747 (Korea, Republic of); Shin, Ki Soon [Department of Biology, Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Kang, Shin Jung, E-mail: sjkang@sejong.ac.kr [Department of Molecular Biology, Sejong University, Seoul 143-747 (Korea, Republic of)

    2013-08-09

    Highlights: •Heptachlor inhibited mitochondrial electron transport chain complex III activity. •Heptachlor promoted generation of reactive oxygen species. •Heptachlor induced Bax activation. •Heptachlor induced mitochondria-mediated and caspase-dependent apoptosis. -- Abstract: Environmental toxins like pesticides have been implicated in the pathogenesis of Parkinson’s disease (PD). Epidemiological studies suggested that exposures to organochlorine pesticides have an association with an increased PD risk. In the present study, we examined the mechanism of toxicity induced by an organochlorine pesticide heptachlor. In a human dopaminergic neuroblastoma SH-SY5Y cells, heptachlor induced both morphological and functional damages in mitochondria. Interestingly, the compound inhibited mitochondrial electron transport chain complex III activity. Rapid generation of reactive oxygen species and the activation of Bax were then detected. Subsequently, mitochondria-mediated, caspase-dependent apoptosis followed. Our results raise a possibility that an organochlorine pesticide heptachlor can act as a neurotoxicant associated with PD.

  11. In vitro formation of the Merkel cell-neurite complex in embryonic mouse whiskers using organotypic co-cultures.

    Science.gov (United States)

    Ishida, Kentaro; Saito, Tetsuichiro; Mitsui, Toshiyuki

    2018-06-01

    A Merkel cell-neurite complex is a touch receptor composed of specialized epithelial cells named Merkel cells and peripheral sensory nerves in the skin. Merkel cells are found in touch-sensitive skin components including whisker follicles. The nerve fibers that innervate Merkel cells of a whisker follicle extend from the maxillary branch of the trigeminal ganglion. Whiskers as a sensory organ attribute to the complicated architecture of the Merkel cell-neurite complex, and therefore it is intriguing how the structure is formed. However, observing the dynamic process of the formation of a Merkel cell-neurite complex in whiskers during embryonic development is still difficult. In this study, we tried to develop an organotypic co-culture method of a whisker pad and a trigeminal ganglion explant to form the Merkel cell-neurite complex in vitro. We initially developed two distinct culture methods of a single whisker row and a trigeminal ganglion explant, and then combined them. By dissecting and cultivating a single row from a whisker pad, the morphogenesis of whisker follicles could be observed under a microscope. After the co-cultivation of the whisker row with a trigeminal ganglion explant, a Merkel cell-neurite complex composed of Merkel cells, which were positive for both cytokeratin 8 and SOX2, Neurofilament-H-positive trigeminal nerve fibers and Schwann cells expressing Nestin, SOX2 and SOX10 was observed via immunohistochemical analyses. These results suggest that the process for the formation of a Merkel cell-neurite complex can be observed under a microscope using our organotypic co-culture method. © 2018 Japanese Society of Developmental Biologists.

  12. Topological recursion for chord diagrams, RNA complexes, and cells in moduli spaces

    DEFF Research Database (Denmark)

    Andersen, Jørgen Ellegaard; Chekhov, Leonid O.; Penner, Robert

    2013-01-01

    and free energies are convergent for small t and all s as a perturbation of the Gaussian potential, which arises for st=0. This perturbation is computed using the formalism of the topological recursion. The corresponding enumeration of chord diagrams gives at once the number of RNA complexes of a given...... topology as well as the number of cells in Riemann's moduli spaces for bordered surfaces. The free energies are computed here in principle for all genera and explicitly for genera less than four....

  13. Differential expression of Mediator complex subunit MED15 in testicular germ cell tumors.

    Science.gov (United States)

    Klümper, Niklas; Syring, Isabella; Offermann, Anne; Shaikhibrahim, Zaki; Vogel, Wenzel; Müller, Stefan C; Ellinger, Jörg; Strauß, Arne; Radzun, Heinz Joachim; Ströbel, Philipp; Brägelmann, Johannes; Perner, Sven; Bremmer, Felix

    2015-09-17

    Testicular germ cell tumors (TGCT) are the most common cancer entities in young men with increasing incidence observed in the last decades. For therapeutic management it is important, that TGCT are divided into several histological subtypes. MED15 is part of the multiprotein Mediator complex which presents an integrative hub for transcriptional regulation and is known to be deregulated in several malignancies, such as prostate cancer and bladder cancer role, whereas the role of the Mediator complex in TGCT has not been investigated so far. Aim of the study was to investigate the implication of MED15 in TGCT development and its stratification into histological subtypes. Immunohistochemical staining (IHC) against Mediator complex subunit MED15 was conducted on a TGCT cohort containing tumor-free testis (n = 35), intratubular germ cell neoplasia unclassified (IGCNU, n = 14), seminomas (SEM, n = 107) and non-seminomatous germ cell tumors (NSGCT, n = 42), further subdivided into embryonic carcinomas (EC, n = 30), yolk sac tumors (YST, n = 5), chorionic carcinomas (CC, n = 5) and teratomas (TER, n = 2). Quantification of MED15 protein expression was performed through IHC followed by semi-quantitative image analysis using the Definiens software. In tumor-free seminiferous tubules, MED15 protein expression was absent or only low expressed in spermatogonia. Interestingly, the precursor lesions IGCNU exhibited heterogeneous but partly very strong MED15 expression. SEM weakly express the Mediator complex subunit MED15, whereas NSGCT and especially EC show significantly enhanced expression compared to tumor-free testis. In conclusion, MED15 is differentially expressed in tumor-free testis and TGCT. While MED15 is absent or low in tumor-free testis and SEM, NSGCT highly express MED15, hinting at the diagnostic potential of this marker to distinguish between SEM and NSGCT. Further, the precursor lesion IGCNU showed increased nuclear MED15

  14. Theoretical Study of Copper Complexes: Molecular Structure, Properties, and Its Application to Solar Cells

    Directory of Open Access Journals (Sweden)

    Jesus Baldenebro-Lopez

    2013-01-01

    Full Text Available We present a theoretical investigation of copper complexes with potential applications as sensitizers for solar cells. The density functional theory (DFT and time-dependent DFT were utilized, using the M06 hybrid meta-GGA functional with the LANL2DZ (D95V on first row and DZVP basis sets. This level of calculation was used to find the optimized molecular structure, the absorption spectra, the molecular orbitals energies, and the chemical reactivity parameters that arise from conceptual DFT. Solvent effects have been taken into account by an implicit approach, namely, the polarizable continuum model (PCM, using the nonequilibrium version of the IEF-PCM model.

  15. Self-potential and Complex Conductivity Monitoring of In Situ Hydrocarbon Remediation in Microbial Fuel Cell

    Science.gov (United States)

    Zhang, C.; Revil, A.; Ren, Z.; Karaoulis, M.; Mendonca, C. A.

    2013-12-01

    Petroleum hydrocarbon contamination of soil and groundwater in both non-aqueous phase liquid and dissolved forms generated from spills and leaks is a wide spread environmental issue. Traditional cleanup of hydrocarbon contamination in soils and ground water using physical, chemical, and biological remedial techniques is often expensive and ineffective. Recent studies show that the microbial fuel cell (MFC) can simultaneously enhance biodegradation of hydrocarbons in soil and groundwater and yield electricity. Non-invasive geophysical techniques such as self-potential (SP) and complex conductivity (induced polarization) have shown the potential to detect and characterize the nature of electron transport mechanism of in situ bioremediation of organic contamination plumes. In this study, we deployed both SP and complex conductivity in lab scale MFCs to monitor time-laps geophysical response of degradation of hydrocarbons by MFC. Two different sizes of MFC reactors were used in this study (DI=15 cm cylinder reactor and 94.5cm x 43.5 cm rectangle reactor), and the initial hydrocarbon concentration is 15 g diesel/kg soil. SP and complex conductivity measurements were measured using non-polarizing Ag/AgCl electrodes. Sensitivity study was also performed using COMSOL Multiphysics to test different electrode configurations. The SP measurements showed stronger anomalies adjacent to the MFC than locations afar, and both real and imaginary parts of complex conductivity are greater in areas close to MFC than areas further away and control samples without MFC. The joint use of SP and complex conductivity could in situ evaluate the dynamic changes of electrochemical parameters during this bioremediation process at spatiotemporal scales unachievable with traditional sampling methods. The joint inversion of these two methods to evaluate the efficiency of MFC enhanced hydrocarbon remediation in the subsurface.

  16. Bio-Photoelectrochemical Solar Cells Incorporating Reaction Center and Reaction Center Plus Light Harvesting Complexes

    Science.gov (United States)

    Yaghoubi, Houman

    Harvesting solar energy can potentially be a promising solution to the energy crisis now and in the future. However, material and processing costs continue to be the most important limitations for the commercial devices. A key solution to these problems might lie within the development of bio-hybrid solar cells that seeks to mimic photosynthesis to harvest solar energy and to take advantage of the low material costs, negative carbon footprint, and material abundance. The bio-photoelectrochemical cell technologies exploit biomimetic means of energy conversion by utilizing plant-derived photosystems which can be inexpensive and ultimately the most sustainable alternative. Plants and photosynthetic bacteria harvest light, through special proteins called reaction centers (RCs), with high efficiency and convert it into electrochemical energy. In theory, photosynthetic RCs can be used in a device to harvest solar energy and generate 1.1 V open circuit voltage and ~1 mA cm-2 short circuit photocurrent. Considering the nearly perfect quantum yield of photo-induced charge separation, efficiency of a protein-based solar cell might exceed 20%. In practice, the efficiency of fabricated devices has been limited mainly due to the challenges in the electron transfer between the protein complex and the device electrodes as well as limited light absorption. The overarching goal of this work is to increase the power conversion efficiency in protein-based solar cells by addressing those issues (i.e. electron transfer and light absorption). This work presents several approaches to increase the charge transfer rate between the photosynthetic RC and underlying electrode as well as increasing the light absorption to eventually enhance the external quantum efficiency (EQE) of bio-hybrid solar cells. The first approach is to decrease the electron transfer distance between one of the redox active sites in the RC and the underlying electrode by direct attachment of the of protein complex

  17. Lipids in host-pathogen interactions: pathogens exploit the complexity of the host cell lipidome.

    Science.gov (United States)

    van der Meer-Janssen, Ynske P M; van Galen, Josse; Batenburg, Joseph J; Helms, J Bernd

    2010-01-01

    Lipids were long believed to have a structural role in biomembranes and a role in energy storage utilizing cellular lipid droplets and plasma lipoproteins. Research over the last decades has identified an additional role of lipids in cellular signaling, membrane microdomain organization and dynamics, and membrane trafficking. These properties make lipids an attractive target for pathogens to modulate host cell processes in order to allow their survival and replication. In this review we will summarize the often ingenious strategies of pathogens to modify the lipid homeostasis of host cells, allowing them to divert cellular processes. To this end pathogens take full advantage of the complexity of the lipidome. The examples are categorized in generalized and emerging principles describing the involvement of lipids in host-pathogen interactions. Several pathogens are described that simultaneously induce multiple changes in the host cell signaling and trafficking mechanisms. Elucidation of these pathogen-induced changes may have important implications for drug development. The emergence of high-throughput lipidomic techniques will allow the description of changes of the host cell lipidome at the level of individual molecular lipid species and the identification of lipid biomarkers.

  18. Endosomal sorting complexes required for ESCRTing cells toward death during neurogenesis, neurodevelopment and neurodegeneration.

    Science.gov (United States)

    Kaul, Zenia; Chakrabarti, Oishee

    2018-03-25

    The endosomal sorting complexes required for transport (ESCRT) proteins help in the recognition, sorting and degradation of ubiquitinated cargoes from the cell surface, long-lived proteins or aggregates, and aged organelles present in the cytosol. These proteins take part in the endo-lysosomal system of degradation. The ESCRT proteins also play an integral role in cytokinesis, viral budding and mRNA transport. Many neurodegenerative diseases are caused by toxic accumulation of cargo in the cell, which causes stress and ultimately leads to neuronal death. This accumulation of cargo occurs because of defects in the endo-lysosomal degradative pathway-loss of function of ESCRTs has been implicated in this mechanism. ESCRTs also take part in many survival processes, lack of which can culminate in neuronal cell death. While the role played by the ESCRT proteins in maintaining healthy neurons is known, their role in neurodegenerative diseases is still poorly understood. In this review, we highlight the importance of ESCRTs in maintaining healthy neurons and then suggest how perturbations in many of the survival mechanisms governed by these proteins could eventually lead to cell death; quite often these correlations are not so obviously laid out. Extensive neuronal death eventually culminates in neurodegeneration. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  19. Immune complexes stimulate CCR7-dependent dendritic cell migration to lymph nodes

    Science.gov (United States)

    Clatworthy, Menna R.; Aronin, Caren E. Petrie; Mathews, Rebeccah J.; Morgan, Nicole; Smith, Kenneth G.C.; Germain, Ronald N.

    2014-01-01

    Antibodies are critical for defence against a variety of microbes but may also be pathogenic in some autoimmune diseases. Many effector functions of antibody are mediated by Fcγ receptors (FcγRs), which are found on most immune cells, including dendritic cells (DCs). DCs are important antigen presenting cells and play a central role in inducing antigen-specific tolerance or immunity1,2. Following antigen acquisition in peripheral tissues, DCs migrate to draining lymph nodes via lymphatics to present antigen to T cells. In this study we demonstrate that FcγR engagement by IgG immune complexes (IC) stimulates DC migration from peripheral tissues to the paracortex of draining lymph nodes. In vitro, IC-stimulated murine and human DCs showed enhanced directional migration in a CCL19 gradient and increased CCR7 expression. Using intravital two-photon microscopy, we observed that local administration of IC resulted in dermal DC mobilisation. We confirmed that dermal DC migration to lymph nodes was CCR7-dependent and increased in the absence of the inhibitory receptor, FcγRIIb. These observations have relevance to autoimmunity, because autoantibody-containing serum from mice and humans with SLE also increased dermal DC migration to lymph nodes in vivo, suggesting that this process may occur in lupus, potentially driving the inappropriate localisation of autoantigen-bearing DCs. PMID:25384086

  20. DNA Replication and Cell Cycle Progression Regulatedby Long Range Interaction between Protein Complexes bound to DNA.

    Science.gov (United States)

    Matsson, L

    2001-12-01

    A nonstationary interaction that controlsDNA replication and the cell cycle isderived from many-body physics in achemically open T cell. The model predictsa long range force F'(ξ) =- (κ/2) ξ(1 - ξ)(2 - ξ)between thepre-replication complexes (pre-RCs) boundby the origins in DNA, ξ = ϕ/N being the relativedisplacement of pre-RCs, ϕ the number of pre-RCs, N the number of replicons to be replicated,and κ the compressibilitymodulus in the lattice of pre-RCs whichbehaves dynamically like an elasticallybraced string. Initiation of DNAreplication is induced at the thresholdϕ = N by a switch ofsign of F''(ξ), fromattraction (-) and assembly in the G(1) phase (0force at ϕ = 2N, from repulsion inS phase back to attraction in G(2), when all primed replicons havebeen duplicated once. F'(0) = 0corresponds to a resting cell in theabsence of driving force at ϕ= 0. The model thus ensures that the DNAcontent in G(2) cells is exactlytwice that of G(1) cells. The switch of interaction at the R-point, at which N pre-RCs have been assembled, starts the release of Rb protein thus also explaining the shift in the Rb phosphorylation from mitogen-dependent cyclinD to mitogen-independent cyclin E.Shape,slope and scale of the response curvesderived agree well with experimental datafrom dividing T cells and polymerising MTs,the variable length of which is due to anonlinear dependence of the growthamplitude on the initial concentrations oftubulin dimers and guanosine-tri-phosphate(GTP). The model also explains the dynamic instabilityin growing MTs.

  1. Interactions of opsonized immune complexes with whole blood cells: binding to erythrocytes restricts complex uptake by leucocyte populations

    DEFF Research Database (Denmark)

    Nielsen, C H; Svehag, S E; Marquart, H V

    1994-01-01

    binding, the main contributors being B cells. E initially inhibited and then later enhanced the IC binding to lymphocytes, suggesting that E promote B cell uptake of C3d,g-covered IC via CR2. Our findings, that E can restrict the IC uptake by circulating leucocytes, and that an IC-induced degranulation...

  2. 25 Years of Tension over Actin Binding to the Cadherin Cell Adhesion Complex: The Devil is in the Details.

    Science.gov (United States)

    Nelson, W James; Weis, William I

    2016-07-01

    Over the past 25 years, there has been a conceptual (re)evolution in understanding how the cadherin cell adhesion complex, which contains F-actin-binding proteins, binds to the actin cytoskeleton. There is now good synergy between structural, biochemical, and cell biological results that the cadherin-catenin complex binds to F-actin under force. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Shared fine specificity between T-cell receptors and an antibody recognizing a peptide/major histocompatibility class I complex

    DEFF Research Database (Denmark)

    Stryhn, A; Andersen, P S; Pedersen, L O

    1996-01-01

    Cytotoxic T cells recognize mosaic structures consisting of target peptides embedded within self-major histocompatibility complex (MHC) class I molecules. This structure has been described in great detail for several peptide-MHC complexes. In contrast, how T-cell receptors recognize peptide...... each other showing that peptide residues 1, 3, 4, 6, and 7 were exposed on the MHC surface and recognized by the T cells. Thus, the majority, and perhaps all, of the side chains of the non-primary anchor residues may be available for T-cell recognition, and contribute to the stringent specificity of T...... cells. A striking similarity between the specificity of the T cells and that of the pSAN antibody was found and most of the peptide residues, which could be recognized by the T cells, could also be recognized by the antibody....

  4. Microencapsulated cells of Lactobacillus paracasei subsp. paracasei in biopolymer complex coacervates and their function in a yogurt matrix.

    Science.gov (United States)

    Bosnea, L A; Moschakis, T; Biliaderis, C G

    2017-02-22

    L. paracasei subsp. paracasei E6 cells were encapsulated by complex coacervation using whey protein isolate (WPI) and gum arabic and introduced in stirred yogurts after fermentation. For comparison purposes, yogurts without addition of L. paracasei and yogurts with free cells of L. paracasei were produced. The survival of free and microencapsulated L. paracasei cells was evaluated during storage of the yogurts for 45 days at 4 °C. In addition, yogurts were exposed to simulated gastric juice and the reduction in viable numbers of L. paracasei cells was assessed. The effect of complex coacervates' addition on the rheological properties of yogurts was also evaluated. Yogurts containing encapsulated L. paracasei cells showed a slightly improved cell survival (≤0.22 log CFU g -1 reduction) during storage when compared to yogurts containing free cells (≤0.64 log CFU g -1 reduction). Moreover, the microencapsulated L. paracasei cells exhibited greater survival compared to free cells upon exposure of the yogurt samples to simulated gastric juice (pH 2.0) for 3 h. Finally, the incorporation of complex coacervates did not significantly affect the rheological properties of yogurts especially when added at concentrations less than 10% w/w. Consequently, the inclusion of microencapsulated bacteria by complex coacervation in yogurts, could become an effective vehicle for successful delivery of probiotics to the gut, and hence contributing to the improvement of the gastrointestinal tract health, without altering the texture of the product.

  5. DNA fragmentation and cell death mediated by T cell antigen receptor/CD3 complex on a leukemia T cell line.

    Science.gov (United States)

    Takahashi, S; Maecker, H T; Levy, R

    1989-10-01

    An anti-T cell receptor (TcR) monoclonal antibody (mAb), LC4, directed against a human leukemic T cell line, SUP-T13, caused DNA fragmentation ("apoptosis") and cell death upon binding to this cell line. Cross-linking of receptor molecules was necessary for this effect since F(ab')2, but not Fab', fragments of LC4 could induce cell death. Five anti-CD3 mAb tested also caused apoptosis, but only when they were presented on a solid phase. Interestingly, soluble anti-CD3 mAb induced calcium flux and had an additive effect on the calcium flux and interleukin 2 receptor expression induced by LC4, but these anti-CD3 mAb reversed the growth inhibition and apoptosis caused by LC4. The calcium ionophore A23187, but not the protein kinase C activator phorbol 12-myristate 13-acetate (PMA), also induced apoptosis, suggesting that protein kinase C activation alone does not cause apoptosis, although PMA is growth inhibitory. These results suggest that two distinct biological phenomena can accompany stimulation of the TcR/CD3 complex. In both cases, calcium flux and interleukin 2 receptor expression is induced, but only in one case is apoptosis and cell death seen. The signal initiating apoptosis can be selectively prevented by binding CD3 portion of the receptor in this cell line. This difference in signals mediated by the TcR/CD3 complex may be important in explaining the process of thymic selection, as well as in choosing anti-TcR mAb for therapeutic use.

  6. Autoantibodies in autoimmune thyroid disease promote immune complex formation with self antigens and increase B cell and CD4+ T cell proliferation in response to self antigens

    DEFF Research Database (Denmark)

    Nielsen, Claus Henrik; Hegedüs, Laszlo; Leslie, Robert Graham Quinton

    2004-01-01

    B cells are centrally involved as antigen-presenting cells in certain autoimmune diseases. To establish whether autoantibodies form immune complexes (IC) with self-antigens in autoimmune thyroid disease (AITD) and promote B cell uptake of self-antigen, sera from patients with Hashimoto......'s thyroiditis (HT), Graves' disease (GD) and healthy controls were incubated with human thyroglobulin (Tg) before adding normal peripheral blood mononuclear cells. The deposition of immunoglobulins and C3 fragments on B cells was then assessed. Inclusion of Tg in serum from HT patients promoted B cell capture...

  7. Autoantibodies in autoimmune thyroid disease promote immune complex formation with self antigens and increase B cell and CD4+ T cell proliferation in response to self antigens

    DEFF Research Database (Denmark)

    Nielsen, Claus Henrik; Hegedüs, Laszlo; Leslie, Robert Graham Quinton

    2004-01-01

    's thyroiditis (HT), Graves' disease (GD) and healthy controls were incubated with human thyroglobulin (Tg) before adding normal peripheral blood mononuclear cells. The deposition of immunoglobulins and C3 fragments on B cells was then assessed. Inclusion of Tg in serum from HT patients promoted B cell capture......B cells are centrally involved as antigen-presenting cells in certain autoimmune diseases. To establish whether autoantibodies form immune complexes (IC) with self-antigens in autoimmune thyroid disease (AITD) and promote B cell uptake of self-antigen, sera from patients with Hashimoto...

  8. Versatile polyion complex micelles for peptide and siRNA vectorization to engineer tolerogenic dendritic cells.

    Science.gov (United States)

    Mebarek, Naila; Vicente, Rita; Aubert-Pouëssel, Anne; Quentin, Julie; Mausset-Bonnefont, Anne-Laure; Devoisselle, Jean-Marie; Jorgensen, Christian; Bégu, Sylvie; Louis-Plence, Pascale

    2015-05-01

    Dendritic cells (DCs) are professional antigen-presenting cells that play a critical role in maintaining the balance between immunity and tolerance and, as such are a promising immunotherapy tool to induce immunity or to restore tolerance. The main challenge to harness the tolerogenic properties of DCs is to preserve their immature phenotype. We recently developed polyion complex micelles, formulated with double hydrophilic block copolymers of poly(methacrylic acid) and poly(ethylene oxide) blocks and able to entrap therapeutic molecules, which did not induce DC maturation. In the current study, the intrinsic destabilizing membrane properties of the polymers were used to optimize endosomal escape property of the micelles in order to propose various strategies to restore tolerance. On the first hand, we showed that high molecular weight (Mw) copolymer-based micelles were efficient to favor the release of the micelle-entrapped peptide into the endosomes, and thus to improve peptide presentation by immature (i) DCs. On the second hand, we put in evidence that low Mw copolymer-based micelles were able to favor the cytosolic release of micelle-entrapped small interfering RNAs, dampening the DCs immunogenicity. Therefore, we demonstrate the versatile use of polyionic complex micelles to preserve tolerogenic properties of DCs. Altogether, our results underscored the potential of such micelle-loaded iDCs as a therapeutic tool to restore tolerance in autoimmune diseases. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Protein-carbohydrate complex reveals circulating metastatic cells in a microfluidic assay

    KAUST Repository

    Simone, Giuseppina

    2013-02-11

    Advances in carbohydrate sequencing technologies reveal the tremendous complexity of the glycome and the role that glycomics might have to bring insight into the biological functions. Carbohydrate-protein interactions, in particular, are known to be crucial to most mammalian physiological processes as mediators of cell adhesion and metastasis, signal transducers, and organizers of protein interactions. An assay is developed here to mimic the multivalency of biological complexes that selectively and sensitively detect carbohydrate-protein interactions. The binding of β-galactosides and galectin-3 - a protein that is correlated to the progress of tumor and metastasis - is examined. The efficiency of the assay is related to the expression of the receptor while anchoring to the interaction\\'s strength. Comparative binding experiments reveal molecular binding preferences. This study establishes that the assay is robust to isolate metastatic cells from colon affected patients and paves the way to personalized medicine. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. MUC1-C activates polycomb repressive complexes and downregulates tumor suppressor genes in human cancer cells.

    Science.gov (United States)

    Rajabi, Hasan; Hiraki, Masayuki; Kufe, Donald

    2018-04-01

    The PRC2 and PRC1 complexes are aberrantly expressed in human cancers and have been linked to decreases in patient survival. MUC1-C is an oncoprotein that is also overexpressed in diverse human cancers and is associated with a poor prognosis. Recent studies have supported a previously unreported function for MUC1-C in activating PRC2 and PRC1 in cancer cells. In the regulation of PRC2, MUC1-C (i) drives transcription of the EZH2 gene, (ii) binds directly to EZH2, and (iii) enhances occupancy of EZH2 on target gene promoters with an increase in H3K27 trimethylation. Regarding PRC1, which is recruited to PRC2 sites in the hierarchical model, MUC1-C induces BMI1 transcription, forms a complex with BMI1, and promotes H2A ubiquitylation. MUC1-C thereby contributes to the integration of PRC2 and PRC1-mediated repression of tumor suppressor genes, such as CDH1, CDKN2A, PTEN and BRCA1. Like PRC2 and PRC1, MUC1-C is associated with the epithelial-mesenchymal transition (EMT) program, cancer stem cell (CSC) state, and acquisition of anticancer drug resistance. In concert with these observations, targeting MUC1-C downregulates EZH2 and BMI1, inhibits EMT and the CSC state, and reverses drug resistance. These findings emphasize the significance of MUC1-C as a therapeutic target for inhibiting aberrant PRC function and reprogramming the epigenome in human cancers.

  11. Protein-carbohydrate complex reveals circulating metastatic cells in a microfluidic assay

    KAUST Repository

    Simone, Giuseppina; Malara, Natalia Maria; Trunzo, Valentina; Perozziello, Gerardo; Neužil, Pavel; Francardi, Marco; Roveda, Laura; Renne, Maria; Prati, Ubaldo; Mollace, Vincenzo; Manz, Andreas; Di Fabrizio, Enzo M.

    2013-01-01

    Advances in carbohydrate sequencing technologies reveal the tremendous complexity of the glycome and the role that glycomics might have to bring insight into the biological functions. Carbohydrate-protein interactions, in particular, are known to be crucial to most mammalian physiological processes as mediators of cell adhesion and metastasis, signal transducers, and organizers of protein interactions. An assay is developed here to mimic the multivalency of biological complexes that selectively and sensitively detect carbohydrate-protein interactions. The binding of β-galactosides and galectin-3 - a protein that is correlated to the progress of tumor and metastasis - is examined. The efficiency of the assay is related to the expression of the receptor while anchoring to the interaction's strength. Comparative binding experiments reveal molecular binding preferences. This study establishes that the assay is robust to isolate metastatic cells from colon affected patients and paves the way to personalized medicine. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Copper Complexes with Tetradentate Ligands for Enhanced Charge Transport in Dye-Sensitized Solar Cells

    Directory of Open Access Journals (Sweden)

    Hannes Michaels

    2018-05-01

    Full Text Available In dye-sensitized solar cells (DSCs, the redox mediator is responsible for the regeneration of the oxidized dye and for the hole transport towards the cathode. Here, we introduce new copper complexes with tetradentate 6,6′-bis(4-(S-isopropyl-2-oxazolinyl-2,2′-bipyridine ligands, Cu(oxabpy, as redox mediators. Copper coordination complexes with a square-planar geometry show low reorganization energies and thus introduce smaller losses in photovoltage. Slow recombination kinetics of excited electrons between the TiO2 and CuII(oxabpy species lead to an exceptionally long electron lifetime, a high Fermi level in the TiO2, and a high photovoltage of 920 mV with photocurrents of 10 mA∙cm−2 and 6.2% power conversion efficiency. Meanwhile, a large driving force remains for the dye regeneration of the Y123 dye with high efficiencies. The square-planar Cu(oxabpy complexes yield viscous gel-like solutions. The unique charge transport characteristics are attributed to a superposition of diffusion and electronic conduction. An enhancement in charge transport performance of 70% despite the higher viscosity is observed upon comparison of Cu(oxabpy to the previously reported Cu(tmby2 redox electrolyte.

  13. A New Method to Develop Human Dental Pulp Cells and Platelet-rich Fibrin Complex.

    Science.gov (United States)

    He, Xuan; Chen, Wen-Xia; Ban, Guifei; Wei, Wei; Zhou, Jun; Chen, Wen-Jin; Li, Xian-Yu

    2016-11-01

    Platelet-rich fibrin (PRF) has been used as a scaffold material in various tissue regeneration studies. In the previous methods to combine seed cells with PRF, the structure of PRF was damaged, and the manipulation time in vitro was also increased. The objective of this in vitro study was to explore an appropriate method to develop a PRF-human dental pulp cell (hDPC) complex to maintain PRF structure integrity and to find out the most efficient part of PRF. The PRF-hDPC complex was developed at 3 different time points during PRF preparation: (1) the before centrifugation (BC) group, the hDPC suspension was added to the venous blood before blood centrifugation; (2) the immediately after centrifugation (IAC) group, the hDPC suspension was added immediately after blood centrifugation; (3) the after centrifugation (AC) group, the hDPC suspension was added 10 minutes after blood centrifugation; and (4) the control group, PRF without hDPC suspension. The prepared PRF-hDPC complexes were cultured for 7 days. The samples were fixed for histologic, immunohistochemistry, and scanning electron microscopic evaluation. Real-time polymerase chain reaction was performed to evaluate messenger RNA expression of alkaline phosphatase and dentin sialophosphoprotein. Enzyme-linked immunosorbent assay quantification for growth factors was performed within the different parts of the PRF. Histologic, immunohistochemistry, and scanning electron microscopic results revealed that hDPCs were only found in the BC group and exhibited favorable proliferation. Real-time polymerase chain reaction revealed that alkaline phosphatase and dentin sialophosphoprotein expression increased in the cultured PRF-hDPC complex. The lower part of the PRF released the maximum quantity of growth factors. Our new method to develop a PRF-hDPCs complex maintained PRF structure integrity. The hDPCs were distributed in the buffy coat, which might be the most efficient part of PRF. Copyright © 2016 American

  14. Forced-rupture of cell-adhesion complexes reveals abrupt switch between two brittle states

    Science.gov (United States)

    Toan, Ngo Minh; Thirumalai, D.

    2018-03-01

    Cell adhesion complexes (CACs), which are activated by ligand binding, play key roles in many cellular functions ranging from cell cycle regulation to mediation of cell extracellular matrix adhesion. Inspired by single molecule pulling experiments using atomic force spectroscopy on leukocyte function-associated antigen-1 (LFA-1), expressed in T-cells, bound to intercellular adhesion molecules (ICAM), we performed constant loading rate (rf) and constant force (F) simulations using the self-organized polymer model to describe the mechanism of ligand rupture from CACs. The simulations reproduce the major experimental finding on the kinetics of the rupture process, namely, the dependence of the most probable rupture forces (f*s) on ln rf (rf is the loading rate) exhibits two distinct linear regimes. The first, at low rf, has a shallow slope, whereas the slope at high rf is much larger, especially for a LFA-1/ICAM-1 complex with the transition between the two occurring over a narrow rf range. Locations of the two transition states (TSs) extracted from the simulations show an abrupt change from a high value at low rf or constant force, F, to a low value at high rf or F. This unusual behavior in which the CACs switch from one brittle (TS position is a constant over a range of forces) state to another brittle state is not found in forced-rupture in other protein complexes. We explain this novel behavior by constructing the free energy profiles, F(Λ)s, as a function of a collective reaction coordinate (Λ), involving many key charged residues and a critical metal ion (Mg2+). The TS positions in F(Λ), which quantitatively agree with the parameters extracted using the Bell-Evans model, change abruptly at a critical force, demonstrating that it, rather than the molecular extension, is a good reaction coordinate. Our combined analyses using simulations performed in both the pulling modes (constant rf and F) reveal a new mechanism for the two loading regimes observed in the

  15. Presynaptic dystroglycan-pikachurin complex regulates the proper synaptic connection between retinal photoreceptor and bipolar cells.

    Science.gov (United States)

    Omori, Yoshihiro; Araki, Fumiyuki; Chaya, Taro; Kajimura, Naoko; Irie, Shoichi; Terada, Koji; Muranishi, Yuki; Tsujii, Toshinori; Ueno, Shinji; Koyasu, Toshiyuki; Tamaki, Yasuhiro; Kondo, Mineo; Amano, Shiro; Furukawa, Takahisa

    2012-05-02

    Dystroglycan (DG) is a key component of the dystrophin-glycoprotein complex (DGC) at the neuromuscular junction postsynapse. In the mouse retina, the DGC is localized at the presynapse of photoreceptor cells, however, the function of presynaptic DGC is poorly understood. Here, we developed and analyzed retinal photoreceptor-specific DG conditional knock-out (DG CKO) mice. We found that the DG CKO retina showed a reduced amplitude and a prolonged implicit time of the ERG b-wave. Electron microscopic analysis revealed that bipolar dendrite invagination into the photoreceptor terminus is perturbed in the DG CKO retina. In the DG CKO retina, pikachurin, a DG ligand in the retina, is markedly decreased at photoreceptor synapses. Interestingly, in the Pikachurin(-/-) retina, the DG signal at the ribbon synaptic terminus was severely reduced, suggesting that pikachurin is required for the presynaptic accumulation of DG at the photoreceptor synaptic terminus, and conversely DG is required for pikachurin accumulation. Furthermore, we found that overexpression of pikachurin induces formation and clustering of a DG-pikachurin complex on the cell surface. The Laminin G repeats of pikachurin, which are critical for its oligomerization and interaction with DG, were essential for the clustering of the DG-pikachurin complex as well. These results suggest that oligomerization of pikachurin and its interaction with DG causes DG assembly on the synapse surface of the photoreceptor synaptic terminals. Our results reveal that the presynaptic interaction of pikachurin with DG at photoreceptor terminals is essential for both the formation of proper photoreceptor ribbon synaptic structures and normal retinal electrophysiology.

  16. The GARP/Latent TGF-β1 complex on Treg cells modulates the induction of peripherally derived Treg cells during oral tolerance.

    Science.gov (United States)

    Edwards, Justin P; Hand, Timothy W; Morais da Fonseca, Denise; Glass, Deborah D; Belkaid, Yasmine; Shevach, Ethan M

    2016-06-01

    Treg cells can secrete latent TGF-β1 (LTGF-β1), but can also utilize an alternative pathway for transport and expression of LTGF-β1 on the cell surface in which LTGF-β1 is coupled to a distinct LTGF-β binding protein termed glycoprotein A repetitions predominant (GARP)/LRRC32. The function of the GARP/LTGF-β1 complex has remained elusive. Here, we examine in vivo the roles of GARP and TGF-β1 in the induction of oral tolerance. When Foxp3(-) OT-II T cells were transferred to wild-type recipient mice followed by OVA feeding, the conversion of Foxp3(-) to Foxp3(+) OT-II cells was dependent on recipient Treg cells. Neutralization of IL-2 in the recipient mice also abrogated this conversion. The GARP/LTGF-β1 complex on recipient Treg cells, but not dendritic cell-derived TGF-β1, was required for efficient induction of Foxp3(+) T cells and for the suppression of delayed hypersensitivity. Expression of the integrin αvβ8 by Treg cells (or T cells) in the recipients was dispensable for induction of Foxp3 expression. Transient depletion of the bacterial flora enhanced the development of oral tolerance by expanding Treg cells with enhanced expression of the GARP/LTGF-β1 complex. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  17. Spatio-temporal diversification of the cell wall matrix materials in the developing stomatal complexes of Zea mays.

    Science.gov (United States)

    Giannoutsou, E; Apostolakos, P; Galatis, B

    2016-11-01

    The matrix cell wall materials, in developing Zea mays stomatal complexes are asymmetrically distributed, a phenomenon appearing related to the local cell wall expansion and deformation, the establishment of cell polarity, and determination of the cell division plane. In cells of developing Zea mays stomatal complexes, definite cell wall regions expand determinately and become locally deformed. This differential cell wall behavior is obvious in the guard cell mother cells (GMCs) and the subsidiary cell mother cells (SMCs) that locally protrude towards the adjacent GMCs. The latter, emitting a morphogenetic stimulus, induce polarization/asymmetrical division in SMCs. Examination of immunolabeled specimens revealed that homogalacturonans (HGAs) with a high degree of de-esterification (2F4- and JIM5-HGA epitopes) and arabinogalactan proteins are selectively distributed in the extending and deformed cell wall regions, while their margins are enriched with rhamnogalacturonans (RGAs) containing highly branched arabinans (LM6-RGA epitope). In SMCs, the local cell wall matrix differentiation constitutes the first structural event, indicating the establishment of cell polarity. Moreover, in the premitotic GMCs and SMCs, non-esterified HGAs (2F4-HGA epitope) are preferentially localized in the cell wall areas outlining the cytoplasm where the preprophase band is formed. In these areas, the forthcoming cell plate fuses with the parent cell walls. These data suggest that the described heterogeneity in matrix cell wall materials is probably involved in: (a) local cell wall expansion and deformation, (b) the transduction of the inductive GMC stimulus, and (c) the determination of the division plane in GMCs and SMCs.

  18. [Progression of nerve fiber layer defects in retrobulbar optic neuritis by the macular ganglion cell complex].

    Science.gov (United States)

    Hong, D; Bosc, C; Chiambaretta, F

    2017-11-01

    Recent studies with SD OCT had shown early axonal damage to the macular ganglion cell complex (which consists of the three innermost layers of the retina: Inner Plexiform Layer [IPL], Ganglion Cell Layer [GCL], Retinal Nerve Fibre layer [RNFL]) in optic nerve pathology. Retrobulbar optic neuritis (RBON), occurring frequently in demyelinating diseases, leads to atrophy of the optic nerve fibers at the level of the ganglion cell axons, previously described in the literature. The goal of this study is to evaluate the progression of optic nerve fiber defects and macular ganglion cell complex defects with the SPECTRALIS OCT via a reproducible method by calculating a mean thickness in each quadrant after an episode of retrobulbar optic neuritis. This is a prospective monocentric observational study including 8 patients at the Clermont-Ferrand university medical center. All patients underwent ocular examination with macular and disc OCT analysis and a Goldmann visual field at the time of inclusion (onset or recurrence of RBON), at 3 months and at 6 months. Patients were 40-years-old on average at the time of inclusion. After 6 months of follow-up, there was progression of the atrophy of the macular ganglion cell complex in the affected eye on (11.5% or 11μm) predominantly inferonasally (13.9% or 16μm) and superonasally (12.9% or 14μm) while the other eye remained stable. The decrease in thickness occurred mainly in the most internal 3 layers of the retina. On average, the loss in thickness of the peripapillary RNFL was predominantly inferotemporal (24.9% or 39μm) and superotemporal (21.8% or 28μm). In 3 months of progression, the loss of optic nerve fibers is already seen on macular and disc OCT after an episode of RBON, especially in inferior quadrants in spite of the improvement in the Goldmann visual field and visual acuity. Segmentation by quadrant was used here to compare the progression of the defect by region compared to the fovea in a global and reproducible

  19. Progesterone from the cumulus cells is the sperm chemoattractant secreted by the rabbit oocyte cumulus complex.

    Directory of Open Access Journals (Sweden)

    Héctor Alejandro Guidobaldi

    Full Text Available Sperm chemotaxis in mammals have been identified towards several female sources as follicular fluid (FF, oviduct fluid, and conditioned medium from the cumulus oophorus (CU and the oocyte (O. Though several substances were confirmed as sperm chemoattractant, Progesterone (P seems to be the best chemoattractant candidate, because: 1 spermatozoa express a cell surface P receptor, 2 capacitated spermatozoa are chemotactically attracted in vitro by gradients of low quantities of P; 3 the CU cells produce and secrete P after ovulation; 4 a gradient of P may be kept stable along the CU; and 5 the most probable site for sperm chemotaxis in vivo could be near and/or inside the CU. The aim of this study was to verify whether P is the sperm chemoattractant secreted by the rabbit oocyte-cumulus complex (OCC in the rabbit, as a mammalian animal model. By means of videomicroscopy and computer image analysis we observed that only the CU are a stable source of sperm attractants. The CU produce and secrete P since the hormone was localized inside these cells by immunocytochemistry and in the conditioned medium by enzyme immunoassay. In addition, rabbit spermatozoa express a cell surface P receptor detected by western blot and localized over the acrosomal region by immunocytochemistry. To confirm that P is the sperm chemoattractant secreted by the CU, the sperm chemotactic response towards the OCC conditioned medium was inhibited by three different approaches: P from the OCC conditioned medium was removed with an anti-P antibody, the attractant gradient of the OCC conditioned medium was disrupted by a P counter gradient, and the sperm P receptor was blocked with a specific antibody. We concluded that only the CU but not the oocyte secretes P, and the latter chemoattract spermatozoa by means of a cell surface receptor. Our findings may be of interest in assisted reproduction procedures in humans, animals of economic importance and endangered species.

  20. 'Special K' and a Loss of Cell-To-Cell Adhesion in Proximal Tubule-Derived Epithelial Cells: Modulation of the Adherens Junction Complex by Ketamine

    Science.gov (United States)

    Hills, Claire E.; Jin, Tianrong; Siamantouras, Eleftherios; Liu, Issac K-K; Jefferson, Kieran P.; Squires, Paul E.

    2013-01-01

    Ketamine, a mild hallucinogenic class C drug, is the fastest growing ‘party drug’ used by 16–24 year olds in the UK. As the recreational use of Ketamine increases we are beginning to see the signs of major renal and bladder complications. To date however, we know nothing of a role for Ketamine in modulating both structure and function of the human renal proximal tubule. In the current study we have used an established model cell line for human epithelial cells of the proximal tubule (HK2) to demonstrate that Ketamine evokes early changes in expression of proteins central to the adherens junction complex. Furthermore we use AFM single-cell force spectroscopy to assess if these changes functionally uncouple cells of the proximal tubule ahead of any overt loss in epithelial cell function. Our data suggests that Ketamine (24–48 hrs) produces gross changes in cell morphology and cytoskeletal architecture towards a fibrotic phenotype. These physical changes matched the concentration-dependent (0.1–1 mg/mL) cytotoxic effect of Ketamine and reflect a loss in expression of the key adherens junction proteins epithelial (E)- and neural (N)-cadherin and β-catenin. Down-regulation of protein expression does not involve the pro-fibrotic cytokine TGFβ, nor is it regulated by the usual increase in expression of Slug or Snail, the transcriptional regulators for E-cadherin. However, the loss in E-cadherin can be partially rescued pharmacologically by blocking p38 MAPK using SB203580. These data provide compelling evidence that Ketamine alters epithelial cell-to-cell adhesion and cell-coupling in the proximal kidney via a non-classical pro-fibrotic mechanism and the data provides the first indication that this illicit substance can have major implications on renal function. Understanding Ketamine-induced renal pathology may identify targets for future therapeutic intervention. PMID:24009666

  1. Examining Rhodium Catalyst complexes for Use with Conducting Polymers Designed for Fuel Cells in Preparing Biosensors

    Energy Technology Data Exchange (ETDEWEB)

    Carpio, M.M.; Kerr, J.B.

    2005-01-01

    Biosensing devices are important because they can detect, record, and transmit information regarding the presence of, or physiological changes in, different chemical or biological materials in the environment. The goal of this research is to prepare a biosensing device that is effective, quick, and low cost. This is done by examining which chemicals will work best when placed in a biosensor. The first study involved experimenting on a rhodium catalyst complexed with ligands such as bipyridine and imidazole. The rhodium catalyst is important because it is reduced from RhIII to RhI, forms a hydride by reaction with water and releases the hydride to react with nicotinamide adenine dinucleotide (NAD+) to selectively produce 1,4-NADH, the reduced form of NAD+. The second study looked at different types of ketones and enzymes for the enzyme-substrate reaction converting a ketone into an alcohol. Preliminary results showed that the rhodium complexed with bipyridine was able to carry out all the reactions, while the rhodium complexed with imidazole was not able to produce and release hydrides. In addition, the most effective ketone to use is benzylacetone with the enzyme alcohol dehydrogenase from baker’s yeast. Future work includes experimenting with bis-imidazole, which mimics the structure of bipyridine to see if it has the capability to reduce and if the reduction rate is comparable to the bipyridine complex. Once all testing is completed, the fastest catalysts will be combined with polymer membranes designed for fuel cells to prepare biosensing devices that can be used in a variety of applications including ones in the medical and environmental fields.

  2. Complex I and complex III inhibition specifically increase cytosolic hydrogen peroxide levels without inducing oxidative stress in HEK293 cells

    NARCIS (Netherlands)

    Forkink, M.; Basit, F.; Teixeira, J.; Swarts, H.G.; Koopman, W.J.H.; Willems, P.H.G.M.

    2015-01-01

    Inhibitor studies with isolated mitochondria demonstrated that complex I (CI) and III (CIII) of the electron transport chain (ETC) can act as relevant sources of mitochondrial reactive oxygen species (ROS). Here we studied ROS generation and oxidative stress induction during chronic (24h) inhibition

  3. Translocation of differently sized and charged polystyrene nanoparticles in in vitro intestinal cell models of increasing complexity

    NARCIS (Netherlands)

    Walczak, A.P.; Kramer, E.; Hendriksen, P.J.M.; Tromp, P.; Helsper, J.P.F.G.; Zande, M. van der; Rietjens, I.M.C.M.; Bouwmeester, H.

    2015-01-01

    Intestinal translocation is a key factor for determining bioavailability of nanoparticles (NPs) after oral uptake. Therefore, we evaluated three in vitro intestinal cell models of increasing complexity which might affect the translocation of NPs: a mono-culture (Caco-2 cells), a co-culture with

  4. T cell responses affected by aminopeptidase N (CD13)-mediated trimming of major histocompatibility complex class II-bound peptides

    DEFF Research Database (Denmark)

    Larsen, S L; Pedersen, L O; Buus, S

    1996-01-01

    Endocytosed protein antigens are believed to be fragmented in what appears to be a balance between proteolysis and MHC-mediated epitope protection, and the resulting peptide-MHC complexes are transported to the surface of the antigen-presenting cells (APC) and presented to T cells. The events tha...

  5. Structural studies of complex carbohydrates of plant cell walls. Progress report, June 15, 1992--June 14, 1993

    Energy Technology Data Exchange (ETDEWEB)

    Darvill, A.G.

    1994-10-01

    This report contains the abstracts of fourteen papers published, in press, or in preparation reporting on research activities to investigate the structure, as well as the function of cell walls in plants. This document also contains research on methods to determine the structure of complex carbohydrates of the cell walls.

  6. Modeling the Effect of APC Truncation on Destruction Complex Function in Colorectal Cancer Cells

    Science.gov (United States)

    Barua, Dipak; Hlavacek, William S.

    2013-01-01

    In colorectal cancer cells, APC, a tumor suppressor protein, is commonly expressed in truncated form. Truncation of APC is believed to disrupt degradation of β—catenin, which is regulated by a multiprotein complex called the destruction complex. The destruction complex comprises APC, Axin, β—catenin, serine/threonine kinases, and other proteins. The kinases and , which are recruited by Axin, mediate phosphorylation of β—catenin, which initiates its ubiquitination and proteosomal degradation. The mechanism of regulation of β—catenin degradation by the destruction complex and the role of truncation of APC in colorectal cancer are not entirely understood. Through formulation and analysis of a rule-based computational model, we investigated the regulation of β—catenin phosphorylation and degradation by APC and the effect of APC truncation on function of the destruction complex. The model integrates available mechanistic knowledge about site-specific interactions and phosphorylation of destruction complex components and is consistent with an array of published data. We find that the phosphorylated truncated form of APC can outcompete Axin for binding to β—catenin, provided that Axin is limiting, and thereby sequester β—catenin away from Axin and the Axin-recruited kinases and . Full-length APC also competes with Axin for binding to β—catenin; however, full-length APC is able, through its SAMP repeats, which bind Axin and which are missing in truncated oncogenic forms of APC, to bring β—catenin into indirect association with Axin and Axin-recruited kinases. Because our model indicates that the positive effects of truncated APC on β—catenin levels depend on phosphorylation of APC, at the first 20-amino acid repeat, and because phosphorylation of this site is mediated by , we suggest that is a potential target for therapeutic intervention in colorectal cancer. Specific inhibition of is predicted to limit binding of β—catenin to truncated

  7. Bacterial actin MreB assembles in complex with cell shape protein RodZ.

    Science.gov (United States)

    van den Ent, Fusinita; Johnson, Christopher M; Persons, Logan; de Boer, Piet; Löwe, Jan

    2010-03-17

    Bacterial actin homologue MreB is required for cell shape maintenance in most non-spherical bacteria, where it assembles into helical structures just underneath the cytoplasmic membrane. Proper assembly of the actin cytoskeleton requires RodZ, a conserved, bitopic membrane protein that colocalises to MreB and is essential for cell shape determination. Here, we present the first crystal structure of bacterial actin engaged with a natural partner and provide a clear functional significance of the interaction. We show that the cytoplasmic helix-turn-helix motif of Thermotoga maritima RodZ directly interacts with monomeric as well as filamentous MreB and present the crystal structure of the complex. In vitro and in vivo analyses of mutant T. maritima and Escherichia coli RodZ validate the structure and reveal the importance of the MreB-RodZ interaction in the ability of cells to propagate as rods. Furthermore, the results elucidate how the bacterial actin cytoskeleton might be anchored to the membrane to help constrain peptidoglycan synthesis in the periplasm.

  8. Spindle cell carcinoma of the breast as complex cystic lesion: a case report

    International Nuclear Information System (INIS)

    Kitada, Masahiro; Hayashi, Satoshi; Matsuda, Yoshinari; Ishibashi, Kei; Oikawa, Keisuke; Miyokawa, Naoyuki

    2014-01-01

    Spindle cell carcinoma of the breast is a rare tumor. This tumor can proliferate rapidly and cause cystic changes because of internal tissue necrosis. We evaluated a 54-year-old woman with right breast lump. Mammography showed a category four mass with a diameter of 2.5 cm. Ultrasonography (US) revealed a complex cystic lesion, and fine-needle aspiration (FNA) cytology demonstrated bloody fluid and malignant cells. Partial breast resection and sentinel lymph node biopsy were performed. Immunohistology revealed spindle cells with positive results for cytokeratin (AE1/AE3) and vimentin, partially positive results for s-100, and negative results for desmin and α-actin. The pathological stage was IIA, and biochemical characterization showed that the tumor was triple negative. Six courses of FEC-100 chemotherapy (5-fluorouracil 500 mg/m 2 , epirubicin 100 mg/m 2 , and cyclophosphamide 500 mg/m 2 ) were administered. Radiotherapy was performed. This case is discussed with reference to the literature

  9. Major histocompatibility complex-restricted self-recognition in responses to trinitrophenyl-Ficoll. A novel cell interaction pathway requiring self-recognition of accessory cell H-2 determinants by both T cells and B cells

    International Nuclear Information System (INIS)

    Hodes, R.J.; Hathcock, K.S.; Singer, A.

    1983-01-01

    In vitro primary antibody responses to limiting concentrations of trinitrophenyl (TNP)-Ficoll were shown to be T cell dependent, requiring the cooperation of T helper (TH) cells, B cells, and accessory cells. Under these conditions, TH cells derived from long-term radiation bone marrow chimeras were major histocompatibility complex (MHC) restricted in their ability to cooperate with accessory cells expressing host-type MHC determinants. The requirement for MHC-restricted self-recognition by TNP-Ficoll-reactive B cells was assessed under these T-dependent conditions. In the presence of competent TH cells, chimeric B cells were found to be MHC restricted, cooperating only with accessory cells that expressed host-type MHC products. In contrast, the soluble products of certain monoclonal T cell lines were able to directly activate B cells in response to TNP-Ficoll, bypassing any requirement for MHC-restricted self-recognition. These findings demonstrate the existence of a novel cell interaction pathway in which B cells as well as TH cells are each required to recognize self-MHC determinants on accessory cells, but are not required to recognize each other. They further demonstrate that the requirement for self-recognition by B cells may be bypassed in certain T-dependent activation pathways

  10. New iridium complex as additive to the spiro-OMeTAD in perovskite solar cells with enhanced stability

    Directory of Open Access Journals (Sweden)

    Laura Badia

    2014-08-01

    Full Text Available A new iridium complex, IrCp*Cl(PyPyz[TFSI], has been synthesized and used as additive for the hole transporter material, spiro-OMeTAD, in perovskite solar cells. The cells prepared with this Ir additive present higher efficiency than reference cells, and similar to cells prepared with Co additive. We have determined that the presence of metal complexes as additives decreases the recombination rate, as it has been observed by impedance spectroscopy. Very interestingly, while the efficiency after 3 months decreases by 22% and 70% for reference cell and cell with Co additive, respectively, the efficiency of devices containing the Ir additive is only decreased by a 4%.

  11. Chemical sporulation and germination: cytoprotective nanocoating of individual mammalian cells with a degradable tannic acid-FeIII complex

    Science.gov (United States)

    Lee, Juno; Cho, Hyeoncheol; Choi, Jinsu; Kim, Doyeon; Hong, Daewha; Park, Ji Hun; Yang, Sung Ho; Choi, Insung S.

    2015-11-01

    Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature.Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature. Electronic supplementary information (ESI) available: Experimental details, LSCM images, and SEM and TEM images. See DOI: 10.1039/c5nr05573c

  12. Cell cycle effects of L-sulforaphane, a major antioxidant from cruciferous vegetables: The role of the anaphase promoting complex.

    Science.gov (United States)

    Shelley, Zhaoping; Royce, Simon G; Ververis, Katherine; Karagiannis, Tom C

    2014-01-01

    L-sulforaphane (LSF) is a natural isothiocyanate found in cruciferous vegetables particularly broccoli. LSF has been identified as a potent antioxidant and anti-cancer agent and is widely known to regulate phase II detoxifying enzymes and induce cell cycle arrest or apoptosis in malignant cells in vitro and in vivo. Previous studies have found significant G2/M cell cycle arrest in response to LSF in various model of cancer and results have mainly been attributed to increased cyclin B1 protein levels and increased p21expression. Using genome-wide mRNA-Seq analysis we provide insights into the molecular mechanisms of action of LSF to identify a key pathway in cell cycle progression - the role of the anaphase promoting complex (APC) pathway. We evaluated gene expression changes in human erythroleukemic K562 cells following treatment with 15 μM LSF for 48h and compared them to immortalized human keratinocytes, human microvascular endothelial cells (HMEC-1) cells and normal human umbilical endothelial cells (HUVEC). We identified disparate gene expression changes in response to LSF between malignant and normal cells and immortalized cell lines. The results highlight significant down-regulation of kinase CDK1 which is suggestive that the existence and activity of APC/CDC20 complex will be inhibited along with its associated down-stream degradation of key cell cycle regulators preventing cell cycle progression from mitotic exit.

  13. Cellular interactions of a water-soluble supramolecular polymer complex of carbon nanotubes with human epithelial colorectal adenocarcinoma cells.

    Science.gov (United States)

    Lee, Yeonju; Geckeler, Kurt E

    2012-08-01

    Water-soluble, PAX-loaded carbon nanotubes are fabricated by employing a synthetic polyampholyte, PDM. To investigate the suitability of the polyampholyte and the nanotubes as drug carriers, different cellular interactions such as the human epithelial Caco-2 cells viability, their effect on the cell growth, and the change in the transepithelial electrical resistance in Caco-2 cells are studied. The resulting complex is found to exhibit an effective anti-cancer effect against colon cancer cells and an increased the reduction of the electrical resistance in the Caco-2 cells when compared to the precursor PAX. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Lymphocyte depletion in thymic nurse cells: a tool to identify in situ lympho-epithelial complexes having thymic nurse cell characteristics

    NARCIS (Netherlands)

    Leene, W.; de Waal Malefijt, R.; Roholl, P. J.; Hoeben, K. A.

    1988-01-01

    In situ pre-existing complexes of epithelial cells and thymocytes having thymic nurse cell characteristics were visualized in the murine thymus cortex using dexamethasone as a potent killer of cortisone-sensitive thymocytes. The degradation and subsequent depletion of cortisone-sensitive thymocytes

  15. Deposition of idiotype-anti-idiotype immune complexes in renal glomeruli after polyclonal B cell activation

    International Nuclear Information System (INIS)

    Goldman, M.; Rose, L.M.; Hochmann, A.; Lambert, P.H.

    1982-01-01

    We investigated the possible role of idiotypic interactions in the pathogenesis of the glomerular lesions observed in mice undergoing polyclonal B cell activation. BALB/c mice were studied for the presence of renal deposits of T15 idiotype-anti-T15 idiotype-immune complexes (IC) after injection of bacterial lipopolysaccharides (LPS). The T15 idiotype is the major idiotype of BALB/c mice anti-phosphorylcholine (PC) antibodies, which are cross-reactive with the idiotype of the TEPC-15 myeloma protein. This model was used because T15 idiotype-anti-T15 idiotype IC have been detected in the circulation of BALB/c mice after polyclonal B cell activation. First, an idiotype-specific immunofluorescence technique allowed us to detect T15 idiotype-bearing immunoglobulins in glomeruli from day 6 to day 28 after LPS injection. Second, fluorescein isothiocyanate-conjugated TEPC-15 myeloma protein was found to localize in the glomeruli after in vivo injection 18 d after LPS administration. This renal localization was shown to be idiotype-specific and could be quantified in a trace-labeling experiment. Third, kidney-deposited immunoglobulins of mice injected with LPS were eluted, radiolabeled, and analyzed by radioimmunoassay. Both T15 idiotype-bearing immunoglobulins and anti-T15 idiotype antibodies were detected in the eluates, providing further evidence for a renal deposition of T15 idiotype-anti-T15 idiotype IC. Polyclonal B cell activation is likely to result in a simultaneous triggering of many idiotypic clones and of corresponding anti-idiotypic clones represented in the B cell repertoire. This could lead to the formation of a variety of idiotype-anti-idiotype IC that could participate in the development of glomerular lesions

  16. Genomic binding profiles of functionally distinct RNA polymerase III transcription complexes in human cells.

    Science.gov (United States)

    Moqtaderi, Zarmik; Wang, Jie; Raha, Debasish; White, Robert J; Snyder, Michael; Weng, Zhiping; Struhl, Kevin

    2010-05-01

    Genome-wide occupancy profiles of five components of the RNA polymerase III (Pol III) machinery in human cells identified the expected tRNA and noncoding RNA targets and revealed many additional Pol III-associated loci, mostly near short interspersed elements (SINEs). Several genes are targets of an alternative transcription factor IIIB (TFIIIB) containing Brf2 instead of Brf1 and have extremely low levels of TFIIIC. Strikingly, expressed Pol III genes, unlike nonexpressed Pol III genes, are situated in regions with a pattern of histone modifications associated with functional Pol II promoters. TFIIIC alone associates with numerous ETC loci, via the B box or a novel motif. ETCs are often near CTCF binding sites, suggesting a potential role in chromosome organization. Our results suggest that human Pol III complexes associate preferentially with regions near functional Pol II promoters and that TFIIIC-mediated recruitment of TFIIIB is regulated in a locus-specific manner.

  17. Coping with complexity: machine learning optimization of cell-free protein synthesis.

    Science.gov (United States)

    Caschera, Filippo; Bedau, Mark A; Buchanan, Andrew; Cawse, James; de Lucrezia, Davide; Gazzola, Gianluca; Hanczyc, Martin M; Packard, Norman H

    2011-09-01

    Biological systems contain complex metabolic pathways with many nonlinearities and synergies that make them difficult to predict from first principles. Protein synthesis is a canonical example of such a pathway. Here we show how cell-free protein synthesis may be improved through a series of iterated high-throughput experiments guided by a machine-learning algorithm implementing a form of evolutionary design of experiments (Evo-DoE). The algorithm predicts fruitful experiments from statistical models of the previous experimental results, combined with stochastic exploration of the experimental space. The desired experimental response, or evolutionary fitness, was defined as the yield of the target product, and new experimental conditions were discovered to have ∼ 350% greater yield than the standard. An analysis of the best experimental conditions discovered indicates that there are two distinct classes of kinetics, thus showing how our evolutionary design of experiments is capable of significant innovation, as well as gradual improvement. Copyright © 2011 Wiley Periodicals, Inc.

  18. Scrapie affects the maturation cycle and immune complex trapping by follicular dendritic cells in mice.

    Directory of Open Access Journals (Sweden)

    Gillian McGovern

    2009-12-01

    Full Text Available Transmissible spongiform encephalopathies (TSEs or prion diseases are infectious neurological disorders of man and animals, characterised by abnormal disease-associated prion protein (PrP(d accumulations in the brain and lymphoreticular system (LRS. Prior to neuroinvasion, TSE agents often accumulate to high levels within the LRS, apparently without affecting immune function. However, our analysis of scrapie-affected sheep shows that PrP(d accumulations within the LRS are associated with morphological changes to follicular dendritic cells (FDCs and tingible body macrophages (TBMs. Here we examined FDCs and TBMs in the mesenteric lymph nodes (MLNs of scrapie-affected mice by light and electron microscopy. In MLNs from uninfected mice, FDCs could be morphologically categorised into immature, mature and regressing forms. However, in scrapie-affected MLNs this maturation cycle was adversely affected. FDCs characteristically trap and retain immune complexes on their surfaces, which they display to B-lymphocytes. In scrapie-affected MLNs, some FDCs were found where areas of normal and abnormal immune complex retention occurred side by side. The latter co-localised with PrP(d plasmalemmal accumulations. Our data suggest this previously unrecognised morphology represents the initial stage of an abnormal FDC maturation cycle. Alterations to the FDCs included PrP(d accumulation, abnormal cell membrane ubiquitin and excess immunoglobulin accumulation. Regressing FDCs, in contrast, appeared to lose their membrane-attached PrP(d. Together, these data suggest that TSE infection adversely affects the maturation and regression cycle of FDCs, and that PrP(d accumulation is causally linked to the abnormal pathology observed. We therefore support the hypothesis that TSEs cause an abnormality in immune function.

  19. Genetic recombination within the human T-cell receptor α-chain gene complex

    International Nuclear Information System (INIS)

    Robinson, M.A.; Kindt, T.J.

    1987-01-01

    Genetic analyses of the human T-cell receptor (TCR) α-chain genes indicate that recombination events may occur frequently within this gene complex. Examination of the inheritance of restriction fragment length polymorphisms (RFLP) detected by using probes for constant or variable region gene segments made it possible to assign TCRα haplotypes to the 16 parents and 43 offspring of eight families studied. A total of six RFLP, three for the constant region and three for variable region segments, were examined in the present studies. Most enzyme and probe combinations tested revealed no polymorphism and those finally selected for the study showed limited polymorphism in that only two or, in one case, three allelic forms of the gene were seen. In spite of limited variability at this level, extensive heterogeneity was observed for the combinations of markers present in haplotypes, suggesting that frequent recombination events have occurred. Most strikingly, multiple combinations of RFLP occurring in close proximity of the TCRα constant region gene were observed in this study. A high recombination frequency for the TCRα gene complex is further supported by the observation that two children, one in each of two families, inherited recombinant TCRα haplotypes

  20. Cadherin complexes recruit mRNAs and RISC to regulate epithelial cell signaling.

    Science.gov (United States)

    Kourtidis, Antonis; Necela, Brian; Lin, Wan-Hsin; Lu, Ruifeng; Feathers, Ryan W; Asmann, Yan W; Thompson, E Aubrey; Anastasiadis, Panos Z

    2017-10-02

    Cumulative evidence demonstrates that most RNAs exhibit specific subcellular distribution. However, the mechanisms regulating this phenomenon and its functional consequences are still under investigation. Here, we reveal that cadherin complexes at the apical zonula adherens (ZA) of epithelial adherens junctions recruit the core components of the RNA-induced silencing complex (RISC) Ago2, GW182, and PABPC1, as well as a set of 522 messenger RNAs (mRNAs) and 28 mature microRNAs (miRNAs or miRs), via PLEKHA7. Top canonical pathways represented by these mRNAs include Wnt/β-catenin, TGF-β, and stem cell signaling. We specifically demonstrate the presence and silencing of MYC, JUN, and SOX2 mRNAs by miR-24 and miR-200c at the ZA. PLEKHA7 knockdown dissociates RISC from the ZA, decreases loading of the ZA-associated mRNAs and miRNAs to Ago2, and results in a corresponding increase of MYC, JUN, and SOX2 protein expression. The present work reveals a mechanism that directly links junction integrity to the silencing of a set of mRNAs that critically affect epithelial homeostasis. © 2017 Kourtidis et al.

  1. Rapid and specific purification of Argonaute-small RNA complexes from crude cell lysates.

    Science.gov (United States)

    Flores-Jasso, C Fabián; Salomon, William E; Zamore, Phillip D

    2013-02-01

    Small interfering RNAs (siRNAs) direct Argonaute proteins, the core components of the RNA-induced silencing complex (RISC), to cleave complementary target RNAs. Here, we describe a method to purify active RISC containing a single, unique small RNA guide sequence. We begin by capturing RISC using a complementary 2'-O-methyl oligonucleotide tethered to beads. Unlike other methods that capture RISC but do not allow its recovery, our strategy purifies active, soluble RISC in good yield. The method takes advantage of the finding that RISC partially paired to a target through its siRNA guide dissociates more than 300 times faster than a fully paired siRNA in RISC. We use this strategy to purify fly Ago1- and Ago2-RISC, as well as mouse AGO2-RISC. The method can discriminate among RISCs programmed with different guide strands, making it possible to deplete and recover specific RISC populations. Endogenous microRNA:Argonaute complexes can also be purified from cell lysates. Our method scales readily and takes less than a day to complete.

  2. Comparison of two binuclear vanadium-catecholate complexes: Synthesis, X-ray structure and effects in cancer cells

    Science.gov (United States)

    Chi, Zixiang; Zhu, Linli; Lu, Xiaoming

    2011-08-01

    Two binuclear vanadium-catecholate complexes [Et 3NH] 2[V VO 2(μ-cat)] 2( 1) and [Et 3NH] 2[V VO 2(μ-N-2,3-D)] 2( 2) (cat = catechol, N-2,3-D = naphthalene-2,3-diol) have been synthesized and characterized by X-ray diffraction, IR, UV-vis spectroscopy and cyclic voltammetry (CV). X-ray analysis reveals that the structures of complexes 1 and 2 are both in the anion form of V. Et 3N works as counter-ions and connects the main frame by hydrogen bonding. The electrochemical behavior of the two complexes is studied in comparison to that of the free ligands and the two complexes display different redox potentials. Pharmaceutical screenings of complexes 1 and 2 have been made against two representative cancer cell-lines A-549 (lung cancer) and Bel-7402 (liver cancer) by MTT assay. The inhibition of cell proliferation was determined 72 h after cells were exposed to the tested compounds at a concentration of 5 μg/mL. Complex 1 exhibits well inhibition ratio against both two cell-lines (76.28% and 75.94%), while 2 displays positive and negative effect (65.36% and -68.82%) respectively. In association with X-ray and electrochemistry, a preliminary analysis about the possible inhibitory mechanism is provided.

  3. Distribution of class ii major histocompatibility complex antigenexpressing cells in human dental pulp with carious lesions

    Directory of Open Access Journals (Sweden)

    Tetiana Haniastuti

    2012-09-01

    Full Text Available Background: Dental caries is a bacterial infection which causes destruction of the hard tissues of the tooth. Exposure of the dentin to the oral environment as a result of caries inevitably results in a cellular response in the pulp. The major histocompatibility complex (MHC is a group of genes that code for cell-surface histocompatibility antigens. Cells expressing class II MHC molecules participate in the initial recognition and the processing of antigenic substances to serve as antigen-presenting cells. Purpose: The aim of the study was to elucidate the alteration in the distribution of class II MHC antigen-expressing cells in human dental pulp as carious lesions progressed toward the pulp. Methods: Fifteen third molars with caries at the occlusal site at various stages of decay and 5 intact third molars were extracted and used in this study. Before decalcifying with 10% EDTA solution (pH 7.4, all the samples were observed by micro-computed tomography to confirm the lesion condition three-dimensionally. The specimens were then processed for cryosection and immunohistochemistry using an anti-MHC class II monoclonal antibody. Results: Class II MHC antigen-expressing cells were found both in normal and carious specimens. In normal tooth, the class II MHC-immunopositive cells were observed mainly at the periphery of the pulp tissue. In teeth with caries, class II MHC-immunopositive cells were located predominantly subjacent to the carious lesions. As the caries progressed, the number of class II MHC antigen-expressing cells was increased. Conclusion: The depth of carious lesions affects the distribution of class II MHC antigen-expressing cells in the dental pulp.Latar belakang: Karies merupakan penyakit infeksi bakteri yang mengakibatkan destruksi jaringan keras gigi. Dentin yang terbuka akibat karies akan menginduksi respon imun seluler pada pulpa. Kompleks histokompatibilitas utama (MHC merupakan sekumpulan gen yang mengkode histokompatibilitas

  4. Proteomic Analysis of Calcium- and Phosphorylation-dependentCalmodulin Complexes in Mammalian Cells

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Deok-Jin; Wang, Daojing

    2006-05-26

    Protein conformational changes due to cofactor binding (e.g. metal ions, heme) and/or posttranslational modifications (e.g. phosphorylation) modulate dynamic protein complexes. Calmodulin (CaM) plays an essential role in regulating calcium (Ca{sup 2+}) signaling and homeostasis. No systematic approach on the identification of phosphorylation-dependent Ca{sup 2+}/CaM binding proteins has been published. Herein, we report a proteome-wide study of phosphorylation-dependent CaM binding proteins from mammalian cells. This method, termed 'Dynamic Phosphoprotein Complex Trapping', 'DPPC Trapping' for short, utilizes a combination of in vivo and in vitro assays. The basic strategy is to drastically shift the equilibrium towards endogenous phosphorylation of Ser, Thr, and Tyr at the global scale by inhibiting corresponding phosphatases in vivo. The phosphorylation-dependent calmodulin-binding proteins are then trapped in vitro in a Ca{sup 2+}-dependent manner by CaM-Sepharose chromatography. Finally, the isolated calmodulin-binding proteins are separated by SDS-PAGE and identified by LC/MS/MS. In parallel, the phosphorylation-dependent binding is visualized by silver staining and/or Western blotting. Using this method, we selectively identified over 120 CaM-associated proteins including many previously uncharacterized. We verified ubiquitin-protein ligase EDD1, inositol 1, 4, 5-triphosphate receptor type 1 (IP{sub 3}R1), and ATP-dependent RNA helicase DEAD box protein 3 (DDX3), as phosphorylation-dependent CaM binding proteins. To demonstrate the utilities of our method in understanding biological pathways, we showed that pSer/Thr of IP{sub 3}R1 in vivo by staurosporine-sensitive kinase(s), but not by PKA/PKG/PKC, significantly reduced the affinity of its Ca{sup 2+}-dependent CaM binding. However, pSer/Thr of IP{sub 3}R1 did not substantially affect its Ca{sup 2+}-independent CaM binding. We further showed that phosphatase PP1, but not PP2A or PP2B

  5. Transition Metal Polypyridine Complexes: Studies of Mediation in Dye-Sensitized Solar Cells and Charge Separation

    Energy Technology Data Exchange (ETDEWEB)

    Elliott, C. Michael [Colorado State Univ., Fort Collins, CO (United States). Dept. of Chemistry; Prieto, Amy L. [Colorado State Univ., Fort Collins, CO (United States). Dept. of Chemistry

    2017-02-08

    The Elliott group has long been supported by DOE for studies of cobalt(II/III) trisbypiridine (DTB) mediator complexes in dye sensitized solar cells. Previous work demonstrated that Co(II/III) chemistry is sensitive to the environment, showing unprecedented electrode-surface and electrolyte dependant voltammetry. In electrolytes that have large lipophilic cations, voltammetry of the [Co(DTB)3]2+/3+ couple is nearly Nernstian in appearance on nominally oxide-free metal surfaces. In contrast, on semiconductor electrodes in electrolytes with small, hard cations such as Li+, the electron transfer rates are so slow that it is difficult to measure any Faradaic current even at overpotentials of ±1 V. These studies are of direct relevance to the operation of cobalt-based mediators in solar cells. The research has also shown that these mediators are compatible with copper phenantroline based dyes, in contrast to I- due to the insolubility of CuI.

  6. Composite cell sheet for periodontal regeneration: crosstalk between different types of MSCs in cell sheet facilitates complex periodontal-like tissue regeneration.

    Science.gov (United States)

    Zhang, Hao; Liu, Shiyu; Zhu, Bin; Xu, Qiu; Ding, Yin; Jin, Yan

    2016-11-14

    Tissue-engineering strategies based on mesenchymal stem cells (MSCs) and cell sheets have been widely used for periodontal tissue regeneration. However, given the complexity in periodontal structure, the regeneration methods using a single species of MSC could not fulfill the requirement for periodontal regeneration. We researched the interaction between the periodontal ligament stem cells (PDLSCs) and jaw bone marrow-derived mesenchymal stem cells (JBMMSCs), and constructed a composite cell sheet comprising both of the above MSCs to regenerate complex periodontium-like structures in nude mice. Our results show that by co-culturing PDLSCs and JBMMSCs, the expressions of bone and extracellular matrix (ECM)-related genes and proteins were significantly improved in both MSCs. Further investigations showed that, compared to the cell sheet using PDLSCs or JBMMSCs, the composite stem cell sheet (CSCS), which comprises these two MSCs, expressed higher levels of bone- and ECM-related genes and proteins, and generated a composite structure more similar to the native periodontal tissue physiologically in vivo. In conclusion, our results demonstrate that the crosstalk between PDLSCs and JBMMSCs in cell sheets facilitate regeneration of complex periodontium-like structures, providing a promising new strategy for physiological and functional regeneration of periodontal tissue.

  7. Poly(amido)amine (PAMAM) dendrimer-cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    Science.gov (United States)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath

    2013-09-01

    Dendrimer-cisplatin complexes were prepared using PAMAM dendrimers with terminal -NH2 and -COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer-cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was 11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC50 values in ovarian cancer cells when compared with carboxylate surface dendrimers ( p cisplatin complexes resulted in a 7.0-fold increase ( p cisplatin chemotherapy of ovarian cancer.

  8. Perivascular Epithelioid Cell Tumor (PEComa of the Uterine Cervix in a Patient with Tuberous Sclerosis Complex: A Literature Review

    Directory of Open Access Journals (Sweden)

    Handan ÇELİK

    2018-01-01

    Full Text Available Perivascular epithelioid cell tumors (PEComa are a rare type of mesenchymal tumor arising from perivascular epithelial cells. These tumor cells are a co-expression of both melanocytic and myogenic antigens, such as HMB 45 and smooth muscle actin, and at least in some patients, are located around vessels. PEComas has been reported at various sites, including visceral organs, soft tissue, the prostate gland and broad ligaments. In the female reproductive system, the uterine corpus is the most common site of involvement. Some cases are related to tuberous sclerosis complex. Cervical PEComa with tuberous sclerosis complex is presented in the case of a 41 year-old and the literature is reviewed. There have been only eight cases of cervical PEComas and only one other case associated with tuberous sclerosis complex reported to date.

  9. Interactions of opsonized immune complexes with whole blood cells: binding to erythrocytes restricts complex uptake by leucocyte populations

    DEFF Research Database (Denmark)

    Nielsen, C H; Svehag, S E; Marquart, H V

    1994-01-01

    -binding to granulocytes (PMN), monocytes and lymphocytes was inhibited by up to 46%, 61% and 48%, respectively, depending on the incubation time and the IC-concentration tested. The E-mediated inhibition of the binding to PMN was found to correlate with the average numbers of CR1 per E during the initial 15 min...... to the findings for PMN, the difference between IC-binding to monocytes in the absence and presence of E increased progressively over the 90 min observation period, suggesting that different mechanisms are involved in the late-phase IC uptake by monocytes and PMN. Lymphocytes were heterogeneous with respect to IC...... binding, the main contributors being B cells. E initially inhibited and then later enhanced the IC binding to lymphocytes, suggesting that E promote B cell uptake of C3d,g-covered IC via CR2. Our findings, that E can restrict the IC uptake by circulating leucocytes, and that an IC-induced degranulation...

  10. Exocytosis and polarity in plant cells: insights by studying cellulose synthase complexes and the exocyst

    NARCIS (Netherlands)

    Ying Zhang, Ying

    2012-01-01

    The work presented in this thesis covers aspects of exocytosis, plant cell growth and cell wall formation. These processes are strongly linked as cell growth and cell wall formation occur simultaneously and exocytosis is the process that delivers cell wall components to the existing cell wall

  11. Effects of hyaluronic acid- chitosan-gelatin complex on the apoptosis and cell cycle of L929 cells

    Institute of Scientific and Technical Information of China (English)

    MAO Jinshu; WANG Xianghui; CUI Yuanlu; YAO Kangde

    2003-01-01

    With the development in the field of tissue engineering, the interaction between biomaterials and cells has been deeply studied. Viewing the cells seeded on the surface of materials as an organic whole, cell cycle and apoptosis are analyzed to deepen the study of cell compatibility on biomaterials, while cellproliferation and differentiation are studied at the same time. In this paper, hyaluronic acid is incorporated into the chitosan-gelatin system. Propidium iodide (PI) was used in cell cycle analysis and the double-staining of cells with annexin-V and PI was applied in cell apoptosis analysis. The results show that incorporated hyaluronic acid shortens the adaptation period of cells on the material surface, and then cells enter the normal cell cycle quickly. In addition, added hyaluronic acid inhibits cell apoptosis triggered by the membranes. Therefore,hyaluronic acid improves the cell compatibility of chitosan-gelatin system and benefits the design of biomimetic materials.

  12. Mapping photothermally induced gene expression in living cells and tissues by nanorod-locked nucleic acid complexes.

    Science.gov (United States)

    Riahi, Reza; Wang, Shue; Long, Min; Li, Na; Chiou, Pei-Yu; Zhang, Donna D; Wong, Pak Kin

    2014-04-22

    The photothermal effect of plasmonic nanostructures has numerous applications, such as cancer therapy, photonic gene circuit, large cargo delivery, and nanostructure-enhanced laser tweezers. The photothermal operation can also induce unwanted physical and biochemical effects, which potentially alter the cell behaviors. However, there is a lack of techniques for characterizing the dynamic cell responses near the site of photothermal operation with high spatiotemporal resolution. In this work, we show that the incorporation of locked nucleic acid probes with gold nanorods allows photothermal manipulation and real-time monitoring of gene expression near the area of irradiation in living cells and animal tissues. The multimodal gold nanorod serves as an endocytic delivery reagent to transport the probes into the cells, a fluorescence quencher and a binding competitor to detect intracellular mRNA, and a plasmonic photothermal transducer to induce cell ablation. We demonstrate the ability of the gold nanorod-locked nucleic acid complex for detecting the spatiotemporal gene expression in viable cells and tissues and inducing photothermal ablation of single cells. Using the gold nanorod-locked nucleic acid complex, we systematically characterize the dynamic cellular heat shock responses near the site of photothermal operation. The gold nanorod-locked nucleic acid complex enables mapping of intracellular gene expressions and analyzes the photothermal effects of nanostructures toward various biomedical applications.

  13. AJUBA LIM Proteins Limit Hippo Activity in Proliferating Cells by Sequestering the Hippo Core Kinase Complex in the Cytosol.

    Science.gov (United States)

    Jagannathan, Radhika; Schimizzi, Gregory V; Zhang, Kun; Loza, Andrew J; Yabuta, Norikazu; Nojima, Hitoshi; Longmore, Gregory D

    2016-10-15

    The Hippo pathway controls organ growth and is implicated in cancer development. Whether and how Hippo pathway activity is limited to sustain or initiate cell growth when needed is not understood. The members of the AJUBA family of LIM proteins are negative regulators of the Hippo pathway. In mammalian epithelial cells, we found that AJUBA LIM proteins limit Hippo regulation of YAP, in proliferating cells only, by sequestering a cytosolic Hippo kinase complex in which LATS kinase is inhibited. At the plasma membranes of growth-arrested cells, AJUBA LIM proteins do not inhibit or associate with the Hippo kinase complex. The ability of AJUBA LIM proteins to inhibit YAP regulation by Hippo and to associate with the kinase complex directly correlate with their capacity to limit Hippo signaling during Drosophila wing development. AJUBA LIM proteins did not influence YAP activity in response to cell-extrinsic or cell-intrinsic mechanical signals. Thus, AJUBA LIM proteins limit Hippo pathway activity in contexts where cell proliferation is needed. Copyright © 2016 Jagannathan et al.

  14. Mycobacterium tuberculosis complex enhances susceptibility of CD4 T cells to HIV through a TLR2-mediated pathway.

    Directory of Open Access Journals (Sweden)

    Seema M Thayil

    Full Text Available Among HIV-infected individuals, co-infection with Mycobacterium tuberculosis is associated with faster progression to AIDS. We investigated the hypothesis that M. bovis BCG and M. tuberculosis (Mtb complex could enhance susceptibility of CD4+ cells to HIV infection. Peripheral blood mononuclear cells (PBMCs collected from healthy donors were stimulated with M. bovis BCG, M. tuberculosis CDC1551 and M. smegmatis MC(2155, and stimulated CD4+ cells were infected with R5-and X4-tropic single replication-competent pseudovirus. CD4+ cells stimulated with Mtb complex showed enhanced infection with R5- and X4-tropic HIV, compared to unstimulated cells or cells stimulated with M. smegmatis (p<0.01. Treatment with TLR2 siRNA reversed the increased susceptibility of CD4+ cells with R5- and X4-tropic virus induced by Mtb complex. These findings suggest that TB infection and/or BCG vaccination may be a risk factor for HIV acquisition.

  15. Interaction between NANOS2 and the CCR4-NOT Deadenylation Complex Is Essential for Male Germ Cell Development in Mouse

    OpenAIRE

    Suzuki, Atsushi; Saba, Rie; Miyoshi, Kei; Morita, Yoshinori; Saga, Yumiko

    2012-01-01

    Nanos is one of the evolutionarily conserved proteins implicated in germ cell development and we have previously shown that it interacts with the CCR4-NOT deadenylation complex leading to the suppression of specific RNAs. However, the molecular mechanism and physiological significance of this interaction have remained elusive. In our present study, we identify CNOT1, a component of the CCR4-NOT deadenylation complex, as a direct factor mediating the interaction with NANOS2. We find that the f...

  16. Cu(II Complexes of Isoniazid Schiff Bases: DNA/BSA Binding and Cytotoxicity Studies on A549 Cell Line

    Directory of Open Access Journals (Sweden)

    Pulipaka Ramadevi

    2014-01-01

    Full Text Available A series of isonicotinoyl hydrazones have been synthesized via template method and were complexed to Cu(II. The ligands are coordinated to Cu(II ion through the enolic oxygen and azomethine nitrogen resulting in a square planar geometry. The CT-DNA and bovine serum albumin binding propensities of the compounds were determined spectrophotometrically, the results of which indicate good binding propensity of complexes to DNA and BSA with high binding constant values. Furthermore, the compounds have been investigated for their cytotoxicities on A549 human lung cancer cell. Also the mode of cell death was examined employing various staining techniques and was found to be apoptotic.

  17. Mitochondrial genome-knockout cells demonstrate a dual mechanism of action for the electron transport complex I inhibitor mycothiazole.

    Science.gov (United States)

    Meyer, Kirsten J; Singh, A Jonathan; Cameron, Alanna; Tan, An S; Leahy, Dora C; O'Sullivan, David; Joshi, Praneta; La Flamme, Anne C; Northcote, Peter T; Berridge, Michael V; Miller, John H

    2012-04-01

    Mycothiazole, a polyketide metabolite isolated from the marine sponge Cacospongia mycofijiensis, is a potent inhibitor of metabolic activity and mitochondrial electron transport chain complex I in sensitive cells, but other cells are relatively insensitive to the drug. Sensitive cell lines (IC(50) 0.36-13.8 nM) include HeLa, P815, RAW 264.7, MDCK, HeLa S3, 143B, 4T1, B16, and CD4/CD8 T cells. Insensitive cell lines (IC(50) 12.2-26.5 μM) include HL-60, LN18, and Jurkat. Thus, there is a 34,000-fold difference in sensitivity between HeLa and HL-60 cells. Some sensitive cell lines show a biphasic response, suggesting more than one mechanism of action. Mitochondrial genome-knockout ρ(0) cell lines are insensitive to mycothiazole, supporting a conditional mitochondrial site of action. Mycothiazole is cytostatic rather than cytotoxic in sensitive cells, has a long lag period of about 12 h, and unlike the complex I inhibitor, rotenone, does not cause G(2)/M cell cycle arrest. Mycothiazole decreases, rather than increases the levels of reactive oxygen species after 24 h. It is concluded that the cytostatic inhibitory effects of mycothiazole on mitochondrial electron transport function in sensitive cell lines may depend on a pre-activation step that is absent in insensitive cell lines with intact mitochondria, and that a second lower-affinity cytotoxic target may also be involved in the metabolic and growth inhibition of cells.

  18. Octamer-binding protein 4 affects the cell biology and phenotypic transition of lung cancer cells involving β-catenin/E-cadherin complex degradation.

    Science.gov (United States)

    Chen, Zhong-Shu; Ling, Dong-Jin; Zhang, Yang-De; Feng, Jian-Xiong; Zhang, Xue-Yu; Shi, Tian-Sheng

    2015-03-01

    Clinical studies have reported evidence for the involvement of octamer‑binding protein 4 (Oct4) in the tumorigenicity and progression of lung cancer; however, the role of Oct4 in lung cancer cell biology in vitro and its mechanism of action remain to be elucidated. Mortality among lung cancer patients is more frequently due to metastasis rather than their primary tumors. Epithelial‑mesenchymal transition (EMT) is a prominent biological event for the induction of epithelial cancer metastasis. The aim of the present study was to investigate whether Oct4 had the capacity to induce lung cancer cell metastasis via the promoting the EMT in vitro. Moreover, the effect of Oct4 on the β‑catenin/E‑cadherin complex, associated with EMT, was examined using immunofluorescence and immunoprecipitation assays as well as western blot analysis. The results demonstrated that Oct4 enhanced cell invasion and adhesion accompanied by the downregulation of epithelial marker cytokeratin, and upregulation of the mesenchymal markers vimentin and N‑cadherin. Furthermore, Oct4 induced EMT of lung cancer cells by promoting β‑catenin/E‑cadherin complex degradation and regulating nuclear localization of β‑catenin. In conclusion, the present study indicated that Oct4 affected the cell biology of lung cancer cells in vitro through promoting lung cancer cell metastasis via EMT; in addition, the results suggested that the association and degradation of the β‑catenin/E‑cadherin complex was regulated by Oct4 during the process of EMT.

  19. Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

    DEFF Research Database (Denmark)

    Mustafa, A S; Amoudy, H A; Wiker, H G

    1998-01-01

    GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

  20. Charge-Transfer Dynamics in the Lowest Excited State of a Pentacene–Fullerene Complex: Implications for Organic Solar Cells

    KAUST Repository

    Joseph, Saju

    2017-10-02

    We characterize the dynamic nature of the lowest excited state in a pentacene/C60 complex on the femtosecond time scale, via a combination of ab initio molecular dynamics and time-dependent density functional theory. We analyze the correlations between the molecular vibrations of the complex and the oscillations in the electron-transfer character of its lowest excited state, which point to vibration-induced coherences between the (pentacene-based) local-excitation (LE) state and the complex charge-transfer (CT) state. We discuss the implications of our results on this model system for the exciton-dissociation process in organic solar cells.

  1. Mast cells contribute to the mucosal adjuvant effect of CTA1-DD after IgG-complex formation.

    Science.gov (United States)

    Fang, Yu; Larsson, Lisa; Mattsson, Johan; Lycke, Nils; Xiang, Zou

    2010-09-01

    Mast cell activation is one of the most dramatic immune-mediated responses the body can encounter. In the worst scenario (i.e., anaphylaxis), this response is fatal. However, the importance of mast cells as initiators and effectors of both innate and adaptive immunity in healthy individuals has recently been appreciated. It was reported that mast cell activation can be used as an adjuvant to promote Ag-specific humoral immune responses upon vaccination. In this study, we have used a clinically relevant mucosal adjuvant, cholera toxin A1 subunit (CTA1)-DD, which is a fusion protein composed of CTA1, the ADP-ribosylating part of cholera toxin, and DD, two Ig-binding domains derived from Staphylococcus aureus protein A. CTA1-DD in combination with polyclonal IgG induced degranulation and production of TNF-alpha from mouse mast cells. Furthermore, CTA1-DD and polyclonal IgG complex induced mast cell degranulation in mouse skin tissue and nasal mucosa. We also found that intranasal immunization with hapten (4-hydroxy-3-nitrophenyl) acetyl (NP) coupled to chicken gammaglobulin admixed with CTA1-DD complexed with polyclonal IgG greatly enhanced serum IgG anti-NP Ab responses and stimulated higher numbers of NP-specific plasma cells in the bone marrow as compared with that observed in mice immunized with NP-chicken gammaglobulin with CTA1-DD alone. This CTA1-DD/IgG complex-mediated enhancement was mast cell dependent because it was absent in mast cell-deficient Kit(W-sh/W-sh) mice. In conclusion, our data suggest that a clinically relevant adjuvant, CTA1-DD, exerts additional augmenting effects through activation of mucosal mast cells, clearly demonstrating that mast cells could be further exploited for improving the efficacy of mucosal vaccines.

  2. Putative supramolecular complexes formed by carotenoids and xanthophylls with ascorbic acid to reverse multidrug resistance in cancer cells.

    Science.gov (United States)

    Molnár, József; Serly, Julianna; Pusztai, Rozália; Vincze, Irén; Molnár, Péter; Horváth, Györgyi; Deli, József; Maoka, Takashi; Zalatnai, Attila; Tokuda, Harukuni; Nishino, Hoyoku

    2012-02-01

    The molecular basis of interaction of selected carotenoids and xanthophylls with ascorbic acid on cancer cells was studied to determine their anticancer effects. Drug accumulation was measured in a human ABCB1 gene-transfected mouse lymphoma cell line and in a human lung cancer cell line by flow cytometry; furthermore, their anticancer effects were determined in mice in vivo. Several carotenoids inhibited the multidrug resistance of cancer cells. Ascorbic acid improved the effect of certain xanthophylls, but the effect of capsanthin was not modified. Capsanthin had weak (12%) but capsorubin (85%) had a remarkable antiproliferative effect on A549 lung cancer cells. Capsorubin reduced immediate-early tumor antigen expression, while capsanthin was not effective. Capsorubin accumulates selectively in the nuclei of cancer cells. The Authors suggest a special complex formation between membrane-bound capsorubin and ascorbic acid, which can be exploited in experimental chemotherapy.

  3. Peripheral Light-Harvesting LH2 Complex Can Be Assembled in Cells of Nonsulfur Purple Bacterium Rhodoblastus acidophilus without Carotenoids.

    Science.gov (United States)

    Bol'shakov, M A; Ashikhmin, A A; Makhneva, Z K; Moskalenko, A A

    2015-09-01

    The effect of carotenoids on the assembly of LH2 complex in cells of the purple nonsulfur bacterium Rhodoblastus acidophilus was investigated. For this purpose, the bacterial culture was cultivated with an inhibitor of carotenoid biosynthesis - 71 µM diphenylamine (DPA). The inhibitor decreased the level of biosynthesis of the colored carotenoids in membranes by ~58%. It was found that a large amount of phytoene was accumulated in them. This carotenoid precursor was bound nonspecifically to LH2 complex and did not stabilize its structure. Thermostability testing of the isolated LH2 complex together with analysis of carotenoid composition revealed that the population of this complex was heterogeneous with respect to carotenoid composition. One fraction of the LH2 complex with carotenoid content around 90% remains stable and was not destroyed under heating for 15 min at 50°C. The other fraction of LH2 complex containing on average less than one molecule of carotenoid per complex was destroyed under heating, forming a zone of free pigments (and polypeptides). The data suggest that a certain part of the LH2 complexes is assembled without carotenoids in cells of the nonsulfur bacterium Rbl. acidophilus grown with DPA. These data contradict the fact that the LH2 complex from nonsulfur bacteria cannot be assembled without carotenoids, but on the other hand, they are in good agreement with the results demonstrated in our earlier studies of the sulfur bacteria Allochromatium minutissimum and Ectothiorhodospira haloalkaliphila. Carotenoidless LH2 complex was obtained from these bacteria with the use of DPA (Moskalenko, A. A., and Makhneva, Z. K. (2012) J. Photochem. Photobiol., 108, 1-7; Ashikhmin, A., et al. (2014) Photosynth. Res., 119, 291-303).

  4. Response of single bacterial cells to stress gives rise to complex history dependence at the population level

    Science.gov (United States)

    Mathis, Roland; Ackermann, Martin

    2016-01-01

    Most bacteria live in ever-changing environments where periods of stress are common. One fundamental question is whether individual bacterial cells have an increased tolerance to stress if they recently have been exposed to lower levels of the same stressor. To address this question, we worked with the bacterium Caulobacter crescentus and asked whether exposure to a moderate concentration of sodium chloride would affect survival during later exposure to a higher concentration. We found that the effects measured at the population level depended in a surprising and complex way on the time interval between the two exposure events: The effect of the first exposure on survival of the second exposure was positive for some time intervals but negative for others. We hypothesized that the complex pattern of history dependence at the population level was a consequence of the responses of individual cells to sodium chloride that we observed: (i) exposure to moderate concentrations of sodium chloride caused delays in cell division and led to cell-cycle synchronization, and (ii) whether a bacterium would survive subsequent exposure to higher concentrations was dependent on the cell-cycle state. Using computational modeling, we demonstrated that indeed the combination of these two effects could explain the complex patterns of history dependence observed at the population level. Our insight into how the behavior of single cells scales up to processes at the population level provides a perspective on how organisms operate in dynamic environments with fluctuating stress exposure. PMID:26960998

  5. Major histocompatibility complex-unrestricted cytolytic activity of human T cells: analysis of precursor frequency and effector phenotype

    International Nuclear Information System (INIS)

    Patel, S.S.; Thiele, D.L.; Lipsky, P.E.

    1987-01-01

    The frequency and phenotype of human T cells that mediate major histocompatibility complex (MHC)-unrestricted cytolysis were analyzed. T cell clones were generated by culturing adherent cell-depleted peripheral blood mononuclear cells at a density of 0.3 cell/well with phytohemagglutinin, recombinant interleukin 2 (rIL-2), and irradiated autologous peripheral blood mononuclear cells and/or Epstein-Barr virus-transformed lymphoblastoid cell lines. All of the 198 clones generated by this method were T cells (CD2 + , CD3 + , CD4 + or CD2 + , CD3 + , CD8 + ) that possessed potent lytic activity against K562, an erythroleukemia line sensitive to lysis by human natural killer cells, and Cur, a renal carcinoma cell line resistant to human natural killer activity. Cytolysis, measured by 51 Cr release, was MHC-unrestricted, since the clones were able to lyse MHC class I or class II negative targets, as well as MHC class I and class II negative targets. Although the clones produced tissue necrosis factor/lymphotoxin-like molecules, lysis of Cur of K562 was not mediated by a soluble factor secreted by the clones. These data indicate that the capacity for MHC-unrestricted tumoricidal activity and expression of NKH1 and CD11b, but not CD 16, are properties common to all or nearly all human peripheral blood-derived T cell clones regardless of CD4 or CD8 phenotype

  6. Hyaluronan activates Hyal-2/WWOX/Smad4 signaling and causes bubbling cell death when the signaling complex is overexpressed

    Science.gov (United States)

    Hsu, Li-Jin; Hong, Qunying; Chen, Shur-Tzu; Kuo, Hsiang-Lin; Schultz, Lori; Heath, John; Lin, Sing-Ru; Lee, Ming-Hui; Li, Dong-Zhang; Li, Zih-Ling; Cheng, Hui-Ching; Armand, Gerard; Chang, Nan-Shan

    2017-01-01

    Malignant cancer cells frequently secrete significant amounts of transforming growth factor beta (TGF-β), hyaluronan (HA) and hyaluronidases to facilitate metastasizing to target organs. In a non-canonical signaling, TGF-β binds membrane hyaluronidase Hyal-2 for recruiting tumor suppressors WWOX and Smad4, and the resulting Hyal-2/WWOX/Smad4 complex is accumulated in the nucleus to enhance SMAD-promoter dependent transcriptional activity. Yeast two-hybrid analysis showed that WWOX acts as a bridge to bind both Hyal-2 and Smad4. When WWOX-expressing cells were stimulated with high molecular weight HA, an increased formation of endogenous Hyal-2/WWOX/Smad4 complex occurred rapidly, followed by relocating to the nuclei in 20-40 min. In WWOX-deficient cells, HA failed to induce Smad2/3/4 relocation to the nucleus. To prove the signaling event, we designed a real time tri-molecular FRET analysis and revealed that HA induces the signaling pathway from ectopic Smad4 to WWOX and finally to p53, as well as from Smad4 to Hyal-2 and then to WWOX. An increased binding of the Smad4/Hyal-2/WWOX complex occurs with time in the nucleus that leads to bubbling cell death. In contrast, HA increases the binding of Smad4/WWOX/p53, which causes membrane blebbing but without cell death. In traumatic brain injury-induced neuronal death, the Hyal-2/WWOX complex was accumulated in the apoptotic nuclei of neurons in the rat brains in 24 hr post injury, as determined by immunoelectron microscopy. Together, HA activates the Hyal-2/WWOX/Smad4 signaling and causes bubbling cell death when the signaling complex is overexpressed. PMID:27845895

  7. Hyaluronan activates Hyal-2/WWOX/Smad4 signaling and causes bubbling cell death when the signaling complex is overexpressed.

    Science.gov (United States)

    Hsu, Li-Jin; Hong, Qunying; Chen, Shur-Tzu; Kuo, Hsiang-Lin; Schultz, Lori; Heath, John; Lin, Sing-Ru; Lee, Ming-Hui; Li, Dong-Zhang; Li, Zih-Ling; Cheng, Hui-Ching; Armand, Gerard; Chang, Nan-Shan

    2017-03-21

    Malignant cancer cells frequently secrete significant amounts of transforming growth factor beta (TGF-β), hyaluronan (HA) and hyaluronidases to facilitate metastasizing to target organs. In a non-canonical signaling, TGF-β binds membrane hyaluronidase Hyal-2 for recruiting tumor suppressors WWOX and Smad4, and the resulting Hyal-2/WWOX/Smad4 complex is accumulated in the nucleus to enhance SMAD-promoter dependent transcriptional activity. Yeast two-hybrid analysis showed that WWOX acts as a bridge to bind both Hyal-2 and Smad4. When WWOX-expressing cells were stimulated with high molecular weight HA, an increased formation of endogenous Hyal-2/WWOX/Smad4 complex occurred rapidly, followed by relocating to the nuclei in 20-40 min. In WWOX-deficient cells, HA failed to induce Smad2/3/4 relocation to the nucleus. To prove the signaling event, we designed a real time tri-molecular FRET analysis and revealed that HA induces the signaling pathway from ectopic Smad4 to WWOX and finally to p53, as well as from Smad4 to Hyal-2 and then to WWOX. An increased binding of the Smad4/Hyal-2/WWOX complex occurs with time in the nucleus that leads to bubbling cell death. In contrast, HA increases the binding of Smad4/WWOX/p53, which causes membrane blebbing but without cell death. In traumatic brain injury-induced neuronal death, the Hyal-2/WWOX complex was accumulated in the apoptotic nuclei of neurons in the rat brains in 24 hr post injury, as determined by immunoelectron microscopy. Together, HA activates the Hyal-2/WWOX/Smad4 signaling and causes bubbling cell death when the signaling complex is overexpressed.

  8. Non-canonical TAF complexes regulate active promoters in human embryonic stem cells.

    Science.gov (United States)

    Maston, Glenn A; Zhu, Lihua Julie; Chamberlain, Lynn; Lin, Ling; Fang, Minggang; Green, Michael R

    2012-11-13

    The general transcription factor TFIID comprises the TATA-box-binding protein (TBP) and approximately 14 TBP-associated factors (TAFs). Here we find, unexpectedly, that undifferentiated human embryonic stem cells (hESCs) contain only six TAFs (TAFs 2, 3, 5, 6, 7 and 11), whereas following differentiation all TAFs are expressed. Directed and global chromatin immunoprecipitation analyses reveal an unprecedented promoter occupancy pattern: most active genes are bound by only TAFs 3 and 5 along with TBP, whereas the remaining active genes are bound by TBP and all six hESC TAFs. Consistent with these results, hESCs contain a previously undescribed complex comprising TAFs 2, 6, 7, 11 and TBP. Altering the composition of hESC TAFs, either by depleting TAFs that are present or ectopically expressing TAFs that are absent, results in misregulated expression of pluripotency genes and induction of differentiation. Thus, the selective expression and use of TAFs underlies the ability of hESCs to self-renew.DOI:http://dx.doi.org/10.7554/eLife.00068.001.

  9. Retinal nerve fiber layer and ganglion cell complex thickness in patients with type 2 diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Mehmet Demir

    2014-01-01

    Full Text Available Aim: The aim of the following study is to evaluate the retinal nerve fiber layer (RNFL and ganglion cell complex (GCC thickness in patients with type 2 diabetes mellitus (DM. Materials and Methods: Average, inferior, and superior values of RNFL and GCC thickness were measured in 123 patients using spectral domain optical coherence tomography. The values of participants with DM were compared to controls. Diabetic patients were collected in Groups 1, 2 and 3. Group 1 = 33 participants who had no diabetic retinopathy (DR; Group 2 = 30 participants who had mild nonproliferative DR and Group 3 = 30 participants who had moderate non-proliferative DR. The 30 healthy participants collected in Group 4. Analysis of variance test and a multiple linear regression analysis were used for statistical analysis. Results: The values of RNFL and GCC in the type 2 diabetes were thinner than controls, but this difference was not statistically significant. Conclusions: This study showed that there is a nonsignificant loss of RNFL and GCC in patients with type 2 diabetes.

  10. RNA-Based TWIST1 Inhibition via Dendrimer Complex to Reduce Breast Cancer Cell Metastasis

    Directory of Open Access Journals (Sweden)

    James Finlay

    2015-01-01

    Full Text Available Breast cancer is the leading cause of cancer-related deaths among women in the United States, and survival rates are lower for patients with metastases and/or triple-negative breast cancer (TNBC; ER, PR, and Her2 negative. Understanding the mechanisms of cancer metastasis is therefore crucial to identify new therapeutic targets and develop novel treatments to improve patient outcomes. A potential target is the TWIST1 transcription factor, which is often overexpressed in aggressive breast cancers and is a master regulator of cellular migration through epithelial-mesenchymal transition (EMT. Here, we demonstrate an siRNA-based TWIST1 silencing approach with delivery using a modified poly(amidoamine (PAMAM dendrimer. Our results demonstrate that SUM1315 TNBC cells efficiently take up PAMAM-siRNA complexes, leading to significant knockdown of TWIST1 and EMT-related target genes. Knockdown lasts up to one week after transfection and leads to a reduction in migration and invasion, as determined by wound healing and transwell assays. Furthermore, we demonstrate that PAMAM dendrimers can deliver siRNA to xenograft orthotopic tumors and siRNA remains in the tumor for at least four hours after treatment. These results suggest that further development of dendrimer-based delivery of siRNA for TWIST1 silencing may lead to a valuable adjunctive therapy for patients with TNBC.

  11. Non-SMC condensin I complex proteins control chromosome segregation and survival of proliferating cells in the zebrafish neural retina

    Directory of Open Access Journals (Sweden)

    Harris William A

    2009-07-01

    Full Text Available Abstract Background The condensation of chromosomes and correct sister chromatid segregation during cell division is an essential feature of all proliferative cells. Structural maintenance of chromosomes (SMC and non-SMC proteins form the condensin I complex and regulate chromosome condensation and segregation during mitosis. However, due to the lack of appropriate mutants, the function of the condensin I complex during vertebrate development has not been described. Results Here, we report the positional cloning and detailed characterization of retinal phenotypes of a zebrafish mutation at the cap-g locus. High resolution live imaging reveals that the progression of mitosis between prometa- to telophase is delayed and that sister chromatid segregation is impaired upon loss of CAP-G. CAP-G associates with chromosomes between prometa- and telophase of the cell cycle. Loss of the interaction partners CAP-H and CAP-D2 causes cytoplasmic mislocalization of CAP-G throughout mitosis. DNA content analysis reveals increased genomic imbalances upon loss of non-SMC condensin I subunits. Within the retina, loss of condensin I function causes increased rates of apoptosis among cells within the proliferative ciliary marginal zone (CMZ whereas postmitotic retinal cells are viable. Inhibition of p53-mediated apoptosis partially rescues cell numbers in cap-g mutant retinae and allows normal layering of retinal cell types without alleviating their aberrant nuclear sizes. Conclusion Our findings indicate that the condensin I complex is particularly important within rapidly amplifying progenitor cell populations to ensure faithful chromosome segregation. In contrast, differentiation of postmitotic retinal cells is not impaired upon polyploidization.

  12. Modeling two-phase flow in three-dimensional complex flow-fields of proton exchange membrane fuel cells

    Science.gov (United States)

    Kim, Jinyong; Luo, Gang; Wang, Chao-Yang

    2017-10-01

    3D fine-mesh flow-fields recently developed by Toyota Mirai improved water management and mass transport in proton exchange membrane (PEM) fuel cell stacks, suggesting their potential value for robust and high-power PEM fuel cell stack performance. In such complex flow-fields, Forchheimer's inertial effect is dominant at high current density. In this work, a two-phase flow model of 3D complex flow-fields of PEMFCs is developed by accounting for Forchheimer's inertial effect, for the first time, to elucidate the underlying mechanism of liquid water behavior and mass transport inside 3D complex flow-fields and their adjacent gas diffusion layers (GDL). It is found that Forchheimer's inertial effect enhances liquid water removal from flow-fields and adds additional flow resistance around baffles, which improves interfacial liquid water and mass transport. As a result, substantial improvements in high current density cell performance and operational stability are expected in PEMFCs with 3D complex flow-fields, compared to PEMFCs with conventional flow-fields. Higher current density operation required to further reduce PEMFC stack cost per kW in the future will necessitate optimizing complex flow-field designs using the present model, in order to efficiently remove a large amount of product water and hence minimize the mass transport voltage loss.

  13. Complexes of γ-tubulin with nonreceptor protein tyrosine kinases Src and Fyn in differentiating P19 embryonal carcinoma cells

    International Nuclear Information System (INIS)

    Kukharskyy, Vitaliy; Sulimenko, Vadym; Macurek, Libor; Sulimenko, Tetyana; Draberova, Eduarda; Draber, Pavel

    2004-01-01

    Nonreceptor protein tyrosine kinases of the Src family have been shown to play an important role in signal transduction as well as in regulation of microtubule protein interactions. Here we show that γ-tubulin (γ-Tb) in P19 embryonal carcinoma cells undergoing neuronal differentiation is phosphorylated and forms complexes with protein tyrosine kinases of the Src family, Src and Fyn. Elevated expression of both kinases during differentiation corresponded with increased level of proteins phosphorylated on tyrosine. Immunoprecipitation experiments with antibodies against Src, Fyn, γ-tubulin, and with anti-phosphotyrosine antibody revealed that γ-tubulin appeared in complexes with these kinases. In vitro kinase assays showed tyrosine phosphorylation of proteins in γ-tubulin complexes isolated from differentiated cells. Pretreatment of cells with Src family selective tyrosine kinase inhibitor PP2 reduced the amount of phosphorylated γ-tubulin in the complexes. Binding experiments with recombinant SH2 and SH3 domains of Src and Fyn kinases revealed that protein complexes containing γ-tubulin bound to SH2 domains and that these interactions were of SH2-phosphotyrosine type. The combined data suggest that Src family kinases might have an important role in the regulation of γ-tubulin interaction with tubulin dimers or other proteins during neurogenesis

  14. Rescue of cell death and inflammation of a mouse model of complex 1-mediated vision loss by repurposed drug molecules.

    Science.gov (United States)

    Yu, Alfred K; Datta, Sandipan; McMackin, Marissa Z; Cortopassi, Gino A

    2017-12-15

    Inherited mitochondrial optic neuropathies, such as Leber's hereditary optic neuropathy (LHON) and Autosomal dominant optic atrophy (ADOA) are caused by mutant mitochondrial proteins that lead to defects in mitochondrial complex 1-driven ATP synthesis, and cause specific retinal ganglion cell (RGC) loss. Complex 1 defects also occur in patients with primary open angle glaucoma (POAG), in which there is specific RGC loss. The treatment of mitochondrial optic neuropathy in the US is only supportive. The Ndufs4 knockout (Ndufs4 KO) mouse is a mitochondrial complex 1-deficient model that leads to RGC loss and rapid vision loss and allows for streamlined testing of potential therapeutics. Preceding RGC loss in the Ndufs4 KO is the loss of starburst amacrine cells, which may be an important target in the mechanism of complex 1-deficient vision loss. Papaverine and zolpidem were recently shown to be protective of bioenergetic loss in cell models of optic neuropathy. Treatment of Ndufs4 KO mice with papaverine, zolpidem, and rapamycin-suppressed inflammation, prevented cell death, and protected from vision loss. Thus, in the Ndufs4 KO mouse model of mitochondrial optic neuropathy, papaverine and zolpidem provided significant protection from multiple pathophysiological features, and as approved drugs in wide human use could be considered for the novel indication of human optic neuropathy. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  15. Max-Min SINR in Large-Scale Single-Cell MU-MIMO: Asymptotic Analysis and Low Complexity Transceivers

    KAUST Repository

    Sifaou, Houssem; Kammoun, Abla; Sanguinetti, Luca; Debbah, Merouane; Alouini, Mohamed-Slim

    2016-01-01

    This work focuses on the downlink and uplink of large-scale single-cell MU-MIMO systems in which the base station (BS) endowed with M antennas communicates with K single-antenna user equipments (UEs). Particularly, we aim at reducing the complexity

  16. Increased podophyllotoxin production in Podophyllum hexandrum cell suspension cultures after feeding coniferyl alcohol as a β-cyclodextrin complex

    NARCIS (Netherlands)

    Woerdenbag, H.J.; Van Uden, W.; Frijlink, H.W.; Lerk, C.F.; Pras, N.; Malingre, T.M.

    1990-01-01

    Cell suspension cultures, derived from roots of Podophyllum hexandrum Royle (Berberidaceae), accumulate podophyllotoxin. In this study the use of β-cyclodextrin in feeding the poorly water-soluble precursor coniferyl alcohol to these cultures is described. By complexation with β-cyclodextrin, a

  17. Increased podophyllotoxin production in Podophyllum hexandrum cell suspension cultures after feeding coniferyl alcohol as a β-cyclodextrin complex

    NARCIS (Netherlands)

    Woerdenbag, H J; van Uden, W; Frijlink, H W; Lerk, C F; Pras, N; Malingré, T M

    Cell suspension cultures, derived from roots of Podophyllum hexandrum Royle (Berberidaceae), accumulate podophyllotoxin. In this study the use of β-cyclodextrin in feeding the poorly water-soluble precursor coniferyl alcohol to these cultures is described. By complexation with β-cyclodextrin, a

  18. Mitochondrially targeted vitamin E succinate efficiently kills breast tumour-initiating cells in a complex II-dependent manner

    Czech Academy of Sciences Publication Activity Database

    Yan, B.; Stantic, M.; Zobalová, Renata; Bezawork-Geleta, A.; Stapelberg, M.; Stursa, J.; Prokopová, Kateřina; Dong, L.; Neužil, Jiří

    2015-01-01

    Roč. 15, č. 401 (2015) ISSN 1471-2407 R&D Projects: GA MZd NT14078; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:86652036 Keywords : Tumour-initiating cells * Mitochondrially targeted vitamin E succinate * Complex II Subject RIV: FD - Oncology ; Hematology Impact factor: 3.265, year: 2015

  19. Caprolactone-based bipyridine macroligands for novel ruthenium polypyridyl complexes for applications in dye-sensitized solar cells

    NARCIS (Netherlands)

    Marin, V.N.; Holder, E.; Hoogenboom, R.; Schubert, U.S.

    2004-01-01

    In an attempt to reduce electrolyte loss in solar cells, the use of sold or quasi-solid state electrolytes is advantageous. The authors synthesized a metal contg. polymer via two synthetic pathways. In the first, a bipyridine-bisphenanthroline ruthenium metal complex with a hydroxy functionality was

  20. Targeting Alpha-Fetoprotein (AFP)-MHC Complex with CAR T-Cell Therapy for Liver Cancer.

    Science.gov (United States)

    Liu, Hong; Xu, Yiyang; Xiang, Jingyi; Long, Li; Green, Shon; Yang, Zhiyuan; Zimdahl, Bryan; Lu, Jingwei; Cheng, Neal; Horan, Lucas H; Liu, Bin; Yan, Su; Wang, Pei; Diaz, Juan; Jin, Lu; Nakano, Yoko; Morales, Javier F; Zhang, Pengbo; Liu, Lian-Xing; Staley, Binnaz K; Priceman, Saul J; Brown, Christine E; Forman, Stephen J; Chan, Vivien W; Liu, Cheng

    2017-01-15

    The majority of tumor-specific antigens are intracellular and/or secreted and therefore inaccessible by conventional chimeric antigen receptor (CAR) T-cell therapy. Given that all intracellular/secreted proteins are processed into peptides and presented by class I MHC on the surface of tumor cells, we used alpha-fetoprotein (AFP), a specific liver cancer marker, as an example to determine whether peptide-MHC complexes can be targets for CAR T-cell therapy against solid tumors. We generated a fully human chimeric antigen receptor, ET1402L1-CAR (AFP-CAR), with exquisite selectivity and specificity for the AFP 158-166 peptide complexed with human leukocyte antigen (HLA)-A*02:01. We report that T cells expressing AFP-CAR selectively degranulated, released cytokines, and lysed liver cancer cells that were HLA-A*02:01 + /AFP + while sparing cells from multiple tissue types that were negative for either expressed proteins. In vivo, intratumoral injection of AFP-CAR T cells significantly regressed both Hep G2 and AFP 158 -expressing SK-HEP-1 tumors in SCID-Beige mice (n = 8 for each). Moreover, intravenous administration of AFP-CAR T cells in Hep G2 tumor-bearing NSG mice lead to rapid and profound tumor growth inhibition (n = 6). Finally, in an established intraperitoneal liver cancer xenograft model, AFP-CAR T cells showed robust antitumor activity (n = 6). This study demonstrates that CAR T-cell immunotherapy targeting intracellular/secreted solid tumor antigens can elicit a potent antitumor response. Our approach expands the spectrum of antigens available for redirected T-cell therapy against solid malignancies and offers a promising new avenue for liver cancer immunotherapy. Clin Cancer Res; 23(2); 478-88. ©2016 AACR. ©2016 American Association for Cancer Research.

  1. Complex N-Glycans Influence the Spatial Arrangement of Voltage Gated Potassium Channels in Membranes of Neuronal-Derived Cells.

    Directory of Open Access Journals (Sweden)

    M Kristen Hall

    Full Text Available The intrinsic electrical properties of a neuron depend on expression of voltage gated potassium (Kv channel isoforms, as well as their distribution and density in the plasma membrane. Recently, we showed that N-glycosylation site occupancy of Kv3.1b modulated its placement in the cell body and neurites of a neuronal-derived cell line, B35 neuroblastoma cells. To extrapolate this mechanism to other N-glycosylated Kv channels, we evaluated the impact of N-glycosylation occupancy of Kv3.1a and Kv1.1 channels. Western blots revealed that wild type Kv3.1a and Kv1.1 α-subunits had complex and oligomannose N-glycans, respectively, and that abolishment of the N-glycosylation site(s generated Kv proteins without N-glycans. Total internal reflection fluorescence microscopy images revealed that N-glycans of Kv3.1a contributed to its placement in the cell membrane while N-glycans had no effect on the distribution of Kv1.1. Based on particle analysis of EGFP-Kv proteins in the adhered membrane, glycosylated forms of Kv3.1a, Kv1.1, and Kv3.1b had differences in the number, size or density of Kv protein clusters in the cell membrane of neurites and cell body of B35 cells. Differences were also observed between the unglycosylated forms of the Kv proteins. Cell dissociation assays revealed that cell-cell adhesion was increased by the presence of complex N-glycans of Kv3.1a, like Kv3.1b, whereas cell adhesion was similar in the oligomannose and unglycosylated Kv1.1 subunit containing B35 cells. Our findings provide direct evidence that N-glycans of Kv3.1 splice variants contribute to the placement of these glycoproteins in the plasma membrane of neuronal-derived cells while those of Kv1.1 were absent. Further when the cell membrane distribution of the Kv channel was modified by N-glycans then the cell-cell adhesion properties were altered. Our study demonstrates that N-glycosylation of Kv3.1a, like Kv3.1b, provides a mechanism for the distribution of these

  2. Clostridium botulinum serotype D neurotoxin and toxin complex bind to bovine aortic endothelial cells via sialic acid.

    Science.gov (United States)

    Yoneyama, Tohru; Miyata, Keita; Chikai, Tomoyuki; Mikami, Akifumi; Suzuki, Tomonori; Hasegawa, Kimiko; Ikeda, Toshihiko; Watanabe, Toshihiro; Ohyama, Tohru; Niwa, Koichi

    2008-12-01

    Botulinum neurotoxin (BoNT) is produced as a large toxin complex (L-TC) associated with nontoxic nonhemagglutinin (NTNHA) and three hemagglutinin subcomponents (HA-70, -33 and -17). The binding properties of BoNT to neurons and L-TC to intestinal epithelial cells are well documented, while those to other tissues are largely unknown. Here, to obtain novel insights into the pathogenesis of foodborne botulism, we examine whether botulinum toxins bind to vascular endothelial cells. BoNT and 750 kDa L-TC (a complex of BoNT, NTNHA and HAs) of Clostridium botulinum serotype D were incubated with bovine aortic endothelial cells (BAECs), and binding to the cells was assessed using sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot. Both BoNT and L-TC bound to BAECs, with L-TC showing stronger binding. Binding of BoNT and L-TC to BAECs was significantly inhibited by N-acetyl neuraminic acid in the cell culture medium or by treatment of the cells with neuraminidase. However, galactose, lactose or N-acetyl galactosamine did not significantly inhibit toxin binding to the cells. This is the first report demonstrating that BoNT and L-TC bind to BAECs via sialic acid, and this mechanism may be important in the trafficking pathway of BoNT in foodborne botulism.

  3. Role of the multichain IL-2 receptor complex in the control of normal and malignant T-cell proliferation

    International Nuclear Information System (INIS)

    Waldmann, T.A.

    1987-01-01

    Antigen-induced activation of resting T-cells induces the synthesis of interleukin-2 (IL-2), as well as the expression of specific cell surface receptors for this lymphokine. There are at least two forms of the cellular receptors for IL-2, one with a very high affinity and the other with a lower affinity. The authors have identified two IL-2 binding peptides, a 55-kd peptide reactive with the anti-Tac monoclonal antibody, and a novel 75-kd non-Tac IL-2 binding peptide. Cell lines bearing either the p55, Tac, or the p75 peptide along manifested low-affinity IL-2 binding, whereas cell lines bearing both peptides manifested both high- and low-affinity receptors. Fusion of cell membranes from low-affinity IL-2 binding cells bearing the Tac peptide alone with membranes from a cell line bearing the p75 peptide alone generates hybrid membranes bearing high-affinity receptors. They propose a multichain model for the high-affinity IL-2 receptor in which both the Tac and the p75 IL-2 binding peptides are associated in a receptor complex. In contrast to resting T-cells, human T-cell lymphotropic virus I-associated adult T-cell leukemia cells constitutively express large numbers of IL-2 receptors. Because IL-2 receptors are present on the malignant T-cells but not on normal resting cells, clinical trials have been initiated in which patients with adult T-cell leukemia are being treated with either unmodified or toxin-conjugated forms of anti-Tac monoclonal antibody directed toward this growth factor receptor. Cross-linking studies were done using [ 125 I] IL-2

  4. Stem cell factor induces phosphatidylinositol 3'-kinase-dependent Lyn/Tec/Dok-1 complex formation in hematopoietic cells

    NARCIS (Netherlands)

    T.B. van Dijk (Thamar); M. Parren-Van Amelsvoort (Martine); H. Mano; M.M. von Lindern (Marieke); B. Löwenberg (Bob); E. van den Akker (Emile)

    2000-01-01

    textabstractStem cell factor (SCF) has an important role in the proliferation, differentiation, survival, and migration of hematopoietic cells. SCF exerts its effects by binding to cKit, a receptor with intrinsic tyrosine kinase activity. Activation of

  5. Complex mutual regulation of facilitates chromatin transcription (FACT) subunits on both mRNA and protein levels in human cells.

    Science.gov (United States)

    Safina, Alfiya; Garcia, Henry; Commane, Mairead; Guryanova, Olga; Degan, Seamus; Kolesnikova, Kateryna; Gurova, Katerina V

    2013-08-01

    Facilitates chromatin transcription (FACT) is a chromatin remodeling complex with two subunits: SSRP1 and SPT16. Mechanisms controlling FACT levels are of interest, since the complex is not expressed in most differentiated cells, but is frequently upregulated in cancer, particularly in poorly differentiated, aggressive tumors. Moreover, inhibition of FACT expression or function in tumor cells interferes with their survival. Here we demonstrate that SSRP1 and SPT16 protein levels decline upon induction of cellular differentiation or senescence in vitro and that similar declines in protein levels for both SSRP1 and SPT16 occur upon RNAi-mediated knockdown of either SSRP1 or SPT16. The interdependence of SSRP1 and SPT16 protein levels was found to be due to their association with SSRP1 and SPT16 mRNAs, which stabilizes the proteins. In particular, presence of SSRP1 mRNA is critical for SPT16 protein stability. In addition, binding of SSRP1 and SPT16 mRNAs to the FACT complex increases the stability and efficiency of translation of the mRNAs. These data support a model in which the FACT complex is stable when SSRP1 mRNA is present, but quickly degrades when SSRP1 mRNA levels drop. In the absence of FACT complex, SSRP1 and SPT16 mRNAs are unstable and inefficiently translated, making reactivation of FACT function unlikely in normal cells. Thus, we have described a complex and unusual mode of regulation controlling cellular FACT levels that results in amplified and stringent control of FACT activity. The FACT dependence of tumor cells suggests that mechanisms controlling FACT levels could be targeted for anticancer therapy.

  6. Antiproliferative and apoptosis-inducing activity of an oxidovanadium(IV) complex with the flavonoid silibinin against osteosarcoma cells.

    Science.gov (United States)

    Leon, I E; Porro, V; Di Virgilio, A L; Naso, L G; Williams, P A M; Bollati-Fogolín, M; Etcheverry, S B

    2014-01-01

    Flavonoids are a large family of polyphenolic compounds synthesized by plants. They display interesting biological effects mainly related to their antioxidant properties. On the other hand, vanadium compounds also exhibit different biological and pharmacological effects in cell culture and in animal models. Since coordination of ligands to metals can improve or change the pharmacological properties, we report herein, for the first time, a detailed study of the mechanisms of action of an oxidovanadium(IV) complex with the flavonoid silibinin, Na2[VO(silibinin)2]·6H2O (VOsil), in a model of the human osteosarcoma derived cell line MG-63. The complex inhibited the viability of osteosarcoma cells in a dose-dependent manner with a greater potency than that of silibinin and oxidovanadium(IV) (p cell cycle arrest and activated caspase 3, triggering apoptosis as determined by flow cytometry. As a whole, these results show the main mechanisms of the deleterious effects of VOsil in the osteosarcoma cell line, demonstrating that this complex is a promising compound for cancer treatments.

  7. Nuclear Materials Characterization in the Materials and Fuels Complex Analytical Hot Cells

    International Nuclear Information System (INIS)

    Rodriquez, Michael

    2009-01-01

    As energy prices skyrocket and interest in alternative, clean energy sources builds, interest in nuclear energy has increased. This increased interest in nuclear energy has been termed the 'Nuclear Renaissance'. The performance of nuclear fuels, fuels and reactor materials and waste products are becoming a more important issue as the potential for designing new nuclear reactors is more immediate. The Idaho National Laboratory (INL) Materials and Fuels Complex (MFC) Analytical Laboratory Hot Cells (ALHC) are rising to the challenge of characterizing new reactor materials, byproducts and performance. The ALHC is a facility located near Idaho Falls, Idaho at the INL Site. It was built in 1958 as part of the former Argonne National Laboratory West Complex to support the operation of the second Experimental Breeder Reactor (EBR-II). It is part of a larger analytical laboratory structure that includes wet chemistry, instrumentation and radiochemistry laboratories. The purpose of the ALHC is to perform analytical chemistry work on highly radioactive materials. The primary work in the ALHC has traditionally been dissolution of nuclear materials so that less radioactive subsamples (aliquots) could be transferred to other sections of the laboratory for analysis. Over the last 50 years though, the capabilities within the ALHC have also become independent of other laboratory sections in a number of ways. While dissolution, digestion and subdividing samples are still a vitally important role, the ALHC has stand alone capabilities in the area of immersion density, gamma scanning and combustion gas analysis. Recent use of the ALHC for immersion density shows that extremely fine and delicate operations can be performed with the master-slave manipulators by qualified operators. Twenty milligram samples were tested for immersion density to determine the expansion of uranium dioxide after irradiation in a nuclear reactor. The data collected confirmed modeling analysis with very tight

  8. Protein C inhibits endocytosis of thrombin-thrombomodulin complexes in A549 lung cancer cells and human umbilical vein endothelial cells

    International Nuclear Information System (INIS)

    Maruyama, I.; Majerus, P.W.

    1987-01-01

    We investigated the effect of protein C on the endocytosis of thrombin-thrombomodulin complexes. We previously showed that exposure of umbilical vein endothelial cells to thrombin stimulated the internalization and degradation of thrombin. A similar internalization was stimulated by a monoclonal antithrombomodulin antibody. We have repeated these studies in the presence of protein C and found that endocytosis of 125 I-thrombin-thrombomodulin complexes, but not 125 I-antithrombomodulin-thrombomodulin complexes, is inhibited. Activated protein C did not inhibit endocytosis of thrombin-thrombomodulin complexes. Protein C inhibited both internalization and degradation of 125 I-thrombin and diisopropylphosphoryl (DIP) 125 I-thrombin in human lung cancer cells (A549). These effects were observed at protein C concentrations found in human plasma. Protein S had no effect on the inhibition of endocytosis of thrombin-thrombomodulin complexes by protein C. We propose that protein C may regulate the rate of endocytosis of thrombin-thrombomodulin complexes in vivo and thereby control the capacity for endothelium to activate protein C

  9. Human decidua-derived mesenchymal stem cells differentiate into functional alveolar type II-like cells that synthesize and secrete pulmonary surfactant complexes.

    Directory of Open Access Journals (Sweden)

    Alejandro Cerrada

    Full Text Available Lung alveolar type II (ATII cells are specialized in the synthesis and secretion of pulmonary surfactant, a lipid-protein complex that reduces surface tension to minimize the work of breathing. Surfactant synthesis, assembly and secretion are closely regulated and its impairment is associated with severe respiratory disorders. At present, well-established ATII cell culture models are not available. In this work, Decidua-derived Mesenchymal Stem Cells (DMSCs have been differentiated into Alveolar Type II- Like Cells (ATII-LCs, which display membranous cytoplasmic organelles resembling lamellar bodies, the organelles involved in surfactant storage and secretion by native ATII cells, and accumulate disaturated phospholipid species, a surfactant hallmark. Expression of characteristic ATII cells markers was demonstrated in ATII-LCs at gene and protein level. Mimicking the response of ATII cells to secretagogues, ATII-LCs were able to exocytose lipid-rich assemblies, which displayed highly surface active capabilities, including faster interfacial adsorption kinetics than standard native surfactant, even in the presence of inhibitory agents. ATII-LCs could constitute a highly useful ex vivo model for the study of surfactant biogenesis and the mechanisms involved in protein processing and lipid trafficking, as well as the packing and storage of surfactant complexes.

  10. Complexation of c6-ceramide with cholesteryl phosphocholine - a potent solvent-free ceramide delivery formulation for cells in culture.

    Directory of Open Access Journals (Sweden)

    Pramod Sukumaran

    Full Text Available Ceramides are potent bioactive molecules in cells. However, they are very hydrophobic molecules, and difficult to deliver efficiently to cells. We have made fluid bilayers from a short-chain D-erythro-ceramide (C6-Cer and cholesteryl phosphocholine (CholPC, and have used this as a formulation to deliver ceramide to cells. C6-Cer complexed with CholPC led to much larger biological effects in cultured cells (rat thyroid FRTL-5 and human HeLa cells in culture compared to C6-Cer dissolved in dimethyl sulfoxide (DMSO. Inhibition of cell proliferation and induction of apoptosis was significantly more efficient by C6-Cer/CholPC compared to C6-Cer dissolved in DMSO. C6-Cer/CholPC also permeated cell membranes and caused mitochondrial Ca(2+ influx more efficiently than C6-Cer in DMSO. Even though CholPC was taken up by cells to some extent (from C6-Cer/CholPC bilayers, and was partially hydrolyzed to free cholesterol (about 9%, none of the antiproliferative effects were due to CholPC or excess cholesterol. The ceramide effect was not limited to D-erythro-C6-Cer, since L-erythro-C6-Cer and D-erythro-C6-dihydroCer also inhibited cell priolifereation and affected Ca(2+ homeostasis. We conclude that C6-Cer complexed to CholPC increased the bioavailability of the short-chain ceramide for cells, and potentiated its effects in comparison to solvent-dissolved C6-Cer. This new ceramide formulation appears to be superior to previous solvent delivery approaches, and may even be useful with longer-chain ceramides.

  11. Bifacial dye-sensitized solar cells from covalent-bonded polyaniline-multiwalled carbon nanotube complex counter electrodes

    Science.gov (United States)

    Zhang, Huihui; He, Benlin; Tang, Qunwei; Yu, Liangmin

    2015-02-01

    Exploration of cost-effective counter electrodes (CEs) and enhancement of power conversion efficiency have been two persistent objectives for dye-sensitized solar cells (DSSCs). In the current work, polyaniline-multiwalled carbon nanotube (PANi-MWCNT) complexes are synthesized by a reflux method and employed as CE materials for bifacial DSSCs. Owing to the high optical transparency of PANi-MWCNT complex CE, the incident light from rear side can compensate for the incident light from TiO2 anode. The charge-transfer ability and electrochemical behaviors demonstrate the potential utilization of PANi-MWCNT complex CEs in robust bifacial DSSCs. The electrochemical properties as well as photovoltaic performances are optimized by adjusting MWCNT dosages. A maximum power conversion efficiency of 9.24% is recorded from the bifacial DSSC employing PANi-8 wt‰ MWCNT complex CE for both irradiation, which is better than 8.08% from pure PANi CE.

  12. Complex cells decrease errors for the Müller-Lyer illusion in a model of the visual ventral stream.

    Science.gov (United States)

    Zeman, Astrid; Obst, Oliver; Brooks, Kevin R

    2014-01-01

    To improve robustness in object recognition, many artificial visual systems imitate the way in which the human visual cortex encodes object information as a hierarchical set of features. These systems are usually evaluated in terms of their ability to accurately categorize well-defined, unambiguous objects and scenes. In the real world, however, not all objects and scenes are presented clearly, with well-defined labels and interpretations. Visual illusions demonstrate a disparity between perception and objective reality, allowing psychophysicists to methodically manipulate stimuli and study our interpretation of the environment. One prominent effect, the Müller-Lyer illusion, is demonstrated when the perceived length of a line is contracted (or expanded) by the addition of arrowheads (or arrow-tails) to its ends. HMAX, a benchmark object recognition system, consistently produces a bias when classifying Müller-Lyer images. HMAX is a hierarchical, artificial neural network that imitates the "simple" and "complex" cell layers found in the visual ventral stream. In this study, we perform two experiments to explore the Müller-Lyer illusion in HMAX, asking: (1) How do simple vs. complex cell operations within HMAX affect illusory bias and precision? (2) How does varying the position of the figures in the input image affect classification using HMAX? In our first experiment, we assessed classification after traversing each layer of HMAX and found that in general, kernel operations performed by simple cells increase bias and uncertainty while max-pooling operations executed by complex cells decrease bias and uncertainty. In our second experiment, we increased variation in the positions of figures in the input images that reduced bias and uncertainty in HMAX. Our findings suggest that the Müller-Lyer illusion is exacerbated by the vulnerability of simple cell operations to positional fluctuations, but ameliorated by the robustness of complex cell responses to such

  13. Engineering the cell surface display of cohesins for assembly of cellulosome-inspired enzyme complexes on Lactococcus lactis

    Directory of Open Access Journals (Sweden)

    Wieczorek Andrew S

    2010-09-01

    Full Text Available Abstract Background The assembly and spatial organization of enzymes in naturally occurring multi-protein complexes is of paramount importance for the efficient degradation of complex polymers and biosynthesis of valuable products. The degradation of cellulose into fermentable sugars by Clostridium thermocellum is achieved by means of a multi-protein "cellulosome" complex. Assembled via dockerin-cohesin interactions, the cellulosome is associated with the cell surface during cellulose hydrolysis, forming ternary cellulose-enzyme-microbe complexes for enhanced activity and synergy. The assembly of recombinant cell surface displayed cellulosome-inspired complexes in surrogate microbes is highly desirable. The model organism Lactococcus lactis is of particular interest as it has been metabolically engineered to produce a variety of commodity chemicals including lactic acid and bioactive compounds, and can efficiently secrete an array of recombinant proteins and enzymes of varying sizes. Results Fragments of the scaffoldin protein CipA were functionally displayed on the cell surface of Lactococcus lactis. Scaffolds were engineered to contain a single cohesin module, two cohesin modules, one cohesin and a cellulose-binding module, or only a cellulose-binding module. Cell toxicity from over-expression of the proteins was circumvented by use of the nisA inducible promoter, and incorporation of the C-terminal anchor motif of the streptococcal M6 protein resulted in the successful surface-display of the scaffolds. The facilitated detection of successfully secreted scaffolds was achieved by fusion with the export-specific reporter staphylococcal nuclease (NucA. Scaffolds retained their ability to associate in vivo with an engineered hybrid reporter enzyme, E. coli β-glucuronidase fused to the type 1 dockerin motif of the cellulosomal enzyme CelS. Surface-anchored complexes exhibited dual enzyme activities (nuclease and β-glucuronidase, and were

  14. Heterogeneity of Purkinje cell simple spike-complex spike interactions: zebrin- and non-zebrin-related variations.

    Science.gov (United States)

    Tang, Tianyu; Xiao, Jianqiang; Suh, Colleen Y; Burroughs, Amelia; Cerminara, Nadia L; Jia, Linjia; Marshall, Sarah P; Wise, Andrew K; Apps, Richard; Sugihara, Izumi; Lang, Eric J

    2017-08-01

    Cerebellar Purkinje cells (PCs) generate two types of action potentials, simple and complex spikes. Although they are generated by distinct mechanisms, interactions between the two spike types exist. Zebrin staining produces alternating positive and negative stripes of PCs across most of the cerebellar cortex. Thus, here we compared simple spike-complex spike interactions both within and across zebrin populations. Simple spike activity undergoes a complex modulation preceding and following a complex spike. The amplitudes of the pre- and post-complex spike modulation phases were correlated across PCs. On average, the modulation was larger for PCs in zebrin positive regions. Correlations between aspects of the complex spike waveform and simple spike activity were found, some of which varied between zebrin positive and negative PCs. The implications of the results are discussed with regard to hypotheses that complex spikes are triggered by rises in simple spike activity for either motor learning or homeostatic functions. Purkinje cells (PCs) generate two types of action potentials, called simple and complex spikes (SSs and CSs). We first investigated the CS-associated modulation of SS activity and its relationship to the zebrin status of the PC. The modulation pattern consisted of a pre-CS rise in SS activity, and then, following the CS, a pause, a rebound, and finally a late inhibition of SS activity for both zebrin positive (Z+) and negative (Z-) cells, though the amplitudes of the phases were larger in Z+ cells. Moreover, the amplitudes of the pre-CS rise with the late inhibitory phase of the modulation were correlated across PCs. In contrast, correlations between modulation phases across CSs of individual PCs were generally weak. Next, the relationship between CS spikelets and SS activity was investigated. The number of spikelets/CS correlated with the average SS firing rate only for Z+ cells. In contrast, correlations across CSs between spikelet numbers and the

  15. Titanium biomaterials with complex surfaces induced aberrant peripheral circadian rhythms in bone marrow mesenchymal stromal cells.

    Science.gov (United States)

    Hassan, Nathaniel; McCarville, Kirstin; Morinaga, Kenzo; Mengatto, Cristiane M; Langfelder, Peter; Hokugo, Akishige; Tahara, Yu; Colwell, Christopher S; Nishimura, Ichiro

    2017-01-01

    Circadian rhythms maintain a high level of homeostasis through internal feed-forward and -backward regulation by core molecules. In this study, we report the highly unusual peripheral circadian rhythm of bone marrow mesenchymal stromal cells (BMSCs) induced by titanium-based biomaterials with complex surface modifications (Ti biomaterial) commonly used for dental and orthopedic implants. When cultured on Ti biomaterials, human BMSCs suppressed circadian PER1 expression patterns, while NPAS2 was uniquely upregulated. The Ti biomaterials, which reduced Per1 expression and upregulated Npas2, were further examined with BMSCs harvested from Per1::luc transgenic rats. Next, we addressed the regulatory relationship between Per1 and Npas2 using BMSCs from Npas2 knockout mice. The Npas2 knockout mutation did not rescue the Ti biomaterial-induced Per1 suppression and did not affect Per2, Per3, Bmal1 and Clock expression, suggesting that the Ti biomaterial-induced Npas2 overexpression was likely an independent phenomenon. Previously, vitamin D deficiency was reported to interfere with Ti biomaterial osseointegration. The present study demonstrated that vitamin D supplementation significantly increased Per1::luc expression in BMSCs, though the presence of Ti biomaterials only moderately affected the suppressed Per1::luc expression. Available in vivo microarray data from femurs exposed to Ti biomaterials in vitamin D-deficient rats were evaluated by weighted gene co-expression network analysis. A large co-expression network containing Npas2, Bmal1, and Vdr was observed to form with the Ti biomaterials, which was disintegrated by vitamin D deficiency. Thus, the aberrant BMSC peripheral circadian rhythm may be essential for the integration of Ti biomaterials into bone.

  16. Retinal ganglion cell complex changes using spectral domain optical coherence tomography in diabetic patients without retinopathy

    Institute of Scientific and Technical Information of China (English)

    Ahmed; I.Hegazy; Rasha; H.Zedan; Tamer; A.Macky; Soheir; M.Esmat

    2017-01-01

    AIM:To assess the ganglion cell complex(GCC)thickness in diabetic eyes without retinopathy. METHODS:Two groups included 45 diabetic eyes without retinopathy and 21 non diabetic eyes. All subjects underwent full medical and ophthalmological history,full ophthalmological examination,measuring GCC thickness and central foveal thickness(CFT)using the RTVue~? spectral domainoptical coherence tomography(SD-OCT),and HbA1C level.RESULTS:GCC focal loss volume(FLV%)was significantly more in diabetic eyes(22.2% below normal)than normal eyes(P=0.024). No statistically significant difference was found between the diabetic group and the control group regarding GCC global loss volume(GLV%)(P=0.160). CFT was positively correlated to the average,superior and inferior GCC(P=0.001,0.000 and 0.001 respectively)and negatively correlated to GLV% and FLV%(P=0.002 and0.031 respectively)in diabetic eyes. C/D ratio in diabetic eyes was negatively correlated to average,superior and inferior GCC(P=0.015,0.007 and 0.017 respectively). The FLV% was negatively correlated to the refraction and level of Hb A1c(P=0.019 and 0.013 respectively)and positively correlated to the best corrected visual acuity(BCVA)in log MAR in diabetic group(P=0.004).CONCLUSION:Significant GCC thinning in diabetes predates retinal vasculopathy,which is mainly focal rather than diffuse. It has no preference to either the superior or inferior halves of the macula. Increase of myopic error is significantly accompanied with increased focal GCC loss. GCC loss is accompanied with increased C/D ratio in diabetic eyes.

  17. Retinal ganglion cell complex changes using spectral domain optical coherence tomography in diabetic patients without retinopathy

    Directory of Open Access Journals (Sweden)

    Ahmed I. Hegazy

    2017-03-01

    Full Text Available AIM: To assess the ganglion cell complex (GCC thickness in diabetic eyes without retinopathy. METHODS: Two groups included 45 diabetic eyes without retinopathy and 21 non diabetic eyes. All subjects underwent full medical and ophthalmological history, full ophthalmological examination, measuring GCC thickness and central foveal thickness (CFT using the RTVue® spectral domain-optical coherence tomography (SD-OCT, and HbA1C level. RESULTS: GCC focal loss volume (FLV% was significantly more in diabetic eyes (22.2% below normal than normal eyes (P=0.024. No statistically significant difference was found between the diabetic group and the control group regarding GCC global loss volume (GLV% (P=0.160. CFT was positively correlated to the average, superior and inferior GCC (P=0.001, 0.000 and 0.001 respectively and negatively correlated to GLV% and FLV% (P=0.002 and 0.031 respectively in diabetic eyes. C/D ratio in diabetic eyes was negatively correlated to average, superior and inferior GCC (P=0.015, 0.007 and 0.017 respectively. The FLV% was negatively correlated to the refraction and level of HbA1c (P=0.019 and 0.013 respectively and positively correlated to the best corrected visual acuity (BCVA in logMAR in diabetic group (P=0.004. CONCLUSION: Significant GCC thinning in diabetes predates retinal vasculopathy, which is mainly focal rather than diffuse. It has no preference to either the superior or inferior halves of the macula. Increase of myopic error is significantly accompanied with increased focal GCC loss. GCC loss is accompanied with increased C/D ratio in diabetic eyes.

  18. Calibrated complex impedance of CHO cells and E. coli bacteria at GHz frequencies using scanning microwave microscopy

    International Nuclear Information System (INIS)

    Tuca, Silviu-Sorin; Gramse, Georg; Kasper, Manuel; Oh, Yoo Jin; Zhu, Rong; Hinterdorfer, Peter; Badino, Giorgio; Brinciotti, Enrico; Rankl, Christian; Kienberger, Ferry

    2016-01-01

    The application of scanning microwave microscopy (SMM) to extract calibrated electrical properties of cells and bacteria in air is presented. From the S _1_1 images, after calibration, complex impedance and admittance images of Chinese hamster ovary cells and E. coli bacteria deposited on a silicon substrate have been obtained. The broadband capabilities of SMM have been used to characterize the bio-samples between 2 GHz and 20 GHz. The resulting calibrated cell and bacteria admittance at 19 GHz were Y _c_e_l_l = 185 μS + j285 μS and Y _b_a_c_t_e_r_i_a = 3 μS + j20 μS, respectively. A combined circuitry-3D finite element method EMPro model has been developed and used to investigate the frequency response of the complex impedance and admittance of the SMM setup. Based on a proposed parallel resistance–capacitance model, the equivalent conductance and parallel capacitance of the cells and bacteria were obtained from the SMM images. The influence of humidity and frequency on the cell conductance was experimentally studied. To compare the cell conductance with bulk water properties, we measured the imaginary part of the bulk water loss with a dielectric probe kit in the same frequency range resulting in a high level of agreement. (paper)

  19. Dectin-1 is essential for reverse transcytosis of glycosylated SIgA-antigen complexes by intestinal M cells.

    Directory of Open Access Journals (Sweden)

    Nicolas Rochereau

    2013-09-01

    Full Text Available Intestinal microfold (M cells possess a high transcytosis capacity and are able to transport a broad range of materials including particulate antigens, soluble macromolecules, and pathogens from the intestinal lumen to inductive sites of the mucosal immune system. M cells are also the primary pathway for delivery of secretory IgA (SIgA to the gut-associated lymphoid tissue. However, although the consequences of SIgA uptake by M cells are now well known and described, the mechanisms whereby SIgA is selectively bound and taken up remain poorly understood. Here we first demonstrate that both the Cα1 region and glycosylation, more particularly sialic acid residues, are involved in M cell-mediated reverse transcytosis. Second, we found that SIgA is taken up by M cells via the Dectin-1 receptor, with the possible involvement of Siglec-5 acting as a co-receptor. Third, we establish that transcytosed SIgA is taken up by mucosal CX3CR1⁺ dendritic cells (DCs via the DC-SIGN receptor. Fourth, we show that mucosal and systemic antibody responses against the HIV p24-SIgA complexes administered orally is strictly dependent on the expression of Dectin-1. Having deciphered the mechanisms leading to specific targeting of SIgA-based Ag complexes paves the way to the use of such a vehicle for mucosal vaccination against various infectious diseases.

  20. TLR2-Modulating Lipoproteins of the Mycobacterium tuberculosis Complex Enhance the HIV Infectivity of CD4+ T Cells.

    Science.gov (United States)

    Skerry, Ciaran; Klinkenberg, Lee G; Page, Kathleen R; Karakousis, Petros C

    2016-01-01

    Co-infection with Mycobacterium tuberculosis accelerates progression from HIV to AIDS. Our previous studies showed that M. tuberculosis complex, unlike M. smegmatis, enhances TLR2-dependent susceptibility of CD4+ T cells to HIV. The M. tuberculosis complex produces multiple TLR2-stimulating lipoproteins, which are absent in M. smegmatis. M. tuberculosis production of mature lipoproteins and TLR2 stimulation is dependent on cleavage by lipoprotein signal peptidase A (LspA). In order to determine the role of potential TLR2-stimulating lipoproteins on mycobacterial-mediated HIV infectivity of CD4+ T cells, we generated M. smegmatis recombinant strains overexpressing genes encoding various M. bovis BCG lipoproteins, as well as a Mycobacterium bovis BCG strain deficient in LspA (ΔlspA). Exposure of human peripheral blood mononuclear cells (PBMC) to M. smegmatis strains overexpressing the BCG lipoproteins, LprF (p<0.01), LprH (p<0.05), LprI (p<0.05), LprP (p<0.001), LprQ (p<0.005), MPT83 (p<0.005), or PhoS1 (p<0.05), resulted in increased HIV infectivity of CD4+ T cells isolated from these PBMC. Conversely, infection of PBMC with ΔlspA reduced HIV infectivity of CD4+ T cells by 40% relative to BCG-infected cells (p<0.05). These results may have important implications for TB vaccination programs in areas with high mother-to-child HIV transmission.

  1. Poly(amido)amine (PAMAM) dendrimer–cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    International Nuclear Information System (INIS)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath

    2013-01-01

    Dendrimer–cisplatin complexes were prepared using PAMAM dendrimers with terminal –NH 2 and –COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer–cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was ∼11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC 50 values in ovarian cancer cells when compared with carboxylate surface dendrimers (p < 0.05). A correlation was observed among cytotoxicity of the complexes, cellular uptake, and platinum–DNA adduct formation. Treatment with dendrimer–cisplatin complexes resulted in a 7.0-fold increase (p < 0.05) in expression of apoptotic genes (Bcl2, Bax, p53) and 13.2- to 27.1-fold increase (p < 0.05) in the activity of caspases 3, 8, and 9 in vitro. Results suggest that PAMAM dendrimers can be used as potential carrier for cisplatin chemotherapy of ovarian cancer

  2. Poly(amido)amine (PAMAM) dendrimer-cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath, E-mail: palakurthi@tamhsc.edu [Texas A and M Health Science Center, Irma Lerma Rangel College of Pharmacy (United States)

    2013-09-15

    Dendrimer-cisplatin complexes were prepared using PAMAM dendrimers with terminal -NH{sub 2} and -COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer-cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was {approx}11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC{sub 50} values in ovarian cancer cells when compared with carboxylate surface dendrimers (p < 0.05). A correlation was observed among cytotoxicity of the complexes, cellular uptake, and platinum-DNA adduct formation. Treatment with dendrimer-cisplatin complexes resulted in a 7.0-fold increase (p < 0.05) in expression of apoptotic genes (Bcl2, Bax, p53) and 13.2- to 27.1-fold increase (p < 0.05) in the activity of caspases 3, 8, and 9 in vitro. Results suggest that PAMAM dendrimers can be used as potential carrier for cisplatin chemotherapy of ovarian cancer.

  3. Major Histocompatibilty Complex-Restricted Adaptive Immune Responses to CT26 Colon Cancer Cell Line in Mixed Allogeneic Chimera.

    Science.gov (United States)

    Lee, K W; Choi, B; Kim, Y M; Cho, C W; Park, H; Moon, J I; Choi, G-S; Park, J B; Kim, S J

    2017-06-01

    Although the induction of mixed allogeneic chimera shows promising clinical tolerance results in organ transplantation, its clinical relevance as an anti-cancer therapy is yet unknown. We introduced a mixed allogenic chimera setting with the use of a murine colon cancer cell line, CT26, by performing double bone marrow transplantation. We analyzed donor- and recipient-restricted anti-cancer T-cell responses, and phenotypes of subpopulations of T cells. The protocol involves challenging 1 × 10 5 cells of CT26 cells intra-hepatically on day 50 after bone marrow transplantation, and, by use of CT26 lysates and an H-2L d -restricted AH1 pentamer, flow cytometric analysis was performed to detect the generation of cancer-specific CD4 + and CD8 + T cells at various time points. We found that immunocompetence against tumors depends heavily on cancer-specific CD8 + T-cell responses in a major histocompatibility complex-restricted manner; the evidence was further supported by the increase of interferon-γ-secreting CD4 + T cells. Moreover, we demonstrated that during the effector immune response to CT26 cancer challenge, there was a presence of central memory cells (CD62L hi CCR7 + ) as well as effector memory cells (CD62L lo CCR7 - ). Moreover, mixed allogeneic chimeras (BALB/c to C56BL/6 or vice versa) showed similar or heightened immune responses to CT26 cells compared with that of wild-type mice. Our results suggest that the responses of primary immunocompetency and of pre-existing memory T cells against allogeneic cancer are sustained and preserved long-term in a mixed allogeneic chimeric environment. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. A bioaccumulative cyclometalated platinum(II) complex with two-photon-induced emission for live cell imaging.

    Science.gov (United States)

    Koo, Chi-Kin; Wong, Ka-Leung; Man, Cornelia Wing-Yin; Lam, Yun-Wah; So, Leo King-Yan; Tam, Hoi-Lam; Tsao, Sai-Wah; Cheah, Kok-Wai; Lau, Kai-Chung; Yang, Yang-Yi; Chen, Jin-Can; Lam, Michael Hon-Wah

    2009-02-02

    The cyclometalated platinum(II) complex [Pt(L)Cl], where HL is a new cyclometalating ligand 2-phenyl-6-(1H-pyrazol-3-yl)pyridine containing C(phenyl), N(pyridyl), and N(pyrazolyl) donor moieties, was found to possess two-photon-induced luminescent properties. The two-photon-absorption cross section of the complex in N,N-dimethylformamide at room temperature was measured to be 20.8 GM. Upon two-photon excitation at 730 nm from a Ti:sapphire laser, bright-green emission was observed. Besides its two-photon-induced luminescent properties, [Pt(L)Cl] was able to be rapidly accumulated in live HeLa and NIH3T3 cells. The two-photon-induced luminescence of the complex was retained after live cell internalization and can be observed by two-photon confocal microscopy. Its bioaccumulation properties enabled time-lapse imaging of the internalization process of the dye into living cells. Cytotoxicity of [Pt(L)Cl] to both tested cell lines was low, according to MTT assays, even at loadings as high as 20 times the dose concentration for imaging for 6 h.

  5. Design of hypoxia-targeting radiopharmaceuticals: selective uptake of copper-64 complexes in hypoxic cells in vitro

    International Nuclear Information System (INIS)

    Dearling, J.L.J.; Lewis, J.S.; Mullen, G.E.D.; Rae, M.T.; Zweit, J.; Blower, P.J.

    1998-01-01

    The well-known perfusion tracer CuPTSM, labelled with 62 Cu or 64 Cu, is believed to be trapped in cells non-selectively by a bioreductive mechanism. It is proposed that by modifying the ligand to increase its electron donor strength (for example by adding alkyl functionality or replacing sulphur ligands with oxygen ligands), the copper complexes will become less easily reduced and tracers with selectivity for hypoxic tissues could thus be developed. The aim of this work was to prepare 64 Cu-labelled complexes of two series of ligands, based on the bis(thiosemicarbazone) (13 ligands) and bis(salicylaldimine) (3 ligands) skeletons, and to evaluate the hypoxia dependence of their uptake in cells. The complexes were incubated with Chinese hamster ovary cells under normoxic and hypoxic conditions, and the cells isolated by centrifugation to determine radioactivity uptake at various time points up to 90 min. Several members of both series demonstrated significant (P 60 Cu, 61 Cu, 62 Cu, 64 Cu) and targeted radiotherapy ( 64 Cu, 67 Cu). (orig.)

  6. Recruitment of Mediator Complex by Cell Type and Stage-Specific Factors Required for Tissue-Specific TAF Dependent Gene Activation in an Adult Stem Cell Lineage.

    Science.gov (United States)

    Lu, Chenggang; Fuller, Margaret T

    2015-12-01

    Onset of terminal differentiation in adult stem cell lineages is commonly marked by robust activation of new transcriptional programs required to make the appropriate differentiated cell type(s). In the Drosophila male germ line stem cell lineage, the switch from proliferating spermatogonia to spermatocyte is accompanied by one of the most dramatic transcriptional changes in the fly, as over 1000 new transcripts turn on in preparation for meiosis and spermatid differentiation. Here we show that function of the coactivator complex Mediator is required for activation of hundreds of new transcripts in the spermatocyte program. Mediator appears to act in a sequential hierarchy, with the testis activating Complex (tMAC), a cell type specific form of the Mip/dREAM general repressor, required to recruit Mediator subunits to the chromatin, and Mediator function required to recruit the testis TAFs (tTAFs), spermatocyte specific homologs of subunits of TFIID. Mediator, tMAC and the tTAFs co-regulate expression of a major set of spermatid differentiation genes. The Mediator subunit Med22 binds the tMAC component Topi when the two are coexpressed in S2 cells, suggesting direct recruitment. Loss of Med22 function in spermatocytes causes meiosis I maturation arrest male infertility, similar to loss of function of the tMAC subunits or the tTAFs. Our results illuminate how cell type specific versions of the Mip/dREAM complex and the general transcription machinery cooperate to drive selective gene activation during differentiation in stem cell lineages.

  7. Recruitment of Mediator Complex by Cell Type and Stage-Specific Factors Required for Tissue-Specific TAF Dependent Gene Activation in an Adult Stem Cell Lineage.

    Directory of Open Access Journals (Sweden)

    Chenggang Lu

    2015-12-01

    Full Text Available Onset of terminal differentiation in adult stem cell lineages is commonly marked by robust activation of new transcriptional programs required to make the appropriate differentiated cell type(s. In the Drosophila male germ line stem cell lineage, the switch from proliferating spermatogonia to spermatocyte is accompanied by one of the most dramatic transcriptional changes in the fly, as over 1000 new transcripts turn on in preparation for meiosis and spermatid differentiation. Here we show that function of the coactivator complex Mediator is required for activation of hundreds of new transcripts in the spermatocyte program. Mediator appears to act in a sequential hierarchy, with the testis activating Complex (tMAC, a cell type specific form of the Mip/dREAM general repressor, required to recruit Mediator subunits to the chromatin, and Mediator function required to recruit the testis TAFs (tTAFs, spermatocyte specific homologs of subunits of TFIID. Mediator, tMAC and the tTAFs co-regulate expression of a major set of spermatid differentiation genes. The Mediator subunit Med22 binds the tMAC component Topi when the two are coexpressed in S2 cells, suggesting direct recruitment. Loss of Med22 function in spermatocytes causes meiosis I maturation arrest male infertility, similar to loss of function of the tMAC subunits or the tTAFs. Our results illuminate how cell type specific versions of the Mip/dREAM complex and the general transcription machinery cooperate to drive selective gene activation during differentiation in stem cell lineages.

  8. Anti-cancer effects of newly developed chemotherapeutic agent, glycoconjugated palladium (II) complex, against cisplatin-resistant gastric cancer cells

    International Nuclear Information System (INIS)

    Tanaka, Mamoru; Kamiya, Takeshi; Joh, Takashi; Kataoka, Hiromi; Yano, Shigenobu; Ohi, Hiromi; Kawamoto, Keisuke; Shibahara, Takashi; Mizoshita, Tsutomu; Mori, Yoshinori; Tanida, Satoshi

    2013-01-01

    Cisplatin (CDDP) is the most frequently used chemotherapeutic agent for various types of advanced cancer, including gastric cancer. However, almost all cancer cells acquire resistance against CDDP, and this phenomenon adversely affects prognosis. Thus, new chemotherapeutic agents that can overcome the CDDP-resistant cancer cells will improve the survival of advanced cancer patients. We synthesized new glycoconjugated platinum (II) and palladium (II) complexes, [PtCl 2 (L)] and [PdCl 2 (L)]. CDDP-resistant gastric cancer cell lines were established by continuous exposure to CDDP, and gene expression in the CDDP-resistant gastric cancer cells was analyzed. The cytotoxicity and apoptosis induced by [PtCl 2 (L)] and [PdCl 2 (L)] in CDDP-sensitive and CDDP-resistant gastric cancer cells were evaluated. DNA double-strand breaks by drugs were assessed by evaluating phosphorylated histone H2AX. Xenograft tumor mouse models were established and antitumor effects were also examined in vivo. CDDP-resistant gastric cancer cells exhibit ABCB1 and CDKN2A gene up-regulation, as compared with CDDP-sensitive gastric cancer cells. In the analyses of CDDP-resistant gastric cancer cells, [PdCl 2 (L)] overcame cross-resistance to CDDP in vitro and in vivo. [PdCl 2 (L)] induced DNA double-strand breaks. These results indicate that [PdCl 2 (L)] is a potent chemotherapeutic agent for CDDP-resistant gastric cancer and may have clinical applications

  9. CLL cells are resistant to smac mimetics because of an inability to form a ripoptosome complex

    NARCIS (Netherlands)

    Maas, C.; Tromp, J. M.; van Laar, J.; Thijssen, R.; Elias, J. A.; Malara, A.; Krippner-Heidenreich, A.; Silke, J.; van Oers, M. H. J.; Eldering, E.

    2013-01-01

    In the lymph node (LN) environment, chronic lymphocytic leukemia (CLL) cells display increased NF-κB activity compared with peripheral blood CLL cells, which contributes to chemoresistance. Antagonists of cellular inhibitor of apoptosis proteins (cIAPs) can induce apoptosis in various cancer cells

  10. Epithelial Cell-Neutrophil Interactions in the Alimentary Tract: A Complex Dialog in Mucosal Surveillance and Inflammation

    Directory of Open Access Journals (Sweden)

    Sean P. Colgan

    2002-01-01

    Full Text Available Inflammatory diseases of mucosal organs as diverse as the lung, kidney, and intestine, inevitably require the intimate interactions of neutrophils with columnar epithelia. The physiologic consequences of such interactions often determine endpoint organ function, and for this reason, much recent interest has developed in identifying mechanisms and novel targets for the treatment of mucosal inflammation. Elegant in vitro model systems incorporating purified human neutrophils and human epithelial cells grown in physiologic orientations have aided in discovery of new and insightful pathways to define basic inflammatory pathways. Here, we will review the recent literature regarding the interactions between columnar epithelial cells and neutrophils, with an emphasis on intestinal epithelial cells, structural aspects of neutrophil transepithelial migration, molecular determinants of neutrophil-epithelial cell interactions, as well as modulation of these pathways. These recent studies highlight the dynamic nature of these pathways and lend insight into the complexity of treating mucosal inflammation.

  11. Stem cell factor induces phosphatidylinositol 3'-kinase-dependent Lyn/Tec/Dok-1 complex formation in hematopoietic cells

    NARCIS (Netherlands)

    van Dijk, T. B.; van den Akker, E.; Amelsvoort, M. P.; Mano, H.; Löwenberg, B.; von Lindern, M.

    2000-01-01

    Stem cell factor (SCF) has an important role in the proliferation, differentiation, survival, and migration of hematopoietic cells. SCF exerts its effects by binding to cKit, a receptor with intrinsic tyrosine kinase activity. Activation of phosphatidylinositol 3'-kinase (PI3-K) by cKit was

  12. Biologically Complex Planar Cell Plasma Membranes Supported on Polyelectrolyte Cushions Enhance Transmembrane Protein Mobility and Retain Native Orientation.

    Science.gov (United States)

    Liu, Han-Yuan; Chen, Wei-Liang; Ober, Christopher K; Daniel, Susan

    2018-01-23

    Reconstituted supported lipid bilayers (SLB) are widely used as in vitro cell-surface models because they are compatible with a variety of surface-based analytical techniques. However, one of the challenges of using SLBs as a model of the cell surface is the limited complexity in membrane composition, including the incorporation of transmembrane proteins and lipid diversity that may impact the activity of those proteins. Additionally, it is challenging to preserve the transmembrane protein native orientation, function, and mobility in SLBs. Here, we leverage the interaction between cell plasma membrane vesicles and polyelectrolyte brushes to create planar bilayers from cell plasma membrane vesicles that have budded from the cell surface. This approach promotes the direct incorporation of membrane proteins and other species into the planar bilayer without using detergent or reconstitution and preserves membrane constituents. Furthermore, the structure of the polyelectrolyte brush serves as a cushion between the planar bilayer and rigid supporting surface, limiting the interaction of the cytosolic domains of membrane proteins with this surface. Single particle tracking was used to analyze the motion of GPI-linked yellow fluorescent proteins (GPI-YFP) and neon-green fused transmembrane P2X2 receptors (P2X2-neon) and shows that this platform retains over 75% mobility of multipass transmembrane proteins in its native membrane environment. An enzyme accessibility assay confirmed that the protein orientation is preserved and results in the extracellular domain facing toward the bulk phase and the cytosolic side facing the support. Because the platform presented here retains the complexity of the cell plasma membrane and preserves protein orientation and mobility, it is a better representative mimic of native cell surfaces, which may find many applications in biological assays aimed at understanding cell membrane phenomena.

  13. Complex cells decrease errors for the Müller-Lyer illusion in a model of the visual ventral stream

    Directory of Open Access Journals (Sweden)

    Astrid eZeman

    2014-09-01

    Full Text Available To improve robustness in object recognition, many artificial visual systems imitate the way in which the human visual cortex encodes object information as a hierarchical set of features. These systems are usually evaluated in terms of their ability to accurately categorize well-defined, unambiguous objects and scenes. In the real world, however, not all objects and scenes are presented clearly, with well-defined labels and interpretations. Visual illusions demonstrate a disparity between perception and objective reality, allowing psychophysicists to methodically manipulate stimuli and study our interpretation of the environment. One prominent effect, the Müller-Lyer illusion, is demonstrated when the perceived length of a line is contracted (or expanded by the addition of arrowheads (or arrow-tails to its ends. HMAX, a benchmark object recognition system, consistently produces a bias when classifying Müller-Lyer images. HMAX is a hierarchical, artificial neural network that imitates the ‘simple’ and ‘complex’ cell layers found in the visual ventral stream. In this study, we perform two experiments to explore the Müller-Lyer illusion in HMAX, asking: 1 How do simple versus complex cell operations within HMAX affect illusory bias and precision? 2 How does varying the position of the figures in the input image affect classification using HMAX? In our first experiment, we assessed classification after traversing each layer of HMAX and found that in general, kernel operations performed by simple cells increase bias and uncertainty while max-pooling operations executed by complex cells decrease bias and uncertainty. In our second experiment, we increased variation in the positions of figures in the input that reduced bias and uncertainty in HMAX. Our findings suggest that the Müller-Lyer illusion is exacerbated by the vulnerability of simple cell operations to positional fluctuations, but ameliorated by the robustness of complex cell

  14. Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

    DEFF Research Database (Denmark)

    Mustafa, A S; Amoudy, H A; Wiker, H G

    1998-01-01

    We have screened peripheral blood mononuclear cells (PBMC) from tuberculosis (TB) patients for proliferative reactivity and interferon-gamma (IFN-gamma) secretion against a panel of purified recombinant (r) and natural (n) culture filtrate (rESAT-6, nMPT59, nMPT64 and nMPB70) and somatic-derived (r......GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

  15. Dynamic Recruitment of Functionally Distinct Swi/Snf Chromatin Remodeling Complexes Modulates Pdx1 Activity in Islet β Cells

    Directory of Open Access Journals (Sweden)

    Brian McKenna

    2015-03-01

    Full Text Available Pdx1 is a transcription factor of fundamental importance to pancreas formation and adult islet β cell function. However, little is known about the positive- and negative-acting coregulators recruited to mediate transcriptional control. Here, we isolated numerous Pdx1-interacting factors possessing a wide range of cellular functions linked with this protein, including, but not limited to, coregulators associated with transcriptional activation and repression, DNA damage response, and DNA replication. Because chromatin remodeling activities are essential to developmental lineage decisions and adult cell function, our analysis focused on investigating the influence of the Swi/Snf chromatin remodeler on Pdx1 action. The two mutually exclusive and indispensable Swi/Snf core ATPase subunits, Brg1 and Brm, distinctly affected target gene expression in β cells. Furthermore, physiological and pathophysiological conditions dynamically regulated Pdx1 binding to these Swi/Snf complexes in vivo. We discuss how context-dependent recruitment of coregulatory complexes by Pdx1 could impact pancreas cell development and adult islet β cell activity.

  16. miR-375 is highly expressed and possibly transactivated by achaete-scute complex homolog 1 in small-cell lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    Huijie Zhao; Lei Zhu; Yujuan Jin; Hongbin Ji; Xiumin Yan; Xueliang Zhu

    2012-01-01

    In this study,we identified five miRNAs highly expressed in the small-cell lung cancer (SCLC) cell line NCI-H209.Among them,the expression levels of miR-375 were dramatically elevated in all SCLC cell lines examined,coincident with the expression of the transcription factor achaete-scute complex homolog 1 (ASCL1).Moreover,miR-375 was upregulated and correlated with ASCL1 in the cell lines generated from mouse SCLC-like tumors as well.Dual-luciferase assays further showed that ASCL1 activated the expression of miR-375 by binding to the three E-box elements in the miR-375 promoter.These results imply a role of ASCL1 in SCLC via the upregulation of miR-375.

  17. CRISPR-Cas9 mediated genetic engineering for the purification of the endogenous integrator complex from mammalian cells.

    Science.gov (United States)

    Baillat, David; Russell, William K; Wagner, Eric J

    2016-12-01

    The Integrator Complex (INT) is a large multi-subunit protein complex, containing at least 14 subunits and a host of associated factors. These protein components have been established through pulldowns of overexpressed epitope tagged subunits or by using antibodies raised against specific subunits. Here, we utilize CRISPR/Cas9 gene editing technology to introduce N-terminal FLAG epitope tags into the endogenous genes that encode Integrator subunit 4 and 11 within HEK293T cells. We provide specific details regarding design, approaches for facile screening, and our observed frequency of successful recombination. Finally, using silver staining, Western blotting and LC-MS/MS we compare the components of INT of purifications from CRISPR derived lines to 293T cells overexpressing FLAG-INTS11 to define a highly resolved constituency of mammalian INT. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. The Cell Wall Teichuronic Acid Synthetase (TUAS Is an Enzyme Complex Located in the Cytoplasmic Membrane of Micrococcus luteus

    Directory of Open Access Journals (Sweden)

    Lingyi Lynn Deng

    2010-01-01

    composed of disaccharide repeating units [-4-β-D-ManNAcAp-(1→6α-D-Glcp−1-]n, which is covalently anchored to the peptidoglycan on the inner cell wall and extended to the outer surface of the cell envelope. An enzyme complex responsible for the TUA chain biosynthesis was purified and characterized. The 440 kDa enzyme complex, named teichuronic acid synthetase (TUAS, is an octomer composed of two kinds of glycosyltransferases, Glucosyltransferase, and ManNAcA-transferase, which is capable of catalyzing the transfer of disaccharide glycosyl residues containing both glucose and the N-acetylmannosaminuronic acid residues. TUAS displays hydrophobic properties and is found primarily associated with the cytoplasmic membrane. The purified TUAS contains carotinoids and lipids. TUAS activity is diminished by phospholipase digestion. We propose that TUAS serves as a multitasking polysaccharide assembling station on the bacterial membrane.

  19. NADPH oxidase complex-derived reactive oxygen species, the actin cytoskeleton, and rho GTPases in cell migration

    DEFF Research Database (Denmark)

    Stanley, Alanna; Thompson, Kerry; Hynes, Ailish

    2014-01-01

    Abstract Significance: Rho GTPases are historically known to be central regulators of actin cytoskeleton reorganization. This affects many processes including cell migration. In addition, members of the Rac subfamily are known to be involved in reactive oxygen species (ROS) production through...... mediating cytoskeletal reorganization. Critical Issues: The role of the actin cytoskeleton in providing a scaffold for components of the Nox complex needs to be examined in the light of these new advances. During cell migration, Rho GTPases, ROS, and cytoskeletal organization appear to function as a complex...... compartments. This in conjunction with the analysis of tissues lacking specific Rho GTPases, and Nox components will facilitate a detailed examination of the interactions of these structures with the actin cytoskeleton. In combination with the analysis of ROS production, including its subcellular location...

  20. Generation of tooth-periodontium complex structures using high-odontogenic potential dental epithelium derived from mouse embryonic stem cells.

    Science.gov (United States)

    Zhang, Yancong; Li, Yongliang; Shi, Ruirui; Zhang, Siqi; Liu, Hao; Zheng, Yunfei; Li, Yan; Cai, Jinglei; Pei, Duanqing; Wei, Shicheng

    2017-06-08

    A number of studies have shown that tooth-like structures can be regenerated using induced pluripotent stem cells and mouse embryonic stem (mES) cells. However, few studies have reported the regeneration of tooth-periodontium complex structures, which are more suitable for clinical tooth transplantation. We established an optimized approach to induce high-odontogenic potential dental epithelium derived from mES cells by temporally controlling bone morphogenic protein 4 (BMP4) function and regenerated tooth-periodontium complex structures in vivo. First, immunofluorescence and quantitative reverse transcription-polymerase chain reaction were used to identify the watershed of skin and the oral ectoderm. LDN193189 was then used to inhibit the BMP4 receptor around the watershed, followed by the addition of exogenous BMP4 to promote BMP4 function. The generated dental epithelium was confirmed by western blot analysis and immunofluorescence. The generated epithelium was ultimately combined with embryonic day 14.5 mouse mesenchyme and transplanted into the renal capsules of nude mice. After 4 weeks, the tooth-periodontium complex structure was examined by micro-computed tomography (CT) and hematoxylin and eosin (H&E) staining. Our study found that the turning point of oral ectoderm differentiation occurred around day 3 after the embryoid body was transferred to a common culture plate. Ameloblastin-positive dental epithelial cells were detected following the temporal regulation of BMP4. Tooth-periodontium complex structures, which included teeth, a periodontal membrane, and alveolar bone, were formed when this epithelium was combined with mouse dental mesenchyme and transplanted into the renal capsules of nude mice. Micro-CT and H&E staining revealed that the generated tooth-periodontium complex structures shared a similar histological structure with normal mouse teeth. An optimized induction method was established to promote the differentiation of mES cells into dental

  1. The LH2 complexes are assembled in the cells of purple sulfur bacterium Ectothiorhodospira haloalkaliphila with inhibition of carotenoid biosynthesis.

    Science.gov (United States)

    Ashikhmin, Aleksandr; Makhneva, Zoya; Moskalenko, Andrey

    2014-03-01

    The effect of the inhibitor of carotenoid (Car) biosynthesis, diphenylamine (DPA), on the cells of the purple sulfur bacterium Ectothiorhodospira (Ect.) haloalkaliphila has been studied. There occurs an inhibition of the biosynthesis of colored Cars (≥99 %) at 71 μM DPA. Considering "empty" Car pockets (Moskalenko and Makhneva 2012) the content of Cars in the DPA-treated samples is first calculated more correctly. The total content of the colored Cars in the sample at 71 μM DPA does not exceed 1 % of the wild type. In the DPA-treated cells (membranes) a complete set of pigment-protein complexes is retained. The LH2 complex at 71 μM DPA is isolated, which is identical to the LH2 complex of the wild type in near IR absorption spectra. This suggests that the principles for assembling this LH2 complex in vivo in the absence of colored Cars remain the same. These results are in full agreement with the data obtained earlier for Allochromatium (Alc.) minutissimum (Moskalenko and Makhneva 2012). They are as follows: (1) DPA almost entirely inhibits the biosynthesis of the colored Cars in Ect. haloalkaliphila cells. (2) In the DPA-treated samples non-colored Cars are detected at 53.25 μM DPA (as traces) and at 71 μM DPA. (3) DPA may affect both phytoene synthase (at ≤71 μM DPA) and phytoene desaturase (at ≥53.25 μM DPA). (4) The assembly of LH2 complex does occur without any colored Cars.

  2. Modeling the Excess Cell Surface Stored in a Complex Morphology of Bleb-Like Protrusions.

    Directory of Open Access Journals (Sweden)

    Maryna Kapustina

    2016-03-01

    Full Text Available Cells transition from spread to rounded morphologies in diverse physiological contexts including mitosis and mesenchymal-to-amoeboid transitions. When these drastic shape changes occur rapidly, cell volume and surface area are approximately conserved. Consequently, the rounded cells are suddenly presented with a several-fold excess of cell surface whose area far exceeds that of a smooth sphere enclosing the cell volume. This excess is stored in a population of bleb-like protrusions (BLiPs, whose size distribution is shown by electron micrographs to be skewed. We introduce three complementary models of rounded cell morphologies with a prescribed excess surface area. A 2D Hamiltonian model provides a mechanistic description of how discrete attachment points between the cell surface and cortex together with surface bending energy can generate a morphology that satisfies a prescribed excess area and BLiP number density. A 3D random seed-and-growth model simulates efficient packing of BLiPs over a primary rounded shape, demonstrating a pathway for skewed BLiP size distributions that recapitulate 3D morphologies. Finally, a phase field model (2D and 3D posits energy-based constitutive laws for the cell membrane, nematic F-actin cortex, interior cytosol, and external aqueous medium. The cell surface is equipped with a spontaneous curvature function, a proxy for the cell surface-cortex couple, that is a priori unknown, which the model "learns" from the thin section transmission electron micrograph image (2D or the "seed and growth" model image (3D. Converged phase field simulations predict self-consistent amplitudes and spatial localization of pressure and stress throughout the cell for any posited stationary morphology target and cell compartment constitutive properties. The models form a general framework for future studies of cell morphological dynamics in a variety of biological contexts.

  3. Long-term drug modification to the surface of mesenchymal stem cells by the avidin-biotin complex method.

    Science.gov (United States)

    Takayama, Yukiya; Kusamori, Kosuke; Hayashi, Mika; Tanabe, Noriko; Matsuura, Satoru; Tsujimura, Mari; Katsumi, Hidemasa; Sakane, Toshiyasu; Nishikawa, Makiya; Yamamoto, Akira

    2017-12-05

    Mesenchymal stem cells (MSCs) have various functions, making a significant contribution to tissue repair. On the other hand, the viability and function of MSCs are not lasting after an in vivo transplant, and the therapeutic effects of MSCs are limited. Although various chemical modification methods have been applied to MSCs to improve their viability and function, most of conventional drug modification methods are short-term and unstable and cause cytotoxicity. In this study, we developed a method for long-term drug modification to C3H10T1/2 cells, murine mesenchymal stem cells, without any damage, using the avidin-biotin complex method (ABC method). The modification of NanoLuc luciferase (Nluc), a reporter protein, to C3H10T1/2 cells by the ABC method lasted for at least 14 days in vitro without major effects on the cellular characteristics (cell viability, cell proliferation, migration ability, and differentiation ability). Moreover, in vivo, the surface Nluc modification to C3H10T1/2 cells by the ABC method lasted for at least 7 days. Therefore, these results indicate that the ABC method may be useful for long-term surface modification of drugs and for effective MSC-based therapy.

  4. Stage-specific damage to synaptonemal complexes and metaphase chromosomes induced by X rays in male mouse germ cells

    International Nuclear Information System (INIS)

    Backer, L.C.; Sontag, M.R.; Allen, J.W.

    1991-01-01

    Synaptonemal complexes (SCs) reveal mutagen-induced effects in germ cell meiotic chromosomes. The study was aimed at characterizing relationships between SC and metaphase I chromosome damage following radiation exposure at various stages of spermatogenesis. Male mice were irradiated with doses of 0, 2, or 4 Gy, and spermatocytes were harvested at times consistent with earlier exposures as spermatogonial stem cells, preleptotene cells (premeiotic DNA synthesis), or meiotic prophase cells. After stem-cell exposure, twice as many rearrangements were observed in SCs as in metaphase I chromosomes. Irradiation during premeiotic DNA synthesis resulted in dose-related increases in SC breakage and rearrangements (including novel forms) and in metaphase chromosomal aberrations. Following prophase exposure, various types and levels of SC and metaphase damage were observed. Irradiation of zygotene cells led to high frequencies of chromosome multivalents in metaphase I without a correspondingly high level of damage in preceding prophase SCs. Thus, irradiation of premeiotic and meiotic cells results in variable relationships between SC and metaphase chromosome damage

  5. Sedimentation properties of DNA-membrane complexes and yield of DNA breaks at irradiation of mammalian cells

    International Nuclear Information System (INIS)

    Erzgraber, G.; Kozubek, S.; Lapidus, I.L.

    1985-01-01

    The dependence of the relative sedimentation velocity of DNA-membrane complexes on the dose of irradiation and time of incubation of Chinese Hamster cells is analysed. It is concluded that the initial part of the curve provides the information on the occurrence of single strand breaks in DNA; the position of the local maximum allows us to calculate the yield of DNA double strand breaks. The reparation decay constant can be estimated as well

  6. Biooxidation of 2-phenylethanol to phenylacetic acid by whole-cell Gluconobacter oxydans biocatalyst immobilized in polyelectrolyte complex capsules

    Czech Academy of Sciences Publication Activity Database

    Bertóková, A.; Vikartovská, A.; Bučko, M.; Gemeiner, P.; Tkáč, J.; Chorvát, D.; Štefuca, V.; Neděla, Vilém

    2015-01-01

    Roč. 33, č. 2 (2015), s. 111-120 ISSN 1024-2422 R&D Projects: GA ČR(CZ) GA14-22777S Institutional support: RVO:68081731 Keywords : Gluconobacter oxydans * natural flavors * phenylacetic acid * immobilized whole-cell biocatalyst * polyelectrolyte complex capsules * environmental scanning electron microscopy Subject RIV: JA - Electronics ; Optoelectronics, Electrical Engineering Impact factor: 0.892, year: 2015

  7. Self-assembling complexes of quantum dots and scFv antibodies for cancer cell targeting and imaging.

    Directory of Open Access Journals (Sweden)

    Tatiana A Zdobnova

    Full Text Available Semiconductor quantum dots represent a novel class of fluorophores with unique physical and chemical properties which could enable a remarkable broadening of the current applications of fluorescent imaging and optical diagnostics. Complexes of quantum dots and antibodies are promising visualising agents for fluorescent detection of selective biomarkers overexpressed in tumor tissues. Here we describe the construction of self-assembling fluorescent complexes of quantum dots and anti-HER1 or anti-HER2/neu scFv antibodies and their interactions with cultured tumor cells. A binding strategy based on a very specific non-covalent interaction between two proteins, barnase and barstar, was used to connect quantum dots and the targeting antibodies. Such a strategy allows combining the targeting and visualization functions simply by varying the corresponding modules of the fluorescent complex.

  8. The RSV F and G glycoproteins interact to form a complex on the surface of infected cells

    International Nuclear Information System (INIS)

    Low, Kit-Wei; Tan, Timothy; Ng, Ken; Tan, Boon-Huan; Sugrue, Richard J.

    2008-01-01

    In this study, the interaction between the respiratory syncytial virus (RSV) fusion (F) protein, attachment (G) protein, and small hydrophobic (SH) proteins was examined. Immunoprecipitation analysis suggested that the F and G proteins exist as a protein complex on the surface of RSV-infected cells, and this conclusion was supported by ultracentrifugation analysis that demonstrated co-migration of surface-expressed F and G proteins. Although our analysis provided evidence for an interaction between the G and SH proteins, no evidence was obtained for a single protein complex involving all three of the virus proteins. These data suggest the existence of multiple virus glycoprotein complexes within the RSV envelope. Although the stimulus that drives RSV-mediated membrane fusion is unknown, the association between the G and F proteins suggest an indirect role for the G protein in this process

  9. More sensitive correlation of afferent pupillary defect with ganglion cell complex

    Directory of Open Access Journals (Sweden)

    Eulogio Besada

    2018-04-01

    Full Text Available Purpose: This study investigated the correlation between the relative afferent pupillary defect (RAPD and retinal nerve fiber layer thickness (RNFLT in optic neuropathy. Methods: RAPD assessment was performed using a log unit neutral density filter bar. Spectral domain optical coherence tomography RTVue-100 (Optovue was used to examine the subjects. The optic nerve head pattern (ONH was subdivided and identified for the purpose of the study into circumpapillary RNFLT (cpRNFLT and peripheral circumpapillary RNFLT (pcpRNFLT. The cpRNFLT, pcpRNFLT and ganglion cell complex (GCC parameters were analyzed. Results: Eighteen females and twenty three males with asymmetric optic neuropathy and a RAPD participated. Thirty-three subjects had glaucoma and eight had optic neuropathy other than glaucoma. Significant correlations (p < 0.02 were obtained for the RAPD and the percentage difference loss of the GCC and RNFLT parameters. The grouped mean percentage difference loss for RNFLT was significantly different from that of the GCC (p < 0.001. At a 0.6 log unit RAPD, the average mean percentage difference loss was 23% for the CRNFLT, 15% for the GCC, 12% for the global loss volume percentage and 6% for the focal loss volume percentage (FLV%. Conclusions: Significant correlations between RNFLT loss for cpRNFLT, pcpRNFLT and GCC parameters with RAPD were observed. Approximately a 35% higher sensitivity was obtained using GCC compared to CRNFL parameters. The expected change in GCC average for every 0.3 log unit increment was approximately 8.49 μm. The FLV% corresponded more sensitively to a RAPD but appeared to be influenced by disease severity. Resumen: Objetivo: Este estudio investigó la correlación entre el defecto pupilar aferente relativo (DPAR y el grosor de la capa de fibras nerviosas de la retina (RNFLT en la neuropatía óptica. Métodos: La valoración del DPAR se realizó utilizando una barra de filtro de densidad neutra de unidades logar

  10. Deliberate ROS production and auxin synergistically trigger the asymmetrical division generating the subsidiary cells in Zea mays stomatal complexes.

    Science.gov (United States)

    Livanos, Pantelis; Galatis, Basil; Apostolakos, Panagiotis

    2016-07-01

    Subsidiary cell generation in Poaceae is an outstanding example of local intercellular stimulation. An inductive stimulus emanates from the guard cell mother cells (GMCs) towards their laterally adjacent subsidiary cell mother cells (SMCs) and triggers the asymmetrical division of the latter. Indole-3-acetic acid (IAA) immunolocalization in Zea mays protoderm confirmed that the GMCs function as local sources of auxin and revealed that auxin is polarly accumulated between GMCs and SMCs in a timely-dependent manner. Besides, staining techniques showed that reactive oxygen species (ROS) exhibit a closely similar, also time-dependent, pattern of appearance suggesting ROS implication in subsidiary cell formation. This phenomenon was further investigated by using the specific NADPH-oxidase inhibitor diphenylene iodonium, the ROS scavenger N-acetyl-cysteine, menadione which leads to ROS overproduction, and H2O2. Treatments with diphenylene iodonium, N-acetyl-cysteine, and menadione specifically blocked SMC polarization and asymmetrical division. In contrast, H2O2 promoted the establishment of SMC polarity and subsequently subsidiary cell formation in "younger" protodermal areas. Surprisingly, H2O2 favored the asymmetrical division of the intervening cells of the stomatal rows leading to the creation of extra apical subsidiary cells. Moreover, H2O2 altered IAA localization, whereas synthetic auxin analogue 1-napthaleneacetic acid enhanced ROS accumulation. Combined treatments with ROS modulators along with 1-napthaleneacetic acid or 2,3,5-triiodobenzoic acid, an auxin efflux inhibitor, confirmed the crosstalk between ROS and auxin functioning during subsidiary cell generation. Collectively, our results demonstrate that ROS are critical partners of auxin during development of Z. mays stomatal complexes. The interplay between auxin and ROS seems to be spatially and temporarily regulated.

  11. Mitochondrially targeted vitamin E succinate efficiently kills breast tumour-initiating cells in a complex II-dependent manner

    International Nuclear Information System (INIS)

    Yan, Bing; Stantic, Marina; Zobalova, Renata; Bezawork-Geleta, Ayenachew; Stapelberg, Michael; Stursa, Jan; Prokopova, Katerina; Dong, Lanfeng; Neuzil, Jiri

    2015-01-01

    Accumulating evidence suggests that breast cancer involves tumour-initiating cells (TICs), which play a role in initiation, metastasis, therapeutic resistance and relapse of the disease. Emerging drugs that target TICs are becoming a focus of contemporary research. Mitocans, a group of compounds that induce apoptosis of cancer cells by destabilising their mitochondria, are showing their potential in killing TICs. In this project, we investigated mitochondrially targeted vitamin E succinate (MitoVES), a recently developed mitocan, for its in vitro and in vivo efficacy against TICs. The mammosphere model of breast TICs was established by culturing murine NeuTL and human MCF7 cells as spheres. This model was verified by stem cell marker expression, tumour initiation capacity and chemotherapeutic resistance. Cell susceptibility to MitoVES was assessed and the cell death pathway investigated. In vivo efficacy was studied by grafting NeuTL TICs to form syngeneic tumours. Mammospheres derived from NeuTL and MCF7 breast cancer cells were enriched in the level of stemness, and the sphere cells featured altered mitochondrial function. Sphere cultures were resistant to several established anti-cancer agents while they were susceptible to MitoVES. Killing of mammospheres was suppressed when the mitochondrial complex II, the molecular target of MitoVES, was knocked down. Importantly, MitoVES inhibited progression of syngeneic HER2 high tumours derived from breast TICs by inducing apoptosis in tumour cells. These results demonstrate that using mammospheres, a plausible model for studying TICs, drugs that target mitochondria efficiently kill breast tumour-initiating cells. The online version of this article (doi:10.1186/s12885-015-1394-7) contains supplementary material, which is available to authorized users

  12. Substrate-mediated delivery of gene complex nanoparticles via polydopamine coating for enhancing competitiveness of endothelial cells.

    Science.gov (United States)

    Li, Bo-Chao; Chang, Hao; Ren, Ke-Feng; Ji, Jian

    2016-11-01

    Substrate-mediated delivery of functional plasmid DNA (pDNA) has been proven to be a promising strategy to promote competitiveness of endothelial cells (ECs) over smooth muscle cells (SMCs), which is beneficial to inducing fast endothelialization of implanted vascular devices. Thus, it is of great importance to develop universal approaches with simplicity and easiness to immobilize DNA complex nanoparticles on substrates. In this study, the bioinspired polydopamine (PDA) coating was employed in immobilization of DNA complex nanoparticles, which were composed of protamine (PrS) and plasmid DNA encoding with hepatocyte growth factor (HGF-pDNA) gene. We demonstrated that the DNA complex nanoparticles can be successfully immobilized onto the PDA surface. Consequently, the HGF expression of both ECs and SMCs were significantly improved when they cultured on the DNA complex nanoparticles-immobilized substrates. Furthermore, EC proliferation was specifically promoted due to bioactivity of HGF, leading to an enhancement of EC competitiveness over SMCs. Our findings demonstrated the substrate-mediated functional gene nanoparticle delivery through PDA coating as a simple and efficient approach. It may hold great potential in the field of interventional cardiovascular implants. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. The elongin complex antagonizes the chromatin factor Corto for vein versus intervein cell identity in Drosophila wings.

    Science.gov (United States)

    Rougeot, Julien; Renard, Myrtille; Randsholt, Neel B; Peronnet, Frédérique; Mouchel-Vielh, Emmanuèle

    2013-01-01

    Drosophila wings mainly consist of two cell types, vein and intervein cells. Acquisition of either fate depends on specific expression of genes that are controlled by several signaling pathways. The nuclear mechanisms that translate signaling into regulation of gene expression are not completely understood, but they involve chromatin factors from the Trithorax (TrxG) and Enhancers of Trithorax and Polycomb (ETP) families. One of these is the ETP Corto that participates in intervein fate through interaction with the Drosophila EGF Receptor--MAP kinase ERK pathway. Precise mechanisms and molecular targets of Corto in this process are not known. We show here that Corto interacts with the Elongin transcription elongation complex. This complex, that consists of three subunits (Elongin A, B, C), increases RNA polymerase II elongation rate in vitro by suppressing transient pausing. Analysis of phenotypes induced by EloA, B, or C deregulation as well as genetic interactions suggest that the Elongin complex might participate in vein vs intervein specification, and antagonizes corto as well as several TrxG genes in this process. Chromatin immunoprecipitation experiments indicate that Elongin C and Corto bind the vein-promoting gene rhomboid in wing imaginal discs. We propose that Corto and the Elongin complex participate together in vein vs intervein fate, possibly through tissue-specific transcriptional regulation of rhomboid.

  14. The elongin complex antagonizes the chromatin factor Corto for vein versus intervein cell identity in Drosophila wings.

    Directory of Open Access Journals (Sweden)

    Julien Rougeot

    Full Text Available Drosophila wings mainly consist of two cell types, vein and intervein cells. Acquisition of either fate depends on specific expression of genes that are controlled by several signaling pathways. The nuclear mechanisms that translate signaling into regulation of gene expression are not completely understood, but they involve chromatin factors from the Trithorax (TrxG and Enhancers of Trithorax and Polycomb (ETP families. One of these is the ETP Corto that participates in intervein fate through interaction with the Drosophila EGF Receptor--MAP kinase ERK pathway. Precise mechanisms and molecular targets of Corto in this process are not known. We show here that Corto interacts with the Elongin transcription elongation complex. This complex, that consists of three subunits (Elongin A, B, C, increases RNA polymerase II elongation rate in vitro by suppressing transient pausing. Analysis of phenotypes induced by EloA, B, or C deregulation as well as genetic interactions suggest that the Elongin complex might participate in vein vs intervein specification, and antagonizes corto as well as several TrxG genes in this process. Chromatin immunoprecipitation experiments indicate that Elongin C and Corto bind the vein-promoting gene rhomboid in wing imaginal discs. We propose that Corto and the Elongin complex participate together in vein vs intervein fate, possibly through tissue-specific transcriptional regulation of rhomboid.

  15. Precise Temporal Profiling of Signaling Complexes in Primary Cells Using SWATH Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Etienne Caron

    2017-03-01

    Full Text Available Spatiotemporal organization of protein interactions in cell signaling is a fundamental process that drives cellular functions. Given differential protein expression across tissues and developmental stages, the architecture and dynamics of signaling interaction proteomes is, likely, highly context dependent. However, current interaction information has been almost exclusively obtained from transformed cells. In this study, we applied an advanced and robust workflow combining mouse genetics and affinity purification (AP-SWATH mass spectrometry to profile the dynamics of 53 high-confidence protein interactions in primary T cells, using the scaffold protein GRB2 as a model. The workflow also provided a sufficient level of robustness to pinpoint differential interaction dynamics between two similar, but functionally distinct, primary T cell populations. Altogether, we demonstrated that precise and reproducible quantitative measurements of protein interaction dynamics can be achieved in primary cells isolated from mammalian tissues, allowing resolution of the tissue-specific context of cell-signaling events.

  16. A Strategy for Rapid Construction of Blood Vessel-Like Structures with Complex Cell Alignments.

    Science.gov (United States)

    Wang, Nuoxin; Peng, Yunhu; Zheng, Wenfu; Tang, Lixue; Cheng, Shiyu; Yang, Junchuan; Liu, Shaoqin; Zhang, Wei; Jiang, Xingyu

    2018-04-17

    A method is developed that can rapidly produce blood vessel-like structures by bonding cell-laden electrospinning (ES) films layer by layer using fibrin glue within 90 min. This strategy allows control of cell type, cell orientation, and material composition in separate layers. Furthermore, ES films with thicker fibers (polylactic-co-glycolic acid, fiber diameter: ≈3.7 µm) are used as cell-seeding layers to facilitate the cell in-growth; those with thinner fibers (polylactic acid, fiber diameter: ≈1.8 µm) are used as outer reinforcing layers to improve the mechanical strength and reduce the liquid leakage of the scaffold. Cells grow, proliferate, and migrate well in the multilayered structure. This design aims at a new type of blood vessel substitute with flexible control of parameters and implementation of functions. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Preparation of antisera specific for human B cells by immunization of rabbits with immune complexes

    International Nuclear Information System (INIS)

    Welsh, K.I.; Turner, M.J.

    1976-01-01

    Three rabbit antisera are described which are specific without absorption (titer 1:100) for separated human B cells, as measured by complement and non-complement fixing assays. The method of production of these sera involved injections of rabbits with precipitin lines formed between 10μ1 of three separate detergent solubilized membrane preparations and 4μ1 aliquots of rabbit antisera to human B cells. In addition to being B cell specific, the three sera block the MLC reaction, inhibit aggregated IgG binding to B cells, and show differential degrees of B cell lysis when tested on a panel of separated B and T cells. These and other properties suggest that the target specificities of the antibodies are the human equivalent of the murine Ia antigens. (author)

  18. Mitochondria are the primary target in the induction of apoptosis by chiral ruthenium(II) polypyridyl complexes in cancer cells.

    Science.gov (United States)

    Wang, Jin-Quan; Zhang, Ping-Yu; Qian, Chen; Hou, Xiao-Juan; Ji, Liang-Nian; Chao, Hui

    2014-03-01

    A series of novel chiral ruthenium(II) polypyridyl complexes (Δ-Ru1, Λ-Ru1, Δ-Ru2, Λ-Ru2, Δ-Ru3, Λ-Ru3) were synthesized and evaluated to determine their antiproliferative activities. Colocalization, inductively coupled plasma mass spectrometry, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay studies showed that these ruthenium(II) complexes accumulated preferentially in the mitochondria and exhibited cytotoxicity against various cancer cells in vitro. The complex Δ-Ru1 is of particular interest because it was found to have half-maximal inhibitory concentrations comparable to those of cisplatin and better activity than cisplatin against a cisplatin-resistant cell line, A549-CP/R. Δ-Ru1 induced alterations in the mitochondrial membrane potential and triggered intrinsic mitochondria-mediated apoptosis in HeLa cells, which involved the regulation of Bcl-2 family members and the activation of caspases. Taken together, these data suggest that Δ-Ru1 may be a novel mitochondria-targeting anticancer agent.

  19. Aquation Is a Crucial Activation Step for Anticancer Action of Ruthenium(II) Polypyridyl Complexes to Trigger Cancer Cell Apoptosis.

    Science.gov (United States)

    Li, Meng; Lai, Lanhai; Zhao, Zhennan; Chen, Tianfeng

    2016-01-01

    Aquation has been proposed as crucial chemical action step for ruthenium (Ru) complexes, but its effects on the action mechanisms remain elusive. Herein, we have demonstrated the aquation process of a potent Ru polypyridyl complex (RuBmp=[Ru(II) (bmbp)(phen)Cl]ClO4 , bmbp=2,6-bis(6-methylbenzimidazol-2-yl) pyridine, phen=phenanthroline) with a chloride ligand, and revealed that aquation of RuBmp effectively enhanced its hydrophilicity and cellular uptake, thus significantly increasing its anticancer efficacy. The aquation products (H-RuBmp=[Ru(II) (bmbp)(phen)Cl]ClO4 , [Ru(II) (bmbp)(phen)(H2 O)]ClO4 , bmbp) exhibited a much higher apoptosis-inducing ability than the intact complex, with involvement of caspase activation, mitochondria dysfunction, and interaction with cell membrane death receptors. H-RuBmp demonstrated a higher interaction potency with the cell membrane and induced higher levels of ROS overproduction in cancer cells to regulate the AKT, MAPK, and p53 signaling pathways. Taken together, this study could provide useful information for fine-tuning the rational design of next-generation metal medicines. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Intracellular distribution of TM4SF1 and internalization of TM4SF1-antibody complex in vascular endothelial cells

    International Nuclear Information System (INIS)

    Sciuto, Tracey E.; Merley, Anne; Lin, Chi-Iou; Richardson, Douglas; Liu, Yu; Li, Dan; Dvorak, Ann M.; Dvorak, Harold F.; Jaminet, Shou-Ching S.

    2015-01-01

    Transmembrane-4 L-six family member-1 (TM4SF1) is a small plasma membrane-associated glycoprotein that is highly and selectively expressed on the plasma membranes of tumor cells, cultured endothelial cells, and, in vivo, on tumor-associated endothelium. Immunofluorescence microscopy also demonstrated TM4SF1 in cytoplasm and, tentatively, within nuclei. With monoclonal antibody 8G4, and the finer resolution afforded by immuno-nanogold transmission electron microscopy, we now demonstrate TM4SF1 in uncoated cytoplasmic vesicles, nuclear pores and nucleoplasm. Because of its prominent surface location on tumor cells and tumor-associated endothelium, TM4SF1 has potential as a dual therapeutic target using an antibody drug conjugate (ADC) approach. For ADC to be successful, antibodies reacting with cell surface antigens must be internalized for delivery of associated toxins to intracellular targets. We now report that 8G4 is efficiently taken up into cultured endothelial cells by uncoated vesicles in a dynamin-dependent, clathrin-independent manner. It is then transported along microtubules through the cytoplasm and passes through nuclear pores into the nucleus. These findings validate TM4SF1 as an attractive candidate for cancer therapy with antibody-bound toxins that have the capacity to react with either cytoplasmic or nuclear targets in tumor cells or tumor-associated vascular endothelium. - Highlights: • Anti-TM4SF1 antibody 8G4 was efficiently taken up by cultured endothelial cells. • TM4SF1–8G4 internalization is dynamin-dependent but clathrin-independent. • TM4SF1–8G4 complexes internalize along microtubules to reach the perinuclear region. • Internalized TM4SF1–8G4 complexes pass through nuclear pores into the nucleus. • TM4SF1 is an attractive candidate for ADC cancer therapy

  1. Understanding how the complex molecular architecture of mannan-degrading hydrolases contributes to plant cell wall degradation.

    Science.gov (United States)

    Zhang, Xiaoyang; Rogowski, Artur; Zhao, Lei; Hahn, Michael G; Avci, Utku; Knox, J Paul; Gilbert, Harry J

    2014-01-24

    Microbial degradation of plant cell walls is a central component of the carbon cycle and is of increasing importance in environmentally significant industries. Plant cell wall-degrading enzymes have a complex molecular architecture consisting of catalytic modules and, frequently, multiple non-catalytic carbohydrate binding modules (CBMs). It is currently unclear whether the specificities of the CBMs or the topology of the catalytic modules are the primary drivers for the specificity of these enzymes against plant cell walls. Here, we have evaluated the relationship between CBM specificity and their capacity to enhance the activity of GH5 and GH26 mannanases and CE2 esterases against intact plant cell walls. The data show that cellulose and mannan binding CBMs have the greatest impact on the removal of mannan from tobacco and Physcomitrella cell walls, respectively. Although the action of the GH5 mannanase was independent of the context of mannan in tobacco cell walls, a significant proportion of the polysaccharide was inaccessible to the GH26 enzyme. The recalcitrant mannan, however, was fully accessible to the GH26 mannanase appended to a cellulose binding CBM. Although CE2 esterases display similar specificities against acetylated substrates in vitro, only CjCE2C was active against acetylated mannan in Physcomitrella. Appending a mannan binding CBM27 to CjCE2C potentiated its activity against Physcomitrella walls, whereas a xylan binding CBM reduced the capacity of esterases to deacetylate xylan in tobacco walls. This work provides insight into the biological significance for the complex array of hydrolytic enzymes expressed by plant cell wall-degrading microorganisms.

  2. Intracellular distribution of TM4SF1 and internalization of TM4SF1-antibody complex in vascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Sciuto, Tracey E.; Merley, Anne; Lin, Chi-Iou [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Richardson, Douglas [Department of Molecular and Cellular Biology, Harvard University (United States); Liu, Yu [Department of Pharmacology, Shanxi Medical University, Xinjiannanlu 56, Shanxi Province, Taiyuan 030001 (China); Li, Dan; Dvorak, Ann M. [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Dvorak, Harold F., E-mail: hdvorak@bidmc.harvard.edu [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States); Jaminet, Shou-Ching S., E-mail: sjaminet@bidmc.harvard.edu [Center for Vascular Biology Research and Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School (United States)

    2015-09-25

    Transmembrane-4 L-six family member-1 (TM4SF1) is a small plasma membrane-associated glycoprotein that is highly and selectively expressed on the plasma membranes of tumor cells, cultured endothelial cells, and, in vivo, on tumor-associated endothelium. Immunofluorescence microscopy also demonstrated TM4SF1 in cytoplasm and, tentatively, within nuclei. With monoclonal antibody 8G4, and the finer resolution afforded by immuno-nanogold transmission electron microscopy, we now demonstrate TM4SF1 in uncoated cytoplasmic vesicles, nuclear pores and nucleoplasm. Because of its prominent surface location on tumor cells and tumor-associated endothelium, TM4SF1 has potential as a dual therapeutic target using an antibody drug conjugate (ADC) approach. For ADC to be successful, antibodies reacting with cell surface antigens must be internalized for delivery of associated toxins to intracellular targets. We now report that 8G4 is efficiently taken up into cultured endothelial cells by uncoated vesicles in a dynamin-dependent, clathrin-independent manner. It is then transported along microtubules through the cytoplasm and passes through nuclear pores into the nucleus. These findings validate TM4SF1 as an attractive candidate for cancer therapy with antibody-bound toxins that have the capacity to react with either cytoplasmic or nuclear targets in tumor cells or tumor-associated vascular endothelium. - Highlights: • Anti-TM4SF1 antibody 8G4 was efficiently taken up by cultured endothelial cells. • TM4SF1–8G4 internalization is dynamin-dependent but clathrin-independent. • TM4SF1–8G4 complexes internalize along microtubules to reach the perinuclear region. • Internalized TM4SF1–8G4 complexes pass through nuclear pores into the nucleus. • TM4SF1 is an attractive candidate for ADC cancer therapy.

  3. Cell adhesion in Drosophila: versatility of cadherin and integrin complexes during development

    OpenAIRE

    Bulgakova, Natalia A.; Klapholz, Benjamin; Brown, Nicholas H.

    2012-01-01

    We highlight recent progress in understanding cadherin and integrin function in the model organism Drosophila. New functions for these adhesion receptors continue to be discovered in this system, emphasising the importance of cell adhesion within the developing organism and showing that the requirement for cell adhesion changes between cell types. New ways to control adhesion have been discovered, including controlling the expression and recruitment of adhesion components, their posttranslati...

  4. Evaluation of polyelectrolyte complex-based scaffolds for mesenchymal stem cell therapy in cardiac ischemia treatment

    OpenAIRE

    Ceccaldi, Caroline; Bushkalova, Raya; Alfarano, Chiara; Lairez, Olivier; Calise, Denis; Bourin, Philippe; Frugier, Céline; Rouzaud-Laborde, Charlotte; Cussac, Daniel; Parini, Angelo; Sallerin, Brigitte; Girod Fullana, Sophie

    2014-01-01

    Three-dimensional (3D) scaffolds hold great potential for stem cell-based therapies. Indeed, recent results have shown that biomimetic scaffolds may enhance cell survival and promote an increase in the concentration of therapeutic cells at the injury site. The aim of this work was to engineer an original polymeric scaffold based on the respective beneficial effects of alginate and chitosan. Formulations were made from various alginate/chitosan ratios to form opposite-charge polyelectrolyte co...

  5. p53-dependent control of cell death by nicastrin: lack of requirement for presenilin-dependent gamma-secretase complex.

    Science.gov (United States)

    Pardossi-Piquard, Raphaëlle; Dunys, Julie; Giaime, Emilie; Guillot-Sestier, Marie-Victoire; St George-Hyslop, Peter; Checler, Frédéric; Alves da Costa, Cristine

    2009-04-01

    Nicastrin (NCT) is a component of the presenilin (PS)-dependent gamma-secretase complexes that liberate amyloid beta-peptides from the beta-Amyloid Precursor Protein. Several lines of evidence indicate that the members of these complexes could also contribute to the control of cell death. Here we show that over-expression of NCT increases the viability of human embryonic kidney (HEK293) cells and decreases staurosporine (STS)- and thapsigargin (TPS)-induced caspase-3 activation in various cell lines from human and neuronal origins by Akt-dependent pathway. NCT lowers p53 expression, transcriptional activity and promoter transactivation and reduces p53 phosphorylation. NCT-associated protection against STS-stimulated cell death was completely abolished by p53 deficiency. Conversely, the depletion of NCT drastically enhances STS-induced caspase-3 activation and p53 pathway and favored p53 nuclear translocation. We examined whether NCT protective function depends on PS-dependent gamma-secretase activity. First, a 29-amino acid deletion known to reduce NCT-dependent amyloid beta-peptide production did not affect NCT-associated protective phenotype. Second, NCT still reduces STS-induced caspase-3 activation in fibroblasts lacking PS1 and PS2. Third, the gamma-secretase inhibitor DFK167 did not affect NCT-mediated reduction of p53 activity. Altogether, our study indicates that NCT controls cell death via phosphoinositide 3-kinase/Akt and p53-dependent pathways and that this function remains independent of the activity and molecular integrity of the gamma-secretase complexes.

  6. Molecular Basis of Natural Killer Cell Tumor Target Recognition: The NKr/MHC Class I Complex

    National Research Council Canada - National Science Library

    Hasemann, Charles

    1999-01-01

    .... We have pursued this via the heterologous expression of wild type and mutant NK receptors for the purpose of the determination of the atomic structure of an NK receptor/ class I MHC complex via X-ray crystallography...

  7. Proliferating cell nuclear antigen (PCNA)-associated KIAA0101/PAF15 protein is a cell cycle-regulated anaphase-promoting complex/cyclosome substrate.

    Science.gov (United States)

    Emanuele, Michael J; Ciccia, Alberto; Elia, Andrew E H; Elledge, Stephen J

    2011-06-14

    The anaphase-promoting complex/cyclosome (APC/C) is a cell cycle-regulated E3 ubiquitin ligase that controls the degradation of substrate proteins at mitotic exit and throughout the G1 phase. We have identified an APC/C substrate and cell cycle-regulated protein, KIAA0101/PAF15. PAF15 protein levels peak in the G2/M phase of the cell cycle and drop rapidly at mitotic exit in an APC/C- and KEN-box-dependent fashion. PAF15 associates with proliferating cell nuclear antigen (PCNA), and depletion of PAF15 decreases the number of cells in S phase, suggesting a role for it in cell cycle regulation. Following irradiation, PAF15 colocalized with γH2AX foci at sites of DNA damage through its interaction with PCNA. Finally, PAF15 depletion led to an increase in homologous recombination-mediated DNA repair, and overexpression caused sensitivity to UV-induced DNA damage. We conclude that PAF15 is an APC/C-regulated protein involved in both cell cycle progression and the DNA damage response.

  8. Simultaneous inhibition of mTOR-containing complex 1 (mTORC1) and MNK induces apoptosis of cutaneous T-cell lymphoma (CTCL) cells

    DEFF Research Database (Denmark)

    Marzec, Michal Tomasz; Liu, Xiaobin; Wysocka, Maria

    2011-01-01

    mTOR kinase forms the mTORC1 complex by associating with raptor and other proteins and affects a number of key cell functions. mTORC1 activates p70S6kinase 1 (p70S6K1) and inhibits 4E-binding protein 1 (4E-BP1). In turn, p70S6K1 phosphorylates a S6 protein of the 40S ribosomal subunit (S6rp) and 4E...

  9. APC/β-catenin-rich complexes at membrane protrusions regulate mammary tumor cell migration and mesenchymal morphology

    International Nuclear Information System (INIS)

    Odenwald, Matthew A; Prosperi, Jenifer R; Goss, Kathleen H

    2013-01-01

    The APC tumor suppressor is mutated or downregulated in many tumor types, and is prominently localized to punctate clusters at protrusion tips in migratory cells, such as in astrocytes where it has been implicated in directed cell motility. Although APC loss is considered an initiating event in colorectal cancer, for example, it is less clear what role APC plays in tumor cell motility and whether loss of APC might be an important promoter of tumor progression in addition to initiation. The localization of APC and β-catenin was analyzed in multiple cell lines, including non-transformed epithelial lines treated with a proteasome inhibitor or TGFβ to induce an epithelial-to-mesenchymal transition (EMT), as well as several breast cancer lines, by immunofluorescence. APC expression was knocked down in 4T07 mammary tumor cells using lentiviral-mediated delivery of APC-specific short-hairpin (sh) RNAs, and assessed using quantitative (q) reverse-transcriptase (RT)-PCR and western blotting. Tumor cell motility was analyzed by performing wound-filling assays, and morphology via immunofluorescence (IF) and phase-contrast microscopy. Additionally, proliferation was measured using BrdU incorporation, and TCF reporter assays were performed to determine β-catenin/TCF-mediated transcriptional activity. APC/β-catenin-rich complexes were observed at protrusion ends of migratory epithelial cells treated with a proteasome inhibitor or when EMT has been induced and in tumor cells with a mesenchymal, spindle-like morphology. 4T07 tumor cells with reduced APC levels were significantly less motile and had a more rounded morphology; yet, they did not differ significantly in proliferation or β-catenin/TCF transcriptional activity. Furthermore, we found that APC/β-catenin-rich complexes at protrusion ends were dependent upon an intact microtubule cytoskeleton. These findings indicate that membrane protrusions with APC/β-catenin-containing puncta control the migratory potential and

  10. APC/β-catenin-rich complexes at membrane protrusions regulate mammary tumor cell migration and mesenchymal morphology

    Science.gov (United States)

    2013-01-01

    Background The APC tumor suppressor is mutated or downregulated in many tumor types, and is prominently localized to punctate clusters at protrusion tips in migratory cells, such as in astrocytes where it has been implicated in directed cell motility. Although APC loss is considered an initiating event in colorectal cancer, for example, it is less clear what role APC plays in tumor cell motility and whether loss of APC might be an important promoter of tumor progression in addition to initiation. Methods The localization of APC and β-catenin was analyzed in multiple cell lines, including non-transformed epithelial lines treated with a proteasome inhibitor or TGFβ to induce an epithelial-to-mesenchymal transition (EMT), as well as several breast cancer lines, by immunofluorescence. APC expression was knocked down in 4T07 mammary tumor cells using lentiviral-mediated delivery of APC-specific short-hairpin (sh) RNAs, and assessed using quantitative (q) reverse-transcriptase (RT)-PCR and western blotting. Tumor cell motility was analyzed by performing wound-filling assays, and morphology via immunofluorescence (IF) and phase-contrast microscopy. Additionally, proliferation was measured using BrdU incorporation, and TCF reporter assays were performed to determine β-catenin/TCF-mediated transcriptional activity. Results APC/β-catenin-rich complexes were observed at protrusion ends of migratory epithelial cells treated with a proteasome inhibitor or when EMT has been induced and in tumor cells with a mesenchymal, spindle-like morphology. 4T07 tumor cells with reduced APC levels were significantly less motile and had a more rounded morphology; yet, they did not differ significantly in proliferation or β-catenin/TCF transcriptional activity. Furthermore, we found that APC/β-catenin-rich complexes at protrusion ends were dependent upon an intact microtubule cytoskeleton. Conclusions These findings indicate that membrane protrusions with APC/β-catenin-containing puncta

  11. Structure of the complex between teicoplanin and a bacterial cell-wall peptide: use of a carrier-protein approach

    International Nuclear Information System (INIS)

    Economou, Nicoleta J.; Zentner, Isaac J.; Lazo, Edwin; Jakoncic, Jean; Stojanoff, Vivian; Weeks, Stephen D.; Grasty, Kimberly C.; Cocklin, Simon; Loll, Patrick J.

    2013-01-01

    Using a carrier-protein strategy, the structure of teicoplanin bound to its bacterial cell-wall target has been determined. The structure reveals the molecular determinants of target recognition, flexibility in the antibiotic backbone and intrinsic radiation sensitivity of teicoplanin. Multidrug-resistant bacterial infections are commonly treated with glycopeptide antibiotics such as teicoplanin. This drug inhibits bacterial cell-wall biosynthesis by binding and sequestering a cell-wall precursor: a d-alanine-containing peptide. A carrier-protein strategy was used to crystallize the complex of teicoplanin and its target peptide by fusing the cell-wall peptide to either MBP or ubiquitin via native chemical ligation and subsequently crystallizing the protein–peptide–antibiotic complex. The 2.05 Å resolution MBP–peptide–teicoplanin structure shows that teicoplanin recognizes its ligand through a combination of five hydrogen bonds and multiple van der Waals interactions. Comparison of this teicoplanin structure with that of unliganded teicoplanin reveals a flexibility in the antibiotic peptide backbone that has significant implications for ligand recognition. Diffraction experiments revealed an X-ray-induced dechlorination of the sixth amino acid of the antibiotic; it is shown that teicoplanin is significantly more radiation-sensitive than other similar antibiotics and that ligand binding increases radiosensitivity. Insights derived from this new teicoplanin structure may contribute to the development of next-generation antibacterials designed to overcome bacterial resistance

  12. Evidence that the respiratory syncytial virus polymerase complex associates with lipid rafts in virus-infected cells: a proteomic analysis

    International Nuclear Information System (INIS)

    McDonald, Terence P.; Pitt, Andrew R.; Brown, Gaie; Rixon, Helen W. McL.; Sugrue, Richard J.

    2004-01-01

    The interaction between the respiratory syncytial virus (RSV) polymerase complex and lipid rafts was examined in HEp2 cells. Lipid-raft membranes were prepared from virus-infected cells and their protein content was analysed by Western blotting and mass spectrometry. This analysis revealed the presence of the N, P, L, M2-1 and M proteins. However, these proteins appeared to differ from one another in their association with these structures, with the M2-1 protein showing a greater partitioning into raft membranes compared to that of the N, P or M proteins. Determination of the polymerase activity profile of the gradient fractions revealed that 95% of the detectable viral enzyme activity was associated with lipid-raft membranes. Furthermore, analysis of virus-infected cells by confocal microscopy suggested an association between these proteins and the raft-lipid, GM1. Together, these results provide evidence that the RSV polymerase complex is able to associate with lipid rafts in virus-infected cells

  13. Fabrication and characterization of organic solar cells using metal complex of phthalocyanines

    Energy Technology Data Exchange (ETDEWEB)

    Kida, Tomoyasu, E-mail: suzuki@mat.usp.ac.jp; Suzuki, Atsushi, E-mail: suzuki@mat.usp.ac.jp; Akiyama, Tsuyoshi, E-mail: suzuki@mat.usp.ac.jp; Oku, Takeo, E-mail: suzuki@mat.usp.ac.jp [Department of Materials Science, The University of Shiga Prefecture 2500 Hassaka, Hikone, Shiga 522-8533 (Japan)

    2015-02-27

    Fabrication and characterization of organic solar cells using shuttle-cock-type phthalocyanines were carried out. Photovoltaic properties of the solar cells with inverted structures were investigated by current density-voltage characteristics. Effects of phase transition between H and J aggregates on the photovoltaic and optical properties were investigated. The photovoltaic mechanisms, energy levels and band gap of active layers were discussed.

  14. Complex molecular mechanisms cooperate to mediate histone deacetylase inhibitors anti-tumour activity in neuroblastoma cells

    Directory of Open Access Journals (Sweden)

    Nardou Katya

    2008-06-01

    Full Text Available Abstract Background Histone deacetylase inhibitors (HDACi are a new class of promising anti-tumour agent inhibiting cell proliferation and survival in tumour cells with very low toxicity toward normal cells. Neuroblastoma (NB is the second most common solid tumour in children still associated with poor outcome in higher stages and, thus NB strongly requires novel treatment modalities. Results We show here that the HDACi Sodium Butyrate (NaB, suberoylanilide hydroxamic acid (SAHA and Trichostatin A (TSA strongly reduce NB cells viability. The anti-tumour activity of these HDACi involved the induction of cell cycle arrest in the G2/M phase, followed by the activation of the intrinsic apoptotic pathway, via the activation of the caspases cascade. Moreover, HDACi mediated the activation of the pro-apoptotic proteins Bid and BimEL and the inactivation of the anti-apoptotic proteins XIAP, Bcl-xL, RIP and survivin, that further enhanced the apoptotic signal. Interestingly, the activity of these apoptosis regulators was modulated by several different mechanisms, either by caspases dependent proteolytic cleavage or by degradation via the proteasome pathway. In addition, HDACi strongly impaired the hypoxia-induced secretion of VEGF by NB cells. Conclusion HDACi are therefore interesting new anti-tumour agents for targeting highly malignant tumours such as NB, as these agents display a strong toxicity toward aggressive NB cells and they may possibly reduce angiogenesis by decreasing VEGF production by NB cells.

  15. In Vitro Activation of the IκB Kinase Complex by Human T-cell Leukemia Virus Type-1 Tax*

    Science.gov (United States)

    Mukherjee, Sohini; Negi, Veera S.; Keitany, Gladys; Tanaka, Yuetsu; Orth, Kim

    2008-01-01

    Human T-cell leukemia virus type-I expresses Tax, a 40-kDa oncoprotein that activates IκB kinase (IKK), resulting in constitutive activation of NFκB. Herein, we have developed an in vitro signaling assay to analyze IKK complex activation by recombinant Tax. Using this assay in combination with reporter assays, we demonstrate that Tax-mediated activation of IKK is independent of phosphatases. We show that sustained activation of the Tax-mediated activation of the NFκB pathway is dependent on an intact Hsp90-IKK complex. By acetylating and thereby preventing activation of the IKK complex by the Yersinia effector YopJ, we demonstrate that Tax-mediated activation of the IKK complex requires a phosphorylation step. Our characterization of an in vitro signaling assay system for the mechanism of Tax-mediated activation of the IKK complex with a variety of mutants and inhibitors results in a working model for the biochemical mechanism of Tax-induced activation. PMID:18223255

  16. Accessing complexity: the dynamics of virus-specific T cell responses

    DEFF Research Database (Denmark)

    Doherty, P C; Christensen, Jan Pravsgaard

    2000-01-01

    -specific CD8(+ )T cells. Analysis to date with both naturally acquired and experimentally induced infections has established that the numbers of virus-specific CD8(+) T cells present during both the acute and memory phases of the host response are more than tenfold in excess of previously suspected values....... The levels are such that the virus-specific CD8(+) set is readily detected in the human peripheral blood lymphocyte compartment, particularly during persistent infections. Experimentally, it is now possible to measure the extent of cycling for tetramer (+)CD8(+) T cells during the acute and memory phases...... of the host response to viruses. Dissection of the phenotypic, functional, and molecular diversity of CD8(+) T cell populations has been greatly facilitated. It is hoped it will also soon be possible to analyze CD4(+) T cell populations in this way. Though these are early days and there is an enormous amount...

  17. Phosphorylation of Ribosomal Protein S6 Mediates Mammalian Target of Rapamycin Complex 1-Induced Parathyroid Cell Proliferation in Secondary Hyperparathyroidism.

    Science.gov (United States)

    Volovelsky, Oded; Cohen, Gili; Kenig, Ariel; Wasserman, Gilad; Dreazen, Avigail; Meyuhas, Oded; Silver, Justin; Naveh-Many, Tally

    2016-04-01

    Secondary hyperparathyroidism is characterized by increased serum parathyroid hormone (PTH) level and parathyroid cell proliferation. However, the molecular pathways mediating the increased parathyroid cell proliferation remain undefined. Here, we found that the mTOR pathway was activated in the parathyroid of rats with secondary hyperparathyroidism induced by either chronic hypocalcemia or uremia, which was measured by increased phosphorylation of ribosomal protein S6 (rpS6), a downstream target of the mTOR pathway. This activation correlated with increased parathyroid cell proliferation. Inhibition of mTOR complex 1 by rapamycin decreased or prevented parathyroid cell proliferation in secondary hyperparathyroidism rats and in vitro in uremic rat parathyroid glands in organ culture. Knockin rpS6(p-/-) mice, in which rpS6 cannot be phosphorylated because of substitution of all five phosphorylatable serines with alanines, had impaired PTH secretion after experimental uremia- or folic acid-induced AKI. Uremic rpS6(p-/-) mice had no increase in parathyroid cell proliferation compared with a marked increase in uremic wild-type mice. These results underscore the importance of mTOR activation and rpS6 phosphorylation for the pathogenesis of secondary hyperparathyroidism and indicate that mTORC1 is a significant regulator of parathyroid cell proliferation through rpS6. Copyright © 2016 by the American Society of Nephrology.

  18. Low energy costs of F1Fo ATP synthase reversal in colon carcinoma cells deficient in mitochondrial complex IV.

    Science.gov (United States)

    Zhdanov, Alexander V; Andreev, Dmitry E; Baranov, Pavel V; Papkovsky, Dmitri B

    2017-05-01

    Mitochondrial polarisation is paramount for a variety of cellular functions. Under ischemia, mitochondrial membrane potential (ΔΨm) and proton gradient (ΔpH) are maintained via a reversal of mitochondrial F1Fo ATP synthase (mATPase), which can rapidly deplete ATP and drive cells into energy crisis. We found that under normal conditions in cells with disassembled cytochrome c oxidase complex (COX-deficient HCT116), mATPase maintains ΔΨm at levels only 15-20% lower than in WT cells, and for this utilises relatively little ATP. For a small energy expenditure, mATPase enables mitochondrial ΔpH, protein import, Ca 2+ turnover, and supports free radical detoxication machinery enlarged to protect the cells from oxidative damage. Whereas in COX-deficient cells the main source of ATP is glycolysis, the ΔΨm is still maintained upon inhibition of the adenine nucleotide translocators with bongkrekic acid and carboxyatractyloside, indicating that the role of ANTs is redundant, and matrix substrate level phosphorylation alone or in cooperation with ATP-Mg/P i carriers can continuously support the mATPase activity. Intriguingly, we found that mitochondrial complex III is active, and it contributes not only to free radical production, but also to ΔΨm maintenance and energy budget of COX-deficient cells. Overall, this study demonstrates that F1Fo ATP synthase can support general mitochondrial and cellular functions, working in extremely efficient 'energy saving' reverse mode and flexibly recruiting free radical detoxication and ATP producing / transporting pathways. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. TLR2-Modulating Lipoproteins of the Mycobacterium tuberculosis Complex Enhance the HIV Infectivity of CD4+ T Cells.

    Directory of Open Access Journals (Sweden)

    Ciaran Skerry

    Full Text Available Co-infection with Mycobacterium tuberculosis accelerates progression from HIV to AIDS. Our previous studies showed that M. tuberculosis complex, unlike M. smegmatis, enhances TLR2-dependent susceptibility of CD4+ T cells to HIV. The M. tuberculosis complex produces multiple TLR2-stimulating lipoproteins, which are absent in M. smegmatis. M. tuberculosis production of mature lipoproteins and TLR2 stimulation is dependent on cleavage by lipoprotein signal peptidase A (LspA. In order to determine the role of potential TLR2-stimulating lipoproteins on mycobacterial-mediated HIV infectivity of CD4+ T cells, we generated M. smegmatis recombinant strains overexpressing genes encoding various M. bovis BCG lipoproteins, as well as a Mycobacterium bovis BCG strain deficient in LspA (ΔlspA. Exposure of human peripheral blood mononuclear cells (PBMC to M. smegmatis strains overexpressing the BCG lipoproteins, LprF (p<0.01, LprH (p<0.05, LprI (p<0.05, LprP (p<0.001, LprQ (p<0.005, MPT83 (p<0.005, or PhoS1 (p<0.05, resulted in increased HIV infectivity of CD4+ T cells isolated from these PBMC. Conversely, infection of PBMC with ΔlspA reduced HIV infectivity of CD4+ T cells by 40% relative to BCG-infected cells (p<0.05. These results may have important implications for TB vaccination programs in areas with high mother-to-child HIV transmission.

  20. Targeting Common but Complex Proteoglycans on Breast Cancer Cells and Stem Cells Using Evolutionary Refined Malaria Proteins

    Science.gov (United States)

    2016-08-01

    T24 Bladder transitional cell carcinoma ++ UM-UC-6 Bladder transitional cell carcinoma +++ Cancer of mesenchymal lineage CW9019 Rhabdomyosarcoma...Expression (median) nd Bladder cancer (n=32) -3 -2 321-1 0 BCAN CA9 CCR10 CD44 THBD GPC3 GPC5 CSPG5 PODXL2 PTPRG S100A9 SDC1 SDC4 TGFBR3 TMEFF1 TMEFF2...in various other tumours including breast cancers . CCR10 Chemokine receptor 10 Not known Chemokine receptor . Overexpressed in lymphoma

  1. Organometallic tris(8-hydroxyquinoline)aluminum complexes as buffer layers and dopants in inverted organic solar cells

    International Nuclear Information System (INIS)

    Tolkki, Antti; Kaunisto, Kimmo; Heiskanen, Juha P.; Omar, Walaa A.E.; Huttunen, Kirsi; Lehtimäki, Suvi; Hormi, Osmo E.O.; Lemmetyinen, Helge

    2012-01-01

    Tris(8-hydroxyquinoline)aluminum (Alq 3 ) is a frequently used material for organic light emitting diodes. The electronic properties and solubility can be tuned by chemical tailoring of the quinoline part, which makes it an interesting candidate for organic solar cells. Steady-state absorption and fluorescence, as well as time-resolved fluorescence properties of the parent Alq 3 and a series of complexes consisting of derivatives, such as 4-substituted pyrazol, methylpyrazol, arylvinyl, and pyridinoanthrene moieties, of the quinoline ligand, were studied in solutions and in thin films. Suitability of the complexes as anodic buffer layers or dopants in inverted organic solar cells based on the well known bulk heterojunction of poly(3-hexylthiophene) (P3HT) and phenyl-C 61 -butyric acid methyl ester (PCBM) was tested. The devices equipped with the derivatives showed higher power conversion efficiency (η) compared to the photocells containing the parent Alq 3 . Open circuit voltage (V oc ) was increased when the derivatives were utilized as the anodic buffer layer. Doping of the P3HT:PCBM with a small amount of Alq 3 or its derivative improved short circuit current density, V oc , fill factor, and η, while the series resistance decreased. In addition, the devices were stable in air over several weeks without encapsulation. Possible mechanisms leading to the improvements in the photovoltaic performance by using the parent Alq 3 or its derivative as buffer layer or dopant are discussed. - Highlights: ► Tris(8-hydroxyquinoline)aluminum (Alq 3 ) complexes in inverted organic solar cells. ► The Alq 3 complexes were used as an anodic buffer layer and as a dopant. ► Efficiency increased and the derivatives revealed varying open circuit voltage. ► Photovoltaic performance was stable after storage in a dark ambient atmosphere.

  2. Organometallic tris(8-hydroxyquinoline)aluminum complexes as buffer layers and dopants in inverted organic solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Tolkki, Antti, E-mail: antti.tolkki@tut.fi [Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, FI-33101, Tampere (Finland); Kaunisto, Kimmo [Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, FI-33101, Tampere (Finland); Heiskanen, Juha P. [Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, FI-33101, Tampere (Finland); Department of Chemistry, University of Oulu, P.O. Box 3000, FI-90014, Oulu (Finland); Omar, Walaa A.E. [Department of Chemistry, University of Oulu, P.O. Box 3000, FI-90014, Oulu (Finland); Chemistry Branch, Department of Science and Mathematics, Suez Canal University, Suez 43721 (Egypt); Huttunen, Kirsi; Lehtimaeki, Suvi [Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, FI-33101, Tampere (Finland); Hormi, Osmo E.O. [Department of Chemistry, University of Oulu, P.O. Box 3000, FI-90014, Oulu (Finland); Lemmetyinen, Helge [Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, FI-33101, Tampere (Finland)

    2012-04-30

    Tris(8-hydroxyquinoline)aluminum (Alq{sub 3}) is a frequently used material for organic light emitting diodes. The electronic properties and solubility can be tuned by chemical tailoring of the quinoline part, which makes it an interesting candidate for organic solar cells. Steady-state absorption and fluorescence, as well as time-resolved fluorescence properties of the parent Alq{sub 3} and a series of complexes consisting of derivatives, such as 4-substituted pyrazol, methylpyrazol, arylvinyl, and pyridinoanthrene moieties, of the quinoline ligand, were studied in solutions and in thin films. Suitability of the complexes as anodic buffer layers or dopants in inverted organic solar cells based on the well known bulk heterojunction of poly(3-hexylthiophene) (P3HT) and phenyl-C{sub 61}-butyric acid methyl ester (PCBM) was tested. The devices equipped with the derivatives showed higher power conversion efficiency ({eta}) compared to the photocells containing the parent Alq{sub 3}. Open circuit voltage (V{sub oc}) was increased when the derivatives were utilized as the anodic buffer layer. Doping of the P3HT:PCBM with a small amount of Alq{sub 3} or its derivative improved short circuit current density, V{sub oc}, fill factor, and {eta}, while the series resistance decreased. In addition, the devices were stable in air over several weeks without encapsulation. Possible mechanisms leading to the improvements in the photovoltaic performance by using the parent Alq{sub 3} or its derivative as buffer layer or dopant are discussed. - Highlights: Black-Right-Pointing-Pointer Tris(8-hydroxyquinoline)aluminum (Alq{sub 3}) complexes in inverted organic solar cells. Black-Right-Pointing-Pointer The Alq{sub 3} complexes were used as an anodic buffer layer and as a dopant. Black-Right-Pointing-Pointer Efficiency increased and the derivatives revealed varying open circuit voltage. Black-Right-Pointing-Pointer Photovoltaic performance was stable after storage in a dark ambient

  3. Expression of rat class I major histocompatibility complex (MHC) alloantigens and hepatocytes and hepatoma cells

    International Nuclear Information System (INIS)

    Hunt, J.M.; Desai, P.A.; Chakraborty, S.

    1986-01-01

    Altered expression of Class I MHC alloantigens has been reported for murine tumors, and may be associated with the tumorigenic phenotype of tumor cells. To characterize MHC Class I alloantigen expression on a chemically-induced transplantable rat hepatoma cell line, 17X, derived from a (WF x F344) F 1 rat, polyvalent anti-F344 and anti-WF rat alloantisera were first used to immunoprecipitate the rat RT1.A Class I MHC alloantigens expressed on primary (WF x F344) F 1 hepatocyptes in short-term monolayer cultures. Two-dimensional isoelectric focusing and SDS-PAGE of immunoprecipitates from 35 S-methionine-labeled (WF x F344) F 1 hepatocytes clearly resolved the RT1.A/sup u/ (WF) and RT1.A/sup LvI/ (F344) parental alloantigens. Identical radiolabeling and immunoprecipitation failed to detect either parental alloantigen on the 17X hepatoma cells. However, indirect immunofluorescence and immunoblot analyses demonstrated the presence of parental alloantigens on the 17X cells. Immunization of F344 rats but not of WF rats with 17X cells resulted in antibodies cytotoxic for normal (WF X F344) F 1 spleen cells in the presence of complement. These findings indicate that a combination of detection techniques will be necessary to characterize altered alloantigen expression on rat hepatoma cells

  4. Self-production of tissue factor-coagulation factor VII complex by ovarian cancer cells.

    Science.gov (United States)

    Yokota, N; Koizume, S; Miyagi, E; Hirahara, F; Nakamura, Y; Kikuchi, K; Ruf, W; Sakuma, Y; Tsuchiya, E; Miyagi, Y

    2009-12-15

    Thromboembolic events are a major complication in ovarian cancer patients. Tissue factor (TF) is frequently overexpressed in ovarian cancer tissue and correlates with intravascular thrombosis. TF binds to coagulation factor VII (fVII), changing it to its active form, fVIIa. This leads to activation of the extrinsic coagulation cascade. fVII is produced by the liver and believed to be supplied from blood plasma at the site of coagulation. However, we recently showed that ovarian cancer cells express fVII transcripts under normoxia and that this transcription is inducible under hypoxia. These findings led us to hypothesise that ovarian cancer cells are intrinsically associated with TF-fVIIa coagulation activity, which could result in thrombosis. In this study, we examined whether ectopically expressed fVII could cause thrombosis by means of immunohistochemistry, RT-PCR, western blotting and flow cytometry. Ectopic fVII expression occurs frequently in ovarian cancers, particularly in clear cell carcinoma. We further showed that ovarian cancer cells express TF-fVIIa on the cell surface under normoxia and that this procoagulant activity is enhanced by hypoxic stimuli. Moreover, we showed that ovarian cancer cells secrete microparticles (MPs) with TF-fVIIa activity. Production of this procoagulant secretion is enhanced under hypoxia. These results raise the possibility that cancer cell-derived TF-fVIIa could cause thrombotic events in ovarian cancer patients.

  5. Complex interactions of caffeine and its structural analogs with ultraviolet light in cell killing

    International Nuclear Information System (INIS)

    Chan, G.L.; Little, J.B.

    1981-01-01

    We measured the clonogenic survival response of cultured mouse 10 Tsup(1/2) cells exposed to UV light and caffeine post-treatment. When 0.5 and 1 mM caffeine were present for 24 h immediately following UV, the D 0 values of the biphasic survival curves suggest that one subpopulation was sensitized and one subpopulation was protected from killing by UV light. A cloned survivor from the radioprotected subpopulation responded to UV plus caffeine in identical manner as the parent cells. When the caffeine exposure was prolonged to 48 h, only the radiosensitizing effect was observed. Two demethylated analogs of caffeine were also tested. The response of 10 Tsup(1/2) cells to 1 mM theophylline present for 24 h after UV irradiation was approximately the same as that for the same treatment with 1 mM caffeine. However, prolonging the theophylline exposure to 48 h failed to produce the same kind of potentiation of cell killing as that observed for caffeine. Xanthine by itself was a toxic to 10 Tsup(1/2) cells as caffeine, but had no synergistic effect as caffeine when given to UV-irradiated cells for 24 or 48 h. It is therefore unlikely that all the effects of caffeine on UV-irradiated cells are mediated by its demethylated metabolites. (orig.)

  6. Plant-based oral tolerance to hemophilia therapy employs a complex immune regulatory response including LAP+CD4+ T cells

    OpenAIRE

    Wang, Xiaomei; Su, Jin; Sherman, Alexandra; Rogers, Geoffrey L.; Liao, Gongxian; Hoffman, Brad E.; Leong, Kam W.; Terhorst, Cox; Daniell, Henry; Herzog, Roland W.

    2015-01-01

    Coadministering FIX orally and systemically induces tolerance via complex immune regulation, involving tolerogenic dendritic and T-cell subsets.Induced CD4+CD25−LAP+ regulatory T cells with increased IL-10 and TGF-β expression and CD4+CD25+ regulatory T cells suppress antibody formation against FIX.

  7. High-density growth arrest in Ras-transformed cells: low Cdk kinase activities in spite of absence of p27Kip Cdk-complexes

    DEFF Research Database (Denmark)

    Groth, Anja; Willumsen, Berthe Marie

    2005-01-01

    The ras oncogene transforms immortalized, contact-inhibited non-malignant murine fibroblasts into cells that are focus forming, exhibit increased saturation density, and are malignant in suitable hosts. Here, we examined changes in cell cycle control complexes as normal and Ras-transformed cells...

  8. miRNAs in lung cancer - Studying complex fingerprints in patient's blood cells by microarray experiments

    Directory of Open Access Journals (Sweden)

    Huwer Hanno

    2009-10-01

    Full Text Available Abstract Background Deregulated miRNAs are found in cancer cells and recently in blood cells of cancer patients. Due to their inherent stability miRNAs may offer themselves for blood based tumor diagnosis. Here we addressed the question whether there is a sufficient number of miRNAs deregulated in blood cells of cancer patients to be able to distinguish between cancer patients and controls. Methods We synthesized 866 human miRNAs and miRNA star sequences as annotated in the Sanger miRBase onto a microarray designed by febit biomed gmbh. Using the fully automated Geniom Real Time Analyzer platform, we analyzed the miRNA expression in 17 blood cell samples of patients with non-small cell lung carcinomas (NSCLC and in 19 blood samples of healthy controls. Results Using t-test, we detected 27 miRNAs significantly deregulated in blood cells of lung cancer patients as compared to the controls. Some of these miRNAs were validated using qRT-PCR. To estimate the value of each deregulated miRNA, we grouped all miRNAs according to their diagnostic information that was measured by Mutual Information. Using a subset of 24 miRNAs, a radial basis function Support Vector Machine allowed for discriminating between blood cellsamples of tumor patients and controls with an accuracy of 95.4% [94.9%-95.9%], a specificity of 98.1% [97.3%-98.8%], and a sensitivity of 92.5% [91.8%-92.5%]. Conclusion Our findings support the idea that neoplasia may lead to a deregulation of miRNA expression in blood cells of cancer patients compared to blood cells of healthy individuals. Furthermore, we provide evidence that miRNA patterns can be used to detect human cancers from blood cells.

  9. Complex T Cell Interactions Contribute to Helicobacter pylori Gastritis in Mice

    Science.gov (United States)

    Gray, Brian M.; Fontaine, Clinton A.; Poe, Sara A.

    2013-01-01

    Disease due to the gastric pathogen Helicobacter pylori varies in severity from asymptomatic to peptic ulcer disease and cancer. Accumulating evidence suggests that one source of this variation is an abnormal host response. The goal of this study was to use a mouse model of H. pylori gastritis to investigate the roles of regulatory T cells (Treg) as well as proinflammatory T cells (Th1 and Th17) in gastritis, gastric T cell engraftment, and gastric cytokine production. Our results support published data indicating that severe gastritis in T cell recipient mice is due to failure of Treg engraftment, that Treg ameliorate gastritis, and that the proinflammatory response is attributable to interactions between several cell subsets and cytokines. We confirmed that gamma interferon (IFN-γ) is essential for induction of gastritis but showed that IFN-γ-producing CD4 T cells are not necessary. Interleukin 17A (IL-17A) also contributed to gastritis, but to a lesser extent than IFN-γ. Tumor necrosis factor alpha (TNF-α) and IL-17F were also elevated in association with disease. These results indicate that while H. pylori-specific CD4+ T cells and IFN-γ are both essential for induction of gastritis due to H. pylori, IFN-γ production by T cells is not essential. It is likely that other proinflammatory cytokines, such as IL-17F and TNF-α, shown to be elevated in this model, also contribute to the induction of disease. We suggest that gastritis due to H. pylori is associated with loss of immunoregulation and alteration of several cytokines and cell subsets and cannot be attributed to a single immune pathway. PMID:23264048

  10. Synthesis and Characterization of New Palladium(II) Thiosemicarbazone Complexes and Their Cytotoxic Activity against Various Human Tumor Cell Lines

    Science.gov (United States)

    Hernández, Wilfredo; Paz, Juan; Carrasco, Fernando; Spodine, Evgenia; Manzur, Jorge; Sieler, Joachim; Blaurock, Steffen; Beyer, Lothar

    2013-01-01

    The palladium(II) bis-chelate complexes of the type [Pd(TSC1-5)2] (6–10), with their corresponding ligands 4-phenyl-1-(acetone)-thiosemicarbazone, HTSC1 (1), 4-phenyl-1-(2′-chloro-benzaldehyde)-thiosemicarbazone, HTSC2 (2), 4-phenyl-1-(3′-hydroxy-benzaldehyde)-thiosemicarbazone, HTSC3 (3), 4-phenyl-1-(2′-naphthaldehyde)-thiosemicarbazone, HTSC4 (4), and 4-phenyl-1-(1′-nitro-2′-naphthaldehyde)-thiosemicarbazone, HTSC5 (5), were synthesized and characterized by elemental analysis and spectroscopic techniques (IR and 1H- and 13C-NMR). The molecular structure of HTSC3, HTSC4, and [Pd(TSC1)2] (6) have been determined by single crystal X-ray crystallography. Complex 6 shows a square planar geometry with two deprotonated ligands coordinated to PdII through the azomethine nitrogen and thione sulfur atoms in a cis arrangement. The in vitro cytotoxic activity measurements indicate that the palladium(II) complexes (IC50 = 0.01–9.87 μM) exhibited higher antiproliferative activity than their free ligands (IC50 = 23.48–70.86 and >250 μM) against different types of human tumor cell lines. Among all the studied palladium(II) complexes, the [Pd(TSC3)2] (8) complex exhibited high antitumor activity on the DU145 prostate carcinoma and K562 chronic myelogenous leukemia cells, with low values of the inhibitory concentration (0.01 and 0.02 μM, resp.). PMID:24391528

  11. Planar Perovskite Solar Cells with High Open-Circuit Voltage Containing a Supramolecular Iron Complex as Hole Transport Material Dopant.

    Science.gov (United States)

    Saygili, Yasemin; Turren-Cruz, Silver-Hamill; Olthof, Selina; Saes, Bartholomeus Wilhelmus Henricus; Pehlivan, Ilknur Bayrak; Saliba, Michael; Meerholz, Klaus; Edvinsson, Tomas; Zakeeruddin, Shaik M; Grätzel, Michael; Correa-Baena, Juan-Pablo; Hagfeldt, Anders; Freitag, Marina; Tress, Wolfgang

    2018-04-26

    In perovskite solar cells (PSCs), the most commonly used hole transport material (HTM) is spiro-OMeTAD, which is typically doped by metalorganic complexes, for example, based on Co, to improve charge transport properties and thereby enhance the photovoltaic performance of the device. In this study, we report a new hemicage-structured iron complex, 1,3,5-tris(5'-methyl-2,2'-bipyridin-5-yl)ethylbenzene Fe(III)-tris(bis(trifluoromethylsulfonyl)imide), as a p-type dopant for spiro-OMeTAD. The formal redox potential of this compound was measured as 1.29 V vs. the standard hydrogen electrode, which is slightly (20 mV) more positive than that of the commercial cobalt dopant FK209. Photoelectron spectroscopy measurements confirm that the iron complex acts as an efficient p-dopant, as evidenced in an increase of the spiro-OMeTAD work function. When fabricating planar PSCs with the HTM spiro-OMeTAD doped by 5 mol % of the iron complex, a power conversion efficiency of 19.5 % (AM 1.5G, 100 mW cm -2 ) is achieved, compared to 19.3 % for reference devices with FK209. Open circuit voltages exceeding 1.2 V at 1 sun and reaching 1.27 V at 3 suns indicate that recombination at the perovskite/HTM interface is low when employing this iron complex. This work contributes to recent endeavors to reduce recombination losses in perovskite solar cells. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Analysis the macular ganglion cell complex thickness in monocular strabismic amblyopia patients by Fourier-domain OCT

    Directory of Open Access Journals (Sweden)

    Hong-Wei Deng

    2014-11-01

    Full Text Available AIM: To detect the macular ganglion cell complex thickness in monocular strabismus amblyopia patients, in order to explore the relationship between the degree of amblyopia and retinal ganglion cell complex thickness, and found out whether there is abnormal macular ganglion cell structure in strabismic amblyopia. METHODS: Using a fourier-domain optical coherence tomography(FD-OCTinstrument iVue®(Optovue Inc, Fremont, CA, Macular ganglion cell complex(mGCCthickness was measured and statistical the relation rate with the best vision acuity correction was compared Gman among 26 patients(52 eyesincluded in this study. RESULTS: The mean thickness of the mGCC in macular was investigated into three parts: centrial, inner circle(3mmand outer circle(6mm. The mean thicknesses of mGCC in central, inner and outer circle was 50.74±21.51μm, 101.4±8.51μm, 114.2±9.455μm in the strabismic amblyopia eyes(SAE, and 43.79±11.92μm,92.47±25.01μm, 113.3±12.88μm in the contralateral sound eyes(CSErespectively. There was no statistically significant difference among the eyes(P>0.05. But the best corrected vision acuity had a good correlation rate between mGcc thicknesses, which was better relative for the lower part than the upper part.CONCLUSION:There is a relationship between the amblyopia vision acuity and the mGCC thickness. Although there has not statistically significant difference of the mGCC thickness compared with the SAE and CSE. To measure the macular center mGCC thickness in clinic may understand the degree of amblyopia.

  13. SmgGDS is a transient nucleolar protein that protects cells from nucleolar stress and promotes the cell cycle by regulating DREAM complex gene expression.

    Science.gov (United States)

    Gonyo, P; Bergom, C; Brandt, A C; Tsaih, S-W; Sun, Y; Bigley, T M; Lorimer, E L; Terhune, S S; Rui, H; Flister, M J; Long, R M; Williams, C L

    2017-12-14

    The chaperone protein and guanine nucleotide exchange factor SmgGDS (RAP1GDS1) is a key promoter of cancer cell proliferation and tumorigenesis. SmgGDS undergoes nucleocytoplasmic shuttling, suggesting that it has both cytoplasmic and nuclear functions that promote cancer. Previous studies indicate that SmgGDS binds cytoplasmic small GTPases and promotes their trafficking to the plasma membrane. In contrast, little is known about the functions of SmgGDS in the nucleus, or how these nuclear functions might benefit cancer cells. Here we show unique nuclear localization and regulation of gene transcription pathways by SmgGDS. Strikingly, SmgGDS depletion significantly reduces expression of over 600 gene products that are targets of the DREAM complex, which is a transcription factor complex that regulates expression of proteins controlling the cell cycle. The cell cycle regulators E2F1, MYC, MYBL2 (B-Myb) and FOXM1 are among the DREAM targets that are diminished by SmgGDS depletion. E2F1 is well known to promote G1 cell cycle progression, and the loss of E2F1 in SmgGDS-depleted cells provides an explanation for previous reports that SmgGDS depletion characteristically causes a G1 cell cycle arrest. We show that SmgGDS localizes in nucleoli, and that RNAi-mediated depletion of SmgGDS in cancer cells disrupts nucleolar morphology, signifying nucleolar stress. We show that nucleolar SmgGDS interacts with the RNA polymerase I transcription factor upstream binding factor (UBF). The RNAi-mediated depletion of UBF diminishes nucleolar localization of SmgGDS and promotes proteasome-mediated degradation of SmgGDS, indicating that nucleolar sequestration of SmgGDS by UBF stabilizes SmgGDS protein. The ability of SmgGDS to interact with UBF and localize in the nucleolus is diminished by expressing DiRas1 or DiRas2, which are small GTPases that bind SmgGDS and act as tumor suppressors. Taken together, our results support a novel nuclear role for SmgGDS in protecting malignant

  14. PRM1 and KAR5 function in cell-cell fusion and karyogamy to drive distinct bisexual and unisexual cycles in the Cryptococcus pathogenic species complex.

    Directory of Open Access Journals (Sweden)

    Ci Fu

    2017-11-01

    Full Text Available Sexual reproduction is critical for successful evolution of eukaryotic organisms in adaptation to changing environments. In the opportunistic human fungal pathogens, the Cryptococcus pathogenic species complex, C. neoformans primarily undergoes bisexual reproduction, while C. deneoformans undergoes both unisexual and bisexual reproduction. During both unisexual and bisexual cycles, a common set of genetic circuits regulates a yeast-to-hyphal morphological transition, that produces either monokaryotic or dikaryotic hyphae. As such, both the unisexual and bisexual cycles can generate genotypic and phenotypic diversity de novo. Despite the similarities between these two cycles, genetic and morphological differences exist, such as the absence of an opposite mating-type partner and monokaryotic instead of dikaryotic hyphae during C. deneoformans unisexual cycle. To better understand the similarities and differences between these modes of sexual reproduction, we focused on two cellular processes involved in sexual reproduction: cell-cell fusion and karyogamy. We identified orthologs of the plasma membrane fusion protein Prm1 and the nuclear membrane fusion protein Kar5 in both Cryptococcus species, and demonstrated their conserved roles in cell fusion and karyogamy during C. deneoformans α-α unisexual reproduction and C. deneoformans and C. neoformans a-α bisexual reproduction. Notably, karyogamy occurs inside the basidum during bisexual reproduction in C. neoformans, but often occurs earlier following cell fusion during bisexual reproduction in C. deneoformans. Characterization of these two genes also showed that cell fusion is dispensable for solo unisexual reproduction in C. deneoformans. The blastospores produced along hyphae during C. deneoformans unisexual reproduction are diploid, suggesting that diploidization occurs early during hyphal development, possibly through either an endoreplication pathway or cell fusion-independent karyogamy

  15. PEGylated anticancer-carbon nanotubes complex targeting mitochondria of lung cancer cells

    Science.gov (United States)

    Kim, Sang-Woo; Lee, Yeon Kyung; Lee, Jong Yeon; Hong, Jeong Hee; Khang, Dongwoo

    2017-11-01

    Although activating apoptosis in cancer cells by targeting the mitochondria is an effective strategy for cancer therapy, insufficient targeting of the mitochondria in cancer cells restricts the availability in clinical treatment. Here, we report on a polyethylene glycol-coated carbon nanotube (CNT)-ABT737 nanodrug that improves the mitochondrial targeting of lung cancer cells. The polyethylene glycol-coated CNT-ABT737 nanodrug internalized into the early endosomes via macropinocytosis and clathrin-mediated endocytosis in advance of early endosomal escape and delivered into the mitochondria. Cytosol release of the nanodrug led to apoptosis of lung cancer cells by abruption of the mitochondrial membrane potential, inducing Bcl-2-mediated apoptosis and generating intracellular reactive oxygen species. As such, this study provides an effective strategy for increasing the anti-lung cancer efficacy by increasing mitochondria accumulation rate of cytosol released anticancer nanodrugs.

  16. Reconstructing each cell's genome within complex microbial communities-dream or reality?

    Science.gov (United States)

    Clingenpeel, Scott; Clum, Alicia; Schwientek, Patrick; Rinke, Christian; Woyke, Tanja

    2014-01-01

    As the vast majority of microorganisms have yet to be cultivated in a laboratory setting, access to their genetic makeup has largely been limited to cultivation-independent methods. These methods, namely metagenomics and more recently single-cell genomics, have become cornerstones for microbial ecology and environmental microbiology. One ultimate goal is the recovery of genome sequences from each cell within an environment to move toward a better understanding of community metabolic potential and to provide substrate for experimental work. As single-cell sequencing has the ability to decipher all sequence information contained in an individual cell, this method holds great promise in tackling such challenge. Methodological limitations and inherent biases however do exist, which will be discussed here based on environmental and benchmark data, to assess how far we are from reaching this goal.

  17. Back to the drawing board: Understanding the complexity of hepatic innate lymphoid cells.

    Science.gov (United States)

    Marotel, Marie; Hasan, Uzma; Viel, Sébastien; Marçais, Antoine; Walzer, Thierry

    2016-09-01

    Recent studies of immune populations in nonlymphoid organs have highlighted the great diversity of the innate lymphoid system. It has also become apparent that mouse and human innate lymphoid cells (ILCs) have distinct phenotypes and properties. In this issue of the European Journal of Immunology, Harmon et al. [Eur. J. Immunol. 2016. 46: 2111-2120] characterized human hepatic NK-cell subsets. The authors report that hepatic CD56(bright) NK cells resemble mouse liver ILC1s in that they express CXCR6 and have an immature phenotype. However, unlike mouse ILC1s, they express high levels of Eomes and low levels of T-bet, and upon stimulation with tumor cells, secrete low amounts of cytokines. These unexpected findings further support the differences between human and mouse immune populations and prompt the study of the role of hepatic ILC subsets in immune responses. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Complexity of the influence of gangliosides on histamine release from human basophils and rat mast cells

    DEFF Research Database (Denmark)

    Jensen, C; Svendsen, U G; Thastrup, Ole

    1987-01-01

    The influence of exogenous addition of gangliosides on histamine release from human basophils and rat mast cells was examined in vitro. Gangliosides dose-dependently inhibited histamine release, and this inhibition was dependent on the ganglioside sialic acid content, since GT1b, having 3 sialic...... was reflected in the sensitivity of the cells to extracellular calcium, since inhibition of the release could be counteracted by increasing the extracellular concentration of calcium....

  19. The distribution of a new /sup 111/In-Bleomycin complex in tumor cells by autoradiography

    International Nuclear Information System (INIS)

    Hou, D.Y.; Maruyama, Y.

    1987-01-01

    A new radioactive form of Bleomycin (/sup 111/In-BLMC) was effective for tumor imaging and therapy in mouse glioma and human small cell lung cancer (SCLC) cells. The distribution of drug in tumor cells was investigated by autoradiography. Human small cell lung cancer (N417 and H526, NCI) were exposed to /sup 111/ InCl/sub 3/ and (25-150 μCi/ml) or /sup 111/In-BLMC (25-150 μCi) carried by 15-25 μg BLM/ml) in 37 0 C for 1 hr, 3 hr or 24 hr, washed with fresh medium, and spread. The slides were smeared with NTB/sub 2/ or NTB/sub 3/ emulsion by using wet-mounting or dry-mounting technique and developed 3-14 days. The /sup 111/In-BLMC localized on the cell nucleus (47.8%) and nuclear membrane (29.2%); /sup 111/InCl/sub 3/ located mainly in the cytoplasm (45.8%). This indicates that the mechanism of killing of tumor cells may be related to the drug uptake and distribution of /sup 111/In-BLMC. A nuclear and nuclear membrane localization would favor damage to chromosomes and DNA

  20. Oxidative Stress Regulation on Endothelial Cells by Hydrophilic Astaxanthin Complex: Chemical, Biological, and Molecular Antioxidant Activity Evaluation

    Directory of Open Access Journals (Sweden)

    M. Zuluaga

    2017-01-01

    Full Text Available An imbalance in the reactive oxygen species (ROS homeostasis is involved in the pathogenesis of oxidative stress-related diseases. Astaxanthin, a xanthophyll carotenoid with high antioxidant capacities, has been shown to prevent the first stages of oxidative stress. Here, we evaluate the antioxidant capacities of astaxanthin included within hydroxypropyl-beta-cyclodextrin (CD-A to directly and indirectly reduce the induced ROS production. First, chemical methods were used to corroborate the preservation of astaxanthin antioxidant abilities after inclusion. Next, antioxidant scavenging properties of CD-A to inhibit the cellular and mitochondrial ROS by reducing the disturbance in the redox state of the cell and the infiltration of lipid peroxidation radicals were evaluated. Finally, the activation of endogenous antioxidant PTEN/AKT, Nrf2/HO-1, and NQOI gene and protein expression supported the protective effect of CD-A complex on human endothelial cells under stress conditions. Moreover, a nontoxic effect on HUVEC was registered after CD-A complex supplementation. The results reported here illustrate the need to continue exploring the interesting properties of this hydrophilic antioxidant complex to assist endogenous systems to counteract the ROS impact on the induction of cellular oxidative stress state.

  1. ["Point by point" approach to structure-function correlation of glaucoma on the ganglion cell complex in the posterior pole].

    Science.gov (United States)

    Zeitoun, M

    2017-01-01

    To try to establish a "point by point" relationship between the local thickness of the retinal ganglion cell complex and its sensitivity. In total, 104 glaucomatous eyes of 89 patients with a confirmed 24-2 visual field, were measured by superimposing the visual field, using imaging software, with the Wide 40° by 30° measurements of retinal ganglion cell complex obtained from the Topcon © 3D 2000 OCT, after upward adjustment, inversion and scaling. Visual fields were classified into two groups according to the extent of the disease: 58 mild to moderate (MD up to -12dB), and 46 severe (MD beyond -12dB). The 6mm by 6mm central region, equipped with a normative database, was studied, corresponding to 16 points in the visual field. These points were individually matched one by one to the local ganglion cell complex, which was classified into 2 groups depending on whether it was greater or less than 70 microns. The normative database confirmed the pathological nature of the thin areas, with a significance of 95 to 99%. Displacement of central retinal ganglion cells was compensated for. Of 1664 points (16 central points for 104 eyes), 283 points were found to be "borderline" and excluded. Of the 1381 analyzed points, 727 points were classified as "over 70 microns" and 654 points "under 70 microns". (1) For all stages combined, 85.8% of the 727 points which were greater than 70 microns had a deviation between -3 and +3dB: areas above 70 microns had no observable loss of light sensitivity. (2) In total, 92.5% of the 428 points having a gap ranging from -6 to -35dB were located on ganglion cell complex areas below 70 microns: functional visual loss was identified in thin areas, which were less than 70 microns. (3) Areas which were less than 70 microns, that is 654 points, had quite variable sensitivity and can be divided into three groups: the first with preserved sensitivity, another with obliterated sensitivity, and an intermediate group connecting

  2. Celastrol targets mitochondrial respiratory chain complex I to induce reactive oxygen species-dependent cytotoxicity in tumor cells

    Directory of Open Access Journals (Sweden)

    Xu Yuanji

    2011-05-01

    Full Text Available Abstract Background Celastrol is an active ingredient of the traditional Chinese medicinal plant Tripterygium Wilfordii, which exhibits significant antitumor activity in different cancer models in vitro and in vivo; however, the lack of information on the target and mechanism of action of this compound have impeded its clinical application. In this study, we sought to determine the mode of action of celastrol by focusing on the processes that mediate its anticancer activity. Methods The downregulation of heat shock protein 90 (HSP90 client proteins, phosphorylation of c-Jun NH2-terminal kinase (JNK, and cleavage of PARP, caspase 9 and caspase 3 were detected by western blotting. The accumulation of reactive oxygen species (ROS was analyzed by flow cytometry and fluorescence microscopy. Cell cycle progression, mitochondrial membrane potential (MMP and apoptosis were determined by flow cytometry. Absorption spectroscopy was used to determine the activity of mitochondrial respiratory chain (MRC complexes. Results Celastrol induced ROS accumulation, G2-M phase blockage, apoptosis and necrosis in H1299 and HepG2 cells in a dose-dependent manner. N-acetylcysteine (NAC, an antioxidative agent, inhibited celastrol-induced ROS accumulation and cytotoxicity. JNK phosphorylation induced by celastrol was suppressed by NAC and JNK inhibitor SP600125 (SP. Moreover, SP significantly inhibited celastrol-induced loss of MMP, cleavage of PARP, caspase 9 and caspase 3, mitochondrial translocation of Bad, cytoplasmic release of cytochrome c, and cell death. However, SP did not inhibit celastrol-induced ROS accumulation. Celastrol downregulated HSP90 client proteins but did not disrupt the interaction between HSP90 and cdc37. NAC completely inhibited celastrol-induced decrease of HSP90 client proteins, catalase and thioredoxin. The activity of MRC complex I was completely inhibited in H1299 cells treated with 6 μM celastrol in the absence and presence of NAC

  3. IL-32 expression in the airway epithelial cells of patients with Mycobacterium avium complex lung disease.

    NARCIS (Netherlands)

    Bai, X.; Ovrutsky, A.R.; Kartalija, M.; Chmura, K.; Kamali, A.; Honda, J.R.; Oberley-Deegan, R.E.; Dinarello, C.A.; Crapo, J.D.; Chang, L.Y.; Chan, E.D.

    2011-01-01

    Lung disease due to Mycobacterium avium complex (MAC) organisms is increasing. A greater understanding of the host immune response to MAC organisms will provide a foundation to develop novel therapies for these recalcitrant infections. IL-32 is a newly described pro-inflammatory cytokine that

  4. Genomewide effects of peroxisome proliferator-activated receptor gamma in macrophages and dendritic cells--revealing complexity through systems biology.

    Science.gov (United States)

    Cuaranta-Monroy, Ixchelt; Kiss, Mate; Simandi, Zoltan; Nagy, Laszlo

    2015-09-01

    Systems biology approaches have become indispensable tools in biomedical and basic research. These data integrating bioinformatic methods gained prominence after high-throughput technologies became available to investigate complex cellular processes, such as transcriptional regulation and protein-protein interactions, on a scale that had not been studied before. Immunology is one of the medical fields that systems biology impacted profoundly due to the plasticity of cell types involved and the accessibility of a wide range of experimental models. In this review, we summarize the most important recent genomewide studies exploring the function of peroxisome proliferator-activated receptor γ in macrophages and dendritic cells. PPARγ ChIP-seq experiments were performed in adipocytes derived from embryonic stem cells to complement the existing data sets and to provide comparators to macrophage data. Finally, lists of regulated genes generated from such experiments were analysed with bioinformatics and system biology approaches. We show that genomewide studies utilizing high-throughput data acquisition methods made it possible to gain deeper insights into the role of PPARγ in these immune cell types. We also demonstrate that analysis and visualization of data using network-based approaches can be used to identify novel genes and functions regulated by the receptor. The example of PPARγ in macrophages and dendritic cells highlights the crucial importance of systems biology approaches in establishing novel cellular functions for long-known signaling pathways. © 2015 Stichting European Society for Clinical Investigation Journal Foundation.

  5. Fission yeast Sec3 and Exo70 are transported on actin cables and localize the exocyst complex to cell poles.

    Directory of Open Access Journals (Sweden)

    Felipe O Bendezú

    Full Text Available The exocyst complex is essential for many exocytic events, by tethering vesicles at the plasma membrane for fusion. In fission yeast, polarized exocytosis for growth relies on the combined action of the exocyst at cell poles and myosin-driven transport along actin cables. We report here the identification of fission yeast Schizosaccharomyces pombe Sec3 protein, which we identified through sequence homology of its PH-like domain. Like other exocyst subunits, sec3 is required for secretion and cell division. Cells deleted for sec3 are only conditionally lethal and can proliferate when osmotically stabilized. Sec3 is redundant with Exo70 for viability and for the localization of other exocyst subunits, suggesting these components act as exocyst tethers at the plasma membrane. Consistently, Sec3 localizes to zones of growth independently of other exocyst subunits but depends on PIP(2 and functional Cdc42. FRAP analysis shows that Sec3, like all other exocyst subunits, localizes to cell poles largely independently of the actin cytoskeleton. However, we show that Sec3, Exo70 and Sec5 are transported by the myosin V Myo52 along actin cables. These data suggest that the exocyst holocomplex, including Sec3 and Exo70, is present on exocytic vesicles, which can reach cell poles by either myosin-driven transport or random walk.

  6. Interaction of cationic carbosilane dendrimers and their complexes with siRNA with erythrocytes and red blood cell ghosts.

    Science.gov (United States)

    Wrobel, Dominika; Kolanowska, Katarzyna; Gajek, Arkadiusz; Gomez-Ramirez, Rafael; de la Mata, Javier; Pedziwiatr-Werbicka, Elżbieta; Klajnert, Barbara; Waczulikova, Iveta; Bryszewska, Maria

    2014-03-01

    We have investigated the interactions between cationic NN16 and BDBR0011 carbosilane dendrimers with red blood cells or their cell membranes. The carbosilane dendrimers used possess 16 cationic functional groups. Both the dendrimers are made of water-stable carbon-silicon bonds, but NN16 possesses some oxygen-silicon bonds that are unstable in water. The nucleic acid used in the experiments was targeted against GAG-1 gene from the human immunodeficiency virus, HIV-1. By binding to the outer leaflet of the membrane, carbosilane dendrimers decreased the fluidity of the hydrophilic part of the membrane but increased the fluidity of the hydrophobic interior. They induced hemolysis, but did not change the morphology of the cells. Increasing concentrations of dendrimers induced erythrocyte aggregation. Binding of short interfering ribonucleic acid (siRNA) to a dendrimer molecule decreased the availability of cationic groups and diminished their cytotoxicity. siRNA-dendrimer complexes changed neither the fluidity of biological membranes nor caused cell hemolysis. Addition of dendriplexes to red blood cell suspension induced echinocyte formation. Copyright © 2013 Elsevier B.V. All rights reserved.

  7. Platinum(IV) complex LA-12 induces cell cycle phase specific apoptosis in colon carcinoma cell line HCT-116

    Czech Academy of Sciences Publication Activity Database

    Vondálová Blanářová, Olga; Jelínková, Iva; Jendželovský, R.; Souček, Karel; Hofmanová, Jiřina; Sova, P.; Kozubík, Alois

    2009-01-01

    Roč. 276, č. 1 (2009), s. 256-257 ISSN 1742-464X. [34th FEBS Congress. 04.07.2009-09.07.2009, Prague] R&D Projects: GA AV ČR(CZ) 1QS500040507; GA ČR(CZ) GA301/07/1557 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : platinum drugs * cell cycle apoptosis Subject RIV: BO - Biophysics

  8. CRIM1 Complexes with ß-catenin and Cadherins, Stabilizes Cell-Cell Junctions and Is Critical for Neural Morphogenesis

    OpenAIRE

    Ponferrada, Virgilio G.; Fan, Jieqing; Vallance, Jefferson E.; Hu, Shengyong; Mamedova, Aygun; Rankin, Scott A.; Kofron, Matthew; Zorn, Aaron M.; Hegde, Rashmi S.; Lang, Richard A.

    2012-01-01

    In multicellular organisms, morphogenesis is a highly coordinated process that requires dynamically regulated adhesion between cells. An excellent example of cellular morphogenesis is the formation of the neural tube from the flattened epithelium of the neural plate. Cysteine-rich motor neuron protein 1 (CRIM1) is a single-pass (type 1) transmembrane protein that is expressed in neural structures beginning at the neural plate stage. In the frog Xenopus laevis, loss of function studies using C...

  9. A holistic approach to dissecting SPARC family protein complexity reveals FSTL-1 as an inhibitor of pancreatic cancer cell growth.

    Science.gov (United States)

    Viloria, Katrina; Munasinghe, Amanda; Asher, Sharan; Bogyere, Roberto; Jones, Lucy; Hill, Natasha J

    2016-11-25

    SPARC is a matricellular protein that is involved in both pancreatic cancer and diabetes. It belongs to a wider family of proteins that share structural and functional similarities. Relatively little is known about this extended family, but evidence of regulatory interactions suggests the importance of a holistic approach to their study. We show that Hevin, SPOCKs, and SMOCs are strongly expressed within islets, ducts, and blood vessels, suggesting important roles for these proteins in the normal pancreas, while FSTL-1 expression is localised to the stromal compartment reminiscent of SPARC. In direct contrast to SPARC, however, FSTL-1 expression is reduced in pancreatic cancer. Consistent with this, FSTL-1 inhibited pancreatic cancer cell proliferation. The complexity of SPARC family proteins is further revealed by the detection of multiple cell-type specific isoforms that arise due to a combination of post-translational modification and alternative splicing. Identification of splice variants lacking a signal peptide suggests the existence of novel intracellular isoforms. This study underlines the importance of addressing the complexity of the SPARC family and provides a new framework to explain their controversial and contradictory effects. We also demonstrate for the first time that FSTL-1 suppresses pancreatic cancer cell growth.

  10. Regulation of the Min Cell Division Inhibition Complex by the Rcs Phosphorelay in Proteus mirabilis.

    Science.gov (United States)

    Howery, Kristen E; Clemmer, Katy M; Şimşek, Emrah; Kim, Minsu; Rather, Philip N

    2015-08-01

    A key regulator of swarming in Proteus mirabilis is the Rcs phosphorelay, which represses flhDC, encoding the master flagellar regulator FlhD4C2. Mutants in rcsB, the response regulator in the Rcs phosphorelay, hyperswarm on solid agar and differentiate into swarmer cells in liquid, demonstrating that this system also influences the expression of genes central to differentiation. To gain a further understanding of RcsB-regulated genes involved in swarmer cell differentiation, transcriptome sequencing (RNA-Seq) was used to examine the RcsB regulon. Among the 133 genes identified, minC and minD, encoding cell division inhibitors, were identified as RcsB-activated genes. A third gene, minE, was shown to be part of an operon with minCD. To examine minCDE regulation, the min promoter was identified by 5' rapid amplification of cDNA ends (5'-RACE), and both transcriptional lacZ fusions and quantitative real-time reverse transcriptase (qRT) PCR were used to confirm that the minCDE operon was RcsB activated. Purified RcsB was capable of directly binding the minC promoter region. To determine the role of RcsB-mediated activation of minCDE in swarmer cell differentiation, a polar minC mutation was constructed. This mutant formed minicells during growth in liquid, produced shortened swarmer cells during differentiation, and exhibited decreased swarming motility. This work describes the regulation and role of the MinCDE cell division system in P. mirabilis swarming and swarmer cell elongation. Prior to this study, the mechanisms that inhibit cell division and allow swarmer cell elongation were unknown. In addition, this work outlines for the first time the RcsB regulon in P. mirabilis. Taken together, the data presented in this study begin to address how P. mirabilis elongates upon contact with a solid surface. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  11. Interaction between NANOS2 and the CCR4-NOT deadenylation complex is essential for male germ cell development in mouse.

    Directory of Open Access Journals (Sweden)

    Atsushi Suzuki

    Full Text Available Nanos is one of the evolutionarily conserved proteins implicated in germ cell development and we have previously shown that it interacts with the CCR4-NOT deadenylation complex leading to the suppression of specific RNAs. However, the molecular mechanism and physiological significance of this interaction have remained elusive. In our present study, we identify CNOT1, a component of the CCR4-NOT deadenylation complex, as a direct factor mediating the interaction with NANOS2. We find that the first 10 amino acids (AAs of NANOS2 are required for this binding. We further observe that a NANOS2 mutant lacking these first 10 AAs (NANOS2-ΔN10 fails to rescue defects in the Nanos2-null mouse. Our current data thus indicate that the interaction with the CCR4-NOT deadenylation complex is essential for NANOS2 function. In addition, we further demonstrate that NANOS2-ΔN10 can associate with specific mRNAs as well as wild-type NANOS2, suggesting the existence of other NANOS2-associated factor(s that determine the specificity of RNA-binding independently of the CCR4-NOT deadenylation complex.

  12. Allogeneic major histocompatibility complex-mismatched equine bone marrow-derived mesenchymal stem cells are targeted for death by cytotoxic anti-major histocompatibility complex antibodies.

    Science.gov (United States)

    Berglund, A K; Schnabel, L V

    2017-07-01

    Allogeneic mesenchymal stem cells (MSCs) are a promising cell source for treating musculoskeletal injuries in horses. Controversy exists, however, over whether major histocompatibility complex (MHC)-mismatched MSCs are recognised by the recipient immune system and targeted for death by a cytotoxic antibody response. To determine if cytotoxic anti-MHC antibodies generated in vivo following MHC-mismatched MSC injections are capable of initiating complement-dependent cytotoxicity of MSCs. Experimental controlled study. Antisera previously collected at Days 0, 7, 14 and 21 post-injection from 4 horses injected with donor MHC-mismatched equine leucocyte antigen (ELA)-A2 haplotype MSCs and one control horse injected with donor MHC-matched ELA-A2 MSCs were utilised in this study. Antisera were incubated with ELA-A2 MSCs before adding complement in microcytotoxicity assays and cell death was analysed via eosin dye exclusion. ELA-A2 peripheral blood leucocytes (PBLs) were used in the assays as a positive control. Antisera from all 4 horses injected with MHC-mismatched MSCs contained antibodies that caused the death of ELA-A2 haplotype MSCs in the microcytotoxicity assays. In 2 of the 4 horses, antibodies were present as early as Day 7 post-injection. MSC death was consistently equivalent to that of ELA-A2 haplotype PBL death at all time points and antisera dilutions. Antisera from the control horse that was injected with MHC-matched MSCs did not contain cytotoxic ELA-A2 antibodies at any of the time points examined. This study examined MSC death in vitro only and utilized antisera from a small number of horses. The cytotoxic antibody response induced in recipient horses following injection with donor MHC-mismatched MSCs is capable of killing donor MSCs in vitro. These results suggest that the use of allogeneic MHC-mismatched MSCs must be cautioned against, not only for potential adverse events, but also for reduced therapeutic efficacy due to targeted MSC death. © 2016 The

  13. Interaction of Cosup(III) complexes with radiation on 'in vitro' mammalian cells and on molecular systems

    International Nuclear Information System (INIS)

    Alvarez, M.V.; Cabildo, P.; Lopez Zumel, M.C.

    1978-01-01

    Cosup(III) complexes could be considered as potential radiosensitizers in dependence of the electronic configuration of the metal. Due to the high redox state that causes its powerful electrophilia they possibly act through electron-affinic mechanisms. In this paper the effect of [Co(NH 3 ) 5 Cl]Cl 2 on TC.SV-40 hamster cells is reported, in aerobic and hypoxic conditions, and its action, at a non-toxic concentration, in a combined treatment with X-rays. Results show the small protection induced by the product in the survival curves. As a contribution to the knowledge of its possible mechanism of action some molecular proofs and the interaction test with -SHNP cellular groups are made. A high degree of reaction is observed as is the case with good radiosensitizers but at molecular level the behaviour of the complex formed between DNA and the product reveals mutual protection due to their corresponding radiodegradation. (orig.) [de

  14. Graph Cellular Automata with Relation-Based Neighbourhoods of Cells for Complex Systems Modelling: A Case of Traffic Simulation

    Directory of Open Access Journals (Sweden)

    Krzysztof Małecki

    2017-12-01

    Full Text Available A complex system is a set of mutually interacting elements for which it is possible to construct a mathematical model. This article focuses on the cellular automata theory and the graph theory in order to compare various types of cellular automata and to analyse applications of graph structures together with cellular automata. It proposes a graph cellular automaton with a variable configuration of cells and relation-based neighbourhoods (r–GCA. The developed mechanism enables modelling of phenomena found in complex systems (e.g., transport networks, urban logistics, social networks taking into account the interaction between the existing objects. As an implementation example, modelling of moving vehicles has been made and r–GCA was compared to the other cellular automata models simulating the road traffic and used in the computer simulation process.

  15. Evaluation of umbilical cord mesenchymal stem cells labeling with superparamagnetic iron oxide nanoparticles coated with dextran and complexed with Poly-L-Lysine

    International Nuclear Information System (INIS)

    Sibov, Tatiana Tais; Mamani, Javier Bustamante; Pavon, Lorena Favaro; Cardenas, Walter Humberto; Gamarra, Lionel Fernel; Miyaki, Liza Aya Mabuchi; Marti, Luciana Cavalheiro; Sardinha, Luiz Roberto; Oliveira, Daniela Mara de

    2012-01-01

    Objective: The objective of this study was to evaluate the effect of the labeling of umbilical cord vein derived mesenchymal stem cells with superparamagnetic iron oxide nanoparticles coated with dextran and complexed to a non-viral transfector agent transfector poly-L-lysine. Methods: The labeling of mesenchymal stem cells was performed using the superparamagnetic iron oxide nanoparticles/dextran complexed and not complexed to poly-L-lysine. Superparamagnetic iron oxide nanoparticles/dextran was incubated with poly-L-lysine in an ultrasonic sonicator at 37 deg C for 10 minutes for complex formation superparamagnetic iron oxide nanoparticles/dextran/poly-L-lysine by electrostatic interaction. Then, the mesenchymal stem cells were incubated overnight with the complex superparamagnetic iron oxide nanoparticles/dextran/poly-L-lysine and superparamagnetic iron oxide nanoparticles/dextran. After the incubation period the mesenchymal stem cells were evaluated by internalization of the complex superparamagnetic iron oxide nanoparticles/dextran/polyL-lysine and superparamagnetic iron oxide nanoparticles/dextran by Prussian Blue stain. Cellular viability of labeled mesenchymal stem cells was evaluated by cellular proliferation assay using 5,6-carboxyfluorescein-succinimidyl ester method and apoptosis detection by Annexin V- Propidium Iodide assay. Results: mesenchymal stem cells labeled with superparamagnetic iron oxide nanoparticles/ dextran without poly-L-lysine not internalized efficiently the superparamagnetic iron oxide nanoparticles due to its low presence detected within cells. Mesenchymal stem cells labeled with the complex superparamagnetic iron oxide nanoparticles/dextran/polyL-lysine efficiently internalized the superparamagnetic iron oxide nanoparticles due to greater presence in the cells interior. The viability and apoptosis assays demonstrated that the mesenchymal stem cells labeled and not labeled respectively with the superparamagnetic iron oxide

  16. Thermal coupling of a high temperature PEM fuel cell with a complex hydride tank

    DEFF Research Database (Denmark)

    Pfeifer, P.; Wall, C.; Jensen, Jens Oluf

    2009-01-01

    the possibilities of a thermal coupling of a high temperature PEM fuel cell operating at 160-200 degrees C. The starting temperatures and temperature hold-times before starting fuel cell operation, the heat transfer characteristics of the hydride storage tanks, system temperature, fuel cell electrical power......Sodium alanate doped with cerium catalyst has been proven to have fast kinetics for hydrogen ab- and de-sorption as well as a high gravimetric storage density around 5 wt%. The kinetics of hydrogen sorption can be improved by preparing the alanate as nanocrystalline material. However, the second...... decomposition step, i.e. the decomposition of the hexahydride to sodium hydride and aluminium which refers to 1.8 wt% hydrogen is supposed to happen above 110 degrees C. The discharge of the material is thus limited by the level of heat supplied to the hydride storage tank. Therefore, we evaluated...

  17. Regulation of the expression of GARP/latent-TGF-β1 complexes on mouse T cells and their role in Regulatory T Cell and Th17 differentiation1

    Science.gov (United States)

    Edwards, Justin P.; Fujii, Hodaka; Zhou, Angela X.; Creemers, John; Unutmaz, Derya; Shevach, Ethan M.

    2013-01-01

    GARP/LRRC32 has previously been defined as a marker of activated human regulatory T-cells (Tregs) that is responsible for surface localization of latent TGF-β1. We find that GARP and latent TGF-β1 are also found on mouse Tregs activated via TCR stimulation, but in contrast to human Tregs, GARP is also expressed at a low level on resting Tregs. The expression of GARP can be upregulated on mouse Tregs by IL-2 or IL-4 exposure in the absence of TCR signaling. GARP is expressed at a low level on Tregs within the thymus and Treg precursors from the thymus concomitantly express GARP and Foxp3 upon exposure to IL-2. The expression of GARP is independent of TGF-β1 and TGF-β1 loading into GARP and is independent of furin-mediated processing of pro-TGF-β1 to latent TGF-β1. Specific deletion of GARP in CD4+ T cells results in lack of expression of latent-TGF-β1 on activated Tregs. GARP-deficient Tregs develop normally, are present in normal numbers in peripheral tissues, and are fully competent suppressors of the activation of T conventional cells in vitro. Activated Tregs expressing GARP/latent-TGF-β1 complexes are potent inducers of Th17 differentiation in the presence of exogenous IL-6 and inducers of Treg in the presence of IL-2. Induction of both Th17 producing cells and Treg is preferentially induced by Tregs expressing the latent-TGF-β1/GARP complex on their cell surface rather than by secreted latent-TGF-β1. PMID:23645881

  18. Regulation of the expression of GARP/latent TGF-β1 complexes on mouse T cells and their role in regulatory T cell and Th17 differentiation.

    Science.gov (United States)

    Edwards, Justin P; Fujii, Hodaka; Zhou, Angela X; Creemers, John; Unutmaz, Derya; Shevach, Ethan M

    2013-06-01

    GARP/LRRC32 was defined as a marker of activated human regulatory T cells (Tregs) that is responsible for surface localization of latent TGF-β1. We find that GARP and latent TGF-β1 are also found on mouse Tregs activated via TCR stimulation; however, in contrast to human Tregs, GARP is also expressed at a low level on resting Tregs. The expression of GARP can be upregulated on mouse Tregs by IL-2 or IL-4 exposure in the absence of TCR signaling. GARP is expressed at a low level on Tregs within the thymus, and Treg precursors from the thymus concomitantly express GARP and Foxp3 upon exposure to IL-2. The expression of GARP is independent of TGF-β1 and TGF-β1 loading into GARP and is independent of furin-mediated processing of pro-TGF-β1 to latent TGF-β1. Specific deletion of GARP in CD4(+) T cells results in lack of expression of latent TGF-β1 on activated Tregs. GARP-deficient Tregs develop normally, are present in normal numbers in peripheral tissues, and are fully competent suppressors of the activation of conventional T cells in vitro. Activated Tregs expressing GARP/latent TGF-β1 complexes are potent inducers of Th17 differentiation in the presence of exogenous IL-6 and inducers of Treg in the presence of IL-2. Induction of both Th17-producing cells and Tregs is caused preferentially by Tregs expressing the latent TGF-β1/GARP complex on their cell surface rather than by secreted latent TGF-β1.

  19. Mapping the Complex Morphology of Cell Interactions with Nanowire Substrates Using FIB-SEM

    DEFF Research Database (Denmark)

    Wierzbicki, Rafal; Købler, Carsten; Jensen, Mikkel Ravn Boye

    2013-01-01

    Using high resolution focused ion beam scanning electron microscopy (FIB-SEM) we study the details of cell-nanostructure interactions using serial block face imaging. 3T3 Fibroblast cellular monolayers are cultured on flat glass as a control surface and on two types of nanostructured scaffold...... substrates made from silicon black (Nanograss) with low- and high nanowire density. After culturing for 72 hours the cells were fixed, heavy metal stained, embedded in resin, and processed with FIB-SEM block face imaging without removing the substrate. The sample preparation procedure, image acquisition...

  20. Directional Migration in Esophageal Squamous Cell Carcinoma (ESCC) is Epigenetically Regulated by SET Nuclear Oncogene, a Member of the Inhibitor of Histone Acetyltransferase Complex

    OpenAIRE

    Xiang Yuan; Xinshuai Wang; Bianli Gu; Yingjian Ma; Yiwen Liu; Man Sun; Jinyu Kong; Wei Sun; Huizhi Wang; Fuyou Zhou; Shegan Gao

    2017-01-01

    Directional cell migration is of fundamental importance to a variety of biological events, including metastasis of malignant cells. Herein, we specifically investigated SET oncoprotein, a subunit of the recently identified inhibitor of acetyltransferases (INHAT) complex and identified its role in the establishment of front–rear cell polarity and directional migration in Esophageal Squamous Cell Carcinoma (ESCC). We further define the molecular circuits that govern these processes by showing t...

  1. Interaction proteomics analysis of polycomb proteins defines distinct PRC1 complexes in mammalian cells

    DEFF Research Database (Denmark)

    Vandamme, Julien; Völkel, Pamela; Rosnoblet, Claire

    2011-01-01

    Polycomb group (PcG) proteins maintain transcriptional repression of hundreds of genes involved in development, signaling or cancer using chromatin-based epigenetic mechanisms. Biochemical studies in Drosophila have revealed that PcG proteins associate in at least two classes of protein complexes...... known as Polycomb repressive complexes 1 and 2 (PRC1 and PRC2). Drosophila core PRC1 is composed of four subunits, Polycomb (Pc), Sex combs extra (Sce), Polyhomeotic (Ph), and Posterior sex combs (Psc). Each of these proteins has multiple orthologs in vertebrates classified respectively as the CBX, RING...... in order to identify interacting partners of CBX family proteins under the same experimental conditions. Our analysis identified with high confidence about 20 proteins co-eluted with CBX2 and CBX7 tagged proteins, about 40 with CBX4, and around 60 with CBX6 and CBX8. We provide evidences that the CBX...

  2. Multifunctional Pt(II) Reagents: Covalent Modifications of Pt Complexes Enable Diverse Structural Variation and In-Cell Detection.

    Science.gov (United States)

    White, Jonathan D; Haley, Michael M; DeRose, Victoria J

    2016-01-19

    To enhance the functionality of Pt-based reagents, several strategies have been developed that utilize Pt compounds modified with small, reactive handles. This Account encapsulates work done by us and other groups regarding the use of Pt(II) compounds with reactive handles for subsequent elaboration with fluorophores or other functional moieties. Described strategies include the incorporation of substituents for well-known condensation or nucleophilic displacement-type reactions and their use, for example, to tether spectroscopic handles to Pt reagents for in vivo investigation. Other chief uses of displacement-type reactions have included tethering various small molecules exhibiting pharmacological activity directly to Pt, thus adding synergistic effects. Click chemistry-based ligation techniques have also been applied, primarily with azide- and alkyne-appended Pt complexes. Orthogonally reactive click chemistry reactions have proven invaluable when more traditional nucleophilic displacement reactions induce side-reactivity with the Pt center or when systematic functionalization of a larger number of Pt complexes is desired. Additionally, a diverse assortment of Pt-fluorophore conjugates have been tethered via click chemistry conjugation. In addition to providing a convenient synthetic path for diversifying Pt compounds, the use of click-capable Pt complexes has proved a powerful strategy for postbinding covalent modification and detection with fluorescent probes. This strategy bypasses undesirable influences of the fluorophore camouflaged as reactivity due to Pt that may be present when detecting preattached Pt-fluorophore conjugates. Using postbinding strategies, Pt reagent distributions in HeLa and lung carcinoma (NCI-H460) cell cultures were observed with two different azide-modified Pt compounds, a monofunctional Pt(II)-acridine type and a difunctional Pt(II)-neutral complex. In addition, cellular distribution was observed with an alkyne-appended difunctional

  3. Core Transcriptional Regulatory Circuit Controlled by the TAL1 Complex in Human T Cell Acute Lymphoblastic Leukemia

    OpenAIRE

    Sanda, Takaomi; Lawton, Lee N.; Barrasa, M. Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A.; Jamieson, Catriona H.M.; Staudt, Louis M.; Young, Richard A.; Look, A. Thomas

    2012-01-01

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T-cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3 and RUNX1. We show that TAL1 forms a positive interconnected auto-regulatory loop with GATA3 and RUNX1, and that the TAL1 complex directly activates the MY...

  4. Human development VIII: a theory of "deep" quantum chemistry and cell consciousness: quantum chemistry controls genes and biochemistry to give cells and higher organisms consciousness and complex behavior.

    Science.gov (United States)

    Ventegodt, Søren; Hermansen, Tyge Dahl; Flensborg-Madsen, Trine; Nielsen, Maj Lyck; Merrick, Joav

    2006-11-14

    Deep quantum chemistry is a theory of deeply structured quantum fields carrying the biological information of the cell, making it able to remember, intend, represent the inner and outer world for comparison, understand what it "sees", and make choices on its structure, form, behavior and division. We suggest that deep quantum chemistry gives the cell consciousness and all the qualities and abilities related to consciousness. We use geometric symbolism, which is a pre-mathematical and philosophical approach to problems that cannot yet be handled mathematically. Using Occam's razor we have started with the simplest model that works; we presume this to be a many-dimensional, spiral fractal. We suggest that all the electrons of the large biological molecules' orbitals make one huge "cell-orbital", which is structured according to the spiral fractal nature of quantum fields. Consciousness of single cells, multi cellular structures as e.g. organs, multi-cellular organisms and multi-individual colonies (like ants) and human societies can thus be explained by deep quantum chemistry. When biochemical activity is strictly controlled by the quantum-mechanical super-orbital of the cell, this orbital can deliver energetic quanta as biological information, distributed through many fractal levels of the cell to guide form and behavior of an individual single or a multi-cellular organism. The top level of information is the consciousness of the cell or organism, which controls all the biochemical processes. By this speculative work inspired by Penrose and Hameroff we hope to inspire other researchers to formulate more strict and mathematically correct hypothesis on the complex and coherence nature of matter, life and consciousness.

  5. G9a coordinates with the RPA complex to promote DNA damage repair and cell survival.

    Science.gov (United States)

    Yang, Qiaoyan; Zhu, Qian; Lu, Xiaopeng; Du, Yipeng; Cao, Linlin; Shen, Changchun; Hou, Tianyun; Li, Meiting; Li, Zhiming; Liu, Chaohua; Wu, Di; Xu, Xingzhi; Wang, Lina; Wang, Haiying; Zhao, Ying; Yang, Yang; Zhu, Wei-Guo

    2017-07-25

    Histone methyltransferase G9a has critical roles in promoting cancer-cell growth and gene suppression, but whether it is also associated with the DNA damage response is rarely studied. Here, we report that loss of G9a impairs DNA damage repair and enhances the sensitivity of cancer cells to radiation and chemotherapeutics. In response to DNA double-strand breaks (DSBs), G9a is phosphorylated at serine 211 by casein kinase 2 (CK2) and recruited to chromatin. The chromatin-enriched G9a can then directly interact with replication protein A (RPA) and promote loading of the RPA and Rad51 recombinase to DSBs. This mechanism facilitates homologous recombination (HR) and cell survival. We confirmed the interaction between RPA and G9a to be critical for RPA foci formation and HR upon DNA damage. Collectively, our findings demonstrate a regulatory pathway based on CK2-G9a-RPA that permits HR in cancer cells and provide further rationale for the use of G9a inhibitors as a cancer therapeutic.

  6. Revealing the Differences Between Free and Complexed Enzyme Mechanisms and Factors Contributing to Cell Wall Recalcitrance

    Energy Technology Data Exchange (ETDEWEB)

    Resch, Michael G.; Donohoe, Byron; Ciesielski, Peter; Nill, Jennifer; McKinney, Kellene; Mittal, Ashutosh; Katahira, Rui; Himmel, Michael; Biddy, Mary; Beckham, Gregg; Decker, Steve

    2014-09-08

    Enzymatic depolymerization of polysaccharides is a key step in the production of fuels and chemicals from lignocellulosic biomass, and discovery of synergistic biomass-degrading enzyme paradigms will enable improved conversion processes. Historically, revealing insights into enzymatic saccharification mechanisms on plant cell walls has been hindered by uncharacterized substrates and low resolution.

  7. The Allelic Landscape of Human Blood Cell Trait Variation and Links to Common Complex Disease

    NARCIS (Netherlands)

    Astle, William J.; Elding, Heather; Jiang, Tao; Allen, Dave; Ruklisa, Dace; Mann, Alice L.; Mead, Daniel; Bouman, Heleen; Riveros-Mckay, Fernando; Kostadima, Myrto A.; Lambourne, John J.; Sivapalaratnam, Suthesh; Downes, Kate; Kundu, Kousik; Bomba, Lorenzo; Berentsen, Kim; Bradley, John R.; Daugherty, Louise C.; Delaneau, Olivier; Freson, Kathleen; Garner, Stephen F.; Grassi, Luigi; Guerrero, Jose; Haimel, Matthias; Janssen-Megens, Eva M.; Kaan, Anita; Kamat, Mihir; Kim, Bowon; Mandoli, Amit; Marchini, Jonathan; Martens, Joost H. A.; Meacham, Stuart; Megy, Karyn; O'Connell, Jared; Petersen, Romina; Sharifi, Nilofar; Sheard, Simon M.; Staley, James R.; Tuna, Salih; van der Ent, Martijn; Walter, Klaudia; Wang, Shuang-Yin; Wheeler, Eleanor; Wilder, Steven P.; Iotchkova, Valentina; Moore, Carmel; Sambrook, Jennifer; Stunnenberg, Hendrik G.; Di Angelantonio, Emanuele; Kaptoge, Stephen; Kuijpers, Taco W.; Carrillo-de-Santa-Pau, Enrique; Juan, David; Rico, Daniel; Valencia, Alfonso; Chen, Lu; Ge, Bing; Vasquez, Louella; Kwan, Tony; Garrido-Martín, Diego; Watt, Stephen; Yang, Ying; Guigo, Roderic; Beck, Stephan; Paul, Dirk S.; Pastinen, Tomi; Bujold, David; Bourque, Guillaume; Frontini, Mattia; Danesh, John; Roberts, David J.; Ouwehand, Willem H.; Butterworth, Adam S.; Soranzo, Nicole

    2016-01-01

    Many common variants have been associated with hematological traits, but identification of causal genes and pathways has proven challenging. We performed a genome-wide association analysis in the UK Biobank and INTERVAL studies, testing 29.5 million genetic variants for association with 36 red cell,

  8. Cytoplasmic organelles determine complexity and specificity of calcium signalling in adrenal chromaffin cells

    Czech Academy of Sciences Publication Activity Database

    Garsia-Sancho, J.; Verkhratsky, Alexei

    2008-01-01

    Roč. 192, č. 2 (2008), s. 263-271 ISSN 1748-1708 Institutional research plan: CEZ:AV0Z50390512 Keywords : Ca2+ signalling * calcium microdomains * chromaffin cells Subject RIV: JE - Non-nuclear Energetics, Energy Consumption ; Use Impact factor: 2.455, year: 2008

  9. Peptide specific expansion of CD8(+) T cells by recombinant plate bound MHC/peptide complexes

    DEFF Research Database (Denmark)

    Schmidt, Esben G W; Buus, Soren; Thorn, Mette

    2009-01-01

    to in vitro T cell stimulation was investigated. By use of an antigenic peptide derived from the cytomegalovirus (CMVp) we tested the stimulatory efficacy of recombinant plate bound MHC molecules (PB-MHC), being immobilized in culture plates. A single stimulation of non-adherent peripheral blood mononuclear...

  10. Degradation chemistry of RuLL´(NCS)2 complexes in the Dye-sensitized solar cell

    DEFF Research Database (Denmark)

    Lund, Torben

    will present and overview of our degradation investigations of the ruthenium dyes N719, Z907 and C106 with the general structure RuLL´(NCS)2 and show how detailed degradation mechanistic knowledge is important in the developing of DSC cells with improved thermal dye stability. The various ruthenium dye...

  11. Core transcriptional regulatory circuit controlled by the TAL1 complex in human T cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Sanda, Takaomi; Lawton, Lee N; Barrasa, M Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A; Jamieson, Catriona H M; Staudt, Louis M; Young, Richard A; Look, A Thomas

    2012-08-14

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3, and RUNX1. We show that TAL1 forms a positive interconnected autoregulatory loop with GATA3 and RUNX1 and that the TAL1 complex directly activates the MYB oncogene, forming a positive feed-forward regulatory loop that reinforces and stabilizes the TAL1-regulated oncogenic program. One of the critical downstream targets in this circuitry is the TRIB2 gene, which is oppositely regulated by TAL1 and E2A/HEB and is essential for the survival of T-ALL cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Entamoeba histolytica: a beta 1 integrin-like fibronectin receptor assembles a signaling complex similar to those of mammalian cells.

    Science.gov (United States)

    Flores-Robles, Donaciano; Rosales, Carlos; Rosales-Encina, José Luis; Talamás-Rohana, Patricia

    2003-01-01

    During tissue invasion, Entamoeba histolytica trophozoites interact with endothelial cells and extracellular matrix (ECM) proteins such as fibronectin (FN), collagen, and laminin. It has been demonstrated that trophozoites interact with FN through a beta1 integrin-like FN receptor (beta 1EhFNR), activating tyrosine kinases. In order to characterize the signaling process triggered by the amoebic receptor, activation, and association of tyrosine kinases and structural proteins were determined. As a result of FN binding by the beta 1EhFNR, the receptor itself, FAK, and paxillin were phosphorylated in tyrosine. Co-immunoprecipitation experiments showed that a multimolecular signaling complex was formed by the amoebic FN receptor, FAK, paxillin, and vinculin. These results strongly suggest that a signaling pathway, similar to the one used in mammalian cells, is activated when E. histolytica trophozoites adhere to FN.

  13. A newly identified essential complex, Dre2-Tah18, controls mitochondria integrity and cell death after oxidative stress in yeast.

    Directory of Open Access Journals (Sweden)

    Laurence Vernis

    Full Text Available A mutated allele of the essential gene TAH18 was previously identified in our laboratory in a genetic screen for new proteins interacting with the DNA polymerase delta in yeast [1]. The present work shows that Tah18 plays a role in response to oxidative stress. After exposure to lethal doses of H(2O(2, GFP-Tah18 relocalizes to the mitochondria and controls mitochondria integrity and cell death. Dre2, an essential Fe/S cluster protein and homologue of human anti-apoptotic Ciapin1, was identified as a molecular partner of Tah18 in the absence of stress. Moreover, Ciapin1 is able to replace yeast Dre2 in vivo and physically interacts with Tah18. Our results are in favour of an oxidative stress-induced cell death in yeast that involves mitochondria and is controlled by the newly identified Dre2-Tah18 complex.

  14. Control of flowering and cell fate by LIF2, an RNA binding partner of the polycomb complex component LHP1.

    Directory of Open Access Journals (Sweden)

    David Latrasse

    Full Text Available Polycomb Repressive Complexes (PRC modulate the epigenetic status of key cell fate and developmental regulators in eukaryotes. The chromo domain protein like heterochromatin protein1 (LHP1 is a subunit of a plant PRC1-like complex in Arabidopsis thaliana and recognizes histone H3 lysine 27 trimethylation, a silencing epigenetic mark deposited by the PRC2 complex. We have identified and studied an LHP1-Interacting Factor2 (LIF2. LIF2 protein has RNA recognition motifs and belongs to the large hnRNP protein family, which is involved in RNA processing. LIF2 interacts in vivo, in the cell nucleus, with the LHP1 chromo shadow domain. Expression of LIF2 was detected predominantly in vascular and meristematic tissues. Loss-of-function of LIF2 modifies flowering time, floral developmental homeostasis and gynoecium growth determination. lif2 ovaries have indeterminate growth and produce ectopic inflorescences with severely affected flowers showing proliferation of ectopic stigmatic papillae and ovules in short-day conditions. To look at how LIF2 acts relative to LHP1, we conducted transcriptome analyses in lif2 and lhp1 and identified a common set of deregulated genes, which showed significant enrichment in stress-response genes. By comparing expression of LHP1 targets in lif2, lhp1 and lif2 lhp1 mutants we showed that LIF2 can either antagonize or act with LHP1. Interestingly, repression of the FLC floral transcriptional regulator in lif2 mutant is accompanied by an increase in H3K27 trimethylation at the locus, without any change in LHP1 binding, suggesting that LHP1 is targeted independently from LIF2 and that LHP1 binding does not strictly correlate with gene expression. LIF2, involved in cell identity and cell fate decision, may modulate the activity of LHP1 at specific loci, during specific developmental windows or in response to environmental cues that control cell fate determination. These results highlight a novel link between plant RNA

  15. Failure to synthesize the human T-cell CD3-zeta chain and its consequence for the T-cell receptor-CD3 complex expression

    DEFF Research Database (Denmark)

    Geisler, C; Kuhlmann, J; Plesner, T

    1989-01-01

    components, the human T-cell tumour line Jurkat was chemically mutagenized followed by negative selection with F101.01 (a monoclonal antibody against the TcR-CD3 complex), and cloning. Growing clones were analysed for TcR-CD3 expression by immunofluorescence. One clone, J79, was found to express greatly...... diminished levels of TcR-CD3. This clone produced all the TcR-CD3 components except the CD3-zeta, as demonstrated by metabolic labelling and immunoprecipitation followed by one- and two-dimensional sodium dodecyl sulphate-polyacrylamide gel electrophoresis. These data indicate that the CD3-zeta determines...

  16. Tripartite polyionic complex (PIC) micelles as non-viral vectors for mesenchymal stem cell siRNA transfection.

    Science.gov (United States)

    Raisin, Sophie; Morille, Marie; Bony, Claire; Noël, Danièle; Devoisselle, Jean-Marie; Belamie, Emmanuel

    2017-08-22

    In the context of regenerative medicine, the use of RNA interference mechanisms has already proven its efficiency in targeting specific gene expression with the aim of enhancing, accelerating or, more generally, directing stem cell differentiation. However, achievement of good transfection levels requires the use of a gene vector. For in vivo applications, synthetic vectors are an interesting option to avoid possible issues associated with viral vectors (safety, production costs, etc.). Herein, we report on the design of tripartite polyionic complex micelles as original non-viral polymeric vectors suited for mesenchymal stem cell transfection with siRNA. Three micelle formulations were designed to exhibit pH-triggered disassembly in an acidic pH range comparable to that of endosomes. One formulation was selected as the most promising with the highest siRNA loading capacity while clearly maintaining pH-triggered disassembly properties. A thorough investigation of the internalization pathway of micelles into cells with tagged siRNA was made before showing an efficient inhibition of Runx2 expression in primary bone marrow-derived stem cells. This work evidenced PIC micelles as promising synthetic vectors that allow efficient MSC transfection and control over their behavior, from the perspective of their clinical use.

  17. A complex between contactin-1 and the protein tyrosine phosphatase PTPRZ controls the development of oligodendrocyte precursor cells

    Energy Technology Data Exchange (ETDEWEB)

    Lamprianou, Smaragda; Chatzopoulou, Elli; Thomas, Jean-Léon; Bouyain, Samuel; Harroch, Sheila (IP-Korea); (UPMC); (UMKC)

    2013-09-23

    The six members of the contactin (CNTN) family of neural cell adhesion molecules are involved in the formation and maintenance of the central nervous system (CNS) and have been linked to mental retardation and neuropsychiatric disorders such as autism. Five of the six CNTNs bind to the homologous receptor protein tyrosine phosphatases gamma (PTPRG) and zeta (PTPRZ), but the biological roles of these interactions remain unclear. We report here the cocrystal structure of the carbonic anhydrase-like domain of PTPRZ bound to tandem Ig repeats of CNTN1 and combine these structural data with binding assays to show that PTPRZ binds specifically to CNTN1 expressed at the surface of oligodendrocyte precursor cells. Furthermore, analyses of glial cell populations in wild-type and PTPRZ-deficient mice show that the binding of PTPRZ to CNTN1 expressed at the surface of oligodendrocyte precursor cells inhibits their proliferation and promotes their development into mature oligodendrocytes. Overall, these results implicate the PTPRZ/CNTN1 complex as a previously unknown modulator of oligodendrogenesis.

  18. A complex regulatory network coordinating cell cycles during C. elegans development is revealed by a genome-wide RNAi screen.

    Science.gov (United States)

    Roy, Sarah H; Tobin, David V; Memar, Nadin; Beltz, Eleanor; Holmen, Jenna; Clayton, Joseph E; Chiu, Daniel J; Young, Laura D; Green, Travis H; Lubin, Isabella; Liu, Yuying; Conradt, Barbara; Saito, R Mako

    2014-02-28

    The development and homeostasis of multicellular animals requires precise coordination of cell division and differentiation. We performed a genome-wide RNA interference screen in Caenorhabditis elegans to reveal the components of a regulatory network that promotes developmentally programmed cell-cycle quiescence. The 107 identified genes are predicted to constitute regulatory networks that are conserved among higher animals because almost half of the genes are represented by clear human orthologs. Using a series of mutant backgrounds to assess their genetic activities, the RNA interference clones displaying similar properties were clustered to establish potential regulatory relationships within the network. This approach uncovered four distinct genetic pathways controlling cell-cycle entry during intestinal organogenesis. The enhanced phenotypes observed for animals carrying compound mutations attest to the collaboration between distinct mechanisms to ensure strict developmental regulation of cell cycles. Moreover, we characterized ubc-25, a gene encoding an E2 ubiquitin-conjugating enzyme whose human ortholog, UBE2Q2, is deregulated in several cancers. Our genetic analyses suggested that ubc-25 acts in a linear pathway with cul-1/Cul1, in parallel to pathways employing cki-1/p27 and lin-35/pRb to promote cell-cycle quiescence. Further investigation of the potential regulatory mechanism demonstrated that ubc-25 activity negatively regulates CYE-1/cyclin E protein abundance in vivo. Together, our results show that the ubc-25-mediated pathway acts within a complex network that integrates the actions of multiple molecular mechanisms to control cell cycles during development. Copyright © 2014 Roy et al.

  19. Nuclear pore complex assembly and maintenance in POM121- and gp210-deficient cells

    DEFF Research Database (Denmark)

    Stavru, Fabrizia; Nautrup-Pedersen, Gitte; Cordes, Volker C

    2006-01-01

    So far, POM121 and gp210 are the only known anchoring sites of vertebrate nuclear pore complexes (NPCs) within the lipid bilayer of the nuclear envelope (NE) and, thus, are excellent candidates for initiating the NPC assembly process. Indeed, we demonstrate that POM121 can recruit several...... as depletion of POM121 from human fibroblasts, which do not express gp210, further suggest that NPCs can assemble or at least persist in a POM121- and gp210-free form. This points to extensive redundancies in protein-protein interactions within NPCs and suggests that vertebrate NPCs contain additional membrane...

  20. The interaction of platinum complexes with low doses of X-rays in hypoxic cells

    International Nuclear Information System (INIS)

    Skov, K.A.

    1992-01-01

    It is not clear why electron affinic compounds (O2, nitroimidazoles) should be more effective sensitizers at high doses (modify double events, 6, 16) while platinum complexes interact to a greater extent at low doses (modification of single-events). The possibility that crosslinks are involved requires further consideration. While DNA intrastrand crosslinks do not appear essential, future low dose experiments are planned to determine the role of interstrand and DNA-protein crosslinks in modifying response at clinical doses. (author). 31 refs., 2 tabs

  1. DNA interactions of dinuclear RuII arene antitumor complexes in cell-free media

    Czech Academy of Sciences Publication Activity Database

    Nováková, Olga; Nazarov, A.A.; Hartinger, Ch.G.; Keppler, B.K.; Brabec, Viktor

    2009-01-01

    Roč. 77, č. 3 (2009), s. 364-374 ISSN 0006-2952 R&D Projects: GA MŠk(CZ) LC06030; GA MŠk(CZ) ME08017; GA MŠk(CZ) OC08003; GA AV ČR(CZ) 1QS500040581; GA AV ČR(CZ) KAN200200651 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : dinuclear ruthenium complex * DNA * cross-links Subject RIV: BO - Biophysics Impact factor: 4.254, year: 2009

  2. A novel role for the cell cycle regulatory complex cyclin D1-CDK4 in gluconeogenesis

    OpenAIRE

    Hosooka, Tetsuya; Ogawa, Wataru

    2016-01-01

    Dysregulation of gluconeogenesis is a key pathological feature of type 2 diabetes. However, the molecular mechanisms underlying the regulation of gluconeogenesis remain unclear. Bhalla et?al. recently reported that cyclin D1 suppresses hepatic gluconeogenesis through CDK4?dependent phosphorylation of PGC1alpha and consequent inhibition of its activity. The cyclin D1?CDK4 might thus serve as an important link between the cell cycle and control of energy metabolism through modulation of PGC1alp...

  3. Cut loose and run: The complex role of ADAM proteases during neural crest cell development.

    Science.gov (United States)

    Alfandari, Dominique; Taneyhill, Lisa A

    2018-02-24

    ADAM metalloproteases have been shown to play critical roles during development. In this review, we will describe functional evidence that implicates ADAM proteins during the genesis, migration and differentiation of neural crest cells. We will restrict our analysis to the transmembrane ADAMs as other reviews have addressed the role of extracellular metalloproteases (Christian et al. [2013] Critical Reviews in Biochemistry and Molecular Biology 48:544-560). This review will describe advances that have been obtained mainly through the use of two vertebrate model systems, the frog, and avian embryos. The role of the principal substrates of ADAMs, the cadherins, has been extensively described in other reviews, most recently in (Cousin [1997] Mechanisms of Development 148:79-88; Taneyhill and Schiffmacher [2017] Genesis, 55). The function of ADAMs in the migration of other cell types, including the immune system, wound healing and cancer has been described previously in (Dreymueller et al. [2017] Mediators of Inflammation 2017: 9621724). Our goal is to illustrate both the importance of ADAMs in controlling neural crest behavior and how neural crest cells have helped us understand the molecular interactions, substrates, and functions of ADAM proteins in vivo. © 2018 Wiley Periodicals, Inc.

  4. Studies on radiosensitization of Escherichia coli cells by cis-platinum complexes

    International Nuclear Information System (INIS)

    Zimbrick, J.D.; Sukrochana, A.; Richmond, R.C.

    1979-01-01

    We recently reported that the antitumor drug cis-Pt(NH 3 ) 2 Cl 2 (cis-DDP) produces significant radiosensitization of anoxic E coli C cells. We have extended these studies to three other platinum drugs, all of which have been shown to be more effective antitumor drugs than cis-DDP. The drugs are: cis-dichloro bis(ethylene imine) Pt(II) (cis-DEP); cis-dichlorobicyclopentylamine Pt(II) (cis-PAD); and Pt-thymine blue (cis-PTB). Survival curve studies indicate that these drugs all produce greater anoxic radiosensitization of E coli C than cis-DDP at concentrations which are less toxic to the cells than similar concentrations of cis-DDP. If the cells are treated with any one of these drugs for two hours and then washed to remove the drug before irradiation, no detectable radiosensitization is found. We conclude that these drugs have the potential for being useful agents in combined modality therapy and that they warrant further study in mammalian systems

  5. Cell cycle regulation of the BRCA1/acetyl-CoA-carboxylase complex.

    Science.gov (United States)

    Ray, H; Suau, F; Vincent, A; Dalla Venezia, N

    2009-01-16

    Germ-line alterations in BRCA1 are associated with an increased susceptibility to breast and ovarian cancer. The BRCA1 protein has been implicated in multiple cellular functions. We have recently demonstrated that BRCA1 reduces acetyl-CoA-carboxylase alpha (ACCA) activity through its phospho-dependent binding to ACCA, and further established that the phosphorylation of the Ser1263 of ACCA is required for this interaction. Here, to gain more insight into the cellular conditions that trigger the BRCA1/ACCA interaction, we designed an anti-pSer1263 antibody and demonstrated that the Ser1263 of ACCA is phosphorylated in vivo, in a cell cycle-dependent manner. We further showed that the interaction between BRCA1 and ACCA is regulated during cell cycle progression. Taken together, our findings reveal a novel mechanism of regulation of ACCA distinct from the previously described phosphorylation of Ser79, and provide new insights into the control of lipogenesis through the cell cycle.

  6. Machine learning-based screening of complex molecules for polymer solar cells

    Science.gov (United States)

    Jørgensen, Peter Bjørn; Mesta, Murat; Shil, Suranjan; García Lastra, Juan Maria; Jacobsen, Karsten Wedel; Thygesen, Kristian Sommer; Schmidt, Mikkel N.

    2018-06-01

    Polymer solar cells admit numerous potential advantages including low energy payback time and scalable high-speed manufacturing, but the power conversion efficiency is currently lower than for their inorganic counterparts. In a Phenyl-C_61-Butyric-Acid-Methyl-Ester (PCBM)-based blended polymer solar cell, the optical gap of the polymer and the energetic alignment of the lowest unoccupied molecular orbital (LUMO) of the polymer and the PCBM are crucial for the device efficiency. Searching for new and better materials for polymer solar cells is a computationally costly affair using density functional theory (DFT) calculations. In this work, we propose a screening procedure using a simple string representation for a promising class of donor-acceptor polymers in conjunction with a grammar variational autoencoder. The model is trained on a dataset of 3989 monomers obtained from DFT calculations and is able to predict LUMO and the lowest optical transition energy for unseen molecules with mean absolute errors of 43 and 74 meV, respectively, without knowledge of the atomic positions. We demonstrate the merit of the model for generating new molecules with the desired LUMO and optical gap energies which increases the chance of finding suitable polymers by more than a factor of five in comparison to the randomised search used in gathering the training set.

  7. Max-min SINR low complexity transceiver design for single cell massive MIMO

    KAUST Repository

    Sifaou, Houssem

    2016-08-11

    This work focuses on large scale multi-user MIMO systems in which the base station (BS) outfitted with M antennas communicates with K single antenna user equipments (UEs). In particular, we aim at designing the linear precoder and receiver that maximizes the minimum signal-to-interference-plus-noise ratio (SINR) subject to a given power constraint. To gain insights into the structure of the optimal precoder and receiver as well as to reduce the computational complexity for their implementation, we analyze the asymptotic regime where M and K grow large with a given ratio and make use of random matrix theory (RMT) tools to compute accurate approximations. Although simpler, the implementation of the asymptotic precoder and receiver requires fast inversions of large matrices in every coherence period. To overcome this issue, we apply the truncated polynomial expansion (TPE) technique to the precoding and receiving vector of each UE and make use of RMT to determine the optimal weighting coefficients that asymptotically solve the max-min SINR problem. Numerical results are used to show that the proposed TPE-based precoder and receiver almost achieve the same performance as the optimal ones while requiring a lower complexity.

  8. Initiation of poliovirus plus-strand RNA synthesis in a membrane complex of infected HeLa cells

    International Nuclear Information System (INIS)

    Takeda, N.; Kuhn, R.J.; Yang, C.F.; Takegami, T.; Wimmer, E.

    1986-01-01

    An in vitro poliovirus RNA-synthesizing system derived from a crude membrance fraction of infected HeLa cells was used to analyze the mechanism of initiation of poliovirus plus-strand RNA synthesis. This system contains an activity that synthesizes the nucleotidyl proteins VPg-pU and VPg-pUpU. These molecules represent the 5'-terminal structure of nascent RNA molecules and of virion RNA. The membranous replication complex is also capable of synthesizing mucleotidyl proteins containing nine or more of the poliovirus 5'-proximal nucleotides as assayed by the formation of the RNase T 1 -resistant oligonucleotide VPg-pUUAAAACAGp or by fingerprint analysis of the in vitro-synthesized 32 P-RNA. Incubation of preformed VPg-pUpU with unlabeled nucleoside triphosphates resulted in the formation of VPg-pUUAAAACAGp. This reaction, which appeared to be an elongation of VPg-pUpU, was stimulated by the addition of a soluble fraction (S-10) obtained from uninfected HeLa cells. Preformed VPg-pU could be chased into VPg-pUpU in the presence of UTP. The data are consistent with a model that VPg-pU can function as a primer for poliovirus plus-strand RNA synthesis in the membranous replication complex and that the elongation reaction may be stimulated by a host cellular factor

  9. The influence of venous blood flow on the retinal ganglion cell complex in patients with primary open angle glaucoma

    Directory of Open Access Journals (Sweden)

    N. I. Kurysheva

    2014-07-01

    Full Text Available Purpose: To study the influence of venous blood flow on the ganglion cell complex (GCC in patients with preperimetric and perimetric open angle glaucoma.Methods: 74 patients were included in the research. 59 eyes and 62 eyes were diagnosed with preperimetric and perimetric open angle glaucoma respectively. The mean age was 56.5±10.5 years. 22 (12 female and 10 male healthy individuals constituted the control group. The ganglion cell complex and retinal nerve fibre layer were evaluated with the help of optical coherence tomography (RTVue-100 OCT, Optovue, Inc., Fremont, CA. Ocular blood flow was measured by Color Doppler Imaging (multifunctional VOLUSON 730 ProSystem. The statistical analysis included correlation between GCC and RNFL thickness in both glaucoma groups.Results: The results showed a statistically significant reduction of venous blood flow velocity in both glaucoma groups compared to the control group. No difference in venous blood flow parameters between two glaucoma groups was found, except resistance index, which was higher in perimetric group in comparison to preperimetric group. A correlation was also obtained between venous blood flow parameters and GCC and RNFL thickness in both glaucoma groups.Conclusion: Early GCC damage in glaucoma might occur due to venous blood flow reduction. This fact may be of great value in understanding glaucoma pathogenesis and search for novel treatment options.

  10. Bimolecular Complementation to Visualize Filovirus VP40-Host Complexes in Live Mammalian Cells: Toward the Identification of Budding Inhibitors

    Directory of Open Access Journals (Sweden)

    Yuliang Liu

    2011-01-01

    Full Text Available Virus-host interactions play key roles in promoting efficient egress of many RNA viruses, including Ebola virus (EBOV or “e” and Marburg virus (MARV or “m”. Late- (L- domains conserved in viral matrix proteins recruit specific host proteins, such as Tsg101 and Nedd4, to facilitate the budding process. These interactions serve as attractive targets for the development of broad-spectrum budding inhibitors. A major gap still exists in our understanding of the mechanism of filovirus budding due to the difficulty in detecting virus-host complexes and mapping their trafficking patterns in the natural environment of the cell. To address this gap, we used a bimolecular complementation (BiMC approach to detect, localize, and follow the trafficking patterns of eVP40-Tsg101 complexes in live mammalian cells. In addition, we used the BiMC approach along with a VLP budding assay to test small molecule inhibitors identified by in silico screening for their ability to block eVP40 PTAP-mediated interactions with Tsg101 and subsequent budding of eVP40 VLPs. We demonstrated the potential broad spectrum activity of a lead candidate inhibitor by demonstrating its ability to block PTAP-dependent binding of HIV-1 Gag to Tsg101 and subsequent egress of HIV-1 Gag VLPs.

  11. Design of magnetic gene complexes as effective and serum resistant gene delivery systems for mesenchymal stem cells.

    Science.gov (United States)

    Zhang, Tian-Yuan; Wu, Jia-He; Xu, Qian-Hao; Wang, Xia-Rong; Lu, Jingxiong; Hu, Ying; Jo, Jun-Ichiro; Yamamoto, Masaya; Ling, Daishun; Tabata, Yasuhiko; Gao, Jian-Qing

    2017-03-30

    Gene engineered mesenchymal stem cells (MSCs) have been proposed as promising tools for their various applications in biomedicine. Nevertheless, the lack of an effective and safe way to genetically modify these stem cells is still a major obstacle in the current studies. Herein, we designed novel magnetic complexes by assembling cationized pullulan derivatives with magnetic iron oxide nanoparticles for delivering target genes to MSCs. Results showed that this complexes achieved effective gene expression with the assistance of external magnetic field, and resisted the adverse effect induced by serum proteins on the gene delivery. Moreover, neither significant cytotoxicity nor the interference on the osteogenic differentiation to MSCs were observed after magnetofection. Further studies revealed that this effective and serum resistant gene transfection was partly due to the accelerated and enhanced intracellular uptake process driven by external magnetic field. To conclude, the current study presented a novel option for genetic modification of MSCs in an effective, relatively safe and serum compatible way. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. A Rhodium(III) Complex as an Inhibitor of Neural Precursor Cell Expressed, Developmentally Down-Regulated 8-Activating Enzyme with in Vivo Activity against Inflammatory Bowel Disease.

    Science.gov (United States)

    Zhong, Hai-Jing; Wang, Wanhe; Kang, Tian-Shu; Yan, Hui; Yang, Yali; Xu, Lipeng; Wang, Yuqiang; Ma, Dik-Lung; Leung, Chung-Hang

    2017-01-12

    We report herein the identification of the rhodium(III) complex [Rh(phq) 2 (MOPIP)] + (1) as a potent and selective ATP-competitive neural precursor cell expressed, developmentally down-regulated 8 (NEDD8)-activating enzyme (NAE) inhibitor. Structure-activity relationship analysis indicated that the overall organometallic design of complex 1 was important for anti-inflammatory activity. Complex 1 showed promising anti-inflammatory activity in vivo for the potential treatment of inflammatory bowel disease.

  13. Three-dimensional models of cancer for pharmacology and cancer cell biology: capturing tumor complexity in vitro/ex vivo.

    Science.gov (United States)

    Hickman, John A; Graeser, Ralph; de Hoogt, Ronald; Vidic, Suzana; Brito, Catarina; Gutekunst, Matthias; van der Kuip, Heiko

    2014-09-01

    Cancers are complex and heterogeneous pathological "organs" in a dynamic interplay with their host. Models of human cancer in vitro, used in cancer biology and drug discovery, are generally highly reductionist. These cancer models do not incorporate complexity or heterogeneity. This raises the question as to whether the cancer models' biochemical circuitry (not their genome) represents, with sufficient fidelity, a tumor in situ. Around 95% of new anticancer drugs eventually fail in clinical trial, despite robust indications of activity in existing in vitro pre-clinical models. Innovative models are required that better capture tumor biology. An important feature of all tissues, and tumors, is that cells grow in three dimensions. Advances in generating and characterizing simple and complex (with added stromal components) three-dimensional in vitro models (3D models) are reviewed in this article. The application of stirred bioreactors to permit both scale-up/scale-down of these cancer models and, importantly, methods to permit controlled changes in environment (pH, nutrients, and oxygen) are also described. The challenges of generating thin tumor slices, their utility, and potential advantages and disadvantages are discussed. These in vitro/ex vivo models represent a distinct move to capture the realities of tumor biology in situ, but significant characterization work still remains to be done in order to show that their biochemical circuitry accurately reflects that of a tumor. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Hard wiring of T cell receptor specificity for the major histocompatibility complex is underpinned by TCR adaptability

    Energy Technology Data Exchange (ETDEWEB)

    Burrows, Scott R.; Chen, Zhenjun; Archbold, Julia K.; Tynan, Fleur E.; Beddoe, Travis; Kjer-Nielsen, Lars; Miles, John J.; Khanna, Rajiv; Moss, Denis J.; Liu, Yu Chih; Gras, Stephanie; Kostenko, Lyudmila; Brennan, Rebekah M.; Clements, Craig S.; Brooks, Andrew G.; Purcell, Anthony W.; McCluskey, James; Rossjohn, Jamie (Monash); (Queensland Inst. of Med. Rsrch.); (Melbourne)

    2010-07-07

    {alpha}{beta} T cell receptors (TCRs) are genetically restricted to corecognize peptide antigens bound to self-major histocompatibility complex (pMHC) molecules; however, the basis for this MHC specificity remains unclear. Despite the current dogma, evaluation of the TCR-pMHC-I structural database shows that the nongermline-encoded complementarity-determining region (CDR)-3 loops often contact the MHC-I, and the germline-encoded CDR1 and -2 loops frequently participate in peptide-mediated interactions. Nevertheless, different TCRs adopt a roughly conserved docking mode over the pMHC-I, in which three MHC-I residues (65, 69, and 155) are invariably contacted by the TCR in one way or another. Nonetheless, the impact of mutations at these three positions, either individually or together, was not uniformly detrimental to TCR recognition of pHLA-B*0801 or pHLA-B*3508. Moreover, when TCR-pMHC-I recognition was impaired, this could be partially restored by expression of the CD8 coreceptor. The structure of a TCR-pMHC-I complex in which these three (65, 69, and 155) MHC-I positions were all mutated resulted in shifting of the TCR footprint relative to the cognate complex and formation of compensatory interactions. Collectively, our findings reveal the inherent adaptability of the TCR in maintaining peptide recognition while accommodating changes to the central docking site on the pMHC-I.

  15. Positioning cell wall synthetic complexes by the bacterial morphogenetic proteins MreB and MreD.

    Science.gov (United States)

    White, Courtney L; Kitich, Aleksandar; Gober, James W

    2010-05-01

    In Caulobacter crescentus, intact cables of the actin homologue, MreB, are required for the proper spatial positioning of MurG which catalyses the final step in peptidoglycan precursor synthesis. Similarly, in the periplasm, MreC controls the spatial orientation of the penicillin binding proteins and a lytic transglycosylase. We have now found that MreB cables are required for the organization of several other cytosolic murein biosynthetic enzymes such as MraY, MurB, MurC, MurE and MurF. We also show these proteins adopt a subcellular pattern of localization comparable to MurG, suggesting the existence of cytoskeletal-dependent interactions. Through extensive two-hybrid analyses, we have now generated a comprehensive interaction map of components of the bacterial morphogenetic complex. In the cytosol, this complex contains both murein biosynthetic enzymes and morphogenetic proteins, including RodA, RodZ and MreD. We show that the integral membrane protein, MreD, is essential for lateral peptidoglycan synthesis, interacts with the precursor synthesizing enzymes MurG and MraY, and additionally, determines MreB localization. Our results suggest that the interdependent localization of MreB and MreD functions to spatially organize a complex of peptidoglycan precursor synthesis proteins, which is required for propagation of a uniform cell shape and catalytically efficient peptidoglycan synthesis.

  16. Haplotyping the human T-cell receptor β-chain gene complex by use of restriction fragment length polymorphisms

    International Nuclear Information System (INIS)

    Charmley, P.; Chao, A.; Gatti, R.A.; Concannon, P.; Hood, L.

    1990-01-01

    The authors have studied the genetic segregation of human T-cell receptor β-chain (TCRβ) genes on chromosome 7q in 40 CEPH (Centre d'Etude du Polymorphisme Humain) families by using restriction fragment length polymorphisms (RFLPs). They constructed haplotypes from eight RFLPs by using variable- and constant-region cDNA probes, which detect polymorphisms that span more than 600 kilobases of the TCRβ gene complex. Analysis of allele distributions between TCRβ genes revealed significant linkage disequilibrium between only 6 of the 28 different pairs of RFLPs. This linkage disequilibrium strongly influences the most efficient order to proceed for typing of these RFLPs in order to achieve maximum genetic informativeness, which in this study revealed a 97.3% level of heterozygosity within the TCRβ gene complex. The results should provide new insight into recent reports of disease associations with the TCRβ gene complex and should assist in designing future experiments to detect or confirm the existence of disease-susceptibility loci in this region of the human genome

  17. Redox reactions of the α-synuclein-Cu(2+) complex and their effects on neuronal cell viability.

    Science.gov (United States)

    Wang, Chengshan; Liu, Lin; Zhang, Lin; Peng, Yong; Zhou, Feimeng

    2010-09-21

    α-Synuclein (α-syn), a presynaptic protein believed to play an important role in neuropathology in Parkinson's disease (PD), is known to bind Cu(2+). Cu(2+) has been shown to accelerate the aggregation of α-syn to form various toxic aggregates in vitro. Copper is also a redox-active metal whose complexes with amyloidogenic proteins/peptides have been linked to oxidative stress in major neurodegenerative diseases. In this work, the formation of the Cu(2+) complex with α-syn or with an N-terminal peptide, α-syn(1-19), was confirmed with electrospray-mass spectrometry (ES-MS). The redox potentials of the Cu(2+) complex with α-syn (α-syn-Cu(2+)) and α-syn(1-19) were determined to be 0.018 and 0.053 V, respectively. Furthermore, the Cu(2+) center(s) can be readily reduced to Cu(+), and possible reactions of α-syn-Cu(2+) with cellular species (e.g., O(2), ascorbic acid, and dopamine) were investigated. The occurrence of a redox reaction can be rationalized by comparing the redox potential of the α-syn-Cu(2+) complex to that of the specific cellular species. For example, ascorbic acid can directly reduce α-syn-Cu(2+) to α-syn-Cu(+), setting up a redox cycle in which O(2) is reduced to H(2)O(2) and cellular redox species is continuously exhausted. In addition, the H(2)O(2) generated was demonstrated to reduce viability of the neuroblastoma SY-HY5Y cells. Although our results ruled out the direct oxidation of dopamine by α-syn-Cu(2+), the H(2)O(2) generated in the presence of α-syn-Cu(2+) can oxidize dopamine. Our results suggest that oxidative stress is at least partially responsible for the loss of dopaminergic cells in PD brain and reveal the multifaceted role of the α-syn-Cu(2+) complex in oxidative stress associated with PD symptoms.

  18. Aberrant ERK 1/2 complex activation and localization in scrapie-infected GT1-1 cells

    Directory of Open Access Journals (Sweden)

    Didonna Alessandro

    2010-08-01

    Full Text Available Abstract Background Fatal neurodegenerative disorders such as Creutzfeldt-Jakob and Gerstmann-Sträussler-Scheinker diseases in humans, scrapie and bovine spongiform encephalopathy in animals, are characterized by the accumulation in the brain of a pathological form of the prion protein (PrP denominated PrPSc. The latter derives from the host cellular form, PrPC, through a process whereby portions of its α-helical and coil structures are refolded into β-sheet structures. Results In this work, the widely known in vitro model of prion replication, hypothalamic GT1-1 cell line, was used to investigate cellular and molecular responses to prion infection. The MAP kinase cascade was dissected to assess the phosphorylation levels of src, MEK 1/2 and ERK 1/2 signaling molecules, both before and after prion infection. Our findings suggest that prion replication leads to a hyper-activation of this pathway. Biochemical analysis was complemented with immunofluorescence studies to map the localization of the ERK complex within the different cellular compartments. We showed how the ERK complex relocates in the cytosol upon prion infection. We correlated these findings with an impairment of cell growth in prion-infected GT1-1 cells as probed by MTT assay. Furthermore, given the persistent urgency in finding compounds able to cure prion infected cells, we tested the effects on the ERK cascade of two molecules known to block prion replication in vitro, quinacrine and Fab D18. We were able to show that while these two compounds possess similar effects in curing prion infection, they affect the MAP kinase cascade differently. Conclusions Taken together, our results help shed light on the molecular events involved in neurodegeneration and neuronal loss in prion infection and replication. In particular, the combination of chronic activation and aberrant localization of the ERK complex may lead to a lack of essential neuroprotective and survival factors

  19. The dynamic behavior of thin-film ionic transition metal complex-based light-emitting electrochemical cells

    Energy Technology Data Exchange (ETDEWEB)

    Meier, Sebastian B., E-mail: sebastian.meier@belectric.com, E-mail: wiebke.sarfert@siemens.com [Department of Materials Science VI: Materials for Electronics and Energy Technology, Friedrich-Alexander-University of Erlangen-Nuremberg, 91058 Erlangen (Germany); Siemens AG, Corporate Technology, CT RTC MAT IEC-DE, 91058 Erlangen (Germany); Hartmann, David; Sarfert, Wiebke, E-mail: sebastian.meier@belectric.com, E-mail: wiebke.sarfert@siemens.com [Siemens AG, Corporate Technology, CT RTC MAT IEC-DE, 91058 Erlangen (Germany); Winnacker, Albrecht [Department of Materials Science VI: Materials for Electronics and Energy Technology, Friedrich-Alexander-University of Erlangen-Nuremberg, 91058 Erlangen (Germany)

    2014-09-14

    Light-emitting electrochemical cells (LECs) have received increasing attention during recent years due to their simple architecture, based on solely air-stabile materials, and ease of manufacture in ambient atmosphere, using solution-based technologies. The LEC's active layer offers semiconducting, luminescent as well as ionic functionality resulting in device physical processes fundamentally different as compared with organic light-emitting diodes. During operation, electrical double layers (EDLs) form at the electrode interfaces as a consequence of ion accumulation and electrochemical doping sets in leading to the in situ development of a light-emitting p-i-n junction. In this paper, we comment on the use of impedance spectroscopy in combination with complex nonlinear squares fitting to derive key information about the latter events in thin-film ionic transition metal complex-based light-emitting electrochemical cells based on the model compound bis-2-phenylpyridine 6-phenyl-2,2´-bipyridine iridium(III) hexafluoridophosphate ([Ir(ppy)₂(pbpy)][PF₆]). At operating voltages below the bandgap potential of the ionic complex used, we obtain the dielectric constant of the active layer, the conductivity of mobile ions, the transference numbers of electrons and ions, and the thickness of the EDLs, whereas the transient thickness of the p-i-n junction is determined at voltages above the bandgap potential. Most importantly, we find that charge transport is dominated by the ions when carrier injection from the electrodes is prohibited, that ion movement is limited by the presence of transverse internal interfaces and that the width of the intrinsic region constitutes almost 60% of the total active layer thickness in steady state at a low operating voltage.

  20. A Potential Bone-Targeting Hypotoxic Platinum(II) Complex with an Unusual Cytostatic Mechanism toward Osteosarcoma Cells.

    Science.gov (United States)

    Zhang, Zhenqin; Zhu, Zhenzhu; Luo, Cheng; Zhu, Chengcheng; Zhang, Changli; Guo, Zijian; Wang, Xiaoyong

    2018-03-19

    Osteosarcoma (OS) is the most common primary pediatric bone tumor lethal to children and adolescents. Chemotherapeutic agents such as cisplatin are not effective for OS because of their poor accessibility to this cancer and severe systemic toxicity. In this study, a lipophilic platinum(II) complex bearing a bisphosphonate bone-targeting moiety, cis-[PtL(NH 3 ) 2 Cl]NO 3 {BPP; L = tetraethyl [2-(pyridin-2-yl)ethane-1,1-diyl]bisphosphonate}, was prepared and characterized by NMR, electrospray ionization mass spectrometry, and single-crystal X-ray crystallography. The cytotoxicity of BPP toward OS cell lines U2OS and MG-63 was tested by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. BPP exhibits moderate inhibition against U2OS cells through a mechanism involving both DNA binding and a mevalonate pathway. The acute toxicity of BPP to mice is 7-fold lower than that of cisplatin. The relative low systemic toxicity may result from the steric hindrance of the ligand, which blocks BPP approaching the bases of DNA. The results suggest that incorporating bisphosphonates into a platinum complex not only enhances its bone-targeting property but also minimizes its reactivity toward DNA and thereby lowers the systematic toxicity of the complex. The diminished cytotoxicity of BPP could be compensated for by increasing the therapeutic dose with marginal harm. This strategy provides a new possibility for overcoming the ineffectiveness and systemic toxicity of platinum drugs in the treatment of OS.

  1. Glycosylinositol phosphorylceramides from Rosa cell cultures are boron-bridged in the plasma membrane and form complexes with rhamnogalacturonan II.

    Science.gov (United States)

    Voxeur, Aline; Fry, Stephen C

    2014-07-01

    Boron (B) is essential for plant cell-wall structure and membrane functions. Compared with its role in cross-linking the pectic domain rhamnogalacturonan II (RG-II), little information is known about the biological role of B in membranes. Here, we investigated the involvement of glycosylinositol phosphorylceramides (GIPCs), major components of lipid rafts, in the membrane requirement for B. Using thin-layer chromatography and mass spectrometry, we first characterized GIPCs from Rosa cell culture. The major GIPC has one hexose residue, one hexuronic acid residue, inositol phosphate, and a ceramide moiety with a C18 trihydroxylated mono-unsaturated long-chain base and a C24 monohydroxylated saturated fatty acid. Disrupting B bridging (by B starvation in vivo or by treatment with cold dilute HCl or with excess borate in vitro) enhanced the GIPCs' extractability. As RG-II is the main B-binding site in plants, we investigated whether it could form a B-centred complex with GIPCs. Using high-voltage paper electrophoresis, we showed that addition of GIPCs decreased the electrophoretic mobility of radiolabelled RG-II, suggesting formation of a GIPC-B-RG-II complex. Last, using polyacrylamide gel electrophoresis, we showed that added GIPCs facilitate RG-II dimerization in vitro. We conclude that B plays a structural role in the plasma membrane. The disruption of membrane components by high borate may account for the phytotoxicity of excess B. Moreover, the in-vitro formation of a GIPC-B-RG-II complex gives the first molecular explanation of the wall-membrane attachment sites observed in vivo. Finally, our results suggest a role for GIPCs in the RG-II dimerization process. © 2014 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

  2. Genetic complexity of miscanthus cell wall composition and biomass quality for biofuels.

    Science.gov (United States)

    van der Weijde, Tim; Kamei, Claire L Alvim; Severing, Edouard I; Torres, Andres F; Gomez, Leonardo D; Dolstra, Oene; Maliepaard, Chris A; McQueen-Mason, Simon J; Visser, Richard G F; Trindade, Luisa M

    2017-05-25

    Miscanthus sinensis is a high yielding perennial grass species with great potential as a bioenergy feedstock. One of the challenges that currently impedes commercial cellulosic biofuel production is the technical difficulty to efficiently convert lignocellulosic biomass into biofuel. The development of feedstocks with better biomass quality will improve conversion efficiency and the sustainability of the value-chain. Progress in the genetic improvement of biomass quality may be substantially expedited by the development of genetic markers associated to quality traits, which can be used in a marker-assisted selection program. To this end, a mapping population was developed by crossing two parents of contrasting cell wall composition. The performance of 182 F1 offspring individuals along with the parents was evaluated in a field trial with a randomized block design with three replicates. Plants were phenotyped for cell wall composition and conversion efficiency characters in the second and third growth season after establishment. A new SNP-based genetic map for M. sinensis was built using a genotyping-by-sequencing (GBS) approach, which resulted in 464 short-sequence uniparental markers that formed 16 linkage groups in the male map and 17 linkage groups in the female map. A total of 86 QTLs for a variety of biomass quality characteristics were identified, 20 of which were detected in both growth seasons. Twenty QTLs were directly associated to different conversion efficiency characters. Marker sequences were aligned to the sorghum reference genome to facilitate cross-species comparisons. Analyses revealed that for some traits previously identified QTLs in sorghum occurred in homologous regions on the same chromosome. In this work we report for the first time the genetic mapping of cell wall composition and bioconversion traits in the bioenergy crop miscanthus. These results are a first step towards the development of marker-assisted selection programs in miscanthus

  3. Fetal colon cell line FHC exhibits tumorigenic phenotype, complex karyotype, and TP53 gene mutation

    Czech Academy of Sciences Publication Activity Database

    Souček, Karel; Jirsová, Pavla; Brázdová, Marie; Hýžďalová, Martina; Kočí, Lenka; Vydra, D.; Trojanec, R.; Pernicová, Zuzana; Lentvorská, L.; Hajdúch, M.; Hofmanová, Jiřina; Kozubík, Alois

    2010-01-01

    Roč. 197, č. 2 (2010), s. 107-116 ISSN 0165-4608 R&D Projects: GA MŠk ME 919; GA ČR(CZ) GA204/07/0834; GA ČR(CZ) GA204/08/1560; GA AV ČR(CZ) 1QS500040507; GA MZd NS9600 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : colon epithelial cells * TP53 * MYC Subject RIV: BO - Biophysics Impact factor: 1.551, year: 2010

  4. Charge transfer complex states in diketopyrrolopyrrole polymers and fullerene blends: Implications for organic solar cell efficiency

    Science.gov (United States)

    Moghe, D.; Yu, P.; Kanimozhi, C.; Patil, S.; Guha, S.

    2011-12-01

    The spectral photocurrent characteristics of two donor-acceptor diketopyrrolopyrrole (DPP)-based copolymers (PDPP-BBT and TDPP-BBT) blended with a fullerene derivative [6,6]-phenyl C61-butyric acid methyl ester (PCBM) were studied using Fourier-transform photocurrent spectroscopy (FTPS) and monochromatic photocurrent (PC) method. PDPP-BBT:PCBM shows the onset of the lowest charge transfer complex (CTC) state at 1.42 eV, whereas TDPP-BBT:PCBM shows no evidence of the formation of a midgap CTC state. The FTPS and PC spectra of P3HT:PCBM are also compared. The larger singlet state energy difference of TDPP-BBT and PCBM compared to PDPP-BBT/P3HT and PCBM obliterates the formation of a midgap CTC state resulting in an enhanced photovoltaic efficiency over PDPP-BBT:PCBM.

  5. Nuclear pore complex protein mediated nuclear localization of dicer protein in human cells.

    Directory of Open Access Journals (Sweden)

    Yoshinari Ando

    Full Text Available Human DICER1 protein cleaves double-stranded RNA into small sizes, a crucial step in production of single-stranded RNAs which are mediating factors of cytoplasmic RNA interference. Here, we clearly demonstrate that human DICER1 protein localizes not only to the cytoplasm but also to the nucleoplasm. We also find that human DICER1 protein associates with the NUP153 protein, one component of the nuclear pore complex. This association is detected predominantly in the cytoplasm but is also clearly distinguishable at the nuclear periphery. Additional characterization of the NUP153-DICER1 association suggests NUP153 plays a crucial role in the nuclear localization of the DICER1 protein.

  6. Computational Molecular Nanoscience Study of the Properties of Copper Complexes for Dye-Sensitized Solar Cells

    Science.gov (United States)

    Baldenebro-López, Jesús; Castorena-González, José; Flores-Holguín, Norma; Almaral-Sánchez, Jorge; Glossman-Mitnik, Daniel

    2012-01-01

    In this work, we studied a copper complex-based dye, which is proposed for potential photovoltaic applications and is named Cu (I) biquinoline dye. Results of electron affinities and ionization potentials have been used for the correlation between different levels of calculation used in this study, which are based on The Density Functional Theory (DFT) and time-dependent (TD) DFT. Further, the maximum absorption wavelengths of our theoretical calculations were compared with the experimental data. It was found that the M06/LANL2DZ + DZVP level of calculation provides the best approximation. This level of calculation was used to find the optimized molecular structure and to predict the main molecular vibrations, the molecular orbitals energies, dipole moment, isotropic polarizability and the chemical reactivity parameters that arise from Conceptual DFT. PMID:23443107

  7. Heterogeneous nuclear ribonucleoprotein K upregulates the kinetochore complex component NUF2 and promotes the tumorigenicity of colon cancer cells

    International Nuclear Information System (INIS)

    Sugimasa, Hironobu; Taniue, Kenzui; Kurimoto, Akiko; Takeda, Yasuko; Kawasaki, Yoshihiro; Akiyama, Tetsu

    2015-01-01

    Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multi-functional protein involved in transcription, mRNA splicing, mRNA stabilization and translation. Although hnRNP K has been suggested to play a role in the development of many cancers, its molecular function in colorectal cancer has remained elusive. Here we show that hnRNP K plays an important role in the mitotic process in HCT116 colon cancer cells. Furthermore, we demonstrate that hnRNP K directly transactivates the NUF2 gene, the product of which is a component of the NDC80 kinetochore complex and which is known to be critical for a stable spindle microtubule-kinetochore attachment. In addition, knockdown of both hnRNP K and NUF2 caused failure in metaphase chromosome alignment and drastic decrease in the growth of colon cancer cells. These results suggest that the hnRNP K-NUF2 axis is important for the mitotic process and proliferation of colon cancer cells and that this axis could be a target for the therapy of colon cancer. - Highlights: • hnRNP K is required for the tumorigenicity of colon cancer cells. • hnRNP K binds to the promoter region of NUF2 and activates its transcription. • NUF2 expression is correlated with hnRNP K expression in colorectal cancer tissue. • hnRNP K and NUF2 are required for metaphase chromosome alignment. • The hnRNP K-NUF2 axis is important for the proliferation of colon cancer cells

  8. Heterogeneous nuclear ribonucleoprotein K upregulates the kinetochore complex component NUF2 and promotes the tumorigenicity of colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Sugimasa, Hironobu; Taniue, Kenzui [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Kurimoto, Akiko [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Oncology Research Laboratories, Daiichi Sankyo Co., Ltd, 1-2-58, Hiromachi, Shinagawa-ku, Tokyo, 140-8710 (Japan); Takeda, Yasuko; Kawasaki, Yoshihiro [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Akiyama, Tetsu, E-mail: akiyama@iam.u-tokyo.ac.jp [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan)

    2015-03-27

    Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multi-functional protein involved in transcription, mRNA splicing, mRNA stabilization and translation. Although hnRNP K has been suggested to play a role in the development of many cancers, its molecular function in colorectal cancer has remained elusive. Here we show that hnRNP K plays an important role in the mitotic process in HCT116 colon cancer cells. Furthermore, we demonstrate that hnRNP K directly transactivates the NUF2 gene, the product of which is a component of the NDC80 kinetochore complex and which is known to be critical for a stable spindle microtubule-kinetochore attachment. In addition, knockdown of both hnRNP K and NUF2 caused failure in metaphase chromosome alignment and drastic decrease in the growth of colon cancer cells. These results suggest that the hnRNP K-NUF2 axis is important for the mitotic process and proliferation of colon cancer cells and that this axis could be a target for the therapy of colon cancer. - Highlights: • hnRNP K is required for the tumorigenicity of colon cancer cells. • hnRNP K binds to the promoter region of NUF2 and activates its transcription. • NUF2 expression is correlated with hnRNP K expression in colorectal cancer tissue. • hnRNP K and NUF2 are required for metaphase chromosome alignment. • The hnRNP K-NUF2 axis is important for the proliferation of colon cancer cells.

  9. Follicular dermal papilla structures by organization of epithelial and mesenchymal cells in interfacial polyelectrolyte complex fibers.

    Science.gov (United States)

    Lim, Tze Chiun; Leong, Meng Fatt; Lu, Hongfang; Du, Chan; Gao, Shujun; Wan, Andrew C A; Ying, Jackie Y

    2013-09-01

    The hair follicle is a regenerating organ that produces a new hair shaft during each growth cycle. Development and cycling of the hair follicle is governed by interactions between the epithelial and mesenchymal components. Therefore, development of an engineered 3D hair follicle would be useful for studying these interactions to identify strategies for treatment of hair loss. We have developed a technique suitable for assembly of different cell types in close proximity in fibrous hydrogel scaffolds with resolutions of ∼50 μm. By assembly of dermal papilla (DP) and keratinocytes, structures similar to the native hair bulb arrangement are formed. Gene expression of these constructs showed up-regulation of molecules involved in epithelial-mesenchymal interactions of the hair follicle. Implantation of the follicular structures in SCID mice led to the formation of hair follicle-like structures, thus demonstrating their hair inductive ability. The transparency of the fiber matrix and the small dimensions of the follicular structures allowed the direct quantitation of DP cell proliferation by confocal microscopy, clearly illustrating the promoting or inhibitory effects of hair growth regulating agents. Collectively, our results suggested a promising application of these 3D engineered follicular structures for in vitro screening and testing of drugs for hair growth therapy. Copyright © 2013 Elsevier Ltd. All rights reserved.

  10. A Novel Interaction of Ecdysoneless (ECD) Protein with R2TP Complex Component RUVBL1 Is Required for the Functional Role of ECD in Cell Cycle Progression.

    Science.gov (United States)

    Mir, Riyaz A; Bele, Aditya; Mirza, Sameer; Srivastava, Shashank; Olou, Appolinaire A; Ammons, Shalis A; Kim, Jun Hyun; Gurumurthy, Channabasavaiah B; Qiu, Fang; Band, Hamid; Band, Vimla

    2015-12-28

    Ecdysoneless (ECD) is an evolutionarily conserved protein whose germ line deletion is embryonic lethal. Deletion of Ecd in cells causes cell cycle arrest, which is rescued by exogenous ECD, demonstrating a requirement of ECD for normal mammalian cell cycle progression. However, the exact mechanism by which ECD regulates cell cycle is unknown. Here, we demonstrate that ECD protein levels and subcellular localization are invariant during cell cycle progression, suggesting a potential role of posttranslational modifications or protein-protein interactions. Since phosphorylated ECD was recently shown to interact with the PIH1D1 adaptor component of the R2TP cochaperone complex, we examined the requirement of ECD phosphorylation in cell cycle progression. Notably, phosphorylation-deficient ECD mutants that failed to bind to PIH1D1 in vitro fully retained the ability to interact with the R2TP complex and yet exhibited a reduced ability to rescue Ecd-deficient cells from cell cycle arrest. Biochemical analyses demonstrated an additional phosphorylation-independent interaction of ECD with the RUVBL1 component of the R2TP complex, and this interaction is essential for ECD's cell cycle progression function. These studies demonstrate that interaction of ECD with RUVBL1, and its CK2-mediated phosphorylation, independent of its interaction with PIH1D1, are important for its cell cycle regulatory function. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  11. Structure and function of human α-lactalbumin made lethal to tumor cells (HAMLET)-type complexes.

    Science.gov (United States)

    Mossberg, Ann-Kristin; Hun Mok, Kenneth; Morozova-Roche, Ludmilla A; Svanborg, Catharina

    2010-11-01

    Human α-lactalbumin made lethal to tumor cells (HAMLET) and equine lysozyme with oleic acid (ELOA) are complexes consisting of protein and fatty acid that exhibit cytotoxic activities, drastically differing from the activity of their respective proteinaceous compounds. Since the discovery of HAMLET in the 1990s, a wealth of information has been accumulated, illuminating the structural, functional and therapeutic properties of protein complexes with oleic acid, which is summarized in this review. In vitro, both HAMLET and ELOA are produced by using ion-exchange columns preconditioned with oleic acid. However, the complex of human α-lactalbumin with oleic acid with the antitumor activity of HAMLET was found to be naturally present in the acidic fraction of human milk, where it was discovered by serendipity. Structural studies have shown that α-lactalbumin in HAMLET and lysozyme in ELOA are partially unfolded, 'molten-globule'-like, thereby rendering the complexes dynamic and in conformational exchange. HAMLET exists in the monomeric form, whereas ELOA mostly exists as oligomers and the fatty acid stoichiometry varies, with HAMLET holding an average of approximately five oleic acid molecules, whereas ELOA contains a considerably larger number (11- 48). Potent tumoricidal activity is found in both HAMLET and ELOA, and HAMLET has also shown strong potential as an antitumor drug in different in vivo animal models and clinical studies. The gain of new, beneficial function upon partial protein unfolding and fatty acid binding is a remarkable phenomenon, and may reflect a significant generic route of functional diversification of proteins via varying their conformational states and associated ligands. © 2010 The Authors Journal compilation © 2010 FEBS.

  12. Chicken major histocompatibility complex-encoded B-G antigens are found on many cell types that are important for the immune system

    DEFF Research Database (Denmark)

    Salomonsen, J; Dunon, D; Skjødt, K

    1991-01-01

    B-G antigens are a polymorphic multigene family of cell surface molecules encoded by the chicken major histocompatibility complex (MHC). They have previously been described only on cells of the erythroid lineage. By using flow cytometry, section staining, and immunoprecipitation with monoclonal a...

  13. Identification of a rhodium(iii) complex as a Wee1 inhibitor against TP53-mutated triple-negative breast cancer cells.

    Science.gov (United States)

    Yang, Guan-Jun; Zhong, Hai-Jing; Ko, Chung-Nga; Wong, Suk-Yu; Vellaisamy, Kasipandi; Ye, Min; Ma, Dik-Lung; Leung, Chung-Hang

    2018-03-06

    The rhodium(iii) complex 1 was identified as a potent Wee1 inhibitor in vitro and in cellulo. It decreased Wee1 activity and unscheduled mitotic entry, and induced cell damage and death in TP53-mutated triple-negative breast cancer cells. 1 represents a promising scaffold for further development of more potent metal-based Wee1 antagonists.

  14. Obesity-Associated Autoantibody Production Requires AIM to Retain the Immunoglobulin M Immune Complex on Follicular Dendritic Cells

    Directory of Open Access Journals (Sweden)

    Satoko Arai

    2013-04-01

    Full Text Available Natural immunoglobulin M (IgM is reactive to autoantigens and is believed to be important for autoimmunity. Blood pentameric IgM loaded with antigens forms a large immune complex (IC that contains various elements, including apoptosis inhibitor of macrophage (AIM. Here we demonstrate that this IgM-AIM association contributes to autoantibody production under obese conditions. In mice fed a high-fat diet, natural IgM increased through B cell TLR4 stimulation. AIM associated with IgM and protected AIM from renal excretion, increasing blood AIM levels along with the obesity-induced IgM augmentation. Meanwhile, the AIM association inhibited IgM binding to the Fcα/μ receptor on splenic follicular dendritic cells, thereby protecting the IgM IC from Fcα/μ receptor-mediated internalization. This supported IgM-dependent autoantigen presentation to B cells, stimulating IgG autoantibody production. Accordingly, in obese AIM-deficient (AIM−/− mice, the increase of multiple IgG autoantibodies observed in obese wild-type mice was abrogated. Thus, the AIM-IgM association plays a critical role in the obesity-associated autoimmune process.

  15. Dynamic differentiation of GABAA-sensitive influences on orientation selectivity of complex cells in the cat striate cortex.

    Science.gov (United States)

    Pfleger, B; Bonds, A B

    1995-01-01

    The influence of GABAA receptors on orientation selectivity of cat complex cells was tested by iontophoresis of the GABAA receptor blockers bicuculline and N-methyl-bicuculline while stimulating with drifting sinusoidal gratings. Reduction of orientation tuning was markedly less than reported in previous studies that used drifting bars as visual stimuli. Only 3/31 cells lost orientation selectivity, with an average increase in bandwidth of 33%, as opposed to half the cells losing selectivity and a bandwidth increase for the remainder of 47% as reported previously. Infusion of GABAA blockers revealed a prominent stimulus onset transient response, lasting about 120 ms, that showed a broadening of orientation selectivity comparable to that found using drifting bars under similar circumstances. We believe that drifting gratings emphasize a steady-state response component that retains, in the presence of GABAA blockers, significant orientation selectivity. Because the onset transient is initially unselective for orientation, we suggest that the steady-state, orientation-selective response component develops from an alternate inhibitory mechanism, possibly mediated by GABAB receptors.

  16. Effect of the LHCII pigment-protein complex aggregation on photovoltaic properties of sensitized TiO2 solar cells.

    Science.gov (United States)

    Yang, Yiqun; Jankowiak, Ryszard; Lin, Chen; Pawlak, Krzysztof; Reus, Michael; Holzwarth, Alfred R; Li, Jun

    2014-10-14

    A modified dye-sensitized solar cell consisting of a thin TiO2 barrier layer sensitized with natural trimeric light-harvesting complex II (LHCII) from spinach was used as a biomimetic model to study the effects of LHCII aggregation on the photovoltaic properties. The aggregation of individual trimers induced molecular reorganization, which dramatically increased the photocurrent. The morphology of small- and large-size LHCII aggregates deposited on a surface was confirmed by atomic force microscopy. Enhanced LHCII immobilization was accomplished via electrostatic interaction with amine-functionalized photoanodes. The photocurrent responses of the assembled solar cells under illumination at three characteristic wavelength bands in the UV-Vis absorption spectra of LHCII solutions confirmed that a significant photocurrent was generated by LHCII photosensitizers. The enhanced photocurrent by large aggregated LHCII is shown to correlate with the quenching in the far-red fluorescence deriving from chlorophyll-chlorophyll charge transfer states that are effectively coupled with the TiO2 surface and thus inject electrons into the TiO2 conduction band. The large aggregated LHCII with more chlorophyll-chlorophyll charge transfer states is a much better sensitizer since it injects electrons more efficiently into the conduction band of TiO2 than the small aggregated LHCII mostly consisting of unquenched chlorophyll excited state. The assembled solar cells demonstrated remarkable stability in both aqueous buffer and acetonitrile electrolytes over 30 days.

  17. Comparison of complex effluent treatability in different bench scale microbial electrolysis cells

    KAUST Repository

    Ullery, Mark L.

    2014-10-01

    A range of wastewaters and substrates were examined using mini microbial electrolysis cells (mini MECs) to see if they could be used to predict the performance of larger-scale cube MECs. COD removals and coulombic efficiencies corresponded well between the two reactor designs for individual samples, with 66-92% of COD removed for all samples. Current generation was consistent between the reactor types for acetate (AC) and fermentation effluent (FE) samples, but less consistent with industrial (IW) and domestic wastewaters (DW). Hydrogen was recovered from all samples in cube MECs, but gas composition and volume varied significantly between samples. Evidence for direct conversion of substrate to methane was observed with two of the industrial wastewater samples (IW-1 and IW-3). Overall, mini MECs provided organic treatment data that corresponded well with larger scale reactor results, and therefore it was concluded that they can be a useful platform for screening wastewater sources. © 2014 Elsevier Ltd.

  18. Particle-in-cell vs straight line Gaussian calculations for an area of complex topography

    International Nuclear Information System (INIS)

    Lange, R.; Sherman, C.

    1977-01-01

    Two numerical models for the calculation of time integrated air concentraton and ground deposition of airborne radioactive effluent releases are compared. The time dependent Particle-in-Cell (PIC) model and the steady state Gaussian plume model were used for the simulation. The area selected for the comparison was the Hudson River Valley, New York. Input for the models was synthesized from meteorological data gathered in previous studies by various investigators. It was found that the PIC model more closely simulated the three-dimensional effects of the meteorology and topography. Overall, the Gaussian model calculated higher concentrations under stable conditions. In addition, because of its consideration of exposure from the returning plume after flow reversal, the PIC model calculated air concentrations over larger areas than did the Gaussian model

  19. Leaf-cutting ant fungi produce cell wall degrading pectinase complexes reminiscent of phytopathogenic fungi.

    Science.gov (United States)

    Schiøtt, Morten; Rogowska-Wrzesinska, Adelina; Roepstorff, Peter; Boomsma, Jacobus J

    2010-12-31

    Leaf-cutting (attine) ants use their own fecal material to manure fungus gardens, which consist of leaf material overgrown by hyphal threads of the basidiomycete fungus Leucocoprinus gongylophorus that lives in symbiosis with the ants. Previous studies have suggested that the fecal droplets contain proteins that are produced by the fungal symbiont to pass unharmed through the digestive system of the ants, so they can enhance new fungus garden growth. We tested this hypothesis by using proteomics methods to determine the gene sequences of fecal proteins in Acromyrmex echinatior leaf-cutting ants. Seven (21%) of the 33 identified proteins were pectinolytic enzymes that originated from the fungal symbiont and which were still active in the fecal droplets produced by the ants. We show that these enzymes are found in the fecal material only when the ants had access to fungus garden food, and we used quantitative polymerase chain reaction analysis to show that the expression of six of these enzyme genes was substantially upregulated in the fungal gongylidia. These unique structures serve as food for the ants and are produced only by the evolutionarily advanced garden symbionts of higher attine ants, but not by the fungi reared by the basal lineages of this ant clade. Pectinolytic enzymes produced in the gongylidia of the fungal symbiont are ingested but not digested by Acromyrmex leaf-cutting ants so that they end up in the fecal fluid and become mixed with new garden substrate. Substantial quantities of pectinolytic enzymes are typically found in pathogenic fungi that attack live plant tissue, where they are known to breach the cell walls to allow the fungal mycelium access to the cell contents. As the leaf-cutting ant symbionts are derived from fungal clades that decompose dead plant material, our results suggest that their pectinolytic enzymes represent secondarily evolved adaptations that are convergent to those normally found in phytopathogens.

  20. Leaf-cutting ant fungi produce cell wall degrading pectinase complexes reminiscent of phytopathogenic fungi

    Directory of Open Access Journals (Sweden)

    Boomsma Jacobus J

    2010-12-01

    Full Text Available Abstract Background Leaf-cutting (attine ants use their own fecal material to manure fungus gardens, which consist of leaf material overgrown by hyphal threads of the basidiomycete fungus Leucocoprinus gongylophorus that lives in symbiosis with the ants. Previous studies have suggested that the fecal droplets contain proteins that are produced by the fungal symbiont to pass unharmed through the digestive system of the ants, so they can enhance new fungus garden growth. Results We tested this hypothesis by using proteomics methods to determine the gene sequences of fecal proteins in Acromyrmex echinatior leaf-cutting ants. Seven (21% of the 33 identified proteins were pectinolytic enzymes that originated from the fungal symbiont and which were still active in the fecal droplets produced by the ants. We show that these enzymes are found in the fecal material only when the ants had access to fungus garden food, and we used quantitative polymerase chain reaction analysis to show that the expression of six of these enzyme genes was substantially upregulated in the fungal gongylidia. These unique structures serve as food for the ants and are produced only by the evolutionarily advanced garden symbionts of higher attine ants, but not by the fungi reared by the basal lineages of this ant clade. Conclusions Pectinolytic enzymes produced in the gongylidia of the fungal symbiont are ingested but not digested by Acromyrmex leaf-cutting ants so that they end up in the fecal fluid and become mixed with new garden substrate. Substantial quantities of pectinolytic enzymes are typically found in pathogenic fungi that attack live plant tissue, where they are known to breach the cell walls to allow the fungal mycelium access to the cell contents. As the leaf-cutting ant symbionts are derived from fungal clades that decompose dead plant material, our results suggest that their pectinolytic enzymes represent secondarily evolved adaptations that are convergent to

  1. Complex Epigenetic Regulation of Chemotherapy Resistance and Biology in Esophageal Squamous Cell Carcinoma via MicroRNAs

    Directory of Open Access Journals (Sweden)

    Kirsten Lindner

    2018-02-01

    Full Text Available Background: Resistance towards chemotherapy is a major obstacle in the treatment of esophageal squamous cell carcinoma (ESCC. We investigated the role of specific microRNAs in chemotherapy resistance and tumor biology. Methods: We selected three microRNAs from characteristic microRNA signatures of resistant ESCC (hsa-miR-125a-5p, hsa-miR-130a-3p, hsa-miR-1226-3p, and hsa-miR-148a-3p. Effects on chemotherapy, adhesion, migration, apoptosis and cell cycle were assessed in six ESCC cell lines. Target analyses were performed using Western blotting and luciferase techniques. Results: MiR-130a-3p sensitized cells towards cisplatin in 100% of cell lines, miR-148a-3p in 83%, miR-125a-5p in 67%, miR-1226-3p in 50% (p ≤ 0.04. MiR-130a-3p sensitized 83% of cell lines towards 5-FU, miR-148a-3p/miR-125a-5p/miR-1226-3p only 33% (p ≤ 0.015. Several resistance-relevant pathways seem to be targeted on various levels. Bcl-2 was confirmed as a direct target of miR-130a-3p and miR-148a-3p, and p53 as a target of miR-125a-5p. All microRNAs decreased migration and adhesion, except miR-130a-3p, and increased apoptosis. Simultaneous manipulation of two microRNAs exhibited additive sensitizing effects towards cisplatin in 50% (miR-125a-5p/miR-148a-3p, and 75% (miR-148a-3p/miR-130a-3p of cell lines (p ≤ 0.006. Conclusion: Our data present strong evidence that specific microRNA signatures are responsible for drug resistance and aggressiveness of ESCC. Final functional readout of these complex processes appears to be more important than single microRNA-target interactions.

  2. Photochemistry of monodentate and bidentate carbonato complexes of rhodium (3). [applications to spacecraft fuel cells

    Science.gov (United States)

    Sheridan, P. S.

    1980-01-01

    A scheme for the photochemical fixation of water is proposed which involves a five-step reaction sequence; the first step involves the 2 electron reduction of a metal by a coordinated carbonate ligand, with corresponding oxidation of the carbonate to CO2 and O2. Ligand field photolysis of trans- (RH(en)2 H2O CO3) ClO4, and (Rh(en)2 CO3) CLO4 have been studied in the solid state and in aqueous solution at various pH values. Both salts are photoinert in the solid phase, but are quite photoreactive in aqueous solution. In solution, the monodentate ion undergoes efficient isomerization to a mixture of cis and trans - (Rh(en)2 H2O CO3)+, presumably with water exchange. A minor pH increase upon photolysis is evidence of inefficient carbonate (CO3 =) release, with formation of (Rh(en)2 (H2O)2)3+. In contrast, aqueous solutions of the bidentate carbonato complex undergo efficient pH decrease upon ligand field photolysis. Changes in the electronic spectrum (200-500 nm) and pH changes indicate that the desired redox is occurring. The pH increase is due to the aqueous behavior of CO2.

  3. Evidence of active transport of cadmium complexing dithiocarbamates into renal and hepatic cells in vivo

    International Nuclear Information System (INIS)

    Gale, G.R.; Smith, A.B.; Jones, M.M.; Singh, P.K.

    1992-01-01

    A study was made of the effects of certain inhibitors of transport systems on the actions of four cadmium (Cd) complexing N,N-disubstituted dithiocarbamates (DTCs) in mobilizing murine renal and hepatic Cd in vivo. Probenecid, the prototypical antagonist of organic anion transport in the kidney, when given 1 hr prior to each DTC, sharply suppressed the DTC-induced reduction of renal Cd but was virtually without effect on mobilization of Cd from liver. Sulfinpyrazone, which blocks tubular reabsorption of uric acid and also inhibits transport of a variety of organic acids, inhibited markedly the mobilization of both renal and hepatic Cd by DTCs. Phlorizin, an inhibitor of tubular sugar reabsorption, did not affect the Cd mobilizing actions of DTCs in any consistent fashion. We propose that the high degree of selectivity of DTCs in mobilizing renal hepatic Cd is dependent, at lest in part, upon active transport of DTCs into these tissues via the organic anion transport systems. This report presents the first evidence that compounds of the (R) 2 NCSS - class may gain access to intracellular space by an active, carrier-mediated process. (au)

  4. Low-Complexity Interference-Free Downlink Channel Assignment with Improved Performance in Coordinated Small Cells

    KAUST Repository

    Radaydeh, Redha M.

    2015-05-01

    This paper proposes a low-complexity interference-free channel assignment scheme with improved desired downlink performance in coordinated multi-antenna small-coverage access points (APs) that employ the open-access control strategy. The adopted system treats the case when each user can be granted an access to one of the available channels at a time. Moreover, each receive terminal can suppress a limited number of resolvable interfering sources via its highly-correlated receive array. On the other hand, the operation of the deployed APs can be coordinated to serve active users, and the availability of multiple physical channels and the use of uncorrelated transmit antennas at each AP are exploited to improve the performance of supported users. The analysis provides new approaches to use the transmit antenna array at each AP, the multiple physical channels, the receive antenna array at each user in order to identify interference-free channels per each user, and then to select a downlink channel that provides the best possible improved performance. The event of concurrent interference-free channel identification by different users is also treated to further improve the desired link associated with the scheduled user. The analysis considers the practical scenario of imperfect identification of interference-free channel by an active user and/or the imperfectness in scheduling concurrent users requests on the same channel. The developed formulations can be used to study any performance metric and they are applicable for any statistical and geometric channel models. © 2015 IEEE.

  5. Hyperoxia decreases glycolytic capacity, glycolytic reserve and oxidative phosphorylation in MLE-12 cells and inhibits complex I and II function, but not complex IV in isolated mouse lung mitochondria.

    Directory of Open Access Journals (Sweden)

    Kumuda C Das

    Full Text Available High levels of oxygen (hyperoxia are frequently used in critical care units and in conditions of respiratory insufficiencies in adults, as well as in infants. However, hyperoxia has been implicated in a number of pulmonary disorders including bronchopulmonary dysplasia (BPD and adult respiratory distress syndrome (ARDS. Hyperoxia increases the generation of reactive oxygen species (ROS in the mitochondria that could impair the function of the mitochondrial electron transport chain. We analyzed lung mitochondrial function in hyperoxia using the XF24 analyzer (extracellular flux and optimized the assay for lung epithelial cells and mitochondria isolated from lungs of mice. Our data show that hyperoxia decreases basal oxygen consumption rate (OCR, spare respiratory capacity, maximal respiration and ATP turnover in MLE-12 cells. There was significant decrease in glycolytic capacity and glycolytic reserve in MLE-12 cells exposed to hyperoxia. Using mitochondria isolated from lungs of mice exposed to hyperoxia or normoxia we have shown that hyperoxia decreased the basal, state 3 and state3 μ (respiration in an uncoupled state respirations. Further, using substrate or inhibitor of a specific complex we show that the OCR via complex I and II, but not complex IV was decreased, demonstrating that complexes I and II are specific targets of hyperoxia. Further, the activities of complex I (NADH dehydrogenase, NADH-DH and complex II (succinate dehydrogenase, SDH were decreased in hyperoxia, but the activity of complex IV (cytochrome oxidase, COX remains unchanged. Taken together, our study show that hyperoxia impairs glycolytic and mitochondrial energy metabolism in in tact cells, as well as in lungs of mice by selectively inactivating components of electron transport system.

  6. Involvement of F-Actin in Chaperonin-Containing t-Complex 1 Beta Regulating Mouse Mesangial Cell Functions in a Glucose-Induction Cell Model

    Directory of Open Access Journals (Sweden)

    Jin-Shuen Chen

    2011-01-01

    Full Text Available The aim of this study is to investigate the role of chaperonin-containing t-complex polypeptide 1 beta (CCT2 in the regulation of mouse mesangial cell (mMC contraction, proliferation, and migration with filamentous/globular-(F/G- actin ratio under high glucose induction. A low CCT2 mMC model induced by treatment of small interference RNA was established. Groups with and without low CCT2 induction examined in normal and high (H glucose conditions revealed the following major results: (1 low CCT2 or H glucose showed the ability to attenuate F/G-actin ratio; (2 groups with low F/G-actin ratio all showed less cell contraction; (3 suppression of CCT2 may reduce the proliferation and migration which were originally induced by H glucose. In conclusion, CCT2 can be used as a specific regulator for mMC contraction, proliferation, and migration affected by glucose, which mechanism may involve the alteration of F-actin, particularly for cell contraction.

  7. Development of pyridine-containing macrocyclic copper(II) complexes: potential role in the redox modulation of oxaliplatin toxicity in human breast cells.

    Science.gov (United States)

    Fernandes, Ana S; Costa, Judite; Gaspar, Jorge; Rueff, José; Cabral, M Fátima; Cipriano, Madalena; Castro, Matilde; Oliveira, Nuno G

    2012-09-01

    The unique redox and catalytic chemistry of Cu has justified the development of novel Cu complexes for different therapeutic uses including cancer therapy. In this work, four pyridine-containing aza-macrocyclic copper(II) complexes were prepared (CuL1-CuL4) varying in ring size and/or substituents and their superoxide scavenging activity evaluated. CuL3, the most active superoxide scavenger, was further studied as a modulator of the cytotoxicity of oxaliplatin in epithelial breast MCF10A cells and in MCF7 breast cancer cells. Our results show that CuL3 enhances the therapeutic window of oxaliplatin, by both protecting non-tumour cells and increasing its cytotoxic effect in breast carcinoma cells. CuL3 is thus a promising complex to be further studied and to be used as a lead compound for the optimization of novel chemotherapy sensitizers.

  8. VEGF-A/NRP1 stimulates GIPC1 and Syx complex formation to promote RhoA activation and proliferation in skin cancer cells

    Directory of Open Access Journals (Sweden)

    Ayumi Yoshida

    2015-09-01

    Full Text Available Neuropilin-1 (NRP1 has been identified as a VEGF-A receptor. DJM-1, a human skin cancer cell line, expresses endogenous VEGF-A and NRP1. In the present study, the RNA interference of VEGF-A or NRP1 suppressed DJM-1 cell proliferation. Furthermore, the overexpression of the NRP1 wild type restored shNRP1-treated DJM-1 cell proliferation, whereas NRP1 cytoplasmic deletion mutants did not. A co-immunoprecipitation analysis revealed that VEGF-A induced interactions between NRP1 and GIPC1, a scaffold protein, and complex formation between GIPC1 and Syx, a RhoGEF. The knockdown of GIPC1 or Syx reduced active RhoA and DJM-1 cell proliferation without affecting the MAPK or Akt pathway. C3 exoenzyme or Y27632 inhibited the VEGF-A-induced proliferation of DJM-1 cells. Conversely, the overexpression of the constitutively active form of RhoA restored the proliferation of siVEGF-A-treated DJM-1 cells. Furthermore, the inhibition of VEGF-A/NRP1 signaling upregulated p27, a CDK inhibitor. A cell-penetrating oligopeptide that targeted GIPC1/Syx complex formation inhibited the VEGF-A-induced activation of RhoA and suppressed DJM-1 cell proliferation. In conclusion, this new signaling pathway of VEGF-A/NRP1 induced cancer cell proliferation by forming a GIPC1/Syx complex that activated RhoA to degrade the p27 protein.

  9. Cell phone ringtone, but not landline phone ringtone, affects complex reaction time

    Directory of Open Access Journals (Sweden)

    Radosław Zajdel

    2013-02-01

    Full Text Available Introduction: Legislation systems of most countries prohibited using the handheld mobile phone while driving due to the fact that it disturbs concentration and causes hand involvement. Every phone owner is accustomed to the ringtone of his phone and almost involuntarily endeavors to pick it up or check who calls. This engages one’s psychomotor skills, which in our opinion contributes to the attenuation of reaction time needed for performing other crucial functions. Objectives: The aim of the study was: (1 to evaluate the infl uence of the sound of a ringing mobile phone on the complex reaction time (RT score in healthy subjects (owners, and (2 to check if there are any differences in RT when a landline phone and mobile phone ring. Methods: To assess RT we used our system and protocol of examination, previously validated. The examination conditions were standardized. All tests were performed in the same room with the same light and general acoustic con