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Sample records for regulate cell spreading

  1. Deciphering the combinatorial roles of geometric, mechanical, and adhesion cues in regulation of cell spreading.

    Directory of Open Access Journals (Sweden)

    Greg M Harris

    Full Text Available Significant effort has gone towards parsing out the effects of surrounding microenvironment on macroscopic behavior of stem cells. Many of the microenvironmental cues, however, are intertwined, and thus, further studies are warranted to identify the intricate interplay among the conflicting downstream signaling pathways that ultimately guide a cell response. In this contribution, by patterning adhesive PEG (polyethylene glycol hydrogels using Dip Pen Nanolithography (DPN, we demonstrate that substrate elasticity, subcellular elasticity, ligand density, and topography ultimately define mesenchymal stem cells (MSCs spreading and shape. Physical characteristics are parsed individually with 7 kilopascal (kPa hydrogel islands leading to smaller, spindle shaped cells and 105 kPa hydrogel islands leading to larger, polygonal cell shapes. In a parallel effort, a finite element model was constructed to characterize and confirm experimental findings and aid as a predictive tool in modeling cell microenvironments. Signaling pathway inhibition studies suggested that RhoA is a key regulator of cell response to the cooperative effect of the tunable substrate variables. These results are significant for the engineering of cell-extra cellular matrix interfaces and ultimately decoupling matrix bound cues presented to cells in a tissue microenvironment for regenerative medicine.

  2. Myo1c regulates lipid raft recycling to control cell spreading, migration and Salmonella invasion.

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    Brandstaetter, Hemma; Kendrick-Jones, John; Buss, Folma

    2012-04-15

    A balance between endocytosis and membrane recycling regulates the composition and dynamics of the plasma membrane. Internalization and recycling of cholesterol- and sphingolipid-enriched lipid rafts is an actin-dependent process that is mediated by a specialized Arf6-dependent recycling pathway. Here, we identify myosin1c (Myo1c) as the first motor protein that drives the formation of recycling tubules emanating from the perinuclear recycling compartment. We demonstrate that the single-headed Myo1c is a lipid-raft-associated motor protein that is specifically involved in recycling of lipid-raft-associated glycosylphosphatidylinositol (GPI)-linked cargo proteins and their delivery to the cell surface. Whereas Myo1c overexpression increases the levels of these raft proteins at the cell surface, in cells depleted of Myo1c function through RNA interference or overexpression of a dominant-negative mutant, these tubular transport carriers of the recycling pathway are lost and GPI-linked raft markers are trapped in the perinuclear recycling compartment. Intriguingly, Myo1c only selectively promotes delivery of lipid raft membranes back to the cell surface and is not required for recycling of cargo, such as the transferrin receptor, which is mediated by parallel pathways. The profound defect in lipid raft trafficking in Myo1c-knockdown cells has a dramatic impact on cell spreading, cell migration and cholesterol-dependent Salmonella invasion; processes that require lipid raft transport to the cell surface to deliver signaling components and the extra membrane essential for cell surface expansion and remodeling. Thus, Myo1c plays a crucial role in the recycling of lipid raft membrane and proteins that regulate plasma membrane plasticity, cell motility and pathogen entry.

  3. The F-BAR domain protein PACSIN2 associates with Rac1 and regulates cell spreading and migration

    NARCIS (Netherlands)

    de Kreuk, Bart-Jan; Nethe, Micha; Fernandez-Borja, Mar; Anthony, Eloise C.; Hensbergen, Paul J.; Deelder, Andre M.; Plomann, Markus; Hordijk, Peter L.

    2011-01-01

    The Rac1 GTPase controls cytoskeletal dynamics and is a key regulator of cell spreading and migration mediated by signaling through effector proteins, such as the PAK kinases and the Scar and WAVE proteins. We previously identified a series of regulatory proteins that associate with Rac1 through its

  4. Tyrosine Phosphorylation of Rac1: A Role in Regulation of Cell Spreading

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    Chang, Fumin; Lemmon, Christopher; Lietha, Daniel; Eck, Michael; Romer, Lewis

    2011-01-01

    Rac1 influences a multiplicity of vital cellular- and tissue-level control functions, making it an important candidate for targeted therapeutics. The activity of the Rho family member Cdc42 has been shown to be modulated by tyrosine phosphorylation at position 64. We therefore investigated consequences of the point mutations Y64F and Y64D in Rac1. Both mutations altered cell spreading from baseline in the settings of wild type, constitutively active, or dominant negative Rac1 expression, and were accompanied by differences in Rac1 targeting to focal adhesions. Rac1-Y64F displayed increased GTP-binding, increased association with βPIX, and reduced binding with RhoGDI as compared with wild type Rac1. Rac1-Y64D had less binding to PAK than Rac1-WT or Rac1-64F. In vitro assays demonstrated that Y64 in Rac1 is a target for FAK and Src. Taken together, these data suggest a mechanism for the regulation of Rac1 activity by non-receptor tyrosine kinases, with consequences for membrane extension. PMID:22163037

  5. The regulated secretory pathway in CD4(+ T cells contributes to human immunodeficiency virus type-1 cell-to-cell spread at the virological synapse.

    Directory of Open Access Journals (Sweden)

    Clare Jolly

    2011-09-01

    Full Text Available Direct cell-cell spread of Human Immunodeficiency Virus type-1 (HIV-1 at the virological synapse (VS is an efficient mode of dissemination between CD4(+ T cells but the mechanisms by which HIV-1 proteins are directed towards intercellular contacts is unclear. We have used confocal microscopy and electron tomography coupled with functional virology and cell biology of primary CD4(+ T cells from normal individuals and patients with Chediak-Higashi Syndrome and report that the HIV-1 VS displays a regulated secretion phenotype that shares features with polarized secretion at the T cell immunological synapse (IS. Cell-cell contact at the VS re-orientates the microtubule organizing center (MTOC and organelles within the HIV-1-infected T cell towards the engaged target T cell, concomitant with polarization of viral proteins. Directed secretion of proteins at the T cell IS requires specialized organelles termed secretory lysosomes (SL and we show that the HIV-1 envelope glycoprotein (Env localizes with CTLA-4 and FasL in SL-related compartments and at the VS. Finally, CD4(+ T cells that are disabled for regulated secretion are less able to support productive cell-to-cell HIV-1 spread. We propose that HIV-1 hijacks the regulated secretory pathway of CD4(+ T cells to enhance its dissemination.

  6. Lipid raft regulates the initial spreading of melanoma A375 cells by modulating β1 integrin clustering.

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    Wang, Ruifei; Bi, Jiajia; Ampah, Khamal Kwesi; Zhang, Chunmei; Li, Ziyi; Jiao, Yang; Wang, Xiaoru; Ba, Xueqing; Zeng, Xianlu

    2013-08-01

    Cell adhesion and spreading require integrins-mediated cell-extracellular matrix interaction. Integrins function through binding to extracellular matrix and subsequent clustering to initiate focal adhesion formation and actin cytoskeleton rearrangement. Lipid raft, a liquid ordered plasma membrane microdomain, has been reported to play major roles in membrane motility by regulating cell surface receptor function. Here, we identified that lipid raft integrity was required for β1 integrin-mediated initial spreading of melanoma A375 cells on fibronectin. We found that lipid raft disruption with methyl-β-cyclodextrin led to the inability of focal adhesion formation and actin cytoskeleton rearrangement by preventing β1 integrin clustering. Furthermore, we explored the possible mechanism by which lipid raft regulates β1 integrin clustering and demonstrated that intact lipid raft could recruit and modify some adaptor proteins, such as talin, α-actinin, vinculin, paxillin and FAK. Lipid raft could regulate the location of these proteins in lipid raft fractions and facilitate their binding to β1 integrin, which may be crucial for β1 integrin clustering. We also showed that lipid raft disruption impaired A375 cell migration in both transwell and wound healing models. Together, these findings provide a new insight for the relationship between lipid raft and the regulation of integrins. Copyright © 2013 Elsevier Ltd. All rights reserved.

  7. Laminin isoforms differentially regulate adhesion, spreading, proliferation, and ERK activation of β1 integrin-null cells

    International Nuclear Information System (INIS)

    Kikkawa, Yamato; Yu, Hao; Genersch, Elke; Sanzen, Noriko; Sekiguchi, Kiyotoshi; Faessler, Reinhard; Campbell, Kevin P.; Talts, Jan F.; Ekblom, Peter

    2004-01-01

    The presence of many laminin receptors of the β1 integrin family on most cells makes it difficult to define the biological functions of other major laminin receptors such as integrin α6β4 and dystroglycan. We therefore tested the binding of a β1 integrin-null cell line GD25 to four different laminin variants. The cells were shown to produce dystroglycan, which based on affinity chromatography bound to laminin-1, -2/4, and -10/11, but not to laminin-5. The cells also expressed the integrin α6Aβ4A variant. GD25 β1 integrin-null cells are known to bind poorly to laminin-1, but we demonstrate here that these cells bind avidly to laminin-2/4, -5, and -10/11. The initial binding at 20 min to each of these laminins could be inhibited by an integrin α6 antibody, but not by a dystroglycan antibody. Hence, integrin α6Aβ4A of GD25 cells was identified as a major receptor for initial GD25 cell adhesion to three out of four tested laminin isoforms. Remarkably, cell adhesion to laminin-5 failed to promote cell spreading, proliferation, and extracellular signal-regulated kinase (ERK) activation, whereas all these responses occurred in response to adhesion to laminin-2/4 or -10/11. The data establish GD25 cells as useful tools to define the role integrin α6Aβ4A and suggest that laminin isoforms have distinctly different capacities to promote cell adhesion and signaling via integrin α6Aβ4A

  8. PDGF-regulated rab4-dependent recycling of alphavbeta3 integrin from early endosomes is necessary for cell adhesion and spreading.

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    Roberts, M; Barry, S; Woods, A; van der Sluijs, P; Norman, J

    2001-09-18

    It has been postulated that the regulation of integrin vesicular traffic facilitates cell migration by internalizing integrins at the rear of the cell and transporting them forward within vesicles for exocytosis at the leading edge to form new contacts with the extracellular matrix. The rab family of GTPases control key targeting events in the endo/exocytic pathway; therefore, these GTPases may be involved in the regulation of cell-matrix contact assembly. The endo/exocytic cycle of alphavbeta3 and alpha5beta1 integrins was studied using mouse 3T3 fibroblast cell lines. In serum-starved cells, internalized integrins were transported through rab4-positive, early endosomes and arrived at the rab11-positive, perinuclear recycling compartment approximately 30 min after endocytosis. From the recycling compartment, integrins were recycled to the plasma membrane in a rab11-dependent fashion. Following treatment with PDGF, alphavbeta3 integrin, but not alpha5beta1, was rapidly recycled directly back to the plasma membrane from the early endosomes via a rab4-dependent mechanism without the involvement of rab11. This rapid recycling pathway directed alphavbeta3 to numerous small puncta distributed evenly across the dorsal surface of the cell, and the integrin only became localized into focal complexes at later times following PDGF addition. Interestingly, inhibition of PDGF-stimulated alphavbeta3 recycling using dominant-negative rab4 mutants compromised cell adhesion and spreading on vitronectin (a ligand for alphavbeta3), but adhesion to fibronectin (a ligand for alpha5beta1 and alphavbeta3) was unchanged. We propose that growth factor-regulated, rab4-dependent recycling of alphavbeta3 integrin from early endosomes to the plasma membrane is a critical upstream event in the assembly of cell-matrix contacts.

  9. Free energy analysis of cell spreading.

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    McEvoy, Eóin; Deshpande, Vikram S; McGarry, Patrick

    2017-10-01

    In this study we present a steady-state adaptation of the thermodynamically motivated stress fiber (SF) model of Vigliotti et al. (2015). We implement this steady-state formulation in a non-local finite element setting where we also consider global conservation of the total number of cytoskeletal proteins within the cell, global conservation of the number of binding integrins on the cell membrane, and adhesion limiting ligand density on the substrate surface. We present a number of simulations of cell spreading in which we consider a limited subset of the possible deformed spread-states assumed by the cell in order to examine the hypothesis that free energy minimization drives the process of cell spreading. Simulations suggest that cell spreading can be viewed as a competition between (i) decreasing cytoskeletal free energy due to strain induced assembly of cytoskeletal proteins into contractile SFs, and (ii) increasing elastic free energy due to stretching of the mechanically passive components of the cell. The computed minimum free energy spread area is shown to be lower for a cell on a compliant substrate than on a rigid substrate. Furthermore, a low substrate ligand density is found to limit cell spreading. The predicted dependence of cell spread area on substrate stiffness and ligand density is in agreement with the experiments of Engler et al. (2003). We also simulate the experiments of Théry et al. (2006), whereby initially circular cells deform and adhere to "V-shaped" and "Y-shaped" ligand patches. Analysis of a number of different spread states reveals that deformed configurations with the lowest free energy exhibit a SF distribution that corresponds to experimental observations, i.e. a high concentration of highly aligned SFs occurs along free edges, with lower SF concentrations in the interior of the cell. In summary, the results of this study suggest that cell spreading is driven by free energy minimization based on a competition between decreasing

  10. Cortactin Tyrosine Phosphorylation Promotes Its Deacetylation and Inhibits Cell Spreading

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    Meiler, Eugenia; Nieto-Pelegrín, Elvira; Martinez-Quiles, Narcisa

    2012-01-01

    Background Cortactin is a classical Src kinase substrate that participates in actin cytoskeletal dynamics by activating the Arp2/3 complex and interacting with other regulatory proteins, including FAK. Cortactin has various domains that may contribute to the assembly of different protein platforms to achieve process specificity. Though the protein is known to be regulated by post-translational modifications such as phosphorylation and acetylation, how tyrosine phosphorylation regulates cortactin activity is poorly understood. Since the basal level of tyrosine phosphorylation is low, this question must be studied using stimulated cell cultures, which are physiologically relevant but unreliable and difficult to work with. In fact, their unreliability may be the cause of some contradictory findings about the dynamics of tyrosine phosphorylation of cortactin in different processes. Methodology/Principal Findings In the present study, we try to overcome these problems by using a Functional Interaction Trap (FIT) system, which involves cotransfecting cells with a kinase (Src) and a target protein (cortactin), both of which are fused to complementary leucine-zipper domains. The FIT system allowed us to control precisely the tyrosine phosphorylation of cortactin and explore its relationship with cortactin acetylation. Conclusions/Significance Using this system, we provide definitive evidence that a competition exists between acetylation and tyrosine phosphorylation of cortactin and that phosphorylation inhibits cell spreading. We confirmed the results from the FIT system by examining endogenous cortactin in different cell types. Furthermore, we demonstrate that cell spreading promotes the association of cortactin and FAK and that tyrosine phosphorylation of cortactin disrupts this interaction, which may explain how it inhibits cell spreading. PMID:22479425

  11. Live Cell Imaging of Alphaherpes Virus Anterograde Transport and Spread

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    Taylor, Matthew P.; Kratchmarov, Radomir; Enquist, Lynn W.

    2013-01-01

    Advances in live cell fluorescence microscopy techniques, as well as the construction of recombinant viral strains that express fluorescent fusion proteins have enabled real-time visualization of transport and spread of alphaherpes virus infection of neurons. The utility of novel fluorescent fusion proteins to viral membrane, tegument, and capsids, in conjunction with live cell imaging, identified viral particle assemblies undergoing transport within axons. Similar tools have been successfully employed for analyses of cell-cell spread of viral particles to quantify the number and diversity of virions transmitted between cells. Importantly, the techniques of live cell imaging of anterograde transport and spread produce a wealth of information including particle transport velocities, distributions of particles, and temporal analyses of protein localization. Alongside classical viral genetic techniques, these methodologies have provided critical insights into important mechanistic questions. In this article we describe in detail the imaging methods that were developed to answer basic questions of alphaherpes virus transport and spread. PMID:23978901

  12. Dynamics of cell area and force during spreading.

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    Brill-Karniely, Yifat; Nisenholz, Noam; Rajendran, Kavitha; Dang, Quynh; Krishnan, Ramaswamy; Zemel, Assaf

    2014-12-16

    Experiments on human pulmonary artery endothelial cells are presented to show that cell area and the force exerted on a substrate increase simultaneously, but with different rates during spreading; rapid-force increase systematically occurred several minutes past initial spreading. We examine this theoretically and present three complementary mechanisms that may accompany the development of lamellar stress during spreading and underlie the observed behavior. These include: 1), the dynamics of cytoskeleton assembly at the cell basis; 2), the strengthening of acto-myosin forces in response to the generated lamellar stresses; and 3), the passive strain-stiffening of the cytoskeleton. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  13. [SPREADING OF NCTC CLONE 929 CELLS AFTER RESEEDING].

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    Petrov, Yu P; Negulyaev, Yu A; Tsupkina, N V

    2015-01-01

    The period (1 h after reseeding) of behaviour of mouse NCTC clone 929 cells to the conditions of artificial cultivation was studied. The time-lapse imaging followed the processing of the cells with ImageJ program was applied. To characterize the parametres cell status we used the cell area (projection of the cell on substrate) and Rp/Ra ratio introduced earlier as a spreading coefficient (Kuz'minykh, Petrov, 2004). After attaching a substratum, cells have a form of sphere (the phase "sphere") as the daughter cells after a mitosis. We revealed however that after this phase the reseeded cells do not start usual spreading and migration along substratum. They pass a phase of equally spreading in all directions and shaping their area as a circle (phase "circle"). This phase is absent of the daughter cells spreading after mitosis. We assume that the phase "circle" is a result of adaptation of the cells to reseedings at artificial cultivation. It is necessary for formation of a substrate composed of own extracellular matrix components (ECM) of the cells. Own ECM facilitates transition of the cells to their usual spreading and migration along substratum.

  14. Trigeminal perineural spread of renal cell carcinoma

    International Nuclear Information System (INIS)

    Hornik, Alejandro; Rosenblum, Jordan; Biller, Jose

    2012-01-01

    A 55-year-old man had a five-day history of “pins and needles” sensation on the left chin. Examination showed decreased pinprick sensation on the territory of the left mandibular branch of the trigeminal nerve. Brain magnetic resonance imaging (MRI) with gadolinium showed enhancement involving the left mandibular branch. Computed tomography (CT) of the chest, abdomen, and pelvis showed a left kidney mass diagnosed as renal carcinoma following nephrectomy. The “numb-chin” syndrome heralds or accompanies systemic malignancies. Trigeminal perineural spread has been well-documented in head and neck neoplasms, however, to our knowledge, it has not been reported in renal neoplasms. (author)

  15. Modeling universal dynamics of cell spreading on elastic substrates.

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    Fan, Houfu; Li, Shaofan

    2015-11-01

    A three-dimensional (3D) multiscale moving contact line model is combined with a soft matter cell model to study the universal dynamics of cell spreading over elastic substrates. We have studied both the early stage and the late stage cell spreading by taking into account the actin tension effect. In this work, the cell is modeled as an active nematic droplet, and the substrate is modeled as a St. Venant Kirchhoff elastic medium. A complete 3D simulation of cell spreading has been carried out. The simulation results show that the spreading area versus spreading time at different stages obeys specific power laws, which is in good agreement with experimental data and theoretical prediction reported in the literature. Moreover, the simulation results show that the substrate elasticity may affect force dipole distribution inside the cell. The advantage of this approach is that it combines the hydrodynamics of actin retrograde flow with moving contact line model so that it can naturally include actin tension effect resulting from actin polymerization and actomyosin contraction, and thus it might be capable of simulating complex cellular scale phenomenon, such as cell spreading or even crawling.

  16. Contact enhancement of locomotion in spreading cell colonies

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    D'Alessandro, Joseph; Solon, Alexandre P.; Hayakawa, Yoshinori; Anjard, Christophe; Detcheverry, François; Rieu, Jean-Paul; Rivière, Charlotte

    2017-10-01

    The dispersal of cells from an initially constrained location is a crucial aspect of many physiological phenomena, ranging from morphogenesis to tumour spreading. In such processes, cell-cell interactions may deeply alter the motion of single cells, and in turn the collective dynamics. While contact phenomena like contact inhibition of locomotion are known to come into play at high densities, here we focus on the little explored case of non-cohesive cells at moderate densities. We fully characterize the spreading of micropatterned colonies of Dictyostelium discoideum cells from the complete set of individual trajectories. From data analysis and simulation of an elementary model, we demonstrate that contact interactions act to speed up the early population spreading by promoting individual cells to a state of higher persistence, which constitutes an as-yet unreported contact enhancement of locomotion. Our findings also suggest that the current modelling paradigm of memoryless active particles may need to be extended to account for the history-dependent internal state of motile cells.

  17. RNAi screen reveals host cell kinases specifically involved in Listeria monocytogenes spread from cell to cell.

    Directory of Open Access Journals (Sweden)

    Ryan Chong

    Full Text Available Intracellular bacterial pathogens, such as Listeria monocytogenes and Rickettsia conorii display actin-based motility in the cytosol of infected cells and spread from cell to cell through the formation of membrane protrusions at the cell cortex. Whereas the mechanisms supporting cytosolic actin-based motility are fairly well understood, it is unclear whether specific host factors may be required for supporting the formation and resolution of membrane protrusions. To address this gap in knowledge, we have developed high-throughput fluorescence microscopy and computer-assisted image analysis procedures to quantify pathogen spread in human epithelial cells. We used the approach to screen a siRNA library covering the human kinome and identified 7 candidate kinases whose depletion led to severe spreading defects in cells infected with L. monocytogenes. We conducted systematic validation procedures with redundant silencing reagents and confirmed the involvement of the serine/threonine kinases, CSNK1A1 and CSNK2B. We conducted secondary assays showing that, in contrast with the situation observed in CSNK2B-depleted cells, L. monocytogenes formed wild-type cytosolic tails and displayed wild-type actin-based motility in the cytosol of CSNK1A1-depleted cells. Furthermore, we developed a protrusion formation assay and showed that the spreading defect observed in CSNK1A1-depleted cells correlated with the formation of protrusion that did not resolve into double-membrane vacuoles. Moreover, we developed sending and receiving cell-specific RNAi procedures and showed that CSNK1A was required in the sending cells, but was dispensable in the receiving cells, for protrusion resolution. Finally, we showed that the observed defects were specific to Listeria monocytogenes, as Rickettsia conorii displayed wild-type cell-to-cell spread in CSNK1A1- and CSNK2B-depleted cells. We conclude that, in addition to the specific host factors supporting cytosolic actin

  18. Local and Regional Spread of Primary Conjunctival Squamous Cell Carcinoma.

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    Desai, Shilpa J; Pruzan, Noelle L; Geske, Michael J; Jeng, Bennie H; Bloomer, Michele M; Vagefi, M Reza

    2016-04-06

    Two cases of biopsy-proven conjunctival squamous cell carcinoma (SCC) that developed local and regional spread are described. The cases involved a 65-year-old woman and a 79-year-old man who were initially treated at outside institutions for SCC of the conjunctiva. The patients did not have a history of immune compromise. The female patient presented with direct extension into the lacrimal gland but deferred recommended exenteration. Despite eventual exenteration, she developed metastasis to a neck node 6 months later, which was treated with radiotherapy. The male patient presented with local recurrence and a parotid node metastasis treated with exenteration, parotidectomy, selective neck dissection, and postoperative radiotherapy. Review of the outside pathology of both cases revealed positive tumor margins at the time of original resection. Local control of conjunctival SCC is of critical importance to reduce the risk of orbital extension and regional spread.

  19. Neutralisation of HIV-1 cell-cell spread by human and llama antibodies.

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    McCoy, Laura E; Groppelli, Elisabetta; Blanchetot, Christophe; de Haard, Hans; Verrips, Theo; Rutten, Lucy; Weiss, Robin A; Jolly, Clare

    2014-10-02

    Direct cell-cell spread of HIV-1 is a very efficient mode of viral dissemination, with increasing evidence suggesting that it may pose a considerable challenge to controlling viral replication in vivo. Much current vaccine research involves the study of broadly neutralising antibodies (bNabs) that arise during natural infection with the aims of eliciting such antibodies by vaccination or incorporating them into novel therapeutics. However, whether cell-cell spread of HIV-1 can be effectively targeted by bNabs remains unclear, and there is much interest in identifying antibodies capable of efficiently neutralising virus transmitted by cell-cell contact. In this study we have tested a panel of bNAbs for inhibition of cell-cell spread, including some not previously evaluated for inhibition of this mode of HIV-1 transmission. We found that three CD4 binding site antibodies, one from an immunised llama (J3) and two isolated from HIV-1-positive patients (VRC01 and HJ16) neutralised cell-cell spread between T cells, while antibodies specific for glycan moieties (2G12, PG9, PG16) and the MPER (2F5) displayed variable efficacy. Notably, while J3 displayed a high level of potency during cell-cell spread we found that the small size of the llama heavy chain-only variable region (VHH) J3 is not required for efficient neutralisation since recombinant J3 containing a full-length human heavy chain Fc domain was significantly more potent. J3 and J3-Fc also neutralised cell-cell spread of HIV-1 from primary macrophages to CD4+ T cells. In conclusion, while bNabs display variable efficacy at preventing cell-cell spread of HIV-1, we find that some CD4 binding site antibodies can inhibit this mode of HIV-1 dissemination and identify the recently described llama antibody J3 as a particularly potent inhibitor. Effective neutralisation of cell-cell spread between physiologically relevant cell types by J3 and J3-Fc supports the development of VHH J3 nanobodies for therapeutic or

  20. The requirements for herpes simplex virus type 1 cell-cell spread via nectin-1 parallel those for virus entry.

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    Even, Deborah L; Henley, Allison M; Geraghty, Robert J

    2006-08-01

    Herpes simplex virus type 1 (HSV-1) spreads from an infected cell to an uninfected cell by virus entry, virus-induced cell fusion, and cell-cell spread. The three forms of virus spread require the viral proteins gB, gD, and gH-gL, as well as a cellular gD receptor. The mutual requirement for the fusion glycoproteins and gD receptor suggests that virus entry, cell fusion, and cell-cell spread occur by a similar mechanism. The goals of this study were to examine the role of the nectin-1alpha transmembrane domain and cytoplasmic tail in cell-cell spread and to obtain a better understanding of the receptor-dependent events occurring at the plasma membrane during cell-cell spread. We determined that an intact nectin-1alpha V-like domain was required for cell-cell spread, while a membrane-spanning domain and cytoplasmic tail were not. Chimeric forms of nectin-1 that were non-functional for virus entry did not mediate cell-cell spread regardless of whether they could mediate cell fusion. Also, cell-cell spread of syncytial isolates was dependent upon nectin-1alpha expression and occurred through a nectin-1-dependent mechanism. Taken together, our results indicate that nectin-1-dependent events occurring at the plasma membrane during cell-cell spread were equivalent to those for virus entry.

  1. NCAM regulates cell motility

    DEFF Research Database (Denmark)

    Prag, Søren; Lepekhin, Eugene A; Kolkova, Kateryna

    2002-01-01

    Cell migration is required during development of the nervous system. The regulatory mechanisms for this process, however, are poorly elucidated. We show here that expression of or exposure to the neural cell adhesion molecule (NCAM) strongly affected the motile behaviour of glioma cells...... independently of homophilic NCAM interactions. Expression of the transmembrane 140 kDa isoform of NCAM (NCAM-140) caused a significant reduction in cellular motility, probably through interference with factors regulating cellular attachment, as NCAM-140-expressing cells exhibited a decreased attachment...... to a fibronectin substratum compared with NCAM-negative cells. Ectopic expression of the cytoplasmic part of NCAM-140 also inhibited cell motility, presumably via the non-receptor tyrosine kinase p59(fyn) with which NCAM-140 interacts. Furthermore, we showed that the extracellular part of NCAM acted as a paracrine...

  2. Inhibition of HSV cell-to-cell spread by lactoferrin and lactoferricin.

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    Jenssen, Håvard; Sandvik, Kjersti; Andersen, Jeanette H; Hancock, Robert E W; Gutteberg, Tore J

    2008-09-01

    The milk protein lactoferrin (Lf) has multiple functions, including immune stimulation and antiviral activity towards herpes simplex virus 1 and 2 (HSV-1 and HSV-2); antiviral activity has also been reported for the N-terminal pepsin-derived fragment lactoferricin (Lfcin). The anti-HSV mode of action of Lf and Lfcin is assumed to involve, in part, their interaction with the cell surface glycosaminoglycan heparan sulfate, thereby blocking of viral entry. In this study we investigated the ability of human and bovine Lf and Lfcin to inhibit viral cell-to-cell spread as well as the involvement of cell surface glycosaminoglycans during viral cell-to-cell spread. Lf and Lfcin from both human and bovine origin, inhibited cell-to-cell spread of both HSV-1 and HSV-2. Inhibition of cell-to-cell spread by bovine Lfcin involved cell surface chondroitin sulfate. Based on transmission electron microscopy studies, human Lfcin, like bovine Lfcin, was randomly distributed intracellularly, thus differences in their antiviral activity could not be explained by differences in their distribution. In contrast, the cellular localization of iron-saturated (holo)-Lf appeared to differ from that of apo-Lf, indicating that holo- and apo-Lf may exhibit different antiviral mechanisms.

  3. Bacterial spread from cell to cell: beyond actin-based motility.

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    Kuehl, Carole J; Dragoi, Ana-Maria; Talman, Arthur; Agaisse, Hervé

    2015-09-01

    Several intracellular pathogens display the ability to propagate within host tissues by displaying actin-based motility in the cytosol of infected cells. As motile bacteria reach cell-cell contacts they form plasma membrane protrusions that project into adjacent cells and resolve into vacuoles from which the pathogen escapes, thereby achieving spread from cell to cell. Seminal studies have defined the bacterial and cellular factors that support actin-based motility. By contrast, the mechanisms supporting the formation of protrusions and their resolution into vacuoles have remained elusive. Here, we review recent advances in the field showing that Listeria monocytogenes and Shigella flexneri have evolved pathogen-specific mechanisms of bacterial spread from cell to cell. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. TiO2-enriched polymeric powder coatings support human mesenchymal cell spreading and osteogenic differentiation

    International Nuclear Information System (INIS)

    Mozumder, Mohammad Sayem; Zhu, Jesse; Perinpanayagam, Hiran

    2011-01-01

    Novel polymeric powder coatings (PPC) were prepared by ultrafine powder coating technology and shown to support human mesenchymal cell attachment and growth. PPC surfaces enriched with nano-TiO 2 (nTiO 2 ) showed enhanced cellular responses, and were compared to commercially pure titanium (cpTi). After cell attachment and growth, osteogenic differentiation and bone matrix formation ensures osseointegration for implantable biomaterials. Therefore, the objective of this study was to determine if mesenchymal cells grown on PPC could undergo osteogenic differentiation by inducing Runx2 and bone matrix proteins, and then initiate mineralization. Atomic force microscopy revealed intricate three-dimensional micro-topographies, and the measures of nano-roughness and porosity were similar for all PPC surfaces. Scanning electron microscopy showed that the cells attached and spread out over all of the surfaces. After 1 week in osteogenic media, RT-PCR analysis showed the induction of Runx2, the up-regulation of type I collagen, and the initial detection of alkaline phosphatase and bone sialoprotein. After 4 weeks, Alizarin Red staining showed mineral deposition. However, cell spreading and osteogenic differentiation were significantly (P 2 additives may enhance their performance.

  5. HIV Cell-to-Cell Spread Results in Earlier Onset of Viral Gene Expression by Multiple Infections per Cell.

    Directory of Open Access Journals (Sweden)

    Mikaël Boullé

    2016-11-01

    Full Text Available Cell-to-cell spread of HIV, a directed mode of viral transmission, has been observed to be more rapid than cell-free infection. However, a mechanism for earlier onset of viral gene expression in cell-to-cell spread was previously uncharacterized. Here we used time-lapse microscopy combined with automated image analysis to quantify the timing of the onset of HIV gene expression in a fluorescent reporter cell line, as well as single cell staining for infection over time in primary cells. We compared cell-to-cell spread of HIV to cell-free infection, and limited both types of transmission to a two-hour window to minimize differences due to virus transit time to the cell. The mean time to detectable onset of viral gene expression in cell-to-cell spread was accelerated by 19% in the reporter cell line and by 35% in peripheral blood mononuclear cells relative to cell-free HIV infection. Neither factors secreted by infected cells, nor contact with infected cells in the absence of transmission, detectably changed onset. We recapitulated the earlier onset by infecting with multiple cell-free viruses per cell. Surprisingly, the acceleration in onset of viral gene expression was not explained by cooperativity between infecting virions. Instead, more rapid onset was consistent with a model where the fastest expressing virus out of the infecting virus pool sets the time for infection independently of the other co-infecting viruses.

  6. Semaphorin SEMA3F and VEGF Have Opposing Effects on Cell Attachment and Spreading

    Directory of Open Access Journals (Sweden)

    Patrick Nasarre

    2003-01-01

    Full Text Available SEMA3F, isolated from a 3p21.3 deletion, has antitumor activity in transfected cells, and protein expression correlates with tumor stage and histology. In primary tumors, SEMA3F and VEGF surface staining is inversely correlated. Coupled with SEMA3F at the leading edge of motile cells, we previously suggested that both proteins competitively regulate cell motility and adhesion. We have investigated this using the breast cancer cell line, MCF7. SEMA3F inhibited cell attachment and spreading as evidenced by loss of lamellipodia extensions, membrane ruffling, and cell-cell contacts, with cells eventually rounding-up and detaching. In contrast, VEGF had opposite effects. Although SEMA3F binds NRP2 with 10-fold greater affinity than NRP1, the effects in MCF7 were mediated by NRP1. This was determined by receptor expression and blocking of anti-NRP1 antibodies. Similar effects, but through NRP2, were observed in the C100 breast cancer cell line. Although we were unable to demonstrate changes in total GTPbound Rac1 or RhoA, we did observe changes in the localization of Rac1-GFP using time lapse microscopy. Following SEMA3F, Rac1 moved to the base of lamellipodia and — with their collapse — to the membrane. These results support the concept that SEMA3F and VEGF have antagonistic actions affecting motility in primary tumor cell.

  7. SNARE-mediated trafficking of α5β1 integrin is required for spreading in CHO cells

    International Nuclear Information System (INIS)

    Skalski, Michael; Coppolino, Marc G.

    2005-01-01

    In this study, the role of SNARE-mediated membrane traffic in regulating integrin localization was examined and the requirement for SNARE function in cellular spreading was quantitatively assessed. Membrane traffic was inhibited with the VAMP-specific catalytic light chain from tetanus toxin (TeTx-LC), a dominant-negative form (E329Q) of N-ethylmaleimide-sensitive fusion protein (NSF), and brefeldin A (BfA). Inhibition of membrane traffic with either E329Q-NSF or TeTx-LC, but not BfA, significantly inhibited spreading of CHO cells on fibronectin. Spreading was rescued in TeTx-LC-expressing cells by co-transfection with a TeTx-resistant cellubrevin/VAMP3. E329Q-NSF, a general inhibitor of SNARE function, was a more potent inhibitor of cell spreading than TeTx-LC, suggesting that tetanus toxin-insensitive SNAREs contribute to adhesion. It was found that E329Q-NSF prevented trafficking of α 5 β 1 integrins from a central Rab11-containing compartment to sites of protrusion during cell adhesion, while TeTx-LC delayed this trafficking. These results are consistent with a model of cellular adhesion that implicates SNARE function as an important component of integrin trafficking during the process of cell spreading

  8. Molecular mechanisms of cell-cell spread of intracellular bacterial pathogens.

    Science.gov (United States)

    Ireton, Keith

    2013-07-17

    Several bacterial pathogens, including Listeria monocytogenes, Shigella flexneri and Rickettsia spp., have evolved mechanisms to actively spread within human tissues. Spreading is initiated by the pathogen-induced recruitment of host filamentous (F)-actin. F-actin forms a tail behind the microbe, propelling it through the cytoplasm. The motile pathogen then encounters the host plasma membrane, forming a bacterium-containing protrusion that is engulfed by an adjacent cell. Over the past two decades, much progress has been made in elucidating mechanisms of F-actin tail formation. Listeria and Shigella produce tails of branched actin filaments by subverting the host Arp2/3 complex. By contrast, Rickettsia forms tails with linear actin filaments through a bacterial mimic of eukaryotic formins. Compared with F-actin tail formation, mechanisms controlling bacterial protrusions are less well understood. However, recent findings have highlighted the importance of pathogen manipulation of host cell-cell junctions in spread. Listeria produces a soluble protein that enhances bacterial protrusions by perturbing tight junctions. Shigella protrusions are engulfed through a clathrin-mediated pathway at 'tricellular junctions'--specialized membrane regions at the intersection of three epithelial cells. This review summarizes key past findings in pathogen spread, and focuses on recent developments in actin-based motility and the formation and internalization of bacterial protrusions.

  9. Monitoring of living cell attachment and spreading using reverse symmetry waveguide sensing

    DEFF Research Database (Denmark)

    Horvath, R.; Pedersen, H.C.; Skivesen, N.

    2005-01-01

    The effect of the attachment and spreading of living cells on the modes of a grating coupled reverse symmetry waveguide sensor is investigated in real time. The reverse symmetry design has an increased probing depth into the sample making it well suited for the monitoring of cell morphology....... As a result, significant changes in the incoupling peak height and peak shape were observed during cell attachment and spreading. It is suggested that the area under the incoupling peaks reflects the initial cell attachment process, while the mean peak position is mostly governed by the spreading of the cells...

  10. Regulation of beta cell replication

    DEFF Research Database (Denmark)

    Lee, Ying C; Nielsen, Jens Høiriis

    2008-01-01

    Beta cell mass, at any given time, is governed by cell differentiation, neogenesis, increased or decreased cell size (cell hypertrophy or atrophy), cell death (apoptosis), and beta cell proliferation. Nutrients, hormones and growth factors coupled with their signalling intermediates have been...... suggested to play a role in beta cell mass regulation. In addition, genetic mouse model studies have indicated that cyclins and cyclin-dependent kinases that determine cell cycle progression are involved in beta cell replication, and more recently, menin in association with cyclin-dependent kinase...... inhibitors has been demonstrated to be important in beta cell growth. In this review, we consider and highlight some aspects of cell cycle regulation in relation to beta cell replication. The role of cell cycle regulation in beta cell replication is mostly from studies in rodent models, but whether...

  11. Quantitative measurement of changes in adhesion force involving focal adhesion kinase during cell attachment, spread, and migration

    International Nuclear Information System (INIS)

    Wu, C.-C.; Su, H.-W.; Lee, C.-C.; Tang, M.-J.; Su, F.-C.

    2005-01-01

    Focal adhesion kinase (FAK) is a critical protein for the regulation of integrin-mediated cellular functions and it can enhance cell motility in Madin-Darby canine kidney (MDCK) cells by hepatocyte growth factor (HGF) induction. We utilized optical trapping and cytodetachment techniques to measure the adhesion force between pico-Newton and nano-Newton (nN) for quantitatively investigating the effects of FAK on adhesion force during initial binding (5 s), beginning of spreading (30 min), spreadout (12 h), and migration (induced by HGF) in MDCK cells with overexpressed FAK (FAK-WT), FAK-related non-kinase (FRNK), as well as normal control cells. Optical tweezers was used to measure the initial binding force between a trapped cell and glass coverslide or between a trapped bead and a seeded cell. In cytodetachment, the commercial atomic force microscope probe with an appropriate spring constant was used as a cyto-detacher to evaluate the change of adhesion force between different FAK expression levels of cells in spreading, spreadout, and migrating status. The results demonstrated that FAK-WT significantly increased the adhesion forces as compared to FRNK cells throughout all the different stages of cell adhesion. For cells in HGF-induced migration, the adhesion force decreased to almost the same level (∼600 nN) regardless of FAK levels indicating that FAK facilitates cells to undergo migration by reducing the adhesion force. Our results suggest FAK plays a role of enhancing cell adhesive ability in the binding and spreading, but an appropriate level of adhesion force is required for HGF-induced cell migration

  12. Graphene as transparent and current spreading electrode in silicon solar cell

    Energy Technology Data Exchange (ETDEWEB)

    Behura, Sanjay K., E-mail: sanjaybehura@gmail.com; Nayak, Sasmita; Jani, Omkar [Solar Energy Research Wing, Gujarat Energy Research and Management Institute - Research, Innovation and Incubation Centre, Gandhinagar 382007, Gujarat (India); Mahala, Pramila [School of Solar Energy, Pandit Deendayal Petroleum University, Gandhinagar 382007, Gujarat (India)

    2014-11-15

    Fabricated bi-layer graphene (BLG) has been studied as transparent and current spreading electrode (TCSE) for silicon solar cell, using TCAD-Silvaco 2D simulation. We have carried out comparative study using both Ag grids and BLG as current spreading electrode (CSE) and TCSE, respectively. Our study reveals that BLG based solar cell shows better efficiency of 24.85% than Ag-based cell (21.44%), in all of the critical aspects, including generation rate, recombination rate, electric field, potential and quantum efficiency. Further BLG based cell exhibits pronounce rectifying behavior, low saturation current, and good turn-on voltage while studying in dark.

  13. Graphene as transparent and current spreading electrode in silicon solar cell

    Directory of Open Access Journals (Sweden)

    Sanjay K. Behura

    2014-11-01

    Full Text Available Fabricated bi-layer graphene (BLG has been studied as transparent and current spreading electrode (TCSE for silicon solar cell, using TCAD-Silvaco 2D simulation. We have carried out comparative study using both Ag grids and BLG as current spreading electrode (CSE and TCSE, respectively. Our study reveals that BLG based solar cell shows better efficiency of 24.85% than Ag-based cell (21.44%, in all of the critical aspects, including generation rate, recombination rate, electric field, potential and quantum efficiency. Further BLG based cell exhibits pronounce rectifying behavior, low saturation current, and good turn-on voltage while studying in dark.

  14. Cell spreading on titanium dioxide film formed and modified with aerosol beam and femtosecond laser

    Energy Technology Data Exchange (ETDEWEB)

    Shinonaga, Togo, E-mail: togo@jwri.osaka-u.ac.jp [Graduate School of Engineering, Osaka University, 1-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Tsukamoto, Masahiro [Joining and Welding Research Institute, Osaka University, 11-1 Mihogaoka, Ibaraki, Osaka 567-0047 (Japan); Nagai, Akiko; Yamashita, Kimihiro; Hanawa, Takao [Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, 2-3-10 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-0062 (Japan); Matsushita, Nobuhiro [Materials and Structures Laboratory, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8503 (Japan); Xie, Guoqiang [Institute for Materials Research, Tohoku University, 2-1-1 Karahira, Aoba-ku, Sendai, Miyagi 980-8577 (Japan); Abe, Nobuyuki [Joining and Welding Research Institute, Osaka University, 11-1 Mihogaoka, Ibaraki, Osaka 567-0047 (Japan)

    2014-01-01

    Titanium (Ti) is widely used in biomaterials because of its excellent anti-corrosion properties and high strength. However, Ti has no biological function, so its bioactivity must be improved. Coating a titanium dioxide (TiO{sub 2}) film on a Ti plate surface has been shown to improve the biocompatibility of Ti plates. If periodic nanostructures were formed on the film surface, the direction of cell spreading might be controlled by the direction of the grooves. Controlling cell spreading on biomaterials would contribute to the creation of advanced biomaterials. In this paper, a TiO{sub 2} film was formed on a Ti plate with an aerosol beam composed of sub micron-sized TiO{sub 2} particles and helium gas. Periodic nanostructures, lying perpendicular to the laser electric field polarization vector, were formed on the film by scanning the femtosecond laser focusing spot. The period and height of the periodic nanostructures were about 230 nm and 150 nm, respectively. In a cell test, cell spreading was observed along the grooves of the periodic nanostructures; in contrast, cell spreading did not show a definite direction on TiO{sub 2} a film without periodic nanostructures. These results suggest that the direction of cell spreading on the film can be controlled by periodic nanostructure formation generated using a femtosecond laser.

  15. Cell spreading on titanium dioxide film formed and modified with aerosol beam and femtosecond laser

    International Nuclear Information System (INIS)

    Shinonaga, Togo; Tsukamoto, Masahiro; Nagai, Akiko; Yamashita, Kimihiro; Hanawa, Takao; Matsushita, Nobuhiro; Xie, Guoqiang; Abe, Nobuyuki

    2014-01-01

    Titanium (Ti) is widely used in biomaterials because of its excellent anti-corrosion properties and high strength. However, Ti has no biological function, so its bioactivity must be improved. Coating a titanium dioxide (TiO 2 ) film on a Ti plate surface has been shown to improve the biocompatibility of Ti plates. If periodic nanostructures were formed on the film surface, the direction of cell spreading might be controlled by the direction of the grooves. Controlling cell spreading on biomaterials would contribute to the creation of advanced biomaterials. In this paper, a TiO 2 film was formed on a Ti plate with an aerosol beam composed of sub micron-sized TiO 2 particles and helium gas. Periodic nanostructures, lying perpendicular to the laser electric field polarization vector, were formed on the film by scanning the femtosecond laser focusing spot. The period and height of the periodic nanostructures were about 230 nm and 150 nm, respectively. In a cell test, cell spreading was observed along the grooves of the periodic nanostructures; in contrast, cell spreading did not show a definite direction on TiO 2 a film without periodic nanostructures. These results suggest that the direction of cell spreading on the film can be controlled by periodic nanostructure formation generated using a femtosecond laser.

  16. Heparanase facilitates cell adhesion and spreading by clustering of cell surface heparan sulfate proteoglycans.

    Directory of Open Access Journals (Sweden)

    Flonia Levy-Adam

    2008-06-01

    Full Text Available Heparanase is a heparan sulfate (HS degrading endoglycosidase participating in extracellular matrix degradation and remodeling. Apart of its well characterized enzymatic activity, heparanase was noted to exert also enzymatic-independent functions. Non-enzymatic activities of heparanase include enhanced adhesion of tumor-derived cells and primary T-cells. Attempting to identify functional domains of heparanase that would serve as targets for drug development, we have identified heparin binding domains of heparanase. A corresponding peptide (residues Lys(158-Asp(171, termed KKDC was demonstrated to physically associate with heparin and HS, and to inhibit heparanase enzymatic activity. We hypothesized that the pro-adhesive properties of heparanase are mediated by its interaction with cell surface HS proteoglycans, and utilized the KKDC peptide to examine this possibility. We provide evidence that the KKDC peptide interacts with cell membrane HS, resulting in clustering of syndecan-1 and syndecan-4. We applied classical analysis of cell morphology, fluorescent and time-lapse microscopy and demonstrated that the KKDC peptide efficiently stimulates the adhesion and spreading of various cell types, mediated by PKC, Src, and the small GTPase Rac1. These results support, and further substantiate the notion that heparanase function is not limited to its enzymatic activity.

  17. Regulation of cell wall biosynthesis.

    Science.gov (United States)

    Zhong, Ruiqin; Ye, Zheng-Hua

    2007-12-01

    Plant cell walls differ in their amount and composition among various cell types and even in different microdomains of the wall of a given cell. Plants must have evolved regulatory mechanisms controlling biosynthesis, targeted secretion, and assembly of wall components to achieve the heterogeneity in cell walls. A number of factors, including hormones, the cytoskeleton, glycosylphosphatidylinositol-anchored proteins, phosphoinositides, and sugar nucleotide supply, have been implicated in the regulation of cell wall biosynthesis or deposition. In the past two years, there have been important discoveries in transcriptional regulation of secondary wall biosynthesis. Several transcription factors in the NAC and MYB families have been shown to be the key switches for activation of secondary wall biosynthesis. These studies suggest a transcriptional network comprised of a hierarchy of transcription factors is involved in regulating secondary wall biosynthesis. Further investigation and integration of the regulatory players participating in the making of cell walls will certainly lead to our understanding of how wall amounts and composition are controlled in a given cell type. This may eventually allow custom design of plant cell walls on the basis of our needs.

  18. Cell Based GIS as Cellular Automata for Disaster Spreading Predictions and Required Data Systems

    Directory of Open Access Journals (Sweden)

    Kohei Arai

    2013-03-01

    Full Text Available A method for prediction and simulation based on the Cell Based Geographic Information System(GIS as Cellular Automata (CA is proposed together with required data systems, in particular metasearch engine usage in an unified way. It is confirmed that the proposed cell based GIS as CA has flexible usage of the attribute information that is attached to the cell in concert with location information and does work for disaster spreading simulation and prediction.

  19. Regulators of Tfh cell differentiation

    Directory of Open Access Journals (Sweden)

    Gajendra Motiram Jogdand

    2016-11-01

    Full Text Available The follicular helper T (Tfh cells help is critical for activation of B cells, antibody class switching and germinal center formation. The Tfh cells are characterized by the expression of CXCR5, ICOS, PD-1, Bcl-6, and IL-21. They are involved in clearing infections and are adversely linked with autoimmune diseases and also have a role in viral replication as well as clearance. Tfh cells are generated from naïve CD4 T cells with sequential steps involving cytokine signaling (IL-21, IL-6, IL-12, activin A, migration and positioning in the germinal center by CXCR5, surface receptors (ICOS/ICOSL, SAP/SLAM as well as transcription factor (Bcl-6, c-Maf, STAT3 signaling and repressor miR155. On the other hand Tfh generation is negatively regulated at specific steps of Tfh generation by specific cytokine (IL-2, IL-7, surface receptor (PD-1, CTLA-4, transcription factors Blimp-1, STAT5, T-bet, KLF-2 signaling and repressor miR 146a. Interestingly, miR 17-92 and FOXO1 acts as a positive as well as a negative regulator of Tfh differentiation depending on the time of expression and disease specificity. Tfh cells are also generated from the conversion of other effector T cells as exemplified by Th1 cells converting into Tfh during viral infection. The mechanistic details of effector T cells conversion into Tfh are yet to be clear. To manipulate Tfh cells for therapeutic implication and or for effective vaccination strategies, it is important to know positive and negative regulators of Tfh generation. Hence, in this review we have highlighted and interlinked molecular signaling from cytokines, surface receptors, transcription factors, ubiquitin Ligase and miRNA as positive and negative regulators for Tfh differentiation.

  20. Spreading of a prion domain from cell-to-cell by vesicular transport in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Carmen I Nussbaum-Krammer

    2013-03-01

    Full Text Available Prion proteins can adopt self-propagating alternative conformations that account for the infectious nature of transmissible spongiform encephalopathies (TSEs and the epigenetic inheritance of certain traits in yeast. Recent evidence suggests a similar propagation of misfolded proteins in the spreading of pathology of neurodegenerative diseases including Alzheimer's or Parkinson's disease. Currently there is only a limited number of animal model systems available to study the mechanisms that underlie the cell-to-cell transmission of aggregation-prone proteins. Here, we have established a new metazoan model in Caenorhabditis elegans expressing the prion domain NM of the cytosolic yeast prion protein Sup35, in which aggregation and toxicity are dependent upon the length of oligopeptide repeats in the glutamine/asparagine (Q/N-rich N-terminus. NM forms multiple classes of highly toxic aggregate species and co-localizes to autophagy-related vesicles that transport the prion domain from the site of expression to adjacent tissues. This is associated with a profound cell autonomous and cell non-autonomous disruption of mitochondrial integrity, embryonic and larval arrest, developmental delay, widespread tissue defects, and loss of organismal proteostasis. Our results reveal that the Sup35 prion domain exhibits prion-like properties when expressed in the multicellular organism C. elegans and adapts to different requirements for propagation that involve the autophagy-lysosome pathway to transmit cytosolic aggregation-prone proteins between tissues.

  1. Vinculin promotes cell spreading by mechanically coupling integrins to the cytoskeleton

    Science.gov (United States)

    Ezzell, R. M.; Goldmann, W. H.; Wang, N.; Parasharama, N.; Ingber, D. E.

    1997-01-01

    Mouse F9 embryonic carcinoma 5.51 cells that lack the cytoskeletal protein vinculin spread poorly on extracellular matrix compared with wild-type F9 cells or two vinculin-transfected clones (5.51Vin3 and Vin4; Samuels et al., 1993, J. Cell Biol. 121, 909-921). In the present study, we used this model system to determine how the presence of vinculin promotes cytoskeletal alterations and associated changes in cell shape. Microscopic analysis of cell spreading at early times, revealed that 5.51 cells retained the ability to form filopodia; however, they could not form lamellipodia, assemble stress fibers, or efficiently spread over the culture substrate. Detergent (Triton X-100) studies revealed that these major differences in cell morphology and cytoskeletal organization did not result from differences in levels of total polymerized or cross-linked actin. Biochemical studies showed that 5.51 cells, in addition to lacking vinculin, exhibited slightly reduced levels of alpha-actinin and paxillin in their detergent-insoluble cytoskeleton. The absence of vinculin correlated with a decrease in the mechanical stiffness of the integrin-cytoskeleton linkage, as measured using cell magnetometry. Furthermore, when vinculin was replaced by transfection in 5.51Vin3 and 5.51Vin4 cells, the levels of cytoskeletal-associated alpha-actinin and paxillin, the efficiency of transmembrane mechanical coupling, and the formation of actin stress fibers were all restored to near wild-type levels. These findings suggest that vinculin may promote cell spreading by stabilizing focal adhesions and transferring mechanical stresses that drive cytoskeletal remodeling, rather than by altering the total level of actin polymerization or cross-linking.

  2. A Role for Human Skin Mast Cells in Dengue Virus Infection and Systemic Spread.

    Science.gov (United States)

    Troupin, Andrea; Shirley, Devon; Londono-Renteria, Berlin; Watson, Alan M; McHale, Cody; Hall, Alex; Hartstone-Rose, Adam; Klimstra, William B; Gomez, Gregorio; Colpitts, Tonya M

    2016-12-01

    Dengue virus (DENV) is a mosquito-borne flavivirus that causes serious global human disease and mortality. Skin immune cells are an important component of initial DENV infection and systemic spread. Here, we show that mast cells are a target of DENV in human skin and that DENV infection of skin mast cells induces degranulation and alters cytokine and growth factor expression profiles. Importantly, to our knowledge, we also demonstrate for the first time that DENV localizes within secretory granules in infected skin mast cells. In addition, DENV within extracellular granules was infectious in vitro and in vivo, trafficking through lymph to draining lymph nodes in mice. We demonstrate an important role for human skin mast cells in DENV infection and identify a novel mechanism for systemic spread of DENV infection from the initial peripheral mosquito injection site. Copyright © 2016 by The American Association of Immunologists, Inc.

  3. The HSV-1 mechanisms of cell-to-cell spread and fusion are critically dependent on host PTP1B.

    Directory of Open Access Journals (Sweden)

    Jillian C Carmichael

    2018-05-01

    Full Text Available All herpesviruses have mechanisms for passing through cell junctions, which exclude neutralizing antibodies and offer a clear path to neighboring, uninfected cells. In the case of herpes simplex virus type 1 (HSV-1, direct cell-to-cell transmission takes place between epithelial cells and sensory neurons, where latency is established. The spreading mechanism is poorly understood, but mutations in four different HSV-1 genes can dysregulate it, causing neighboring cells to fuse to produce syncytia. Because the host proteins involved are largely unknown (other than the virus entry receptor, we were intrigued by an earlier discovery that cells infected with wild-type HSV-1 will form syncytia when treated with salubrinal. A biotinylated derivative of this drug was used to pull down cellular complexes, which were analyzed by mass spectrometry. One candidate was a protein tyrosine phosphatase (PTP1B, and although it ultimately proved not to be the target of salubrinal, it was found to be critical for the mechanism of cell-to-cell spread. In particular, a highly specific inhibitor of PTP1B (CAS 765317-72-4 blocked salubrinal-induced fusion, and by itself resulted in a dramatic reduction in the ability of HSV-1 to spread in the presence of neutralizing antibodies. The importance of this phosphatase was confirmed in the absence of drugs by using PTP1B-/- cells. Importantly, replication assays showed that virus titers were unaffected when PTP1B was inhibited or absent. Only cell-to-cell spread was altered. We also examined the effects of salubrinal and the PTP1B inhibitor on the four Syn mutants of HSV-1, and strikingly different responses were found. That is, both drugs individually enhanced fusion for some mutants and reduced fusion for others. PTP1B is the first host factor identified to be specifically required for cell-to-cell spread, and it may be a therapeutic target for preventing HSV-1 reactivation disease.

  4. The HSV-1 mechanisms of cell-to-cell spread and fusion are critically dependent on host PTP1B.

    Science.gov (United States)

    Carmichael, Jillian C; Yokota, Hiroki; Craven, Rebecca C; Schmitt, Anthony; Wills, John W

    2018-05-01

    All herpesviruses have mechanisms for passing through cell junctions, which exclude neutralizing antibodies and offer a clear path to neighboring, uninfected cells. In the case of herpes simplex virus type 1 (HSV-1), direct cell-to-cell transmission takes place between epithelial cells and sensory neurons, where latency is established. The spreading mechanism is poorly understood, but mutations in four different HSV-1 genes can dysregulate it, causing neighboring cells to fuse to produce syncytia. Because the host proteins involved are largely unknown (other than the virus entry receptor), we were intrigued by an earlier discovery that cells infected with wild-type HSV-1 will form syncytia when treated with salubrinal. A biotinylated derivative of this drug was used to pull down cellular complexes, which were analyzed by mass spectrometry. One candidate was a protein tyrosine phosphatase (PTP1B), and although it ultimately proved not to be the target of salubrinal, it was found to be critical for the mechanism of cell-to-cell spread. In particular, a highly specific inhibitor of PTP1B (CAS 765317-72-4) blocked salubrinal-induced fusion, and by itself resulted in a dramatic reduction in the ability of HSV-1 to spread in the presence of neutralizing antibodies. The importance of this phosphatase was confirmed in the absence of drugs by using PTP1B-/- cells. Importantly, replication assays showed that virus titers were unaffected when PTP1B was inhibited or absent. Only cell-to-cell spread was altered. We also examined the effects of salubrinal and the PTP1B inhibitor on the four Syn mutants of HSV-1, and strikingly different responses were found. That is, both drugs individually enhanced fusion for some mutants and reduced fusion for others. PTP1B is the first host factor identified to be specifically required for cell-to-cell spread, and it may be a therapeutic target for preventing HSV-1 reactivation disease.

  5. Time-series observation of the spreading out of microvessel endothelial cells with atomic force microscopy

    International Nuclear Information System (INIS)

    Han Dong; Ma Wanyun; Liao Fulong; Yeh Meiling; Ouyang Zhigang; Sun Yunxu

    2003-01-01

    The spreading out of microvessel endothelial cells plays a key role in angiogenesis and the post-injury healing of endothelial cells. In our study, a physical force applied with an atomic force microscopic (AFM) cantilever tip in contact mode partly broke the peripheral adhesion that just-confluent cultured rat cerebral microvessel endothelial cells had formed with basal structures and resulted in the cells actively withdrawing from the stimulated area. Time-series changes in cell extension were imaged using tapping mode AFM, in conjunction with total internal reflection fluorescence microscopy, intensified charge-coupled device and field emission scanning electron microscopy. We also interpreted phase images of living endothelial cells. The results showed that formation of a fibronectin molecule monolayer is key to the spreading out of the cells. Lamellipods as well as filopods would spread out in temporal and spatial distribution following the formation of fibronectin layer. In addition, a lattice-like meshwork of filopods formed in the regions leading lamellipods, which would possibly provide a fulcrum for the filaments of the cytoskeleton within the leading cell body periphery

  6. Microfluidic high viability neural cell separation using viscoelastically tuned hydrodynamic spreading

    DEFF Research Database (Denmark)

    Wu, Zhigang; Hjort, Klas; Wicher, Grzegorz

    2008-01-01

    polymer solution of alginic sodium, the spreading behavior was investigated at different polymer concentrations and flow rates. Particle separation was studied in the same detail for 9.9 microm and 1.9 microm latex beads. Using buffered aqueous solutions and further surface treatments to protect from cell...

  7. Establishment and characterization of a new cell line derived from human colorectal laterally spreading tumor

    Science.gov (United States)

    Wang, Xin-Ying; Lai, Zhou-Sheng; Yeung, Chung-Man; Wang, Ji-De; Deng, Wen; Li, Hoi Yee; Han, Yu-Jing; Kung, Hsiang-Fu; Jiang, Bo; Lin, Marie Chia-mi

    2008-01-01

    AIM: To study the molecular mechanism of laterally spreading tumor (LST), a cell line [Laterally Spreading Tumor-Rectum 1 (LST-R1)] was derived and the characteristics of this cell line were investigated. METHODS: A new cell line (LST-R1) originated from laterally spreading tumor was established. Properties of the cell line were characterized using scanning and transmission electron microscopy, immunohistochemistry method, cytogenetic analysis and nude mice xenograft experiments. In vitro invasion assay, cDNA microarray and Western blotting were used to compare the difference between the LST-R1 and other colorectal cancer cell lines derived from prudent colon cancer. RESULTS: Our study demonstrated that both epithelial special antigen (ESA) and cytokeratin-20 (CK20) were expressed in LST-R1. The cells presented microvilli and tight junction with large nuclei. The karyotypic analysis showed hyperdiploid features with structural chromosome aberrations. The in vivo tumorigenicity was also demonstrated in nude mice xenograft experiments. The invasion assay suggested this cell line has a higher invasive ability. cDNA microarray and Western blotting show the loss of the expression of E-cadherin in LST-R1 cells. CONCLUSION: We established and characterized a colorectal cancer cell line, LST-R1 and LST-R1 has an obvious malignant tendency, which maybe partially attributed to the changes of the expression of some adhesion molecules, such as E-cadherin. It is also a versatile tool for exploring the original and progressive mechanisms of laterally spreading tumor and the early colon cancer genesis. PMID:18300345

  8. TiO{sub 2}-enriched polymeric powder coatings support human mesenchymal cell spreading and osteogenic differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Mozumder, Mohammad Sayem; Zhu, Jesse [Department of Chemical and Biochemical Engineering, University of Western Ontario, London, Ontario, N6A 5B9 (Canada); Perinpanayagam, Hiran, E-mail: Hiran.Perinpanayagam@schulich.uwo.ca [Schulich School of Medicine and Dentistry, University of Western Ontario, London, Ontario, N6A 5C1 (Canada)

    2011-06-15

    Novel polymeric powder coatings (PPC) were prepared by ultrafine powder coating technology and shown to support human mesenchymal cell attachment and growth. PPC surfaces enriched with nano-TiO{sub 2} (nTiO{sub 2}) showed enhanced cellular responses, and were compared to commercially pure titanium (cpTi). After cell attachment and growth, osteogenic differentiation and bone matrix formation ensures osseointegration for implantable biomaterials. Therefore, the objective of this study was to determine if mesenchymal cells grown on PPC could undergo osteogenic differentiation by inducing Runx2 and bone matrix proteins, and then initiate mineralization. Atomic force microscopy revealed intricate three-dimensional micro-topographies, and the measures of nano-roughness and porosity were similar for all PPC surfaces. Scanning electron microscopy showed that the cells attached and spread out over all of the surfaces. After 1 week in osteogenic media, RT-PCR analysis showed the induction of Runx2, the up-regulation of type I collagen, and the initial detection of alkaline phosphatase and bone sialoprotein. After 4 weeks, Alizarin Red staining showed mineral deposition. However, cell spreading and osteogenic differentiation were significantly (P < 0.05) higher on the cpTi controls than on the PPC surfaces. Furthermore, spreading and differentiation were consistently higher on the titanium-enriched PPC-2, -3 and -4 than on the titanium-free PPC-1. Therefore, despite the presence of complex micro-topographies and nano-features, titanium-enrichment enhanced the cellular response, and pure titanium still provided the best substrate. These findings confirm the cytocompatibility of these novel polymeric coatings and suggest that titanium-enrichment and nTiO{sub 2} additives may enhance their performance.

  9. Effect of chondroitin sulfate proteoglycans on neuronal cell adhesion, spreading and neurite growth in culture

    Directory of Open Access Journals (Sweden)

    Jingyu Jin

    2018-01-01

    Full Text Available As one major component of extracellular matrix (ECM in the central nervous system, chondroitin sulfate proteoglycans (CSPGs have long been known as inhibitors enriched in the glial scar that prevent axon regeneration after injury. Although many studies have shown that CSPGs inhibited neurite outgrowth in vitro using different types of neurons, the mechanism by which CSPGs inhibit axonal growth remains poorly understood. Using cerebellar granule neuron (CGN culture, in this study, we evaluated the effects of different concentrations of both immobilized and soluble CSPGs on neuronal growth, including cell adhesion, spreading and neurite growth. Neurite length decreased while CSPGs concentration arised, meanwhile, a decrease in cell density accompanied by an increase in cell aggregates formation was observed. Soluble CSPGs also showed an inhibition on neurite outgrowth, but it required a higher concentration to induce cell aggregates formation than coated CSPGs. We also found that growth cone size was significantly reduced on CSPGs and neuronal cell spreading was restrained by CSPGs, attributing to an inhibition on lamellipodial extension. The effect of CSPGs on neuron adhesion was further evidenced by interference reflection microscopy (IRM which directly demonstrated that both CGNs and cerebral cortical neurons were more loosely adherent to a CSPG substrate. These data demonstrate that CSPGs have an effect on cell adhesion and spreading in addition to neurite outgrowth.

  10. HCMV spread and cell tropism are determined by distinct virus populations.

    Directory of Open Access Journals (Sweden)

    Laura Scrivano

    Full Text Available Human cytomegalovirus (HCMV can infect many different cell types in vivo. Two gH/gL complexes are used for entry into cells. gH/gL/pUL(128,130,131A shows no selectivity for its host cell, whereas formation of a gH/gL/gO complex only restricts the tropism mainly to fibroblasts. Here, we describe that depending on the cell type in which virus replication takes place, virus carrying the gH/gL/pUL(128,130,131A complex is either released or retained cell-associated. We observed that virus spread in fibroblast cultures was predominantly supernatant-driven, whereas spread in endothelial cell (EC cultures was predominantly focal. This was due to properties of virus released from fibroblasts and EC. Fibroblasts released virus which could infect both fibroblasts and EC. In contrast, EC released virus which readily infected fibroblasts, but was barely able to infect EC. The EC infection capacities of virus released from fibroblasts or EC correlated with respectively high or low amounts of gH/gL/pUL(128,130,131A in virus particles. Moreover, we found that focal spread in EC cultures could be attributed to EC-tropic virus tightly associated with EC and not released into the supernatant. Preincubation of fibroblast-derived virus progeny with EC or beads coated with pUL131A-specific antibodies depleted the fraction that could infect EC, and left a fraction that could predominantly infect fibroblasts. These data strongly suggest that HCMV progeny is composed of distinct virus populations. EC specifically retain the EC-tropic population, whereas fibroblasts release EC-tropic and non EC-tropic virus. Our findings offer completely new views on how HCMV spread may be controlled by its host cells.

  11. An Optimized Injectable Hydrogel Scaffold Supports Human Dental Pulp Stem Cell Viability and Spreading

    Directory of Open Access Journals (Sweden)

    T. D. Jones

    2016-01-01

    Full Text Available Introduction. HyStem-C™ is a commercially available injectable hydrogel composed of polyethylene glycol diacrylate (PEGDA, hyaluronan (HA, and gelatin (Gn. These components can be mechanically tuned to enhance cell viability and spreading. Methods. The concentration of PEGDA with an added disulfide bond (PEGSSDA was varied from 0.5 to 8.0% (w/v to determine the optimal concentration for injectable clinical application. We evaluated the cell viability of human dental pulp stem cells (hDPSCs embedded in 2% (w/v PEGSSDA-HA-Gn hydrogels. Volume ratios of HA : Gn from 100 : 0 to 25 : 75 were varied to encourage hDPSC spreading. Fibronectin (Fn was added to our model to determine the effect of extracellular matrix protein concentration on hDPSC behavior. Results. Our preliminary data suggests that the hydrogel gelation time decreased as the PEGSSDA cross-linker concentration increased. The PEGSSDA-HA-Gn was biocompatible with hDPSCs, and increased ratios of HA : Gn enhanced cell viability for 14 days. Additionally, cell proliferation with added fibronectin increased significantly over time at concentrations of 1.0 and 10.0 μg/mL in PEGDA-HA-Gn hydrogels, while cell spreading significantly increased at Fn concentrations of 0.1 μg/mL. Conclusions. This study demonstrates that PEG-based injectable hydrogels maintain hDPSC viability and facilitate cell spreading, mainly in the presence of extracellular matrix (ECM proteins.

  12. Spread of white-nose syndrome on a network regulated by geography and climate.

    Science.gov (United States)

    Maher, Sean P; Kramer, Andrew M; Pulliam, J Tomlin; Zokan, Marcus A; Bowden, Sarah E; Barton, Heather D; Magori, Krisztian; Drake, John M

    2012-01-01

    Wildlife and plant diseases can reduce biodiversity, disrupt ecosystem services and threaten human health. Emerging pathogens have displayed a variety of spatial spread patterns due to differences in host ecology, including diffusive spread from an epicentre (West Nile virus), jump dispersal on a network (foot-and-mouth disease), or a combination of these (Sudden oak death). White-nose syndrome is a highly pathogenic infectious disease of bats currently spreading across North America. Understanding how bat ecology influences this spread is crucial to management of infected and vulnerable populations. Here we show that white-nose syndrome spread is not diffusive but rather mediated by patchily distributed habitat and large-scale gradients in winter climate. Simulations predict rapid expansion and infection of most counties with caves in the contiguous United States by winter 2105-2106. Our findings show the unique pattern of white-nose syndrome spread corresponds to ecological traits of the host and suggest hypotheses for transmission mechanisms acting at the local scale.

  13. Cell swelling and volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay

    1992-01-01

    The extracellular space in the brain is typically 20% of the tissue volume and is reduced to at least half its size under conditions of neural insult. Whether there is a minimum size to the extracellular space was discussed. A general model for cell volume regulation was presented, followed...... by a discussion on how many of the generally involved mechanisms are identified in neural cells and (or) in astrocytes. There seems to be clear evidence suggesting that parallel K+ and Cl- channels mediate regulatory volume decrease in primary cultures of astrocytes, and a stretch-activated cation channel has...... been reported. The role of the different channels was discussed. A taurine leak pathway is clearly activated after cell swelling both in astrocytes and in neurones. The relations between the effect of glutamate and cell swelling were discussed. Discussion on the clearance of potassium from...

  14. Stress relaxing hyaluronic acid-collagen hydrogels promote cell spreading, fiber remodeling, and focal adhesion formation in 3D cell culture.

    Science.gov (United States)

    Lou, Junzhe; Stowers, Ryan; Nam, Sungmin; Xia, Yan; Chaudhuri, Ovijit

    2018-02-01

    The physical and architectural cues of the extracellular matrix (ECM) play a critical role in regulating important cellular functions such as spreading, migration, proliferation, and differentiation. Natural ECM is a complex viscoelastic scaffold composed of various distinct components that are often organized into a fibrillar microstructure. Hydrogels are frequently used as synthetic ECMs for 3D cell culture, but are typically elastic, due to covalent crosslinking, and non-fibrillar. Recent work has revealed the importance of stress relaxation in viscoelastic hydrogels in regulating biological processes such as spreading and differentiation, but these studies all utilize synthetic ECM hydrogels that are non-fibrillar. Key mechanotransduction events, such as focal adhesion formation, have only been observed in fibrillar networks in 3D culture to date. Here we present an interpenetrating network (IPN) hydrogel system based on HA crosslinked with dynamic covalent bonds and collagen I that captures the viscoelasticity and fibrillarity of ECM in tissues. The IPN hydrogels exhibit two distinct processes in stress relaxation, one from collagen and the other from HA crosslinking dynamics. Stress relaxation in the IPN hydrogels can be tuned by modulating HA crosslinker affinity, molecular weight of the HA, or HA concentration. Faster relaxation in the IPN hydrogels promotes cell spreading, fiber remodeling, and focal adhesion (FA) formation - behaviors often inhibited in other hydrogel-based materials in 3D culture. This study presents a new, broadly adaptable materials platform for mimicking key ECM features of viscoelasticity and fibrillarity in hydrogels for 3D cell culture and sheds light on how these mechanical and structural cues regulate cell behavior. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Cell-to-Cell Measles Virus Spread between Human Neurons Is Dependent on Hemagglutinin and Hyperfusogenic Fusion Protein.

    Science.gov (United States)

    Sato, Yuma; Watanabe, Shumpei; Fukuda, Yoshinari; Hashiguchi, Takao; Yanagi, Yusuke; Ohno, Shinji

    2018-03-15

    Measles virus (MV) usually causes acute infection but in rare cases persists in the brain, resulting in subacute sclerosing panencephalitis (SSPE). Since human neurons, an important target affected in the disease, do not express the known MV receptors (signaling lymphocyte activation molecule [SLAM] and nectin 4), how MV infects neurons and spreads between them is unknown. Recent studies have shown that many virus strains isolated from SSPE patients possess substitutions in the extracellular domain of the fusion (F) protein which confer enhanced fusion activity. Hyperfusogenic viruses with such mutations, unlike the wild-type MV, can induce cell-cell fusion even in SLAM- and nectin 4-negative cells and spread efficiently in human primary neurons and the brains of animal models. We show here that a hyperfusogenic mutant MV, IC323-F(T461I)-EGFP (IC323 with a fusion-enhancing T461I substitution in the F protein and expressing enhanced green fluorescent protein), but not the wild-type MV, spreads in differentiated NT2 cells, a widely used human neuron model. Confocal time-lapse imaging revealed the cell-to-cell spread of IC323-F(T461I)-EGFP between NT2 neurons without syncytium formation. The production of virus particles was strongly suppressed in NT2 neurons, also supporting cell-to-cell viral transmission. The spread of IC323-F(T461I)-EGFP was inhibited by a fusion inhibitor peptide as well as by some but not all of the anti-hemagglutinin antibodies which neutralize SLAM- or nectin-4-dependent MV infection, suggesting the presence of a distinct neuronal receptor. Our results indicate that MV spreads in a cell-to-cell manner between human neurons without causing syncytium formation and that the spread is dependent on the hyperfusogenic F protein, the hemagglutinin, and the putative neuronal receptor for MV. IMPORTANCE Measles virus (MV), in rare cases, persists in the human central nervous system (CNS) and causes subacute sclerosing panencephalitis (SSPE) several

  16. Cell intrinsic immunity spreads to bystander cells via the intercellular transfer of cGAMP.

    Science.gov (United States)

    Ablasser, Andrea; Schmid-Burgk, Jonathan L; Hemmerling, Inga; Horvath, Gabor L; Schmidt, Tobias; Latz, Eicke; Hornung, Veit

    2013-11-28

    The innate immune defence of multicellular organisms against microbial pathogens requires cellular collaboration. Information exchange allowing immune cells to collaborate is generally attributed to soluble protein factors secreted by pathogen-sensing cells. Cytokines, such as type I interferons (IFNs), serve to alert non-infected cells to the possibility of pathogen challenge. Moreover, in conjunction with chemokines they can instruct specialized immune cells to contain and eradicate microbial infection. Several receptors and signalling pathways exist that couple pathogen sensing to the induction of cytokines, whereas cytosolic recognition of nucleic acids seems to be exquisitely important for the activation of type I IFNs, master regulators of antiviral immunity. Cytosolic DNA is sensed by the receptor cyclic GMP-AMP (cGAMP) synthase (cGAS), which catalyses the synthesis of the second messenger cGAMP(2'-5'). This molecule in turn activates the endoplasmic reticulum (ER)-resident receptor STING, thereby inducing an antiviral state and the secretion of type I IFNs. Here we find in murine and human cells that cGAS-synthesized cGAMP(2'-5') is transferred from producing cells to neighbouring cells through gap junctions, where it promotes STING activation and thus antiviral immunity independently of type I IFN signalling. In line with the limited cargo specificity of connexins, the proteins that assemble gap junction channels, most connexins tested were able to confer this bystander immunity, thus indicating a broad physiological relevance of this local immune collaboration. Collectively, these observations identify cGAS-triggered cGAMP(2'-5') transfer as a novel host strategy that serves to rapidly convey antiviral immunity in a transcription-independent, horizontal manner.

  17. Controlling Gel Structure to Modulate Cell Adhesion and Spreading on the Surface of Microcapsules.

    Science.gov (United States)

    Zheng, Huizhen; Gao, Meng; Ren, Ying; Lou, Ruyun; Xie, Hongguo; Yu, Weiting; Liu, Xiudong; Ma, Xiaojun

    2016-08-03

    The surface properties of implanted materials or devices play critical roles in modulating cell behavior. However, the surface properties usually affect cell behaviors synergetically so that it is still difficult to separately investigate the influence of a single property on cell behavior in practical applications. In this study, alginate-chitosan (AC) microcapsules with a dense or loose gel structure were fabricated to understand the effect of gel structure on cell behavior. Cells preferentially adhered and spread on the loose gel structure microcapsules rather than on the dense ones. The two types of microcapsules exhibited nearly identical surface positive charges, roughness, stiffness, and hydrophilicity; thus, the result suggested that the gel structure was the principal factor affecting cell behavior. X-ray photoelectron spectroscopy analyses demonstrated that the overall percentage of positively charged amino groups was similar on both microcapsules. The different gel structures led to different states and distributions of the positively charged amino groups of chitosan, so we conclude that the loose gel structure facilitated greater cell adhesion and spreading mainly because more protonated amino groups remained unbound and exposed on the surface of these microcapsules.

  18. Smooth muscle cell rigidity and extracellular matrix organization influence endothelial cell spreading and adhesion formation in coculture.

    Science.gov (United States)

    Wallace, Charles S; Strike, Sophie A; Truskey, George A

    2007-09-01

    Efforts to develop functional tissue-engineered blood vessels have focused on improving the strength and mechanical properties of the vessel wall, while the functional status of the endothelium within these vessels has received less attention. Endothelial cell (EC) function is influenced by interactions between its basal surface and the underlying extracellular matrix. In this study, we utilized a coculture model of a tissue-engineered blood vessel to evaluate EC attachment, spreading, and adhesion formation to the extracellular matrix on the surface of quiescent smooth muscle cells (SMCs). ECs attached to and spread on SMCs primarily through the alpha(5)beta(1)-integrin complex, whereas ECs used either alpha(5)beta(1)- or alpha(v)beta(3)-integrin to spread on fibronectin (FN) adsorbed to plastic. ECs in coculture lacked focal adhesions, but EC alpha(5)beta(1)-integrin bound to fibrillar FN on the SMC surface, promoting rapid fibrillar adhesion formation. As assessed by both Western blot analysis and quantitative real-time RT-PCR, coculture suppressed the expression of focal adhesion proteins and mRNA, whereas tensin protein and mRNA expression were elevated. When attached to polyacrylamide gels with similar elastic moduli as SMCs, focal adhesion formation and the rate of cell spreading increased relative to ECs in coculture. Thus, the elastic properties are only one factor contributing to EC spreading and focal adhesion formation in coculture. The results suggest that the softness of the SMCs and the fibrillar organization of FN inhibit focal adhesions and reduce cell spreading while promoting fibrillar adhesion formation. These changes in the type of adhesions may alter EC signaling pathways in tissue-engineered blood vessels.

  19. The cysteine-rich domain of human ADAM 12 supports cell adhesion through syndecans and triggers signaling events that lead to beta1 integrin-dependent cell spreading

    DEFF Research Database (Denmark)

    Iba, K; Albrechtsen, R; Gilpin, B

    2000-01-01

    The ADAMs (a disintegrin and metalloprotease) family of proteins is involved in a variety of cellular interactions, including cell adhesion and ecto- domain shedding. Here we show that ADAM 12 binds to cell surface syndecans. Three forms of recombinant ADAM 12 were used in these experiments......-dependent manner attach to ADAM 12 via members of the syndecan family. After binding to syndecans, mesenchymal cells spread and form focal adhesions and actin stress fibers. Integrin beta1 was responsible for cell spreading because function-blocking monoclonal antibodies completely inhibited cell spreading......, and chondroblasts lacking beta1 integrin attached but did not spread. These data suggest that mesenchymal cells use syndecans as the initial receptor for the ADAM 12 cysteine-rich domain-mediated cell adhesion, and then the beta1 integrin to induce cell spreading. Interestingly, carcinoma cells attached but did...

  20. EMMPRIN regulates cytoskeleton reorganization and cell adhesion in prostate cancer.

    Science.gov (United States)

    Zhu, Haining; Zhao, Jun; Zhu, Beibei; Collazo, Joanne; Gal, Jozsef; Shi, Ping; Liu, Li; Ström, Anna-Lena; Lu, Xiaoning; McCann, Richard O; Toborek, Michal; Kyprianou, Natasha

    2012-01-01

    Proteins on cell surface play important roles during cancer progression and metastasis via their ability to mediate cell-to-cell interactions and navigate the communication between cells and the microenvironment. In this study a targeted proteomic analysis was conducted to identify the differential expression of cell surface proteins in human benign (BPH-1) versus malignant (LNCaP and PC-3) prostate epithelial cells. We identified EMMPRIN (extracellular matrix metalloproteinase inducer) as a key candidate and shRNA functional approaches were subsequently applied to determine the role of EMMPRIN in prostate cancer cell adhesion, migration, invasion as well as cytoskeleton organization. EMMPRIN was found to be highly expressed on the surface of prostate cancer cells compared to BPH-1 cells, consistent with a correlation between elevated EMMPRIN and metastasis found in other tumors. No significant changes in cell proliferation, cell cycle progression, or apoptosis were detected in EMMPRIN knockdown cells compared to the scramble controls. Furthermore, EMMPRIN silencing markedly decreased the ability of PC-3 cells to form filopodia, a critical feature of invasive behavior, while it increased expression of cell-cell adhesion and gap junction proteins. Our results suggest that EMMPRIN regulates cell adhesion, invasion, and cytoskeleton reorganization in prostate cancer cells. This study identifies a new function for EMMPRIN as a contributor to prostate cancer cell-cell communication and cytoskeleton changes towards metastatic spread, and suggests its potential value as a marker of prostate cancer progression to metastasis. Copyright © 2011 Wiley Periodicals, Inc.

  1. Ion Channels Involved in Cell Volume Regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay

    2011-01-01

    regulatory ion channels involved, and the mechanisms (cellular signalling pathways) that regulate these channels. Finally, I shall also briefly review current investigations in these two cell lines that focuses on how changes in cell volume can regulate cell functions such as cell migration, proliferation......This mini review outlines studies of cell volume regulation in two closely related mammalian cell lines: nonadherent Ehrlich ascites tumour cells (EATC) and adherent Ehrlich Lettre ascites (ELA) cells. Focus is on the regulatory volume decrease (RVD) that occurs after cell swelling, the volume...

  2. Cell adhesion and spreading on polymer surfaces micropatterned by ion beams

    International Nuclear Information System (INIS)

    Satriano, C.; Carnazza, S.; Licciardello, A.; Guglielmino, S.; Marletta, G.

    2003-01-01

    The cell adhesion and spreading behavior on surfaces of poly(ethyleneterephtalate) and poly(hydroxymethylsiloxane) micropatterned by focused 15 keV Ga + beams has been studied. It has been found that while no modification in the cell adhesion process could be observed for unirradiated and irradiated areas on the patterned surfaces, in the case of polyhydroxymethylsiloxane the cell adhesion process is basically confined within the irradiated areas and a clear dependence of the cell ordering on the lateral size of the irradiated areas is observed. The results are discussed in terms of the specific spatially resolved chemical modification induced by Ga + irradiation onto the two different polymers. Thus, the irradiation-induced modification of composition, functional groups concentration, surface free energy, and nanoscale morphology have been studied by means of x-ray photoelectron spectroscopy, time-of-flight secondary ion mass spectrometry, contact angle, and atomic force microscopy. The cell adhesion and spreading behavior was found to nicely correlate with the increase of the acid-base component γ AB of the surface free energy and more particularly with the dramatic increase of the Lewis basic electron-donor term

  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. The Role of Mesenchymal Stem Cells in Promoting Ovarian Cancer Growth and Spread

    Science.gov (United States)

    2014-12-01

    Immune Response Arm Cells MSC effects Innate Dendritic Cells (APC) Inhibition of maturation (CD80/86 expression) by STAT3 and IL10 (Beyth...are better understood. It is known that once MDSCs are 9 activated, they accumulate in lymphoid organs and tumors where they exert specific T cell 10...expressed on leukocyte 32 subsets and non-immune cells and may regulate important aspects of innate and adaptive 33 immune responses (Mempel, Voelcker et al

  5. Quantifying rates of cell migration and cell proliferation in co-culture barrier assays reveals how skin and melanoma cells interact during melanoma spreading and invasion.

    Science.gov (United States)

    Haridas, Parvathi; Penington, Catherine J; McGovern, Jacqui A; McElwain, D L Sean; Simpson, Matthew J

    2017-06-21

    Malignant spreading involves the migration of cancer cells amongst other native cell types. For example, in vivo melanoma invasion involves individual melanoma cells migrating through native skin, which is composed of several distinct subpopulations of cells. Here, we aim to quantify how interactions between melanoma and fibroblast cells affect the collective spreading of a heterogeneous population of these cells in vitro. We perform a suite of circular barrier assays that includes: (i) monoculture assays with fibroblast cells; (ii) monoculture assays with SK-MEL-28 melanoma cells; and (iii) a series of co-culture assays initiated with three different ratios of SK-MEL-28 melanoma cells and fibroblast cells. Using immunostaining, detailed cell density histograms are constructed to illustrate how the two subpopulations of cells are spatially arranged within the spreading heterogeneous population. Calibrating the solution of a continuum partial differential equation to the experimental results from the monoculture assays allows us to estimate the cell diffusivity and the cell proliferation rate for the melanoma and the fibroblast cells, separately. Using the parameter estimates from the monoculture assays, we then make a prediction of the spatial spreading in the co-culture assays. Results show that the parameter estimates obtained from the monoculture assays lead to a reasonably accurate prediction of the spatial arrangement of the two subpopulations in the co-culture assays. Overall, the spatial pattern of spreading of the melanoma cells and the fibroblast cells is very similar in monoculture and co-culture conditions. Therefore, we find no clear evidence of any interactions other than cell-to-cell contact and crowding effects. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Performance Analysis of Cell-Phone Worm Spreading in Cellular Networks through Opportunistic Communications

    Directory of Open Access Journals (Sweden)

    YAHUI, W.

    2012-05-01

    Full Text Available Worms spreading directly between cell-phones over short-range radio (Bluetooth, WiFi, etc. are increasing rapidly. Communication by these technologies is opportunistic and has very close relation with the social characteristics of the phone carriers. In this paper, we try to evaluate the impact of different characteristics on the spreading performance of worms. On the other hand, the behaviors of worms may have certain impact, too. For example, worms may make phones be completely dysfunctional and these phones can be seen as killed. We study the impact of the killing speed. Using the Markov model, we propose some theoretical models to evaluate the spreading performance in different cases. Simulation results show the accuracy of our models. Numerical results show that if users do not believe the data coming from others easily, the worms may bring less damage. Surprisingly, if the users are more willing to install the anti-virus software, the worms may bring bigger damage when the software becomes to be outdated with high probability. Though the worms can bring big damage on the network temporarily by killing phones rapidly, numerical results show that this behavior may decrease the total damage in the long time. Therefore, killing nodes more rapidly may be not optimal for worms.

  7. Simultaneous quantification of oxidative stress and cell spreading using 5-(and-6)-chloromethyl-2',7'-dichlorofluorescein.

    NARCIS (Netherlands)

    Koopman, W.J.H.; Verkaart, S.A.J.; Emst-de Vries, S.E. van; Grefte, S.; Smeitink, J.A.M.; Willems, P.H.G.M.

    2006-01-01

    BACKGROUND: Mitochondrial dysfunction may lead to increased oxidative stress and consequent changes in cell spreading. Here, we describe and validate a novel method for simultaneous quantification of these two parameters. METHODS: Human skin fibroblasts were loaded with

  8. Unusual Spread of Renal Cell Carcinoma to the Clivus with Cranial Nerve Deficit

    OpenAIRE

    Okudo, Jerome; Anusim, Nwabundo

    2016-01-01

    Renal cell carcinoma (RCC) has unusual presentation affecting elderly males with a smoking history. The incidence of RCC varies while the incidence of spread of RCC to the clivus is rare. The typicality of RCC presentation includes hematuria, flank pain, and a palpable flank mass; however, RCC can also present with clival metastasis. The unique path of the abducens nerve in the clivus makes it susceptible to damage in metastasis. We report a case of a 54-year-old African American female that ...

  9. Water reservoir maintained by cell growth fuels the spreading of a bacterial swarm.

    Science.gov (United States)

    Wu, Yilin; Berg, Howard C

    2012-03-13

    Flagellated bacteria can swim across moist surfaces within a thin layer of fluid, a means for surface colonization known as swarming. This fluid spreads with the swarm, but how it does so is unclear. We used micron-sized air bubbles to study the motion of this fluid within swarms of Escherichia coli. The bubbles moved diffusively, with drift. Bubbles starting at the swarm edge drifted inward for the first 5 s and then moved outward. Bubbles starting 30 μm from the swarm edge moved inward for the first 20 s, wandered around in place for the next 40 s, and then moved outward. Bubbles starting at 200 or 300 μm from the edge moved outward or wandered around in place, respectively. So the general trend was inward near the outer edge of the swarm and outward farther inside, with flows converging on a region about 100 μm from the swarm edge. We measured cellular metabolic activities with cells expressing a short-lived GFP and cell densities with cells labeled with a membrane fluorescent dye. The fluorescence plots were similar, with peaks about 80 μm from the swarm edge and slopes that mimicked the particle drift rates. These plots suggest that net fluid flow is driven by cell growth. Fluid depth is largest in the multilayered region between approximately 30 and 200 μm from the swarm edge, where fluid agitation is more vigorous. This water reservoir travels with the swarm, fueling its spreading. Intercellular communication is not required; cells need only grow.

  10. HIV-associated disruption of tight and adherens junctions of oral epithelial cells facilitates HSV-1 infection and spread.

    Directory of Open Access Journals (Sweden)

    Irna Sufiawati

    Full Text Available Herpes simplex virus (HSV types 1 and 2 are the most common opportunistic infections in HIV/AIDS. In these immunocompromised individuals, HSV-1 reactivates and replicates in oral epithelium, leading to oral disorders such as ulcers, gingivitis, and necrotic lesions. Although the increased risk of HSV infection may be mediated in part by HIV-induced immune dysfunction, direct or indirect interactions of HIV and HSV at the molecular level may also play a role. In this report we show that prolonged interaction of the HIV proteins tat and gp120 and cell-free HIV virions with polarized oral epithelial cells leads to disruption of tight and adherens junctions of epithelial cells through the mitogen-activated protein kinase signaling pathway. HIV-induced disruption of oral epithelial junctions facilitates HSV-1 paracellular spread between the epithelial cells. Furthermore, HIV-associated disruption of adherens junctions exposes sequestered nectin-1, an adhesion protein and critical receptor for HSV envelope glycoprotein D (gD. Exposure of nectin-1 facilitates binding of HSV-1 gD, which substantially increases HSV-1 infection of epithelial cells with disrupted junctions over that of cells with intact junctions. Exposed nectin-1 from disrupted adherens junctions also increases the cell-to-cell spread of HSV-1 from infected to uninfected oral epithelial cells. Antibodies to nectin-1 and HSV-1 gD substantially reduce HSV-1 infection and cell-to-cell spread, indicating that HIV-promoted HSV infection and spread are mediated by the interaction of HSV gD with HIV-exposed nectin-1. Our data suggest that HIV-associated disruption of oral epithelial junctions may potentiate HSV-1 infection and its paracellular and cell-to-cell spread within the oral mucosal epithelium. This could be one of the possible mechanisms of rapid development of HSV-associated oral lesions in HIV-infected individuals.

  11. Transient Oral Human Cytomegalovirus Infections Indicate Inefficient Viral Spread from Very Few Initially Infected Cells.

    Science.gov (United States)

    Mayer, Bryan T; Krantz, Elizabeth M; Swan, David; Ferrenberg, James; Simmons, Karen; Selke, Stacy; Huang, Meei-Li; Casper, Corey; Corey, Lawrence; Wald, Anna; Schiffer, Joshua T; Gantt, Soren

    2017-06-15

    Cytomegalovirus (CMV) is acquired by the oral route in children, and primary infection is associated with abundant mucosal replication, as well as the establishment of latency in myeloid cells that results in lifelong infection. The efficiency of primary CMV infection in humans following oral exposure, however, is unknown. We consistently detected self-limited, low-level oral CMV shedding events, which we termed transient CMV infections, in a prospective birth cohort of 30 highly exposed CMV-uninfected infants. We estimated the likelihood of transient oral CMV infections by comparing their observed frequency to that of established primary infections, characterized by persistent high-level shedding, viremia, and seroconversion. We developed mathematical models of viral dynamics upon initial oral CMV infection and validated them using clinical shedding data. Transient infections comprised 76 to 88% of oral CMV shedding events. For this high percentage of transient infections to occur, we identified two mathematical prerequisites: a very small number of initially infected oral cells (1 to 4) and low viral infectivity (<1.5 new cells infected/cell). These observations indicate that oral CMV infection in infants typically begins with a single virus that spreads inefficiently to neighboring cells. Thus, although the incidence of CMV infection is high during infancy, our data provide a mechanistic framework to explain why multiple CMV exposures are typically required before infection is successfully established. These findings imply that a sufficiently primed immune response could prevent CMV from establishing latent infection in humans and support the achievability of a prophylactic CMV vaccine. IMPORTANCE CMV infects the majority of the world's population and is a major cause of birth defects. Developing a vaccine to prevent CMV infection would be extremely valuable but would be facilitated by a better understanding of how natural human CMV infection is acquired. We

  12. Unusual Spread of Renal Cell Carcinoma to the Clivus with Cranial Nerve Deficit

    Directory of Open Access Journals (Sweden)

    Jerome Okudo

    2016-01-01

    Full Text Available Renal cell carcinoma (RCC has unusual presentation affecting elderly males with a smoking history. The incidence of RCC varies while the incidence of spread of RCC to the clivus is rare. The typicality of RCC presentation includes hematuria, flank pain, and a palpable flank mass; however, RCC can also present with clival metastasis. The unique path of the abducens nerve in the clivus makes it susceptible to damage in metastasis. We report a case of a 54-year-old African American female that was evaluated for back pain, weakness, numbness, and tingling of bilateral lower extremities and subsequently disconjugate gaze and diplopia. Brain MRI confirmed metastasis to the clivus. She was started on radiotherapy and was planned for chemotherapy and transfer to a nursing home. When a patient presents with sudden unusual cranial nerve pathology, the possibility of metastatic RCC should be sought.

  13. Unusual Spread of Renal Cell Carcinoma to the Clivus with Cranial Nerve Deficit.

    Science.gov (United States)

    Okudo, Jerome; Anusim, Nwabundo

    2016-01-01

    Renal cell carcinoma (RCC) has unusual presentation affecting elderly males with a smoking history. The incidence of RCC varies while the incidence of spread of RCC to the clivus is rare. The typicality of RCC presentation includes hematuria, flank pain, and a palpable flank mass; however, RCC can also present with clival metastasis. The unique path of the abducens nerve in the clivus makes it susceptible to damage in metastasis. We report a case of a 54-year-old African American female that was evaluated for back pain, weakness, numbness, and tingling of bilateral lower extremities and subsequently disconjugate gaze and diplopia. Brain MRI confirmed metastasis to the clivus. She was started on radiotherapy and was planned for chemotherapy and transfer to a nursing home. When a patient presents with sudden unusual cranial nerve pathology, the possibility of metastatic RCC should be sought.

  14. Materials as stem cell regulators

    Science.gov (United States)

    Murphy, William L.; McDevitt, Todd C.; Engler, Adam J.

    2014-01-01

    The stem cell/material interface is a complex, dynamic microenvironment in which the cell and the material cooperatively dictate one another's fate: the cell by remodelling its surroundings, and the material through its inherent properties (such as adhesivity, stiffness, nanostructure or degradability). Stem cells in contact with materials are able to sense their properties, integrate cues via signal propagation and ultimately translate parallel signalling information into cell fate decisions. However, discovering the mechanisms by which stem cells respond to inherent material characteristics is challenging because of the highly complex, multicomponent signalling milieu present in the stem cell environment. In this Review, we discuss recent evidence that shows that inherent material properties may be engineered to dictate stem cell fate decisions, and overview a subset of the operative signal transduction mechanisms that have begun to emerge. Further developments in stem cell engineering and mechanotransduction are poised to have substantial implications for stem cell biology and regenerative medicine. PMID:24845994

  15. Loss of an actin crosslinker uncouples cell spreading from cell stiffening on gels with a gradient of stiffness

    Science.gov (United States)

    Wen, Qi; Byfield, Fitzroy J.; Nordstrom, Kerstin; Arratia, Paulo E.; Miller, R. Tyler; Janmey, Paul A.

    2009-03-01

    We use microfluidics techniques to produce gels with a gradient of stiffness to show the essential function of the actin crosslinker filamin A in cell responses to mechanical stimuli. M2 melanoma cells null for filamin A do not alter their adherent area in response to increased substrate stiffness when they link to the substrate only through collagen receptors, but change adherent area normally when bound through fibronectin receptors. In contrast, filamin A-replete A7 cells change adherent area on both substrates and respond more strongly to collagen 1-coated gels than to fibronectin-coated gels. A7 cells alter their stiffness, as measured by atomic force microscopy, to match the elastic modulus of the substrate immediately adjacent to them on the gradient. M2 cells, in contrast, maintain a constant stiffness on all substrates that is as low as that of A7 cells on the softest gels achievable (1000 Pa). By contrasting the responses of these cell types to different adhesive substrates, cell spreading can be dissociated from stiffening.

  16. Mobilization of HIV Spread by Diaphanous 2 Dependent Filopodia in Infected Dendritic Cells

    Science.gov (United States)

    Aggarwal, Anupriya; Iemma, Tina L.; Shih, Ivy; Newsome, Timothy P.; McAllery, Samantha; Cunningham, Anthony L.; Turville, Stuart G.

    2012-01-01

    Paramount to the success of persistent viral infection is the ability of viruses to navigate hostile environments en route to future targets. In response to such obstacles, many viruses have developed the ability of establishing actin rich-membrane bridges to aid in future infections. Herein through dynamic imaging of HIV infected dendritic cells, we have observed how viral high-jacking of the actin/membrane network facilitates one of the most efficient forms of HIV spread. Within infected DC, viral egress is coupled to viral filopodia formation, with more than 90% of filopodia bearing immature HIV on their tips at extensions of 10 to 20 µm. Live imaging showed HIV filopodia routinely pivoting at their base, and projecting HIV virions at µm.sec−1 along repetitive arc trajectories. HIV filopodial dynamics lead to up to 800 DC to CD4 T cell contacts per hour, with selection of T cells culminating in multiple filopodia tethering and converging to envelope the CD4 T-cell membrane with budding HIV particles. Long viral filopodial formation was dependent on the formin diaphanous 2 (Diaph2), and not a dominant Arp2/3 filopodial pathway often associated with pathogenic actin polymerization. Manipulation of HIV Nef reduced HIV transfer 25-fold by reducing viral filopodia frequency, supporting the potency of DC HIV transfer was dependent on viral filopodia abundance. Thus our observations show HIV corrupts DC to CD4 T cell interactions by physically embedding at the leading edge contacts of long DC filopodial networks. PMID:22685410

  17. Mobilization of HIV spread by diaphanous 2 dependent filopodia in infected dendritic cells.

    Directory of Open Access Journals (Sweden)

    Anupriya Aggarwal

    Full Text Available Paramount to the success of persistent viral infection is the ability of viruses to navigate hostile environments en route to future targets. In response to such obstacles, many viruses have developed the ability of establishing actin rich-membrane bridges to aid in future infections. Herein through dynamic imaging of HIV infected dendritic cells, we have observed how viral high-jacking of the actin/membrane network facilitates one of the most efficient forms of HIV spread. Within infected DC, viral egress is coupled to viral filopodia formation, with more than 90% of filopodia bearing immature HIV on their tips at extensions of 10 to 20 µm. Live imaging showed HIV filopodia routinely pivoting at their base, and projecting HIV virions at µm.sec⁻¹ along repetitive arc trajectories. HIV filopodial dynamics lead to up to 800 DC to CD4 T cell contacts per hour, with selection of T cells culminating in multiple filopodia tethering and converging to envelope the CD4 T-cell membrane with budding HIV particles. Long viral filopodial formation was dependent on the formin diaphanous 2 (Diaph2, and not a dominant Arp2/3 filopodial pathway often associated with pathogenic actin polymerization. Manipulation of HIV Nef reduced HIV transfer 25-fold by reducing viral filopodia frequency, supporting the potency of DC HIV transfer was dependent on viral filopodia abundance. Thus our observations show HIV corrupts DC to CD4 T cell interactions by physically embedding at the leading edge contacts of long DC filopodial networks.

  18. Immune regulation by mast cells

    NARCIS (Netherlands)

    Suurmond, Jolien

    2016-01-01

    The objective of this PhD thesis is to understand mast cell (and basophil) functions and their role in autoimmune disease by focusing on three main aims: 1. To characterize the interaction between innate and Fc receptor triggers on mast cell and basophil function 2. To analyze the interaction

  19. Cell Cycle Regulation of Stem Cells by MicroRNAs.

    Science.gov (United States)

    Mens, Michelle M J; Ghanbari, Mohsen

    2018-06-01

    MicroRNAs (miRNAs) are a class of small non-coding RNA molecules involved in the regulation of gene expression. They are involved in the fine-tuning of fundamental biological processes such as proliferation, differentiation, survival and apoptosis in many cell types. Emerging evidence suggests that miRNAs regulate critical pathways involved in stem cell function. Several miRNAs have been suggested to target transcripts that directly or indirectly coordinate the cell cycle progression of stem cells. Moreover, previous studies have shown that altered expression levels of miRNAs can contribute to pathological conditions, such as cancer, due to the loss of cell cycle regulation. However, the precise mechanism underlying miRNA-mediated regulation of cell cycle in stem cells is still incompletely understood. In this review, we discuss current knowledge of miRNAs regulatory role in cell cycle progression of stem cells. We describe how specific miRNAs may control cell cycle associated molecules and checkpoints in embryonic, somatic and cancer stem cells. We further outline how these miRNAs could be regulated to influence cell cycle progression in stem cells as a potential clinical application.

  20. The Arabidopsis synaptotagmin SYTA regulates the cell-to-cell movement of diverse plant viruses

    Directory of Open Access Journals (Sweden)

    Asako eUchiyama

    2014-11-01

    Full Text Available Synaptotagmins are a large gene family in animals that have been extensively characterized due to their role as calcium sensors to regulate synaptic vesicle exocytosis and endocytosis in neurons, and dense core vesicle exocytosis for hormone secretion from neuroendocrine cells. Thought to be exclusive to animals, synaptotagmins have recently been characterized in Arabidopsis thaliana, in which they comprise a five gene family. Using infectivity and leaf-based functional assays, we have shown that Arabidopsis SYTA regulates endocytosis and marks an endosomal vesicle recycling pathway to regulate movement protein-mediated trafficking of the Begomovirus Cabbage leaf curl virus (CaLCuV and the Tobamovirus Tobacco mosaic virus (TMV through plasmodesmata (Lewis and Lazarowitz, 2010. To determine whether SYTA has a central role in regulating the cell-to-cell trafficking of a wider range of diverse plant viruses, we extended our studies here to examine the role of SYTA in the cell-to-cell movement of additional plant viruses that employ different modes of movement, namely the Potyvirus Turnip mosaic virus (TuMV, the Caulimovirus Cauliflower mosaic virus (CaMV and the Tobamovirus Turnip vein clearing virus (TVCV, which in contrast to TMV does efficiently infect Arabidopsis. We found that both TuMV and TVCV systemic infection, and the cell-to-cell trafficking of the their movement proteins, were delayed in the Arabidopsis Col-0 syta-1 knockdown mutant. In contrast, CaMV systemic infection was not inhibited in syta-1. Our studies show that SYTA is a key regulator of plant virus intercellular movement, being necessary for the ability of diverse cell-to-cell movement proteins encoded by Begomoviruses (CaLCuV MP, Tobamoviruses (TVCV and TMV 30K protein and Potyviruses (TuMV P3N-PIPO to alter PD and thereby mediate virus cell-to-cell spread.

  1. Cell Size Regulation in Bacteria

    Science.gov (United States)

    Amir, Ariel

    2014-05-01

    Various bacteria such as the canonical gram negative Escherichia coli or the well-studied gram positive Bacillus subtilis divide symmetrically after they approximately double their volume. Their size at division is not constant, but is typically distributed over a narrow range. Here, we propose an analytically tractable model for cell size control, and calculate the cell size and interdivision time distributions, as well as the correlations between these variables. We suggest ways of extracting the model parameters from experimental data, and show that existing data for E. coli supports partial size control, and a particular explanation: a cell attempts to add a constant volume from the time of initiation of DNA replication to the next initiation event. This hypothesis accounts for the experimentally observed correlations between mother and daughter cells as well as the exponential dependence of size on growth rate.

  2. The helicase, DDX3X, interacts with poly(A)-binding protein 1 (PABP1) and caprin-1 at the leading edge of migrating fibroblasts and is required for efficient cell spreading.

    Science.gov (United States)

    Copsey, Alice C; Cooper, Simon; Parker, Robert; Lineham, Ella; Lapworth, Cuzack; Jallad, Deema; Sweet, Steve; Morley, Simon J

    2017-08-30

    DDX3X, a helicase, can interact directly with mRNA and translation initiation factors, regulating the selective translation of mRNAs that contain a structured 5' untranslated region. This activity modulates the expression of mRNAs controlling cell cycle progression and mRNAs regulating actin dynamics, contributing to cell adhesion and motility. Previously, we have shown that ribosomes and translation initiation factors localise to the leading edge of migrating fibroblasts in loci enriched with actively translating ribosomes, thereby promoting steady-state levels of ArpC2 and Rac1 proteins at the leading edge of cells during spreading. As DDX3X can regulate Rac1 levels, cell motility and metastasis, we have examined DDX3X protein interactions and localisation using many complementary approaches. We now show that DDX3X can physically interact and co-localise with poly(A)-binding protein 1 and caprin-1 at the leading edge of spreading cells. Furthermore, as depletion of DDX3X leads to decreased cell motility, this provides a functional link between DDX3X, caprin-1 and initiation factors at the leading edge of migrating cells to promote cell migration and spreading. © 2017 The Author(s).

  3. Cell fate regulation in the shoot meristem.

    Science.gov (United States)

    Laux, T; Mayer, K F

    1998-04-01

    The shoot meristem is a proliferative centre containing pluripotent stem cells that are the ultimate source of all cells and organs continuously added to the growing shoot. The progeny of the stem cells have two developmental options, either to renew the stem cell population or to leave the meristem and to differentiate, possibly according to signals from more mature tissue. The destiny of each cell depends on its position within the dynamic shoot meristem. Genetic data suggest a simple model in which graded positional information is provided by antagonistic gene functions and is interpreted by genes which regulate cell fate.

  4. Biophysical regulation of stem cell differentiation.

    Science.gov (United States)

    Govey, Peter M; Loiselle, Alayna E; Donahue, Henry J

    2013-06-01

    Bone adaptation to its mechanical environment, from embryonic through adult life, is thought to be the product of increased osteoblastic differentiation from mesenchymal stem cells. In parallel with tissue-scale loading, these heterogeneous populations of multipotent stem cells are subject to a variety of biophysical cues within their native microenvironments. Bone marrow-derived mesenchymal stem cells-the most broadly studied source of osteoblastic progenitors-undergo osteoblastic differentiation in vitro in response to biophysical signals, including hydrostatic pressure, fluid flow and accompanying shear stress, substrate strain and stiffness, substrate topography, and electromagnetic fields. Furthermore, stem cells may be subject to indirect regulation by mechano-sensing osteocytes positioned to more readily detect these same loading-induced signals within the bone matrix. Such paracrine and juxtacrine regulation of differentiation by osteocytes occurs in vitro. Further studies are needed to confirm both direct and indirect mechanisms of biophysical regulation within the in vivo stem cell niche.

  5. Physiology of cell volume regulation in vertebrates

    DEFF Research Database (Denmark)

    Hoffmann, Else K; Lambert, Ian H; Pedersen, Stine F

    2009-01-01

    and their regulation by, e.g., membrane deformation, ionic strength, Ca(2+), protein kinases and phosphatases, cytoskeletal elements, GTP binding proteins, lipid mediators, and reactive oxygen species, upon changes in cell volume. We also discuss the nature of the upstream elements in volume sensing in vertebrate...... organisms. Importantly, cell volume impacts on a wide array of physiological processes, including transepithelial transport; cell migration, proliferation, and death; and changes in cell volume function as specific signals regulating these processes. A discussion of this issue concludes the review.......The ability to control cell volume is pivotal for cell function. Cell volume perturbation elicits a wide array of signaling events, leading to protective (e.g., cytoskeletal rearrangement) and adaptive (e.g., altered expression of osmolyte transporters and heat shock proteins) measures and, in most...

  6. Regulation of the cell cycle by irradiation

    International Nuclear Information System (INIS)

    Akashi, Makoto

    1995-01-01

    The molecular mechanism of cell proliferation is extremely complex; deregulation results in neoplastic transformation. In eukaryotes, proliferation of cells is finely regulated through the cell cycle. Studies have shown that the cell cycle is regulated by s series of enzymes known as cyclin-dependent kinases (CDKs). The activities of CDKs are controlled by their association with regulatory subunits, cyclins; the expression of cyclins and the activation of the different cyclin-CDK complexes are required for the cell to cycle. Thus, the cell cycle is regulated by activating and inhibiting phosphorylation of the CDK subunits and this program has internal check points at different stages of the cell cycle. When cells are exposed to external insults such as DNA damaging agents, negative regulation of the cell cycle occurs; arrest in either G1 or G2 stage is induced to prevent the cells from prematurely entering into the next stage before DNA is repaired. Recently, a potent inhibitor of CDKs, which inhibits the phosphorylation of retinoblastoma susceptibility (Rb) gene product by cyclin A-CDK2, cyclin E-CDK2, cyclin D1-CDK4, and cyclin D2-CDK4 complexes has been identified. This protein named WAF1, Sdi1, Cip1, or p21 (a protein of Mr 21,000) contains a p53-binding site in its promoter and studies have reported that the expression of WAF1 was directly regulated by p53; cells with loss of p53 activity due to mutational alteration were unable to induce WAF1. This chapter will be focused on the mechanisms of the cell cycle including inhibitors of CDKs, and the induction of WAF1 by irradiation through a pathway independent of p53 will be also described. (author)

  7. Deposition of cellular fibronectin and desorption of human serum albumin during adhesion and spreading of human endothelial cells on polymers

    NARCIS (Netherlands)

    Dekker, A.; Dekker, A.; Beugeling, T.; Beugeling, T.; Wind, H.; Poot, Andreas A.; Bantjes, A.; Bantjes, A.; Feijen, Jan; van Aken, W.G.

    1991-01-01

    More insight into the mechanism of adhesion of human endothelial cells (HEC) on to polymeric surfaces may lead to the development of improved small-diameter vascular grafts. HEC suspended in 20% human serum-containing culture medium adhere and spread well on moderately water-wettable polymers such

  8. Insight into the epidemiology of cutaneous squamous cell carcinoma with perineural spread.

    Science.gov (United States)

    Warren, Timothy A; Whiteman, David C; Porceddu, Sandro V; Panizza, Benedict J

    2016-09-01

    Perineural spread (PNS) of cutaneous squamous cell carcinoma of the head and neck (SCCHN) can be associated with poor outcomes. Disease understanding and awareness is limited leading to delayed diagnosis and treatment. The purpose of this study was to identify epidemiological features of patients with PNS of cutaneous SCCHN. Tumor characteristics and demographics of patients with PNS of cutaneous SCCHN managed through a single institution were collected between 1998 and 2013. One hundred twenty patients were included in this study. The majority had a history of skin cancer (85.8%). The median time from primary tumor treatment to PNS symptom onset was 16 months (range, 1-86 months). A total of 34.2% had no perineural invasion (PNI) detected in the primary, and 22.5% had no known primary tumor. Only 5.8% of the patients had nodal involvement at presentation. Patients can present with PNS from cutaneous SCCHN with no known primary tumor or with primary tumors without PNI. The majority of patients presented without regional nodal involvement. © 2016 Wiley Periodicals, Inc. Head Neck 38: 1416-1420, 2016. © 2016 Wiley Periodicals, Inc.

  9. Signaling hierarchy regulating human endothelial cell development

    Science.gov (United States)

    Our present knowledge of the regulation of mammalian endothelial cell differentiation has been largely derived from studies of mouse embryonic development. However, unique mechanisms and hierarchy of signals that govern human endothelial cell development are unknown and, thus, explored in these stud...

  10. Cell adhesion and spreading at a charged interface: Insight into the mechanism using surface techniques and mathematical modelling

    International Nuclear Information System (INIS)

    DeNardis, Nadica Ivošević; Ilić, Jadranka Pečar; Ružić, Ivica; Pletikapić, Galja

    2015-01-01

    Highlights: • Kinetics of adhesion and spreading of the algal cell at a charged interface is explored. • Amperometric signals are analyzed using extended methodology and the reaction kinetics model. • The model reconstructs and quantifies individual states of the three-step adhesion process. • Adhesion kinetics of the algal cell is slower than that of its plasma membrane vesicle. • Slow spreading of organic film at the interface could be due to the attenuated effect of the potential. - Abstract: We study the kinetics of adhesion and spreading of an algal cell and its plasma membrane vesicle at the charged interface. A simple system of an isolated plasma membrane vesicle without internal content has been developed and characterized by atomic force microscopy (AFM). We extend the methodology based on the reaction kinetics model and empirical fitting for the analysis of amperometric signals, and demonstrate its validity and pertinence in a wide range of surface charge densities. Adhesion kinetics of the algal cell is slower than that of its plasma membrane vesicle. Isolated plasma membrane contributes about one quarter to the cell contact area. The model reconstructs and quantifies individual states of the three-step adhesion process of the algal cell and makes it possible to associate them with various features of amperometric signal. At the time of current amplitude, the ruptured state predominates and the cell spread contact area is larger than its initial area as well as the contact area of the plasma membrane vesicle. These results suggest that a major structural disruption of the cell membrane, collapse of cytoskeleton and leakage of intracellular material could appear close to the time of current amplitude. Further, slow kinetics of the organic film spreading at the interface to its maximal extent is considered as the rate determining step, which could be a consequence of the attenuated effect of potential at the modified interface, stronger

  11. Lipids in the cell: organisation regulates function.

    Science.gov (United States)

    Santos, Ana L; Preta, Giulio

    2018-06-01

    Lipids are fundamental building blocks of all cells and play important roles in the pathogenesis of different diseases, including inflammation, autoimmune disease, cancer, and neurodegeneration. The lipid composition of different organelles can vary substantially from cell to cell, but increasing evidence demonstrates that lipids become organised specifically in each compartment, and this organisation is essential for regulating cell function. For example, lipid microdomains in the plasma membrane, known as lipid rafts, are platforms for concentrating protein receptors and can influence intra-cellular signalling. Lipid organisation is tightly regulated and can be observed across different model organisms, including bacteria, yeast, Drosophila, and Caenorhabditis elegans, suggesting that lipid organisation is evolutionarily conserved. In this review, we summarise the importance and function of specific lipid domains in main cellular organelles and discuss recent advances that investigate how these specific and highly regulated structures contribute to diverse biological processes.

  12. Regulation of cell cycle progression by cell-cell and cell-matrix forces

    NARCIS (Netherlands)

    Uroz, Marina; Wistorf, Sabrina; Serra-Picamal, Xavier; Conte, Vito; Sales-Pardo, Marta; Roca-Cusachs, Pere; Guimerà, Roger; Trepat, Xavier

    2018-01-01

    It has long been proposed that the cell cycle is regulated by physical forces at the cell-cell and cell-extracellular matrix (ECM) interfaces 1-12 . However, the evolution of these forces during the cycle has never been measured in a tissue, and whether this evolution affects cell cycle progression

  13. Matrix rigidity regulates cancer cell growth and cellular phenotype.

    Directory of Open Access Journals (Sweden)

    Robert W Tilghman

    2010-09-01

    Full Text Available The mechanical properties of the extracellular matrix have an important role in cell growth and differentiation. However, it is unclear as to what extent cancer cells respond to changes in the mechanical properties (rigidity/stiffness of the microenvironment and how this response varies among cancer cell lines.In this study we used a recently developed 96-well plate system that arrays extracellular matrix-conjugated polyacrylamide gels that increase in stiffness by at least 50-fold across the plate. This plate was used to determine how changes in the rigidity of the extracellular matrix modulate the biological properties of tumor cells. The cell lines tested fall into one of two categories based on their proliferation on substrates of differing stiffness: "rigidity dependent" (those which show an increase in cell growth as extracellular rigidity is increased, and "rigidity independent" (those which grow equally on both soft and stiff substrates. Cells which grew poorly on soft gels also showed decreased spreading and migration under these conditions. More importantly, seeding the cell lines into the lungs of nude mice revealed that the ability of cells to grow on soft gels in vitro correlated with their ability to grow in a soft tissue environment in vivo. The lung carcinoma line A549 responded to culture on soft gels by expressing the differentiated epithelial marker E-cadherin and decreasing the expression of the mesenchymal transcription factor Slug.These observations suggest that the mechanical properties of the matrix environment play a significant role in regulating the proliferation and the morphological properties of cancer cells. Further, the multiwell format of the soft-plate assay is a useful and effective adjunct to established 3-dimensional cell culture models.

  14. Matrix Rigidity Regulates Cancer Cell Growth and Cellular Phenotype

    Science.gov (United States)

    Tilghman, Robert W.; Cowan, Catharine R.; Mih, Justin D.; Koryakina, Yulia; Gioeli, Daniel; Slack-Davis, Jill K.; Blackman, Brett R.; Tschumperlin, Daniel J.; Parsons, J. Thomas

    2010-01-01

    Background The mechanical properties of the extracellular matrix have an important role in cell growth and differentiation. However, it is unclear as to what extent cancer cells respond to changes in the mechanical properties (rigidity/stiffness) of the microenvironment and how this response varies among cancer cell lines. Methodology/Principal Findings In this study we used a recently developed 96-well plate system that arrays extracellular matrix-conjugated polyacrylamide gels that increase in stiffness by at least 50-fold across the plate. This plate was used to determine how changes in the rigidity of the extracellular matrix modulate the biological properties of tumor cells. The cell lines tested fall into one of two categories based on their proliferation on substrates of differing stiffness: “rigidity dependent” (those which show an increase in cell growth as extracellular rigidity is increased), and “rigidity independent” (those which grow equally on both soft and stiff substrates). Cells which grew poorly on soft gels also showed decreased spreading and migration under these conditions. More importantly, seeding the cell lines into the lungs of nude mice revealed that the ability of cells to grow on soft gels in vitro correlated with their ability to grow in a soft tissue environment in vivo. The lung carcinoma line A549 responded to culture on soft gels by expressing the differentiated epithelial marker E-cadherin and decreasing the expression of the mesenchymal transcription factor Slug. Conclusions/Significance These observations suggest that the mechanical properties of the matrix environment play a significant role in regulating the proliferation and the morphological properties of cancer cells. Further, the multiwell format of the soft-plate assay is a useful and effective adjunct to established 3-dimensional cell culture models. PMID:20886123

  15. Tip cells: master regulators of tubulogenesis?

    Science.gov (United States)

    Weavers, Helen; Skaer, Helen

    2014-07-01

    The normal development of an organ depends on the coordinated regulation of multiple cell activities. Focusing on tubulogenesis, we review the role of specialised cells or groups of cells that are selected from within tissue primordia and differentiate at the outgrowing tips or leading edge of developing tubules. Tip or leading cells develop distinctive patterns of gene expression that enable them to act both as sensors and transmitters of intercellular signalling. This enables them to explore the environment, respond to both tissue intrinsic signals and extrinsic cues from surrounding tissues and to regulate the behaviour of their neighbours, including the setting of cell fate, patterning cell division, inducing polarity and promoting cell movement and cell rearrangements by neighbour exchange. Tip cells are also able to transmit mechanical tension to promote tissue remodelling and, by interacting with the extracellular matrix, they can dictate migratory pathways and organ shape. Where separate tubular structures fuse to form networks, as in the airways of insects or the vascular system of vertebrates, specialised fusion tip cells act to interconnect disparate elements of the developing network. Finally, we consider their importance in the maturation of mature physiological function and in the development of disease. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  16. Redox regulation of plant stem cell fate.

    Science.gov (United States)

    Zeng, Jian; Dong, Zhicheng; Wu, Haijun; Tian, Zhaoxia; Zhao, Zhong

    2017-10-02

    Despite the importance of stem cells in plant and animal development, the common mechanisms of stem cell maintenance in both systems have remained elusive. Recently, the importance of hydrogen peroxide (H 2 O 2 ) signaling in priming stem cell differentiation has been extensively studied in animals. Here, we show that different forms of reactive oxygen species (ROS) have antagonistic roles in plant stem cell regulation, which were established by distinct spatiotemporal patterns of ROS-metabolizing enzymes. The superoxide anion (O2·-) is markedly enriched in stem cells to activate WUSCHEL and maintain stemness, whereas H 2 O 2 is more abundant in the differentiating peripheral zone to promote stem cell differentiation. Moreover, H 2 O 2 negatively regulates O2·- biosynthesis in stem cells, and increasing H 2 O 2 levels or scavenging O2·- leads to the termination of stem cells. Our results provide a mechanistic framework for ROS-mediated control of plant stem cell fate and demonstrate that the balance between O2·- and H 2 O 2 is key to stem cell maintenance and differentiation. © 2017 The Authors.

  17. Matrix regulators in neural stem cell functions.

    Science.gov (United States)

    Wade, Anna; McKinney, Andrew; Phillips, Joanna J

    2014-08-01

    Neural stem/progenitor cells (NSPCs) reside within a complex and dynamic extracellular microenvironment, or niche. This niche regulates fundamental aspects of their behavior during normal neural development and repair. Precise yet dynamic regulation of NSPC self-renewal, migration, and differentiation is critical and must persist over the life of an organism. In this review, we summarize some of the major components of the NSPC niche and provide examples of how cues from the extracellular matrix regulate NSPC behaviors. We use proteoglycans to illustrate the many diverse roles of the niche in providing temporal and spatial regulation of cellular behavior. The NSPC niche is comprised of multiple components that include; soluble ligands, such as growth factors, morphogens, chemokines, and neurotransmitters, the extracellular matrix, and cellular components. As illustrated by proteoglycans, a major component of the extracellular matrix, the NSPC, niche provides temporal and spatial regulation of NSPC behaviors. The factors that control NSPC behavior are vital to understand as we attempt to modulate normal neural development and repair. Furthermore, an improved understanding of how these factors regulate cell proliferation, migration, and differentiation, crucial for malignancy, may reveal novel anti-tumor strategies. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Cell volume regulation: physiology and pathophysiology

    DEFF Research Database (Denmark)

    Lambert, I H; Hoffmann, E K; Pedersen, Stine Helene Falsig

    2008-01-01

    are sensed are still far from clear, significant progress has been made with respect to the nature of the sensors, transducers and effectors that convert a change in cell volume into a physiological response. In the present review, we summarize recent major developments in the field, and emphasize......Cell volume perturbation initiates a wide array of intracellular signalling cascades, leading to protective and adaptive events and, in most cases, activation of volume-regulatory osmolyte transport, water loss, and hence restoration of cell volume and cellular function. Cell volume is challenged....../hypernatremia. On the other hand, it has recently become clear that an increase or reduction in cell volume can also serve as a specific signal in the regulation of physiological processes such as transepithelial transport, cell migration, proliferation and death. Although the mechanisms by which cell volume perturbations...

  19. Regulation of Murine Natural Killer Cell Commitment

    Directory of Open Access Journals (Sweden)

    Nicholas D Huntington

    2013-01-01

    Full Text Available NK cells can derive from the same precursors as B and T cells, however to achieve lineage specificity, several transcription factors need to be activated or annulled. While a few important transcription factors have identified for NK genesis the mechanisms of how this is achieved is far from resolved. Adding to the complexity of this, NK cells are found and potentially develop in diverse locations in vivo and it remains to be addressed if a common NK cell precursor seeds diverse niches and how transcription factors may differentially regulate NK cell commitment in distinct microenvironments. Here we will summarise some recent findings in NK cell commitment and discuss how a NK cell transcriptional network might be organised, while addressing some misconceptions and anomalies along the way.

  20. Mast cell activators as novel immune regulators.

    Science.gov (United States)

    Johnson-Weaver, Brandi; Choi, Hae Woong; Abraham, Soman N; Staats, Herman F

    2018-05-26

    Mast cells are an important cell type of the innate immune system that when activated, play a crucial role in generating protective innate host responses after bacterial and viral infection. Additionally, activated mast cells influence lymph node composition to regulate the induction of adaptive immune responses. The recognition that mast cells play a beneficial role in host responses to microbial infection and induction of adaptive immunity has provided the rationale to evaluate mast cell activators for use as antimicrobials or vaccine adjuvants. This review summarizes the role of mast cell activators in antimicrobial responses while also discussing the use of different classes of mast cell activators as potent vaccine adjuvants that enhance the induction of protective immune responses. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Signaling hierarchy regulating human endothelial cell development.

    Science.gov (United States)

    Kelly, Melissa A; Hirschi, Karen K

    2009-05-01

    Our present knowledge of the regulation of mammalian endothelial cell differentiation has been largely derived from studies of mouse embryonic development. However, unique mechanisms and hierarchy of signals that govern human endothelial cell development are unknown and, thus, explored in these studies. Using human embryonic stem cells as a model system, we were able to reproducibly and robustly generate differentiated endothelial cells via coculture on OP9 marrow stromal cells. We found that, in contrast to studies in the mouse, bFGF and VEGF had no specific effects on the initiation of human vasculogenesis. However, exogenous Ihh promoted endothelial cell differentiation, as evidenced by increased production of cells with cobblestone morphology that coexpress multiple endothelial-specific genes and proteins, form lumens, and exhibit DiI-AcLDL uptake. Inhibition of BMP signaling using Noggin or BMP4, specifically, using neutralizing antibodies suppressed endothelial cell formation; whereas, addition of rhBMP4 to cells treated with the hedgehog inhibitor cyclopamine rescued endothelial cell development. Our studies revealed that Ihh promoted human endothelial cell differentiation from pluripotent hES cells via BMP signaling, providing novel insights applicable to modulating human endothelial cell formation and vascular regeneration for human clinical therapies.

  2. Adhesive and mechanical regulation of mesenchymal stem cell differentiation in human bone marrow and periosteum-derived progenitor cells

    Directory of Open Access Journals (Sweden)

    Jeroen Eyckmans

    2012-08-01

    It has previously been demonstrated that cell shape can influence commitment of human bone marrow-derived mesenchymal stem cells (hBMCs to adipogenic, osteogenic, chondrogenic, and other lineages. Human periosteum-derived cells (hPDCs exhibit multipotency similar to hBMCs, but hPDCs may offer enhanced potential for osteogenesis and chondrogenesis given their apparent endogenous role in bone and cartilage repair in vivo. Here, we examined whether hPDC differentiation is regulated by adhesive and mechanical cues comparable to that reported for hBMC differentiation. When cultured in the appropriate induction media, hPDCs at high cell seeding density demonstrated enhanced levels of adipogenic or chondrogenic markers as compared with hPDCs at low cell seeding density. Cell seeding density correlated inversely with projected area of cell spreading, and directly limiting cell spreading with micropatterned substrates promoted adipogenesis or chondrogenesis while substrates promoting cell spreading supported osteogenesis. Interestingly, cell seeding density influenced differentiation through both changes in cell shape and non-shape-mediated effects: density-dependent adipogenesis and chondrogenesis were regulated primarily by cell shape whereas non-shape effects strongly influenced osteogenic potential. Inhibition of cytoskeletal contractility by adding the Rho kinase inhibitor Y27632 further enhanced adipogenic differentiation and discouraged osteogenic differentiation of hPDCs. Together, our results suggest that multipotent lineage decisions of hPDCs are impacted by cell adhesive and mechanical cues, though to different extents than hBMCs. Thus, future studies of hPDCs and other primary stem cell populations with clinical potential should consider varying biophysical metrics for more thorough optimization of stem cell differentiation.

  3. Rac function is crucial for cell migration but is not required for spreading and focal adhesion formation

    DEFF Research Database (Denmark)

    Steffen, Anika; Ladwein, Markus; Dimchev, Georgi A

    2013-01-01

    can be potently stimulated by Rho GTPases of the Rac subfamily, but also by RhoG or Cdc42. Here we describe viable fibroblast cell lines genetically deficient for Rac1 that lack detectable levels of Rac2 and Rac3. Rac-deficient cells were devoid of apparent lamellipodia, but these structures were...... filopodia and established focal adhesions. Spreading in these cells was achieved by the extension of filopodia followed by the advancement of cytoplasmic veils between them. The number and size of focal adhesions as well as their intensity were largely unaffected by genetic removal of Rac1. However, Rac...

  4. Cell cycle regulation of hematopoietic stem or progenitor cells.

    Science.gov (United States)

    Hao, Sha; Chen, Chen; Cheng, Tao

    2016-05-01

    The highly regulated process of blood production is achieved through the hierarchical organization of hematopoietic stem cell (HSC) subsets and their progenies, which differ in self-renewal and differentiation potential. Genetic studies in mice have demonstrated that cell cycle is tightly controlled by the complex interplay between extrinsic cues and intrinsic regulatory pathways involved in HSC self-renewal and differentiation. Deregulation of these cellular programs may transform HSCs or hematopoietic progenitor cells (HPCs) into disease-initiating stem cells, and can result in hematopoietic malignancies such as leukemia. While previous studies have shown roles for some cell cycle regulators and related signaling pathways in HSCs and HPCs, a more complete picture regarding the molecular mechanisms underlying cell cycle regulation in HSCs or HPCs is lacking. Based on accumulated studies in this field, the present review introduces the basic components of the cell cycle machinery and discusses their major cellular networks that regulate the dormancy and cell cycle progression of HSCs. Knowledge on this topic would help researchers and clinicians to better understand the pathogenesis of relevant blood disorders and to develop new strategies for therapeutic manipulation of HSCs.

  5. Triiodothyronine regulates cell growth and survival in renal cell cancer.

    Science.gov (United States)

    Czarnecka, Anna M; Matak, Damian; Szymanski, Lukasz; Czarnecka, Karolina H; Lewicki, Slawomir; Zdanowski, Robert; Brzezianska-Lasota, Ewa; Szczylik, Cezary

    2016-10-01

    Triiodothyronine plays an important role in the regulation of kidney cell growth, differentiation and metabolism. Patients with renal cell cancer who develop hypothyreosis during tyrosine kinase inhibitor (TKI) treatment have statistically longer survival. In this study, we developed cell based model of triiodothyronine (T3) analysis in RCC and we show the different effects of T3 on renal cell cancer (RCC) cell growth response and expression of the thyroid hormone receptor in human renal cell cancer cell lines from primary and metastatic tumors along with human kidney cancer stem cells. Wild-type thyroid hormone receptor is ubiquitously expressed in human renal cancer cell lines, but normalized against healthy renal proximal tube cell expression its level is upregulated in Caki-2, RCC6, SKRC-42, SKRC-45 cell lines. On the contrary the mRNA level in the 769-P, ACHN, HKCSC, and HEK293 cells is significantly decreased. The TRβ protein was abundant in the cytoplasm of the 786-O, Caki-2, RCC6, and SKRC-45 cells and in the nucleus of SKRC-42, ACHN, 769-P and cancer stem cells. T3 has promoting effect on the cell proliferation of HKCSC, Caki-2, ASE, ACHN, SK-RC-42, SMKT-R2, Caki-1, 786-0, and SK-RC-45 cells. Tyrosine kinase inhibitor, sunitinib, directly inhibits proliferation of RCC cells, while thyroid hormone receptor antagonist 1-850 (CAS 251310‑57-3) has less significant inhibitory impact. T3 stimulation does not abrogate inhibitory effect of sunitinib. Renal cancer tumor cells hypostimulated with T3 may be more responsive to tyrosine kinase inhibition. Moreover, some tumors may be considered as T3-independent and present aggressive phenotype with thyroid hormone receptor activated independently from the ligand. On the contrary proliferation induced by deregulated VHL and or c-Met pathways may transgress normal T3 mediated regulation of the cell cycle.

  6. Redox regulation in cancer stem cells

    Science.gov (United States)

    Reactive oxygen species (ROS) and ROS-dependent (redox regulation) signaling pathways and transcriptional activities are thought to be critical in stem cell self-renewal and differentiation during growth and organogenesis. Aberrant ROS burst and dysregulation of those ROS-dependent cellular processe...

  7. The regulation of apoptotic cell death

    Directory of Open Access Journals (Sweden)

    Amarante-Mendes G.P.

    1999-01-01

    Full Text Available Apoptosis is a fundamental biological phenomenon in which the death of a cell is genetically and biochemically regulated. Different molecules are involved in the regulation of the apoptotic process. Death receptors, coupled to distinct members of the caspases as well as other adapter molecules, are involved in the initiation of the stress signals (The Indictment. Members of the Bcl-2 family control at the mitochondrial level the decision between life and death (The Judgement. The effector caspases are responsible for all morphological and biochemical changes related to apoptosis including the "eat-me" signals perceived by phagocytes and neighboring cells (The Execution. Finally, apoptosis would have little biological significance without the recognition and removal of the dying cells (The Burial.

  8. Dynamic ubiquitin signaling in cell cycle regulation.

    Science.gov (United States)

    Gilberto, Samuel; Peter, Matthias

    2017-08-07

    The cell division cycle is driven by a collection of enzymes that coordinate DNA duplication and separation, ensuring that genomic information is faithfully and perpetually maintained. The activity of the effector proteins that perform and coordinate these biological processes oscillates by regulated expression and/or posttranslational modifications. Ubiquitylation is a cardinal cellular modification and is long known for driving cell cycle transitions. In this review, we emphasize emerging concepts of how ubiquitylation brings the necessary dynamicity and plasticity that underlie the processes of DNA replication and mitosis. New studies, often focusing on the regulation of chromosomal proteins like DNA polymerases or kinetochore kinases, are demonstrating that ubiquitylation is a versatile modification that can be used to fine-tune these cell cycle events, frequently through processes that do not involve proteasomal degradation. Understanding how the increasing variety of identified ubiquitin signals are transduced will allow us to develop a deeper mechanistic perception of how the multiple factors come together to faithfully propagate genomic information. Here, we discuss these and additional conceptual challenges that are currently under study toward understanding how ubiquitin governs cell cycle regulation. © 2017 Gilberto and Peter.

  9. Regulation of satellite cell function in sarcopenia

    Directory of Open Access Journals (Sweden)

    Stephen E Alway

    2014-09-01

    Full Text Available The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell function that is impacted by the environment (niche of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia, and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration. While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function.

  10. Regulation of Satellite Cell Function in Sarcopenia

    Science.gov (United States)

    Alway, Stephen E.; Myers, Matthew J.; Mohamed, Junaith S.

    2014-01-01

    The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell) function that is impacted by the environment (niche) of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse, or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins, and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration). While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function. PMID:25295003

  11. Modeling potential Emerald Ash Borer spread through GIS/cell-based/gravity models with data bolstered by web-based inputs

    Science.gov (United States)

    Louis R. Iverson; Anantha M. Prasad; Davis Sydnor; Jonathan Bossenbroek; Mark W. Schwartz; Mark W. Schwartz

    2006-01-01

    We model the susceptibility and potential spread of the organism across the eastern United States and especially through Michigan and Ohio using Forest Inventory and Analysis (FIA) data. We are also developing a cell-based model for the potential spread of the organism. We have developed a web-based tool for public agencies and private individuals to enter the...

  12. Disruption of the novel gene fad104 causes rapid postnatal death and attenuation of cell proliferation, adhesion, spreading and migration

    International Nuclear Information System (INIS)

    Nishizuka, Makoto; Kishimoto, Keishi; Kato, Ayumi; Ikawa, Masahito; Okabe, Masaru; Sato, Ryuichiro; Niida, Hiroyuki; Nakanishi, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2009-01-01

    The molecular mechanisms at the beginning of adipogenesis remain unknown. Previously, we identified a novel gene, fad104 (factor for adipocyte differentiation 104), transiently expressed at the early stage of adipocyte differentiation. Since the knockdown of the expression of fad104 dramatically repressed adipogenesis, it is clear that fad104 plays important roles in adipocyte differentiation. However, the physiological roles of fad104 are still unknown. In this study, we generated fad104-deficient mice by gene targeting. Although the mice were born in the expected Mendelian ratios, all died within 1 day of birth, suggesting fad104 to be crucial for survival after birth. Furthermore, analyses of mouse embryonic fibroblasts (MEFs) prepared from fad104-deficient mice provided new insights into the functions of fad104. Disruption of fad104 inhibited adipocyte differentiation and cell proliferation. In addition, cell adhesion and wound healing assays using fad104-deficient MEFs revealed that loss of fad104 expression caused a reduction in stress fiber formation, and notably delayed cell adhesion, spreading and migration. These results indicate that fad104 is essential for the survival of newborns just after birth and important for cell proliferation, adhesion, spreading and migration

  13. Monte Carlo calculated microdosimetric spread for cell nucleus-sized targets exposed to brachytherapy 125I and 192Ir sources and 60Co cell irradiation.

    Science.gov (United States)

    Villegas, Fernanda; Tilly, Nina; Ahnesjö, Anders

    2013-09-07

    The stochastic nature of ionizing radiation interactions causes a microdosimetric spread in energy depositions for cell or cell nucleus-sized volumes. The magnitude of the spread may be a confounding factor in dose response analysis. The aim of this work is to give values for the microdosimetric spread for a range of doses imparted by (125)I and (192)Ir brachytherapy radionuclides, and for a (60)Co source. An upgraded version of the Monte Carlo code PENELOPE was used to obtain frequency distributions of specific energy for each of these radiation qualities and for four different cell nucleus-sized volumes. The results demonstrate that the magnitude of the microdosimetric spread increases when the target size decreases or when the energy of the radiation quality is reduced. Frequency distributions calculated according to the formalism of Kellerer and Chmelevsky using full convolution of the Monte Carlo calculated single track frequency distributions confirm that at doses exceeding 0.08 Gy for (125)I, 0.1 Gy for (192)Ir, and 0.2 Gy for (60)Co, the resulting distribution can be accurately approximated with a normal distribution. A parameterization of the width of the distribution as a function of dose and target volume of interest is presented as a convenient form for the use in response modelling or similar contexts.

  14. Cell cycle regulation in human embryonic stem cells: links to adaptation to cell culture.

    Science.gov (United States)

    Barta, Tomas; Dolezalova, Dasa; Holubcova, Zuzana; Hampl, Ales

    2013-03-01

    Cell cycle represents not only a tightly orchestrated mechanism of cell replication and cell division but it also plays an important role in regulation of cell fate decision. Particularly in the context of pluripotent stem cells or multipotent progenitor cells, regulation of cell fate decision is of paramount importance. It has been shown that human embryonic stem cells (hESCs) show unique cell cycle characteristics, such as short doubling time due to abbreviated G1 phase; these properties change with the onset of differentiation. This review summarizes the current understanding of cell cycle regulation in hESCs. We discuss cell cycle properties as well as regulatory machinery governing cell cycle progression of undifferentiated hESCs. Additionally, we provide evidence that long-term culture of hESCs is accompanied by changes in cell cycle properties as well as configuration of several cell cycle regulatory molecules.

  15. Radiotherapy and chemotherapy change vessel tree geometry and metastatic spread in a small cell lung cancer xenograft mouse tumor model.

    Directory of Open Access Journals (Sweden)

    Thorsten Frenzel

    Full Text Available Tumor vasculature is critical for tumor growth, formation of distant metastases and efficiency of radio- and chemotherapy treatments. However, how the vasculature itself is affected during cancer treatment regarding to the metastatic behavior has not been thoroughly investigated. Therefore, the aim of this study was to analyze the influence of hypofractionated radiotherapy and cisplatin chemotherapy on vessel tree geometry and metastasis formation in a small cell lung cancer xenograft mouse tumor model to investigate the spread of malignant cells during different treatments modalities.The biological data gained during these experiments were fed into our previously developed computer model "Cancer and Treatment Simulation Tool" (CaTSiT to model the growth of the primary tumor, its metastatic deposit and also the influence on different therapies. Furthermore, we performed quantitative histology analyses to verify our predictions in xenograft mouse tumor model.According to the computer simulation the number of cells engrafting must vary considerably to explain the different weights of the primary tumor at the end of the experiment. Once a primary tumor is established, the fractal dimension of its vasculature correlates with the tumor size. Furthermore, the fractal dimension of the tumor vasculature changes during treatment, indicating that the therapy affects the blood vessels' geometry. We corroborated these findings with a quantitative histological analysis showing that the blood vessel density is depleted during radiotherapy and cisplatin chemotherapy. The CaTSiT computer model reveals that chemotherapy influences the tumor's therapeutic susceptibility and its metastatic spreading behavior.Using a system biological approach in combination with xenograft models and computer simulations revealed that the usage of chemotherapy and radiation therapy determines the spreading behavior by changing the blood vessel geometry of the primary tumor.

  16. Renal intercalated cells and blood pressure regulation

    Directory of Open Access Journals (Sweden)

    Susan M. Wall

    2017-12-01

    Full Text Available Type B and non-A, non-B intercalated cells are found within the connecting tubule and the cortical collecting duct. Of these cell types, type B intercalated cells are known to mediate Cl⁻ absorption and HCO₃⁻ secretion largely through pendrin-dependent Cl⁻/HCO₃⁻ exchange. This exchange is stimulated by angiotensin II administration and is also stimulated in models of metabolic alkalosis, for instance after aldosterone or NaHCO₃ administration. In some rodent models, pendrin-mediated HCO₃⁻ secretion modulates acid-base balance. However, the role of pendrin in blood pressure regulation is likely of more physiological or clinical significance. Pendrin regulates blood pressure not only by mediating aldosterone-sensitive Cl⁻ absorption, but also by modulating the aldosterone response for epithelial Na⁺ channel (ENaC-mediated Na⁺ absorption. Pendrin regulates ENaC through changes in open channel of probability, channel surface density, and channels subunit total protein abundance. Thus, aldosterone stimulates ENaC activity through both direct and indirect effects, the latter occurring through its stimulation of pendrin expression and function. Therefore, pendrin contributes to the aldosterone pressor response. Pendrin may also modulate blood pressure in part through its action in the adrenal medulla, where it modulates the release of catecholamines, or through an indirect effect on vascular contractile force. This review describes how aldosterone and angiotensin II-induced signaling regulate pendrin and the contributory role of pendrin in distal nephron function and blood pressure.

  17. Spreading Sequence Design for Multiple Cell Synchronous DS-CDMA Systems under Total Weighted Squared Correlation Criterion

    Directory of Open Access Journals (Sweden)

    Cotae Paul

    2004-01-01

    Full Text Available An algorithm for designing spreading sequences for an overloaded multicellular synchronous DS-CDMA system on uplink is introduced. The criterion used to measure the optimality of the design is the total weighted square correlation (TWSC assuming the channel state information known perfectly at both transmitter and receiver. By using this algorithm it is possible to obtain orthogonal generalized WBE sequences sets for any processing gain. The bandwidth of initial generalized WBE signals of each cell is preserved in the extended signal space associated to multicellular system. Mathematical formalism is illustrated by selected numerical examples.

  18. Cell-free DNA: A Neglected Source for Antibiotic Resistance Genes Spreading from WWTPs.

    Science.gov (United States)

    Zhang, Yan; Li, Aolin; Dai, Tianjiao; Li, Feifei; Xie, Hui; Chen, Lujun; Wen, Donghui

    2018-01-02

    Cell-associated ARGs in wastewater treatment plants (WWTPs) has been concerned, however, cell-free ARGs in WWTPs was rarely studied. In this study, the abundances of four representative ARGs, sulII, tetC, bla PSE-1 , and ermB, in a large municipal WWTP were investigated in both cell-associated and cell-free fractions. Cell-associated ARGs was the dominant ARGs fraction in the raw wastewater. After biological treatment, sludge settling, membrane filtration, and disinfection, cell-associated ARGs were substantially reduced, though the ratios of ARG/16S rRNA gene were increased with disinfection. Cell-free ARGs persisted in the WWTP with a removal of 0.36 log to 2.68 logs, which was much lower than the removal of cell-associated ARGs (3.21 logs to 4.14 logs). Therefore, the abundance ratio of cell-free ARGs to cell-associated ARGs increased from 0.04-1.59% to 2.00-1895.08% along the treatment processes. After 25-day-storage, cell-free ARGs in both biological effluent and disinfection effluent increased by 0.14 log to 1.99 logs and 0.12 log to 1.77 logs respectively, reflecting the persistence and low decay rate of cell-free ARGs in the discharge water. Therefore, cell-free ARGs might be a kind of important but previously neglected pollutant from WWTPs, which added potential risks to the effluent receiving environments.

  19. Nanomaterials for regulating cancer and stem cell fate

    Science.gov (United States)

    Shah, Birju P.

    The realm of nanomedicine has grown exponentially over the past few decades. However, there are several obstacles that need to be overcome, prior to the wide-spread clinical applications of these nanoparticles, such as (i) developing well-defined nanoparticles of varying size, morphology and composition to enable various clinical applications; (ii) overcome various physiological barriers encountered in order to deliver the therapeutics to the target location; and (iii) real-time monitoring of the nano-therapeutics within the human body for tracking their uptake, localization and effect. Hence, this dissertation focuses on developing multimodal nanotechnology-based approaches to overcome the above-mentioned challenges and thus enable regulation of cancer and stem cell fate. The initial part of this dissertation describes the development of multimodal magnetic core-shell nanoparticles (MCNPs), comprised of a highly magnetic core surrounded by a thin gold shell, thus combining magnetic and plasmonic properties. These nanoparticles were utilized for mainly two applications: (i) Magnetically-facilitated delivery of siRNA and plasmid DNA for effective stem cell differentiation and imaging and (ii) Combined hyperthermia and targeted delivery of a mitochondria-targeting peptide for enhancing apoptosis in cancer cells. The following part of this dissertation presents the generation of a multi-functional cyclodextrin-conjugated polymeric delivery platform (known as DexAMs), for co-delivery of anticancer drugs and siRNAs in a target-specific manner to brain tumor cells. This combined delivery of chemotherapeutics and siRNA resulted in a synergistic effect on the apoptosis of brain tumor cells, as compared to the individual treatments. The final part of this thesis presents development of stimuli-responsive uorescence resonance energy transfer (FRET)-based mesoporous silica nanoparticles for real-time monitoring of drug release in cells. The stimuli-responsive behavior of

  20. Cell shape regulates global histone acetylation in human mammaryepithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Le Beyec, Johanne; Xu, Ren; Lee, Sun-Young; Nelson, Celeste M.; Rizki, Aylin; Alcaraz, Jordi; Bissell, Mina J.

    2007-02-28

    Extracellular matrix (ECM) regulates cell morphology and gene expression in vivo; these relationships are maintained in three-dimensional (3D) cultures of mammary epithelial cells. In the presence of laminin-rich ECM (lrECM), mammary epithelial cells round up and undergo global histone deacetylation, a process critical for their functional differentiation. However, it remains unclear whether lrECM-dependent cell rounding and global histone deacetylation are indeed part of a common physical-biochemical pathway. Using 3D cultures as well as nonadhesive and micropatterned substrata, here we showed that the cell 'rounding' caused by lrECM was sufficient to induce deacetylation of histones H3 and H4 in the absence of biochemical cues. Microarray and confocal analysis demonstrated that this deacetylation in 3D culture is associated with a global increase in chromatin condensation and a reduction in gene expression. Whereas cells cultured on plastic substrata formed prominent stress fibers, cells grown in 3D lrECM or on micropatterns lacked these structures. Disruption of the actin cytoskeleton with cytochalasin D phenocopied the lrECM-induced cell rounding and histone deacetylation. These results reveal a novel link between ECM-controlled cell shape and chromatin structure, and suggest that this link is mediated by changes in the actin cytoskeleton.

  1. The V domain of dog PVRL4 (nectin-4) mediates canine distemper virus entry and virus cell-to-cell spread.

    Science.gov (United States)

    Delpeut, Sebastien; Noyce, Ryan S; Richardson, Christopher D

    2014-04-01

    The entry of canine distemper virus (CDV) is a multistep process that involves the attachment of CDV hemagglutinin (H) to its cellular receptor, followed by fusion between virus and cell membranes. Our laboratory recently identified PVRL4 (nectin-4) to be the epithelial receptor for measles and canine distemper viruses. In this study, we demonstrate that the V domain of PVRL4 is critical for CDV entry and virus cell-to-cell spread. Furthermore, four key amino acid residues within the V domain of dog PVRL4 and two within the CDV hemagglutinin were shown to be essential for receptor-mediated virus entry. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. EMMPRIN regulates β1 integrin-mediated adhesion through Kindlin-3 in human melanoma cells.

    Science.gov (United States)

    Delyon, Julie; Khayati, Farah; Djaafri, Ibtissem; Podgorniak, Marie-Pierre; Sadoux, Aurélie; Setterblad, Niclas; Boutalbi, Zineb; Maouche, Kamel; Maskos, Uwe; Menashi, Suzanne; Lebbé, Céleste; Mourah, Samia

    2015-06-01

    EMMPRIN is known to promote tumor invasion through extracellular matrix (ECM) degradation. Here we report that EMMPRIN can regulate melanoma cell adhesion to the ECM through an interaction with β1 integrin involving kindlin-3. In this study, EMMPRIN knockdown in the human melanoma cell line M10 using siRNA decreased cell invasion and significantly increased cell adhesion and spreading. A morphological change from a round to a spread shape was observed associated with enhanced phalloidin-labelled actin staining. In situ proximity ligation assay and co-immunoprecipitation revealed that EMMPRIN silencing increased the interaction of β1 integrin with kindlin-3, a focal adhesion protein. This was associated with an increase in β1 integrin activation and a decrease in the phosphorylation of the downstream integrin kinase FAK. Moreover, the expression at both the transcript and protein level of kindlin-3 and of β1 integrin was inversely regulated by EMMPRIN. EMMPRIN did not regulate either talin expression or its interaction with β1 integrin. These results are consistent with our in vivo demonstration that EMMPRIN inhibition increased β1 integrin activation and its interaction with kindlin-3. To conclude, these findings reveal a new role of EMMPRIN in tumor cell migration through ß1 integrin/kindlin-3-mediated adhesion pathway. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Substrate Curvature Regulates Cell Migration -A Computational Study

    Science.gov (United States)

    He, Xiuxiu; Jiang, Yi

    Cell migration in host microenvironment is essential to cancer etiology, progression and metastasis. Cellular processes of adhesion, cytoskeletal polymerization, contraction, and matrix remodeling act in concert to regulate cell migration, while local extracellular matrix architecture modulate these processes. In this work we study how stromal microenvironment with native and cell-derived curvature at micron-meter scale regulate cell motility pattern. We developed a 3D model of single cell migration on a curved substrate. Mathematical analysis of cell morphological adaption to the cell-substrate interface shows that cell migration on convex surfaces deforms more than on concave surfaces. Both analytical and simulation results show that curved surfaces regulate the cell motile force for cell's protruding front through force balance with focal adhesion and cell contraction. We also found that cell migration on concave substrates is more persistent. These results offer a novel biomechanical explanation to substrate curvature regulation of cell migration. NIH 1U01CA143069.

  4. Regulated Mucin Secretion from Airway Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Kenneth Bruce Adler

    2013-09-01

    Full Text Available Secretory epithelial cells of the proximal airways synthesize and secrete gel-forming polymeric mucins. The secreted mucins adsorb water to form mucus that is propelled by neighboring ciliated cells, providing a mobile barrier which removes inhaled particles and pathogens from the lungs. Several features of the intracellular trafficking of mucins make the airway secretory cell an interesting comparator for the cell biology of regulated exocytosis. Polymeric mucins are exceedingly large molecules (up to 3x10^6 D per monomer whose folding and initial polymerization in the ER requires the protein disulfide isomerase Agr2. In the Golgi, mucins further polymerize to form chains and possibly branched networks comprising more than 20 monomers. The large size of mucin polymers imposes constraints on their packaging into transport vesicles along the secretory pathway. Sugar side chains account for >70% of the mass of mucins, and their attachment to the protein core by O-glycosylation occurs in the Golgi. Mature polymeric mucins are stored in large secretory granules ~1 um in diameter. These are translocated to the apical membrane to be positioned for exocytosis by cooperative interactions among MARCKS, cysteine string protein (CSP, HSP70 and the cytoskeleton. Mucin granules undergo exocytic fusion with the plasma membrane at a low basal rate and a high stimulated rate. Both rates are mediated by a regulated exocytic mechanism as indicated by phenotypes in both basal and stimulated secretion in mice lacking Munc13-2, a sensor of the second messengers calcium and diacylglycerol (DAG. Basal secretion is induced by low levels of activation of P2Y2 purinergic and A3 adenosine receptors by extracellular ATP released in paracrine fashion and its metabolite adenosine. Stimulated secretion is induced by high levels of the same ligands, and possibly by inflammatory mediators as well. Activated receptors are coupled to phospholipase C by Gq, resulting in the

  5. Epigenetic regulation of hematopoietic stem cell aging

    International Nuclear Information System (INIS)

    Beerman, Isabel; Rossi, Derrick J.

    2014-01-01

    Aging is invariably associated with alterations of the hematopoietic stem cell (HSC) compartment, including loss of functional capacity, altered clonal composition, and changes in lineage contribution. Although accumulation of DNA damage occurs during HSC aging, it is unlikely such consistent aging phenotypes could be solely attributed to changes in DNA integrity. Another mechanism by which heritable traits could contribute to the changes in the functional potential of aged HSCs is through alterations in the epigenetic landscape of adult stem cells. Indeed, recent studies on hematopoietic stem cells have suggested that altered epigenetic profiles are associated with HSC aging and play a key role in modulating the functional potential of HSCs at different stages during ontogeny. Even small changes of the epigenetic landscape can lead to robustly altered expression patterns, either directly by loss of regulatory control or through indirect, additive effects, ultimately leading to transcriptional changes of the stem cells. Potential drivers of such changes in the epigenetic landscape of aged HSCs include proliferative history, DNA damage, and deregulation of key epigenetic enzymes and complexes. This review will focus largely on the two most characterized epigenetic marks – DNA methylation and histone modifications – but will also discuss the potential role of non-coding RNAs in regulating HSC function during aging

  6. Prostaglandin E2 regulates hematopoietic stem cell

    International Nuclear Information System (INIS)

    Wang Yingying; Zhou Daohong; Meng Aimin

    2013-01-01

    Prostaglandin E2 (PGE2) is a bioactive lipid molecule produced by cyclooxygenase (COX), which plays an important role on hematopoiesis. While it can block differentiation of myeloid progenitors but enhance proliferation of erythroid progenitors. Recent research found that PGE2 have the effects on hematopoietic stem cell (HSC) function and these effects were independent from effects on progenitor cells. Exposure of HSC cells to PGE2 in vitro can increase homing efficiency of HSC to the murine bone marrow compartment and decrease HSC apoptosis, meanwhile increase long-term stem cell engraftment. In-vivo treatment with PGE2 expands short-term HSC and engraftment in murine bone marrow but not long-term HSC.In addition, PGE2 increases HSC survival after radiation injury and enhance hematopoietic recovery, resulting maintains hematopoietic homeostasis. PGE2 regulates HSC homeostasis by reactive oxygen species and Wnt pathway. Clinical beneficial of 16, 16-dimethyl-prostaglandin E2 treatment to enhance engraftment of umbilical cord blood suggest important improvements to therapeutic strategies. (authors)

  7. Epigenetic regulation of hematopoietic stem cell aging

    Energy Technology Data Exchange (ETDEWEB)

    Beerman, Isabel, E-mail: isabel.beerman@childrens.harvard.edu [Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138 (United States); Department of Pediatrics, Harvard Medical School, Boston, MA 02115 (United States); Program in Cellular and Molecular Medicine, Division of Hematology/Oncology, Boston Children' s Hospital, MA 02116 (United States); Rossi, Derrick J. [Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138 (United States); Department of Pediatrics, Harvard Medical School, Boston, MA 02115 (United States); Program in Cellular and Molecular Medicine, Division of Hematology/Oncology, Boston Children' s Hospital, MA 02116 (United States)

    2014-12-10

    Aging is invariably associated with alterations of the hematopoietic stem cell (HSC) compartment, including loss of functional capacity, altered clonal composition, and changes in lineage contribution. Although accumulation of DNA damage occurs during HSC aging, it is unlikely such consistent aging phenotypes could be solely attributed to changes in DNA integrity. Another mechanism by which heritable traits could contribute to the changes in the functional potential of aged HSCs is through alterations in the epigenetic landscape of adult stem cells. Indeed, recent studies on hematopoietic stem cells have suggested that altered epigenetic profiles are associated with HSC aging and play a key role in modulating the functional potential of HSCs at different stages during ontogeny. Even small changes of the epigenetic landscape can lead to robustly altered expression patterns, either directly by loss of regulatory control or through indirect, additive effects, ultimately leading to transcriptional changes of the stem cells. Potential drivers of such changes in the epigenetic landscape of aged HSCs include proliferative history, DNA damage, and deregulation of key epigenetic enzymes and complexes. This review will focus largely on the two most characterized epigenetic marks – DNA methylation and histone modifications – but will also discuss the potential role of non-coding RNAs in regulating HSC function during aging.

  8. Spreading convulsions, spreading depolarization and epileptogenesis in human cerebral cortex

    DEFF Research Database (Denmark)

    Dreier, Jens P; Major, Sebastian; Pannek, Heinz-Wolfgang

    2012-01-01

    Spreading depolarization of cells in cerebral grey matter is characterized by massive ion translocation, neuronal swelling and large changes in direct current-coupled voltage recording. The near-complete sustained depolarization above the inactivation threshold for action potential generating...... stimulations. Eventually, epileptic field potentials were recorded during the period that had originally seen spreading depression of activity. Such spreading convulsions are characterized by epileptic field potentials on the final shoulder of the large slow potential change of spreading depolarization. We...

  9. Electrically Reversible Redox-Switchable Polydopamine Films for Regulating Cell Behavior

    International Nuclear Information System (INIS)

    Tan, Guoxin; Liu, Yan; Wu, Yuxuan; Ouyang, Kongyou; Zhou, Lei; Yu, Peng; Liao, Jinwen; Ning, Chengyun

    2017-01-01

    Highlights: • The phenolic/quinone groups on polydopamine can redox-switchable reversible under electrical stimulation. • The quinone groups on PDA (oxidized PDA) enhanced cell spreading and proliferation. • The phenolic groups on PDA (reduced PDA) induced cell differentiation. - Abstract: Switchable surfaces that respond to external stimuli are important for regulating cell behavior. The results herein suggest that the redox process of polydopamine (PDA) is a switching reaction between oxidized polydopamine and reduced polydopamine, involving an interconversion of coupled two-proton (2H + ) and two-electron (2e − ) processes. The redox-switchable reversible surface potential arising from the potential-tunable redox reaction of the phenolic and quinone groups on PDA on titanium induced both cell adhesion and spreading. In vitro experiments demonstrated that the quinone groups on PDA greatly enhanced pre-osteoblasts MC3T3-E1 cell spreading and proliferation. Phenolic groups enhanced the induction of differentiation. The proposed methodology may allow further investigation of switchable surfaces for biological and medical applications.

  10. The V domain of dog PVRL4 (nectin-4) mediates canine distemper virus entry and virus cell-to-cell spread

    International Nuclear Information System (INIS)

    Delpeut, Sebastien; Noyce, Ryan S.; Richardson, Christopher D.

    2014-01-01

    The entry of canine distemper virus (CDV) is a multistep process that involves the attachment of CDV hemagglutinin (H) to its cellular receptor, followed by fusion between virus and cell membranes. Our laboratory recently identified PVRL4 (nectin-4) to be the epithelial receptor for measles and canine distemper viruses. In this study, we demonstrate that the V domain of PVRL4 is critical for CDV entry and virus cell-to-cell spread. Furthermore, four key amino acid residues within the V domain of dog PVRL4 and two within the CDV hemagglutinin were shown to be essential for receptor-mediated virus entry. - Highlights: • PVRL4 (nectin-4) is the epithelial cell receptor for measles and canine distemper viruses. • V domain of PVRL4 is critical for CDV entry, cell-to-cell spread, and syncytia formation. • Chimeric PVRL1 backbone substituted with the V domain of PVRL4 can function as a receptor. • Amino acids (F132/P133/A134/G135) within the V domain are essential for PVRL4 receptor activity. • Amino acids (P493/Y539) within CDV H protein are essential for PVRL4 receptor interaction

  11. The V domain of dog PVRL4 (nectin-4) mediates canine distemper virus entry and virus cell-to-cell spread

    Energy Technology Data Exchange (ETDEWEB)

    Delpeut, Sebastien; Noyce, Ryan S. [The Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5 (Canada); IWK Health Centre, Canadian Center for Vaccinology, Goldbloom Pavilion, Halifax, Nova Scotia, Canada B3H 1X5 (Canada); Richardson, Christopher D., E-mail: chris.richardson@dal.ca [The Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5 (Canada); IWK Health Centre, Canadian Center for Vaccinology, Goldbloom Pavilion, Halifax, Nova Scotia, Canada B3H 1X5 (Canada); The Department of Pediatrics, Dalhousie University, Halifax, Nova Scotia (Canada)

    2014-04-15

    The entry of canine distemper virus (CDV) is a multistep process that involves the attachment of CDV hemagglutinin (H) to its cellular receptor, followed by fusion between virus and cell membranes. Our laboratory recently identified PVRL4 (nectin-4) to be the epithelial receptor for measles and canine distemper viruses. In this study, we demonstrate that the V domain of PVRL4 is critical for CDV entry and virus cell-to-cell spread. Furthermore, four key amino acid residues within the V domain of dog PVRL4 and two within the CDV hemagglutinin were shown to be essential for receptor-mediated virus entry. - Highlights: • PVRL4 (nectin-4) is the epithelial cell receptor for measles and canine distemper viruses. • V domain of PVRL4 is critical for CDV entry, cell-to-cell spread, and syncytia formation. • Chimeric PVRL1 backbone substituted with the V domain of PVRL4 can function as a receptor. • Amino acids (F132/P133/A134/G135) within the V domain are essential for PVRL4 receptor activity. • Amino acids (P493/Y539) within CDV H protein are essential for PVRL4 receptor interaction.

  12. N-wasp is essential for the negative regulation of B cell receptor signaling.

    Directory of Open Access Journals (Sweden)

    Chaohong Liu

    2013-11-01

    Full Text Available Negative regulation of receptor signaling is essential for controlling cell activation and differentiation. In B-lymphocytes, the down-regulation of B-cell antigen receptor (BCR signaling is critical for suppressing the activation of self-reactive B cells; however, the mechanism underlying the negative regulation of signaling remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we demonstrate that neuronal Wiskott-Aldrich syndrome protein (N-WASP, which is coexpressed with WASP in all immune cells, is a critical negative regulator of B-cell signaling. B-cell-specific N-WASP gene deletion causes enhanced and prolonged BCR signaling and elevated levels of autoantibodies in the mouse serum. The increased signaling in N-WASP knockout B cells is concurrent with increased accumulation of F-actin at the B-cell surface, enhanced B-cell spreading on the antigen-presenting membrane, delayed B-cell contraction, inhibition in the merger of signaling active BCR microclusters into signaling inactive central clusters, and a blockage of BCR internalization. Upon BCR activation, WASP is activated first, followed by N-WASP in mouse and human primary B cells. The activation of N-WASP is suppressed by Bruton's tyrosine kinase-induced WASP activation, and is restored by the activation of SH2 domain-containing inositol 5-phosphatase that inhibits WASP activation. Our results reveal a new mechanism for the negative regulation of BCR signaling and broadly suggest an actin-mediated mechanism for signaling down-regulation.

  13. Defined α-synuclein prion-like molecular assemblies spreading in cell culture.

    Science.gov (United States)

    Aulić, Suzana; Le, Tran Thanh Nhat; Moda, Fabio; Abounit, Saïda; Corvaglia, Stefania; Casalis, Loredana; Gustincich, Stefano; Zurzolo, Chiara; Tagliavini, Fabrizio; Legname, Giuseppe

    2014-06-04

    α-Synuclein (α-syn) plays a central role in the pathogenesis of synucleinopathies, a group of neurodegenerative disorders that includes Parkinson disease, dementia with Lewy bodies and multiple system atrophy. Several findings from cell culture and mouse experiments suggest intercellular α-syn transfer. Through a methodology used to obtain synthetic mammalian prions, we tested whether recombinant human α-syn amyloids can promote prion-like accumulation in neuronal cell lines in vitro. A single exposure to amyloid fibrils of human α-syn was sufficient to induce aggregation of endogenous α-syn in human neuroblastoma SH-SY5Y cells. Remarkably, endogenous wild-type α-syn was sufficient for the formation of these aggregates, and overexpression of the protein was not required. Our results provide compelling evidence that endogenous α-syn can accumulate in cell culture after a single exposure to exogenous α-syn short amyloid fibrils. Importantly, using α-syn short amyloid fibrils as seed, endogenous α-syn aggregates and accumulates over several passages in cell culture, providing an excellent tool for potential therapeutic screening of pathogenic α-syn aggregates.

  14. Molecular regulation of plant cell wall extensibility

    Science.gov (United States)

    Cosgrove, D. J.

    1998-01-01

    Gravity responses in plants often involve spatial and temporal changes in cell growth, which is regulated primarily by controlling the ability of the cell wall to extend. The wall is thought to be a cellulose-hemicellulose network embedded in a hydrated matrix of complex polysaccharides and a small amount of structural protein. The wall extends by a form of polymer creep, which is mediated by expansins, a novel group of wall-loosening proteins. Expansins were discovered during a molecular dissection of the "acid growth" behavior of cell walls. Expansin alters the rheology of plant walls in profound ways, yet its molecular mechanism of action is still uncertain. It lacks detectable hydrolytic activity against the major components of the wall, but it is able to disrupt noncovalent adhesion between wall polysaccharides. The discovery of a second family of expansins (beta-expansins) sheds light on the biological role of a major group of pollen allergens and implies that expansins have evolved for diverse developmental functions. Finally, the contribution of other processes to wall extensibility is briefly summarized.

  15. Spread of human T-cell leukemia virus (HTLV-I) in the Dutch homosexual community

    NARCIS (Netherlands)

    Goudsmit, J.; de Wolf, F.; van de Wiel, B.; Smit, L.; Bakker, M.; Albrecht-van Lent, N.; Coutinho, R. A.

    1987-01-01

    Sequential sera of 697 homosexual men, participating in a prospective study (1984-1986) of the risk to acquire human immunodeficiency virus (HIV) or AIDS, were tested for antibodies to human T-cell leukaemia virus (HTLV-I) by particle agglutination and immunoblotting. No intravenous drug users were

  16. Redox Regulation of Endothelial Cell Fate

    Science.gov (United States)

    Song, Ping; Zou, Ming-Hui

    2014-01-01

    Endothelial cells (ECs) are present throughout blood vessels and have variable roles in both physiological and pathological settings. EC fate is altered and regulated by several key factors in physiological or pathological conditions. Reactive nitrogen species and reactive oxygen species derived from NAD(P)H oxidases, mitochondria, or nitric oxide-producing enzymes are not only cytotoxic but also compose a signaling network in the redox system. The formation, actions, key molecular interactions, and physiological and pathological relevance of redox signals in ECs remain unclear. We review the identities, sources, and biological actions of oxidants and reductants produced during EC function or dysfunction. Further, we discuss how ECs shape key redox sensors and examine the biological functions, transcriptional responses, and post-translational modifications evoked by the redox system in ECs. We summarize recent findings regarding the mechanisms by which redox signals regulate the fate of ECs and address the outcome of altered EC fate in health and disease. Future studies will examine if the redox biology of ECs can be targeted in pathophysiological conditions. PMID:24633153

  17. Integrating physiological regulation with stem cell and tissue homeostasis

    Science.gov (United States)

    Nakada, Daisuke; Levi, Boaz P.; Morrison, Sean J.

    2015-01-01

    Summary Stem cells are uniquely able to self-renew, to undergo multilineage differentiation, and to persist throughout life in a number of tissues. Stem cells are regulated by a combination of shared and tissue-specific mechanisms and are distinguished from restricted progenitors by differences in transcriptional and epigenetic regulation. Emerging evidence suggests that other aspects of cellular physiology, including mitosis, signal transduction, and metabolic regulation also differ between stem cells and their progeny. These differences may allow stem cells to be regulated independently of differentiated cells in response to circadian rhythms, changes in metabolism, diet, exercise, mating, aging, infection, and disease. This allows stem cells to sustain homeostasis or to remodel relevant tissues in response to physiological change. Stem cells are therefore not only regulated by short-range signals that maintain homeostasis within their tissue of origin, but also by long-range signals that integrate stem cell function with systemic physiology. PMID:21609826

  18. Regulation of Arabidopsis Early Anther Development by Putative Cell-Cell Signaling Molecules and Transcriptional Regulators

    Institute of Scientific and Technical Information of China (English)

    Yu-Jin Sun; Carey LH Hord; Chang-Bin Chen; Hong Ma

    2007-01-01

    Anther development in flowering plants involves the formation of several cell types, including the tapetal and pollen mother cells. The use of genetic and molecular tools has led to the identification and characterization of genes that are critical for normal cell division and differentiation in Arabidopsis early anther development. We review here several recent studies on these genes, including the demonstration that the putative receptor protein kinases BAM1 and BAM2 together play essential roles in the control of early cell division and differentiation. In addition, we discuss the hypothesis that BAM1/2 may form a positive-negative feedback regulatory loop with a previously identified key regulator, SPOROCYTELESS (also called NOZZLE),to control the balance between sporogenous and somatic cell types in the anther. Furthermore, we summarize the isolation and functional analysis of the DYSFUNCTIONAL TAPETUM1 (DYT1) gene in promoting proper tapetal cell differentiation. Our finding that DYT1 encodes a putative transcription factor of the bHLH family, as well as relevant expression analyses, strongly supports a model that DYT1 serves as a critical link between upstream factors and downstream target genes that are critical for normal tapetum development and function. These studies, together with other recently published works, indicate that cell-cell communication and transcriptional control are key processes essential for cell fate specification in anther development.

  19. Preparation and regulating cell adhesion of anion-exchangeable layered double hydroxide micropatterned arrays.

    Science.gov (United States)

    Yao, Feng; Hu, Hao; Xu, Sailong; Huo, Ruijie; Zhao, Zhiping; Zhang, Fazhi; Xu, Fujian

    2015-02-25

    We describe a reliable preparation of MgAl-layered double hydroxide (MgAl-LDH) micropatterned arrays on gold substrate by combining SO3(-)-terminated self-assembly monolayer and photolithography. The synthesis route is readily extended to prepare LDH arrays on the SO3(-)-terminated polymer-bonded glass substrate amenable for cell imaging. The anion-exchangeable MgAl-LDH micropattern can act both as bioadhesive region for selective cell adhesion and as nanocarrier for drug molecules to regulate cell behaviors. Quantitative analysis of cell adhesion shows that selective HepG2 cell adhesion and spreading are promoted by the micropatterned MgAl-LDH, and also suppressed by methotrexate drug released from the LDH interlayer galleries.

  20. Authoritative regulation and the stem cell debate.

    Science.gov (United States)

    Capps, Benjamin

    2008-01-01

    In this paper I argue that liberal democratic communities are justified in regulating the activities of their members because of the inevitable existence of conflicting conceptions of what is considered as morally right. This will often lead to tension and disputes, and in such circumstances, reliance on peaceful or orderly co-existence will not normally suffice. In such pluralistic societies, the boundary between permissible and impermissible activities will be unclear; and this becomes a particular concern in controversial issues which raise specific anxieties and uncertainty. One context that has repeatedly raised issues in this regard is that of biotechnology and, in particular, the recent stem cell debate, on which this paper concentrates. While such developments have the potential to make significant improvements to therapeutic progress, we should also be sceptical because predicting the impact of these developments remains uncertain and complex. For the sake of socio-political stability, it will therefore be necessary to enact and enforce rules which limit these competing claims in public policy but which may not be compatible with what individual moral commitments ideally permit. One way to achieve this is to establish procedural frameworks to resolve potential disputes in the public sphere about what is right, wrong, or permissible conduct. I argue that for one to commit to authoritative regulation, an idea of harm prevention through state intervention is necessary; and that this requires optimum mechanisms of procedure which allow the individual the opportunity to compromise and yet to continue to oppose or fight for changes as demanded by his or her moral position.

  1. Perineural Spread of Renal Cell Carcinoma: A Case Illustration with a Proposed Anatomic Mechanism and a Review of the Literature.

    Science.gov (United States)

    Capek, Stepan; Krauss, William E; Amrami, Kimberly K; Parisi, Joseph E; Spinner, Robert J

    2016-05-01

    Perineural spread (PNS) is an unusual mechanism of tumor extension and has been typically reported in squamous cell carcinoma, adenocystic carcinoma, and desmoplastic melanoma. Our group has previously demonstrated PNS in rectal, prostate, bladder, and cervical cancer from the primary site along the autonomic nerves to the major somatic nerves and even intradurally. We believe similar principles apply to renal cell carcinoma (RCC) as well, despite the different anatomy. We performed a retrospective search to identify cases of intradural-extramedullary metastases of RCC caused by PNS. Strict anatomic and imaging inclusion criteria were defined: only lesions located between T6 and L3 were included, and PNS as a potential cause had to be supported by imaging evidence. Although 3 cases of spinal intradural metastases were identified, only one met our strict inclusion criteria. A 61-year-old woman developed a late intradural-extramedullary metastasis of RCC 16 years after the original diagnosis that we believe represents an example of visceral organ PNS. RCC can propagate via PNS from the primary tumor along the autonomic nerves to the aorticorenal, celiac, and mesenteric ganglia and then along the thoracic and lumbar splanchnic nerves to the corresponding spinal nerves and intradurally. We present radiologic evidence together with the review of the literature to support the premise that PNS of RCC not only occurs but goes unrecognized. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. Regulation of Water in Plant Cells

    Science.gov (United States)

    Kowles, Richard V.

    2010-01-01

    Cell water relationships are important topics to be included in cell biology courses. Differences exist in the control of water relationships in plant cells relative to control in animal cells. One important reason for these differences is that turgor pressure is a consideration in plant cells. Diffusion and osmosis are the underlying factors…

  3. Innate lymphoid cells in autoimmunity: emerging regulators in rheumatic diseases

    NARCIS (Netherlands)

    Shikhagaie, Medya M.; Germar, Kristine; Bal, Suzanne M.; Ros, Xavier Romero; Spits, Hergen

    2017-01-01

    Innate lymphoid cells (ILCs) are important in the regulation of barrier homeostasis. These cells do not express T cell receptors but share many functional similarities with T helper cells and cytotoxic CD8(+) T lymphocytes. ILCs are divided into three groups, namely group 1 ILCs, group 2 ILCs and

  4. Mitochondrial regulation of cell death: a phylogenetically conserved control

    Directory of Open Access Journals (Sweden)

    Lorenzo Galluzzi

    2016-02-01

    Full Text Available Mitochondria are fundamental for eukaryotic cells as they participate in critical catabolic and anabolic pathways. Moreover, mitochondria play a key role in the signal transduction cascades that precipitate many (but not all regulated variants of cellular demise. In this short review, we discuss the differential implication of mitochondria in the major forms of regulated cell death.

  5. Regulation of Autophagy by Glucose in Mammalian Cells

    OpenAIRE

    Moruno, Félix; Pérez-Jiménez, Eva; Knecht, Erwin

    2012-01-01

    Autophagy is an evolutionarily conserved process that contributes to maintain cell homeostasis. Although it is strongly regulated by many extracellular factors, induction of autophagy is mainly produced by starvation of nutrients. In mammalian cells, the regulation of autophagy by amino acids, and also by the hormone insulin, has been extensively investigated, but knowledge about the effects of other autophagy regulators, including another nutrient, glucose, is more limited. Here we will focu...

  6. LHP1 Regulates H3K27me3 Spreading and Shapes the Three-Dimensional Conformation of the Arabidopsis Genome

    KAUST Repository

    Veluchamy, Alaguraj; Jé gu, Teddy; Ariel, Federico; Latrasse, David; Mariappan, Kiruthiga Gayathri; Kim, SoonKap; Crespi, Martin; Hirt, Heribert; Bergounioux, Catherine; Raynaud, Cé cile; Benhamed, Moussa

    2016-01-01

    Precise expression patterns of genes in time and space are essential for proper development of multicellular organisms. Dynamic chromatin conformation and spatial organization of the genome constitute a major step in this regulation to modulate developmental outputs. Polycomb repressive complexes (PRCs) mediate stable or flexible gene repression in response to internal and environmental cues. In Arabidopsis thaliana, LHP1 co-localizes with H3K27me3 epigenetic marks throughout the genome and interacts with PRC1 and PRC2 members as well as with a long noncoding RNA. Here, we show that LHP1 is responsible for the spreading of H3K27me3 towards the 3’ end of the gene body. We also identified a subset of LHP1-activated genes and demonstrated that LHP1 shapes local chromatin topology in order to control transcriptional co-regulation. Our work reveals a general role of LHP1 from local to higher conformation levels of chromatin configuration to determine its accessibility to define gene expression patterns.

  7. LHP1 Regulates H3K27me3 Spreading and Shapes the Three-Dimensional Conformation of the Arabidopsis Genome.

    Directory of Open Access Journals (Sweden)

    Alaguraj Veluchamy

    Full Text Available Precise expression patterns of genes in time and space are essential for proper development of multicellular organisms. Dynamic chromatin conformation and spatial organization of the genome constitute a major step in this regulation to modulate developmental outputs. Polycomb repressive complexes (PRCs mediate stable or flexible gene repression in response to internal and environmental cues. In Arabidopsis thaliana, LHP1 co-localizes with H3K27me3 epigenetic marks throughout the genome and interacts with PRC1 and PRC2 members as well as with a long noncoding RNA. Here, we show that LHP1 is responsible for the spreading of H3K27me3 towards the 3' end of the gene body. We also identified a subset of LHP1-activated genes and demonstrated that LHP1 shapes local chromatin topology in order to control transcriptional co-regulation. Our work reveals a general role of LHP1 from local to higher conformation levels of chromatin configuration to determine its accessibility to define gene expression patterns.

  8. LHP1 Regulates H3K27me3 Spreading and Shapes the Three-Dimensional Conformation of the Arabidopsis Genome

    KAUST Repository

    Veluchamy, Alaguraj

    2016-07-13

    Precise expression patterns of genes in time and space are essential for proper development of multicellular organisms. Dynamic chromatin conformation and spatial organization of the genome constitute a major step in this regulation to modulate developmental outputs. Polycomb repressive complexes (PRCs) mediate stable or flexible gene repression in response to internal and environmental cues. In Arabidopsis thaliana, LHP1 co-localizes with H3K27me3 epigenetic marks throughout the genome and interacts with PRC1 and PRC2 members as well as with a long noncoding RNA. Here, we show that LHP1 is responsible for the spreading of H3K27me3 towards the 3’ end of the gene body. We also identified a subset of LHP1-activated genes and demonstrated that LHP1 shapes local chromatin topology in order to control transcriptional co-regulation. Our work reveals a general role of LHP1 from local to higher conformation levels of chromatin configuration to determine its accessibility to define gene expression patterns.

  9. Regulation of T cell responses in atherosclerosis

    NARCIS (Netherlands)

    Puijvelde, Gijsbrecht Henricus Maria van

    2007-01-01

    One of the most important characteristics of atherosclerosis is the chronic inflammatory response in which T cells and NKT cells are very important. In this thesis several methods to modulate the activity of these T and NKT cells in atherosclerosis are described. The induction of regulatory T cells

  10. Effect of plasma surface functionalization on preosteoblast cells spreading and adhesion on a biomimetic hydroxyapatite layer formed on a titanium surface

    International Nuclear Information System (INIS)

    Myung, Sung Woon; Ko, Yeong Mu; Kim, Byung Hoon

    2013-01-01

    This study examined the plasma surface modification of biomimetic hydroxyapatite (HAp) formed on a titanium (Ti) surface as well as its influence on the behavior of preosteoblast cells. Ti substrates pre-treated with a plasma-polymerized thin film rich in carboxyl groups were subjected to a biomimetic process in a simulated body fluid solution to synthesize the HAp. The HAp layer grown on Ti substrate was then coated with two types of plasma polymerized acrylic acid and allyl amine thin film. The different types of Ti substrates were characterized by attenuated total reflection Fourier transform infrared spectroscopy, energy dispersive spectroscopy and X-ray diffraction. HAp with a Ca/P ratio from 1.25 to 1.38 was obtained on the Ti substrate and hydrophilic carboxyl (-COOH) and amine (-NH 2 ) functional groups were introduced to its surface. Scanning electron microscopy was used to observe the surface of the HAp coatings and the morphology of MC3T3-E1 cells. These results showed that the -COOH-modified HAp surfaces promoted the cell spreading synergistically by changing the surface morphology and chemical state.-NH 2 modified HAp had the lowest cell spreading and proliferation compared to HAp and -COOH-modified HAp. These results correspond to fluorescein analysis, which showed many more cell spreading of COOH/HAp/Ti surface compared to HAp and NH 2 modified HAp. A MTT assay was used to evaluate cell proliferation. The results showed that the proliferation of MC3T3-E1 cells increased in the order of COOH/HAp/Ti > HAp/Ti > NH 2 /Ti > Ti, corresponding to the effect of cell spreading for 6 days. The change in morphology and the chemical surface properties of the biomaterial via plasma polymerization can affect the behavior of MC3T3-E1 cells.

  11. Effect of plasma surface functionalization on preosteoblast cells spreading and adhesion on a biomimetic hydroxyapatite layer formed on a titanium surface

    Energy Technology Data Exchange (ETDEWEB)

    Myung, Sung Woon; Ko, Yeong Mu; Kim, Byung Hoon, E-mail: kim5055@chosun.ac.kr

    2013-12-15

    This study examined the plasma surface modification of biomimetic hydroxyapatite (HAp) formed on a titanium (Ti) surface as well as its influence on the behavior of preosteoblast cells. Ti substrates pre-treated with a plasma-polymerized thin film rich in carboxyl groups were subjected to a biomimetic process in a simulated body fluid solution to synthesize the HAp. The HAp layer grown on Ti substrate was then coated with two types of plasma polymerized acrylic acid and allyl amine thin film. The different types of Ti substrates were characterized by attenuated total reflection Fourier transform infrared spectroscopy, energy dispersive spectroscopy and X-ray diffraction. HAp with a Ca/P ratio from 1.25 to 1.38 was obtained on the Ti substrate and hydrophilic carboxyl (-COOH) and amine (-NH{sub 2}) functional groups were introduced to its surface. Scanning electron microscopy was used to observe the surface of the HAp coatings and the morphology of MC3T3-E1 cells. These results showed that the -COOH-modified HAp surfaces promoted the cell spreading synergistically by changing the surface morphology and chemical state.-NH{sub 2} modified HAp had the lowest cell spreading and proliferation compared to HAp and -COOH-modified HAp. These results correspond to fluorescein analysis, which showed many more cell spreading of COOH/HAp/Ti surface compared to HAp and NH{sub 2} modified HAp. A MTT assay was used to evaluate cell proliferation. The results showed that the proliferation of MC3T3-E1 cells increased in the order of COOH/HAp/Ti > HAp/Ti > NH{sub 2}/Ti > Ti, corresponding to the effect of cell spreading for 6 days. The change in morphology and the chemical surface properties of the biomaterial via plasma polymerization can affect the behavior of MC3T3-E1 cells.

  12. Cell shape and spreading of stromal (mesenchymal) stem cells cultured on fibronectin coated gold and hydroxyapatite surfaces

    DEFF Research Database (Denmark)

    Dolatshahi-Pirouz, A; Jensen, Thomas Hartvig Lindkjær; Kolind, Kristian

    2011-01-01

    In order to identify the cellular mechanisms leading to the biocompatibility of hydroxyapatite implants, we studied the interaction of human bone marrow derived stromal (mesenchymal) stem cells (hMSCs) with fibronectin-coated gold (Au) and hydroxyapatite (HA) surfaces. The adsorption of fibronectin...

  13. Mitochondrial apoptotic pathways induced by Drosophila programmed cell death regulators

    International Nuclear Information System (INIS)

    Claveria, Cristina; Torres, Miguel

    2003-01-01

    Multicellular organisms eliminate unwanted or damaged cells by cell death, a process essential to the maintenance of tissue homeostasis. Cell death is a tightly regulated event, whose alteration by excess or defect is involved in the pathogenesis of many diseases such as cancer, autoimmune syndromes, and neurodegenerative processes. Studies in model organisms, especially in the nematode Caenorhabditis elegans, have been crucial in identifying the key molecules implicated in the regulation and execution of programmed cell death. In contrast, the study of cell death in Drosophila melanogaster, often an excellent model organism, has identified regulators and mechanisms not obviously conserved in other metazoans. Recent molecular and cellular analyses suggest, however, that the mechanisms of action of the main programmed cell death regulators in Drosophila include a canonical mitochondrial pathway

  14. Redox regulation of cell proliferation: Bioinformatics and redox proteomics approaches to identify redox-sensitive cell cycle regulators.

    Science.gov (United States)

    Foyer, Christine H; Wilson, Michael H; Wright, Megan H

    2018-03-29

    Plant stem cells are the foundation of plant growth and development. The balance of quiescence and division is highly regulated, while ensuring that proliferating cells are protected from the adverse effects of environment fluctuations that may damage the genome. Redox regulation is important in both the activation of proliferation and arrest of the cell cycle upon perception of environmental stress. Within this context, reactive oxygen species serve as 'pro-life' signals with positive roles in the regulation of the cell cycle and survival. However, very little is known about the metabolic mechanisms and redox-sensitive proteins that influence cell cycle progression. We have identified cysteine residues on known cell cycle regulators in Arabidopsis that are potentially accessible, and could play a role in redox regulation, based on secondary structure and solvent accessibility likelihoods for each protein. We propose that redox regulation may function alongside other known posttranslational modifications to control the functions of core cell cycle regulators such as the retinoblastoma protein. Since our current understanding of how redox regulation is involved in cell cycle control is hindered by a lack of knowledge regarding both which residues are important and how modification of those residues alters protein function, we discuss how critical redox modifications can be mapped at the molecular level. Crown Copyright © 2018. Published by Elsevier Inc. All rights reserved.

  15. Regulation of Stem Cell Differentiation by Histone Methyltransferases and Demethylases

    DEFF Research Database (Denmark)

    Pasini, D; Bracken, A P; Agger, K

    2008-01-01

    The generation of different cell types from stem cells containing identical genetic information and their organization into tissues and organs during development is a highly complex process that requires defined transcriptional programs. Maintenance of such programs is epigenetically regulated...... and the factors involved in these processes are often essential for development. The activities required for cell-fate decisions are frequently deregulated in human tumors, and the elucidation of the molecular mechanisms that regulate these processes is therefore important for understanding both developmental...

  16. Retinoic acid signalling in thymocytes regulates T cell development

    DEFF Research Database (Denmark)

    Wendland, Kerstin; Sitnik, Katarzyna Maria; Kotarsky, Knut

    in the regulatory regions of targetgenes. RA has been reported to play a direct role in regulating multiple aspects of peripheralT cell responses1, but whether endogenous RA signalling occurs in developingthymocytes and the potential impact of such signals in regulating T cell developmentremains unclear. To address......RARα. This blocks RA signalling in developing thymocytes from the DN3/4 stageonwards and thus allows us to study the role of RA in T cell development...

  17. Extracellular Matrix as a Regulator of Epidermal Stem Cell Fate.

    Science.gov (United States)

    Chermnykh, Elina; Kalabusheva, Ekaterina; Vorotelyak, Ekaterina

    2018-03-27

    Epidermal stem cells reside within the specific anatomic location, called niche, which is a microenvironment that interacts with stem cells to regulate their fate. Regulation of many important processes, including maintenance of stem cell quiescence, self-renewal, and homeostasis, as well as the regulation of division and differentiation, are common functions of the stem cell niche. As it was shown in multiple studies, extracellular matrix (ECM) contributes a lot to stem cell niches in various tissues, including that of skin. In epidermis, ECM is represented, primarily, by a highly specialized ECM structure, basement membrane (BM), which separates the epidermal and dermal compartments. Epidermal stem cells contact with BM, but when they lose the contact and migrate to the overlying layers, they undergo terminal differentiation. When considering all of these factors, ECM is of fundamental importance in regulating epidermal stem cells maintenance, proper mobilization, and differentiation. Here, we summarize the remarkable progress that has recently been made in the research of ECM role in regulating epidermal stem cell fate, paying special attention to the hair follicle stem cell niche. We show that the destruction of ECM components impairs epidermal stem cell morphogenesis and homeostasis. A deep understanding of ECM molecular structure as well as the development of in vitro system for stem cell maintaining by ECM proteins may bring us to developing new approaches for regenerative medicine.

  18. Expression profiling of genes regulated by TGF-beta: Differential regulation in normal and tumour cells

    Directory of Open Access Journals (Sweden)

    Takahashi Takashi

    2007-04-01

    Full Text Available Abstract Background TGF-beta is one of the key cytokines implicated in various disease processes including cancer. TGF-beta inhibits growth and promotes apoptosis in normal epithelial cells and in contrast, acts as a pro-tumour cytokine by promoting tumour angiogenesis, immune-escape and metastasis. It is not clear if various actions of TGF-beta on normal and tumour cells are due to differential gene regulations. Hence we studied the regulation of gene expression by TGF-beta in normal and cancer cells. Results Using human 19 K cDNA microarrays, we show that 1757 genes are exclusively regulated by TGF-beta in A549 cells in contrast to 733 genes exclusively regulated in HPL1D cells. In addition, 267 genes are commonly regulated in both the cell-lines. Semi-quantitative and real-time qRT-PCR analysis of some genes agrees with the microarray data. In order to identify the signalling pathways that influence TGF-beta mediated gene regulation, we used specific inhibitors of p38 MAP kinase, ERK kinase, JNK kinase and integrin signalling pathways. The data suggest that regulation of majority of the selected genes is dependent on at least one of these pathways and this dependence is cell-type specific. Interestingly, an integrin pathway inhibitor, RGD peptide, significantly affected TGF-beta regulation of Thrombospondin 1 in A549 cells. Conclusion These data suggest major differences with respect to TGF-beta mediated gene regulation in normal and transformed cells and significant role of non-canonical TGF-beta pathways in the regulation of many genes by TGF-beta.

  19. Fibulin-1 suppression of fibronectin-regulated cell adhesion and motility.

    Science.gov (United States)

    Twal, W O; Czirok, A; Hegedus, B; Knaak, C; Chintalapudi, M R; Okagawa, H; Sugi, Y; Argraves, W S

    2001-12-01

    Fibulin-1 is an extracellular matrix protein often associated with fibronectin (FN) in vivo. In this study, the ability of fibulin-1 to modulate adhesion, spreading and motility-promoting activities of FN was investigated. Fibulin-1 was found to have pronounced inhibitory effects on the cell attachment and spreading promoted by FN. Fibulin-1 was also found to inhibit the motility of a variety of cell types on FN substrata. For example, the FN-dependent haptotactic motility of breast carcinoma (MDA MB231) cells, epidermal carcinoma (A431), melanoma (A375 SM), rat pulmonary aortic smooth muscle cells (PAC1) and Chinese hamster ovary (CHO) cells was inhibited by the presence of fibulin-1 bound to FN-coated Boyden chamber membranes. Cells transfected to overproduce fibulin-1 displayed reduced velocity, distance of movement and persistence time on FN substrata. Similarly, the incorporation of fibulin-1 into FN-containing type I collagen gels inhibited the invasion of endocardial cushion mesenchymal cells migrating from cultured embryonic heart explants. By contrast, incorporation of fibulin-1 into collagen gels lacking FN had no effect on the migration of endocardial cushion cells. These results suggest that the motility-suppressive effects of fibulin-1 might be FN specific. Furthermore, such effects are cell-type specific, in that the migration of gingival fibroblasts and endothelial cells on FN substrata is not responsive to fibulin-1. Additional studies found that the mechanism for the motility-suppressive effects of fibulin-1 does not involve perturbations of interactions between alpha5beta1 or alpha4 integrins, or heparan sulfate proteoglycans with FN. However, fibulin-1 was found to inhibit extracellular signal regulated kinase (ERK) activation and to suppress phosphorylation of myosin heavy chain. This ability to influence signal transduction cascades that modulate the actin-myosin motor complex might be the basis for the effects of fibulin-1 on adhesion and

  20. Streptomyces sporulation - Genes and regulators involved in bacterial cell differentiation

    OpenAIRE

    Larsson, Jessica

    2010-01-01

    Streptomycetes are Gram-positive bacteria with a complex developmental life cycle. They form spores on specialized cells called aerial hyphae, and this sporulation involves alterations in growth, morphogenesis and cell cycle processes like cell division and chromosome segregation. Understanding the developmental mechanisms that streptomycetes have evolved for regulating for example cell division is of general interest in bacterial cell biology. It can also be valuable in the design of new dru...

  1. Cytokinetics and Regulation of Progenitor Cells

    Energy Technology Data Exchange (ETDEWEB)

    Lajtha, L. G. [Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (United Kingdom)

    1967-07-15

    Full text: In spite of great differences in the life-span of fully differentiated haemic cells, the cellular kinetics of their production appears to be similar. Recent evidence indicates a common ultimate stem cell for most of the cells in the peripheral blood. The various pathways of differentiation, however, result in transient dividing and differentiating cell populations which differ from each other not only in their specific biochemical processes but also in the manner of control and kinetic pattern of their proliferation. The population best understood is the erythroid progenitor series of cells, primarily because it has the greatest number of experimentally measurable parameters at the present. This will be discussed in detail and comparisons will be made with the myeloid and lymphoid progenitor populations. The fine structure of the bone-marrow stem cell population will be examined in particular, with regard to the suitability or otherwise of the current stem cell models to explain the kinetic pattern of all the peripheral blood elements after perturbations of their steady-state values. Four different assay methods of bone-marrow stem cells have been examined with regard to the kinetic pattern following perturbation of the steady-state system, e.g. by irradiation. Basically, the stem cell assays fall into two categories: those depending on grafting haemopoietic cells into suitably treated recipients, and those in which recovery of the population is allowed in the animal in which the perturbation was produced, without handling the cells. Evidence is accumulating which indicates that in the grafting techniques, a selective loss of stem cells may occur, . especially stem cells in cell cycle, hence in early stages of recovery of the population unduly low numerical values might be noted. In view of this observation, the concept of the colony-forming cell may have to be revised and instead the colony-forming property of the stem cell introduced. (author)

  2. [Genetic regulation of plant shoot stem cells].

    Science.gov (United States)

    Al'bert, E V; Ezhova, T A

    2013-02-01

    This article describes the main features of plant stem cells and summarizes the results of studies of the genetic control of stem cell maintenance in the apical meristem of the shoot. It is demonstrated that the WUS-CLV gene system plays a key role in the maintenance of shoot apical stem cells and the formation of adventitious buds and somatic embryos. Unconventional concepts of plant stem cells are considered.

  3. Cytokinetics and Regulation of Progenitor Cells

    International Nuclear Information System (INIS)

    Lajtha, L.G.

    1967-01-01

    Full text: In spite of great differences in the life-span of fully differentiated haemic cells, the cellular kinetics of their production appears to be similar. Recent evidence indicates a common ultimate stem cell for most of the cells in the peripheral blood. The various pathways of differentiation, however, result in transient dividing and differentiating cell populations which differ from each other not only in their specific biochemical processes but also in the manner of control and kinetic pattern of their proliferation. The population best understood is the erythroid progenitor series of cells, primarily because it has the greatest number of experimentally measurable parameters at the present. This will be discussed in detail and comparisons will be made with the myeloid and lymphoid progenitor populations. The fine structure of the bone-marrow stem cell population will be examined in particular, with regard to the suitability or otherwise of the current stem cell models to explain the kinetic pattern of all the peripheral blood elements after perturbations of their steady-state values. Four different assay methods of bone-marrow stem cells have been examined with regard to the kinetic pattern following perturbation of the steady-state system, e.g. by irradiation. Basically, the stem cell assays fall into two categories: those depending on grafting haemopoietic cells into suitably treated recipients, and those in which recovery of the population is allowed in the animal in which the perturbation was produced, without handling the cells. Evidence is accumulating which indicates that in the grafting techniques, a selective loss of stem cells may occur, . especially stem cells in cell cycle, hence in early stages of recovery of the population unduly low numerical values might be noted. In view of this observation, the concept of the colony-forming cell may have to be revised and instead the colony-forming property of the stem cell introduced. (author)

  4. Equine herpesvirus type 1 (EHV-1) glycoprotein K is required for efficient cell-to-cell spread and virus egress

    International Nuclear Information System (INIS)

    Neubauer, Antonie; Osterrieder, Nikolaus

    2004-01-01

    The function of the equine herpesvirus type 1 (EHV-1) glycoprotein K (gK) homologue was investigated. Deletion of 88% of the UL53-homologous open reading frame in EHV-1 strain RacH resulted in a severe growth defect of the gK-negative virus (HΔgK) as reflected by a significant decrease in the production of infectious virus progeny on RK13 cells. The HΔgK virus induced only minute plaques, was unable to form syncytia, and its penetration efficiency into RK13 cells was reduced by approximately 40%. To further analyze gK function and intracellular trafficking, gK of strain RacH was replaced by a C-terminally truncated gK-green fluorescent protein fusion protein (gK-GFP). The generated recombinant virus was shown to replicate well on non-complementing cells, and virus penetration and syncytium formation were comparable to parental RacH. A reduction in plaque size and slightly decreased intra- and extracellular virus titers, however, were observed. The gK-GFP fusion protein was expressed with early-late kinetics, and multiple forms of the protein exhibiting M r s between 50,000 and 85,000 were detected by Western blot analysis. The various gK-GFP forms were shown to be N-glycosylated, associated with membranes of the Golgi apparatus, and were incorporated into extracellular virions. Complete processing of gK-GFP was only observed within the context of viral infection. From the results, we concluded that EHV-1 gK is required for efficient virus growth in vitro and that the carboxy-terminal amino acids are not required for its function, because the gK-GFP fusion protein was able to complement for EHV-1 growth in the absence of authentic gK

  5. T-cell regulation in lepromatous leprosy.

    Directory of Open Access Journals (Sweden)

    Kidist Bobosha

    2014-04-01

    Full Text Available Regulatory T (Treg cells are known for their role in maintaining self-tolerance and balancing immune reactions in autoimmune diseases and chronic infections. However, regulatory mechanisms can also lead to prolonged survival of pathogens in chronic infections like leprosy and tuberculosis (TB. Despite high humoral responses against Mycobacterium leprae (M. leprae, lepromatous leprosy (LL patients have the characteristic inability to generate T helper 1 (Th1 responses against the bacterium. In this study, we investigated the unresponsiveness to M. leprae in peripheral blood mononuclear cells (PBMC of LL patients by analysis of IFN-γ responses to M. leprae before and after depletion of CD25+ cells, by cell subsets analysis of PBMC and by immunohistochemistry of patients' skin lesions. Depletion of CD25+ cells from total PBMC identified two groups of LL patients: 7/18 (38.8% gained in vitro responsiveness towards M. leprae after depletion of CD25+ cells, which was reversed to M. leprae-specific T-cell unresponsiveness by addition of autologous CD25+ cells. In contrast, 11/18 (61.1% remained anergic in the absence of CD25+ T-cells. For both groups mitogen-induced IFN-γ was, however, not affected by depletion of CD25+ cells. In M. leprae responding healthy controls, treated lepromatous leprosy (LL and borderline tuberculoid leprosy (BT patients, depletion of CD25+ cells only slightly increased the IFN-γ response. Furthermore, cell subset analysis showed significantly higher (p = 0.02 numbers of FoxP3+ CD8+CD25+ T-cells in LL compared to BT patients, whereas confocal microscopy of skin biopsies revealed increased numbers of CD68+CD163+ as well as FoxP3+ cells in lesions of LL compared to tuberculoid and borderline tuberculoid leprosy (TT/BT lesions. Thus, these data show that CD25+ Treg cells play a role in M. leprae-Th1 unresponsiveness in LL.

  6. Fibronectin-calcium phosphate composite layer on hydroxyapatite to enhance adhesion, cell spread and osteogenic differentiation of human mesenchymal stem cells in vitro

    International Nuclear Information System (INIS)

    Sogo, Yu; Ito, Atsuo; Matsuno, Tomonori; Oyane, Ayako; Tamazawa, Gaku; Satoh, Tazuko; Yamazaki, Atsushi; Uchimura, Eiji; Ohno, Tadao

    2007-01-01

    Fibronectin (Fn) and type I collagen (Col) were immobilized on a surface of a hydroxyapatite (HAP) ceramic by coprecipitation with calcium phosphate in a supersaturated calcium phosphate solution prepared by mixing clinically approved infusion fluids. These proteins and the calcium phosphate precipitate formed a composite surface layer. As a result, the proteins were immobilized firmly as not to be released completely for 3 d in a physiological salt solution. When human mesenchymal stem cells (hMSCs) were cultured on a HAP ceramic in a differentiation medium supplemented with dexamethasone, β-glycerophosphate and ascorbic acid, hMSCs spread well within 1 h. The alkaline phosphatase (ALP) activity of hMSCs cultured on the Fn-calcium phosphate composite layer significantly increased compared with that of hMSCs cultured on the untreated HAP ceramic. On the other hand, Col did not increase the ALP activity of hMSCs and no synergy between Fn and Col was observed. Therefore, the Fn-calcium phosphate composite layer formed on the HAP is useful for the enhancement of the spreading and osteogenic differentiation of hMSCs in vitro

  7. Hemidesmosomal linker proteins regulate cell motility, invasion and tumorigenicity in oral squamous cell carcinoma derived cells.

    Science.gov (United States)

    Chaudhari, Pratik Rajeev; Charles, Silvania Emlit; D'Souza, Zinia Charlotte; Vaidya, Milind Murlidhar

    2017-11-15

    BPAG1e and Plectin are hemidesmosomal linker proteins which anchor intermediate filament proteins to the cell surface through β4 integrin. Recent reports indicate that these proteins play a role in various cellular processes apart from their known anchoring function. However, the available literature is inconsistent. Further, the previous study from our laboratory suggested that Keratin8/18 pair promotes cell motility and tumor progression by deregulating β4 integrin signaling in oral squamous cell carcinoma (OSCC) derived cells. Based on these findings, we hypothesized that linker proteins may have a role in neoplastic progression of OSCC. Downregulation of hemidesmosomal linker proteins in OSCC derived cells resulted in reduced cell migration accompanied by alterations in actin organization. Further, decreased MMP9 activity led to reduced cell invasion in linker proteins knockdown cells. Moreover, loss of these proteins resulted in reduced tumorigenic potential. SWATH analysis demonstrated upregulation of N-Myc downstream regulated gene 1 (NDRG1) in linker proteins downregulated cells as compared to vector control cells. Further, the defects in phenotype upon linker proteins ablation were rescued upon loss of NDRG1 in linker proteins knockdown background. These data together indicate that hemidesmosomal linker proteins regulate cell motility, invasion and tumorigenicity possibly through NDRG1 in OSCC derived cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Notch1-Dll4 signaling and mechanical force regulate leader cell formation during collective cell migration

    OpenAIRE

    Riahi, Reza; Sun, Jian; Wang, Shue; Long, Min; Zhang, Donna D.; Wong, Pak Kin

    2015-01-01

    At the onset of collective cell migration, a subset of cells within an initially homogenous population acquires a distinct “leader” phenotype with characteristic morphology and motility. However, the factors driving leader cell formation as well as the mechanisms regulating leader cell density during the migration process remain to be determined. Here, we use single cell gene expression analysis and computational modeling to show that leader cell identity is dynamically regulated by Dll4 sign...

  9. Regulation of Germinal Center Reactions by B and T Cells

    Directory of Open Access Journals (Sweden)

    Yeonseok Chung

    2013-10-01

    Full Text Available Break of B cell tolerance to self-antigens results in the development of autoantibodies and, thus, leads to autoimmunity. How B cell tolerance is maintained during active germinal center (GC reactions is yet to be fully understood. Recent advances revealed several subsets of T cells and B cells that can positively or negatively regulate GC B cell responses in vivo. IL-21-producing CXCR5+ CD4+ T cells comprise a distinct lineage of helper T cells—termed follicular helper T cells (TFH—that can provide help for the development of GC reactions where somatic hypermutation and affinity maturation take place. Although the function of TFH cells is beneficial in generating high affinity antibodies against infectious agents, aberrant activation of TFH cell or B cell to self-antigens results in autoimmunity. At least three subsets of immune cells have been proposed as regulatory cells that can limit such antibody-mediated autoimmunity, including follicular regulatory T cells (TFR, Qa-1 restricted CD8+ regulatory T cells (CD8+TREG, and regulatory B cells (BREG. In this review, we will discuss our current understanding of GC B cell regulation with specific emphasis on the newly identified immune cell subsets involved in this process.

  10. Comparison between ultrasonography and MR imaging for discriminating squamous cell carcinoma nodes with extranodal spread in the neck

    International Nuclear Information System (INIS)

    Katayama, Ikuo; Sasaki, Miho; Kimura, Yasuo; Hotokezaka, Yuka; Eida, Sato; Tashiro, Shigeki; Sumi, Misa; Nakamura, Takashi

    2012-01-01

    Objective: To compare the diagnostic ability of ultrasonography (US) and MR imaging for discriminating squamous cell carcinoma (SCC) nodes with extranodal spread (ENS) in the neck. Methods: US and MR imaging was retrospectively evaluated for differentiating ENS-positive (n = 28) from ENS-negative (n = 26) SCC nodes (>10 mm short-axis diameter) in 50 patients with head and neck SCCs. We assessed nodal size on US and MR images; irregular nodal margin on US; and vanishing nodal border, flare, and shaggy nodal margin signs on T1-, fat-suppressed T2-, and contrast-enhanced T1-weighted MR images, respectively. US and MR images were analyzed by 3 radiologists in consensus and the results were compared between ENS-positive and ENS-negative SCC nodes. Results: The nodal sizes of ENS-positive nodes (21 ± 9 mm) were significantly larger than those of ENS-negative SCC nodes (14 ± 4 mm) (p < 0.001). Irregular nodal margins were more frequently observed in ENS-positive SCC nodes (75%) than in ENS-negative SCC nodes (12%). The vanishing nodal margin, flare, and shaggy nodal margin signs were more frequently observed in ENS-positive SCC nodes (93%, 89%, and 82%, respectively) than in ENS-negative nodes (46%, 19%, and 19%, respectively). A combination of size (≥22 mm) and imaging criteria (irregular margin or flare sign) best discriminated ENS-positive SCC nodes with 82% sensitivity, 89% specificity, and 85% accuracy for US and 89% sensitivity, 81% specificity, and 85% accuracy for MR imaging. Conclusion: US discriminated ENS-positive from ENS-negative SCC nodes with comparable accuracy and higher specificity than MR imaging.

  11. Regulated portals of entry into the cell

    Science.gov (United States)

    Conner, Sean D.; Schmid, Sandra L.

    2003-03-01

    The plasma membrane is the interface between cells and their harsh environment. Uptake of nutrients and all communication among cells and between cells and their environment occurs through this interface. `Endocytosis' encompasses several diverse mechanisms by which cells internalize macromolecules and particles into transport vesicles derived from the plasma membrane. It controls entry into the cell and has a crucial role in development, the immune response, neurotransmission, intercellular communication, signal transduction, and cellular and organismal homeostasis. As the complexity of molecular interactions governing endocytosis are revealed, it has become increasingly clear that it is tightly coordinated and coupled with overall cell physiology and thus, must be viewed in a broader context than simple vesicular trafficking.

  12. European regulation for therapeutic use of stem cells.

    Science.gov (United States)

    Ferry, Nicolas

    2017-01-01

    The regulation for the use of stem cells has evolved during the past decade with the aim of ensuring a high standard of quality and safety for human derived products throughout Europe to comply with the provision of the Lisbon treaty. To this end, new regulations have been issued and the regulatory status of stem cells has been revised. Indeed, stem cells used for therapeutic purposes can now be classified as a cell preparation, or as advanced therapy medicinal products depending on the clinical indication and on the procedure of cell preparation. Furthermore, exemptions to the European regulation are applicable for stem cells prepared and used within the hospital. The aim of this review is to give the non-specialized reader a broad overview of this particular regulatory landscape.

  13. Ornithine decarboxylase and extracellular polyamines regulate microvascular sprouting and actin cytoskeleton dynamics in endothelial cells

    International Nuclear Information System (INIS)

    Kucharzewska, Paulina; Welch, Johanna E.; Svensson, Katrin J.; Belting, Mattias

    2010-01-01

    The polyamines are essential for cancer cell proliferation during tumorigenesis. Targeted inhibition of ornithine decarboxylase (ODC), i.e. a key enzyme of polyamine biosynthesis, by α-difluoromethylornithine (DFMO) has shown anti-neoplastic activity in various experimental models. This activity has mainly been attributed to the anti-proliferative effect of DFMO in cancer cells. Here, we provide evidence that unperturbed ODC activity is a requirement for proper microvessel sprouting ex vivo as well as the migration of primary human endothelial cells. DFMO-mediated ODC inhibition was reversed by extracellular polyamine supplementation, showing that anti-angiogenic effects of DFMO were specifically related to polyamine levels. ODC inhibition was associated with an abnormal morphology of the actin cytoskeleton during cell spreading and migration. Moreover, our data suggest that de-regulated actin cytoskeleton dynamics in DFMO treated endothelial cells may be related to constitutive activation of the small GTPase CDC42, i.e. a well-known regulator of cell motility and actin cytoskeleton remodeling. These insights into the potential role of polyamines in angiogenesis should stimulate further studies testing the combined anti-tumor effect of polyamine inhibition and established anti-angiogenic therapies in vivo.

  14. Nanotechnology in the regulation of stem cell behavior

    International Nuclear Information System (INIS)

    Wu, King-Chuen; Tseng, Ching-Li; Wu, Chi-Chang; Wang, Yang-Kao; Kao, Feng-Chen; Tu, Yuan-Kun; C So, Edmund

    2013-01-01

    Stem cells are known for their potential to repair damaged tissues. The adhesion, growth and differentiation of stem cells are likely controlled by the surrounding microenvironment which contains both chemical and physical cues. Physical cues in the microenvironment, for example, nanotopography, were shown to play important roles in stem cell fate decisions. Thus, controlling stem cell behavior by nanoscale topography has become an important issue in stem cell biology. Nanotechnology has emerged as a new exciting field and research from this field has greatly advanced. Nanotechnology allows the manipulation of sophisticated surfaces/scaffolds which can mimic the cellular environment for regulating cellular behaviors. Thus, we summarize recent studies on nanotechnology with applications to stem cell biology, including the regulation of stem cell adhesion, growth, differentiation, tracking and imaging. Understanding the interactions of nanomaterials with stem cells may provide the knowledge to apply to cell–scaffold combinations in tissue engineering and regenerative medicine. (review)

  15. Molecular regulation of human hematopoietic stem cells

    NARCIS (Netherlands)

    van Galen, P.L.J.

    2014-01-01

    Peter van Galen focuses on understanding the determinants that maintain the stem cell state. Using human hematopoietic stem cells (HSCs) as a model, processes that govern self-renewal and tissue regeneration were investigated. Specifically, a role for microRNAs in balancing the human HSC

  16. Retinoic Acid Signaling in Thymic Epithelial Cells Regulates Thymopoiesis

    DEFF Research Database (Denmark)

    Wendland, Kerstin; Niss, Kristoffer; Kotarsky, Knut

    2018-01-01

    Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis. In the abse......Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis...

  17. BMP signaling regulates satellite cell-dependent postnatal muscle growth.

    Science.gov (United States)

    Stantzou, Amalia; Schirwis, Elija; Swist, Sandra; Alonso-Martin, Sonia; Polydorou, Ioanna; Zarrouki, Faouzi; Mouisel, Etienne; Beley, Cyriaque; Julien, Anaïs; Le Grand, Fabien; Garcia, Luis; Colnot, Céline; Birchmeier, Carmen; Braun, Thomas; Schuelke, Markus; Relaix, Frédéric; Amthor, Helge

    2017-08-01

    Postnatal growth of skeletal muscle largely depends on the expansion and differentiation of resident stem cells, the so-called satellite cells. Here, we demonstrate that postnatal satellite cells express components of the bone morphogenetic protein (BMP) signaling machinery. Overexpression of noggin in postnatal mice (to antagonize BMP ligands), satellite cell-specific knockout of Alk3 (the gene encoding the BMP transmembrane receptor) or overexpression of inhibitory SMAD6 decreased satellite cell proliferation and accretion during myofiber growth, and ultimately retarded muscle growth. Moreover, reduced BMP signaling diminished the adult satellite cell pool. Abrogation of BMP signaling in satellite cell-derived primary myoblasts strongly diminished cell proliferation and upregulated the expression of cell cycle inhibitors p21 and p57 In conclusion, these results show that BMP signaling defines postnatal muscle development by regulating satellite cell-dependent myofiber growth and the generation of the adult muscle stem cell pool. © 2017. Published by The Company of Biologists Ltd.

  18. Roquin Paralogs Differentially Regulate Functional NKT Cell Subsets.

    Science.gov (United States)

    Drees, Christoph; Vahl, J Christoph; Bortoluzzi, Sabrina; Heger, Klaus D; Fischer, Julius C; Wunderlich, F Thomas; Peschel, Christian; Schmidt-Supprian, Marc

    2017-04-01

    NKT cells represent a small subset of glycolipid-recognizing T cells that are heavily implicated in human allergic, autoimmune, and malignant diseases. In the thymus, precursor cells recognize self-glycolipids by virtue of their semi-invariant TCR, which triggers NKT cell lineage commitment and maturation. During their development, NKT cells are polarized into the NKT1, NKT2, and NKT17 subsets, defined through their cytokine-secretion patterns and the expression of key transcription factors. However, we have largely ignored how the differentiation into the NKT cell subsets is regulated. In this article, we describe the mRNA-binding Roquin-1 and -2 proteins as central regulators of murine NKT cell fate decisions. In the thymus, T cell-specific ablation of the Roquin paralogs leads to a dramatic expansion of NKT17 cells, whereas peripheral mature NKT cells are essentially absent. Roquin-1/2-deficient NKT17 cells show exaggerated lineage-specific expression of nearly all NKT17-defining proteins tested. We show through mixed bone marrow chimera experiments that NKT17 polarization is mediated through cell-intrinsic mechanisms early during NKT cell development. In contrast, the loss of peripheral NKT cells is due to cell-extrinsic factors. Surprisingly, Roquin paralog-deficient NKT cells are, in striking contrast to conventional T cells, compromised in their ability to secrete cytokines. Altogether, we show that Roquin paralogs regulate the development and function of NKT cell subsets in the thymus and periphery. Copyright © 2017 by The American Association of Immunologists, Inc.

  19. Regulation of pulmonary inflammation by mesenchymal cells

    NARCIS (Netherlands)

    Alkhouri, Hatem; Poppinga, Wilfred Jelco; Tania, Navessa Padma; Ammit, Alaina; Schuliga, Michael

    2014-01-01

    Pulmonary inflammation and tissue remodelling are common elements of chronic respiratory diseases such as asthma, chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF), and pulmonary hypertension (PH). In disease, pulmonary mesenchymal cells not only contribute to tissue

  20. Mast Cell Interactions and Crosstalk in Regulating Allergic Inflammation.

    Science.gov (United States)

    Velez, Tania E; Bryce, Paul J; Hulse, Kathryn E

    2018-04-17

    This review summarizes recent findings on mast cell biology with a focus on IgE-independent roles of mast cells in regulating allergic responses. Recent studies have described novel mast cell-derived molecules, both secreted and membrane-bound, that facilitate cross-talk with a variety of immune effector cells to mediate type 2 inflammatory responses. Mast cells are complex and dynamic cells that are persistent in allergy and are capable of providing signals that lead to the initiation and persistence of allergic mechanisms.

  1. Sonoporation of adherent cells under regulated ultrasound cavitation conditions.

    Science.gov (United States)

    Muleki Seya, Pauline; Fouqueray, Manuela; Ngo, Jacqueline; Poizat, Adrien; Inserra, Claude; Béra, Jean-Christophe

    2015-04-01

    A sonoporation device dedicated to the adherent cell monolayer has been implemented with a regulation process allowing the real-time monitoring and control of inertial cavitation activity. Use of the cavitation-regulated device revealed first that adherent cell sonoporation efficiency is related to inertial cavitation activity, without inducing additional cell mortality. Reproducibility is enhanced for the highest sonoporation rates (up to 17%); sonoporation efficiency can reach 26% when advantage is taken of the standing wave acoustic configuration by applying a frequency sweep with ultrasound frequency tuned to the modal acoustic modes of the cavity. This device allows sonoporation of adherent and suspended cells, and the use of regulation allows some environmental parameters such as the temperature of the medium to be overcome, resulting in the possibility of cell sonoporation even at ambient temperature. Copyright © 2015 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

  2. Biophysical force regulation in 3D tumor cell invasion

    Science.gov (United States)

    Wu, Mingming

    When embedded within 3D extracellular matrices (ECM), animal cells constantly probe and adapt to the ECM locally (at cell length scale) and exert forces and communicate with other cells globally (up to 10 times of cell length). It is now well accepted that mechanical crosstalk between animal cells and their microenvironment critically regulate cell function such as migration, proliferation and differentiation. Disruption of the cell-ECM crosstalk is implicated in a number of pathologic processes including tumor progression and fibrosis. Central to the problem of cell-ECM crosstalk is the physical force that cells generate. By measuring single cell generated force within 3D collagen matrices, we revealed a mechanical crosstalk mechanism between the tumor cells and the ECM. Cells generate sufficient force to stiffen collagen fiber network, and stiffer matrix, in return promotes larger cell force generation. Our work highlights the importance of fibrous nonlinear elasticity in regulating tumor cell-ECM interaction, and results may have implications in the rapid tissue stiffening commonly found in tumor progression and fibrosis. This work is partially supported by NIH Grants R21RR025801 and R21GM103388.

  3. MHC class II molecules regulate growth in human T cells

    DEFF Research Database (Denmark)

    Nielsen, M; Odum, Niels; Bendtzen, K

    1994-01-01

    MHC-class-II-positive T cells are found in tissues involved in autoimmune disorders. Stimulation of class II molecules by monoclonal antibodies (mAbs) or bacterial superantigens induces protein tyrosine phosphorylation through activation of protein tyrosine kinases in T cells, and class II signals...... lines tested. Only one of three CD4+, CD45RAhigh, ROhigh T cells responded to class II costimulation. There was no correlation between T cell responsiveness to class II and the cytokine production profile of the T cell in question. Thus, T cell lines producing interferon (IFN)-gamma but not IL-4 (TH1...... modulate several T cell responses. Here, we studied further the role of class II molecules in the regulation of T cell growth. Costimulation of class II molecules by immobilized HLA-DR mAb significantly enhanced interleukin (IL)-2-supported T cell growth of the majority of CD4+, CD45RAlow, ROhigh T cell...

  4. Quantitative regulation of B cell division destiny by signal strength.

    Science.gov (United States)

    Turner, Marian L; Hawkins, Edwin D; Hodgkin, Philip D

    2008-07-01

    Differentiation to Ab secreting and isotype-switched effector cells is tightly linked to cell division and therefore the degree of proliferation strongly influences the nature of the immune response. The maximum number of divisions reached, termed the population division destiny, is stochastically distributed in the population and is an important parameter in the quantitative outcome of lymphocyte responses. In this study, we further assessed the variables that regulate B cell division destiny in vitro in response to T cell- and TLR-dependent stimuli. Both the concentration and duration of stimulation were able to regulate the average maximum number of divisions undergone for each stimulus. Notably, a maximum division destiny was reached during provision of repeated saturating stimulation, revealing that an intrinsic limit to proliferation exists even under these conditions. This limit was linked directly to division number rather than time of exposure to stimulation and operated independently of the survival regulation of the cells. These results demonstrate that a B cell population's division destiny is regulable by the stimulatory conditions up to an inherent maximum value. Division destiny is a crucial parameter in regulating the extent of B cell responses and thereby also the nature of the immune response mounted.

  5. Understanding cell cycle and cell death regulation provides novel weapons against human diseases.

    Science.gov (United States)

    Wiman, K G; Zhivotovsky, B

    2017-05-01

    Cell division, cell differentiation and cell death are the three principal physiological processes that regulate tissue homoeostasis in multicellular organisms. The growth and survival of cells as well as the integrity of the genome are regulated by a complex network of pathways, in which cell cycle checkpoints, DNA repair and programmed cell death have critical roles. Disruption of genomic integrity and impaired regulation of cell death may both lead to uncontrolled cell growth. Compromised cell death can also favour genomic instability. It is becoming increasingly clear that dysregulation of cell cycle and cell death processes plays an important role in the development of major disorders such as cancer, cardiovascular disease, infection, inflammation and neurodegenerative diseases. Research achievements in these fields have led to the development of novel approaches for treatment of various conditions associated with abnormalities in the regulation of cell cycle progression or cell death. A better understanding of how cellular life-and-death processes are regulated is essential for this development. To highlight these important advances, the Third Nobel Conference entitled 'The Cell Cycle and Cell Death in Disease' was organized at Karolinska Institutet in 2016. In this review we will summarize current understanding of cell cycle progression and cell death and discuss some of the recent advances in therapeutic applications in pathological conditions such as cancer, neurological disorders and inflammation. © 2017 The Association for the Publication of the Journal of Internal Medicine.

  6. Effects of air polishing and an amino acid buffered hypochlorite solution to dentin surfaces and periodontal ligament cell survival, attachment, and spreading.

    Science.gov (United States)

    Schmidlin, Patrick R; Fujioka-Kobayashi, Masako; Mueller, Heinz-Dieter; Sculean, Anton; Lussi, Adrian; Miron, Richard J

    2017-06-01

    The aim of this study is to examine morphological changes of dentin surfaces following air polishing or amino acid buffered hypochlorite solution application and to assess their influence on periodontal ligament (PDL) cell survival, attachment, and spreading to dentin discs in vitro. Bovine dentin discs were treated with either (i) Classic, (ii) Plus, or (iii) Perio powder (EMS). Furthermore, Perisolv® a hypochlorite solution buffered with various amino acids was investigated. Untreated dentin discs served as controls. Morphological changes to dentin discs were assessed using scanning electron microscopy (SEM). Human PDL cells were seeded onto the respectively treated discs, and samples were then investigated for PDL cell survival, attachment, and spreading using a live/dead assay, adhesion assay, and SEM imaging, respectively. Both control and Perisolv®-rinsed dentin discs demonstrated smooth surfaces at low and high magnifications. The Classic powders demonstrated the thickest coating followed by the Powder Plus. The Perio powder demonstrated marked alterations of dentin discs by revealing the potential to open dentinal tubules even before rinsing. Seeding of PDL cells demonstrated an almost 100 % survival rate on all samples demonstrating very high biocompatibility for all materials. Significantly higher PDL cell numbers were observed on samples treated with the Perio powder and the Perisolv® solution (approximately 40 % more cells; p air polishing or application with Perisolv®. Future in vitro and animal testing is necessary to further characterize the beneficial effects of either system in a clinical setting. The use of air polishing or application with Perisolv amino acid buffered hypochlorite solution was effective in treating root surfaces and allowed for near 100 % PDL cell survival, attachment, and spreading onto all root surfaces.

  7. Activation of ion transport systems during cell volume regulation

    International Nuclear Information System (INIS)

    Eveloff, J.L.; Warnock, D.G.

    1987-01-01

    This review discusses the activation of transport pathways during volume regulation, including their characteristics, the possible biochemical pathways that may mediate the activation of transport pathways, and the relations between volume regulation and transepithelial transport in renal cells. Many cells regulate their volume when exposed to an anisotonic medium. The changes in cell volume are caused by activation of ion transport pathways, plus the accompanying osmotically driven water movement such that cell volume returns toward normal levels. The swelling of hypertonically shrunken cells is termed regulatory volume increase (RVI) and involves an influx of NaCl into the cell via either activation of Na-Cl, Na-K-2Cl cotransport systems, or Na + -H + and Cl - -HCO 3 - exchangers. The reshrinking of hypotonically swollen cells is termed regulatory volume decrease (RVD) and involves an efflux of KCl and water from the cell by activation of either separate K + and Cl - conductances, a K-Cl cotransport system, or parallel K + -H + and Cl - -HCO 3 - exchangers. The biochemical mechanisms involved in the activation of transport systems are largely unknown, however, the phosphoinositide pathway may be implicated in RVI; phorbol esters, cGMP, and Ca 2+ affect the process of volume regulation. Renal tubular cells, as well as the blood cells that transverse the medulla, are subjected to increasing osmotic gradients from the corticomedullary junction to the papillary tip, as well as changing interstitial and tubule fluid osmolarity, depending on the diuretic state of the animal. Medullary cells from the loop of Henle and the papilla can volume regulate by activating Na-K-2Cl cotransport or Na + -H + and Cl - -HCO 3 - exchange systems

  8. Protein kinase C signaling and cell cycle regulation

    Directory of Open Access Journals (Sweden)

    Adrian R Black

    2013-01-01

    Full Text Available A link between T cell proliferation and the protein kinase C (PKC family of serine/threonine kinases has been recognized for about thirty years. However, despite the wealth of information on PKC-mediated control of T cell activation, understanding of the effects of PKCs on the cell cycle machinery in this cell type remains limited. Studies in other systems have revealed important cell cycle-specific effects of PKC signaling that can either positively or negatively impact proliferation. The outcome of PKC activation is highly context-dependent, with the precise cell cycle target(s and overall effects determined by the specific isozyme involved, the timing of PKC activation, the cell type, and the signaling environment. Although PKCs can regulate all stages of the cell cycle, they appear to predominantly affect G0/G1 and G2. PKCs can modulate multiple cell cycle regulatory molecules, including cyclins, cyclin-dependent kinases (cdks, cdk inhibitors and cdc25 phosphatases; however, evidence points to Cip/Kip cdk inhibitors and D-type cyclins as key mediators of PKC-regulated cell cycle-specific effects. Several PKC isozymes can target Cip/Kip proteins to control G0/G1→S and/or G2→M transit, while effects on D-type cyclins regulate entry into and progression through G1. Analysis of PKC signaling in T cells has largely focused on its roles in T cell activation; thus, observed cell cycle effects are mainly positive. A prominent role is emerging for PKCθ, with non-redundant functions of other isozymes also described. Additional evidence points to PKCδ as a negative regulator of the cell cycle in these cells. As in other cell types, context-dependent effects of individual isozymes have been noted in T cells, and Cip/Kip cdk inhibitors and D-type cyclins appear to be major PKC targets. Future studies are anticipated to take advantage of the similarities between these various systems to enhance understanding of PKC-mediated cell cycle regulation in

  9. ASIC PROTEINS REGULATE SMOOTH MUSCLE CELL MIGRATION

    OpenAIRE

    Grifoni, Samira C.; Jernigan, Nikki L.; Hamilton, Gina; Drummond, Heather A.

    2007-01-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated Epithelial Na+ Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration, however the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence indi...

  10. Mitochondrial fission proteins regulate programmed cell death in yeast.

    Science.gov (United States)

    Fannjiang, Yihru; Cheng, Wen-Chih; Lee, Sarah J; Qi, Bing; Pevsner, Jonathan; McCaffery, J Michael; Hill, R Blake; Basañez, Gorka; Hardwick, J Marie

    2004-11-15

    The possibility that single-cell organisms undergo programmed cell death has been questioned in part because they lack several key components of the mammalian cell death machinery. However, yeast encode a homolog of human Drp1, a mitochondrial fission protein that was shown previously to promote mammalian cell death and the excessive mitochondrial fragmentation characteristic of apoptotic mammalian cells. In support of a primordial origin of programmed cell death involving mitochondria, we found that the Saccharomyces cerevisiae homolog of human Drp1, Dnm1, promotes mitochondrial fragmentation/degradation and cell death following treatment with several death stimuli. Two Dnm1-interacting factors also regulate yeast cell death. The WD40 repeat protein Mdv1/Net2 promotes cell death, consistent with its role in mitochondrial fission. In contrast to its fission function in healthy cells, Fis1 unexpectedly inhibits Dnm1-mediated mitochondrial fission and cysteine protease-dependent cell death in yeast. Furthermore, the ability of yeast Fis1 to inhibit mitochondrial fission and cell death can be functionally replaced by human Bcl-2 and Bcl-xL. Together, these findings indicate that yeast and mammalian cells have a conserved programmed death pathway regulated by a common molecular component, Drp1/Dnm1, that is inhibited by a Bcl-2-like function.

  11. Correlation between podoplanin expression and extracapsular spread in squamous cell carcinoma of the oral cavity using subjective immunoreactivity scores and semiquantitative image analysis.

    Science.gov (United States)

    Mermod, Maxime; Bongiovanni, Massimo; Petrova, Tatiana V; Dubikovskaya, Elena A; Simon, Christian; Tolstonog, Genrich; Monnier, Yan

    2017-01-01

    The correlation between podoplanin expression and extracapsular spread in head and neck squamous cell carcinoma (HNSCC) has never been reported. The purpose of this study was to assess the predictive value of podoplanin expression for this parameter. Subjective immunoreactivity scores and semiquantitative image analysis of podoplanin expression were performed in 67 patients with primary oral squamous cell carcinoma and in their corresponding lymph nodes. Neck classification showed 34 cases (51%) of pN0 and 33 cases (49%) of pN+. Correlation between the levels of podoplanin expression and the histopathological data was established. In lymph nodes, a high level of podoplanin expression correlated with the presence of extracapsular spread by multivariate analysis (p = .03). A strong correlation between subjective and semiquantitative image analysis was observed (r = 0.77; p < .001). A high level of podoplanin expression in lymph node metastases of oral squamous cell carcinoma is independently associated with extracapsular spread. © 2016 Wiley Periodicals, Head Neck 39: 98-108, 2017. © 2016 Wiley Periodicals, Inc.

  12. BMP signalling differentially regulates distinct haematopoietic stem cell types

    NARCIS (Netherlands)

    M. Crisan (Mihaela); P. Solaimani Kartalaei (Parham); C.S. Vink (Chris); T. Yamada-Inagawa (Tomoko); K. Bollerot (Karine); W.F.J. van IJcken (Wilfred); R. Van Der Linden (Reinier); S.C. de Sousa Lopes (Susana Chuva); R. Monteiro (Rui); C.L. Mummery (Christine); E.A. Dzierzak (Elaine)

    2015-01-01

    textabstractAdult haematopoiesis is the outcome of distinct haematopoietic stem cell (HSC) subtypes with self-renewable repopulating ability, but with different haematopoietic cell lineage outputs. The molecular basis for this heterogeneity is largely unknown. BMP signalling regulates HSCs as they

  13. Gamma Delta T-Cells Regulate Inflammatory Cell Infiltration of the Lung after Trauma-Hemorrhage

    Science.gov (United States)

    2015-06-01

    suggesting a role for this T- cell subset in both innate and acquired immunity (7, 8). Studies have shown that +% T cells are required for both controlled...increased infiltration of both lymphoid and myeloid cells in WT mice after TH-induced ALI. In parallel to +% T cells , myeloid cells (i.e., monocytes...GAMMA DELTA T CELLS REGULATE INFLAMMATORY CELL INFILTRATION OF THE LUNG AFTER TRAUMA-HEMORRHAGE Meenakshi Rani,* Qiong Zhang,* Richard F. Oppeltz

  14. Lin28a regulates germ cell pool size and fertility

    Science.gov (United States)

    Shinoda, Gen; de Soysa, T. Yvanka; Seligson, Marc T.; Yabuuchi, Akiko; Fujiwara, Yuko; Huang, Pei Yi; Hagan, John P.; Gregory, Richard I.; Moss, Eric G.; Daley, George Q.

    2013-01-01

    Overexpression of LIN28A is associated with human germ cell tumors and promotes primordial germ cell (PGC) development from embryonic stem cells in vitro and in chimeric mice. Knockdown of Lin28a inhibits PGC development in vitro, but how constitutional Lin28a deficiency affects the mammalian reproductive system in vivo remains unknown. Here, we generated Lin28a knockout (KO) mice and found that Lin28a deficiency compromises the size of the germ cell pool in both males and females by affecting PGC proliferation during embryogenesis. Interestingly however, in Lin28a KO males the germ cell pool partially recovers during postnatal expansion, while fertility remains impaired in both males and females mated to wild type mice. Embryonic overexpression of let-7, a microRNA negatively regulated by Lin28a, reduces the germ cell pool, corroborating the role of the Lin28a/let-7 axis in regulating the germ lineage. PMID:23378032

  15. 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.

  16. RasGRP3 regulates the migration of glioma cells via interaction with Arp3

    Science.gov (United States)

    Lee, Hae Kyung; Finniss, Susan; Cazacu, Simona; Xiang, Cunli; Poisson, Laila M.; Blumberg, Peter M.; Brodie, Chaya

    2015-01-01

    Glioblastoma (GBM), the most aggressive primary brain tumors, are highly infiltrative. Although GBM express high Ras activity and Ras proteins have been implicated in gliomagenesis, Ras-activating mutations are not frequent in these tumors. RasGRP3, an important signaling protein responsive to diacylglycerol (DAG), increases Ras activation. Here, we examined the expression and functions of RasGRP3 in GBM and glioma cells. RasGRP3 expression was upregulated in GBM specimens and glioma stem cells compared with normal brains and neural stem cells, respectively. RasGRP3 activated Ras and Rap1 in glioma cells and increased cell migration and invasion partially via Ras activation. Using pull-down assay and mass spectroscopy we identified the actin-related protein, Arp3, as a novel interacting protein of RasGRP3. The interaction of RasGRP3 and Arp3 was validated by immunofluorescence staining and co-immunoprecipitation, and PMA, which activates RasGRP3 and induces its translocation to the peri-nuclear region, increased the association of Arp3 and RasGRP3. Arp3 was upregulated in GBM, regulated cell spreading and migration and its silencing partially decreased these effects of RasGRP3 in glioma cells. In summary, RasGRP3 acts as an important integrating signaling protein of the DAG and Ras signaling pathways and actin polymerization and represents an important therapeutic target in GBM. PMID:25682201

  17. TGF-β Signaling Regulates Pancreatic β-Cell Proliferation through Control of Cell Cycle Regulator p27 Expression

    International Nuclear Information System (INIS)

    Suzuki, Tomoyuki; Dai, Ping; Hatakeyama, Tomoya; Harada, Yoshinori; Tanaka, Hideo; Yoshimura, Norio; Takamatsu, Tetsuro

    2013-01-01

    Proliferation of pancreatic β-cells is an important mechanism underlying β-cell mass adaptation to metabolic demands. Increasing β-cell mass by regeneration may ameliorate or correct both type 1 and type 2 diabetes, which both result from inadequate production of insulin by β-cells of the pancreatic islet. Transforming growth factor β (TGF-β) signaling is essential for fetal development and growth of pancreatic islets. In this study, we exposed HIT-T15, a clonal pancreatic β-cell line, to TGF-β signaling. We found that inhibition of TGF-β signaling promotes proliferation of the cells significantly, while TGF-β signaling stimulation inhibits proliferation of the cells remarkably. We confirmed that this proliferative regulation by TGF-β signaling is due to the changed expression of the cell cycle regulator p27. Furthermore, we demonstrated that there is no observed effect on transcriptional activity of p27 by TGF-β signaling. Our data show that TGF-β signaling mediates the cell-cycle progression of pancreatic β-cells by regulating the nuclear localization of CDK inhibitor, p27. Inhibition of TGF-β signaling reduces the nuclear accumulation of p27, and as a result this inhibition promotes proliferation of β-cells

  18. Cellular growth in plants requires regulation of cell wall biochemistry.

    Science.gov (United States)

    Chebli, Youssef; Geitmann, Anja

    2017-02-01

    Cell and organ morphogenesis in plants are regulated by the chemical structure and mechanical properties of the extracellular matrix, the cell wall. The two primary load bearing components in the plant cell wall, the pectin matrix and the cellulose/xyloglucan network, are constantly remodelled to generate the morphological changes required during plant development. This remodelling is regulated by a plethora of loosening and stiffening agents such as pectin methyl-esterases, calcium ions, expansins, and glucanases. The tight spatio-temporal regulation of the activities of these agents is a sine qua non condition for proper morphogenesis at cell and tissue levels. The pectin matrix and the cellulose-xyloglucan network operate in concert and their behaviour is mutually dependent on their chemical, structural and mechanical modifications. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. ASIC proteins regulate smooth muscle cell migration.

    Science.gov (United States)

    Grifoni, Samira C; Jernigan, Nikki L; Hamilton, Gina; Drummond, Heather A

    2008-03-01

    The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated that Epithelial Na(+)Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration; however, the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence individual ASIC expression and determine the importance of ASIC proteins in wound healing and chemotaxis (PDGF-bb)-initiated migration. We found ASIC1, ASIC2, and ASIC3, but not ASIC4, expression in A10 cells. ASIC1, ASIC2, and ASIC3 siRNA molecules significantly suppressed expression of their respective proteins compared to non-targeting siRNA (RISC) transfected controls by 63%, 44%, and 55%, respectively. Wound healing was inhibited by 10, 20, and 26% compared to RISC controls following suppression of ASIC1, ASIC2, and ASIC3, respectively. Chemotactic migration was inhibited by 30% and 45%, respectively, following suppression of ASIC1 and ASIC3. ASIC2 suppression produced a small, but significant, increase in chemotactic migration (4%). Our data indicate that ASIC expression is required for normal migration and may suggest a novel role for ASIC proteins in cellular migration.

  20. Physical model for membrane protrusions during spreading

    International Nuclear Information System (INIS)

    Chamaraux, F; Ali, O; Fourcade, B; Keller, S; Bruckert, F

    2008-01-01

    During cell spreading onto a substrate, the kinetics of the contact area is an observable quantity. This paper is concerned with a physical approach to modeling this process in the case of ameboid motility where the membrane detaches itself from the underlying cytoskeleton at the leading edge. The physical model we propose is based on previous reports which highlight that membrane tension regulates cell spreading. Using a phenomenological feedback loop to mimic stress-dependent biochemistry, we show that the actin polymerization rate can be coupled to the stress which builds up at the margin of the contact area between the cell and the substrate. In the limit of small variation of membrane tension, we show that the actin polymerization rate can be written in a closed form. Our analysis defines characteristic lengths which depend on elastic properties of the membrane–cytoskeleton complex, such as the membrane–cytoskeleton interaction, and on molecular parameters, the rate of actin polymerization. We discuss our model in the case of axi-symmetric and non-axi-symmetric spreading and we compute the characteristic time scales as a function of fundamental elastic constants such as the strength of membrane–cytoskeleton adherence

  1. SOX15 regulates proliferation and migration of endometrial cancer cells.

    Science.gov (United States)

    Rui, Xiaohui; Xu, Yun; Jiang, Xiping; Guo, Caixia; Jiang, Jingting

    2017-10-31

    The study aimed to investigate the effects of Sry-like high mobility group box 15 ( SOX15 ) on proliferation and migration of endometrial cancer (EC) cells. Immunohistochemistry (IHC) was applied to determine the expression of SOX15 in EC tissues and adjacent tissues. We used cell transfection method to construct the HEC-1-A and Ishikawa cell lines with stable overexpression and low expression SOX15 Reverse-transcription quantitative real-time PCR (RT-qPCR) and Western blot were performed to examine expression of SOX15 mRNA and SOX15 protein, respectively. By conducting a series of cell proliferation assay and migration assay, we analyzed the influence of SOX15 overexpression or low expression on EC cell proliferation and migration. The expression of SOX15 mRNA and protein in EC tissues was significantly lower than that in adjacent tissues. After lentivirus-transfecting SOX15 , the expression level of SOX15 mRNA and protein was significantly increased in cells of SOX15 group, and decreased in sh- SOX15 group. Overexpression of SOX15 could suppress cell proliferation, while down-regulation of SOX15 increased cell proliferation. Flow cytometry results indicated that overexpression of SOX15 induced the ratio of cell-cycle arrest in G 1 stage. In addition, Transwell migration assay results showed that SOX15 overexpression significantly inhibited cell migration, and also down-regulation of SOX15 promoted the migration. As a whole, SOX15 could regulate the proliferation and migration of EC cells and up- regulation of SOX15 could be valuable for EC treatment. © 2017 The Author(s).

  2. Human cyclin T1 expression ameliorates a T-cell-specific transcriptional limitation for HIV in transgenic rats, but is not sufficient for a spreading infection of prototypic R5 HIV-1 strains ex vivo

    Directory of Open Access Journals (Sweden)

    Littman Dan R

    2009-01-01

    Full Text Available Abstract Background Cells derived from native rodents have limits at distinct steps of HIV replication. Rat primary CD4 T-cells, but not macrophages, display a profound transcriptional deficit that is ameliorated by transient trans-complementation with the human Tat-interacting protein Cyclin T1 (hCycT1. Results Here, we generated transgenic rats that selectively express hCycT1 in CD4 T-cells and macrophages. hCycT1 expression in rat T-cells boosted early HIV gene expression to levels approaching those in infected primary human T-cells. hCycT1 expression was necessary, but not sufficient, to enhance HIV transcription in T-cells from individual transgenic animals, indicating that endogenous cellular factors are critical co-regulators of HIV gene expression in rats. T-cells from hCD4/hCCR5/hCycT1-transgenic rats did not support productive infection of prototypic wild-type R5 HIV-1 strains ex vivo, suggesting one or more significant limitation in the late phase of the replication cycle in this primary rodent cell type. Remarkably, we identify a replication-competent HIV-1 GFP reporter strain (R7/3 YU-2 Env that displays characteristics of a spreading, primarily cell-to-cell-mediated infection in primary T-cells from hCD4/hCCR5-transgenic rats. Moreover, the replication of this recombinant HIV-1 strain was significantly enhanced by hCycT1 transgenesis. The viral determinants of this so far unique replicative ability are currently unknown. Conclusion Thus, hCycT1 expression is beneficial to de novo HIV infection in a transgenic rat model, but additional genetic manipulations of the host or virus are required to achieve full permissivity.

  3. Molecular biological mechanism II. Molecular mechanisms of cell cycle regulation

    International Nuclear Information System (INIS)

    Jung, T.

    2000-01-01

    The cell cycle in eukaryotes is regulated by central cell cycle controlling protein kinase complexes. These protein kinase complexes consist of a catalytic subunit from the cyclin-dependent protein kinase family (CDK), and a regulatory subunit from the cyclin family. Cyclins are characterised by their periodic cell cycle related synthesis and destruction. Each cell cycle phase is characterised by a specific set of CDKs and cyclins. The activity of CDK/cyclin complexes is mainly regulated on four levels. It is controlled by specific phosphorylation steps, the synthesis and destruction of cyclins, the binding of specific inhibitor proteins, and by active control of their intracellular localisation. At several critical points within the cell cycle, named checkpoints, the integrity of the cellular genome is monitored. If damage to the genome or an unfinished prior cell cycle phase is detected, the cell cycle progression is stopped. These cell cycle blocks are of great importance to secure survival of cells. Their primary importance is to prevent the manifestation and heritable passage of a mutated genome to daughter cells. Damage sensing, DNA repair, cell cycle control and apoptosis are closely linked cellular defence mechanisms to secure genome integrity. Disregulation in one of these defence mechanisms are potentially correlated with an increased cancer risk and therefore in at least some cases with an increased radiation sensitivity. (orig.) [de

  4. Nuclear myosin I regulates cell membrane tension

    Czech Academy of Sciences Publication Activity Database

    Venit, Tomáš; Kalendová, Alžběta; Petr, Martin; Dzijak, Rastislav; Pastorek, Lukáš; Rohožková, Jana; Malohlava, M.; Hozák, Pavel

    2016-01-01

    Roč. 6, AUG 2 (2016), č. článku 30864. ISSN 2045-2322 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109; GA ČR GAP305/11/2232; GA MŠk(CZ) LO1304 Institutional support: RVO:68378050 Keywords : neuronal growth cone * rna-polymerase-ii * cancer cells * phosphatidylinositol 4,5-bisphosphate * myo1c * actin * transcription * complex * motor * afm Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.259, year: 2016

  5. Tilted light sheet microscopy with 3D point spread functions for single-molecule super-resolution imaging in mammalian cells

    Science.gov (United States)

    Gustavsson, Anna-Karin; Petrov, Petar N.; Lee, Maurice Y.; Shechtman, Yoav; Moerner, W. E.

    2018-02-01

    To obtain a complete picture of subcellular nanostructures, cells must be imaged with high resolution in all three dimensions (3D). Here, we present tilted light sheet microscopy with 3D point spread functions (TILT3D), an imaging platform that combines a novel, tilted light sheet illumination strategy with engineered long axial range point spread functions (PSFs) for low-background, 3D super localization of single molecules as well as 3D super-resolution imaging in thick cells. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The axial positions of the single molecules are encoded in the shape of the PSF rather than in the position or thickness of the light sheet, and the light sheet can therefore be formed using simple optics. The result is flexible and user-friendly 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validated TILT3D for 3D superresolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed Tetrapod PSF for fiducial bead tracking and live axial drift correction. We envision TILT3D to become an important tool not only for 3D super-resolution imaging, but also for live whole-cell single-particle and single-molecule tracking.

  6. Tilted Light Sheet Microscopy with 3D Point Spread Functions for Single-Molecule Super-Resolution Imaging in Mammalian Cells.

    Science.gov (United States)

    Gustavsson, Anna-Karin; Petrov, Petar N; Lee, Maurice Y; Shechtman, Yoav; Moerner, W E

    2018-02-01

    To obtain a complete picture of subcellular nanostructures, cells must be imaged with high resolution in all three dimensions (3D). Here, we present tilted light sheet microscopy with 3D point spread functions (TILT3D), an imaging platform that combines a novel, tilted light sheet illumination strategy with engineered long axial range point spread functions (PSFs) for low-background, 3D super localization of single molecules as well as 3D super-resolution imaging in thick cells. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The axial positions of the single molecules are encoded in the shape of the PSF rather than in the position or thickness of the light sheet, and the light sheet can therefore be formed using simple optics. The result is flexible and user-friendly 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validated TILT3D for 3D super-resolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed Tetrapod PSF for fiducial bead tracking and live axial drift correction. We envision TILT3D to become an important tool not only for 3D super-resolution imaging, but also for live whole-cell single-particle and single-molecule tracking.

  7. Regulation of Floral Stem Cell Termination in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Toshiro eIto

    2015-02-01

    Full Text Available In Arabidopsis, floral stem cells are maintained only at the initial stages of flower development, and they are terminated at a specific time to ensure proper development of the reproductive organs. Floral stem cell termination is a dynamic and multi-step process involving many transcription factors, chromatin remodeling factors and signaling pathways. In this review, we discuss the mechanisms involved in floral stem cell maintenance and termination, highlighting the interplay between transcriptional regulation and epigenetic machinery in the control of specific floral developmental genes. In addition, we discuss additional factors involved in floral stem cell regulation, with the goal of untangling the complexity of the floral stem cell regulatory network.

  8. Regulation of Cell and Gene Therapy Medicinal Products in Taiwan.

    Science.gov (United States)

    Lin, Yi-Chu; Wang, Po-Yu; Tsai, Shih-Chih; Lin, Chien-Liang; Tai, Hsuen-Yung; Lo, Chi-Fang; Wu, Shiow-Ing; Chiang, Yu-Mei; Liu, Li-Ling

    2015-01-01

    Owing to the rapid and mature development of emerging biotechnology in the fields of cell culture, cell preservation, and recombinant DNA technology, more and more cell or gene medicinal therapy products have been approved for marketing, to treat serious diseases which have been challenging to treat with current medical practice or medicine. This chapter will briefly introduce the Taiwan Food and Drug Administration (TFDA) and elaborate regulation of cell and gene therapy medicinal products in Taiwan, including regulatory history evolution, current regulatory framework, application and review procedures, and relevant jurisdictional issues. Under the promise of quality, safety, and efficacy of medicinal products, it is expected the regulation and environment will be more flexible, streamlining the process of the marketing approval of new emerging cell or gene therapy medicinal products and providing diverse treatment options for physicians and patients.

  9. EZH2: a pivotal regulator in controlling cell differentiation.

    Science.gov (United States)

    Chen, Ya-Huey; Hung, Mien-Chie; Li, Long-Yuan

    2012-01-01

    Epigenetic regulation plays an important role in stem cell self-renewal, maintenance and lineage differentiation. The epigenetic profiles of stem cells are related to their transcriptional signature. Enhancer of Zeste homlog 2 (EZH2), a catalytic subunit of epigenetic regulator Polycomb repressive complex 2 (PRC2), has been shown to be a key regulator in controlling cellular differentiation. EZH2 is a histone methyltransferase that not only methylates histone H3 on Lys 27 (H3K27me3) but also interacts with and recruits DNA methyltransferases to methylate CpG at certain EZH2 target genes to establish firm repressive chromatin structures, contributing to tumor progression and the regulation of development and lineage commitment both in embryonic stem cells (ESCs) and adult stem cells. In addition to its well-recognized epigenetic gene silencing function, EZH2 also directly methylates nonhistone targets such as the cardiac transcription factor, GATA4, resulting in attenuated GATA4 transcriptional activity and gene repression. This review addresses recent progress toward the understanding of the biological functions and regulatory mechanisms of EZH2 and its targets as well as their roles in stem cell maintenance and cell differentiation.

  10. Regulation of T cell differentiation and function by EZH2

    Directory of Open Access Journals (Sweden)

    THEODOROS KARANTANOS

    2016-05-01

    Full Text Available The enhancer of zeste homologue 2 (EZH2, one of the polycomb group (PcG proteins, is the catalytic subunit of Polycomb-repressive complex 2 (PRC2 and induces the trimethylation of the histone H3 lysine 27 (H3K27me3 promoting epigenetic gene silencing. EZH2 contains a SET domain promoting the methyltransferase activity while the three other protein components of PRC2, namely EED, SUZ12 and RpAp46/48 induce compaction of the chromatin permitting EZH2 enzymatic activity. Numerous studies highlight the role of this evolutionary conserved protein as a master regulator of differentiation in humans involved in the repression of the homeotic (Hox gene and the inactivation of X-chromosome. Through its effects in the epigenetic regulation of critical genes, EZH2 has been strongly linked to cell cycle progression, stem cell pluripotency and cancer biology. Most recently, EZH2 has been associated with hematopoietic stem cell proliferation and differentiation, thymopoiesis and lymphopoiesis. Several studies have evaluated the role of EZH2 in the regulation of T cell differentiation and plasticity as well as its implications in the development of autoimmune diseases and graft versus host disease (GvHD. In this review we will briefly summarize the current knowledge regarding the role of EZH2 in the regulation of T cell differentiation, effector function and homing in the tumor microenvironment and we will discuss possible therapeutic targeting of EZH2 in order to alter T cell immune functions.

  11. Cystatin F as a regulator of immune cell cytotoxicity.

    Science.gov (United States)

    Kos, Janko; Nanut, Milica Perišić; Prunk, Mateja; Sabotič, Jerica; Dautović, Esmeralda; Jewett, Anahid

    2018-05-10

    Cysteine cathepsins are lysosomal peptidases involved in the regulation of innate and adaptive immune responses. Among the diverse processes, regulation of granule-dependent cytotoxicity of cytotoxic T-lymphocytes (CTLs) and natural killer (NK) cells during cancer progression has recently gained significant attention. The function of cysteine cathepsins is regulated by endogenous cysteine protease inhibitors-cystatins. Whereas other cystatins are generally cytosolic or extracellular proteins, cystatin F is present in endosomes and lysosomes and is thus able to regulate the activity of its target directly. It is delivered to endosomal/lysosomal vesicles as an inactive, disulphide-linked dimer. Proteolytic cleavage of its N-terminal part leads to the monomer, the only form that is a potent inhibitor of cathepsins C, H and L, involved in the activation of granzymes and perforin. In NK cells and CTLs the levels of active cathepsin C and of granzyme B are dependent on the concentration of monomeric, active cystatin F. In tumour microenvironment, inactive dimeric cystatin F can be secreted from tumour cells or immune cells and further taken up by the cytotoxic cells. Subsequent monomerization and inhibition of cysteine cathepsins within the endosomal/lysosomal vesicles impairs granzyme and perforin activation, and provokes cell anergy. Further, the glycosylation pattern has been shown to be important in controlling secretion of cystatin F from target cells, as well as internalization by cytotoxic cells and trafficking to endosomal/lysosomal vesicles. Cystatin F is therefore an important mediator used by bystander cells to reduce NK and T-cell cytotoxicity.

  12. Exercise regulates breast cancer cell viability

    DEFF Research Database (Denmark)

    Dethlefsen, Christine; Lillelund, Christian; Midtgaard, Julie

    2016-01-01

    .003) and cytokines. Yet, these systemic adaptations had no effect on breast cancer cell viability in vitro. During 2 h of acute exercise, increases in serum lactate (6-fold, p ... no impact. Our data question the prevailing dogma that training-dependent baseline reductions in risk factors mediate the protective effect of exercise on breast cancer. Instead, we propose that the cancer protection is driven by accumulative effects of repeated acute exercise responses.......Purpose: Exercise decreases breast cancer risk and disease recurrence, but the underlying mechanisms are unknown. Training adaptations in systemic factors have been suggested as mediating causes. We aimed to examine if systemic adaptations to training over time, or acute exercise responses...

  13. NKT Cell Networks in the Regulation of Tumor Immunity

    Science.gov (United States)

    Robertson, Faith C.; Berzofsky, Jay A.; Terabe, Masaki

    2014-01-01

    CD1d-restricted natural killer T (NKT) cells lie at the interface between the innate and adaptive immune systems and are important mediators of immune responses and tumor immunosurveillance. These NKT cells uniquely recognize lipid antigens, and their rapid yet specific reactions influence both innate and adaptive immunity. In tumor immunity, two NKT subsets (type I and type II) have contrasting roles in which they not only cross-regulate one another, but also impact innate immune cell populations, including natural killer, dendritic, and myeloid lineage cells, as well as adaptive populations, especially CD8+ and CD4+ T cells. The extent to which NKT cells promote or suppress surrounding cells affects the host’s ability to prevent neoplasia and is consequently of great interest for therapeutic development. Data have shown the potential for therapeutic use of NKT cell agonists and synergy with immune response modifiers in both pre-clinical studies and preliminary clinical studies. However, there is room to improve treatment efficacy by further elucidating the biological mechanisms underlying NKT cell networks. Here, we discuss the progress made in understanding NKT cell networks, their consequent role in the regulation of tumor immunity, and the potential to exploit that knowledge in a clinical setting. PMID:25389427

  14. NKT cell networks in the regulation of tumor immunity.

    Science.gov (United States)

    Robertson, Faith C; Berzofsky, Jay A; Terabe, Masaki

    2014-01-01

    CD1d-restricted natural killer T (NKT) cells lie at the interface between the innate and adaptive immune systems and are important mediators of immune responses and tumor immunosurveillance. These NKT cells uniquely recognize lipid antigens, and their rapid yet specific reactions influence both innate and adaptive immunity. In tumor immunity, two NKT subsets (type I and type II) have contrasting roles in which they not only cross-regulate one another, but also impact innate immune cell populations, including natural killer, dendritic, and myeloid lineage cells, as well as adaptive populations, especially CD8(+) and CD4(+) T cells. The extent to which NKT cells promote or suppress surrounding cells affects the host's ability to prevent neoplasia and is consequently of great interest for therapeutic development. Data have shown the potential for therapeutic use of NKT cell agonists and synergy with immune response modifiers in both pre-clinical studies and preliminary clinical studies. However, there is room to improve treatment efficacy by further elucidating the biological mechanisms underlying NKT cell networks. Here, we discuss the progress made in understanding NKT cell networks, their consequent role in the regulation of tumor immunity, and the potential to exploit that knowledge in a clinical setting.

  15. NKT cell networks in the regulation of tumor immunity

    Directory of Open Access Journals (Sweden)

    Faith C Robertson

    2014-10-01

    Full Text Available CD1d-restricted natural killer T (NKT cells lie at the interface between the innate and adaptive immune systems and are important mediators of immune responses and tumor immunosurveillance. These NKT cells uniquely recognize lipid antigens, and their rapid yet specific reactions influence both innate and adaptive immunity. In tumor immunity, two NKT subsets (type I and type II have contrasting roles in which they not only cross-regulate one another, but also impact innate immune cell populations, including natural killer, dendritic and myeloid lineage cells, as well as adaptive populations, especially CD8+ and CD4+ T cells. The extent to which NKT cells promote or suppress surrounding cells affects the host’s ability to prevent neoplasia and is consequently of great interest for therapeutic development. Data have shown the potential for therapeutic use of NKT cell agonists and synergy with immune response modifiers in both pre-clinical studies and preliminary clinical studies. However, there is room to improve treatment efficacy by further elucidating the biological mechanisms underlying NKT cell networks. Here, we discuss the progress made in understanding NKT cell networks, their consequent role in the regulation of tumor immunity, and the potential to exploit that knowledge in a clinical setting.

  16. THE PROGRAMED CELL DEATH REGULATORS OF ISOLATED MODEL SYSTEMS

    Directory of Open Access Journals (Sweden)

    D. V. Vatlitsov

    2016-06-01

    Full Text Available The technology evolution creates the prerequisites for the emergence of new informational concept and approaches to the formation of a fundamentally new principles of biological objects understanding. The aim was to study the activators of the programmed cell death in an isolated system model. Cell culture aging parameters were performed on flow cytometer. It had formed the theory that the changes in the concentrations of metal ions and increase their extracellular concentration had formed a negative gradient into the cells.regulation of cell death. It was shown that the metals ions concentrations.

  17. α-Synuclein Amyloids Hijack Prion Protein to Gain Cell Entry, Facilitate Cell-to-Cell Spreading and Block Prion Replication.

    Science.gov (United States)

    Aulić, Suzana; Masperone, Lara; Narkiewicz, Joanna; Isopi, Elisa; Bistaffa, Edoardo; Ambrosetti, Elena; Pastore, Beatrice; De Cecco, Elena; Scaini, Denis; Zago, Paola; Moda, Fabio; Tagliavini, Fabrizio; Legname, Giuseppe

    2017-08-30

    The precise molecular mechanism of how misfolded α-synuclein (α-Syn) accumulates and spreads in synucleinopathies is still unknown. Here, we show the role of the cellular prion protein (PrP C ) in mediating the uptake and the spread of recombinant α-Syn amyloids. The in vitro data revealed that the presence of PrP C fosters the higher uptake of α-Syn amyloid fibrils, which was also confirmed in vivo in wild type (Prnp +/+ ) compared to PrP knock-out (Prnp -/- ) mice. Additionally, the presence of α-Syn amyloids blocked the replication of scrapie prions (PrP Sc ) in vitro and ex vivo, indicating a link between the two proteins. Indeed, whilst PrP C is mediating the internalization of α-Syn amyloids, PrP Sc is not able to replicate in their presence. This observation has pathological relevance, since several reported case studies show that the accumulation of α-Syn amyloid deposits in Creutzfeldt-Jakob disease patients is accompanied by a longer disease course.

  18. Regulation of Autophagy by Glucose in Mammalian Cells

    Directory of Open Access Journals (Sweden)

    Erwin Knecht

    2012-07-01

    Full Text Available Autophagy is an evolutionarily conserved process that contributes to maintain cell homeostasis. Although it is strongly regulated by many extracellular factors, induction of autophagy is mainly produced by starvation of nutrients. In mammalian cells, the regulation of autophagy by amino acids, and also by the hormone insulin, has been extensively investigated, but knowledge about the effects of other autophagy regulators, including another nutrient, glucose, is more limited. Here we will focus on the signalling pathways by which environmental glucose directly, i.e., independently of insulin and glucagon, regulates autophagy in mammalian cells, but we will also briefly mention some data in yeast. Although glucose deprivation mainly induces autophagy via AMPK activation and the subsequent inhibition of mTORC1, we will also comment other signalling pathways, as well as evidences indicating that, under certain conditions, autophagy can be activated by glucose. A better understanding on how glucose regulates autophagy not only will expand our basic knowledge of this important cell process, but it will be also relevant to understand common human disorders, such as cancer and diabetes, in which glucose levels play an important role.

  19. NFIX Regulates Neural Progenitor Cell Differentiation During Hippocampal Morphogenesis

    Science.gov (United States)

    Heng, Yee Hsieh Evelyn; McLeay, Robert C.; Harvey, Tracey J.; Smith, Aaron G.; Barry, Guy; Cato, Kathleen; Plachez, Céline; Little, Erica; Mason, Sharon; Dixon, Chantelle; Gronostajski, Richard M.; Bailey, Timothy L.; Richards, Linda J.; Piper, Michael

    2014-01-01

    Neural progenitor cells have the ability to give rise to neurons and glia in the embryonic, postnatal and adult brain. During development, the program regulating whether these cells divide and self-renew or exit the cell cycle and differentiate is tightly controlled, and imbalances to the normal trajectory of this process can lead to severe functional consequences. However, our understanding of the molecular regulation of these fundamental events remains limited. Moreover, processes underpinning development of the postnatal neurogenic niches within the cortex remain poorly defined. Here, we demonstrate that Nuclear factor one X (NFIX) is expressed by neural progenitor cells within the embryonic hippocampus, and that progenitor cell differentiation is delayed within Nfix−/− mice. Moreover, we reveal that the morphology of the dentate gyrus in postnatal Nfix−/− mice is abnormal, with fewer subgranular zone neural progenitor cells being generated in the absence of this transcription factor. Mechanistically, we demonstrate that the progenitor cell maintenance factor Sry-related HMG box 9 (SOX9) is upregulated in the hippocampus of Nfix−/− mice and demonstrate that NFIX can repress Sox9 promoter-driven transcription. Collectively, our findings demonstrate that NFIX plays a central role in hippocampal morphogenesis, regulating the formation of neuronal and glial populations within this structure. PMID:23042739

  20. Mast Cells Regulate Wound Healing in Diabetes.

    Science.gov (United States)

    Tellechea, Ana; Leal, Ermelindo C; Kafanas, Antonios; Auster, Michael E; Kuchibhotla, Sarada; Ostrovsky, Yana; Tecilazich, Francesco; Baltzis, Dimitrios; Zheng, Yongjun; Carvalho, Eugénia; Zabolotny, Janice M; Weng, Zuyi; Petra, Anastasia; Patel, Arti; Panagiotidou, Smaro; Pradhan-Nabzdyk, Leena; Theoharides, Theoharis C; Veves, Aristidis

    2016-07-01

    Diabetic foot ulceration is a severe complication of diabetes that lacks effective treatment. Mast cells (MCs) contribute to wound healing, but their role in diabetes skin complications is poorly understood. Here we show that the number of degranulated MCs is increased in unwounded forearm and foot skin of patients with diabetes and in unwounded dorsal skin of diabetic mice (P diabetic mice. Pretreatment with the MC degranulation inhibitor disodium cromoglycate rescues diabetes-associated wound-healing impairment in mice and shifts macrophages to the regenerative M2 phenotype (P diabetic mice deficient in MCs have delayed wound healing compared with their wild-type (WT) controls, implying that some MC mediator is needed for proper healing. MCs are a major source of vascular endothelial growth factor (VEGF) in mouse skin, but the level of VEGF is reduced in diabetic mouse skin, and its release from human MCs is reduced in hyperglycemic conditions. Topical treatment with the MC trigger substance P does not affect wound healing in MC-deficient mice, but improves it in WT mice. In conclusion, the presence of nondegranulated MCs in unwounded skin is required for proper wound healing, and therapies inhibiting MC degranulation could improve wound healing in diabetes. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

  1. Regulation of T Cell Differentiation and Function by EZH2

    Science.gov (United States)

    Karantanos, Theodoros; Christofides, Anthos; Bardhan, Kankana; Li, Lequn; Boussiotis, Vassiliki A.

    2016-01-01

    The enhancer of zeste homolog 2 (EZH2), one of the polycomb-group proteins, is the catalytic subunit of Polycomb-repressive complex 2 (PRC2) and induces the trimethylation of the histone H3 lysine 27 (H3K27me3) promoting epigenetic gene silencing. EZH2 contains a SET domain promoting the methyltransferase activity, while the three other protein components of PRC2, namely EED, SUZ12, and RpAp46/48, induce compaction of the chromatin permitting EZH2 enzymatic activity. Numerous studies highlight the role of this evolutionary conserved protein as a master regulator of differentiation in humans involved in the repression of the homeotic gene and the inactivation of X-chromosome. Through its effects in the epigenetic regulation of critical genes, EZH2 has been strongly linked to cell cycle progression, stem cell pluripotency, and cancer biology, being currently at the cutting edge of research. Most recently, EZH2 has been associated with hematopoietic stem cell proliferation and differentiation, thymopoiesis and lymphopoiesis. Several studies have evaluated the role of EZH2 in the regulation of T cell differentiation and plasticity as well as its implications in the development of autoimmune diseases and graft-versus-host disease (GVHD). The aim of this review is to summarize the current knowledge regarding the role of EZH2 in the regulation of the differentiation and function of T cells focusing on possible applications in various immune-mediated conditions, including autoimmune disorders and GVHD. PMID:27199994

  2. Hedgehog Signaling Regulates the Survival of Gastric Cancer Cells by Regulating the Expression of Bcl-2

    Science.gov (United States)

    Han, Myoung-Eun; Lee, Young-Suk; Baek, Sun-Yong; Kim, Bong-Seon; Kim, Jae-Bong; Oh, Sae-Ock

    2009-01-01

    Gastric cancer is the second most common cause of cancer deaths worldwide. The underlying molecular mechanisms of its carcinogenesis are relatively poorly characterized. Hedgehog (Hh) signaling, which is critical for development of various organs including the gastrointestinal tract, has been associated with gastric cancer. The present study was undertaken to reveal the underlying mechanism by which Hh signaling controls gastric cancer cell proliferation. Treatment of gastric cancer cells with cyclopamine, a specific inhibitor of Hh signaling pathway, reduced proliferation and induced apoptosis of gastric cancer cells. Cyclopamine treatment induced cytochrome c release from mitochondria and cleavage of caspase 9. Moreover, Bcl-2 expression was significantly reduced by cyclopamine treatment. These results suggest that Hh signaling regulates the survival of gastric cancer cells by regulating the expression of Bcl-2. PMID:19742123

  3. Estrogen receptor alpha is cell cycle-regulated and regulates the cell cycle in a ligand-dependent fashion.

    Science.gov (United States)

    JavanMoghadam, Sonia; Weihua, Zhang; Hunt, Kelly K; Keyomarsi, Khandan

    2016-06-17

    Estrogen receptor alpha (ERα) has been implicated in several cell cycle regulatory events and is an important predictive marker of disease outcome in breast cancer patients. Here, we aimed to elucidate the mechanism through which ERα influences proliferation in breast cancer cells. Our results show that ERα protein is cell cycle-regulated in human breast cancer cells and that the presence of 17-β-estradiol (E2) in the culture medium shortened the cell cycle significantly (by 4.5 hours, P cycle duration were observed in the S and G2/M phases, whereas the G1 phase was indistinguishable under liganded and unliganded conditions. In addition, ERα knockdown in MCF-7 cells accelerated mitotic exit, whereas transfection of ERα-negative MDA-MB-231 cells with exogenous ERα significantly shortened the S and G2/M phases (by 9.1 hours, P cycle progression through the S and G2/M phases than fulvestrant does, presumably because of the destabilizing effect of fulvestrant on ERα protein. Together, these results show that ERα modulates breast cancer cell proliferation by regulating events during the S and G2/M phases of the cell cycle in a ligand-dependent fashion. These results provide the rationale for an effective treatment strategy that includes a cell cycle inhibitor in combination with a drug that lowers estrogen levels, such as an aromatase inhibitor, and an antiestrogen that does not result in the degradation of ERα, such as tamoxifen.

  4. NSA2, a novel nucleolus protein regulates cell proliferation and cell cycle

    International Nuclear Information System (INIS)

    Zhang, Heyu; Ma, Xi; Shi, Taiping; Song, Quansheng; Zhao, Hongshan; Ma, Dalong

    2010-01-01

    NSA2 (Nop seven-associated 2) was previously identified in a high throughput screen of novel human genes associated with cell proliferation, and the NSA2 protein is evolutionarily conserved across different species. In this study, we revealed that NSA2 is broadly expressed in human tissues and cultured cell lines, and located in the nucleolus of the cell. Both of the putative nuclear localization signals (NLSs) of NSA2, also overlapped with nucleolar localization signals (NoLSs), are capable of directing nucleolar accumulation. Moreover, over-expression of the NSA2 protein promoted cell growth in different cell lines and regulated the G1/S transition in the cell cycle. SiRNA silencing of the NSA2 transcript attenuated the cell growth and dramatically blocked the cell cycle in G1/S transition. Our results demonstrated that NSA2 is a nucleolar protein involved in cell proliferation and cell cycle regulation.

  5. NK Cell Subtypes as Regulators of Autoimmune Liver Disease

    Directory of Open Access Journals (Sweden)

    Guohui Jiao

    2016-01-01

    Full Text Available As major components of innate immunity, NK cells not only exert cell-mediated cytotoxicity to destroy tumors or infected cells, but also act to regulate the functions of other cells in the immune system by secreting cytokines and chemokines. Thus, NK cells provide surveillance in the early defense against viruses, intracellular bacteria, and cancer cells. However, the effecter function of NK cells must be exquisitely controlled to prevent inadvertent attack against normal “self” cells. In an organ such as the liver, where the distinction between immunotolerance and immune defense against routinely processed pathogens is critical, the plethora of NK cells has a unique role in the maintenance of homeostasis. Once self-tolerance is broken, autoimmune liver disease resulted. NK cells act as a “two-edged weapon” and even play opposite roles with both regulatory and inducer activities in the hepatic environment. That is, NK cells act not only to produce inflammatory cytokines and chemokines, but also to alter the proliferation and activation of associated lymphocytes. However, the precise regulatory mechanisms at work in autoimmune liver diseases remain to be identified. In this review, we focus on recent research with NK cells and their potential role in the development of autoimmune liver disease.

  6. Margination of Stiffened Red Blood Cells Regulated By Vessel Geometry.

    Science.gov (United States)

    Chen, Yuanyuan; Li, Donghai; Li, Yongjian; Wan, Jiandi; Li, Jiang; Chen, Haosheng

    2017-11-10

    Margination of stiffened red blood cells has been implicated in many vascular diseases. Here, we report the margination of stiffened RBCs in vivo, and reveal the crucial role of the vessel geometry in the margination by calculations when the blood is seen as viscoelastic fluid. The vessel-geometry-regulated margination is then confirmed by in vitro experiments in microfluidic devices, and it establishes new insights to cell sorting technology and artificial blood vessel fabrication.

  7. Phosphorylation of Large T Antigen Regulates Merkel Cell Polyomavirus Replication

    International Nuclear Information System (INIS)

    Diaz, Jason; Wang, Xin; Tsang, Sabrina H.; Jiao, Jing; You, Jianxin

    2014-01-01

    Merkel Cell Polyomavirus (MCPyV) was recently discovered as a novel human polyomavirus that is associated with ~80% of Merkel Cell Carcinomas. The Large Tumor antigen (LT) is an early viral protein which has a variety of functions, including manipulation of the cell cycle and initiating viral DNA replication. Phosphorylation plays a critical regulatory role for polyomavirus LT proteins, but no investigation of MCPyV LT phosphorylation has been performed to date. In this report mass spectrometry analysis reveals three unique phosphorylation sites: T271, T297 and T299. In vivo replication assays confirm that phosphorylation of T271 does not play a role in viral replication, while modification at T297 and T299 have dramatic and opposing effects on LT’s ability to initiate replication from the viral origin. We test these mutants for their ability to bind, unwind, and act as a functional helicase at the viral origin. These studies provide a framework for understanding how phosphorylation of LT may dynamically regulate viral replication. Although the natural host cell of MCPyV has not yet been established, this work provides a foundation for understanding how LT activity is regulated and provides tools for better exploring this regulation in both natural host cells and Merkel cells

  8. Phosphorylation of Large T Antigen Regulates Merkel Cell Polyomavirus Replication

    Energy Technology Data Exchange (ETDEWEB)

    Diaz, Jason; Wang, Xin; Tsang, Sabrina H. [Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104 (United States); Jiao, Jing [Department of Pathology and Laboratory Medicine, Children’s Hospital of Philadelphia, Philadelphia, PA 19104 (United States); You, Jianxin, E-mail: jianyou@mail.med.upenn.edu [Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104 (United States)

    2014-07-08

    Merkel Cell Polyomavirus (MCPyV) was recently discovered as a novel human polyomavirus that is associated with ~80% of Merkel Cell Carcinomas. The Large Tumor antigen (LT) is an early viral protein which has a variety of functions, including manipulation of the cell cycle and initiating viral DNA replication. Phosphorylation plays a critical regulatory role for polyomavirus LT proteins, but no investigation of MCPyV LT phosphorylation has been performed to date. In this report mass spectrometry analysis reveals three unique phosphorylation sites: T271, T297 and T299. In vivo replication assays confirm that phosphorylation of T271 does not play a role in viral replication, while modification at T297 and T299 have dramatic and opposing effects on LT’s ability to initiate replication from the viral origin. We test these mutants for their ability to bind, unwind, and act as a functional helicase at the viral origin. These studies provide a framework for understanding how phosphorylation of LT may dynamically regulate viral replication. Although the natural host cell of MCPyV has not yet been established, this work provides a foundation for understanding how LT activity is regulated and provides tools for better exploring this regulation in both natural host cells and Merkel cells.

  9. Curcumin affects cell survival and cell volume regulation in human renal and intestinal cells

    Science.gov (United States)

    Kössler, Sonja; Nofziger, Charity; Jakab, Martin; Dossena, Silvia; Paulmichl, Markus

    2012-01-01

    Curcumin (1,7-bis(4-hydroxy-3-methoxyphenyl)-1E,6E-heptadiene-3,5-dione or diferuloyl methane) is a polyphenol derived from the Curcuma longa plant, commonly known as turmeric. This substance has been used extensively in Ayurvedic medicine for centuries for its anti-oxidant, analgesic, anti-inflammatory and antiseptic activity. More recently curcumin has been found to possess anti-cancer properties linked to its pro-apoptotic and anti-proliferative actions. The underlying mechanisms of these diverse effects are complex, not fully elucidated and subject of intense scientific debate. Despite increasing evidence indicating that different cation channels can be a molecular target for curcumin, very little is known about the effect of curcumin on chloride channels. Since, (i) the molecular structure of curcumin indicates that the substance could potentially interact with chloride channels, (ii) chloride channels play a role during the apoptotic process and regulation of the cell volume, and (iii) apoptosis is a well known effect of curcumin, we set out to investigate whether or not curcumin could (i) exert a modulatory effect (direct or indirect) on the swelling activated chloride current IClswell in a human cell system, therefore (ii) affect cell volume regulation and (iii) ultimately modulate cell survival. The IClswell channels, which are essential for regulating the cell volume after swelling, are also known to be activated under isotonic conditions as an early event in the apoptotic process. Here we show that long-term exposure of a human kidney cell line to extracellular 0.1–10 μM curcumin modulates IClswell in a dose-dependent manner (0.1 μM curcumin is ineffective, 0.5–5.0 μM curcumin increase, while 10 μM curcumin decrease the current), and short-term exposure to micromolar concentrations of curcumin does not affect IClswell neither if applied from the extracellular nor from the intracellular side – therefore, a direct effect of curcumin on

  10. Curcumin affects cell survival and cell volume regulation in human renal and intestinal cells

    International Nuclear Information System (INIS)

    Kössler, Sonja; Nofziger, Charity; Jakab, Martin; Dossena, Silvia; Paulmichl, Markus

    2012-01-01

    Curcumin (1,7-bis(4-hydroxy-3-methoxyphenyl)-1E,6E-heptadiene-3,5-dione or diferuloyl methane) is a polyphenol derived from the Curcuma longa plant, commonly known as turmeric. This substance has been used extensively in Ayurvedic medicine for centuries for its anti-oxidant, analgesic, anti-inflammatory and antiseptic activity. More recently curcumin has been found to possess anti-cancer properties linked to its pro-apoptotic and anti-proliferative actions. The underlying mechanisms of these diverse effects are complex, not fully elucidated and subject of intense scientific debate. Despite increasing evidence indicating that different cation channels can be a molecular target for curcumin, very little is known about the effect of curcumin on chloride channels. Since, (i) the molecular structure of curcumin indicates that the substance could potentially interact with chloride channels, (ii) chloride channels play a role during the apoptotic process and regulation of the cell volume, and (iii) apoptosis is a well known effect of curcumin, we set out to investigate whether or not curcumin could (i) exert a modulatory effect (direct or indirect) on the swelling activated chloride current ICl swell in a human cell system, therefore (ii) affect cell volume regulation and (iii) ultimately modulate cell survival. The ICl swell channels, which are essential for regulating the cell volume after swelling, are also known to be activated under isotonic conditions as an early event in the apoptotic process. Here we show that long-term exposure of a human kidney cell line to extracellular 0.1–10 μM curcumin modulates ICl swell in a dose-dependent manner (0.1 μM curcumin is ineffective, 0.5–5.0 μM curcumin increase, while 10 μM curcumin decrease the current), and short-term exposure to micromolar concentrations of curcumin does not affect ICl swell neither if applied from the extracellular nor from the intracellular side – therefore, a direct effect of curcumin on ICl

  11. Stem cell aging: mechanisms, regulators and therapeutic opportunities

    Science.gov (United States)

    Oh, Juhyun; Lee, Yang David; Wagers, Amy J

    2014-01-01

    Aging tissues experience a progressive decline in homeostatic and regenerative capacities, which has been attributed to degenerative changes in tissue-specific stem cells, stem cell niches and systemic cues that regulate stem cell activity. Understanding the molecular pathways involved in this age-dependent deterioration of stem cell function will be critical for developing new therapies for diseases of aging that target the specific causes of age-related functional decline. Here we explore key molecular pathways that are commonly perturbed as tissues and stem cells age and degenerate. We further consider experimental evidence both supporting and refuting the notion that modulation of these pathways per se can reverse aging phenotypes. Finally, we ask whether stem cell aging establishes an epigenetic ‘memory’ that is indelibly written or one that can be reset. PMID:25100532

  12. Regulation of basophil and mast cell development by transcription factors

    Directory of Open Access Journals (Sweden)

    Haruka Sasaki

    2016-04-01

    Full Text Available Basophils and mast cells play important roles in host defense against parasitic infections and allergic responses. Several progenitor populations, either shared or specific, for basophils and/or mast cells have been identified, thus elucidating the developmental pathways of these cells. Multiple transcription factors essential for their development and the relationships between them have been also revealed. For example, IRF8 induces GATA2 expression to promote the generation of both basophils and mast cells. The STAT5-GATA2 axis induces C/EBPα and MITF expression, facilitating the differentiation into basophils and mast cells, respectively. In addition, C/EBPα and MITF mutually suppress each other's expression. This review provides an overview of recent advances in our understanding of how transcription factors regulate the development of basophils and mast cells.

  13. Drosophila Glypicans Regulate Follicle Stem Cell Maintenance and Niche Competition.

    Science.gov (United States)

    Su, Tsu-Yi; Nakato, Eriko; Choi, Pui Yee; Nakato, Hiroshi

    2018-04-09

    Adult stem cells reside in specialized microenvironments, called niches, which provide signals for stem cells to maintain their undifferentiated and self-renewing state. To maintain stem cell quality, several types of stem cells are known to be regularly replaced by progenitor cells through niche competition. However, the cellular and molecular bases for stem cell competition for niche occupancy are largely unknown. Here, we show that two Drosophila members of the glypican family of heparan sulfate proteoglycans (HSPGs), Dally and Dally-like (Dlp), differentially regulate follicle stem cell (FSC) maintenance and FSC competitiveness for niche occupancy. Lineage analyses of glypican mutant FSC clones showed that dally is essential for normal FSC maintenance. In contrast, dlp is a hyper-competitive mutation: dlp mutant FSC progenitors often eventually occupy the entire epithelial sheet. RNAi knockdown experiments showed that Dally and Dlp play both partially redundant and distinct roles in regulating Jak/Stat, Wg and Hh signaling in FSCs. The Drosophila FSC system offers a powerful genetic model to study the mechanisms by which HSPGs exert specific functions in stem cell replacement and competition. Copyright © 2018, Genetics.

  14. Ascorbate regulates haematopoietic stem cell function and leukaemogenesis.

    Science.gov (United States)

    Agathocleous, Michalis; Meacham, Corbin E; Burgess, Rebecca J; Piskounova, Elena; Zhao, Zhiyu; Crane, Genevieve M; Cowin, Brianna L; Bruner, Emily; Murphy, Malea M; Chen, Weina; Spangrude, Gerald J; Hu, Zeping; DeBerardinis, Ralph J; Morrison, Sean J

    2017-09-28

    Stem-cell fate can be influenced by metabolite levels in culture, but it is not known whether physiological variations in metabolite levels in normal tissues regulate stem-cell function in vivo. Here we describe a metabolomics method for the analysis of rare cell populations isolated directly from tissues and use it to compare mouse haematopoietic stem cells (HSCs) to restricted haematopoietic progenitors. Each haematopoietic cell type had a distinct metabolic signature. Human and mouse HSCs had unusually high levels of ascorbate, which decreased with differentiation. Systemic ascorbate depletion in mice increased HSC frequency and function, in part by reducing the function of Tet2, a dioxygenase tumour suppressor. Ascorbate depletion cooperated with Flt3 internal tandem duplication (Flt3 ITD ) leukaemic mutations to accelerate leukaemogenesis, through cell-autonomous and possibly non-cell-autonomous mechanisms, in a manner that was reversed by dietary ascorbate. Ascorbate acted cell-autonomously to negatively regulate HSC function and myelopoiesis through Tet2-dependent and Tet2-independent mechanisms. Ascorbate therefore accumulates within HSCs to promote Tet activity in vivo, limiting HSC frequency and suppressing leukaemogenesis.

  15. A self-regulating hydrogen generator for micro fuel cells

    Energy Technology Data Exchange (ETDEWEB)

    Moghaddam, Saeed; Pengwang, Eakkachai; Shannon, Mark A. [Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 1206 West Green Street, Urbana, IL 61801 (United States); Masel, Richard I. [Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, 213 Roger Adams Lab, 600 S. Mathews, Urbana, IL 61801 (United States)

    2008-10-15

    The ever-increasing power demands and miniaturization of portable electronics, micro-sensors and actuators, and emerging technologies such as cognitive arthropods have created a significant interest in development of micro fuel cells. One of the major challenges in development of hydrogen micro fuel cells is the fabrication and integration of auxiliary systems for generating, regulating, and delivering hydrogen gas to the membrane electrode assembly (MEA). In this paper, we report the development of a hydrogen gas generator with a micro-scale control system that does not consume any power. The hydrogen generator consists of a hydride reactor and a water reservoir, with a regulating valve separating them. The regulating valve consists of a port from the water reservoir and a movable membrane with via holes that permit water to flow from the reservoir to the hydride reactor. Water flows towards the hydride reactor, but stops within the membrane via holes due to capillary forces. Water vapor then diffuses from the via holes into the hydride reactor resulting in generation of hydrogen gas. When the rate of hydrogen consumed by the MEA is lower than the generation rate, gas pressure builds up inside the hydride reactor, deflecting the membrane, closing the water regulator valve, until the pressure drops, whereby the valve reopens. We have integrated the self-regulating micro hydrogen generator to a MEA and successfully conducted fuel cell tests under varying load conditions. (author)

  16. VMP1 related autophagy and apoptosis in colorectal cancer cells: VMP1 regulates cell death

    Energy Technology Data Exchange (ETDEWEB)

    Qian, Qinyi [Department of Ultrasonograph, Changshu No. 2 People’s Hospital, Changshu (China); Zhou, Hao; Chen, Yan [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Shen, Chenglong [Department of General Surgery, Changshu No. 2 People’s Hospital, Changshu (China); He, Songbing; Zhao, Hua; Wang, Liang [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China); Wan, Daiwei, E-mail: 372710369@qq.com [Department of Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou (China); Gu, Wen, E-mail: 505339704@qq.com [Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou (China)

    2014-01-17

    Highlights: •This research confirmed VMP1 as a regulator of autophagy in colorectal cancer cell lines. •We proved the pro-survival role of VMP1-mediated autophagy in colorectal cancer cell lines. •We found the interaction between VMP1 and BECLIN1 also existing in colorectal cancer cell lines. -- Abstract: Vacuole membrane protein 1 (VMP1) is an autophagy-related protein and identified as a key regulator of autophagy in recent years. In pancreatic cell lines, VMP1-dependent autophagy has been linked to positive regulation of apoptosis. However, there are no published reports on the role of VMP1 in autophagy and apoptosis in colorectal cancers. Therefore, to address this gap of knowledge, we decided to interrogate regulation of autophagy and apoptosis by VMP1. We have studied the induction of autophagy by starvation and rapamycin treatment in colorectal cell lines using electron microscopy, immunofluorescence, and immunoblotting. We found that starvation-induced autophagy correlated with an increase in VMP1 expression, that VMP1 interacted with BECLIN1, and that siRNA mediated down-regulation of VMP1-reduced autophagy. Next, we examined the relationship between VMP1-dependent autophagy and apoptosis and found that VMP1 down-regulation sensitizes cells to apoptosis and that agents that induce apoptosis down-regulate VMP1. In conclusion, similar to its reported role in other cell types, VMP1 is an important regulator of autophagy in colorectal cell lines. However, in contrast to its role in pancreatic cell lines, in colorectal cancer cells, VMP1-dependent autophagy appears to be pro-survival rather than pro-cell death.

  17. Investigating microenvironmental regulation of human chordoma cell behaviour.

    Directory of Open Access Journals (Sweden)

    Priya Patel

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

  18. Electrostatic behavior of the charge-regulated bacterial cell surface.

    Science.gov (United States)

    Hong, Yongsuk; Brown, Derick G

    2008-05-06

    The electrostatic behavior of the charge-regulated surfaces of Gram-negative Escherichia coli and Gram-positive Bacillus brevis was studied using numerical modeling in conjunction with potentiometric titration and electrophoretic mobility data as a function of solution pH and electrolyte composition. Assuming a polyelectrolytic polymeric bacterial cell surface, these experimental and numerical analyses were used to determine the effective site numbers of cell surface acid-base functional groups and Ca(2+) sorption coefficients. Using effective site concentrations determined from 1:1 electrolyte (NaCl) experimental data, the charge-regulation model was able to replicate the effects of 2:1 electrolyte (CaCl(2)), both alone and as a mixture with NaCl, on the measured zeta potential using a single Ca(2+) surface binding constant for each of the bacterial species. This knowledge is vital for understanding how cells respond to changes in solution pH and electrolyte composition as well as how they interact with other surfaces. The latter is especially important due to the widespread use of the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory in the interpretation of bacterial adhesion. As surface charge and surface potential both vary on a charge-regulated surface, accurate modeling of bacterial interactions with surfaces ultimately requires use of an electrostatic model that accounts for the charge-regulated nature of the cell surface.

  19. Zfp206 regulates ES cell gene expression and differentiation.

    Science.gov (United States)

    Zhang, Wen; Walker, Emily; Tamplin, Owen J; Rossant, Janet; Stanford, William L; Hughes, Timothy R

    2006-01-01

    Understanding transcriptional regulation in early developmental stages is fundamental to understanding mammalian development and embryonic stem (ES) cell properties. Expression surveys suggest that the putative SCAN-Zinc finger transcription factor Zfp206 is expressed specifically in ES cells [Zhang,W., Morris,Q.D., Chang,R., Shai,O., Bakowski,M.A., Mitsakakis,N., Mohammad,N., Robinson,M.D., Zirngibl,R., Somogyi,E. et al., (2004) J. Biol., 3, 21; Brandenberger,R., Wei,H., Zhang,S., Lei,S., Murage,J., Fisk,G.J., Li,Y., Xu,C., Fang,R., Guegler,K. et al., (2004) Nat. Biotechnol., 22, 707-716]. Here, we confirm this observation, and we show that ZFP206 expression decreases rapidly upon differentiation of cultured mouse ES cells, and during development of mouse embryos. We find that there are at least six isoforms of the ZFP206 transcript, the longest being predominant. Overexpression and depletion experiments show that Zfp206 promotes formation of undifferentiated ES cell clones, and positively regulates abundance of a very small set of transcripts whose expression is also specific to ES cells and the two- to four-cell stages of preimplantation embryos. This set includes members of the Zscan4, Thoc4, Tcstv1 and eIF-1A gene families, none of which have been functionally characterized in vivo but whose members include apparent transcription factors, RNA-binding proteins and translation factors. Together, these data indicate that Zfp206 is a regulator of ES cell differentiation that controls a set of genes expressed very early in development, most of which themselves appear to be regulators.

  20. Ion channels involved in cell volume regulation: effects on migration, proliferation, and programmed cell death in non adherent EAT cells and adherent ELA cells.

    Science.gov (United States)

    Hoffmann, Else Kay

    2011-01-01

    This mini review outlines studies of cell volume regulation in two closely related mammalian cell lines: nonadherent Ehrlich ascites tumour cells (EATC) and adherent Ehrlich Lettre ascites (ELA) cells. Focus is on the regulatory volume decrease (RVD) that occurs after cell swelling, the volume regulatory ion channels involved, and the mechanisms (cellular signalling pathways) that regulate these channels. Finally, I shall also briefly review current investigations in these two cell lines that focuses on how changes in cell volume can regulate cell functions such as cell migration, proliferation, and programmed cell death. Copyright © 2011 S. Karger AG, Basel.

  1. Nrf2 regulates cellular behaviors and Notch signaling in oral squamous cell carcinoma cells.

    Science.gov (United States)

    Fan, Hong; Paiboonrungruan, Chorlada; Zhang, Xinyan; Prigge, Justin R; Schmidt, Edward E; Sun, Zheng; Chen, Xiaoxin

    2017-11-04

    Oxidative stress is known to play a pivotal role in the development of oral squamous cell carcinoma (OSCC). We have demonstrated that activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway has chemopreventive effects against oxidative stress-associated OSCC. However, Nrf2 have dual roles in cancer development; while it prevents carcinogenesis of normal cells, hyperactive Nrf2 also promotes the survival of cancer cells. This study is aimed to understand the function of Nrf2 in regulating cellular behaviors of OSCC cells, and the potential mechanisms through which Nrf2 facilitates OSCC. We established the Nrf2-overexpressing and Nrf2-knockdown OSCC cell lines, and examined the function of Nrf2 in regulating cell proliferation, migration, invasion, cell cycle and colony formation. Our data showed that Nrf2 overexpression promoted cancer phenotypes in OSCC cells, whereas Nrf2 silencing inhibited these phenotypes. In addition, Nrf2 positively regulated Notch signaling pathway in OSCC cells in vitro. Consistent with this observation, Nrf2 activation in Keap1 -/- mice resulted in not only hyperproliferation of squamous epithelial cells in mouse tongue as evidenced by increased expression of PCNA, but also activation of Notch signaling in these cells as evidenced by increased expression of NICD1 and Hes1. In conclusion, Nrf2 regulates cancer behaviors and Notch signaling in OSCC cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Ikaros limits follicular B cell activation by regulating B cell receptor signaling pathways

    International Nuclear Information System (INIS)

    Heizmann, Beate; Sellars, MacLean; Macias-Garcia, Alejandra; Chan, Susan; Kastner, Philippe

    2016-01-01

    The Ikaros transcription factor is essential for early B cell development, but its effect on mature B cells is debated. We show that Ikaros is required to limit the response of naive splenic B cells to B cell receptor signals. Ikaros deficient follicular B cells grow larger and enter cell cycle faster after anti-IgM stimulation. Unstimulated mutant B cells show deregulation of positive and negative regulators of signal transduction at the mRNA level, and constitutive phosphorylation of ERK, p38, SYK, BTK, AKT and LYN. Stimulation results in enhanced and prolonged ERK and p38 phosphorylation, followed by hyper-proliferation. Pharmacological inhibition of ERK and p38 abrogates the increased proliferative response of Ikaros deficient cells. These results suggest that Ikaros functions as a negative regulator of follicular B cell activation.

  3. Ikaros limits follicular B cell activation by regulating B cell receptor signaling pathways

    Energy Technology Data Exchange (ETDEWEB)

    Heizmann, Beate [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Sellars, MacLean [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Macias-Garcia, Alejandra [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Institute for Medical Engineering and Science at MIT, Cambridge, MA 02139 (United States); Chan, Susan, E-mail: scpk@igbmc.fr [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Kastner, Philippe, E-mail: scpk@igbmc.fr [Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), INSERM U964, CNRS UMR 7104, Université de Strasbourg, 67404 Illkirch (France); Faculté de Médecine, Université de Strasbourg, Strasbourg (France)

    2016-02-12

    The Ikaros transcription factor is essential for early B cell development, but its effect on mature B cells is debated. We show that Ikaros is required to limit the response of naive splenic B cells to B cell receptor signals. Ikaros deficient follicular B cells grow larger and enter cell cycle faster after anti-IgM stimulation. Unstimulated mutant B cells show deregulation of positive and negative regulators of signal transduction at the mRNA level, and constitutive phosphorylation of ERK, p38, SYK, BTK, AKT and LYN. Stimulation results in enhanced and prolonged ERK and p38 phosphorylation, followed by hyper-proliferation. Pharmacological inhibition of ERK and p38 abrogates the increased proliferative response of Ikaros deficient cells. These results suggest that Ikaros functions as a negative regulator of follicular B cell activation.

  4. [Regulation of airway stem cell proliferation in idiopathic pulmonary fibrosis].

    Science.gov (United States)

    Yang, S X; Wu, Q; Sun, X; Li, X; Li, K; Xu, L; Li, Y; Zhang, Q Y; Zhang, Y C; Chen, H Y

    2016-09-01

    To investigate the effect of fibroblasts on regulating airway stem cell proliferation in idiopathic pulmonary fibrosis. Lung cell suspension was prepared from β-actin-GFP mice. Airway stem cells were obtained by fluorescence activated cell sorting and co-cultured with lung fibroblasts. The fibroblasts were treated with TGF-β inhibitor SB43142. The expression of growth factors FGF1/2 and the effect of FGF1/2 on stem cell proliferation were observed. The cloning efficiency of airway stem cells, when co-cultured with normal lung fibroblast cells for 8 days, was (3.5±1.1)%, while the cloning efficiency was reduced to (0.04±0.04)% when co-cultured with lung fibroblasts from idiopathic pulmonary fibrosis patients. The difference between the 2 groups was statistically significant(P=0.002 5). TGF-β receptor inhibitor SB431542 increased lung fibroblast growth factors FGF1/2 expression.FGF1 mRNA expression was increased to the experimental group 0.005 5 from 0.000 2 in the control group.FGF2 mRNA expression of the amount raised to the experimental group 0.000 15 from 0.000 8 in the control group.FGF1/2 promoted the growth of airway stem cells. After FGF1/2 was co-cultured with normal lung fibroblast cells for 8 days, the cloning efficiency of airway stem cells was (0.3±0.1)%. During the development of idiopathic pulmonary fibrosis, fibroblast secreted FGF1/2 regulate airway stem cell proliferation.

  5. Spatio-temporal morphology changes in and quenching effects on the 2D spreading dynamics of cell colonies in both plain and methylcellulose-containing culture media.

    Science.gov (United States)

    Muzzio, N E; Pasquale, M A; Huergo, M A C; Bolzán, A E; González, P H; Arvia, A J

    2016-06-01

    To deal with complex systems, microscopic and global approaches become of particular interest. Our previous results from the dynamics of large cell colonies indicated that their 2D front roughness dynamics is compatible with the standard Kardar-Parisi-Zhang (KPZ) or the quenched KPZ equations either in plain or methylcellulose (MC)-containing gel culture media, respectively. In both cases, the influence of a non-uniform distribution of the colony constituents was significant. These results encouraged us to investigate the overall dynamics of those systems considering the morphology and size, the duplication rate, and the motility of single cells. For this purpose, colonies with different cell populations (N) exhibiting quasi-circular and quasi-linear growth fronts in plain and MC-containing culture media are investigated. For small N, the average radial front velocity and its change with time depend on MC concentration. MC in the medium interferes with cell mitosis, contributes to the local enlargement of cells, and increases the distribution of spatio-temporal cell density heterogeneities. Colony spreading in MC-containing media proceeds under two main quenching effects, I and II; the former mainly depending on the culture medium composition and structure and the latter caused by the distribution of enlarged local cell domains. For large N, colony spreading occurs at constant velocity. The characteristics of cell motility, assessed by measuring their trajectories and the corresponding velocity field, reflect the effect of enlarged, slow-moving cells and the structure of the medium. Local average cell size distribution and individual cell motility data from plain and MC-containing media are qualitatively consistent with the predictions of both the extended cellular Potts models and the observed transition of the front roughness dynamics from a standard KPZ to a quenched KPZ. In this case, quenching effects I and II cooperate and give rise to the quenched

  6. New insights into how trafficking regulates T cell receptor signaling

    Directory of Open Access Journals (Sweden)

    Jieqiong Lou

    2016-07-01

    Full Text Available AbstractThere is emerging evidence that exocytosis plays an important role in regulating T cell receptor (TCR signaling. The trafficking molecules involved in lytic granule (LG secretion in cytotoxic T lymphocytes (CTL have been well studied due to the immune disorder known as familial hemophagocytic lymphohisiocytosis (FHLH. However, the knowledge of trafficking machineries regulating the exocytosis of receptors and signaling molecules remains quite limited. In this review, we summarize the reported trafficking molecules involved in the transport of the TCR and downstream signaling molecules to the cell surface. By combining this information with the known knowledge of LG exocytosis and general exocytic trafficking machinery, we attempt to draw a more complete picture of how the TCR signaling network and exocytic trafficking matrix are interconnected to facilitate T cell activation. This also highlights how membrane compartmentalization facilitates the spatiotemporal organization of cellular responses that are essential for immune functions.

  7. Studies on regulation of the cell cycle in fission yeast.

    Directory of Open Access Journals (Sweden)

    Miroslava Požgajová

    2015-05-01

    Full Text Available All living organisms including plants and animals are composed of millions of cells. These cells perform different functions for the organism although they possess the same chromosomes and carry the same genetic information. Thus, to be able to understand multicellular organism we need to understand the life cycle of individual cells from which the organism comprises. The cell cycle is the life cycle of a single cell in the plant or animal body. It involves series of events in which components of the cell doubles and afterwards equally segregate into daughter cells. Such process ensures growth of the organism, and specialized reductional cell division which leads to production of gamets, assures sexual reproduction. Cell cycle is divided in the G1, S, G2 and M phase. Two gap-phases (G1 and G2 separate S phase (or synthesis and M phase which stays either for mitosis or meiosis. Essential for normal life progression and reproduction is correct chromosome segregation during mitosis and meiosis. Defects in the division program lead to aneuploidy, which in turn leads to birth defects, miscarriages or cancer. Even thou, researchers invented much about the regulation of the cell cycle, there is still long way to understand the complexity of the regulatory machineries that ensure proper segregation of chromosomes. In this paper we would like to describe techniques and materials we use for our studies on chromosome segregation in the model organism Schizosaccharomyces pombe.

  8. Cell volume regulation in epithelial physiology and cancer

    DEFF Research Database (Denmark)

    Pedersen, Stine Helene Falsig; Hoffmann, Else Kay; Novak, Ivana

    2013-01-01

    expression of ion transporters and channels is now recognized as one of the hallmarks of cancer, it is timely to consider this especially for epithelia. Epithelial cells are highly proliferative and epithelial cancers, carcinomas, account for about 90% of all cancers. In this review we will focus on ion...... such as cancer, transepithelial and cell volume regulatory ion transport are dys-regulated. Furthermore, epithelial architecture and coordinated ion transport function are lost, cell survival/death balance is altered, and new interactions with the stroma arise, all contributing to drug resistance. Since altered...... transporters and channels with key physiological functions in epithelia and known roles in the development of cancer in these tissues. Their roles in cell survival, cell cycle progression, and development of drug resistance in epithelial cancers will be discussed....

  9. A quantitative and dynamic model for plant stem cell regulation.

    Directory of Open Access Journals (Sweden)

    Florian Geier

    Full Text Available Plants maintain pools of totipotent stem cells throughout their entire life. These stem cells are embedded within specialized tissues called meristems, which form the growing points of the organism. The shoot apical meristem of the reference plant Arabidopsis thaliana is subdivided into several distinct domains, which execute diverse biological functions, such as tissue organization, cell-proliferation and differentiation. The number of cells required for growth and organ formation changes over the course of a plants life, while the structure of the meristem remains remarkably constant. Thus, regulatory systems must be in place, which allow for an adaptation of cell proliferation within the shoot apical meristem, while maintaining the organization at the tissue level. To advance our understanding of this dynamic tissue behavior, we measured domain sizes as well as cell division rates of the shoot apical meristem under various environmental conditions, which cause adaptations in meristem size. Based on our results we developed a mathematical model to explain the observed changes by a cell pool size dependent regulation of cell proliferation and differentiation, which is able to correctly predict CLV3 and WUS over-expression phenotypes. While the model shows stem cell homeostasis under constant growth conditions, it predicts a variation in stem cell number under changing conditions. Consistent with our experimental data this behavior is correlated with variations in cell proliferation. Therefore, we investigate different signaling mechanisms, which could stabilize stem cell number despite variations in cell proliferation. Our results shed light onto the dynamic constraints of stem cell pool maintenance in the shoot apical meristem of Arabidopsis in different environmental conditions and developmental states.

  10. Vinculin contributes to Cell Invasion by Regulating Contractile Activation

    Science.gov (United States)

    Mierke, Claudia Tanja

    2008-07-01

    Vinculin is a component of the focal adhesion complex and is described as a mechano-coupling protein connecting the integrin receptor and the actin cytoskeleton. Vinculin knock-out (k.o.) cells (vin-/-) displayed increased migration on a 2-D collagen- or fibronectin-coated substrate compared to wildtype cells, but the role of vinculin in cell migration through a 3-D connective tissue is unknown. We determined the invasiveness of established tumor cell lines using a 3-D collagen invasion assay. Gene expression analysis of 4 invasive and 4 non-invasive tumor cell lines revealed that vinculin expression was significantly increased in invasive tumor cell lines. To analyze the mechanisms by which vinculin increased cell invasion in a 3-D gel, we studied mouse embryonic fibroblasts wildtype and vin-/- cells. Wildtype cells were 3-fold more invasive compared vin-/- cells. We hypothesized that the ability to generate sufficient traction forces is a prerequisite for tumor cell migration in a 3-D connective tissue matrix. Using traction microscopy, we found that wildtype exerted 3-fold higher tractions on fibronectin-coated polyacrylamide gels compared to vin-/- cells. These results show that vinculin controls two fundamental functions that lead to opposite effects on cell migration in a 2-D vs. a 3-D environment: On the one hand, vinculin stabilizes the focal adhesions (mechano-coupling function) and thereby reduces motility in 2-D. On the other hand, vinculin is also a potent activator of traction generation (mechano-regulating function) that is important for cell invasion in a 3-D environment.

  11. Phosphoinositides: Tiny Lipids With Giant Impact on Cell Regulation

    Science.gov (United States)

    2013-01-01

    Phosphoinositides (PIs) make up only a small fraction of cellular phospholipids, yet they control almost all aspects of a cell's life and death. These lipids gained tremendous research interest as plasma membrane signaling molecules when discovered in the 1970s and 1980s. Research in the last 15 years has added a wide range of biological processes regulated by PIs, turning these lipids into one of the most universal signaling entities in eukaryotic cells. PIs control organelle biology by regulating vesicular trafficking, but they also modulate lipid distribution and metabolism via their close relationship with lipid transfer proteins. PIs regulate ion channels, pumps, and transporters and control both endocytic and exocytic processes. The nuclear phosphoinositides have grown from being an epiphenomenon to a research area of its own. As expected from such pleiotropic regulators, derangements of phosphoinositide metabolism are responsible for a number of human diseases ranging from rare genetic disorders to the most common ones such as cancer, obesity, and diabetes. Moreover, it is increasingly evident that a number of infectious agents hijack the PI regulatory systems of host cells for their intracellular movements, replication, and assembly. As a result, PI converting enzymes began to be noticed by pharmaceutical companies as potential therapeutic targets. This review is an attempt to give an overview of this enormous research field focusing on major developments in diverse areas of basic science linked to cellular physiology and disease. PMID:23899561

  12. Expression of E-selectin ligands on circulating tumor cells: cross-regulation with cancer stem cell regulatory pathways?

    International Nuclear Information System (INIS)

    Burdick, Monica M.; Henson, Karissa A.; Delgadillo, Luis F.; Choi, Young Eun; Goetz, Douglas J.; Tees, David F. J.; Benencia, Fabian

    2012-01-01

    Although significant progress has been made in the fight against cancer, successful treatment strategies have yet to be developed to combat those tumors that have metastasized to distant organs. Poor characterization of the molecular mechanisms of cancer spread is a major impediment to designing predictive diagnostics and effective clinical interventions against late stage disease. In hematogenous metastasis, it is widely suspected that circulating tumor cells (CTCs) express specific adhesion molecules that actively initiate contact with the vascular endothelium lining the vessel walls of the target organ. This “tethering” is mediated by ligands expressed by CTCs that bind to E-selectin expressed by endothelial cells. However, it is currently unknown whether expression of functional E-selectin ligands on CTCs is related to cancer stem cell regulatory or maintenance pathways, particularly epithelial-to-mesenchymal transition and the reverse, mesenchymal-to-epithelial transition. In this hypothesis and theory article, we explore the potential roles of these mechanisms on the dynamic regulation of selectin ligands mediating CTC trafficking during metastasis.

  13. TCR down-regulation controls T cell homeostasis

    DEFF Research Database (Denmark)

    Boding, Lasse; Bonefeld, Charlotte Menné; Nielsen, Bodil L

    2009-01-01

    TCR and cytokine receptor signaling play key roles in the complex homeostatic mechanisms that maintain a relative stable number of T cells throughout life. Despite the homeostatic mechanisms, a slow decline in naive T cells is typically observed with age. The CD3gamma di-leucine-based motif...... controls TCR down-regulation and plays a central role in fine-tuning TCR expression and signaling in T cells. In this study, we show that the age-associated decline of naive T cells is strongly accelerated in CD3gammaLLAA knock-in mice homozygous for a double leucine to alanine mutation in the CD3gamma di......-leucine-based motif, whereas the number of memory T cells is unaffected by the mutation. This results in premature T cell population senescence with a severe dominance of memory T cells and very few naive T cells in middle-aged to old CD3gamma mutant mice. The reduced number of naive T cells in CD3gamma mutant mice...

  14. Regulation of NKT Cell Localization in Homeostasis and Infection

    Science.gov (United States)

    Slauenwhite, Drew; Johnston, Brent

    2015-01-01

    Natural killer T (NKT) cells are a specialized subset of T lymphocytes that regulate immune responses in the context of autoimmunity, cancer, and microbial infection. Lipid antigens derived from bacteria, parasites, and fungi can be presented by CD1d molecules and recognized by the canonical T cell receptors on NKT cells. Alternatively, NKT cells can be activated through recognition of self-lipids and/or pro-inflammatory cytokines generated during infection. Unlike conventional T cells, only a small subset of NKT cells traffic through the lymph nodes under homeostatic conditions, with the largest NKT cell populations localizing to the liver, lungs, spleen, and bone marrow. This is thought to be mediated by differences in chemokine receptor expression profiles. However, the impact of infection on the tissue localization and function of NKT remains largely unstudied. This review focuses on the mechanisms mediating the establishment of peripheral NKT cell populations during homeostasis and how tissue localization of NKT cells is affected during infection. PMID:26074921

  15. Regulation of NKT Cell Localization in Homeostasis and Infection.

    Science.gov (United States)

    Slauenwhite, Drew; Johnston, Brent

    2015-01-01

    Natural killer T (NKT) cells are a specialized subset of T lymphocytes that regulate immune responses in the context of autoimmunity, cancer, and microbial infection. Lipid antigens derived from bacteria, parasites, and fungi can be presented by CD1d molecules and recognized by the canonical T cell receptors on NKT cells. Alternatively, NKT cells can be activated through recognition of self-lipids and/or pro-inflammatory cytokines generated during infection. Unlike conventional T cells, only a small subset of NKT cells traffic through the lymph nodes under homeostatic conditions, with the largest NKT cell populations localizing to the liver, lungs, spleen, and bone marrow. This is thought to be mediated by differences in chemokine receptor expression profiles. However, the impact of infection on the tissue localization and function of NKT remains largely unstudied. This review focuses on the mechanisms mediating the establishment of peripheral NKT cell populations during homeostasis and how tissue localization of NKT cells is affected during infection.

  16. ZDHHC3 Tyrosine Phosphorylation Regulates Neural Cell Adhesion Molecule Palmitoylation

    Science.gov (United States)

    Lievens, Patricia Marie-Jeanne; Kuznetsova, Tatiana; Kochlamazashvili, Gaga; Cesca, Fabrizia; Gorinski, Natalya; Galil, Dalia Abdel; Cherkas, Volodimir; Ronkina, Natalia; Lafera, Juri; Gaestel, Matthias

    2016-01-01

    The neural cell adhesion molecule (NCAM) mediates cell-cell and cell-matrix adhesion. It is broadly expressed in the nervous system and regulates neurite outgrowth, synaptogenesis, and synaptic plasticity. Previous in vitro studies revealed that palmitoylation of NCAM is required for fibroblast growth factor 2 (FGF2)-stimulated neurite outgrowth and identified the zinc finger DHHC (Asp-His-His-Cys)-containing proteins ZDHHC3 and ZDHHC7 as specific NCAM-palmitoylating enzymes. Here, we verified that FGF2 controlled NCAM palmitoylation in vivo and investigated molecular mechanisms regulating NCAM palmitoylation by ZDHHC3. Experiments with overexpression and pharmacological inhibition of FGF receptor (FGFR) and Src revealed that these kinases control tyrosine phosphorylation of ZDHHC3 and that ZDHHC3 is phosphorylated by endogenously expressed FGFR and Src proteins. By site-directed mutagenesis, we found that Tyr18 is an FGFR1-specific ZDHHC3 phosphorylation site, while Tyr295 and Tyr297 are specifically phosphorylated by Src kinase in cell-based and cell-free assays. Abrogation of tyrosine phosphorylation increased ZDHHC3 autopalmitoylation, enhanced interaction with NCAM, and upregulated NCAM palmitoylation. Expression of ZDHHC3 with tyrosine mutated in cultured hippocampal neurons promoted neurite outgrowth. Our findings for the first time highlight that FGFR- and Src-mediated tyrosine phosphorylation of ZDHHC3 modulates ZDHHC3 enzymatic activity and plays a role in neuronal morphogenesis. PMID:27247265

  17. The regulation of CD5 expression in murine T cells

    Directory of Open Access Journals (Sweden)

    Herzenberg Leonard A

    2001-05-01

    Full Text Available Abstract Background CD5 is a pan-T cell surface marker that is also present on a subset of B cells, B-1a cells.Functional and developmental subsets of T cells express characteristic CD5 levels that vary over roughly a 30-fold range. Previous investigators have cloned a 1.7 Kb fragment containing the CD5 promoter and showed that it can confer similar lymphocyte-specific expression pattern as observed for endogenous CD5 expression. Results We further characterize the CD5 promoter and identify minimal and regulatory regions on the CD5 promoter. Using a luciferase reporter system, we show that a 43 bp region on the CD5 promoter regulates CD5 expression in resting mouse thymoma EL4 T cells and that an Ets binding site within the 43 bp region mediates the CD5 expression. In addition, we show that Ets-1, a member of the Ets family of transcription factors, recognizes the Ets binding site in the electrophoretic mobility shift assay (EMSA. This Ets binding site is directly responsible for the increase in reporter activity when co-transfected with increasing amounts of Ets-1 expression plasmid. We also identify two additional evolutionarily-conserved regions in the CD5 promoter (CD5X and CD5Y and demonstrate the respective roles of the each region in the regulation of CD5 transcription. Conclusion Our studies define a minimal and regulatory promoter for CD5 and show that the CD5 expression level in T cells is at least partially dependent on the level of Ets-1 protein. Based on the findings in this report, we propose a model of CD5 transcriptional regulation in T cells.

  18. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  19. The epigenetic regulation of stem cell factors in hepatic stellate cells.

    Science.gov (United States)

    Reister, Sven; Kordes, Claus; Sawitza, Iris; Häussinger, Dieter

    2011-10-01

    The epigenetic regulation by DNA methylation is an important mechanism to control the expression of stem cell factors as demonstrated in tumor cells. It was recently shown that hepatic stellate cells (HSC) express stem/progenitor cell factors and have a differentiation potential. The aim of this work was to investigate if the expression of stem cell markers is regulated by DNA methylation during activation of rat HSC. It was found that CD133, Notch1, and Notch3 are regulated via DNA methylation in HSC, whereas Nestin shows no DNA methylation in HSC and other undifferentiated cells such as embryonic stem cells and umbilical cord blood stem cells from rats. In contrast to this, DNA methylation controls Nestin expression in differentiated cells like hepatocytes and the hepatoma cell line H4IIE. Demethylation by 5-Aza-2-deoxycytidine was sufficient to induce Nestin in H4IIE cells. In quiescent stellate cells and embryonic stem cells, the Nestin expression was suppressed by histone H3 methylation at lysine 9, which is another epigenetic mechanism. Apart from the known induction of Nestin in cultured HSC, this intermediate filament protein was also induced after partial hepatectomy, indicating activation of HSC during liver regeneration. Taken together, this study demonstrates for the first time that the expression of stem cell-associated factors such as CD133, Notch1, and Notch3 is controlled by DNA methylation in HSC. The regulation of Nestin by DNA methylation seems to be restricted to differentiated cells, whereas undifferentiated cells use different epigenetic mechanisms such as histone H3 methylation to control Nestin expression.

  20. EDR2 negatively regulates salicylic acid-based defenses and cell death during powdery mildew infections of Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Nishimura Marc

    2007-07-01

    Full Text Available Abstract Background The hypersensitive necrosis response (HR of resistant plants to avirulent pathogens is a form of programmed cell death in which the plant sacrifices a few cells under attack, restricting pathogen growth into adjacent healthy tissues. In spite of the importance of this defense response, relatively little is known about the plant components that execute the cell death program or about its regulation in response to pathogen attack. Results We isolated the edr2-6 mutant, an allele of the previously described edr2 mutants. We found that edr2-6 exhibited an exaggerated chlorosis and necrosis response to attack by three pathogens, two powdery mildew and one downy mildew species, but not in response to abiotic stresses or attack by the bacterial leaf speck pathogen. The chlorosis and necrosis did not spread beyond inoculated sites suggesting that EDR2 limits the initiation of cell death rather than its spread. The pathogen-induced chlorosis and necrosis of edr2-6 was correlated with a stimulation of the salicylic acid defense pathway and was suppressed in mutants deficient in salicylic acid signaling. EDR2 encodes a novel protein with a pleckstrin homology and a StAR transfer (START domain as well as a plant-specific domain of unknown function, DUF1336. The pleckstrin homology domain binds to phosphatidylinositol-4-phosphate in vitro and an EDR2:HA:GFP protein localizes to endoplasmic reticulum, plasma membrane and endosomes. Conclusion EDR2 acts as a negative regulator of cell death, specifically the cell death elicited by pathogen attack and mediated by the salicylic acid defense pathway. Phosphatidylinositol-4-phosphate may have a role in limiting cell death via its effect on EDR2. This role in cell death may be indirect, by helping to target EDR2 to the appropriate membrane, or it may play a more direct role.

  1. Role for nectin-1 in herpes simplex virus 1 entry and spread in human retinal pigment epithelial cells

    Science.gov (United States)

    Tiwari, Vaibhav; Oh, Myung-Jin; Kovacs, Maria; Shukla, Shripaad Y.; Valyi-Nagy, Tibor; Shukla, Deepak

    2009-01-01

    Herpes simplex virus 1 (HSV-1) demonstrates a unique ability to infect a variety of host cell types. Retinal pigment epithelial (RPE) cells form the outermost layer of the retina and provide a potential target for viral invasion and permanent vision impairment. Here we examine the initial cellular and molecular mechanisms that facilitate HSV-1 invasion of human RPE cells. High-resolution confocal microscopy demonstrated initial interaction of green fluorescent protein (GFP)-tagged virions with filopodia-like structures present on cell surfaces. Unidirectional movement of the virions on filopodia to the cell body was detected by live cell imaging of RPE cells, which demonstrated susceptibility to pH-dependent HSV-1 entry and replication. Use of RT-PCR indicated expression of nectin-1, herpes virus entry mediator (HVEM) and 3-O-sulfotransferase-3 (as a surrogate marker for 3-O-sulfated heparan sulfate). HVEM and nectin-1 expression was subsequently verified by flow cytometry. Nectin-1 expression in murine retinal tissue was also demonstrated by immunohistochemistry. Antibodies against nectin-1, but not HVEM, were able to block HSV-1 infection. Similar blocking effects were seen with a small interfering RNA construct specifically directed against nectin-1, which also blocked RPE cell fusion with HSV-1 glycoprotein-expressing Chinese hamster ovary (CHO-K1) cells. Anti-nectin-1 antibodies and F-actin depolymerizers were also successful in blocking the cytoskeletal changes that occur upon HSV-1 entry into cells. Our findings shed new light on the cellular and molecular mechanisms that help the virus to enter the cells of the inner eye. PMID:18803666

  2. The cell cycle-regulated genes of Schizosaccharomyces pombe.

    Science.gov (United States)

    Oliva, Anna; Rosebrock, Adam; Ferrezuelo, Francisco; Pyne, Saumyadipta; Chen, Haiying; Skiena, Steve; Futcher, Bruce; Leatherwood, Janet

    2005-07-01

    Many genes are regulated as an innate part of the eukaryotic cell cycle, and a complex transcriptional network helps enable the cyclic behavior of dividing cells. This transcriptional network has been studied in Saccharomyces cerevisiae (budding yeast) and elsewhere. To provide more perspective on these regulatory mechanisms, we have used microarrays to measure gene expression through the cell cycle of Schizosaccharomyces pombe (fission yeast). The 750 genes with the most significant oscillations were identified and analyzed. There were two broad waves of cell cycle transcription, one in early/mid G2 phase, and the other near the G2/M transition. The early/mid G2 wave included many genes involved in ribosome biogenesis, possibly explaining the cell cycle oscillation in protein synthesis in S. pombe. The G2/M wave included at least three distinctly regulated clusters of genes: one large cluster including mitosis, mitotic exit, and cell separation functions, one small cluster dedicated to DNA replication, and another small cluster dedicated to cytokinesis and division. S. pombe cell cycle genes have relatively long, complex promoters containing groups of multiple DNA sequence motifs, often of two, three, or more different kinds. Many of the genes, transcription factors, and regulatory mechanisms are conserved between S. pombe and S. cerevisiae. Finally, we found preliminary evidence for a nearly genome-wide oscillation in gene expression: 2,000 or more genes undergo slight oscillations in expression as a function of the cell cycle, although whether this is adaptive, or incidental to other events in the cell, such as chromatin condensation, we do not know.

  3. The Cell Cycle–Regulated Genes of Schizosaccharomyces pombe

    Science.gov (United States)

    Oliva, Anna; Rosebrock, Adam; Ferrezuelo, Francisco; Pyne, Saumyadipta; Chen, Haiying; Skiena, Steve

    2005-01-01

    Many genes are regulated as an innate part of the eukaryotic cell cycle, and a complex transcriptional network helps enable the cyclic behavior of dividing cells. This transcriptional network has been studied in Saccharomyces cerevisiae (budding yeast) and elsewhere. To provide more perspective on these regulatory mechanisms, we have used microarrays to measure gene expression through the cell cycle of Schizosaccharomyces pombe (fission yeast). The 750 genes with the most significant oscillations were identified and analyzed. There were two broad waves of cell cycle transcription, one in early/mid G2 phase, and the other near the G2/M transition. The early/mid G2 wave included many genes involved in ribosome biogenesis, possibly explaining the cell cycle oscillation in protein synthesis in S. pombe. The G2/M wave included at least three distinctly regulated clusters of genes: one large cluster including mitosis, mitotic exit, and cell separation functions, one small cluster dedicated to DNA replication, and another small cluster dedicated to cytokinesis and division. S. pombe cell cycle genes have relatively long, complex promoters containing groups of multiple DNA sequence motifs, often of two, three, or more different kinds. Many of the genes, transcription factors, and regulatory mechanisms are conserved between S. pombe and S. cerevisiae. Finally, we found preliminary evidence for a nearly genome-wide oscillation in gene expression: 2,000 or more genes undergo slight oscillations in expression as a function of the cell cycle, although whether this is adaptive, or incidental to other events in the cell, such as chromatin condensation, we do not know. PMID:15966770

  4. The cell cycle-regulated genes of Schizosaccharomyces pombe.

    Directory of Open Access Journals (Sweden)

    Anna Oliva

    2005-07-01

    Full Text Available Many genes are regulated as an innate part of the eukaryotic cell cycle, and a complex transcriptional network helps enable the cyclic behavior of dividing cells. This transcriptional network has been studied in Saccharomyces cerevisiae (budding yeast and elsewhere. To provide more perspective on these regulatory mechanisms, we have used microarrays to measure gene expression through the cell cycle of Schizosaccharomyces pombe (fission yeast. The 750 genes with the most significant oscillations were identified and analyzed. There were two broad waves of cell cycle transcription, one in early/mid G2 phase, and the other near the G2/M transition. The early/mid G2 wave included many genes involved in ribosome biogenesis, possibly explaining the cell cycle oscillation in protein synthesis in S. pombe. The G2/M wave included at least three distinctly regulated clusters of genes: one large cluster including mitosis, mitotic exit, and cell separation functions, one small cluster dedicated to DNA replication, and another small cluster dedicated to cytokinesis and division. S. pombe cell cycle genes have relatively long, complex promoters containing groups of multiple DNA sequence motifs, often of two, three, or more different kinds. Many of the genes, transcription factors, and regulatory mechanisms are conserved between S. pombe and S. cerevisiae. Finally, we found preliminary evidence for a nearly genome-wide oscillation in gene expression: 2,000 or more genes undergo slight oscillations in expression as a function of the cell cycle, although whether this is adaptive, or incidental to other events in the cell, such as chromatin condensation, we do not know.

  5. Peripheral serotonin regulates maternal calcium trafficking in mammary epithelial cells during lactation in mice.

    Directory of Open Access Journals (Sweden)

    Jimena Laporta

    Full Text Available Lactation is characterized by massive transcellular flux of calcium, from the basolateral side of the mammary alveolar epithelium (blood into the ductal lumen (milk. Regulation of calcium transport during lactation is critical for maternal and neonatal health. The monoamine serotonin (5-HT is synthesized by the mammary gland and functions as a homeostatic regulation of lactation. Genetic ablation of tryptophan hydroxylase 1 (Tph1, which encodes the rate-limiting enzyme in non-neuronal serotonin synthesis, causes a deficiency in circulating serotonin. As a consequence maternal calcium concentrations decrease, mammary epithelial cell morphology is altered, and cell proliferation is decreased during lactation. Here we demonstrate that serotonin deficiency decreases the expression and disrupts the normal localization of calcium transporters located in the apical (PMCA2 and basolateral (CaSR, ORAI-1 membranes of the lactating mammary gland. In addition, serotonin deficiency decreases the mRNA expression of calcium transporters located in intracellular compartments (SERCA2, SPCA1 and 2. Mammary expression of serotonin receptor isoform 2b and its downstream pathways (PLCβ3, PKC and MAP-ERK1/2 are also decreased by serotonin deficiency, which might explain the numerous phenotypic alterations described above. In most cases, addition of exogenous 5-hydroxy-L-tryptophan to the Tph1 deficient mice rescued the phenotype. Our data supports the hypothesis that serotonin is necessary for proper mammary gland structure and function, to regulate blood and mammary epithelial cell transport of calcium during lactation. These findings can be applicable to the treatment of lactation-induced hypocalcemia in dairy cows and can have profound implications in humans, given the wide-spread use of selective serotonin reuptake inhibitors as antidepressants during pregnancy and lactation.

  6. Leading research on cell proliferation regulation technology; Saibo zoshoku seigyo gijutsu no sendo kenkyu

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-03-01

    For developing intelligent material, animal test alternative model, bio-cell analysis equipment, self-controlling bio-reactor and medical material, development of functional cells was studied by cell proliferation regulation technology. In fiscal 1996, the expression analysis and separation technology of specific gene for cell proliferation, and the intracellular regulation technology were surveyed from the viewpoint of intracellular regulation. The cell proliferation regulation technology by specific regulating material of cells, extracellular matrix, coculture system and embryonic cell was surveyed from the viewpoint of extracellular regulation. In addition, based on these survey results, new cell culture/analysis technology, new bio-material, artificial organ system, energy saving bio-reactor, environment purification microorganism, and animal test alternative model were surveyed as applications to industrial basic technologies from a long-term viewpoint. The approach to cell proliferation regulation requires preparation of a concrete proliferation regulation technology system of cells, and concrete application targets. 268 refs., 43 figs., 4 tabs.

  7. Insulin promotes cell migration by regulating PSA-NCAM

    Energy Technology Data Exchange (ETDEWEB)

    Monzo, Hector J.; Coppieters, Natacha [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Park, Thomas I.H. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dieriks, Birger V.; Faull, Richard L.M. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dragunow, Mike [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Curtis, Maurice A., E-mail: m.curtis@auckland.ac.nz [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand)

    2017-06-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  8. Insulin promotes cell migration by regulating PSA-NCAM

    International Nuclear Information System (INIS)

    Monzo, Hector J.; Coppieters, Natacha; Park, Thomas I.H.; Dieriks, Birger V.; Faull, Richard L.M.; Dragunow, Mike; Curtis, Maurice A.

    2017-01-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  9. Platelet rich plasma promotes skeletal muscle cell migration in association with up-regulation of FAK, paxillin, and F-Actin formation.

    Science.gov (United States)

    Tsai, Wen-Chung; Yu, Tung-Yang; Lin, Li-Ping; Lin, Mioa-Sui; Tsai, Ting-Ta; Pang, Jong-Hwei S

    2017-11-01

    Platelet rich plasma (PRP) contains various cytokines and growth factors which may be beneficial to the healing process of injured muscle. The aim of this study was to investigate the effect and molecular mechanism of PRP on migration of skeletal muscle cells. Skeletal muscle cells intrinsic to Sprague-Dawley rats were treated with PRP. The cell migration was evaluated by transwell filter migration assay and electric cell-substrate impedance sensing. The spreading of cells was evaluated microscopically. The formation of filamentous actin (F-actin) cytoskeleton was assessed by immunofluorescence staining. The protein expressions of paxillin and focal adhesion kinase (FAK) were assessed by Western blot analysis. Transfection of paxillin small-interfering RNA (siRNAs) to muscle cells was performed to validate the role of paxillin in PRP-mediated promotion of cell migration. Dose-dependently PRP promotes migration of and spreading and muscle cells. Protein expressions of paxillin and FAK were up-regulated dose-dependently. F-actin formation was also enhanced by PRP treatment. Furthermore, the knockdown of paxillin expression impaired the effect of PRP to promote cell migration. It was concluded that PRP promoting migration of muscle cells is associated with up-regulation of proteins expression of paxillin and FAK as well as increasing F-actin formation. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2506-2512, 2017. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  10. A dual character of flavonoids in influenza A virus replication and spread through modulating cell-autonomous immunity by MAPK signaling pathways

    Science.gov (United States)

    Dong, Wenjuan; Wei, Xiuli; Zhang, Fayun; Hao, Junfeng; Huang, Feng; Zhang, Chunling; Liang, Wei

    2014-01-01

    Flavonoids are well known as a large class of polyphenolic compounds, which have a variety of physiological activities, including anti-influenza virus activity. The influenza A/WSN/33 infected A549 cells have been used to screen anti-influenza virus drugs from natural flavonoid compounds library. Unexpectedly, some flavonoid compounds significantly inhibited virus replication, while the others dramatically promoted virus replication. In this study, we attempted to understand these differences between flavonoid compounds in their antivirus mechanisms. Hesperidin and kaempferol were chosen as representatives of both sides, each of which exhibited the opposite effects on influenza virus replication. Our investigation revealed that the opposite effects produced by hesperidin and kaempferol on influenza virus were due to inducing the opposite cell-autonomous immune responses by selectively modulating MAP kinase pathways: hesperidin up-regulated P38 and JNK expression and activation, thus resulting in the enhanced cell-autonomous immunity; while kaempferol dramatically down-regulated p38 and JNK expression and activation, thereby suppressing cell-autonomous immunity. In addition, hesperidin restricted RNPs export from nucleus by down-regulating ERK activation, but kaempferol promoted RNPs export by up-regulating ERK activation. Our findings demonstrate that a new generation of anti-influenza virus drugs could be developed based on selective modulation of MAP kinase pathways to stimulate cell-autonomous immunity. PMID:25429875

  11. Mitochondrial peroxiredoxin 3 regulates sensory cell survival in the cochlea.

    Directory of Open Access Journals (Sweden)

    Fu-Quan Chen

    Full Text Available This study delineates the role of peroxiredoxin 3 (Prx3 in hair cell death induced by several etiologies of acquired hearing loss (noise trauma, aminoglycoside treatment, age. In vivo, Prx3 transiently increased in mouse cochlear hair cells after traumatic noise exposure, kanamycin treatment, or with progressing age before any cell loss occurred; when Prx3 declined, hair cell loss began. Maintenance of high Prx3 levels via treatment with the radical scavenger 2,3-dihydroxybenzoate prevented kanamycin-induced hair cell death. Conversely, reducing Prx3 levels with Prx3 siRNA increased the severity of noise-induced trauma. In mouse organ of Corti explants, reactive oxygen species and levels of Prx3 mRNA and protein increased concomitantly at early times of drug challenge. When Prx3 levels declined after prolonged treatment, hair cells began to die. The radical scavenger p-phenylenediamine maintained Prx3 levels and attenuated gentamicin-induced hair cell death. Our results suggest that Prx3 is up-regulated in response to oxidative stress and that maintenance of Prx3 levels in hair cells is a critical factor in their susceptibility to acquired hearing loss.

  12. A Single HIV-1 Cluster and a Skewed Immune Homeostasis Drive the Early Spread of HIV among Resting CD4+ Cell Subsets within One Month Post-Infection

    Science.gov (United States)

    Avettand-Fenoël, Véronique; Nembot, Georges; Mélard, Adeline; Blanc, Catherine; Lascoux-Combe, Caroline; Slama, Laurence; Allegre, Thierry; Allavena, Clotilde; Yazdanpanah, Yazdan; Duvivier, Claudine; Katlama, Christine; Goujard, Cécile; Seksik, Bao Chau Phung; Leplatois, Anne; Molina, Jean-Michel; Meyer, Laurence; Autran, Brigitte; Rouzioux, Christine

    2013-01-01

    Optimizing therapeutic strategies for an HIV cure requires better understanding the characteristics of early HIV-1 spread among resting CD4+ cells within the first month of primary HIV-1 infection (PHI). We studied the immune distribution, diversity, and inducibility of total HIV-DNA among the following cell subsets: monocytes, peripheral blood activated and resting CD4 T cells, long-lived (naive [TN] and central-memory [TCM]) and short-lived (transitional-memory [TTM] and effector-memory cells [TEM]) resting CD4+T cells from 12 acutely-infected individuals recruited at a median 36 days from infection. Cells were sorted for total HIV-DNA quantification, phylogenetic analysis and inducibility, all studied in relation to activation status and cell signaling. One month post-infection, a single CCR5-restricted viral cluster was massively distributed in all resting CD4+ subsets from 88% subjects, while one subject showed a slight diversity. High levels of total HIV-DNA were measured among TN (median 3.4 log copies/million cells), although 10-fold less (p = 0.0005) than in equally infected TCM (4.5), TTM (4.7) and TEM (4.6) cells. CD3−CD4+ monocytes harbored a low viral burden (median 2.3 log copies/million cells), unlike equally infected resting and activated CD4+ T cells (4.5 log copies/million cells). The skewed repartition of resting CD4 subsets influenced their contribution to the pool of resting infected CD4+T cells, two thirds of which consisted of short-lived TTM and TEM subsets, whereas long-lived TN and TCM subsets contributed the balance. Each resting CD4 subset produced HIV in vitro after stimulation with anti-CD3/anti-CD28+IL-2 with kinetics and magnitude varying according to subset differentiation, while IL-7 preferentially induced virus production from long-lived resting TN cells. In conclusion, within a month of infection, a clonal HIV-1 cluster is massively distributed among resting CD4 T-cell subsets with a flexible inducibility, suggesting that

  13. A single HIV-1 cluster and a skewed immune homeostasis drive the early spread of HIV among resting CD4+ cell subsets within one month post-infection.

    Directory of Open Access Journals (Sweden)

    Charline Bacchus

    Full Text Available Optimizing therapeutic strategies for an HIV cure requires better understanding the characteristics of early HIV-1 spread among resting CD4+ cells within the first month of primary HIV-1 infection (PHI. We studied the immune distribution, diversity, and inducibility of total HIV-DNA among the following cell subsets: monocytes, peripheral blood activated and resting CD4 T cells, long-lived (naive [TN] and central-memory [TCM] and short-lived (transitional-memory [TTM] and effector-memory cells [TEM] resting CD4+T cells from 12 acutely-infected individuals recruited at a median 36 days from infection. Cells were sorted for total HIV-DNA quantification, phylogenetic analysis and inducibility, all studied in relation to activation status and cell signaling. One month post-infection, a single CCR5-restricted viral cluster was massively distributed in all resting CD4+ subsets from 88% subjects, while one subject showed a slight diversity. High levels of total HIV-DNA were measured among TN (median 3.4 log copies/million cells, although 10-fold less (p = 0.0005 than in equally infected TCM (4.5, TTM (4.7 and TEM (4.6 cells. CD3-CD4+ monocytes harbored a low viral burden (median 2.3 log copies/million cells, unlike equally infected resting and activated CD4+ T cells (4.5 log copies/million cells. The skewed repartition of resting CD4 subsets influenced their contribution to the pool of resting infected CD4+T cells, two thirds of which consisted of short-lived TTM and TEM subsets, whereas long-lived TN and TCM subsets contributed the balance. Each resting CD4 subset produced HIV in vitro after stimulation with anti-CD3/anti-CD28+IL-2 with kinetics and magnitude varying according to subset differentiation, while IL-7 preferentially induced virus production from long-lived resting TN cells. In conclusion, within a month of infection, a clonal HIV-1 cluster is massively distributed among resting CD4 T-cell subsets with a flexible inducibility

  14. TIM-1 signaling in B cells regulates antibody production

    International Nuclear Information System (INIS)

    Ma, Juan; Usui, Yoshihiko; Takeda, Kazuyoshi; Harada, Norihiro; Yagita, Hideo; Okumura, Ko; Akiba, Hisaya

    2011-01-01

    Highlights: → TIM-1 is highly expressed on anti-IgM + anti-CD40-stimulated B cells. → Anti-TIM-1 mAb enhanced proliferation and Ig production on activated B cell in vitro. → TIM-1 signaling regulates Ab production by response to TI-2 and TD antigens in vivo. -- Abstract: Members of the T cell Ig and mucin (TIM) family have recently been implicated in the control of T cell-mediated immune responses. In this study, we found TIM-1 expression on anti-IgM- or anti-CD40-stimulated splenic B cells, which was further up-regulated by the combination of anti-IgM and anti-CD40 Abs. On the other hand, TIM-1 ligand was constitutively expressed on B cells and inducible on anti-CD3 + anti-CD28-stimulated CD4 + T cells. In vitro stimulation of activated B cells by anti-TIM-1 mAb enhanced proliferation and expression of a plasma cell marker syndecan-1 (CD138). We further examined the effect of TIM-1 signaling on antibody production in vitro and in vivo. Higher levels of IgG2b and IgG3 secretion were detected in the culture supernatants of the anti-TIM-1-stimulated B cells as compared with the control IgG-stimulated B cells. When immunized with T-independent antigen TNP-Ficoll, TNP-specific IgG1, IgG2b, and IgG3 Abs were slightly increased in the anti-TIM-1-treated mice. When immunized with T-dependent antigen OVA, serum levels of OVA-specific IgG2b, IgG3, and IgE Abs were significantly increased in the anti-TIM-1-treated mice as compared with the control IgG-treated mice. These results suggest that TIM-1 signaling in B cells augments antibody production by enhancing B cell proliferation and differentiation.

  15. TIM-1 signaling in B cells regulates antibody production

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Juan [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Usui, Yoshihiko [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Department of Ophthalmology, Tokyo Medical University, 6-7-1 Nishi-shinjuku-ku, Tokyo 160-0023 (Japan); Takeda, Kazuyoshi [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Harada, Norihiro [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Department of Respiratory Medicine, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Research Institute for Diseases of Old Ages, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Yagita, Hideo; Okumura, Ko [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Akiba, Hisaya, E-mail: hisaya@juntendo.ac.jp [Department of Immunology, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan)

    2011-03-11

    Highlights: {yields} TIM-1 is highly expressed on anti-IgM + anti-CD40-stimulated B cells. {yields} Anti-TIM-1 mAb enhanced proliferation and Ig production on activated B cell in vitro. {yields} TIM-1 signaling regulates Ab production by response to TI-2 and TD antigens in vivo. -- Abstract: Members of the T cell Ig and mucin (TIM) family have recently been implicated in the control of T cell-mediated immune responses. In this study, we found TIM-1 expression on anti-IgM- or anti-CD40-stimulated splenic B cells, which was further up-regulated by the combination of anti-IgM and anti-CD40 Abs. On the other hand, TIM-1 ligand was constitutively expressed on B cells and inducible on anti-CD3{sup +} anti-CD28-stimulated CD4{sup +} T cells. In vitro stimulation of activated B cells by anti-TIM-1 mAb enhanced proliferation and expression of a plasma cell marker syndecan-1 (CD138). We further examined the effect of TIM-1 signaling on antibody production in vitro and in vivo. Higher levels of IgG2b and IgG3 secretion were detected in the culture supernatants of the anti-TIM-1-stimulated B cells as compared with the control IgG-stimulated B cells. When immunized with T-independent antigen TNP-Ficoll, TNP-specific IgG1, IgG2b, and IgG3 Abs were slightly increased in the anti-TIM-1-treated mice. When immunized with T-dependent antigen OVA, serum levels of OVA-specific IgG2b, IgG3, and IgE Abs were significantly increased in the anti-TIM-1-treated mice as compared with the control IgG-treated mice. These results suggest that TIM-1 signaling in B cells augments antibody production by enhancing B cell proliferation and differentiation.

  16. Analysis of mammary specific gene locus regulation in differentiated cells derived by somatic cell fusion

    International Nuclear Information System (INIS)

    Robinson, Claire; Kolb, Andreas F.

    2009-01-01

    The transcriptional regulation of a gene is best analysed in the context of its normal chromatin surroundings. However, most somatic cells, in contrast to embryonic stem cells, are refractory to accurate modification by homologous recombination. We show here that it is possible to introduce precise genomic modifications in ES cells and to analyse the phenotypic consequences in differentiated cells by using a combination of gene targeting, site-specific recombination and somatic cell fusion. To provide a proof of principle, we have analysed the regulation of the casein gene locus in mammary gland cells derived from modified murine ES cells by somatic cell fusion. A β-galactosidase reporter gene was inserted in place of the β-casein gene and the modified ES cells, which do not express the reporter gene, were fused with the mouse mammary gland cell line HC11. The resulting cell clones expressed the β-galactosidase gene to a similar extent and with similar hormone responsiveness as the endogenous gene. However, a reporter gene under the control of a minimal β-casein promoter (encompassing the two consensus STAT5 binding sites which mediate the hormone response of the casein genes) was unable to replicate expression levels or hormone responsiveness of the endogenous gene when inserted into the same site of the casein locus. As expected, these results implicate sequences other than the STAT5 sites in the regulation of the β-casein gene

  17. SOX2 regulates acinar cell development in the salivary gland

    Science.gov (United States)

    Emmerson, Elaine; May, Alison J; Nathan, Sara; Cruz-Pacheco, Noel; Lizama, Carlos O; Maliskova, Lenka; Zovein, Ann C; Shen, Yin; Muench, Marcus O; Knox, Sarah M

    2017-01-01

    Acinar cells play an essential role in the secretory function of exocrine organs. Despite this requirement, how acinar cells are generated during organogenesis is unclear. Using the acini-ductal network of the developing human and murine salivary gland, we demonstrate an unexpected role for SOX2 and parasympathetic nerves in generating the acinar lineage that has broad implications for epithelial morphogenesis. Despite SOX2 being expressed by progenitors that give rise to both acinar and duct cells, genetic ablation of SOX2 results in a failure to establish acini but not ducts. Furthermore, we show that SOX2 targets acinar-specific genes and is essential for the survival of acinar but not ductal cells. Finally, we illustrate an unexpected and novel role for peripheral nerves in the creation of acini throughout development via regulation of SOX2. Thus, SOX2 is a master regulator of the acinar cell lineage essential to the establishment of a functional organ. DOI: http://dx.doi.org/10.7554/eLife.26620.001 PMID:28623666

  18. Delineating the regulation of energy homeostasis using hypothalamic cell models.

    Science.gov (United States)

    Wellhauser, Leigh; Gojska, Nicole M; Belsham, Denise D

    2015-01-01

    Attesting to its intimate peripheral connections, hypothalamic neurons integrate nutritional and hormonal cues to effectively manage energy homeostasis according to the overall status of the system. Extensive progress in the identification of essential transcriptional and post-translational mechanisms regulating the controlled expression and actions of hypothalamic neuropeptides has been identified through the use of animal and cell models. This review will introduce the basic techniques of hypothalamic investigation both in vivo and in vitro and will briefly highlight the key advantages and challenges of their use. Further emphasis will be place on the use of immortalized models of hypothalamic neurons for in vitro study of feeding regulation, with a particular focus on cell lines proving themselves most fruitful in deciphering fundamental basics of NPY/AgRP, Proglucagon, and POMC neuropeptide function. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Tumor Response to Radiotherapy Regulated by Endothelial Cell Apoptosis

    Science.gov (United States)

    Garcia-Barros, Monica; Paris, Francois; Cordon-Cardo, Carlos; Lyden, David; Rafii, Shahin; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

    2003-05-01

    About 50% of cancer patients receive radiation therapy. Here we investigated the hypothesis that tumor response to radiation is determined not only by tumor cell phenotype but also by microvascular sensitivity. MCA/129 fibrosarcomas and B16F1 melanomas grown in apoptosis-resistant acid sphingomyelinase (asmase)-deficient or Bax-deficient mice displayed markedly reduced baseline microvascular endothelial apoptosis and grew 200 to 400% faster than tumors on wild-type microvasculature. Thus, endothelial apoptosis is a homeostatic factor regulating angiogenesis-dependent tumor growth. Moreover, these tumors exhibited reduced endothelial apoptosis upon irradiation and, unlike tumors in wild-type mice, they were resistant to single-dose radiation up to 20 grays (Gy). These studies indicate that microvascular damage regulates tumor cell response to radiation at the clinically relevant dose range.

  20. Regulation of nonsmall-cell lung cancer stem cell like cells by neurotransmitters and opioid peptides.

    Science.gov (United States)

    Banerjee, Jheelam; Papu John, Arokya M S; Schuller, Hildegard M

    2015-12-15

    Nonsmall-cell lung cancer (NSCLC) is the leading type of lung cancer and has a poor prognosis. We have shown that chronic stress promoted NSCLC xenografts in mice via stress neurotransmitter-activated cAMP signaling downstream of beta-adrenergic receptors and incidental beta-blocker therapy was reported to improve clinical outcomes in NSCLC patients. These findings suggest that psychological stress promotes NSCLC whereas pharmacologically or psychologically induced decreases in cAMP may inhibit NSCLC. Cancer stem cells are thought to drive the development, progression and resistance to therapy of NSCLC. However, their potential regulation by stress neurotransmitters has not been investigated. In the current study, epinephrine increased the number of cancer stem cell like cells (CSCs) from three NSCLC cell lines in spheroid formation assays while enhancing intracellular cAMP and the stem cell markers sonic hedgehog (SHH), aldehyde dehydrogenase-1 (ALDH-1) and Gli1, effects reversed by GABA or dynorphin B via Gαi -mediated inhibition of cAMP formation. The growth of NSCLC xenografts in a mouse model of stress reduction was significantly reduced as compared with mice maintained under standard conditions. Stress reduction reduced serum levels of corticosterone, norepinephrine and epinephrine while the inhibitory neurotransmitter γ-aminobutyric acid (GABA) and opioid peptides increased. Stress reduction significantly reduced cAMP, VEGF, p-ERK, p-AKT, p-CREB, p-SRc, SHH, ALDH-1 and Gli1 in xenograft tissues whereas cleaved caspase-3 and p53 were induced. We conclude that stress neurotransmitters activate CSCs in NSCLC via multiple cAMP-mediated pathways and that pharmacologically or psychologically induced decreases in cAMP signaling may improve clinical outcomes in NSCLC patients. © 2015 UICC.

  1. Prediction of epigenetically regulated genes in breast cancer cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Loss, Leandro A; Sadanandam, Anguraj; Durinck, Steffen; Nautiyal, Shivani; Flaucher, Diane; Carlton, Victoria EH; Moorhead, Martin; Lu, Yontao; Gray, Joe W; Faham, Malek; Spellman, Paul; Parvin, Bahram

    2010-05-04

    Methylation of CpG islands within the DNA promoter regions is one mechanism that leads to aberrant gene expression in cancer. In particular, the abnormal methylation of CpG islands may silence associated genes. Therefore, using high-throughput microarrays to measure CpG island methylation will lead to better understanding of tumor pathobiology and progression, while revealing potentially new biomarkers. We have examined a recently developed high-throughput technology for measuring genome-wide methylation patterns called mTACL. Here, we propose a computational pipeline for integrating gene expression and CpG island methylation profles to identify epigenetically regulated genes for a panel of 45 breast cancer cell lines, which is widely used in the Integrative Cancer Biology Program (ICBP). The pipeline (i) reduces the dimensionality of the methylation data, (ii) associates the reduced methylation data with gene expression data, and (iii) ranks methylation-expression associations according to their epigenetic regulation. Dimensionality reduction is performed in two steps: (i) methylation sites are grouped across the genome to identify regions of interest, and (ii) methylation profles are clustered within each region. Associations between the clustered methylation and the gene expression data sets generate candidate matches within a fxed neighborhood around each gene. Finally, the methylation-expression associations are ranked through a logistic regression, and their significance is quantified through permutation analysis. Our two-step dimensionality reduction compressed 90% of the original data, reducing 137,688 methylation sites to 14,505 clusters. Methylation-expression associations produced 18,312 correspondences, which were used to further analyze epigenetic regulation. Logistic regression was used to identify 58 genes from these correspondences that showed a statistically signifcant negative correlation between methylation profles and gene expression in the

  2. Brassinosteroid regulates cell elongation by modulating gibberellin metabolism in rice.

    Science.gov (United States)

    Tong, Hongning; Xiao, Yunhua; Liu, Dapu; Gao, Shaopei; Liu, Linchuan; Yin, Yanhai; Jin, Yun; Qian, Qian; Chu, Chengcai

    2014-11-01

    Brassinosteroid (BR) and gibberellin (GA) are two predominant hormones regulating plant cell elongation. A defect in either of these leads to reduced plant growth and dwarfism. However, their relationship remains unknown in rice (Oryza sativa). Here, we demonstrated that BR regulates cell elongation by modulating GA metabolism in rice. Under physiological conditions, BR promotes GA accumulation by regulating the expression of GA metabolic genes to stimulate cell elongation. BR greatly induces the expression of D18/GA3ox-2, one of the GA biosynthetic genes, leading to increased GA1 levels, the bioactive GA in rice seedlings. Consequently, both d18 and loss-of-function GA-signaling mutants have decreased BR sensitivity. When excessive active BR is applied, the hormone mostly induces GA inactivation through upregulation of the GA inactivation gene GA2ox-3 and also represses BR biosynthesis, resulting in decreased hormone levels and growth inhibition. As a feedback mechanism, GA extensively inhibits BR biosynthesis and the BR response. GA treatment decreases the enlarged leaf angles in plants with enhanced BR biosynthesis or signaling. Our results revealed a previously unknown mechanism underlying BR and GA crosstalk depending on tissues and hormone levels, which greatly advances our understanding of hormone actions in crop plants and appears much different from that in Arabidopsis thaliana. © 2014 American Society of Plant Biologists. All rights reserved.

  3. MARCKS-related protein regulates cytoskeletal organization at cell-cell and cell-substrate contacts in epithelial cells.

    Science.gov (United States)

    Van Itallie, Christina M; Tietgens, Amber Jean; Aponte, Angel; Gucek, Marjan; Cartagena-Rivera, Alexander X; Chadwick, Richard S; Anderson, James M

    2018-02-02

    Treatment of epithelial cells with interferon-γ and TNF-α (IFN/TNF) results in increased paracellular permeability. To identify relevant proteins mediating barrier disruption, we performed proximity-dependent biotinylation (BioID) of occludin and found that tagging of MARCKS-related protein (MRP; also known as MARCKSL1) increased ∼20-fold following IFN/TNF administration. GFP-MRP was focused at the lateral cell membrane and its overexpression potentiated the physiological response of the tight junction barrier to cytokines. However, deletion of MRP did not abrogate the cytokine responses, suggesting that MRP is not required in the occludin-dependent IFN/TNF response. Instead, our results reveal a key role for MRP in epithelial cells in control of multiple actin-based structures, likely by regulation of integrin signaling. Changes in focal adhesion organization and basal actin stress fibers in MRP-knockout (KO) cells were reminiscent of those seen in FAK-KO cells. In addition, we found alterations in cell-cell interactions in MRP-KO cells associated with increased junctional tension, suggesting that MRP may play a role in focal adhesion-adherens junction cross talk. Together, our results are consistent with a key role for MRP in cytoskeletal organization of cell contacts in epithelial cells. © 2018. Published by The Company of Biologists Ltd.

  4. Cell cycle-dependent Rho GTPase activity dynamically regulates cancer cell motility and invasion in vivo.

    Science.gov (United States)

    Kagawa, Yoshinori; Matsumoto, Shinji; Kamioka, Yuji; Mimori, Koshi; Naito, Yoko; Ishii, Taeko; Okuzaki, Daisuke; Nishida, Naohiro; Maeda, Sakae; Naito, Atsushi; Kikuta, Junichi; Nishikawa, Keizo; Nishimura, Junichi; Haraguchi, Naotsugu; Takemasa, Ichiro; Mizushima, Tsunekazu; Ikeda, Masataka; Yamamoto, Hirofumi; Sekimoto, Mitsugu; Ishii, Hideshi; Doki, Yuichiro; Matsuda, Michiyuki; Kikuchi, Akira; Mori, Masaki; Ishii, Masaru

    2013-01-01

    The mechanism behind the spatiotemporal control of cancer cell dynamics and its possible association with cell proliferation has not been well established. By exploiting the intravital imaging technique, we found that cancer cell motility and invasive properties were closely associated with the cell cycle. In vivo inoculation of human colon cancer cells bearing fluorescence ubiquitination-based cell cycle indicator (Fucci) demonstrated an unexpected phenomenon: S/G2/M cells were more motile and invasive than G1 cells. Microarray analyses showed that Arhgap11a, an uncharacterized Rho GTPase-activating protein (RhoGAP), was expressed in a cell-cycle-dependent fashion. Expression of ARHGAP11A in cancer cells suppressed RhoA-dependent mechanisms, such as stress fiber formation and focal adhesion, which made the cells more prone to migrate. We also demonstrated that RhoA suppression by ARHGAP11A induced augmentation of relative Rac1 activity, leading to an increase in the invasive properties. RNAi-based inhibition of Arhgap11a reduced the invasion and in vivo expansion of cancers. Additionally, analysis of human specimens showed the significant up-regulation of Arhgap11a in colon cancers, which was correlated with clinical invasion status. The present study suggests that ARHGAP11A, a cell cycle-dependent RhoGAP, is a critical regulator of cancer cell mobility and is thus a promising therapeutic target in invasive cancers.

  5. Cell cycle-dependent Rho GTPase activity dynamically regulates cancer cell motility and invasion in vivo.

    Directory of Open Access Journals (Sweden)

    Yoshinori Kagawa

    Full Text Available The mechanism behind the spatiotemporal control of cancer cell dynamics and its possible association with cell proliferation has not been well established. By exploiting the intravital imaging technique, we found that cancer cell motility and invasive properties were closely associated with the cell cycle. In vivo inoculation of human colon cancer cells bearing fluorescence ubiquitination-based cell cycle indicator (Fucci demonstrated an unexpected phenomenon: S/G2/M cells were more motile and invasive than G1 cells. Microarray analyses showed that Arhgap11a, an uncharacterized Rho GTPase-activating protein (RhoGAP, was expressed in a cell-cycle-dependent fashion. Expression of ARHGAP11A in cancer cells suppressed RhoA-dependent mechanisms, such as stress fiber formation and focal adhesion, which made the cells more prone to migrate. We also demonstrated that RhoA suppression by ARHGAP11A induced augmentation of relative Rac1 activity, leading to an increase in the invasive properties. RNAi-based inhibition of Arhgap11a reduced the invasion and in vivo expansion of cancers. Additionally, analysis of human specimens showed the significant up-regulation of Arhgap11a in colon cancers, which was correlated with clinical invasion status. The present study suggests that ARHGAP11A, a cell cycle-dependent RhoGAP, is a critical regulator of cancer cell mobility and is thus a promising therapeutic target in invasive cancers.

  6. Long Noncoding RNA PANDA Positively Regulates Proliferation of Osteosarcoma Cells.

    Science.gov (United States)

    Kotake, Yojiro; Goto, Taiki; Naemura, Madoka; Inoue, Yasutoshi; Okamoto, Haruna; Tahara, Keiichiro

    2017-01-01

    A long noncoding RNA, p21-associated ncRNA DNA damage-activated (PANDA), associates with nuclear transcription factor Y subunit alpha (NF-YA) and inhibits its binding to promoters of apoptosis-related genes, thereby repressing apoptosis in normal human fibroblasts. Here, we show that PANDA is involved in regulating proliferation in the U2OS human osteosarcoma cell line. U2OS cells were transfected with siRNAs against PANDA 72 h later and they were subjected to reverse transcription-polymerase chain reaction (RT-PCR), quantitative RT-PCR and cell-cycle analysis. PANDA was highly expressed in U2OS cells, and its expression was induced by DNA damage. Silencing PANDA caused arrest at the G 1 phase of the cell cycle, leading to inhibition of cell proliferation. Quantitative RT-PCR showed that silencing PANDA increased mRNA levels of the cyclin-dependent kinase inhibitor p18, which caused G 1 phase arrest. These results suggest that PANDA promotes G 1 -S transition by repressing p18 transcription, and thus promotes U2OS cell proliferation. Copyright© 2017 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  7. Cell Cycle Regulation by Alternative Polyadenylation of CCND1.

    Science.gov (United States)

    Wang, Qiong; He, Guopei; Hou, Mengmeng; Chen, Liutao; Chen, Shangwu; Xu, Anlong; Fu, Yonggui

    2018-05-01

    Global shortening of 3'UTRs by alternative polyadenylation (APA) has been observed in cancer cells. However, the role of APA in cancer remains unknown. CCND1 is a proto-oncogene that regulates progression through the G1-S phase of the cell cycle; moreover, it has been observed to be switching to proximal APA sites in cancer cells. To investigate the biological function of the APA of CCND1, we edited the weak poly(A) signal (PAS) of the proximal APA site to a canonical PAS using the CRISPR/Cas9 method, which can force the cells to use a proximal APA site. Cell cycle profiling and proliferation assays revealed that the proximal APA sites of CCND1 accelerated the cell cycle and promoted cell proliferation, but UTR-APA and CR-APA act via different molecular mechanisms. These results indicate that PAS editing with CRISPR/Cas9 provides a good method by which to study the biological function of APA.

  8. Regulation of T Cell Homeostasis and Responses by Pten

    Directory of Open Access Journals (Sweden)

    Ryan H. Newton

    2012-06-01

    Full Text Available The generation of lipid products catalyzed by PI3K is critical for normal T cell homeostasis and a productive immune response. PI3K can be activated in response to antigen receptor, costimulatory, cytokine and chemokine signals. Moreover, dysregulation of this pathway frequently occurs in T cell lymphomas and is implicated in lymphoproliferative autoimmune disease. Akt acts as a central mediator of PI3K signals, downstream of which is the mTOR pathway, controlling cell growth and metabolism. Members of the Foxo family of transcription factors are also regulated by Akt, thus linking control over homing and migration of T cells, as well cell cycle entry, apoptosis, and DNA damage and oxidative stress responses, to PI3K signaling. PTEN, first identified as a tumor suppressor gene, encodes a lipid phosphatase that, by catalyzing the reverse of the PI3K reaction, directly opposes PI3K signaling. However, PTEN may have other functions as well, and recent reports have suggested roles for PTEN as a tumor suppressor independent of its effects on PI3K signaling. Through the use of models in which Pten is deleted specifically in T cells, it is becoming increasingly clear that control over autoimmunity and lymphomagenesis by PTEN involves multi-faceted functions of this molecule at multiple stages of T cell development.

  9. How Is Mono Spread?

    Science.gov (United States)

    ... How Is Mono Spread? Print My sister has mononucleosis. I drank out of her drink before we ... that I have mono now? – Kyle* Mono, or mononucleosis, is spread through direct contact with saliva. This ...

  10. Volume regulation and shape bifurcation in the cell nucleus.

    Science.gov (United States)

    Kim, Dong-Hwee; Li, Bo; Si, Fangwei; Phillip, Jude M; Wirtz, Denis; Sun, Sean X

    2015-09-15

    Alterations in nuclear morphology are closely associated with essential cell functions, such as cell motility and polarization, and correlate with a wide range of human diseases, including cancer, muscular dystrophy, dilated cardiomyopathy and progeria. However, the mechanics and forces that shape the nucleus are not well understood. Here, we demonstrate that when an adherent cell is detached from its substratum, the nucleus undergoes a large volumetric reduction accompanied by a morphological transition from an almost smooth to a heavily folded surface. We develop a mathematical model that systematically analyzes the evolution of nuclear shape and volume. The analysis suggests that the pressure difference across the nuclear envelope, which is influenced by changes in cell volume and regulated by microtubules and actin filaments, is a major factor determining nuclear morphology. Our results show that physical and chemical properties of the extracellular microenvironment directly influence nuclear morphology and suggest that there is a direct link between the environment and gene regulation. © 2015. Published by The Company of Biologists Ltd.

  11. Chloride channels regulate chondrogenesis in chicken mandibular mesenchymal cells.

    Science.gov (United States)

    Tian, Meiyu; Duan, Yinzhong; Duan, Xiaohong

    2010-12-01

    Voltage gated chloride channels (ClCs) play an important role in the regulation of intracellular pH and cell volume homeostasis. Mutations of these genes result in genetic diseases with abnormal bone deformation and body size, indicating that ClCs may have a role in chondrogenesis. In the present study, we isolated chicken mandibular mesenchymal cells (CMMC) from Hamburg-Hamilton (HH) stage 26 chick embryos and induced chondrocyte maturation by using ascorbic acid and β-glycerophosphate (AA-BGP). We also determined the effect of the chloride channel inhibitor NPPB [5-nitro-2-(3-phenylpropylamino) benzoic acid] on regulation of growth, differentiation, and gene expression in these cells using MTT and real-time PCR assays. We found that CLCN1 and CLCN3-7 mRNA were expressed in CMMC and NPPB reduced expression of CLCN3, CLCN5, and CLCN7 mRNA in these cells. At the same time, NPPB inhibited the growth of the CMMC, but had no effect on the mRNA level of cyclin D1 and cyclin E (P>0.05) with/without AA-BGP treatment. AA-BGP increased markers for early chondrocyte differentiation including type II collagen, aggrecan (Ptype X collagen. NPPB antagonized AA-BGP-induced expression of type II collagen and aggrecan (Ptype X collagen (PType X collagen might function as a target of chloride channel inhibitors during the differentiation process. Copyright © 2010 Elsevier Ltd. All rights reserved.

  12. Epigenetic regulation of open chromatin in pluripotent stem cells

    Science.gov (United States)

    Kobayashi, Hiroshi; Kikyo, Nobuaki

    2014-01-01

    The recent progress in pluripotent stem cell research has opened new avenues of disease modeling, drug screening, and transplantation of patient-specific tissues that had been unimaginable until a decade ago. The central mechanism underlying pluripotency is epigenetic gene regulation; the majority of cell signaling pathways, both extracellular and cytoplasmic, eventually alter the epigenetic status of their target genes during the process of activating or suppressing the genes to acquire or maintain pluripotency. It has long been thought that the chromatin of pluripotent stem cells is globally open to enable the timely activation of essentially all genes in the genome during differentiation into multiple lineages. The current article reviews descriptive observations and the epigenetic machinery relevant to what is supposed to be globally open chromatin in pluripotent stem cells. This includes microscopic appearance, permissive gene transcription, chromatin remodeling complexes, histone modifications, DNA methylation, noncoding RNAs, dynamic movement of chromatin proteins, nucleosome accessibility and positioning, and long-range chromosomal interactions. Detailed analyses of each element, however, have revealed that the globally open chromatin hypothesis is not necessarily supported by some of the critical experimental evidence, such as genome-wide nucleosome accessibility and nucleosome positioning. Further understanding of the epigenetic gene regulation is expected to determine the true nature of the so-called globally open chromatin in pluripotent stem. PMID:24695097

  13. Fetuin-A associates with histones intracellularly and shuttles them to exosomes to promote focal adhesion assembly resulting in rapid adhesion and spreading in breast carcinoma cells.

    Science.gov (United States)

    Nangami, Gladys; Koumangoye, Rainelli; Shawn Goodwin, J; Sakwe, Amos M; Marshall, Dana; Higginbotham, James; Ochieng, Josiah

    2014-11-01

    The present analyses were undertaken to define the mechanisms by which fetuin-A modulates cellular adhesion. FLAG-tagged fetuin-A was expressed in breast carcinoma and HEK-293T cells. We demonstrated by confocal microscopy that fetuin-A co-localizes with histone H2A in the cell nucleus, forms stable complexes with histones such as H2A and H3 in solution, and shuttles histones to exosomes. The rate of cellular adhesion and spreading to either fibronectin or laminin coated wells was accelerated significantly in the presence of either endogenous fetuin-A or serum derived protein. More importantly, the formation of focal adhesion complexes on surfaces coated by laminin or fibronectin was accelerated in the presence of fetuin-A or histone coated exosomes. Cellular adhesion mediated by histone coated exosomes was abrogated by heparin and heparinase III. Heparinase III cleaves heparan sulfate from cell surface heparan sulfate proteoglycans. Lastly, the uptake of histone coated exosomes and subsequent cellular adhesion, was abrogated by heparin. Taken together, the data suggest a mechanism where fetuin-A, either endogenously synthesized or supplied extracellularly can extract histones from the nucleus or elsewhere in the cytosol/membrane and load them on cellular exosomes which then mediate adhesion by interacting with cell surface heparan sulfate proteoglycans via bound histones. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Impact of cycling cells and cell cycle regulation on Hydra regeneration.

    Science.gov (United States)

    Buzgariu, Wanda; Wenger, Yvan; Tcaciuc, Nina; Catunda-Lemos, Ana-Paula; Galliot, Brigitte

    2018-01-15

    Hydra tissues are made from three distinct populations of stem cells that continuously cycle and pause in G2 instead of G1. To characterize the role of cell proliferation after mid-gastric bisection, we have (i) used flow cytometry and classical markers to monitor cell cycle modulations, (ii) quantified the transcriptomic regulations of 202 genes associated with cell proliferation during head and foot regeneration, and (iii) compared the impact of anti-proliferative treatments on regeneration efficiency. We confirm two previously reported events: an early mitotic wave in head-regenerating tips, when few cell cycle genes are up-regulated, and an early-late wave of proliferation on the second day, preceded by the up-regulation of 17 cell cycle genes. These regulations appear more intense after mid-gastric bisection than after decapitation, suggesting a position-dependent regulation of cell proliferation during head regeneration. Hydroxyurea, which blocks S-phase progression, delays head regeneration when applied before but not after bisection. This result is consistent with the fact that the Hydra central region is enriched in G2-paused adult stem cells, poised to divide upon injury, thus forming a necessary constitutive pro-blastema. However a prolonged exposure to hydroxyurea does not block regeneration as cells can differentiate apical structures without traversing S-phase, and also escape in few days the hydroxyurea-induced S-phase blockade. Thus Hydra head regeneration, which is a fast event, is highly plastic, relying on large stocks of adult stem cells paused in G2 at amputation time, which immediately divide to proliferate and/or differentiate apical structures even when S-phase is blocked. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  15. Myosin II dynamics are regulated by tension in intercalating cells.

    Science.gov (United States)

    Fernandez-Gonzalez, Rodrigo; Simoes, Sérgio de Matos; Röper, Jens-Christian; Eaton, Suzanne; Zallen, Jennifer A

    2009-11-01

    Axis elongation in Drosophila occurs through polarized cell rearrangements driven by actomyosin contractility. Myosin II promotes neighbor exchange through the contraction of single cell boundaries, while the contraction of myosin II structures spanning multiple pairs of cells leads to rosette formation. Here we show that multicellular actomyosin cables form at a higher frequency than expected by chance, indicating that cable assembly is an active process. Multicellular cables are sites of increased mechanical tension as measured by laser ablation. Fluorescence recovery after photobleaching experiments show that myosin II is stabilized at the cortex in regions of increased tension. Myosin II is recruited in response to an ectopic force and relieving tension leads to a rapid loss of myosin, indicating that tension is necessary and sufficient for cortical myosin localization. These results demonstrate that myosin II dynamics are regulated by tension in a positive feedback loop that leads to multicellular actomyosin cable formation and efficient tissue elongation.

  16. Wnt/β-catenin signaling regulates cancer stem cells in lung cancer A549 cells

    International Nuclear Information System (INIS)

    Teng, Ying; Wang, Xiuwen; Wang, Yawei; Ma, Daoxin

    2010-01-01

    Wnt/β-catenin signaling plays an important role not only in cancer, but also in cancer stem cells. In this study, we found that β-catenin and OCT-4 was highly expressed in cisplatin (DDP) selected A549 cells. Stimulating A549 cells with lithium chloride (LiCl) resulted in accumulation of β-catenin and up-regulation of a typical Wnt target gene cyclin D1. This stimulation also significantly enhanced proliferation, clone formation, migration and drug resistance abilities in A549 cells. Moreover, the up-regulation of OCT-4, a stem cell marker, was observed through real-time PCR and Western blotting. In a reverse approach, we inhibited Wnt signaling by knocking down the expression of β-catenin using RNA interference technology. This inhibition resulted in down-regulation of the Wnt target gene cyclin D1 as well as the proliferation, clone formation, migration and drug resistance abilities. Meanwhile, the expression of OCT-4 was reduced after the inhibition of Wnt/β-catenin signaling. Taken together, our study provides strong evidence that canonical Wnt signaling plays an important role in lung cancer stem cell properties, and it also regulates OCT-4, a lung cancer stem cell marker.

  17. Autophagy regulates the stemness of cervical cancer stem cells

    Directory of Open Access Journals (Sweden)

    Yang Y

    2017-06-01

    Full Text Available Yi Yang,1,2 Li Yu,1 Jin Li,1 Ya Hong Yuan,1 Xiao Li Wang,1 Shi Rong Yan,1 Dong Sheng Li,1 Yan Ding1 1Hubei Key Laboratory of Embryonic Stem Cell Research, 2Reproductive Center, Taihe Hospital, Hubei University of Medicine, Shiyan, People’s Republic of China Abstract: Cancer stem cells (CSCs are a rare population of multipotent cells with the capacity to self-renew. It has been reported that there are CSCs in cervical cancer cells. Pluripotency-associated (PA transcription factors such as Oct4, Sox2, Nanog and CD44 have been used to isolate CSCs subpopulations. In this study, we showed that autophagy plays an important role in the biological behavior of cervical cancer cells. The expression of the autophagy protein Beclin 1 and LC3B was higher in tumorspheres established from human cervical cancers cell lines (and CaSki than in the parental adherent cells. It was also observed that the basal and starvation-induced autophagy flux was higher in tumorspheres than in the bulk population. Autophagy could regulate the expression level of PA proteins in cervical CSCs. In addition, CRISPR/Cas 9-mediated Beclin 1 knockout enhanced the malignancy of HeLa cells, leading to accumulation of PA proteins and promoted tumorsphere formation. Our findings suggest that autophagy modulates homeostasis of PA proteins, and Beclin 1 is critical for CSC maintenance and tumor development in nude mice. This demonstrates that a prosurvival autophagic pathway is critical for CSC maintenance. Keywords: cervical cancer, autophagy, cancer stem cell, LC3, Oct4

  18. Glucose metabolism regulates T cell activation, differentiation and functions

    Directory of Open Access Journals (Sweden)

    Clovis Steve Palmer

    2015-01-01

    Full Text Available The adaptive immune system is equipped to eliminate both tumors and pathogenic microorganisms. It requires a series of complex and coordinated signals to drive the activation, proliferation and differentiation of appropriate T cell subsets. It is now established that changes in cellular activation are coupled to profound changes in cellular metabolism. In addition, emerging evidence now suggest that specific metabolic alterations associated with distinct T cell subsets may be ancillary to their differentiation and influential in their immune functions. The Warburg effect originally used to describe a phenomenon in which most cancer cells relied on aerobic glycolysis for their growth is a key process that sustain T cell activation and differentiation. Here we review how different aspects of metabolism in T cells influence their functions, focusing on the emerging role of key regulators of glucose metabolism such as HIF-1α. A thorough understanding of the role of metabolism in T cell function could provide insights into mechanisms involved in inflammatory-mediated conditions, with the potential for developing novel therapeutic approaches to treat these diseases.

  19. Angiopoietin-related growth factor (AGF) supports adhesion, spreading, and migration of keratinocytes, fibroblasts, and endothelial cells through interaction with RGD-binding integrins

    International Nuclear Information System (INIS)

    Zhang Yueqing; Hu Xiaobo; Tian Ruiyang; Wei Wangui; Hu Wei; Chen Xia; Han Wei; Chen Huayou; Gong Yi

    2006-01-01

    Angiopoietin-related growth factor (AGF) is a newly identified member of angiopoietin-related proteins (ARPs)/angiopoietin-like proteins (Angptls). AGF has been considered as a novel growth factor in accelerating cutaneous wound healing, as it is capable of stimulating keratinocytes proliferation as well as angiogenesis. But in our paper, we demonstrate that AGF stimulates keratinocytes proliferation only at high protein concentration, however, it can potently promote adhesion, spreading, and migration of keratinocytes, fibroblasts, and endothelial cells. Furthermore, we confirm that the adhesion and migration cellular events are mediated by RGD-binding integrins, most possibly the α v -containing integrins, by in vitro inhibition assays using synthetic competitive peptides. Our results strongly suggest that AGF is an integrin ligand as well as a mitogenic growth factor and theoretically participates in cutaneous wound healing in a more complex mechanism

  20. Improvements to laser wakefield accelerated electron beam stability, divergence, and energy spread using three-dimensional printed two-stage gas cell targets

    International Nuclear Information System (INIS)

    Vargas, M.; Schumaker, W.; He, Z.-H.; Zhao, Z.; Behm, K.; Chvykov, V.; Hou, B.; Krushelnick, K.; Maksimchuk, A.; Yanovsky, V.; Thomas, A. G. R.

    2014-01-01

    High intensity, short pulse lasers can be used to accelerate electrons to ultra-relativistic energies via laser wakefield acceleration (LWFA) [T. Tajima and J. M. Dawson, Phys. Rev. Lett. 43, 267 (1979)]. Recently, it was shown that separating the injection and acceleration processes into two distinct stages could prove beneficial in obtaining stable, high energy electron beams [Gonsalves et al., Nat. Phys. 7, 862 (2011); Liu et al., Phys. Rev. Lett. 107, 035001 (2011); Pollock et al., Phys. Rev. Lett. 107, 045001 (2011)]. Here, we use a stereolithography based 3D printer to produce two-stage gas targets for LWFA experiments on the HERCULES laser system at the University of Michigan. We demonstrate substantial improvements to the divergence, pointing stability, and energy spread of a laser wakefield accelerated electron beam compared with a single-stage gas cell or gas jet target

  1. Improvements to laser wakefield accelerated electron beam stability, divergence, and energy spread using three-dimensional printed two-stage gas cell targets

    Energy Technology Data Exchange (ETDEWEB)

    Vargas, M.; Schumaker, W.; He, Z.-H.; Zhao, Z.; Behm, K.; Chvykov, V.; Hou, B.; Krushelnick, K.; Maksimchuk, A.; Yanovsky, V.; Thomas, A. G. R., E-mail: agrt@umich.edu [Center for Ultrafast Optical Science, University of Michigan, Ann Arbor, Michigan 48109 (United States)

    2014-04-28

    High intensity, short pulse lasers can be used to accelerate electrons to ultra-relativistic energies via laser wakefield acceleration (LWFA) [T. Tajima and J. M. Dawson, Phys. Rev. Lett. 43, 267 (1979)]. Recently, it was shown that separating the injection and acceleration processes into two distinct stages could prove beneficial in obtaining stable, high energy electron beams [Gonsalves et al., Nat. Phys. 7, 862 (2011); Liu et al., Phys. Rev. Lett. 107, 035001 (2011); Pollock et al., Phys. Rev. Lett. 107, 045001 (2011)]. Here, we use a stereolithography based 3D printer to produce two-stage gas targets for LWFA experiments on the HERCULES laser system at the University of Michigan. We demonstrate substantial improvements to the divergence, pointing stability, and energy spread of a laser wakefield accelerated electron beam compared with a single-stage gas cell or gas jet target.

  2. Estrogen regulation of TRPM8 expression in breast cancer cells

    International Nuclear Information System (INIS)

    Chodon, Dechen; Guilbert, Arnaud; Dhennin-Duthille, Isabelle; Gautier, Mathieu; Telliez, Marie-Sophie; Sevestre, Henri; Ouadid-Ahidouch, Halima

    2010-01-01

    The calcium-permeable cation channel TRPM8 (melastatin-related transient receptor potential member 8) is over-expressed in several cancers. The present study aimed at investigating the expression, function and potential regulation of TRPM8 channels by ER alpha (estrogen receptor alpha) in breast cancer. RT-PCR, Western blot, immuno-histochemical, and siRNA techniques were used to investigate TRPM8 expression, its regulation by estrogen receptors, and its expression in breast tissue. To investigate the channel activity in MCF-7 cells, we used the whole cell patch clamp and the calcium imaging techniques. TRPM8 channels are expressed at both mRNA and protein levels in the breast cancer cell line MCF-7. Bath application of the potent TRPM8 agonist Icilin (20 μM) induced a strong outwardly rectifying current at depolarizing potentials, which is associated with an elevation of cytosolic calcium concentration, consistent with established TRPM8 channel properties. RT-PCR experiments revealed a decrease in TRPM8 mRNA expression following steroid deprivation for 48 and 72 hours. In steroid deprived medium, addition of 17-beta-estradiol (E 2 , 10 nM) increased both TRPM8 mRNA expression and the number of cells which respond to Icilin, but failed to affect the Ca 2+ entry amplitude. Moreover, silencing ERα mRNA expression with small interfering RNA reduced the expression of TRPM8. Immuno-histochemical examination of the expression of TRPM8 channels in human breast tissues revealed an over-expression of TRPM8 in breast adenocarcinomas, which is correlated with estrogen receptor positive (ER + ) status of the tumours. Taken together, these results show that TRPM8 channels are expressed and functional in breast cancer and that their expression is regulated by ER alpha

  3. RPS27a promotes proliferation, regulates cell cycle progression and inhibits apoptosis of leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Houcai; Yu, Jing; Zhang, Lixia; Xiong, Yuanyuan; Chen, Shuying; Xing, Haiyan; Tian, Zheng; Tang, Kejing; Wei, Hui; Rao, Qing; Wang, Min; Wang, Jianxiang, E-mail: wangjx@ihcams.ac.cn

    2014-04-18

    Highlights: • RPS27a expression was up-regulated in advanced-phase CML and AL patients. • RPS27a knockdown changed biological property of K562 and K562/G01 cells. • RPS27a knockdown affected Raf/MEK/ERK, P21 and BCL-2 signaling pathways. • RPS27a knockdown may be applicable for new combination therapy in CML patients. - Abstract: Ribosomal protein S27a (RPS27a) could perform extra-ribosomal functions besides imparting a role in ribosome biogenesis and post-translational modifications of proteins. The high expression level of RPS27a was reported in solid tumors, and we found that the expression level of RPS27a was up-regulated in advanced-phase chronic myeloid leukemia (CML) and acute leukemia (AL) patients. In this study, we explored the function of RPS27a in leukemia cells by using CML cell line K562 cells and its imatinib resistant cell line K562/G01 cells. It was observed that the expression level of RPS27a was high in K562 cells and even higher in K562/G01 cells. Further analysis revealed that RPS27a knockdown by shRNA in both K562 and K562G01 cells inhibited the cell viability, induced cell cycle arrest at S and G2/M phases and increased cell apoptosis induced by imatinib. Combination of shRNA with imatinib treatment could lead to more cleaved PARP and cleaved caspase-3 expression in RPS27a knockdown cells. Further, it was found that phospho-ERK(p-ERK) and BCL-2 were down-regulated and P21 up-regulated in RPS27a knockdown cells. In conclusion, RPS27a promotes proliferation, regulates cell cycle progression and inhibits apoptosis of leukemia cells. It appears that drugs targeting RPS27a combining with tyrosine kinase inhibitor (TKI) might represent a novel therapy strategy in TKI resistant CML patients.

  4. Characterization of the myeloid-derived suppressor cell subset regulated by NK cells in malignant lymphoma.

    Science.gov (United States)

    Sato, Yusuke; Shimizu, Kanako; Shinga, Jun; Hidaka, Michihiro; Kawano, Fumio; Kakimi, Kazuhiro; Yamasaki, Satoru; Asakura, Miki; Fujii, Shin-Ichiro

    2015-03-01

    Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population with the ability to suppress immune responses and are currently classified into three distinct MDSC subsets: monocytic, granulocytic and non-monocytic, and non-granulocytic MDSCs. Although NK cells provide an important first-line defense against newly transformed cancer cells, it is unknown whether NK cells can regulate MDSC populations in the context of cancer. In this study, we initially found that the frequency of MDSCs in non-Hodgkin lymphoma (NHL) patients was increased and inversely correlated with that of NK cells, but not that of T cells. To investigate the regulation of MDSC subsets by NK cells, we used an EL4 murine lymphoma model and found the non-monocytic and non-granulocytic MDSC subset, i.e., Gr1 + CD11b + Ly6G med Ly6C med MDSC, is increased after NK cell depletion. The MDSC population that expresses MHC class II, CD80, CD124, and CCR2 is regulated mainly by CD27 + CD11b + NK cells. In addition, this MDSC subset produces some immunosuppressive cytokines, including IL-10 but not nitric oxide (NO) or arginase. We also examined two subsets of MDSCs (CD14 + HLA-DR - and CD14 - HLA-DR - MDSC) in NHL patients and found that higher IL-10-producing CD14 + HLA-DR - MDSC subset can be seen in lymphoma patients with reduced NK cell frequency in peripheral blood. Our analyses of MDSCs in this study may enable a better understanding of how MDSCs manipulate the tumor microenvironment and are regulated by NK cells in patients with lymphoma.

  5. Neuron-NG2 Cell Synapses: Novel Functions for Regulating NG2 Cell Proliferation and Differentiation

    Directory of Open Access Journals (Sweden)

    Qian-Kun Yang

    2013-01-01

    Full Text Available NG2 cells are a population of CNS cells that are distinct from neurons, mature oligodendrocytes, astrocytes, and microglia. These cells can be identified by their NG2 proteoglycan expression. NG2 cells have a highly branched morphology, with abundant processes radiating from the cell body, and express a complex set of voltage-gated channels, AMPA/kainate, and GABA receptors. Neurons notably form classical and nonclassical synapses with NG2 cells, which have varied characteristics and functions. Neuron-NG2 cell synapses could fine-tune NG2 cell activities, including the NG2 cell cycle, differentiation, migration, and myelination, and may be a novel potential therapeutic target for NG2 cell-related diseases, such as hypoxia-ischemia injury and periventricular leukomalacia. Furthermore, neuron-NG2 cell synapses may be correlated with the plasticity of CNS in adulthood with the synaptic contacts passing onto their progenies during proliferation, and synaptic contacts decrease rapidly upon NG2 cell differentiation. In this review, we highlight the characteristics of classical and nonclassical neuron-NG2 cell synapses, the potential functions, and the fate of synaptic contacts during proliferation and differentiation, with the emphasis on the regulation of the NG2 cell cycle by neuron-NG2 cell synapses and their potential underlying mechanisms.

  6. The Chromatin Regulator Brpf1 Regulates Embryo Development and Cell Proliferation*

    Science.gov (United States)

    You, Linya; Yan, Kezhi; Zou, Jinfeng; Zhao, Hong; Bertos, Nicholas R.; Park, Morag; Wang, Edwin; Yang, Xiang-Jiao

    2015-01-01

    With hundreds of chromatin regulators identified in mammals, an emerging issue is how they modulate biological and pathological processes. BRPF1 (bromodomain- and PHD finger-containing protein 1) is a unique chromatin regulator possessing two PHD fingers, one bromodomain and a PWWP domain for recognizing multiple histone modifications. In addition, it binds to the acetyltransferases MOZ, MORF, and HBO1 (also known as KAT6A, KAT6B, and KAT7, respectively) to promote complex formation, restrict substrate specificity, and enhance enzymatic activity. We have recently showed that ablation of the mouse Brpf1 gene causes embryonic lethality at E9.5. Here we present systematic analyses of the mutant animals and demonstrate that the ablation leads to vascular defects in the placenta, yolk sac, and embryo proper, as well as abnormal neural tube closure. At the cellular level, Brpf1 loss inhibits proliferation of embryonic fibroblasts and hematopoietic progenitors. Molecularly, the loss reduces transcription of a ribosomal protein L10 (Rpl10)-like gene and the cell cycle inhibitor p27, and increases expression of the cell-cycle inhibitor p16 and a novel protein homologous to Scp3, a synaptonemal complex protein critical for chromosome association and embryo survival. These results uncover a crucial role of Brpf1 in controlling mouse embryo development and regulating cellular and gene expression programs. PMID:25773539

  7. Regulation of Cell Wall Biogenesis in Saccharomyces cerevisiae: The Cell Wall Integrity Signaling Pathway

    Science.gov (United States)

    Levin, David E.

    2011-01-01

    The yeast cell wall is a strong, but elastic, structure that is essential not only for the maintenance of cell shape and integrity, but also for progression through the cell cycle. During growth and morphogenesis, and in response to environmental challenges, the cell wall is remodeled in a highly regulated and polarized manner, a process that is principally under the control of the cell wall integrity (CWI) signaling pathway. This pathway transmits wall stress signals from the cell surface to the Rho1 GTPase, which mobilizes a physiologic response through a variety of effectors. Activation of CWI signaling regulates the production of various carbohydrate polymers of the cell wall, as well as their polarized delivery to the site of cell wall remodeling. This review article centers on CWI signaling in Saccharomyces cerevisiae through the cell cycle and in response to cell wall stress. The interface of this signaling pathway with other pathways that contribute to the maintenance of cell wall integrity is also discussed. PMID:22174182

  8. FOXO3 regulates CD8 T cell memory by T cell-intrinsic mechanisms.

    Directory of Open Access Journals (Sweden)

    Jeremy A Sullivan

    2012-02-01

    Full Text Available CD8 T cell responses have three phases: expansion, contraction, and memory. Dynamic alterations in proliferation and apoptotic rates control CD8 T cell numbers at each phase, which in turn dictate the magnitude of CD8 T cell memory. Identification of signaling pathways that control CD8 T cell memory is incomplete. The PI3K/Akt signaling pathway controls cell growth in many cell types by modulating the activity of FOXO transcription factors. But the role of FOXOs in regulating CD8 T cell memory remains unknown. We show that phosphorylation of Akt, FOXO and mTOR in CD8 T cells occurs in a dynamic fashion in vivo during an acute viral infection. To elucidate the potentially dynamic role for FOXO3 in regulating homeostasis of activated CD8 T cells in lymphoid and non-lymphoid organs, we infected global and T cell-specific FOXO3-deficient mice with Lymphocytic Choriomeningitis Virus (LCMV. We found that FOXO3 deficiency induced a marked increase in the expansion of effector CD8 T cells, preferentially in the spleen, by T cell-intrinsic mechanisms. Mechanistically, the enhanced accumulation of proliferating CD8 T cells in FOXO3-deficient mice was not attributed to an augmented rate of cell division, but instead was linked to a reduction in cellular apoptosis. These data suggested that FOXO3 might inhibit accumulation of growth factor-deprived proliferating CD8 T cells by reducing their viability. By virtue of greater accumulation of memory precursor effector cells during expansion, the numbers of memory CD8 T cells were strikingly increased in the spleens of both global and T cell-specific FOXO3-deficient mice. The augmented CD8 T cell memory was durable, and FOXO3 deficiency did not perturb any of the qualitative attributes of memory T cells. In summary, we have identified FOXO3 as a critical regulator of CD8 T cell memory, and therapeutic modulation of FOXO3 might enhance vaccine-induced protective immunity against intracellular pathogens.

  9. Angiotensin Converting Enzyme Regulates Cell Proliferation and Migration.

    Directory of Open Access Journals (Sweden)

    Erika Costa de Alvarenga

    Full Text Available The angiotensin-I converting enzyme (ACE plays a central role in the renin-angiotensin system, acting by converting the hormone angiotensin-I to the active peptide angiotensin-II (Ang-II. More recently, ACE was shown to act as a receptor for Ang-II, and its expression level was demonstrated to be higher in melanoma cells compared to their normal counterparts. However, the function that ACE plays as an Ang-II receptor in melanoma cells has not been defined yet.Therefore, our aim was to examine the role of ACE in tumor cell proliferation and migration.We found that upon binding to ACE, Ang-II internalizes with a faster onset compared to the binding of Ang-II to its classical AT1 receptor. We also found that the complex Ang-II/ACE translocates to the nucleus, through a clathrin-mediated process, triggering a transient nuclear Ca2+ signal. In silico studies revealed a possible interaction site between ACE and phospholipase C (PLC, and experimental results in CHO cells, demonstrated that the β3 isoform of PLC is the one involved in the Ca2+ signals induced by Ang-II/ACE interaction. Further studies in melanoma cells (TM-5 showed that Ang-II induced cell proliferation through ACE activation, an event that could be inhibited either by ACE inhibitor (Lisinopril or by the silencing of ACE. In addition, we found that stimulation of ACE by Ang-II caused the melanoma cells to migrate, at least in part due to decreased vinculin expression, a focal adhesion structural protein.ACE activation regulates melanoma cell proliferation and migration.

  10. The Haematopoietic Stem Cell Niche: New Insights into the Mechanisms Regulating Haematopoietic Stem Cell Behaviour

    Directory of Open Access Journals (Sweden)

    Andrew J. Lilly

    2011-01-01

    Full Text Available The concept of the haematopoietic stem cell (HSC niche was formulated by Schofield in the 1970s, as a region within the bone marrow containing functional cell types that can maintain HSC potency throughout life. Since then, ongoing research has identified numerous cell types and a plethora of signals that not only maintain HSCs, but also dictate their behaviour with respect to homeostatic requirements and exogenous stresses. It has been proposed that there are endosteal and vascular niches within the bone marrow, which are thought to regulate different HSC populations. However, recent data depicts a more complicated picture, with functional crosstalk between cells in these two regions. In this review, recent research into the endosteal/vascular cell types and signals regulating HSC behaviour are considered, together with the possibility of a single subcompartmentalised niche.

  11. Regulation of intestinal homeostasis by innate immune cells.

    Science.gov (United States)

    Kayama, Hisako; Nishimura, Junichi; Takeda, Kiyoshi

    2013-12-01

    The intestinal immune system has an ability to distinguish between the microbiota and pathogenic bacteria, and then activate pro-inflammatory pathways against pathogens for host defense while remaining unresponsive to the microbiota and dietary antigens. In the intestine, abnormal activation of innate immunity causes development of several inflammatory disorders such as inflammatory bowel diseases (IBD). Thus, activity of innate immunity is finely regulated in the intestine. To date, multiple innate immune cells have been shown to maintain gut homeostasis by preventing inadequate adaptive immune responses in the murine intestine. Additionally, several innate immune subsets, which promote Th1 and Th17 responses and are implicated in the pathogenesis of IBD, have recently been identified in the human intestinal mucosa. The demonstration of both murine and human intestinal innate immune subsets contributing to regulation of adaptive immunity emphasizes the conserved innate immune functions across species and might promote development of the intestinal innate immunity-based clinical therapy.

  12. Regulation of Innate Lymphoid Cells by Aryl Hydrocarbon Receptor

    Science.gov (United States)

    Li, Shiyang; Bostick, John W.; Zhou, Liang

    2018-01-01

    With striking similarity to their adaptive T helper cell counterparts, innate lymphoid cells (ILCs) represent an emerging family of cell types that express signature transcription factors, including T-bet+ Eomes+ natural killer cells, T-bet+ Eomes− group 1 ILCs, GATA3+ group 2 ILCs, RORγt+ group 3 ILCs, and newly identified Id3+ regulatory ILC. ILCs are abundantly present in barrier tissues of the host (e.g., the lung, gut, and skin) at the interface of host–environment interactions. Active research has been conducted to elucidate molecular mechanisms underlying the development and function of ILCs. The aryl hydrocarbon receptor (Ahr) is a ligand-dependent transcription factor, best known to mediate the effects of xenobiotic environmental toxins and endogenous microbial and dietary metabolites. Here, we review recent progresses regarding Ahr function in ILCs. We focus on the Ahr-mediated cross talk between ILCs and other immune/non-immune cells in host tissues especially in the gut. We discuss the molecular mechanisms of the action of Ahr expression and activity in regulation of ILCs in immunity and inflammation, and the interaction between Ahr and other pathways/transcription factors in ILC development and function with their implication in disease. PMID:29354125

  13. Regulation of Innate Lymphoid Cells by Aryl Hydrocarbon Receptor

    Directory of Open Access Journals (Sweden)

    Shiyang Li

    2018-01-01

    Full Text Available With striking similarity to their adaptive T helper cell counterparts, innate lymphoid cells (ILCs represent an emerging family of cell types that express signature transcription factors, including T-bet+ Eomes+ natural killer cells, T-bet+ Eomes− group 1 ILCs, GATA3+ group 2 ILCs, RORγt+ group 3 ILCs, and newly identified Id3+ regulatory ILC. ILCs are abundantly present in barrier tissues of the host (e.g., the lung, gut, and skin at the interface of host–environment interactions. Active research has been conducted to elucidate molecular mechanisms underlying the development and function of ILCs. The aryl hydrocarbon receptor (Ahr is a ligand-dependent transcription factor, best known to mediate the effects of xenobiotic environmental toxins and endogenous microbial and dietary metabolites. Here, we review recent progresses regarding Ahr function in ILCs. We focus on the Ahr-mediated cross talk between ILCs and other immune/non-immune cells in host tissues especially in the gut. We discuss the molecular mechanisms of the action of Ahr expression and activity in regulation of ILCs in immunity and inflammation, and the interaction between Ahr and other pathways/transcription factors in ILC development and function with their implication in disease.

  14. Regulation of plant cells, cell walls and development by mechanical signals

    Energy Technology Data Exchange (ETDEWEB)

    Meyerowitz, Elliot M. [California Inst. of Technology (CalTech), Pasadena, CA (United States)

    2016-06-14

    The overall goal of the revised scope of work for the final year of funding was to characterize cell wall biosynthesis in developing cotyledons and in the shoot apical meristem of Arabidopsis thaliana, as a way of learning about developmental control of cell wall biosynthesis in plants, and interactions between cell wall biosynthesis and the microtubule cytoskeleton. The proposed work had two parts – to look at the effect of mutation in the SPIRAL2 gene on microtubule organization and reorganization, and to thoroughly characterize the glycosyltransferase genes expressed in shoot apical meristems by RNA-seq experiments, by in situ hybridization of the RNAs expressed in the meristem, and by antibody staining of the products of the glycosyltransferases in meristems. Both parts were completed; the spiral2 mutant was found to speed microtubule reorientation after ablation of adjacent cells, supporting our hypothesis that reorganization correlates with microtubule severing, the rate of which is increased by the mutation. The glycosyltransferase characterization was completed and published as Yang et al. (2016). Among the new things learned was that primary cell wall biosynthesis is strongly controlled both by cell type, and by stage of cell cycle, implying not only that different, even adjacent, cells can have different sugar linkages in their (nonshared) walls, but also that a surprisingly large proportion of glycosyltransferases is regulated in the cell cycle, and therefore that the cell cycle regulates wall maturation to a degree previously unrecognized.

  15. Swelling-activated ion channels: functional regulation in cell-swelling, proliferation and apoptosis

    DEFF Research Database (Denmark)

    Stutzin, A; Hoffmann, E K

    2006-01-01

    Cell volume regulation is one of the most fundamental homeostatic mechanisms and essential for normal cellular function. At the same time, however, many physiological mechanisms are associated with regulatory changes in cell size meaning that the set point for cell volume regulation is under phys...... as key players in the maintenance of normal steady-state cell volume, with particular emphasis on the intracellular signalling pathways responsible for their regulation during hypotonic stress, cell proliferation and apoptosis....

  16. Cell adhesion signaling regulates RANK expression in osteoclast precursors.

    Directory of Open Access Journals (Sweden)

    Ayako Mochizuki

    -adherent condition. These results suggest that cell adhesion signaling regulates RANK expression in osteoclast precursors.

  17. Navigating the transcriptional roadmap regulating plant secondary cell wall deposition

    Directory of Open Access Journals (Sweden)

    Steven Grant Hussey

    2013-08-01

    Full Text Available The current status of lignocellulosic biomass as an invaluable resource in industry, agriculture and health has spurred increased interest in understanding the transcriptional regulation of secondary cell wall (SCW biosynthesis. The last decade of research has revealed an extensive network of NAC, MYB and other families of transcription factors regulating Arabidopsis SCW biosynthesis, and numerous studies have explored SCW-related transcription factors in other dicots and monocots. Whilst the general structure of the Arabidopsis network has been a topic of several reviews, they have not comprehensively represented the detailed protein-DNA and protein-protein interactions described in the literature, and an understanding of network dynamics and functionality has not yet been achieved for SCW formation. Furthermore the methodologies employed in studies of SCW transcriptional regulation have not received much attention, especially in the case of non-model organisms. In this review, we have reconstructed the most exhaustive literature-based network representations to date of SCW transcriptional regulation in Arabidopsis. We include a manipulable Cytoscape representation of the Arabidopsis SCW transcriptional network to aid in future studies, along with a list of supporting literature for each documented interaction. Amongst other topics, we discuss the various components of the network, its evolutionary conservation in plants, putative modules and dynamic mechanisms that may influence network function, and the approaches that have been employed in network inference. Future research should aim to better understand network function and its response to dynamic perturbations, whilst the development and application of genome-wide approaches such as ChIP-seq and systems genetics are in progress for the study of SCW transcriptional regulation in non-model organisms.

  18. Regulating the advertising and promotion of stem cell therapies.

    Science.gov (United States)

    von Tigerstrom, Barbara

    2017-10-01

    There are widespread concerns with the ways in which 'unproven' stem cell therapies are advertised to patients. This article explores the potential and limits of using laws that regulate advertising and promotion as a tool to address these concerns. It examines general consumer protection laws and laws and policies on advertising medical products and services, focusing on the USA, Canada and Australia. The content of existing laws and policies covers most of the marketing practices that cause concern, but several systemic factors are likely to limit enforcement efforts. Potential reforms in Australia that would prevent direct-to-consumer advertising of autologous cell therapies are justified in principle and should be considered by other jurisdictions, but again face important practical limits to their effectiveness.

  19. Apolipoprotein E Regulates Amyloid Formation within Endosomes of Pigment Cells

    Directory of Open Access Journals (Sweden)

    Guillaume van Niel

    2015-10-01

    Full Text Available Accumulation of toxic amyloid oligomers is a key feature in the pathogenesis of amyloid-related diseases. Formation of mature amyloid fibrils is one defense mechanism to neutralize toxic prefibrillar oligomers. This mechanism is notably influenced by apolipoprotein E variants. Cells that produce mature amyloid fibrils to serve physiological functions must exploit specific mechanisms to avoid potential accumulation of toxic species. Pigment cells have tuned their endosomes to maximize the formation of functional amyloid from the protein PMEL. Here, we show that ApoE is associated with intraluminal vesicles (ILV within endosomes and remain associated with ILVs when they are secreted as exosomes. ApoE functions in the ESCRT-independent sorting mechanism of PMEL onto ILVs and regulates the endosomal formation of PMEL amyloid fibrils in vitro and in vivo. This process secures the physiological formation of amyloid fibrils by exploiting ILVs as amyloid nucleating platforms.

  20. Epigenetic regulation of vascular smooth muscle cell function in atherosclerosis.

    Science.gov (United States)

    Findeisen, Hannes M; Kahles, Florian K; Bruemmer, Dennis

    2013-04-01

    Epigenetics involve heritable and acquired changes in gene transcription that occur independently of the DNA sequence. Epigenetic mechanisms constitute a hierarchic upper-level of transcriptional control through complex modifications of chromosomal components and nuclear structures. These modifications include, for example, DNA methylation or post-translational modifications of core histones; they are mediated by various chromatin-modifying enzymes; and ultimately they define the accessibility of a transcriptional complex to its target DNA. Integrating epigenetic mechanisms into the pathophysiologic concept of complex and multifactorial diseases such as atherosclerosis may significantly enhance our understanding of related mechanisms and provide promising therapeutic approaches. Although still in its infancy, intriguing scientific progress has begun to elucidate the role of epigenetic mechanisms in vascular biology, particularly in the control of smooth muscle cell phenotypes. In this review, we will summarize epigenetic pathways in smooth muscle cells, focusing on mechanisms involved in the regulation of vascular remodeling.

  1. Regulation of potassium transport in human lens epithelial cells.

    Science.gov (United States)

    Lauf, Peter K; Warwar, Ronald; Brown, Thomas L; Adragna, Norma C

    2006-01-01

    The major K influx pathways and their response to thiol modification by N-ethylmaleimide (NEM) and protein kinase and phosphatase inhibitors were characterized in human lens epithelial B3 (HLE-B3) cells with Rb as K congener. Ouabain (0.1 mM) and bumetanide (5 microM) discriminated between the Na/K pump ( approximately 35% of total Rb influx) and Na-K-2Cl cotransport (NKCC) ( approximately 50%). Cl-replacement with nitrate or sulfamate revealed 100 microM, activated the Na/K pump and abolished NKCC but did not affect KCC. The data suggest at least partial inverse regulation of KCC and NKCC in HLE-B3 cells by signaling cascades involving serine, threonine and tyrosine phosphorylation/dephosphorylation equilibria.

  2. The Drosophila cell adhesion molecule Neuroglian regulates Lissencephaly-1 localisation in circulating immunosurveillance cells

    Directory of Open Access Journals (Sweden)

    Williams Michael J

    2009-03-01

    Full Text Available Abstract Background When the parasitoid wasp Leptopilina boulardi lays its eggs in Drosophila larvae phagocytic cells called plasmatocytes and specialized cells known as lamellocytes encapsulate the egg. This requires these circulating immunosurveillance cells (haemocytes to change from a non-adhesive to an adhesive state enabling them to bind to the invader. Interestingly, attachment of leukocytes, platelets, and insect haemocytes requires the same adhesion complexes as epithelial and neuronal cells. Results Here evidence is presented showing that the Drosophila L1-type cell adhesion molecule Neuroglian (Nrg is required for haemocytes to encapsulate L. boulardi wasp eggs. The amino acid sequence FIGQY containing a conserved phosphorylated tyrosine is found in the intracellular domain of all L1-type cell adhesion molecules. This conserved tyrosine is phosphorylated at the cell periphery of plasmatocytes and lamellocytes prior to parasitisation, but dephosphorylated after immune activation. Intriguingly, another pool of Nrg located near the nucleus of plasmatocytes remains phosphorylated after parasitisation. In mammalian neuronal cells phosphorylated neurofascin, another L1-type cell adhesion molecule interacts with a nucleokinesis complex containing the microtubule binding protein lissencephaly-1 (Lis1 1. Interestingly in plasmatocytes from Nrg mutants the nucleokinesis regulating protein Lissencephaly-1 (Lis1 fails to localise properly around the nucleus and is instead found diffuse throughout the cytoplasm and at unidentified perinuclear structures. After attaching to the wasp egg control plasmatocytes extend filopodia laterally from their cell periphery; as well as extending lateral filopodia plasmatocytes from Nrg mutants also extend many filopodia from their apical surface. Conclusion The Drosophila cellular adhesion molecule Neuroglian is expressed in haemocytes and its activity is required for the encapsulation of L. boularli eggs. At

  3. [Thiamine and its derivatives in the regulation of cell metabolism].

    Science.gov (United States)

    Tylicki, Adam; Siemieniuk, Magdalena

    2011-07-06

    For over 70 years thiamine (vitamin B1) has aroused the interest of biologists, biochemists and medical doctors because of its multilateral participation in key biochemical and physiological processes. The thiamine molecule is composed of pyrimidine and thiazole rings which are linked by a methylene bridge. It is synthesized by microorganisms, fungi and plants, whereas animals and humans have to obtain it from food. There are several known forms of vitamin B1 inside cells: free thiamine, three phosphate esters (mono-, di-, and triphosphate), and the recently found adenosine thiamine triphosphate. Thiamine has a dual, coenzymatic and non-coenzymatic role. First of all, it is a precursor of thiamin diphosphate, which is a coenzyme for over 20 characterized enzymes which are involved in cell bioenergetic processes leading to the synthesis of ATP. Moreover, these enzymes take part in the biosynthesis of pentose (required for the synthesis of nucleotides), amino acids and other organic compounds of cell metabolism. On the other hand, recent discoveries show the non-coenzymatic role of thiamine derivatives in the process of regulation of gene expression (riboswitches in microorganisms and plants), the stress response, and perhaps so far unknown signal transduction pathways associated with adverse environmental conditions, or transduction of nerve signals with participation of thiamine triphosphate and adenosine thiamine triphosphate. From the clinical point of view thiamine deficiency is related to beri-beri, Parkinson disease, Alzheimer disease, Wernicke-Korsakoff syndrome and other pathologies of the nervous system, and it is successfully applied in medical practice. On the other hand, identifying new synthetic analogues of thiamine which could be used as cytostatics, herbicides or agents preventing deficiency of vitamin B1 is currently the major goal of the research. In this paper we present the current state of knowledge of thiamine and its derivatives, indicating

  4. The cell cycle regulator protein P16 and the cellular senescence of dental follicle cells.

    Science.gov (United States)

    Morsczeck, Christian; Hullmann, Markus; Reck, Anja; Reichert, Torsten E

    2018-02-01

    Cellular senescence is a restricting factor for regenerative therapies with somatic stem cells. We showed previously that the onset of cellular senescence inhibits the osteogenic differentiation in stem cells of the dental follicle (DFCs), although the mechanism remains elusive. Two different pathways are involved in the induction of the cellular senescence, which are driven either by the cell cycle protein P21 or by the cell cycle protein P16. In this study, we investigated the expression of cell cycle proteins in DFCs after the induction of cellular senescence. The induction of cellular senescence was proved by an increased expression of β-galactosidase and an increased population doubling time after a prolonged cell culture. Cellular senescence regulated the expression of cell cycle proteins. The expression of cell cycle protein P16 was up-regulated, which correlates with the induction of cellular senescence markers in DFCs. However, the expression of cyclin-dependent kinases (CDK)2 and 4 and the expression of the cell cycle protein P21 were successively decreased in DFCs. In conclusion, our data suggest that a P16-dependent pathway drives the induction of cellular senescence in DFCs.

  5. Osteokalzinexpression and regulation in hematologic malignancies and in cultured cells

    International Nuclear Information System (INIS)

    Wihlidal, P.

    2010-01-01

    Main issue of this work was to gain further insight into the association of haematopoiesis and osteopoiesis. A crucial cue for that is the fact that haematopoietic stem cells of haematopoietic diseases, which are characterised by c-KIT (CD117) expression, express the osteoblast marker osteocalcin. Thus, attention was focussed on the expression and regulation of osteocalcin, on one hand in blood and bone marrow samples of haematological diseases and on the other hand in leukaemic and osteosarcoma cell lines, i.e., by 1. investigating the expression of osteocalcin (OCN) splicing variants in haematological malignancies. We analysed bone marrow obtained from two patients with chronic myeloid leukaemia (CML), seven patients with other myeloproliferative diseases (MPD) and four patients with acute myeloid leukaemia (AML). RT-PCR analyses were performed in order to assess and quantify spliced (OCNs) and unspliced (OCNu) mRNA, the associated transcription factors (AML1 and AML3) as well as c-KIT, which is a marker for activated stem cells. Our data indicate that OCNs mRNA and OCN protein are expressed in c-KIT positive neoplastic stem cells in haematological malignancies. 2. It has been suggested that the tyrosine kinase inhibitor imatinib mesylate (IM), which has proven anti-proliferative effect, influences osteogenesis and bone turnover in treated patients. Thus, we aimed to quantify OCN mRNA, its splicing variants, the associated Runt-domain transcription factors AML1 and AML3, c-KIT and several metabolic genes to gain evidence about the differentiation state in the HL-60 leukaemia cell line as well as MG63 and U2OS osteosarcoma cells and murine primary osteoblasts MC3T3-E1. Our data indicate that IM induces inhibition of proliferation and synthesis of total OCN-mRNA in all cell lines, but a relative increase of OCNs-mRNA was observed in the human cell lines. On the other hand, differentiation-associated genes appeared to be stimulated. This may also indicate an

  6. Outcomes after surgery and postoperative radiotherapy for perineural spread of head and neck cutaneous squamous cell carcinoma.

    Science.gov (United States)

    Warren, Timothy A; Panizza, Benedict; Porceddu, Sandro V; Gandhi, Mitesh; Patel, Parag; Wood, Martin; Nagle, Christina M; Redmond, Michael

    2016-06-01

    Queensland, Australia, has the highest rates of cutaneous squamous cell carcinoma (SCC). Perineural invasion (PNI) is associated with reduced local control and survival. A retrospective review of a prospective database of patients with clinical PNI from cutaneous SCC of the head and neck (SCCHN) treated with surgery and postoperative radiotherapy (PORT) between 2000 and 2011 and a minimum of 24 months follow-up. Patients were excluded if immunosuppressed, had non-SCC histology, or were treated palliatively. Fifty patients (mean age, 60 years) with median follow-up of 50 months were included in this study. A total of 54.8% of known primary tumors had incidental PNI. Ten percent had nodal disease at presentation. MRI neurogram was positive in 95.8%. Recurrence-free survival (RFS) at 5-years was 62%. Five-year disease-specific survival (DSS) and overall survival (OS) were 75% and 64%, respectively. There were no perioperative deaths. This report demonstrates that long-term survival is achievable in patients with clinical PNI from cutaneous SCCHN after surgery and PORT. © 2015 Wiley Periodicals, Inc. Head Neck 38: 824-831, 2016. © 2015 Wiley Periodicals, Inc.

  7. Fluoxetine regulates cell growth inhibition of interferon-α.

    Science.gov (United States)

    Lin, Yu-Min; Yu, Bu-Chin; Chiu, Wen-Tai; Sun, Hung-Yu; Chien, Yu-Chieh; Su, Hui-Chen; Yen, Shu-Yang; Lai, Hsin-Wen; Bai, Chyi-Huey; Young, Kung-Chia; Tsao, Chiung-Wen

    2016-10-01

    Fluoxetine, a well-known anti-depression agent, may act as a chemosensitizer to assist and promote cancer therapy. However, how fluoxetine regulates cellular signaling to enhance cellular responses against tumor cell growth remains unclear. In the present study, addition of fluoxetine promoted growth inhibition of interferon-alpha (IFN-α) in human bladder carcinoma cells but not in normal uroepithelial cells through lessening the IFN-α-induced apoptosis but switching to cause G1 arrest, and maintaining the IFN-α-mediated reduction in G2/M phase. Activations and signal transducer and transactivator (STAT)-1 and peroxisome proliferator-activated receptor alpha (PPAR-α) were involved in this process. Chemical inhibitions of STAT-1 or PPAR-α partially rescued bladder carcinoma cells from IFN-α-mediated growth inhibition via blockades of G1 arrest, cyclin D1 reduction, p53 downregulation and p27 upregulation in the presence of fluoxetine. However, the functions of both proteins were not involved in the control of fluoxetine over apoptosis and maintained the declined G2/M phase of IFN-α. These results indicated that activation of PPAR-α and STAT-1 participated, at least in part, in growth inhibition of IFN-α in the presence of fluoxetine.

  8. PGC-1α regulates alanine metabolism in muscle cells.

    Science.gov (United States)

    Hatazawa, Yukino; Qian, Kun; Gong, Da-Wei; Kamei, Yasutomi

    2018-01-01

    The skeletal muscle is the largest organ in the human body, depositing energy as protein/amino acids, which are degraded in catabolic conditions such as fasting. Alanine is synthesized and secreted from the skeletal muscle that is used as substrates of gluconeogenesis in the liver. During fasting, the expression of PGC-1α, a transcriptional coactivator of nuclear receptors, is increased in the liver and regulates gluconeogenesis. In the present study, we observed increased mRNA expression of PGC-1α and alanine aminotransferase 2 (ALT2) in the skeletal muscle during fasting. In C2C12 myoblast cells overexpressing PGC-1α, ALT2 expression was increased concomitant with an increased alanine level in the cells and medium. In addition, PGC-1α, along with nuclear receptor ERR, dose-dependently enhanced the ALT2 promoter activity in reporter assay using C2C12 cells. In the absence of glucose in the culture medium, mRNA levels of PGC-1α and ALT2 increased. Endogenous PGC-1α knockdown in C2C12 cells reduced ALT2 gene expression level, induced by the no-glucose medium. Taken together, in the skeletal muscle, PGC-1α activates ALT2 gene expression, and alanine production may play roles in adaptation to fasting.

  9. Regulation of stem cell factor expression in inflammation and asthma

    Directory of Open Access Journals (Sweden)

    Carla A Da Silva

    2005-03-01

    Full Text Available Stem cell factor (SCF is a major mast cell growth factor, which could be involved in the local increase of mast cell number in the asthmatic airways. In vivo, SCF expression increases in asthmatic patients and this is reversed after treatment with glucocorticoids. In vitro in human lung fibroblasts in culture, IL-1beta, a pro-inflammatory cytokine, confirms this increased SCF mRNA and protein expression implying the MAP kinases p38 and ERK1/2 very early post-treatment, and glucocorticoids confirm this decrease. Surprisingly, glucocorticoids potentiate the IL-1beta-enhanced SCF expression at short term treatment, implying increased SCF mRNA stability and SCF gene transcription rate. This potentiation involves p38 and ERK1/2. Transfection experiments with the SCF promoter including intron1 also confirm this increase and decrease of SCF expression by IL-1beta and glucocorticoids, and the potentiation by glucocorticoids of the IL-1beta-induced SCF expression. Deletion of the GRE or kappaB sites abolishes this potentiation, and the effect of IL-1beta or glucocorticoids alone. DNA binding of GR and NF-kappaB are also demonstrated for these effects. In conclusion, this review concerns new mechanisms of regulation of SCF expression in inflammation that could lead to potential therapeutic strategy allowing to control mast cell number in the asthmatic airways.

  10. Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines.

    Science.gov (United States)

    Emadi Baygi, Modjtaba; Soheili, Zahra Soheila; Schmitz, Ingo; Sameie, Shahram; Schulz, Wolfgang A

    2010-12-01

    The epithelial-mesenchymal transition (EMT) is regarded as an important step in cancer metastasis. Snail, a master regulator of EMT, has been recently proposed to act additionally as a cell survival factor and inducer of motility. We have investigated the function of Snail (SNAI1) in prostate cancer cells by downregulating its expression via short (21-mer) interfering RNA (siRNA) and measuring the consequences on EMT markers, cell viability, death, cell cycle, senescence, attachment, and invasivity. Of eight carcinoma cell lines, the prostate carcinoma cell lines LNCaP and PC-3 showed the highest and moderate expression of SNAI1 mRNA, respectively, as measured by quantitative RT-PCR. Long-term knockdown of Snail induced a severe decline in cell numbers in LNCaP and PC-3 and caspase activity was accordingly enhanced in both cell lines. In addition, suppression of Snail expression induced senescence in LNCaP cells. SNAI1-siRNA-treated cells did not tolerate detachment from the extracellular matrix, probably due to downregulation of integrin α6. Expression of E-cadherin, vimentin, and fibronectin was also affected. Invasiveness of PC-3 cells was not significantly diminished by Snail knockdown. Our data suggest that Snail acts primarily as a survival factor and inhibitor of cellular senescence in prostate cancer cell lines. We therefore propose that Snail can act as early driver of prostate cancer progression.

  11. Modeling microenvironmental regulation of glioblastoma stem cells: a biomaterials perspective

    Science.gov (United States)

    Heffernan, John M.; Sirianni, Rachael W.

    2018-02-01

    Following diagnosis of a glioblastoma (GBM) brain tumor, surgical resection, chemotherapy and radiation together yield a median patient survival of only 15 months. Importantly, standard treatments fail to address the dynamic regulation of the brain tumor microenvironment that actively supports tumor progression and treatment resistance. It is becoming increasingly recognized that specialized niches within the tumor microenvironment maintain a population of highly malignant glioblastoma stem-like cells (GSCs). GSCs are resistant to traditional chemotherapy and radiation therapy, suggesting that they may be responsible for the near universal rates of tumor recurrence and associated morbidity in GBM. Thus, disrupting microenvironmental support for GSCs could be critical to developing more effective GBM therapies. Three-dimensional (3D) culture models of the tumor microenvironment are powerful tools for identifying key biochemical and biophysical inputs that impact malignant behaviors. Such systems have been used effectively to identify conditions that regulate GSC proliferation, invasion, stem-specific phenotypes, and treatment resistance. Considering the significant role that GSC microenvironments play in regulating this tumorigenic sub-population, these models may be essential for uncovering mechanisms that limit GSCs malignancy.

  12. Regulation of human renin expression in chorion cell primary cultures

    International Nuclear Information System (INIS)

    Duncan, K.G.; Haidar, M.A.; Baxter, J.D.; Reudelhuber, T.L.

    1990-01-01

    The human renin gene is expressed in the kidney, placenta, and several other sites. The release of renin or its precursor, prorenin, can be affected by several regulatory agents. In this study, primary cultures of human placental cells were used to examine the regulation of prorenin release and renin mRNA levels and of the transfected human renin promoter linked to chloramphenicol acetyltransferase reporter sequences. Treatment of the cultures with a calcium ionophore alone, calcium ionophore plus forskolin (that activates adenylate cyclase), or forskolin plus a phorbol ester increased prorenin release and renin mRNA levels 1.3 endash to 6 endash fold, but several classes of steroids did not affect prorenin secretion or renin RNA levels. These results suggest that (i) the first 584 base pairs of the renin gene 5'endash flanking DNA do not contain functional glucocorticoid or estrogen response elements, (ii) placental prorenin release and renin mRNA are regulated by calcium ion and by the combinations of cAMP with either C kinase or calcium ion, and (iii) the first 100 base pairs of the human renin 5'endash flanking DNA direct accurate initiation of transcription and can be regulated by cAMP. Thus, some control of renin release in the placenta (and by inference in other tissues) occurs via transcriptional influences on its promoter

  13. Dendritic cells regulate angiogenesis associated with liver fibrogenesis.

    Science.gov (United States)

    Blois, Sandra M; Piccioni, Flavia; Freitag, Nancy; Tirado-González, Irene; Moschansky, Petra; Lloyd, Rodrigo; Hensel-Wiegel, Karin; Rose, Matthias; Garcia, Mariana G; Alaniz, Laura D; Mazzolini, Guillermo

    2014-01-01

    During liver fibrogenesis the immune response and angiogenesis process are fine-tuned resulting in activation of hepatic stellate cells that produce an excess of extracellular matrix proteins. Dendritic cells (DC) play a central role modulating the liver immunity and have recently been implicated to favour fibrosis regression; although their ability to influence the development of fibrogenesis is unknown. Therefore, we explored whether the depletion of DC during early stages of liver injury has an impact in the development of fibrogenesis. Using the CD11c.DTR transgenic mice, DC were depleted in two experimental models of fibrosis in vivo. The effect of anti-angiogenic therapy was tested during early stages of liver fibrogenesis. DC depletion accelerates the development of fibrosis and as a consequence, the angiogenesis process is boosted. We observed up-regulation of pro-angiogenic factors together with an enhanced vascular endothelial growth factor (VEGF) bioavailability, mainly evidenced by the decrease of anti-angiogenic VEGF receptor 1 (also known as sFlt-1) levels. Interestingly, fibrogenesis process enhanced the expression of Flt-1 on hepatic DC and administration of sFlt-1 was sufficient to abrogate the acceleration of fibrogenesis upon DC depletion. Thus, DC emerge as novel players during the development of liver fibrosis regulating the angiogenesis process and thereby influencing fibrogenesis.

  14. Cyclic guanosine monophosphate in the regulation of the cell function

    Directory of Open Access Journals (Sweden)

    Małgorzata Zbrojkiewicz

    2016-12-01

    Full Text Available Intracellular concentration of cGMP depends on the activity of guanylate cyclase, responsible for its synthesis, on the activity of cyclic nucleotide degrading enzymes - phosphodiesterases (PDEs. There are two forms of guanylate cyclase: the membrane-bound cyclase and the soluble form. The physiological activators of the membrane guanylate cyclase are natriuretic peptides (NPs, and of the cytosolic guanylate cyclase - nitric oxide (NO and carbon monoxide (CO. Intracellular cGMP signaling pathways arise from its direct effect on the activity of G protein kinases, phosphodiesterases and cyclic nucleotide dependent cation channels. It has been shown in recent years that cGMP can also affect other signal pathways in cell signaling activity involving Wnt proteins and sex hormones. The increased interest in the research on the role of cGMP, resulted also in the discovery of its role in the regulation of phototransduction in the eye, neurotransmission, calcium homeostasis, platelet aggregation, heartbeat, bone remodeling, lipid metabolism and the activity of the cation channels. Better understanding of the mechanisms of action of cGMP in the regulation of cell function can create new opportunities for the cGMP affecting drugs use in the pharmacotherapy.

  15. Wnt/β-Catenin Signaling Regulates the Expression of the Ammonium Permease Gene RHBG in Human Cancer Cells.

    Directory of Open Access Journals (Sweden)

    Ahmad Merhi

    Full Text Available Ammonium is a metabolic waste product mainly detoxified by the liver. Hepatic dysfunction can lead to cytotoxic accumulation of circulating ammonium and to subsequent encephalopathy. Transmembrane ammonium transport is a widely spread process ensured by the highly conserved proteins of the Mep-Amt-Rh superfamily, including the mammalian Rhesus (Rh factors. The regulatory mechanisms involved in the control of RH genes expression remain poorly studied. Here we addressed the expression regulation of one of these factors, RHBG. We identify HepG2 hepatocellular carcinoma cells and SW480 colon adenocarcinoma cells as expressing RHBG and show that its expression relies on β-catenin signaling. siRNA-mediated β-catenin knockdown resulted in significant reduction of RHBG mRNA in both cell lines. Pharmaceutical inhibition of the TCF4/β-catenin interaction or knockdown of the transcription factor TCF4 also downregulated RHBG expression. We identify a minimal RHBG regulatory sequence displaying a promoter activity and show that β-catenin and TCF4 bind to this fragment in vivo. We finally characterize the role of potential TCF4 binding sites in RHBG regulation. Taken together, our results indicate RHBG expression as a direct target of β-catenin regulation, a pathway frequently deregulated in many cancers and associated with tumorigenesis.

  16. Regulation of TFIIIB during F9 cell differentiation.

    Science.gov (United States)

    Athineos, Dimitris; Marshall, Lynne; White, Robert J

    2010-03-12

    Differentiation of F9 embryonal carcinoma (EC) cells into parietal endoderm (PE) provides a tractable model system for studying molecular events during early and inaccessible stages of murine development. PE formation is accompanied by extensive changes in gene expression both in vivo and in culture. One of the most dramatic is the ~10-fold decrease in transcriptional output by RNA polymerase (pol) III. This has been attributed to changes in activity of TFIIIB, a factor that is necessary and sufficient to recruit pol III to promoters. The goal of this study was to identify molecular changes that can account for the low activity of TFIIIB following F9 cell differentiation. Three essential subunits of TFIIIB decrease in abundance as F9 cells differentiate; these are Brf1 and Bdp1, which are pol III-specific, and TBP, which is also used by pols I and II. The decreased levels of Brf1 and Bdp1 proteins can be explained by reduced expression of the corresponding mRNAs. However, this is not the case for TBP, which is regulated post-transcriptionally. In proliferating cells, pol III transcription is stimulated by the proto-oncogene product c-Myc and the mitogen-activated protein kinase Erk, both of which bind to TFIIIB. However, c-Myc levels fall during differentiation and Erk becomes inactive through dephosphorylation. The diminished abundance of TFIIIB is therefore likely to be compounded by changes to these positive regulators that are required for its full activity. In addition, PE cells have elevated levels of the retinoblastoma protein RB, which is known to bind and repress TFIIIB. The low activity of TFIIIB in PE can be attributed to a combination of changes, any one of which could be sufficient to inhibit pol III transcription. Declining levels of essential TFIIIB subunits and of activators that are required for maximal TFIIIB activity are accompanied by an increase in a potent repressor of TFIIIB. These events provide fail-safe guarantees to ensure that pol III

  17. Regulation of stem-cell mediated host immunity by the sphingolipid ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    Regulation of stem-cell mediated host immunity by the sphingolipid pathway ... in the generation of mature immune cells and the functioning of the surrounding ... methods with human cells and genetically engineered mice to examine how the ...

  18. Cyclooxygenase-2 Regulates Th17 Cell Differentiation during Allergic Lung Inflammation

    OpenAIRE

    Li, Hong; Bradbury, J. Alyce; Dackor, Ryan T.; Edin, Matthew L.; Graves, Joan P.; DeGraff, Laura M.; Wang, Ping Ming; Bortner, Carl D.; Maruoka, Shuichiro; Lih, Fred B.; Cook, Donald N.; Tomer, Kenneth B.; Jetten, Anton M.; Zeldin, Darryl C.

    2011-01-01

    Rationale: Th17 cells comprise a distinct lineage of proinflammatory T helper cells that are major contributors to allergic responses. It is unknown whether cyclooxygenase (COX)-derived eicosanoids regulate Th17 cells during allergic lung inflammation.

  19. Fas/Fas ligand regulation mediates cell death in human Ewing's sarcoma cells treated with melatonin

    Science.gov (United States)

    García-Santos, G; Martin, V; Rodríguez-Blanco, J; Herrera, F; Casado-Zapico, S; Sánchez-Sánchez, A M; Antolín, I; Rodríguez, C

    2012-01-01

    Background: Despite recent advances in cancer therapy, the 5-year survival rate for Ewing's sarcoma is still very low, and new therapeutic approaches are necessary. It was found previously that melatonin induces cell death in the Ewing's sarcoma cell line, SK-N-MC, by activating the extrinsic apoptotic pathway. Methods: Melatonin actions were analysed by metabolic viability/survival cell assays, flow cytometry, quantitative PCR for mRNA expression, western blot for protein activation/expression and electrophoretic mobility shift assay for transcription factor activation. Results: Melatonin increases the expression of Fas and its ligand Fas L, this increase being responsible for cell death induced by the indolamine. Melatonin also produces a transient increase in intracellular oxidants and activation of the redox-regulated transcription factor Nuclear factor-kappaB. Inhibition of such activation prevents cell death and Fas/Fas L upregulation. Cytotoxic effect and Fas/Fas L regulation occur in all Ewing's cell lines studied, and do not occur in the other tumour cell lines studied where melatonin does not induce cell death. Conclusion: Our data offers new insights in the study of alternative therapeutic strategies in the treatment of Ewing's sarcoma. Further attention deserves to be given to the differences in the cellular biology of sensitive tumours that could explain the cytotoxic effect of melatonin and the increase in the level of free radicals caused by this molecule, in particular cancer types. PMID:22382690

  20. Cell-cycle-dependent regulation of cell motility and determination of the role of Rac1

    DEFF Research Database (Denmark)

    Walmod, Peter S.; Hartmann-Petersen, Rasmus; Prag, S.

    2004-01-01

    comparable to those of control cells in G1. In contrast, transfection with dominant-negative Rac1 reduced cell speed and resulted in cellular displacements, which were identical in G1 and G2. These observations indicate that migration of cultured cells is regulated in a cell-cycle-dependent manner...... for calculation of three key parameters describing cell motility: speed, persistence time and rate of diffusion. All investigated cell lines demonstrated a lower cell displacement in the G2 phase than in the G1/S phases. This was caused by a decrease in speed and/or persistence time. The decrease in motility...... was accompanied by changes in morphology reflecting the larger volume of cells in G2 than in G1. Furthermore, L-cells and HeLa-cells appeared to be less adherent in the G2 phase. Transfection of L-cells with constitutively active Rac1 led to a general increase in the speed and rate of diffusion in G2 to levels...

  1. Fps/Fes protein-tyrosine kinase regulates mast cell adhesion and migration downstream of Kit and beta1 integrin receptors.

    Science.gov (United States)

    Smith, Julie A; Samayawardhena, Lionel A; Craig, Andrew W B

    2010-03-01

    Activation of Kit receptor protein-tyrosine kinase (PTK) by its ligand Stem Cell Factor (SCF) is required for the development of mast cells, and for the regulation of mast cell proliferation, migration and modulation of inflammatory mediator release. Recent studies have implicated the non-receptor PTK Fps/Fes (hereafter referred to as Fes) in signaling downstream of oncogenic Kit, however, the potential role of Fes in regulating Kit signaling is not well defined. In this study, we show that SCF induces transient tyrosine phosphorylation of wild-type Fes as well as kinase-dead Fes in bone marrow-derived mast cells (BMMCs). The latter finding implicates an upstream kinase acting on Fes, which we identified as Fyn PTK. SCF treatment of BMMCs promoted recruitment of Fes to Kit, potentially via direct interaction of the Fes SH2 domain with phosphorylated Kit. While Fes was not required for SCF-induced signaling to Akt and Erk kinases, Fes-deficient (fes-/-) BMMCs displayed a defect in sustained p38 kinase activation, compared to control cells. SCF-treated Fes-deficient BMMCs also displayed elevated beta1 integrin-mediated cell adhesion and spreading on fibronectin, compared to control cells, and a reduction in cell polarization at later times of SCF treatment. Restoring Fes expression in fes-/- BMMCs by retroviral transduction was sufficient to rescue cell spreading and polarization defects. Interestingly, SCF-induced chemotaxis of BMMCs was also defective in Fes-deficient BMMCs, and restored in Fes-rescue BMMCs. Overall, these results implicate Fes in regulating cross-talk between Kit and beta1 integrins to promote cytoskeletal reorganization and motility of mast cells.

  2. Aebp2 as an epigenetic regulator for neural crest cells.

    Directory of Open Access Journals (Sweden)

    Hana Kim

    Full Text Available Aebp2 is a potential targeting protein for the mammalian Polycomb Repression Complex 2 (PRC2. We generated a mutant mouse line disrupting the transcription of Aebp2 to investigate its in vivo roles. Aebp2-mutant homozygotes were embryonic lethal while heterozygotes survived to adulthood with fertility. In developing mouse embryos, Aebp2 is expressed mainly within cells of neural crest origin. In addition, many heterozygotes display a set of phenotypes, enlarged colon and hypopigmentation, similar to those observed in human patients with Hirschsprung's disease and Waardenburg syndrome. These phenotypes are usually caused by the absence of the neural crest-derived ganglia in hindguts and melanocytes. ChIP analyses demonstrated that the majority of the genes involved in the migration and development process of neural crest cells are downstream target genes of AEBP2 and PRC2. Furthermore, expression analyses confirmed that some of these genes are indeed affected in the Aebp2 heterozygotes. Taken together, these results suggest that Aebp2 may regulate the migration and development of the neural crest cells through the PRC2-mediated epigenetic mechanism.

  3. Distribution of a 69-kD laminin-binding protein in aortic and microvascular endothelial cells: modulation during cell attachment, spreading, and migration

    DEFF Research Database (Denmark)

    Yannariello-Brown, J; Wewer, U; Liotta, L

    1988-01-01

    cells identified this protein in BAEC lysates. In frozen sections, these polyclonal antibodies and monoclonal antibodies raised against human tumor 69-kD stained the endothelium of bovine aorta and the medial smooth muscle cells, but not surrounding connective tissue or elastin fibers. When...... nonpermeabilized BAEC were stained in an in vitro migration assay, there appeared to be apical patches of 69 kD staining in stationary cells. However, when released from contact inhibition, 69 kD was localized to ruffling membranes on cells at the migrating front. Permeabilized BAEC stained for 69 kD diffusely...... in permeabilized cultured microvascular endothelial cells in a continuous staining pattern at 6 h postplating which redistributed to punctate patches along the length of the filaments at confluence (96 h). In addition, 69 kD co-distribution with laminin could also be demonstrated in cultured subconfluent cells...

  4. Nuclear receptor TLX regulates cell cycle progression in neural stem cells of the developing brain.

    Science.gov (United States)

    Li, Wenwu; Sun, Guoqiang; Yang, Su; Qu, Qiuhao; Nakashima, Kinichi; Shi, Yanhong

    2008-01-01

    TLX is an orphan nuclear receptor that is expressed exclusively in vertebrate forebrains. Although TLX is known to be expressed in embryonic brains, the mechanism by which it influences neural development remains largely unknown. We show here that TLX is expressed specifically in periventricular neural stem cells in embryonic brains. Significant thinning of neocortex was observed in embryonic d 14.5 TLX-null brains with reduced nestin labeling and decreased cell proliferation in the germinal zone. Cell cycle analysis revealed both prolonged cell cycles and increased cell cycle exit in TLX-null embryonic brains. Increased expression of a cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin D1 provide a molecular basis for the deficiency of cell cycle progression in embryonic brains of TLX-null mice. Furthermore, transient knockdown of TLX by in utero electroporation led to precocious cell cycle exit and differentiation of neural stem cells followed by outward migration. Together these results indicate that TLX plays an important role in neural development by regulating cell cycle progression and exit of neural stem cells in the developing brain.

  5. Mechanical Regulation in Cell Division and in Neurotransmitter Release

    Science.gov (United States)

    Thiyagarajan, Sathish

    During their lifecycle, cells must produce forces which play important roles in several subcellular processes. Force-producing components are organized into macromolecular assemblies of proteins that are often dynamic, and are constructed or disassembled in response to various signals. The forces themselves may directly be involved in subcellular mechanics, or they may influence mechanosensing proteins either within or outside these structures. These proteins play different roles: they may ensure the stability of the force-producing structure, or they may send signals to a coupled process. The generation and sensing of subcellular forces is an active research topic, and this thesis focusses on the roles of these forces in two key areas: cell division and neurotransmitter release. The first part of the thesis deals with the effect of force on cell wall growth regulation during division in the fission yeast Schizosaccharomyces pombe, a cigar-shaped, unicellular organism. During cytokinesis, the last stage of cell division in which the cell physically divides into two, a tense cytokinetic ring anchored to the cellular membrane assembles and constricts, accompanied by the inward centripetal growth of new cell wall, called septum, in the wake of the inward-moving membrane. The contour of the septum hole maintains its circularity as it reduces in size--an indication of regulated growth. To characterize the cell wall growth process, we performed image analysis on contours of the leading edge of the septum obtained via fluorescence microscopy in the labs of our collaborators. We quantified the deviations from circularity using the edge roughness. The roughness was spatially correlated, suggestive of regulated growth. We hypothesized that the cell wall growers are mechanosensitive and respond to the force exerted by the ring. A mathematical model based on this hypothesis then showed that this leads to corrections of roughness in a curvature-dependent fashion. Thus, one of

  6. Regulation of Taurine transporter activity in cultured rat retinal ganglion cells and rat retinal Muller Cells

    International Nuclear Information System (INIS)

    Eissa, Laila A.; Smith, Sylvia B.; El-sherbeny, Amira A.

    2006-01-01

    Diabetic retinopathy is one of the most common complications of diabetes. The amino acid taurine is believed to play an antioxidant protective role in diabetic retinopathy through the scavenging of the reactive species. It is not well established whether taurine uptake is altered in retina cells during diabetic conditions. Thus, the present study was designed to investigate the changes in taurine transport in cultures of rat retinal Muller cells and rat retinal ganglion cells under conditions associated with diabetes. Taurine was abundantly taken up by retinal Muller cells and rat retinal ganglion cells under normal glycemic condition. Taurine was actively transported to rat Muller cells and rat retinal ganglion cells in a Na and Cl dependant manner. Taurine uptake further significantly elevated in both type of cells after the incubation with high glucose concentration. This effect could be attributed to the increase in osmolarity. Because Nitric Oxide (NO) is a molecule implicated in the pathogenesis of diabetes, we also determined the activity of taurine transporter in cultured rat retinal Muller cells and rat retinal ganglion cells in the presence of the NO donors, SIN-1 and SNAP. Taurine uptake was elevated above control value after 24-h incubation with low concentration of NO donors. We finally investigated the ability of neurotoxic glutamate to change taurine transporter activity in both types of cells. Uptake of taurine was significantly increased in rat retinal ganglion cells when only incubated with high concentration of glutamate. Our data provide evidence that taurine transporter is present in cultured rat retinal ganglion and Muller cells and is regulated by hyperosmolarity. The data are relevant to disease such as diabetes and neuronal degeneration where retinal cell volume may dramatically change. (author)

  7. Co-ordinate regulation of the cytoskeleton in 3T3 cells overexpressing thymosin-beta4.

    Science.gov (United States)

    Golla, R; Philp, N; Safer, D; Chintapalli, J; Hoffman, R; Collins, L; Nachmias, V T

    1997-01-01

    In several cell types, short-term increases in the concentration of the G-actin-sequestering peptide thymosin-beta4 (Tbeta4) cause the disassembly of F-actin bundles. To determine the extent of cell adaptability to these reductions in F-actin, we overexpressed Tbeta4 in NIH 3T3 cells. In cell lines with Tbeta4 levels twice those of vector controls, G-actin increased approximately twofold as expected. However, F-actin did not decrease as in short-term experiments but rather also increased approximately twofold so that the G-F ratio remained constant. Surprisingly, the cytoskeletal proteins myosin IIA, alpha-actinin, and tropomyosin also increased nearly twofold. These increases were specific; DNA, total protein, lactic dehydrogenase, profilin, and actin depolymerizing factor levels were unchanged in the overexpressing cells. The Tbeta4 lines spread more fully and adhered to the dish more strongly than vector controls; this altered phenotype correlated with a twofold increase in talin and alpha5-integrin and a nearly threefold increase in vinculin. Focal adhesions, detected by indirect immunofluorescence with antivinculin, were increased in size over the controls. Northern blotting showed that mRNAs for both beta-actin and vinculin were increased twofold in the overexpressing lines. We conclude that 1) NIH 3T3 cells adapt to increased levels of G-actin sequestered by increased Tbeta4 by increasing their total actin so that the F-actin/G-actin ratio remains constant; 2) these cells coordinately increase several cytoskeletal and adhesion plaque proteins; and 3) at least for actin and vinculin, this regulation is at the transcriptional level. We therefore propose that the proteins of this multimember interacting complex making up the actin-based cytoskeleton, are coordinately regulated by factors that control the expression of several proteins. The mechanism may bear similarities to the control of synthesis of another multimember interacting complex, the myofibril of

  8. Regulation of adult neural progenitor cell functions by purinergic signaling.

    Science.gov (United States)

    Tang, Yong; Illes, Peter

    2017-02-01

    Extracellular purines are signaling molecules in the neurogenic niches of the brain and spinal cord, where they activate cell surface purinoceptors at embryonic neural stem cells (NSCs) and adult neural progenitor cells (NPCs). Although mRNA and protein are expressed at NSCs/NPCs for almost all subtypes of the nucleotide-sensitive P2X/P2Y, and the nucleoside-sensitive adenosine receptors, only a few of those have acquired functional significance. ATP is sequentially degraded by ecto-nucleotidases to ADP, AMP, and adenosine with agonistic properties for distinct receptor-classes. Nucleotides/nucleosides facilitate or inhibit NSC/NPC proliferation, migration and differentiation. The most ubiquitous effect of all agonists (especially of ATP and ADP) appears to be the facilitation of cell proliferation, usually through P2Y1Rs and sometimes through P2X7Rs. However, usually P2X7R activation causes necrosis/apoptosis of NPCs. Differentiation can be initiated by P2Y2R-activation or P2X7R-blockade. A key element in the transduction mechanism of either receptor is the increase of the intracellular free Ca 2+ concentration, which may arise due to its release from intracellular storage sites (G protein-coupling; P2Y) or due to its passage through the receptor-channel itself from the extracellular space (ATP-gated ion channel; P2X). Further research is needed to clarify how purinergic signaling controls NSC/NPC fate and how the balance between the quiescent and activated states is established with fine and dynamic regulation. GLIA 2017;65:213-230. © 2016 Wiley Periodicals, Inc.

  9. TNF-α Regulates Mast Cell Functions by Inhibiting Cell Degranulation

    Directory of Open Access Journals (Sweden)

    Yuwei Gao

    2017-11-01

    Full Text Available Background/Aims: The aim of this study was to investigate the involvement of inducible co-stimulatory ligand (ICOSL expression in stimulation of mast cells (MCs by TNF-α and the ability of TNF-α stimulation of MCs to influence CD4+ T cell differentiation and function. The mechanisms underlying TNF-α stimulation of MCs were also explored. Methods: Mast cells and CD4+ T cells were prepared from C57BL/6 mice (aged 6–8 weeks. ICOSL expression by MCs was measured by real-time PCR and flow cytometry, and levels of IL-4, IL-10 and IFN-γ were measured by ELISA. Results: ICOSL expression by MCs was increased by TNF-α stimulation, and resulted in interaction with CD4+ T cells. The IL-4 and IL-10 levels in the co-culture system increased, while IFN-γ levels decreased. Furthermore, CD4+CD25+Foxp3+ T cell proliferation was induced by co-culture with TNF-α-stimulated MCs. The mechanism by which TNF-α stimulated MCs was dependent on the activation of the MAPK signaling pathway. Conclusion: TNF-α upregulated the expression of ICOSL on mast cells via a mechanism that is dependent on MAPK phosphorylation. TNF-α-treated MCs promoted the differentiation of regulatory T cells and induced a shift in cytokine expression from a Th1 to a Th2 profile by up-regulation ICOSL expression and inhibition of MC degranulation. Our study reveals a novel mechanism by which mast cells regulate T cell function.

  10. Innate Lymphoid Cells: A Promising New Regulator in Fibrotic Diseases.

    Science.gov (United States)

    Zhang, Yi; Tang, Jun; Tian, Zhiqiang; van Velkinburgh, Jennifer C; Song, Jianxun; Wu, Yuzhang; Ni, Bing

    2016-09-02

    Fibrosis is a consequence of chronic inflammation and the persistent accumulation of extracellular matrix, for which the cycle of tissue injury and repair becomes a predominant feature. Both the innate and adaptive immune systems play key roles in the progress of fibrosis. The recently identified subsets of innate lymphoid cells (ILCs), which are mainly localize to epithelial surfaces, have been characterized as regulators of chronic inflammation and tissue remodeling, representing a functional bridge between the innate and adaptive immunity. Moreover, recent research has implicated ILCs as potential contributing factors to several kinds of fibrosis diseases, such as hepatic fibrosis and pulmonary fibrosis. Here, we will summarize and discuss the key roles of ILCs and their related factors in fibrotic diseases and their potential for translation to the clinic.

  11. GATA-1 directly regulates Nanog in mouse embryonic stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Wen-Zhong; Ai, Zhi-Ying [College of Life Sciences, Northwest A& F University, Yangling 712100 (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100 (China); Wang, Zhi-Wei [School of Life Sciences and Medical Center, University of Science and Technology of China, Hefei, Anhui 230027 (China); Chen, Lin-Lin [College of Life Sciences, Northwest A& F University, Yangling 712100 (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100 (China); Guo, Ze-Kun, E-mail: gzknwaf@126.com [College of Veterinary Medicine, Northwest A& F University, Yangling 712100 (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100 (China); Zhang, Yong, E-mail: zylabnwaf@126.com [College of Veterinary Medicine, Northwest A& F University, Yangling 712100 (China); Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A& F University, Yangling 712100 (China)

    2015-09-25

    Nanog safeguards pluripotency in mouse embryonic stem cells (mESCs). Insight into the regulation of Nanog is important for a better understanding of the molecular mechanisms that control pluripotency of mESCs. In a silico analysis, we identify four GATA-1 putative binding sites in Nanog proximal promoter. The Nanog promoter activity can be significantly repressed by ectopic expression of GATA-1 evidenced by a promoter reporter assay. Mutation studies reveal that one of the four putative binding sites counts for GATA-1 repressing Nanog promoter activity. Direct binding of GATA-1 on Nanog proximal promoter is confirmed by electrophoretic mobility shift assay and chromatin immunoprecipitation. Our data provide new insights into the expanded regulatory circuitry that coordinates Nanog expression. - Highlights: • The Nanog proximal promoter conceives functional element for GATA-1. • GATA-1 occupies the Nanog proximal promoter in vitro and in vivo. • GATA-1 transcriptionally suppresses Nanog.

  12. A family business: stem cell progeny join the niche to regulate homeostasis.

    Science.gov (United States)

    Hsu, Ya-Chieh; Fuchs, Elaine

    2012-01-23

    Stem cell niches, the discrete microenvironments in which the stem cells reside, play a dominant part in regulating stem cell activity and behaviours. Recent studies suggest that committed stem cell progeny become indispensable components of the niche in a wide range of stem cell systems. These unexpected niche inhabitants provide versatile feedback signals to their stem cell parents. Together with other heterologous cell types that constitute the niche, they contribute to the dynamics of the microenvironment. As progeny are often located in close proximity to stem cell niches, similar feedback regulations may be the underlying principles shared by different stem cell systems.

  13. Posttranscriptional Regulation of Cyclooxygenase-2 in Rat Intestinal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Zhonghua Zhang

    2000-01-01

    Full Text Available Modulation of cyclooxygenase-2 (COX-2 mRNA stability plays an important role in the regulation of its expression by oncogenic Ras. Here, we evaluate COX-2 mRNA stability in response to treatment with two known endogenous promoters of gastrointestinal cancer, the bile acid (chenodeoxycholate; CD and ceramide. Treatment with CD and ceramide resulted in a 10-fold increase in the level of COX-2 protein and a four-fold lengthening of the half-life of COX-2 mRNA. COX-2 mRNA stability was assessed by Northern blot analysis and by evaluating the AU-rich element located in the COX-2 3′-UTR. A known inhibitor of mitogen-activated protein (MAP/extracellular signal-regulated kinase (ERK kinase (MEK, PD98059, reversed the effects of CD or ceramide to stabilize COX-2 mRNA. Overexpression of a dominant-negative ERK-1 or ERK-2 protein also led to destabilization of COX-2 mRNA. Treatment with a p38 MAPK inhibitor, PD169316, or transfection with a dominant-negative p38 MAPK construct reversed the effect of CD or ceramide to stabilize COX-2 mRNA. Expression of a dominant-negative c-Jun N-terminal kinase (JNK had no effect on COX-2 mRNA stability in cells treated with CD or ceramide. We conclude that posttranscriptional mechanisms play an important role in the regulation of COX-2 expression during carcinogenesis.

  14. Prion protein expression regulates embryonic stem cell pluripotency and differentiation.

    Directory of Open Access Journals (Sweden)

    Alberto Miranda

    2011-04-01

    Full Text Available Cellular prion protein (PRNP is a glycoprotein involved in the pathogenesis of transmissible spongiform encephalopathies (TSEs. Although the physiological function of PRNP is largely unknown, its key role in prion infection has been extensively documented. This study examines the functionality of PRNP during the course of embryoid body (EB differentiation in mouse Prnp-null (KO and WT embryonic stem cell (ESC lines. The first feature observed was a new population of EBs that only appeared in the KO line after 5 days of differentiation. These EBs were characterized by their expression of several primordial germ cell (PGC markers until Day 13. In a comparative mRNA expression analysis of genes playing an important developmental role during ESC differentiation to EBs, Prnp was found to participate in the transcription of a key pluripotency marker such as Nanog. A clear switching off of this gene on Day 5 was observed in the KO line as opposed to the WT line, in which maximum Prnp and Nanog mRNA levels appeared at this time. Using a specific antibody against PRNP to block PRNP pathways, reduced Nanog expression was confirmed in the WT line. In addition, antibody-mediated inhibition of ITGB5 (integrin αvβ5 in the KO line rescued the low expression of Nanog on Day 5, suggesting the regulation of Nanog transcription by Prnp via this Itgb5. mRNA expression analysis of the PRNP-related proteins PRND (Doppel and SPRN (Shadoo, whose PRNP function is known to be redundant, revealed their incapacity to compensate for the absence of PRNP during early ESC differentiation. Our findings provide strong evidence for a relationship between Prnp and several key pluripotency genes and attribute Prnp a crucial role in regulating self-renewal/differentiation status of ESC, confirming the participation of PRNP during early embryogenesis.

  15. Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Colin R Lickwar

    2017-08-01

    Full Text Available The intestinal epithelium serves critical physiologic functions that are shared among all vertebrates. However, it is unknown how the transcriptional regulatory mechanisms underlying these functions have changed over the course of vertebrate evolution. We generated genome-wide mRNA and accessible chromatin data from adult intestinal epithelial cells (IECs in zebrafish, stickleback, mouse, and human species to determine if conserved IEC functions are achieved through common transcriptional regulation. We found evidence for substantial common regulation and conservation of gene expression regionally along the length of the intestine from fish to mammals and identified a core set of genes comprising a vertebrate IEC signature. We also identified transcriptional start sites and other putative regulatory regions that are differentially accessible in IECs in all 4 species. Although these sites rarely showed sequence conservation from fish to mammals, surprisingly, they drove highly conserved IEC expression in a zebrafish reporter assay. Common putative transcription factor binding sites (TFBS found at these sites in multiple species indicate that sequence conservation alone is insufficient to identify much of the functionally conserved IEC regulatory information. Among the rare, highly sequence-conserved, IEC-specific regulatory regions, we discovered an ancient enhancer upstream from her6/HES1 that is active in a distinct population of Notch-positive cells in the intestinal epithelium. Together, these results show how combining accessible chromatin and mRNA datasets with TFBS prediction and in vivo reporter assays can reveal tissue-specific regulatory information conserved across 420 million years of vertebrate evolution. We define an IEC transcriptional regulatory network that is shared between fish and mammals and establish an experimental platform for studying how evolutionarily distilled regulatory information commonly controls IEC development

  16. Regulation of cyclooxygenase expression in cultured vascular cells

    International Nuclear Information System (INIS)

    Pash, J.M.

    1988-01-01

    Arachidonic acid metabolism in vascular tissue results in synthesis of prostacylin. The key enzyme in this synthesis pathway, cyclooxygenase, is down-regulated through self-inactivation. An analogous refractory state is produced by aspirin which irreversibly acetylates the enzyme. To further understand this phenomenon, the inactivation and recovery of cyclooxygenase activity was assayed in cultured ray vascular smooth muscle cells using exogenously added arachidonic acid. Self-inactivation of cyclooxygenase was observed following treatment with micromolar amounts of arachidonic acid. The recovery of cyclooxygenase activity following self-inactivation was analogous to that observed following aspirin-inactivation in that it depended on protein synthesis and required either serum or EGF. Two additional factors, TGF-β and uric acid, were found to enhance the stimulation of cyclooxygenase recovery by EGF. A defined medium containing 10 ng/mL EGF, 1 ng/mL TGFβ and 0.1 mM uric acid duplicated the cyclooxygenase recovery activity of 10% serum. Stimulation of cyclooxygenase activity by EGF and TGF-β was inhibited by cycloheximide but not by actinomycin-D, indicating a link to increased translation of pre-existing mRNA. A lack of significant effect on overall protein synthesis by EGF and TGF-β, measured by [ 35 S]-methionine incorporation under conditions where a multi-fold increase in cyclooxygenase activity was seen, indicates that the translational regulation of a small fraction of total mRNA and possibly cyclooxygenase is occurring

  17. Metastatic spread from squamous cell carcinoma of the hypopharynx to the totally implantable venous access port insertion site: Case report and review of literature.

    Science.gov (United States)

    Mangla, Ankit; Agarwal, Nikki; Mullane, Michael Russell

    2017-12-01

    The totally implantable venous access port plays a crucial role in delivering chemotherapy in the outpatient setting. Here, we report the first case of a patient with hypopharyngeal tumor who developed chest wall metastasis over the totally implantable venous access port inserted in the internal jugular vein. Our patient, a 58-year-old man with a hypopharyngeal tumor presented with a lump over the totally implantable venous access port site. The port was removed and the lump was biopsied. The CT studies showed that the tumor had spread along the catheter from the hypopharynx to the chest wall. The pathology from the biopsy showed squamous cell carcinoma (SCC). The patient had poor performance status and opted for hospice care. We present a novel case of metastasis over the totally implantable venous access port implanted in a patient with a hypopharyngeal tumor. We also reviewed relevant literature comparing the data from percutaneous endoscopic gastrostomy (PEG) tube site metastasis with our patient and other similar case reports. © 2017 Wiley Periodicals, Inc.

  18. Slug/SNAI2 regulates cell proliferation and invasiveness of metastatic prostate cancer cell lines.

    Science.gov (United States)

    Emadi Baygi, Modjtaba; Soheili, Zahra-Soheila; Essmann, Frank; Deezagi, Abdolkhaleg; Engers, Rainer; Goering, Wolfgang; Schulz, Wolfgang A

    2010-08-01

    Many metastatic cancers recapitulate the epithelial-to-mesenchymal transition (EMT) resulting in enhanced cell motility and invasiveness. The EMT is regulated by several transcription factors, including the zinc finger protein SNAI2, also named Slug, which appears to exert additional functions during development and cancer progression. We have studied the function of SNAI2 in prostate cancer cells. Quantitative RT-PCR analysis showed strong SNAI2 expression particularly in the PC-3 and PC3-16 prostate carcinoma cell lines. Knockdown of SNAI2 by specific siRNA induced changes in EMT markers and inhibited invasion of both cell lines into a matrigel matrix. SNAI2 siRNA-treated cells did not tolerate detachment from the culture plates, likely at least in part due to downregulation of integrin alpha6beta4. SNAI2 knockdown disturbed the microtubular and actin cytoskeletons, especially severely in PC-3 cells, resulting in grossly enlarged, flattened, and sometimes multinuclear cells. Knockdown also decreased cell proliferation, with a prominent G0/G1 arrest in PC3-16. Together, our data imply that SNAI2 exerts strong effects on the cytoskeleton and adhesion of those prostate cancer cells that express it and is necessary for their proliferation and invasiveness.

  19. Hypoxic stellate cells of pancreatic cancer stroma regulate extracellular matrix fiber organization and cancer cell motility.

    Science.gov (United States)

    Sada, Masafumi; Ohuchida, Kenoki; Horioka, Kohei; Okumura, Takashi; Moriyama, Taiki; Miyasaka, Yoshihiro; Ohtsuka, Takao; Mizumoto, Kazuhiro; Oda, Yoshinao; Nakamura, Masafumi

    2016-03-28

    Desmoplasia and hypoxia in pancreatic cancer mutually affect each other and create a tumor-supportive microenvironment. Here, we show that microenvironment remodeling by hypoxic pancreatic stellate cells (PSCs) promotes cancer cell motility through alteration of extracellular matrix (ECM) fiber architecture. Three-dimensional (3-D) matrices derived from PSCs under hypoxia exhibited highly organized parallel-patterned matrix fibers compared with 3-D matrices derived from PSCs under normoxia, and promoted cancer cell motility by inducing directional migration of cancer cells due to the parallel fiber architecture. Microarray analysis revealed that procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) in PSCs was the gene that potentially regulates ECM fiber architecture under hypoxia. Stromal PLOD2 expression in surgical specimens of pancreatic cancer was confirmed by immunohistochemistry. RNA interference-mediated knockdown of PLOD2 in PSCs blocked parallel fiber architecture of 3-D matrices, leading to decreased directional migration of cancer cells within the matrices. In conclusion, these findings indicate that hypoxia-induced PLOD2 expression in PSCs creates a permissive microenvironment for migration of cancer cells through architectural regulation of stromal ECM in pancreatic cancer. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  20. An eleven gene molecular signature for extra-capsular spread in oral squamous cell carcinoma serves as a prognosticator of outcome in patients without nodal metastases.

    Science.gov (United States)

    Wang, Weining; Lim, Weng Khong; Leong, Hui Sun; Chong, Fui Teen; Lim, Tony K H; Tan, Daniel S W; Teh, Bin Tean; Iyer, N Gopalakrishna

    2015-04-01

    Extracapsular spread (ECS) is an important prognostic factor for oral squamous cell carcinoma (OSCC) and is used to guide management. In this study, we aimed to identify an expression profile signature for ECS in node-positive OSCC using data derived from two different sources: a cohort of OSCC patients from our institution (National Cancer Centre Singapore) and The Cancer Genome Atlas (TCGA) head and neck squamous cell carcinoma (HNSCC) cohort. We also sought to determine if this signature could serve as a prognostic factor in node negative cancers. Patients with a histological diagnosis of OSCC were identified from an institutional database and fresh tumor samples were retrieved. RNA was extracted and gene expression profiling was performed using the Affymetrix GeneChip Human Genome U133 Plus 2.0 microarray platform. RNA sequence data and corresponding clinical data for the TCGA HNSCC cohort were downloaded from the TCGA Data Portal. All data analyses were conducted using R package and SPSS. We identified an 11 gene signature (GGH, MTFR1, CDKN3, PSRC1, SMIM3, CA9, IRX4, CPA3, ZSCAN16, CBX7 and ZFP3) which was robust in segregating tumors by ECS status. In node negative patients, patients harboring this ECS signature had a significantly worse overall survival (p=0.04). An eleven gene signature for ECS was derived. Our results also suggest that this signature is prognostic in a separate subset of patients with no nodal metastasis Further validation of this signature on other datasets and immunohistochemical studies are required to establish utility of this signature in stratifying early stage OSCC patients. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Regulation of TGFβ in the immune system: An emerging role for integrins and dendritic cells

    OpenAIRE

    Worthington, John J.; Fenton, Thomas M.; Czajkowska, Beata I.; Klementowicz, Joanna E.; Travis, Mark A.

    2012-01-01

    Regulation of an immune response requires complex crosstalk between cells of the innate and adaptive immune systems, via both cell?cell contact and secretion of cytokines. An important cytokine with a broad regulatory role in the immune system is transforming growth factor-? (TGF-?). TGF-? is produced by and has effects on many different cells of the immune system, and plays fundamental roles in the regulation of immune responses during homeostasis, infection and disease. Although many cells ...

  2. Does Erythropoietin Regulate TRPC Channels in Red Blood Cells?

    Directory of Open Access Journals (Sweden)

    Jens Danielczok

    2017-03-01

    Full Text Available Background: Cation channels play an essential role in red blood cells (RBCs ion homeostasis. One set of ion channels are the transient receptor potential channels of canonical type (TRPC channels. The abundance of these channels in primary erythroblasts, erythroid cell lines and RBCs was associated with an increase in intracellular Ca2+ upon stimulation with Erythropoietin (Epo. In contrast two independent studies on Epo-treated patients revealed diminished basal Ca2+ concentration or reduced phosphatidylserine exposure to the outer membrane leaflet. Methods: To resolve the seemingly conflicting reports we challenged mature human and mouse RBCs of several genotypes with Epo and Prostaglandin E2 (PGE2 and recorded the intracellular Ca2+ content. Next Generation Sequencing was utilised to approach a molecular analysis of reticulocytes. Results/Conclusions: Our results allow concluding that Epo and PGE2 regulation of the Ca2+ homeostasis is distinctly different between murine and human RBCs and that changes in intracellular Ca2+ upon Epo treatment is a primary rather than a compensatory effect. In human RBCs, Epo itself has no effect on Ca2+ fluxes but inhibits the PGE2-induced Ca2+ entry. In murine mature RBCs functional evidence indicates TRPC4/C5 mediated Ca2+ entry activated by Epo whereas PGE2 leads to a TRPC independent Ca2+ entry.

  3. Histones Induce the Procoagulant Phenotype of Endothelial Cells through Tissue Factor Up-Regulation and Thrombomodulin Down-Regulation.

    Science.gov (United States)

    Kim, Ji Eun; Yoo, Hyun Ju; Gu, Ja Yoon; Kim, Hyun Kyung

    2016-01-01

    The high circulating levels of histones found in various thrombotic diseases may compromise the anticoagulant barrier of endothelial cells. We determined how histones affect endothelial procoagulant tissue factor (TF) and anticoagulant thrombomodulin (TM). Surface antigens, soluble forms, and mRNA levels of TF and TM were measured by flow cytometry, ELISA, and real-time RT-PCR, respectively. TF and TM activity were measured using procoagulant activity, thrombin generation, or chromogenic assays. Involvement of the toll-like receptor (TLR) was assessed using the neutralizing antibodies. Histones dose-dependently induced surface antigens, activity and mRNA levels of endothelial TF. Histone-treated endothelial cells significantly shortened the lag time and enhanced the endogenous thrombin potential of normal plasma, which was normalized by a TF neutralizing antibody. Histones induced phosphatidylserine and protein-disulfide isomerase expression in endothelial cells. Histones also reduced the surface antigen, activity, and mRNA levels of endothelial TM. Polysialic acid and heparin reversed the histone-induced TF up-regulation and TM down-regulation. Activated protein C did not affect the TF up-regulation, but interrupted TM down-regulation. TLR2, and TLR4 inhibitors partially blocked the TF up-regulation. Histones induced the endothelial procoagulant phenotype through TF up-regulation and TM down-regulation. The effects of histones were partly mediated by TLR2, TLR4. Strategies to inhibit the harmful effects of histones in endothelial cells may be required in order to prevent a thrombotic environment.

  4. Effects of activated fibroblasts on phenotype modulation, EGFR signalling and cell cycle regulation in OSCC cells

    Energy Technology Data Exchange (ETDEWEB)

    Berndt, Alexander, E-mail: alexander.berndt@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Büttner, Robert, E-mail: Robert-Buettner@gmx.net [Institute of Biochemistry and Biophysics, Friedrich Schiller University Jena, 07740 Jena (Germany); Gühne, Stefanie, E-mail: stefanie_guehne@gmx.net [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Gleinig, Anna, E-mail: annagleinig@yahoo.com [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Richter, Petra, E-mail: P.Richter@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Chen, Yuan, E-mail: Yuan.Chen@med.uni-jena.de [Center for Molecular Biomedicine, Institute of Pathology, Jena University Hospital, 07740 Jena (Germany); Franz, Marcus, E-mail: Marcus.Franz@med.uni-jena.de [Clinic of Internal Medicine I, Jena University Hospital, 07740 Jena (Germany); Liebmann, Claus, E-mail: Claus.Liebmann@uni-jena.de [Institute of Biochemistry and Biophysics, Friedrich Schiller University Jena, 07740 Jena (Germany)

    2014-04-01

    Crosstalk between carcinoma associated fibroblasts (CAFs) and oral squamous cell carcinoma (OSCC) cells is suggested to mediate phenotype transition of cancer cells as a prerequisite for tumour progression, to predict patients’ outcome, and to influence the efficacy of EGFR inhibitor therapies. Here we investigate the influence of activated fibroblasts as a model for CAFs on phenotype and EGFR signalling in OSCC cells in vitro. For this, immortalised hTERT-BJ1 fibroblasts were activated with TGFβ1 and PDGFAB to generate a myofibroblast or proliferative phenotype, respectively. Conditioned media (FCM{sub TGF}, FCM{sub PDGF}) were used to stimulate PE/CA-PJ15 OSCC cells. Results were compared to the effect of conditioned media of non-stimulated fibroblasts (FCM{sub B}). FCM{sub TGF} stimulation leads to an up-regulation of vimentin in the OSCC cells and an enhancement of invasive behaviour, indicating EMT-like effects. Similarly, FCM{sub TGF}≫FCM{sub PDGF} induced up-regulation of EGFR, but not of ErbB2/ErbB3. In addition, we detected an increase in basal activities of ERK, PI3K/Akt and Stat3 (FCM{sub TGF}>FCM{sub PDGF}) accompanied by protein interaction of vimentin with pERK. These effects are correlated with an increased proliferation. In summary, our results suggest that the activated myofibroblast phenotype provides soluble factors which are able to induce EMT-like phenomena and to increase EGFR signalling as well as cell proliferation in OSCC cells. Our results indicate a possible influence of activated myofibroblasts on EGFR-inhibitor therapy. Therefore, CAFs may serve as promising novel targets for combined therapy strategies. - Highlights: • A cell culture model for cancer associated fibroblasts is described. • The mutual interaction with OSCC cells leads to up-regulation of EGFR in tumour cells. • mCAF induces EGFR downstream signalling with increased proliferation in OSCC. • Erk activation is associated with protein interaction with vimentin

  5. Effects of activated fibroblasts on phenotype modulation, EGFR signalling and cell cycle regulation in OSCC cells

    International Nuclear Information System (INIS)

    Berndt, Alexander; Büttner, Robert; Gühne, Stefanie; Gleinig, Anna; Richter, Petra; Chen, Yuan; Franz, Marcus; Liebmann, Claus

    2014-01-01

    Crosstalk between carcinoma associated fibroblasts (CAFs) and oral squamous cell carcinoma (OSCC) cells is suggested to mediate phenotype transition of cancer cells as a prerequisite for tumour progression, to predict patients’ outcome, and to influence the efficacy of EGFR inhibitor therapies. Here we investigate the influence of activated fibroblasts as a model for CAFs on phenotype and EGFR signalling in OSCC cells in vitro. For this, immortalised hTERT-BJ1 fibroblasts were activated with TGFβ1 and PDGFAB to generate a myofibroblast or proliferative phenotype, respectively. Conditioned media (FCM TGF , FCM PDGF ) were used to stimulate PE/CA-PJ15 OSCC cells. Results were compared to the effect of conditioned media of non-stimulated fibroblasts (FCM B ). FCM TGF stimulation leads to an up-regulation of vimentin in the OSCC cells and an enhancement of invasive behaviour, indicating EMT-like effects. Similarly, FCM TGF ≫FCM PDGF induced up-regulation of EGFR, but not of ErbB2/ErbB3. In addition, we detected an increase in basal activities of ERK, PI3K/Akt and Stat3 (FCM TGF >FCM PDGF ) accompanied by protein interaction of vimentin with pERK. These effects are correlated with an increased proliferation. In summary, our results suggest that the activated myofibroblast phenotype provides soluble factors which are able to induce EMT-like phenomena and to increase EGFR signalling as well as cell proliferation in OSCC cells. Our results indicate a possible influence of activated myofibroblasts on EGFR-inhibitor therapy. Therefore, CAFs may serve as promising novel targets for combined therapy strategies. - Highlights: • A cell culture model for cancer associated fibroblasts is described. • The mutual interaction with OSCC cells leads to up-regulation of EGFR in tumour cells. • mCAF induces EGFR downstream signalling with increased proliferation in OSCC. • Erk activation is associated with protein interaction with vimentin as sign of EMT. • Results qualify

  6. Glial cell ceruloplasmin and hepcidin differentially regulate iron efflux from brain microvascular endothelial cells.

    Science.gov (United States)

    McCarthy, Ryan C; Kosman, Daniel J

    2014-01-01

    We have used an in vitro model system to probe the iron transport pathway across the brain microvascular endothelial cells (BMVEC) of the blood-brain barrier (BBB). This model consists of human BMVEC (hBMVEC) and C6 glioma cells (as an astrocytic cell line) grown in a transwell, a cell culture system commonly used to quantify metabolite flux across a cell-derived barrier. We found that iron efflux from hBMVEC through the ferrous iron permease ferroportin (Fpn) was stimulated by secretion of the soluble form of the multi-copper ferroxidase, ceruloplasmin (sCp) from the co-cultured C6 cells. Reciprocally, expression of sCp mRNA in the C6 cells was increased by neighboring hBMVEC. In addition, data indicate that C6 cell-secreted hepcidin stimulates internalization of hBMVEC Fpn but only when the end-feet projections characteristic of this glia-derived cell line are proximal to the endothelial cells. This hepcidin-dependent loss of Fpn correlated with knock-down of iron efflux from the hBMVEC; this result was consistent with the mechanism by which hepcidin regulates iron efflux in mammalian cells. In summary, the data support a model of iron trafficking across the BBB in which the capillary endothelium induce the underlying astrocytes to produce the ferroxidase activity needed to support Fpn-mediated iron efflux. Reciprocally, astrocyte proximity modulates the effective concentration of hepcidin at the endothelial cell membrane and thus the surface expression of hBMVEC Fpn. These results are independent of the source of hBMVEC iron (transferrin or non-transferrin bound) indicating that the model developed here is broadly applicable to brain iron homeostasis.

  7. Discovery of a Splicing Regulator Required for Cell Cycle Progression

    Energy Technology Data Exchange (ETDEWEB)

    Suvorova, Elena S.; Croken, Matthew; Kratzer, Stella; Ting, Li-Min; Conde de Felipe, Magnolia; Balu, Bharath; Markillie, Lye Meng; Weiss, Louis M.; Kim, Kami; White, Michael W.

    2013-02-01

    In the G1 phase of the cell division cycle, eukaryotic cells prepare many of the resources necessary for a new round of growth including renewal of the transcriptional and protein synthetic capacities and building the machinery for chromosome replication. The function of G1 has an early evolutionary origin and is preserved in single and multicellular organisms, although the regulatory mechanisms conducting G1 specific functions are only understood in a few model eukaryotes. Here we describe a new G1 mutant from an ancient family of apicomplexan protozoans. Toxoplasma gondii temperature-sensitive mutant 12-109C6 conditionally arrests in the G1 phase due to a single point mutation in a novel protein containing a single RNA-recognition-motif (TgRRM1). The resulting tyrosine to asparagine amino acid change in TgRRM1 causes severe temperature instability that generates an effective null phenotype for this protein when the mutant is shifted to the restrictive temperature. Orthologs of TgRRM1 are widely conserved in diverse eukaryote lineages, and the human counterpart (RBM42) can functionally replace the missing Toxoplasma factor. Transcriptome studies demonstrate that gene expression is downregulated in the mutant at the restrictive temperature due to a severe defect in splicing that affects both cell cycle and constitutively expressed mRNAs. The interaction of TgRRM1 with factors of the tri-SNP complex (U4/U6 & U5 snRNPs) indicate this factor may be required to assemble an active spliceosome. Thus, the TgRRM1 family of proteins is an unrecognized and evolutionarily conserved class of splicing regulators. This study demonstrates investigations into diverse unicellular eukaryotes, like the Apicomplexa, have the potential to yield new insights into important mechanisms conserved across modern eukaryotic kingdoms.

  8. Optimizing Hybrid Spreading in Metapopulations.

    OpenAIRE

    Zhang, C.; Zhou, S.; Miller, J. C.; Cox, I. J.; Chain, B. M.

    2015-01-01

    Epidemic spreading phenomena are ubiquitous in nature and society. Examples include the spreading of diseases, information, and computer viruses. Epidemics can spread by local spreading, where infected nodes can only infect a limited set of direct target nodes and global spreading, where an infected node can infect every other node. In reality, many epidemics spread using a hybrid mixture of both types of spreading. In this study we develop a theoretical framework for studying hybrid epidemic...

  9. Optimizing Hybrid Spreading in Metapopulations

    OpenAIRE

    Zhang, Changwang; Zhou, Shi; Miller, Joel C.; Cox, Ingemar J.; Chain, Benjamin M.

    2014-01-01

    Epidemic spreading phenomena are ubiquitous in nature and society. Examples include the spreading of diseases, information, and computer viruses. Epidemics can spread by local spreading, where infected nodes can only infect a limited set of direct target nodes and global spreading, where an infected node can infect every other node. In reality, many epidemics spread using a hybrid mixture of both types of spreading. In this study we develop a theoretical framework for studying hybrid epidemic...

  10. Identification of transcription factors linked to cell cycle regulation in Arabidopsis

    OpenAIRE

    Dehghan Nayeri, Fatemeh

    2014-01-01

    Cell cycle is an essential process in growth and development of living organisms consists of the replication and mitotic phases separated by 2 gap phases; G1 and G2. It is tightly controlled at the molecular level and especially at the level of transcription. Precise regulation of the cell cycle is of central significance for plant growth and development and transcription factors are global regulators of gene expression playing essential roles in cell cycle regulation. This study has uncovere...

  11. Meningeal mast cell-T cell crosstalk regulates T cell encephalitogenicity.

    Science.gov (United States)

    Russi, Abigail E; Walker-Caulfield, Margaret E; Guo, Yong; Lucchinetti, Claudia F; Brown, Melissa A

    2016-09-01

    GM-CSF is a cytokine produced by T helper (Th) cells that plays an essential role in orchestrating neuroinflammation in experimental autoimmune encephalomyelitis, a rodent model of multiple sclerosis. Yet where and how Th cells acquire GM-CSF expression is unknown. In this study we identify mast cells in the meninges, tripartite tissues surrounding the brain and spinal cord, as important contributors to antigen-specific Th cell accumulation and GM-CSF expression. In the absence of mast cells, Th cells do not accumulate in the meninges nor produce GM-CSF. Mast cell-T cell co-culture experiments and selective mast cell reconstitution of the meninges of mast cell-deficient mice reveal that resident meningeal mast cells are an early source of caspase-1-dependent IL-1β that licenses Th cells to produce GM-CSF and become encephalitogenic. We also provide evidence of mast cell-T cell co-localization in the meninges and CNS of recently diagnosed acute MS patients indicating similar interactions may occur in human demyelinating disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Spread effects - methodology

    International Nuclear Information System (INIS)

    2004-01-01

    Diffusion of technology, environmental effects and rebound effects are the principal effects from the funding of renewable energy and energy economising. It is difficult to estimate the impact of the spread effects both prior to the measures are implemented and after the measures are carried out. Statistical methods can be used to estimate the spread effects, but they are insecure and always need to be complemented with qualitative and subjective evaluations. It is more adequate to evaluate potential spread effects from market and market data surveillance for a selection of technologies and parties. Based on this information qualitative indicators for spread effects can be constructed and used both ex ante and ex post (ml)

  13. Ocean, Spreading Centre

    Digital Repository Service at National Institute of Oceanography (India)

    Krishna, K.S.

    over the global midoceanic ridges have found some explicit relationships between spreading rate, seismic structure, and ridge-axis morphology. Bibliography Detrick, R. S., Buhl, P., Vera, E., Mutter, J., Orcutt, J., Madsen, J., and Brocher, T., 1987...

  14. Y-27632 Increases Sensitivity of PANC-1 Cells to EGCG in Regulating Cell Proliferation and Migration.

    Science.gov (United States)

    Liu, Xing; Bi, Yongyi

    2016-10-03

    BACKGROUND The study aimed to investigate the inhibitory effect of (1R,4r)-4-((R)-1-aminoethyl)-N-(pyridin-4-yl) cyclohexanecarboxamide (Y-27632) and (-)-epigallocatechin-3-gallate (EGCG) on the proliferation and migration of PANC-1 cells. EGCG, found in green tea, has been previously shown to be one of the most abundant and powerful catechins in cancer prevention and treatment. Y-27632, a selective inhibitor of rho-associated protein kinase 1, is widely used in treating cardiovascular disease, inflammation, and cancer. MATERIAL AND METHODS PANC-1 cells, maintained in Dulbecco's Modified Eagle's Medium, were treated with dimethyl sulfoxide (control) as well as different concentrations (20, 40, 60, and 80 μg/mL) of EGCG for 48 h. In addition, PANC-1 cells were treated separately with 60 μg/mL EGCG, 20 μM Y-27632, and EGCG combined with Y-27632 (60 μg/mL EGCG + 20 μM Y-27632) for 48 h. The effect of EGCG and Y-27632 on the proliferation and migration of PANC-1 cells was evaluated using Cell Counting Kit-8 and transwell migration assays. The expression of peroxisome proliferator-activated receptor alpha (PPARα) and Caspase-3 mRNA was determined by Quantitative real-time polymerase chain reaction (RT-qPCR). RESULTS EGCG (20-80 μg/mL) inhibited cell viability in a dose-dependent manner. Y-27632 enhanced the sensitivity of PANC-1 cells to EGCG (by increasing the expression of PPARa and Caspase-3 mRNA) and suppressed cell proliferation. PANC-1 cell migration was inhibited by treatment with a combination of EGCG and Y-27632. CONCLUSIONS Y-27632 increases the sensitivity of PANC-1 cells to EGCG in regulating cell proliferation and migration, which is likely to be related to the expression of PPARa mRNA and Caspase-3 mRNA.

  15. The VULCANO spreading programme

    Energy Technology Data Exchange (ETDEWEB)

    Cognet, G.; Laffont, G.; Jegou, C.; Journeau, C.; Sudreau, F.; Pierre, J.; Ramacciotti, M. [CEA (Atomic Energy Commission), DRN/DER - Bat. 212, CEA Cadarache, 13108 St. Paul Lez Durance (France)

    1999-07-01

    Among the currently studied core-catcher projects, some of them suppose corium spreading before cooling, in particular the EPR (European Pressurized Reactor) core-catcher concept is based on mixing the corium with a special concrete, spreading the molten mixture on a large multi-layer surface cooled from the bottom and subsequently cooling by flooding with water. Therefore, melt spreading deserves intensive investigation in order to determine and quantify key phenomena which govern the stopping of spreading. In France, for some years, the Nuclear Reactor Division of the Atomic Energy Commission (CEA/DRN) has undertaken a large program to improve knowledge on corium behaviour and coolability. This program is based on experimental and theoretical investigations which are finally gathered in scenario and mechanistic computer codes. In this framework, the real material experimental programme, VULCANO, conducted within an European frame, is currently devoted to the study of corium spreading. In 1997 and 1998, several tests have been performed on dry corium spreading with various composition of melts. Although all the observed phenomena, in particular the differences between simulant and real material melts have not been yet totally explained, these tests have already provided a lot of information about: The behaviour of complex mixtures including refractory oxides, silica, iron oxides and in one case iron metal; Spreading progression, which was never stopped in any of these tests by a crust formation at the front; The structure of spread melts (porosity, crusts,...); Physico-chemical interaction between melt and the refractory substratum which was composed of zirconia bricks. (authors)

  16. The VULCANO spreading programme

    International Nuclear Information System (INIS)

    Cognet, G.; Laffont, G.; Jegou, C.; Journeau, C.; Sudreau, F.; Pierre, J.; Ramacciotti, M.

    1999-01-01

    Among the currently studied core-catcher projects, some of them suppose corium spreading before cooling, in particular the EPR (European Pressurized Reactor) core-catcher concept is based on mixing the corium with a special concrete, spreading the molten mixture on a large multi-layer surface cooled from the bottom and subsequently cooling by flooding with water. Therefore, melt spreading deserves intensive investigation in order to determine and quantify key phenomena which govern the stopping of spreading. In France, for some years, the Nuclear Reactor Division of the Atomic Energy Commission (CEA/DRN) has undertaken a large program to improve knowledge on corium behaviour and coolability. This program is based on experimental and theoretical investigations which are finally gathered in scenario and mechanistic computer codes. In this framework, the real material experimental programme, VULCANO, conducted within an European frame, is currently devoted to the study of corium spreading. In 1997 and 1998, several tests have been performed on dry corium spreading with various composition of melts. Although all the observed phenomena, in particular the differences between simulant and real material melts have not been yet totally explained, these tests have already provided a lot of information about: The behaviour of complex mixtures including refractory oxides, silica, iron oxides and in one case iron metal; Spreading progression, which was never stopped in any of these tests by a crust formation at the front; The structure of spread melts (porosity, crusts,...); Physico-chemical interaction between melt and the refractory substratum which was composed of zirconia bricks. (authors)

  17. Regulation of gene expression in mammalian cells following ionizing radiation

    International Nuclear Information System (INIS)

    Boothman, D.A.; Lee, S.W

    1991-01-01

    Mammalian cells use a variety of mechanisms to control the expression of new gene transcrips elicited in response to ionizing radiation. Damage-induced proteins have been found which contain DNA binding sites located within the promoter regions of SV40 and human thymidine kinase genes. DNA binding proteins as well as proteins which bind to specific DNA lesions (e.g., XIP bp 175 binds specifically to X-ray-damaged DNA) may play a role in the initial recognition of DNA damage and may initiate DNA repair processes, along with new transcription. Mammalian gene expression after DNA damage is also regulated via the stabilization of preexisting mRNA transcripts. Stabilized mRNA transcripts are translated into protein products not previously present in the cell due to undefined posttranscriptional modifications. Thus far, the only example of mRNA stabilization following X-irradiation is the immediate induction of tissue-type plasminogen activator. Mammalian cells synthesize new mRNA transcripts indirect response to DNA damage. Using cDNA cloning, Northern RNA blotting and nuclear run-on techniques, the levels of a variety of known and previously unknown genes dramatically increase following X-irradiation. These genes/proteins now include; a) DNA binding transcripts factors, such as the UV-responsive element binding factors, ionizing radiation-induced DNA-binding proteins, and XIP bP 175; b) proto-oncogenes, such as c-fos, c-jun, and c-myc; c) several growth-related genes, (e.g., the gadd genes, protein kinase C, IL-1, and thymidine kinase); and d) a variety of other genes, including proteases, tumor necrosis factor-alpha, and DT diaphorase. Mammalian cells respond to X-irradiation by eliciting a very complex series of events resulting in the appearance of new genes and proteins. These gene products may affect DNA repair, adaptive responses, apoptosis, SOS-type mutagenic response, and/or carcinogenesis. (J.P.N.)

  18. PAX2 regulates ADAM10 expression and mediates anchorage-independent cell growth of melanoma cells.

    Directory of Open Access Journals (Sweden)

    Sophia Boyoung Lee

    Full Text Available PAX transcription factors play an important role during development and carcinogenesis. In this study, we investigated PAX2 protein levels in melanocytes and melanoma cells by Western Blot and immunofluorescence analysis and characterized the role of PAX2 in the pathogenesis of melanoma. In vitro we found weak PAX2 protein expression in keratinocytes and melanocytes. Compared to melanocytes increased PAX2 protein levels were detectable in melanoma cell lines. Interestingly, in tissue sections of melanoma patients nuclear PAX2 expression strongly correlated with nuclear atypia and the degree of prominent nucleoli, indicating an association of PAX2 with a more atypical cellular phenotype. In addition, with chromatin immunoprecipitation assay, PAX2 overexpression and PAX2 siRNA we present compelling evidence that PAX2 can regulate ADAM10 expression, a metalloproteinase known to play important roles in melanoma metastasis. In human tissue samples we found co-expression of PAX2 and ADAM10 in melanocytes of benign nevi and in melanoma cells of patients with malignant melanoma. Importantly, the downregulation of PAX2 by specific siRNA inhibited the anchorage independent cell growth and decreased the migratory and invasive capacity of melanoma cells. Furthermore, the downregulation of PAX2 abrogated the chemoresistance of melanoma cells against cisplatin, indicating that PAX2 expression mediates cell survival and plays important roles during melanoma progression.

  19. Adaptive Regulation of Osteopontin Production by Dendritic Cells Through the Bidirectional Interaction With Mesenchymal Stromal Cells

    Directory of Open Access Journals (Sweden)

    Sara Scutera

    2018-06-01

    Full Text Available Mesenchymal stromal cells (MSCs exert immunosuppressive effects on immune cells including dendritic cells (DCs. However, many details of the bidirectional interaction of MSCs with DCs are still unsolved and information on key molecules by which DCs can modulate MSC functions is limited. Here, we report that osteopontin (OPN, a cytokine involved in homeostatic and pathophysiologic responses, is constitutively expressed by DCs and regulated in the DC/MSC cocultures depending on the activation state of MSCs. Resting MSCs promoted OPN production, whereas the production of OPN was suppressed when MSCs were activated by proinflammatory cytokines (i.e., TNF-α, IL-6, and IL-1β. OPN induction required cell-to-cell contact, mediated at least in part, by β1 integrin (CD29. Conversely, activated MSCs inhibited the release of OPN via the production of soluble factors with a major role played by Prostaglandin E2 (PGE2. Accordingly, pretreatment with indomethacin significantly abrogated the MSC-mediated suppression of OPN while the direct addition of exogenous PGE2 inhibited OPN production by DCs. Furthermore, DC-conditioned medium promoted osteogenic differentiation of MSCs with a concomitant inhibition of adipogenesis. These effects were paralleled by the repression of the adipogenic markers PPARγ, adiponectin, and FABP4, and induction of the osteogenic markers alkaline phosphatase, RUNX2, and of the bone-anabolic chemokine CCL5. Notably, blocking OPN activity with RGD peptides or with an antibody against CD29, one of the OPN receptors, prevented the effects of DC-conditioned medium on MSC differentiation and CCL5 induction. Because MSCs have a key role in maintenance of bone marrow (BM hematopoietic stem cell niche through reciprocal regulation with immune cells, we investigated the possible MSC/DC interaction in human BM by immunohistochemistry. Although DCs (CD1c+ are a small percentage of BM cells, we demonstrated colocalization of CD271+ MSCs with

  20. Biomimetic brain tumor niche regulates glioblastoma cells towards a cancer stem cell phenotype.

    Science.gov (United States)

    Liu, Yung-Chiang; Lee, I-Chi; Chen, Pin-Yuan

    2018-05-01

    Glioblastoma (GBM) is the most malignant primary brain tumor and contains tumorigenic cancer stem cells (CSCs), which support the progression of tumor growth. The selection of CSCs and facilitation of the brain tumor niches may assist the development of novel therapeutics for GBM. Herein, hydrogel materials composed of agarose and hydroxypropyl methyl cellulose (HMC) in different concentrations were established and compared to emulate brain tumor niches and CSC microenvironments within a label-free system. Human GBM cell line, U-87 MG, was cultured on a series of HMC-agarose based culture system. Cell aggregation and spheroids formation were investigated after 4 days of culture, and 2.5% HMC-agarose based culture system demonstrated the largest spheroids number and size. Moreover, CD133 marker expression of GBM cells after 6 days of culture in 2.5% HMC-agarose based culture system was 60%, relatively higher than the control group at only 15%. Additionally, cells on 2.5% HMC-agarose based culture system show the highest chemoresistance, even at the high dose of 500 µM temozolomide for 72 h, the live cell ratio was still > 80%. Furthermore, the results also indicate that the expression of ABCG2 gene was up-regulated after culture in 2.5% HMC-agarose based culture system. Therefore, our results demonstrated that biomimetic brain tumor microenvironment may regulate GBM cells towards the CSC phenotype and expression of CSC characteristics. The microenvironment selection and spheroids formation in HMC-agarose based culture system may provide a label-free CSC selection strategy and drug testing model for future biomedical applications.

  1. Mastoparan-Induced Intracellular Ca2+ Fluxes May Regulate Cell-to-Cell Communication in Plants.

    Science.gov (United States)

    Tucker, E. B.; Boss, W. F.

    1996-06-01

    The relationship of Ca2+ and plasmodesmatal closure was examined in staminal hairs of Setcreasea purpurea by microinjecting cells with active mastoparan (Mas-7), inactive mastoparan (Mas-17), active inositol-1,4,5-trisphosphate (IP3), or inactive IP3. Calcium green dextran 10,000 was used to study cellular free Ca2+, and carboxyfluorescein was used to monitor plasmodesmatal closure. When Mas-7 was microinjected into the cytoplasm of cell 1 (the tip cell of a chain of cells), a rapid increase in calcium green dextran-10,000 fluorescence was observed in the cytoplasmic areas on both sides of the plasmodesmata connecting cells 1 and 2 during the same time that the diffusion of carboxyfluorescein through them was blocked. The inhibition of cell-to-cell diffusion was transient, and the closed plasmodesmata reopened within 30 s. The elevated Ca2+ level near plasmodesmata was also transient and returned to base level in about 1.5 min. The transient increase in Ca2+, once initiated in cell 1, repeated with an oscillatory period of 3 min. Elevated Ca2+ and oscillations of Ca2+ were also observed near interconnecting cell walls throughout the chain of cells, indicating that the signal had been transmitted. Previously, we reported that IP3 closed plasmodesmata; now we report that it stimulated Ca2+ and oscillations similar to Mas-7. The effect was specific for similar concentrations of Mas-7 over Mas-17 and active IP3 over inactive IP3. It is important that the Ca2+ channel blocker La3+ eliminated the responses from Mas-7 and IP3, indicating that an influx of Ca2+ was required. These results support the contention that plasmodesmata functioning is regulated via Ca2+ and that IP3 may be an intermediary between the stimulus and Ca2+ elevations.

  2. Cell recognition molecule L1 promotes embryonic stem cell differentiation through the regulation of cell surface glycosylation

    Energy Technology Data Exchange (ETDEWEB)

    Li, Ying [Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044 (China); Department of Clinical Laboratory, Second Affiliated Hospital of Dalian Medical University, Dalian 116023 (China); Huang, Xiaohua [Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044 (China); Department of Clinical Biochemistry, College of Laboratory Medicine, Dalian Medical University, Dalian 116044 (China); An, Yue [Department of Clinical Laboratory, Second Affiliated Hospital of Dalian Medical University, Dalian 116023 (China); Ren, Feng [Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044 (China); Yang, Zara Zhuyun; Zhu, Hongmei; Zhou, Lei [The Key Laboratory of Stem Cell and Regenerative Medicine, Institute of Molecular and Clinical Medicine, Kunming Medical University, Kunming 650228 (China); Department of Anatomy and Developmental Biology, Monash University, Clayton 3800 (Australia); He, Xiaowen; Schachner, Melitta [Keck Center for Collaborative Neuroscience and Department of Cell Biology and Neuroscience, Rutgers University, New Brunswick, NJ (United States); Xiao, Zhicheng, E-mail: zhicheng.xiao@monash.edu [The Key Laboratory of Stem Cell and Regenerative Medicine, Institute of Molecular and Clinical Medicine, Kunming Medical University, Kunming 650228 (China); Department of Anatomy and Developmental Biology, Monash University, Clayton 3800 (Australia); Ma, Keli, E-mail: makeli666@aliyun.com [Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044 (China); Li, Yali, E-mail: yalilipaper@gmail.com [Department of Biochemistry and Molecular Biology, Dalian Medical University, Dalian 116044 (China); Department of Anatomy, National University of Singapore, Singapore 119078 (Singapore)

    2013-10-25

    Highlights: •Down-regulating FUT9 and ST3Gal4 expression blocks L1-induced neuronal differentiation of ESCs. •Up-regulating FUT9 and ST3Gal4 expression in L1-ESCs depends on the activation of PLCγ. •L1 promotes ESCs to differentiate into neuron through regulating cell surface glycosylation. -- Abstract: Cell recognition molecule L1 (CD171) plays an important role in neuronal survival, migration, differentiation, neurite outgrowth, myelination, synaptic plasticity and regeneration after injury. Our previous study has demonstrated that overexpressing L1 enhances cell survival and proliferation of mouse embryonic stem cells (ESCs) through promoting the expression of FUT9 and ST3Gal4, which upregulates cell surface sialylation and fucosylation. In the present study, we examined whether sialylation and fucosylation are involved in ESC differentiation through L1 signaling. RNA interference analysis showed that L1 enhanced differentiation of ESCs into neurons through the upregulation of FUT9 and ST3Gal4. Furthermore, blocking the phospholipase Cγ (PLCγ) signaling pathway with either a specific PLCγ inhibitor or knockdown PLCγ reduced the expression levels of both FUT9 and ST3Gal4 mRNAs and inhibited L1-mediated neuronal differentiation. These results demonstrate that L1 promotes neuronal differentiation from ESCs through the L1-mediated enhancement of FUT9 and ST3Gal4 expression.

  3. Cell recognition molecule L1 promotes embryonic stem cell differentiation through the regulation of cell surface glycosylation

    International Nuclear Information System (INIS)

    Li, Ying; Huang, Xiaohua; An, Yue; Ren, Feng; Yang, Zara Zhuyun; Zhu, Hongmei; Zhou, Lei; He, Xiaowen; Schachner, Melitta; Xiao, Zhicheng; Ma, Keli; Li, Yali

    2013-01-01

    Highlights: •Down-regulating FUT9 and ST3Gal4 expression blocks L1-induced neuronal differentiation of ESCs. •Up-regulating FUT9 and ST3Gal4 expression in L1-ESCs depends on the activation of PLCγ. •L1 promotes ESCs to differentiate into neuron through regulating cell surface glycosylation. -- Abstract: Cell recognition molecule L1 (CD171) plays an important role in neuronal survival, migration, differentiation, neurite outgrowth, myelination, synaptic plasticity and regeneration after injury. Our previous study has demonstrated that overexpressing L1 enhances cell survival and proliferation of mouse embryonic stem cells (ESCs) through promoting the expression of FUT9 and ST3Gal4, which upregulates cell surface sialylation and fucosylation. In the present study, we examined whether sialylation and fucosylation are involved in ESC differentiation through L1 signaling. RNA interference analysis showed that L1 enhanced differentiation of ESCs into neurons through the upregulation of FUT9 and ST3Gal4. Furthermore, blocking the phospholipase Cγ (PLCγ) signaling pathway with either a specific PLCγ inhibitor or knockdown PLCγ reduced the expression levels of both FUT9 and ST3Gal4 mRNAs and inhibited L1-mediated neuronal differentiation. These results demonstrate that L1 promotes neuronal differentiation from ESCs through the L1-mediated enhancement of FUT9 and ST3Gal4 expression

  4. Enteric neural crest cells regulate vertebrate stomach patterning and differentiation.

    Science.gov (United States)

    Faure, Sandrine; McKey, Jennifer; Sagnol, Sébastien; de Santa Barbara, Pascal

    2015-01-15

    In vertebrates, the digestive tract develops from a uniform structure where reciprocal epithelial-mesenchymal interactions pattern this complex organ into regions with specific morphologies and functions. Concomitant with these early patterning events, the primitive GI tract is colonized by the vagal enteric neural crest cells (vENCCs), a population of cells that will give rise to the enteric nervous system (ENS), the intrinsic innervation of the GI tract. The influence of vENCCs on early patterning and differentiation of the GI tract has never been evaluated. In this study, we report that a crucial number of vENCCs is required for proper chick stomach development, patterning and differentiation. We show that reducing the number of vENCCs by performing vENCC ablations induces sustained activation of the BMP and Notch pathways in the stomach mesenchyme and impairs smooth muscle development. A reduction in vENCCs also leads to the transdifferentiation of the stomach into a stomach-intestinal mixed phenotype. In addition, sustained Notch signaling activity in the stomach mesenchyme phenocopies the defects observed in vENCC-ablated stomachs, indicating that inhibition of the Notch signaling pathway is essential for stomach patterning and differentiation. Finally, we report that a crucial number of vENCCs is also required for maintenance of stomach identity and differentiation through inhibition of the Notch signaling pathway. Altogether, our data reveal that, through the regulation of mesenchyme identity, vENCCs act as a new mediator in the mesenchymal-epithelial interactions that control stomach development. © 2015. Published by The Company of Biologists Ltd.

  5. Vaccine-induced antibodies to herpes simplex virus glycoprotein D epitopes involved in virus entry and cell-to-cell spread correlate with protection against genital disease in guinea pigs.

    Science.gov (United States)

    Hook, Lauren M; Cairns, Tina M; Awasthi, Sita; Brooks, Benjamin D; Ditto, Noah T; Eisenberg, Roselyn J; Cohen, Gary H; Friedman, Harvey M

    2018-05-01

    Herpes simplex virus type 2 (HSV-2) glycoprotein D (gD2) subunit antigen is included in many preclinical candidate vaccines. The rationale for including gD2 is to produce antibodies that block crucial gD2 epitopes involved in virus entry and cell-to-cell spread. HSV-2 gD2 was the only antigen in the Herpevac Trial for Women that protected against HSV-1 genital infection but not HSV-2. In that trial, a correlation was detected between gD2 ELISA titers and protection against HSV-1, supporting the importance of antibodies. A possible explanation for the lack of protection against HSV-2 was that HSV-2 neutralization titers were low, four-fold lower than to HSV-1. Here, we evaluated neutralization titers and epitope-specific antibody responses to crucial gD2 epitopes involved in virus entry and cell-to-cell spread as correlates of immune protection against genital lesions in immunized guinea pigs. We detected a strong correlation between neutralizing antibodies and protection against genital disease. We used a high throughput biosensor competition assay to measure epitope-specific responses to seven crucial gD2 linear and conformational epitopes involved in virus entry and spread. Some animals produced antibodies to most crucial epitopes while others produced antibodies to few. The number of epitopes recognized by guinea pig immune serum correlated with protection against genital lesions. We confirmed the importance of antibodies to each crucial epitope using monoclonal antibody passive transfer that improved survival and reduced genital disease in mice after HSV-2 genital challenge. We re-evaluated our prior study of epitope-specific antibody responses in women in the Herpevac Trial. Humans produced antibodies that blocked significantly fewer crucial gD2 epitopes than guinea pigs, and antibody responses in humans to some linear epitopes were virtually absent. Neutralizing antibody titers and epitope-specific antibody responses are important immune parameters to

  6. Bacterial cell-cell communication in the host via RRNPP peptide-binding regulators

    Directory of Open Access Journals (Sweden)

    David ePerez-Pascual

    2016-05-01

    Full Text Available Human microbiomes are composed of complex and dense bacterial consortia. In these environments, bacteria are able to react quickly to change by coordinating their gene expression at the population level via small signaling molecules. In Gram-positive bacteria, cell-cell communication is mostly mediated by peptides that are released into the extracellular environment. Cell-cell communication based on these peptides is especially widespread in the group Firmicutes, in which they regulate a wide array of biological processes, including functions related to host-microbe interactions. Among the different agents of communication, the RRNPP family of cytoplasmic transcriptional regulators, together with their cognate re-internalized signaling peptides, represents a group of emerging importance. RRNPP members that have been studied so far are found mainly in species of bacilli, streptococci, and enterococci. These bacteria are characterized as both human commensal and pathogenic, and share different niches in the human body with other microorganisms. The goal of this mini-review is to present the current state of research on the biological relevance of RRNPP mechanisms in the context of the host, highlighting their specific roles in commensalism or virulence.

  7. Leucine metabolism in regulation of insulin secretion from pancreatic beta cells

    OpenAIRE

    Yang, Jichun; Chi, Yujing; Burkhardt, Brant R.; Guan, Youfei; Wolf, Bryan A

    2010-01-01

    Leucine, a the branched-chain amino acids that must be supplied in daily diet, plays an important role in controlling protein synthesis and regulating cell metabolism in various cell types. In pancreatic β cells, leucine acutely stimulates insulin secretion by serving as both metabolic fuel and allosteric activator of glutamate dehydrogenase to enhance glutaminolysis. Leucine has also been shown to regulate gene transcription and protein synthesis in pancreatic islet β cells via both mTOR-dep...

  8. Membrane mechanisms and intracellular signalling in cell volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else Kay; Dunham, Philip B.

    1995-01-01

    Volume regulation, Signal transduction, Calcium-calmodulin, Stretch-activated channels, Eicosanoids, Macromolecular crowding, Cytoskeleton, Protein phosphorylation, dephosphorylation.......Volume regulation, Signal transduction, Calcium-calmodulin, Stretch-activated channels, Eicosanoids, Macromolecular crowding, Cytoskeleton, Protein phosphorylation, dephosphorylation....

  9. Distinct mechanisms regulate Lck spatial organization in activated T cells

    Directory of Open Access Journals (Sweden)

    Natasha eKapoor-Kaushik

    2016-03-01

    Full Text Available Phosphorylation of the T cell receptor (TCR by the kinase Lck is the first detectable signaling event upon antigen engagement. The distribution of Lck within the plasma membrane, its conformational state, kinase activity and protein interactions all contribute to determine how efficiently Lck phosphorylates the engaged TCR. Here we used cross-correlation raster image spectroscopy (ccRICS and photoactivated localization microscopy (PALM to identify two mechanisms of Lck clustering: an intrinsic mechanism of Lck clustering induced by locking Lck in its open conformation, and an extrinsic mechanism of clustering controlled by the phosphorylation of tyrosine 192, which regulates the affinity of Lck SH2 domain. Both mechanisms of clustering were differently affected by the absence of the kinase Zap70 or the adaptor Lat. We further observed that the adaptor TSAd bound to and promoted the diffusion of Lck when it is phosphorylated on tyrosine 192. Our data suggest that while Lck open conformation drives aggregation and clustering, the spatial organization of Lck is further controlled by signaling events downstream of TCR phosphorylation.

  10. Cytokine Regulation of Microenvironmental Cells in Myeloproliferative Neoplasms

    Directory of Open Access Journals (Sweden)

    Gregor Hoermann

    2015-01-01

    Full Text Available The term myeloproliferative neoplasms (MPN refers to a heterogeneous group of diseases including not only polycythemia vera (PV, essential thrombocythemia (ET, and primary myelofibrosis (PMF, but also chronic myeloid leukemia (CML, and systemic mastocytosis (SM. Despite the clinical and biological differences between these diseases, common pathophysiological mechanisms have been identified in MPN. First, aberrant tyrosine kinase signaling due to somatic mutations in certain driver genes is common to these MPN. Second, alterations of the bone marrow microenvironment are found in all MPN types and have been implicated in the pathogenesis of the diseases. Finally, elevated levels of proinflammatory and microenvironment-regulating cytokines are commonly found in all MPN-variants. In this paper, we review the effects of MPN-related oncogenes on cytokine expression and release and describe common as well as distinct pathogenetic mechanisms underlying microenvironmental changes in various MPN. Furthermore, targeting of the microenvironment in MPN is discussed. Such novel therapies may enhance the efficacy and may overcome resistance to established tyrosine kinase inhibitor treatment in these patients. Nevertheless, additional basic studies on the complex interplay of neoplastic and stromal cells are required in order to optimize targeting strategies and to translate these concepts into clinical application.

  11. CD44 regulates cell migration in human colon cancer cells via Lyn kinase and AKT phosphorylation.

    Science.gov (United States)

    Subramaniam, Venkateswaran; Vincent, Isabella R; Gardner, Helena; Chan, Emily; Dhamko, Helena; Jothy, Serge

    2007-10-01

    Colon cancer is among the leading causes of cancer death in North America. CD44, an adhesion and antiapoptotic molecule is overexpressed in colon cancer. Cofilin is involved in the directional motility of cells. In the present study, we looked at how CD44 might modulate cell migration in human colon cancer via cofilin. We used a human colon cancer cell line, HT29, which expresses CD44, HT29 where CD44 expression was knocked down by siRNA, SW620, a human colon cancer cell line which does not express CD44, stably transfected exons of CD44 in SW620 cells and the colon from CD44 knockout and wild-type mouse. Western blot analysis of siRNA CD44 lysates showed increased level of AKT phosphorylation and decreased level of cofilin expression. Similar results were also observed with SW620 cells and CD44 knockout mouse colon lysates. Experiments using the AKT phosphorylation inhibitor LY294002 indicate that AKT phosphorylation downregulates cofilin. Immunoprecipitation studies showed CD44 complex formation with Lyn, providing an essential link between CD44 and AKT phosphorylation. LY294002 also stabilized Lyn from phosphorylated AKT, suggesting an interaction between Lyn and AKT phosphorylation. Immunocytochemistry showed that cofilin and Lyn expression were downregulated in siRNA CD44 cells and CD44 knockout mouse colon. siRNA CD44 cells had significantly less migration compared to HT29 vector. Given the well-defined roles of CD44, phosphorylated AKT in apoptosis and cancer, these results indicate that CD44-induced cell migration is dependent on its complex formation with Lyn and its consequent regulation of AKT phosphorylation and cofilin expression.

  12. LOH at 6q and 10q in fractionated circulating DNA of ovarian cancer patients is predictive for tumor cell spread and overall survival

    Directory of Open Access Journals (Sweden)

    Kuhlmann Jan

    2012-07-01

    Full Text Available Abstract Background We recently showed that LOH proximal to M6P/IGF2R locus (D6S1581 in primary ovarian tumors is predictive for the presence of disseminated tumor cells (DTC in the bone marrow (BM. For therapy-monitoring, it would be highly desirable to establish a blood-based biomarker. Therefore, we quantified circulating DNA (cirDNA in sera of 63 ovarian cancer patients before surgery and after chemotherapy, measured incidence of LOH at four cancer-relevant chromosomal loci, correlated LOH with tumor cell spread to the BM and evaluated prognostic significance of LOH. Methods cirDNA was fractionated into high- and low molecular-weight fraction (HMWF, LMWF for LOH-profiling, utilizing PCR-based fluorescence microsatellite analysis. BM aspirates were analyzed for DTC by immunocytochemistry using the pan-cytokeratin antibody A45-B/B3. Results cirDNA levels in the HMWF before surgery were predictive for residual tumor load (p = 0.017. After chemotherapy, we observed a significant decline of cirDNA in the LMWF (p = 0.0001 but not in the HMWF. LOH was prevalently detected in the LMWF with an overall frequency of 67%, only moderately ablating after chemotherapy (45%. Before surgery, LOH in the LMWF at marker D10S1765 and D13S218 significantly correlated with tumor grading and FIGO stage (p = 0.033, p = 0.004, respectively. In both combined fractions, LOH at D6S1581 additionally associated with overall survival (OS (p = 0.030. Moreover, solely LOH at D10S1765 in LMWF after therapy correlated with DTC in BM after therapy (p = 0.017. Conclusion We demonstrate the applicability and necessity of DNA-fractionation prior to analyzing circulating LOH and identify LOH at D10S1765 and D6S1581 as novel blood-based biomarkers for ovarian cancer, being relevant for therapy-monitoring.

  13. Daughter-specific transcription factors regulate cell size control in budding yeast.

    Science.gov (United States)

    Di Talia, Stefano; Wang, Hongyin; Skotheim, Jan M; Rosebrock, Adam P; Futcher, Bruce; Cross, Frederick R

    2009-10-01

    In budding yeast, asymmetric cell division yields a larger mother and a smaller daughter cell, which transcribe different genes due to the daughter-specific transcription factors Ace2 and Ash1. Cell size control at the Start checkpoint has long been considered to be a main regulator of the length of the G1 phase of the cell cycle, resulting in longer G1 in the smaller daughter cells. Our recent data confirmed this concept using quantitative time-lapse microscopy. However, it has been proposed that daughter-specific, Ace2-dependent repression of expression of the G1 cyclin CLN3 had a dominant role in delaying daughters in G1. We wanted to reconcile these two divergent perspectives on the origin of long daughter G1 times. We quantified size control using single-cell time-lapse imaging of fluorescently labeled budding yeast, in the presence or absence of the daughter-specific transcriptional regulators Ace2 and Ash1. Ace2 and Ash1 are not required for efficient size control, but they shift the domain of efficient size control to larger cell size, thus increasing cell size requirement for Start in daughters. Microarray and chromatin immunoprecipitation experiments show that Ace2 and Ash1 are direct transcriptional regulators of the G1 cyclin gene CLN3. Quantification of cell size control in cells expressing titrated levels of Cln3 from ectopic promoters, and from cells with mutated Ace2 and Ash1 sites in the CLN3 promoter, showed that regulation of CLN3 expression by Ace2 and Ash1 can account for the differential regulation of Start in response to cell size in mothers and daughters. We show how daughter-specific transcriptional programs can interact with intrinsic cell size control to differentially regulate Start in mother and daughter cells. This work demonstrates mechanistically how asymmetric localization of cell fate determinants results in cell-type-specific regulation of the cell cycle.

  14. Daughter-Specific Transcription Factors Regulate Cell Size Control in Budding Yeast

    Science.gov (United States)

    Di Talia, Stefano; Wang, Hongyin; Skotheim, Jan M.; Rosebrock, Adam P.; Futcher, Bruce; Cross, Frederick R.

    2009-01-01

    In budding yeast, asymmetric cell division yields a larger mother and a smaller daughter cell, which transcribe different genes due to the daughter-specific transcription factors Ace2 and Ash1. Cell size control at the Start checkpoint has long been considered to be a main regulator of the length of the G1 phase of the cell cycle, resulting in longer G1 in the smaller daughter cells. Our recent data confirmed this concept using quantitative time-lapse microscopy. However, it has been proposed that daughter-specific, Ace2-dependent repression of expression of the G1 cyclin CLN3 had a dominant role in delaying daughters in G1. We wanted to reconcile these two divergent perspectives on the origin of long daughter G1 times. We quantified size control using single-cell time-lapse imaging of fluorescently labeled budding yeast, in the presence or absence of the daughter-specific transcriptional regulators Ace2 and Ash1. Ace2 and Ash1 are not required for efficient size control, but they shift the domain of efficient size control to larger cell size, thus increasing cell size requirement for Start in daughters. Microarray and chromatin immunoprecipitation experiments show that Ace2 and Ash1 are direct transcriptional regulators of the G1 cyclin gene CLN3. Quantification of cell size control in cells expressing titrated levels of Cln3 from ectopic promoters, and from cells with mutated Ace2 and Ash1 sites in the CLN3 promoter, showed that regulation of CLN3 expression by Ace2 and Ash1 can account for the differential regulation of Start in response to cell size in mothers and daughters. We show how daughter-specific transcriptional programs can interact with intrinsic cell size control to differentially regulate Start in mother and daughter cells. This work demonstrates mechanistically how asymmetric localization of cell fate determinants results in cell-type-specific regulation of the cell cycle. PMID:19841732

  15. Daughter-specific transcription factors regulate cell size control in budding yeast.

    Directory of Open Access Journals (Sweden)

    Stefano Di Talia

    2009-10-01

    Full Text Available In budding yeast, asymmetric cell division yields a larger mother and a smaller daughter cell, which transcribe different genes due to the daughter-specific transcription factors Ace2 and Ash1. Cell size control at the Start checkpoint has long been considered to be a main regulator of the length of the G1 phase of the cell cycle, resulting in longer G1 in the smaller daughter cells. Our recent data confirmed this concept using quantitative time-lapse microscopy. However, it has been proposed that daughter-specific, Ace2-dependent repression of expression of the G1 cyclin CLN3 had a dominant role in delaying daughters in G1. We wanted to reconcile these two divergent perspectives on the origin of long daughter G1 times. We quantified size control using single-cell time-lapse imaging of fluorescently labeled budding yeast, in the presence or absence of the daughter-specific transcriptional regulators Ace2 and Ash1. Ace2 and Ash1 are not required for efficient size control, but they shift the domain of efficient size control to larger cell size, thus increasing cell size requirement for Start in daughters. Microarray and chromatin immunoprecipitation experiments show that Ace2 and Ash1 are direct transcriptional regulators of the G1 cyclin gene CLN3. Quantification of cell size control in cells expressing titrated levels of Cln3 from ectopic promoters, and from cells with mutated Ace2 and Ash1 sites in the CLN3 promoter, showed that regulation of CLN3 expression by Ace2 and Ash1 can account for the differential regulation of Start in response to cell size in mothers and daughters. We show how daughter-specific transcriptional programs can interact with intrinsic cell size control to differentially regulate Start in mother and daughter cells. This work demonstrates mechanistically how asymmetric localization of cell fate determinants results in cell-type-specific regulation of the cell cycle.

  16. Go with the Flow: Cerebrospinal Fluid Flow Regulates Neural Stem Cell Proliferation.

    Science.gov (United States)

    Kaneko, Naoko; Sawamoto, Kazunobu

    2018-06-01

    Adult neural stem cells in the wall of brain ventricles make direct contact with cerebrospinal fluid. In this issue of Cell Stem Cell, Petrik et al. (2018) demonstrate that these neural stem cells sense the flow of cerebrospinal fluid through a transmembrane sodium channel, ENaC, which regulates their proliferation. Copyright © 2018 Elsevier Inc. All rights reserved.

  17. Role of Cytokine-Induced Glycosylation Changes in Regulating Cell Interactions and Cell Signaling in Inflammatory Diseases and Cancer

    Directory of Open Access Journals (Sweden)

    Justine H. Dewald

    2016-11-01

    Full Text Available Glycosylation is one of the most important modifications of proteins and lipids, and cell surface glycoconjugates are thought to play important roles in a variety of biological functions including cell-cell and cell-substrate interactions, bacterial adhesion, cell immunogenicity and cell signaling. Alterations of glycosylation are observed in number of diseases such as cancer and chronic inflammation. In that context, pro-inflammatory cytokines have been shown to modulate cell surface glycosylation by regulating the expression of glycosyltransferases involved in the biosynthesis of carbohydrate chains. These changes in cell surface glycosylation are also known to regulate cell signaling and could contribute to disease pathogenesis. This review summarizes our current knowledge of the glycosylation changes induced by pro-inflammatory cytokines, with a particular focus on cancer and cystic fibrosis, and their consequences on cell interactions and signaling.

  18. Induced pluripotent stem cells-derived myeloid-derived suppressor cells regulate the CD8+ T cell response

    Directory of Open Access Journals (Sweden)

    Daniel Joyce

    2018-05-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are markedly increased in cancer patients and tumor-bearing mice and promote tumor growth and survival by inhibiting host innate and adaptive immunity. In this study, we generated and characterized MDSCs from murine-induced pluripotent stem cells (iPSCs. The iPSCs were co-cultured with OP9 cells, stimulated with GM-CSF, and became morphologically heterologous under co-culturing with hepatic stellate cells. Allogeneic and OVA-specific antigen stimulation demonstrated that iPS-MDSCs have a T-cell regulatory function. Furthermore, a popliteal lymph node assay and autoimmune hepatitis model showed that iPS-MDSCs also regulate immune responsiveness in vivo and have a therapeutic effect against hepatitis. Taken together, our results demonstrated a method of generating functional MDSCs from iPSCs and highlighted the potential of iPS-MDSCs as a key cell therapy resource for transplantation and autoimmune diseases. Keywords: Myeloid-derived suppressor cells, Induced pluripotent stem cells, T cell response

  19. Altered epigenetic regulation of homeobox genes in human oral squamous cell carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Marcinkiewicz, Katarzyna M.; Gudas, Lorraine J., E-mail: ljgudas@med.cornell.edu

    2014-01-01

    To gain insight into oral squamous cell carcinogenesis, we performed deep sequencing (RNAseq) of non-tumorigenic human OKF6-TERT1R and tumorigenic SCC-9 cells. Numerous homeobox genes are differentially expressed between OKF6-TERT1R and SCC-9 cells. Data from Oncomine, a cancer microarray database, also show that homeobox (HOX) genes are dysregulated in oral SCC patients. The activity of Polycomb repressive complexes (PRC), which causes epigenetic modifications, and retinoic acid (RA) signaling can control HOX gene transcription. HOXB7, HOXC10, HOXC13, and HOXD8 transcripts are higher in SCC-9 than in OKF6-TERT1R cells; using ChIP (chromatin immunoprecipitation) we detected PRC2 protein SUZ12 and the epigenetic H3K27me3 mark on histone H3 at these genes in OKF6-TERT1R, but not in SCC-9 cells. In contrast, IRX1, IRX4, SIX2 and TSHZ3 transcripts are lower in SCC-9 than in OKF6-TERT1R cells. We detected SUZ12 and the H3K27me3 mark at these genes in SCC-9, but not in OKF6-TERT1R cells. SUZ12 depletion increased HOXB7, HOXC10, HOXC13, and HOXD8 transcript levels and decreased the proliferation of OKF6-TERT1R cells. Transcriptional responses to RA are attenuated in SCC-9 versus OKF6-TERT1R cells. SUZ12 and H3K27me3 levels were not altered by RA at these HOX genes in SCC-9 and OKF6-TERT1R cells. We conclude that altered activity of PRC2 is associated with dysregulation of homeobox gene expression in human SCC cells, and that this dysregulation potentially plays a role in the neoplastic transformation of oral keratinocytes. - Highlights: • RNAseq elucidates differences between non-tumorigenic and tumorigenic oral keratinocytes. • Changes in HOX mRNA in SCC-9 vs. OKF6-TERT1R cells are a result of altered epigenetic regulation. • RNAseq shows that retinoic acid (RA) influences gene expression in both OKF6-TERT1R and SCC-9 cells.

  20. Altered epigenetic regulation of homeobox genes in human oral squamous cell carcinoma cells

    International Nuclear Information System (INIS)

    Marcinkiewicz, Katarzyna M.; Gudas, Lorraine J.

    2014-01-01

    To gain insight into oral squamous cell carcinogenesis, we performed deep sequencing (RNAseq) of non-tumorigenic human OKF6-TERT1R and tumorigenic SCC-9 cells. Numerous homeobox genes are differentially expressed between OKF6-TERT1R and SCC-9 cells. Data from Oncomine, a cancer microarray database, also show that homeobox (HOX) genes are dysregulated in oral SCC patients. The activity of Polycomb repressive complexes (PRC), which causes epigenetic modifications, and retinoic acid (RA) signaling can control HOX gene transcription. HOXB7, HOXC10, HOXC13, and HOXD8 transcripts are higher in SCC-9 than in OKF6-TERT1R cells; using ChIP (chromatin immunoprecipitation) we detected PRC2 protein SUZ12 and the epigenetic H3K27me3 mark on histone H3 at these genes in OKF6-TERT1R, but not in SCC-9 cells. In contrast, IRX1, IRX4, SIX2 and TSHZ3 transcripts are lower in SCC-9 than in OKF6-TERT1R cells. We detected SUZ12 and the H3K27me3 mark at these genes in SCC-9, but not in OKF6-TERT1R cells. SUZ12 depletion increased HOXB7, HOXC10, HOXC13, and HOXD8 transcript levels and decreased the proliferation of OKF6-TERT1R cells. Transcriptional responses to RA are attenuated in SCC-9 versus OKF6-TERT1R cells. SUZ12 and H3K27me3 levels were not altered by RA at these HOX genes in SCC-9 and OKF6-TERT1R cells. We conclude that altered activity of PRC2 is associated with dysregulation of homeobox gene expression in human SCC cells, and that this dysregulation potentially plays a role in the neoplastic transformation of oral keratinocytes. - Highlights: • RNAseq elucidates differences between non-tumorigenic and tumorigenic oral keratinocytes. • Changes in HOX mRNA in SCC-9 vs. OKF6-TERT1R cells are a result of altered epigenetic regulation. • RNAseq shows that retinoic acid (RA) influences gene expression in both OKF6-TERT1R and SCC-9 cells

  1. ATM regulation of IL-8 links oxidative stress to cancer cell migration and invasion.

    Science.gov (United States)

    Chen, Wei-Ta; Ebelt, Nancy D; Stracker, Travis H; Xhemalce, Blerta; Van Den Berg, Carla L; Miller, Kyle M

    2015-06-01

    Ataxia-telangiectasia mutated (ATM) protein kinase regulates the DNA damage response (DDR) and is associated with cancer suppression. Here we report a cancer-promoting role for ATM. ATM depletion in metastatic cancer cells reduced cell migration and invasion. Transcription analyses identified a gene network, including the chemokine IL-8, regulated by ATM. IL-8 expression required ATM and was regulated by oxidative stress. IL-8 was validated as an ATM target by its ability to rescue cell migration and invasion defects in ATM-depleted cells. Finally, ATM-depletion in human breast cancer cells reduced lung tumors in a mouse xenograft model and clinical data validated IL-8 in lung metastasis. These findings provide insights into how ATM activation by oxidative stress regulates IL-8 to sustain cell migration and invasion in cancer cells to promote metastatic potential. Thus, in addition to well-established roles in tumor suppression, these findings identify a role for ATM in tumor progression.

  2. Proximity-Based Differential Single-Cell Analysis of the Niche to Identify Stem/Progenitor Cell Regulators.

    Science.gov (United States)

    Silberstein, Lev; Goncalves, Kevin A; Kharchenko, Peter V; Turcotte, Raphael; Kfoury, Youmna; Mercier, Francois; Baryawno, Ninib; Severe, Nicolas; Bachand, Jacqueline; Spencer, Joel A; Papazian, Ani; Lee, Dongjun; Chitteti, Brahmananda Reddy; Srour, Edward F; Hoggatt, Jonathan; Tate, Tiffany; Lo Celso, Cristina; Ono, Noriaki; Nutt, Stephen; Heino, Jyrki; Sipilä, Kalle; Shioda, Toshihiro; Osawa, Masatake; Lin, Charles P; Hu, Guo-Fu; Scadden, David T

    2016-10-06

    Physiological stem cell function is regulated by secreted factors produced by niche cells. In this study, we describe an unbiased approach based on the differential single-cell gene expression analysis of mesenchymal osteolineage cells close to, and further removed from, hematopoietic stem/progenitor cells (HSPCs) to identify candidate niche factors. Mesenchymal cells displayed distinct molecular profiles based on their relative location. We functionally examined, among the genes that were preferentially expressed in proximal cells, three secreted or cell-surface molecules not previously connected to HSPC biology-the secreted RNase angiogenin, the cytokine IL18, and the adhesion molecule Embigin-and discovered that all of these factors are HSPC quiescence regulators. Therefore, our proximity-based differential single-cell approach reveals molecular heterogeneity within niche cells and can be used to identify novel extrinsic stem/progenitor cell regulators. Similar approaches could also be applied to other stem cell/niche pairs to advance the understanding of microenvironmental regulation of stem cell function. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Proximity-based differential single cell analysis of the niche to identify stem/progenitor cell regulators

    Science.gov (United States)

    Silberstein, Lev; Goncalves, Kevin A; Kharchenko, Peter V; Turcotte, Raphael; Kfoury, Youmna; Mercier, Francois; Baryawno, Ninib; Severe, Nicolas; Bachand, Jacqueline; Spencer, Joel; Papazian, Ani; Lee, Dongjun; Chitteti, Brahmananda Reddy; Srour, Edward F; Hoggatt, Jonathan; Tate, Tiffany; Celso, Cristina Lo; Ono, Noriaki; Nutt, Stephen; Heino, Jyrki; Sipilä, Kalle; Shioda, Toshihiro; Osawa, Masatake; Lin, Charles P; Hu, Guo-fu; Scadden, David T

    2016-01-01

    SUMMARY Physiological stem cell function is regulated by secreted factors produced by niche cells. In this study, we describe an unbiased approach based on differential single-cell gene expression analysis of mesenchymal osteolineage cells close to and further removed from hematopoietic stem/progenitor cells to identify candidate niche factors. Mesenchymal cells displayed distinct molecular profiles based on their relative location. Amongst the genes which were preferentially expressed in proximal cells, we functionally examined three secreted or cell surface molecules not previously connected to HSPC biology: the secreted RNase Angiogenin, the cytokine IL18 and the adhesion molecule Embigin and discovered that all of these factors are HSPC quiescence regulators. Our proximity-based differential single cell approach therefore reveals molecular heterogeneity within niche cells and can be used to identify novel extrinsic stem/progenitor cell regulators. Similar approaches could also be applied to other stem cell/niche pairs to advance understanding of microenvironmental regulation of stem cell function. PMID:27524439

  4. Functioning of spontaneous and induced Con A regulators of T-cell proliferation. Modifying factors

    International Nuclear Information System (INIS)

    Kuz'mina, E.G.

    1989-01-01

    It is shown that active spontaneous non-specific regulators of T-cell proliferation are activated in peripheral blood ''in vivo'' by endogenous metabolites; non-specific regulator action can be induced ''in vitro'' by Con A, FGA. Non-specific regulators suppress and increase lymphocyte proliferation. Cyclic character of their functioning is revealed. 4 refs.; 1 tab

  5. Heat shock transcription factors regulate heat induced cell death in a ...

    Indian Academy of Sciences (India)

    2007-03-29

    Mar 29, 2007 ... Heat shock transcription factors regulate heat induced cell death in a rat ... the synthesis of heat shock proteins (Hsps) which is strictly regulated by ... The lack of Hsp synthesis in these cells was due to a failure in HSF1 DNA ...

  6. Differentiation, distribution and gammadelta T cell-driven regulation of IL-22-producing T cells in tuberculosis.

    Directory of Open Access Journals (Sweden)

    Shuyu Yao

    2010-02-01

    Full Text Available Differentiation, distribution and immune regulation of human IL-22-producing T cells in infections remain unknown. Here, we demonstrated in a nonhuman primate model that M. tuberculosis infection resulted in apparent increases in numbers of T cells capable of producing IL-22 de novo without in vitro Ag stimulation, and drove distribution of these cells more dramatically in lungs than in blood and lymphoid tissues. Consistently, IL-22-producing T cells were visualized in situ in lung tuberculosis (TB granulomas by confocal microscopy and immunohistochemistry, indicating that mature IL-22-producing T cells were present in TB granuloma. Surprisingly, phosphoantigen HMBPP activation of Vgamma2Vdelta2 T cells down-regulated the capability of T cells to produce IL-22 de novo in lymphocytes from blood, lung/BAL fluid, spleen and lymph node. Up-regulation of IFNgamma-producing Vgamma2Vdelta2 T effector cells after HMBPP stimulation coincided with the down-regulated capacity of these T cells to produce IL-22 de novo. Importantly, anti-IFNgamma neutralizing Ab treatment reversed the HMBPP-mediated down-regulation effect on IL-22-producing T cells, suggesting that Vgamma2Vdelta2 T-cell-driven IFNgamma-networking function was the mechanism underlying the HMBPP-mediated down-regulation of the capability of T cells to produce IL-22. These novel findings raise the possibility to ultimately investigate the function of IL-22 producing T cells and to target Vgamma2Vdelta2 T cells for balancing potentially hyper-activating IL-22-producing T cells in severe TB.

  7. Mitochondrial fission proteins regulate programmed cell death in yeast

    OpenAIRE

    Fannjiang, Yihru; Cheng, Wen-Chih; Lee, Sarah J.; Qi, Bing; Pevsner, Jonathan; McCaffery, J. Michael; Hill, R. Blake; Basañez, Gorka; Hardwick, J. Marie

    2004-01-01

    The possibility that single-cell organisms undergo programmed cell death has been questioned in part because they lack several key components of the mammalian cell death machinery. However, yeast encode a homolog of human Drp1, a mitochondrial fission protein that was shown previously to promote mammalian cell death and the excessive mitochondrial fragmentation characteristic of apoptotic mammalian cells. In support of a primordial origin of programmed cell death involving mitochondria, we fo...

  8. EEN regulates the proliferation and survival of multiple myeloma cells by potentiating IGF-1 secretion

    International Nuclear Information System (INIS)

    Huang, Er-Wen; Xue, Sheng-Jiang; Li, Xiao-Yan; Xu, Suo-Wen; Cheng, Jian-Ding; Zheng, Jin-Xiang; Shi, He; Lv, Guo-Li; Li, Zhi-Gang; Li, Yue; Liu, Chang-Hui; Chen, Xiao-Hui; Liu, Hong; Li, Jie; Liu, Chao

    2014-01-01

    Highlights: • Levels of EEN expression paralleled with the rate of cell proliferation. • EEN was involved in the proliferation and survival of multiple myeloma (MM) cells. • EEN regulated the activity of IGF-1-Akt/mTOR pathway. • EEN regulated proliferation and survival of MM cells by enhancing IGF-1 secretion. - Abstract: The molecular mechanisms of multiple myeloma are not well defined. EEN is an endocytosis-regulating molecule. Here we report that EEN regulates the proliferation and survival of multiple myeloma cells, by regulating IGF-1 secretion. In the present study, we observed that EEN expression paralleled with cell proliferation, EEN accelerated cell proliferation, facilitated cell cycle transition from G1 to S phase by regulating cyclin-dependent kinases (CDKs) pathway, and delayed cell apoptosis via Bcl2/Bax-mitochondrial pathway. Mechanistically, we found that EEN was indispensable for insulin-like growth factor-1 (IGF-1) secretion and the activation of protein kinase B-mammalian target of rapamycin (Akt-mTOR) pathway. Exogenous IGF-1 overcame the phenotype of EEN depletion, while IGF-1 neutralization overcame that of EEN over-expression. Collectively, these data suggest that EEN may play a pivotal role in excessive cell proliferation and insufficient cell apoptosis of bone marrow plasma cells in multiple myeloma. Therefore, EEN may represent a potential diagnostic marker or therapeutic target for multiple myeloma

  9. EEN regulates the proliferation and survival of multiple myeloma cells by potentiating IGF-1 secretion

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Er-Wen [Guangzhou Institute of Forensic Science, Guangzhou (China); Department of Forensic Pathology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou (China); Xue, Sheng-Jiang [Department of Forensic Pathology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou (China); Li, Xiao-Yan [Department of Pharmacy, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou (China); Xu, Suo-Wen [Department of Pharmacology and Toxicology, School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou (China); Cheng, Jian-Ding; Zheng, Jin-Xiang [Department of Forensic Pathology, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou (China); Shi, He; Lv, Guo-Li; Li, Zhi-Gang; Li, Yue; Liu, Chang-Hui; Chen, Xiao-Hui; Liu, Hong [Guangzhou Institute of Forensic Science, Guangzhou (China); Li, Jie, E-mail: mdlijie@sina.com [Department of Anaesthesiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou (China); Liu, Chao, E-mail: liuchaogaj@21cn.com [Guangzhou Institute of Forensic Science, Guangzhou (China)

    2014-05-02

    Highlights: • Levels of EEN expression paralleled with the rate of cell proliferation. • EEN was involved in the proliferation and survival of multiple myeloma (MM) cells. • EEN regulated the activity of IGF-1-Akt/mTOR pathway. • EEN regulated proliferation and survival of MM cells by enhancing IGF-1 secretion. - Abstract: The molecular mechanisms of multiple myeloma are not well defined. EEN is an endocytosis-regulating molecule. Here we report that EEN regulates the proliferation and survival of multiple myeloma cells, by regulating IGF-1 secretion. In the present study, we observed that EEN expression paralleled with cell proliferation, EEN accelerated cell proliferation, facilitated cell cycle transition from G1 to S phase by regulating cyclin-dependent kinases (CDKs) pathway, and delayed cell apoptosis via Bcl2/Bax-mitochondrial pathway. Mechanistically, we found that EEN was indispensable for insulin-like growth factor-1 (IGF-1) secretion and the activation of protein kinase B-mammalian target of rapamycin (Akt-mTOR) pathway. Exogenous IGF-1 overcame the phenotype of EEN depletion, while IGF-1 neutralization overcame that of EEN over-expression. Collectively, these data suggest that EEN may play a pivotal role in excessive cell proliferation and insufficient cell apoptosis of bone marrow plasma cells in multiple myeloma. Therefore, EEN may represent a potential diagnostic marker or therapeutic target for multiple myeloma.

  10. Forkhead box P3 regulates ARHGAP15 expression and affects migration of glioma cells through the Rac1 signaling pathway.

    Science.gov (United States)

    Sun, Zhen; Zhang, Biao; Wang, Chen; Fu, Tao; Li, Lianling; Wu, Qiaoli; Cai, Ying; Wang, Jinhuan

    2017-01-01

    Forkhead box P3 (FOXP3) plays a crucial role in the development and function of regulatory T cells and was recently identified as a tumor suppressor in different cancer types. Forkhead box P3 is expressed in normal brain tissues, but is strongly downregulated or absent in glioblastomas. In order to understand the FOXP3 adjustment mechanisms in glioma cells, we performed a DNA microarray in U87 cells overexpressing FOXP3 and validated the differences using quantitative real-time PCR, Western blot analysis, and immunohistochemistry in vitro and in vivo. We found that FOXP3 can regulate the expression of ARHGAP15. Expression of FOXP3 was also correlated with ARHGAP15 in glioma samples. Overexpression of FOXP3 inhibited glioma cell migration through ARHGAP15 upregulation and Rac1 inactivation. Silencing of FOXP3 promoted migration through ARHGAP15 downregulation and Rac1 activation. ARHGAP15, a GTPase-activating protein for Rac1, inhibits small GTPase signaling in a dual negative manner. We found that there is a correlation between expression of ARHGAP15 and glioma level. The small GTPase Rac1 plays an important role in cell migration. In addition, we found that FOXP3 regulates expression of epithelial-mesenchymal transition markers E-cadherin and N-cadherin, which is important given that epithelial-mesenchymal transition is critically involved in tumor spreading and dissemination. Thus, FOXP3 or ARHGAP15 may serve as a new molecular target for antimetastatic therapies in treating glioma. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  11. Redox regulation of stress signals: possible roles of dendritic stellate TRX producer cells (DST cell types).

    Science.gov (United States)

    Yodoi, Junji; Nakamura, Hajime; Masutani, Hiroshi

    2002-01-01

    Thioredoxin (TRX) is a 12 kDa protein with redox-active dithiol (Cys-Gly-Pro-Cys) in the active site. TRX is induced by a variety of stresses including viral infection and inflammation. The promoter sequences of the TRX gene contain a series of stress-responsive elements including ORE, ARE, XRE, CRE and SP-1. TRX promotes DNA binding of transcription factors such as NF-kappaB, AP-1 and p53. TRX interacts with target proteins modulating the activity of those proteins. We have identified TRX binding protein-2 (TBP-2), which was identical to vitamin D3 up-regulated protein 1 (VDUP1). Potential action of TBP-2/VDUP1 as a redox-sensitive tumor suppressor will be discussed. There is accumulating evidence for the involvement of TRX in the protection against infectious and inflammatory disorders. We will discuss the role of TRX-dependent redox regulation of the host defense mechanism, in particular its relation to the emerging concept of constitutive and/or inducible TRX on special cell types with dendritic and stellate morphology in the immune, endocrine and nervous systems, which we provisionally designate as dendritic stellate TRX producer cells (DST cell types).

  12. Signaling pathways in PACAP regulation of VIP gene expression in human neuroblastoma cells

    DEFF Research Database (Denmark)

    Falktoft, B.; Georg, B.; Fahrenkrug, J.

    2009-01-01

    Ganglia expressing the neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) innervate vasoactive intestinal peptide (VIP) containing neurons suggesting a role of PACAP in regulating VIP expression. Human NB-1 neuroblastoma cells were applied to study PACAP regulated VIP gene...... in PACAP regulation of the FOS and VIP gene expressions suggest for the first time a role of FOS in PACAP-induced VIP gene expression in human NB-1 neuroblastoma cells. (C) 2009 Elsevier Ltd. All rights reserved Udgivelsesdato: 2009/10...

  13. Mitochondrial Sirt3 supports cell proliferation by regulating glutamine-dependent oxidation in renal cell carcinoma

    International Nuclear Information System (INIS)

    Choi, Jieun; Koh, Eunjin; Lee, Yu Shin; Lee, Hyun-Woo; Kang, Hyeok Gu; Yoon, Young Eun; Han, Woong Kyu; Choi, Kyung Hwa; Kim, Kyung-Sup

    2016-01-01

    Clear cell renal carcinoma (RCC), the most common malignancy arising in the adult kidney, exhibits increased aerobic glycolysis and low mitochondrial respiration due to von Hippel-Lindau gene defects and constitutive hypoxia-inducible factor-α expression. Sirt3 is a major mitochondrial deacetylase that mediates various types of energy metabolism. However, the role of Sirt3 as a tumor suppressor or oncogene in cancer depends on cell types. We show increased Sirt3 expression in the mitochondrial fraction of human RCC tissues. Sirt3 depletion by lentiviral short-hairpin RNA, as well as the stable expression of the inactive mutant of Sirt3, inhibited cell proliferation and tumor growth in xenograft nude mice, respectively. Furthermore, mitochondrial pyruvate, which was used for oxidation in RCC, might be derived from glutamine, but not from glucose and cytosolic pyruvate, due to depletion of mitochondrial pyruvate carrier and the relatively high expression of malic enzyme 2. Depletion of Sirt3 suppressed glutamate dehydrogenase activity, leading to impaired mitochondrial oxygen consumption. Our findings suggest that Sirt3 plays a tumor-progressive role in human RCC by regulating glutamine-derived mitochondrial respiration, particularly in cells where mitochondrial usage of cytosolic pyruvate is severely compromised. -- Highlights: •Sirt3 is required for the maintenance of RCC cell proliferation. •Mitochondrial usage of cytosolic pyruvate is severely compromised in RCC. •Sirt3 supports glutamine-dependent oxidation in RCC.

  14. Electrophoresis of cell membrane heparan sulfate regulates galvanotaxis in glial cells.

    Science.gov (United States)

    Huang, Yu-Ja; Schiapparelli, Paula; Kozielski, Kristen; Green, Jordan; Lavell, Emily; Guerrero-Cazares, Hugo; Quinones-Hinojosa, Alfredo; Searson, Peter

    2017-08-01

    Endogenous electric fields modulate many physiological processes by promoting directional migration, a process known as galvanotaxis. Despite the importance of galvanotaxis in development and disease, the mechanism by which cells sense and migrate directionally in an electric field remains unknown. Here, we show that electrophoresis of cell surface heparan sulfate (HS) critically regulates this process. HS was found to be localized at the anode-facing side in fetal neural progenitor cells (fNPCs), fNPC-derived astrocytes and brain tumor-initiating cells (BTICs), regardless of their direction of galvanotaxis. Enzymatic removal of HS and other sulfated glycosaminoglycans significantly abolished or reversed the cathodic response seen in fNPCs and BTICs. Furthermore, Slit2, a chemorepulsive ligand, was identified to be colocalized with HS in forming a ligand gradient across cellular membranes. Using both imaging and genetic modification, we propose a novel mechanism for galvanotaxis in which electrophoretic localization of HS establishes cell polarity by functioning as a co-receptor and provides repulsive guidance through Slit-Robo signaling. © 2017. Published by The Company of Biologists Ltd.

  15. MTSS1 is epigenetically regulated in glioma cells and inhibits glioma cell motility

    Directory of Open Access Journals (Sweden)

    Daniel Luxen

    2017-02-01

    Full Text Available Epigenetic silencing by DNA methylation in brain tumors has been reported for many genes, however, their function on pathogenesis needs to be evaluated. We investigated the MTSS1 gene, identified as hypermethylated by differential methylation hybridization (DMH. Fifty-nine glioma tissue samples and seven glioma cell lines were examined for hypermethylation of the MTSS1 promotor, MTSS1 expression levels and gene dosage. GBM cell lines were treated with demethylating agents and interrogated for functional consequences of MTSS1 expression after transient transfection. Hypermethylation was significantly associated with IDH1/2 mutation. Comparative SNP analysis indicates higher incidence of loss of heterozygosity of MTSS1 in anaplastic astrocytomas and secondary glioblastomas as well as hypermethylation of the remaining allele. Reversal of promoter hypermethylation results in an increased MTSS1 expression. Cell motility was significantly inhibited by MTSS1 overexpression without influencing cell growth or apoptosis. Immunofluorescence analysis of MTSS1 in human astrocytes indicates co-localization with actin filaments. MTSS1 is down-regulated by DNA methylation in glioblastoma cell lines and is part of the G-CIMP phenotype in primary glioma tissues. Our data on normal astrocytes suggest a function of MTSS1 at focal contact structures with an impact on migratory capacity but no influence on apoptosis or cellular proliferation.

  16. Mitochondrial Sirt3 supports cell proliferation by regulating glutamine-dependent oxidation in renal cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Jieun; Koh, Eunjin; Lee, Yu Shin; Lee, Hyun-Woo; Kang, Hyeok Gu [Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Institute of Genetic Science, Integrated Genomic Research Center for Metabolic Regulation, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Yoon, Young Eun; Han, Woong Kyu [Department of Urology, Urological Science Institute, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Choi, Kyung Hwa [Department of Urology, CHA Bundang Medical Center, CHA University, Seongnam 463-712 (Korea, Republic of); Kim, Kyung-Sup, E-mail: KYUNGSUP59@yuhs.ac [Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Institute of Genetic Science, Integrated Genomic Research Center for Metabolic Regulation, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of)

    2016-06-03

    Clear cell renal carcinoma (RCC), the most common malignancy arising in the adult kidney, exhibits increased aerobic glycolysis and low mitochondrial respiration due to von Hippel-Lindau gene defects and constitutive hypoxia-inducible factor-α expression. Sirt3 is a major mitochondrial deacetylase that mediates various types of energy metabolism. However, the role of Sirt3 as a tumor suppressor or oncogene in cancer depends on cell types. We show increased Sirt3 expression in the mitochondrial fraction of human RCC tissues. Sirt3 depletion by lentiviral short-hairpin RNA, as well as the stable expression of the inactive mutant of Sirt3, inhibited cell proliferation and tumor growth in xenograft nude mice, respectively. Furthermore, mitochondrial pyruvate, which was used for oxidation in RCC, might be derived from glutamine, but not from glucose and cytosolic pyruvate, due to depletion of mitochondrial pyruvate carrier and the relatively high expression of malic enzyme 2. Depletion of Sirt3 suppressed glutamate dehydrogenase activity, leading to impaired mitochondrial oxygen consumption. Our findings suggest that Sirt3 plays a tumor-progressive role in human RCC by regulating glutamine-derived mitochondrial respiration, particularly in cells where mitochondrial usage of cytosolic pyruvate is severely compromised. -- Highlights: •Sirt3 is required for the maintenance of RCC cell proliferation. •Mitochondrial usage of cytosolic pyruvate is severely compromised in RCC. •Sirt3 supports glutamine-dependent oxidation in RCC.

  17. A plant U-box protein, PUB4, regulates asymmetric cell division and cell proliferation in the root meristem

    NARCIS (Netherlands)

    Kinoshita, A.; Hove, ten C.A.; Tabata, R.; Yamada, M.; Shimizu, N.; Ishida, T.; Yamaguchi, K.; Shigenobu, S.; Takebayashi, Y.; Luchies, J.; Kobayashi, M.; Kurata, T.; Wada, T.; Seo, M.; Hasebe, M.; Blilou, I.; Fukuda, H.; Scheres, B.; Heidstra, R.; Kamiya, Y.; Sawa, S.

    2015-01-01

    The root meristem (RM) is a fundamental structure that is responsible for postembryonic root growth. The RM contains the quiescent center (QC), stem cells and frequently dividing meristematic cells, in which the timing and the frequency of cell division are tightly regulated. In Arabidopsis

  18. Optimizing hybrid spreading in metapopulations.

    Science.gov (United States)

    Zhang, Changwang; Zhou, Shi; Miller, Joel C; Cox, Ingemar J; Chain, Benjamin M

    2015-04-29

    Epidemic spreading phenomena are ubiquitous in nature and society. Examples include the spreading of diseases, information, and computer viruses. Epidemics can spread by local spreading, where infected nodes can only infect a limited set of direct target nodes and global spreading, where an infected node can infect every other node. In reality, many epidemics spread using a hybrid mixture of both types of spreading. In this study we develop a theoretical framework for studying hybrid epidemics, and examine the optimum balance between spreading mechanisms in terms of achieving the maximum outbreak size. We show the existence of critically hybrid epidemics where neither spreading mechanism alone can cause a noticeable spread but a combination of the two spreading mechanisms would produce an enormous outbreak. Our results provide new strategies for maximising beneficial epidemics and estimating the worst outcome of damaging hybrid epidemics.

  19. Regulation of CD4 T cells and their effects on immunopathological inflammation following viral infection.

    Science.gov (United States)

    Bhattacharyya, Mitra; Madden, Patrick; Henning, Nathan; Gregory, Shana; Aid, Malika; Martinot, Amanda J; Barouch, Dan H; Penaloza-MacMaster, Pablo

    2017-10-01

    CD4 T cells help immune responses, but knowledge of how memory CD4 T cells are regulated and how they regulate adaptive immune responses and induce immunopathology is limited. Using adoptive transfer of virus-specific CD4 T cells, we show that naive CD4 T cells undergo substantial expansion following infection, but can induce lethal T helper type 1-driven inflammation. In contrast, memory CD4 T cells exhibit a biased proliferation of T follicular helper cell subsets and were able to improve adaptive immune responses in the context of minimal tissue damage. Our analyses revealed that type I interferon regulates the expansion of primary CD4 T cells, but does not seem to play a critical role in regulating the expansion of secondary CD4 T cells. Strikingly, blockade of type I interferon abrogated lethal inflammation by primary CD4 T cells following viral infection, despite that this treatment increased the numbers of primary CD4 T-cell responses. Altogether, these data demonstrate important aspects of how primary and secondary CD4 T cells are regulated in vivo, and how they contribute to immune protection and immunopathology. These findings are important for rational vaccine design and for improving adoptive T-cell therapies against persistent antigens. © 2017 John Wiley & Sons Ltd.

  20. Rac1 mediates cytokine-stimulated hemocyte spreading via prostaglandin biosynthesis in the beet armyworm, Spodoptera exigua

    Science.gov (United States)

    Cell spreading is an integral component of insect hemocytic immune reactions to infections and invasions. Cell spreading is accomplished by cytoskeleton rearrangement, which is activated by three major immune mediators, biogenic monoamines, plasmatocyte-spreading peptide (PSP), and eicosanoids, part...

  1. Gal-3 regulates the capacity of dendritic cells to promote NKT-cell-induced liver injury.

    Science.gov (United States)

    Volarevic, Vladislav; Markovic, Bojana Simovic; Bojic, Sanja; Stojanovic, Maja; Nilsson, Ulf; Leffler, Hakon; Besra, Gurdyal S; Arsenijevic, Nebojsa; Paunovic, Verica; Trajkovic, Vladimir; Lukic, Miodrag L

    2015-02-01

    Galectin-3 (Gal-3), an endogenous lectin, exhibits pro- and anti-inflammatory effects in various disease conditions. In order to explore the role of Gal-3 in NKT-cell-dependent pathology, we induced hepatitis in C57BL/6 WT and Gal-3-deficient mice by using specific ligand for NKT cells: α-galactosylceramide, glycolipid Ag presented by CD1d. The injection of α-galactosylceramide significantly enhanced expression of Gal-3 in liver NKT and dendritic cells (DCs). Genetic deletion or selective inhibition of Gal-3 (induced by Gal-3-inhibitor TD139) abrogated the susceptibility to NKT-cell-dependent hepatitis. Blood levels of pro-inflammatory cytokines (TNF-α, IFN-γ, IL-12) and their production by liver DCs and NKT cells were also downregulated. Genetic deletion or selective inhibition of Gal-3 alleviated influx of inflammatory CD11c(+) CD11b(+) DCs in the liver and favored tolerogenic phenotype and IL-10 production of liver NKT and DCs. Deletion of Gal-3 attenuated the capacity of DCs to support liver damage in the passive transfer experiments and to produce pro-inflammatory cytokines in vitro. Gal-3-deficient DCs failed to optimally stimulate production of pro-inflammatory cytokines in NKT cells, in vitro and in vivo. In conclusion, Gal-3 regulates the capacity of DCs to support NKT-cell-mediated liver injury, playing an important pro-inflammatory role in acute liver injury. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Regulation of normal B-cell differentiation and malignant B-cell survival by OCT2.

    Science.gov (United States)

    Hodson, Daniel J; Shaffer, Arthur L; Xiao, Wenming; Wright, George W; Schmitz, Roland; Phelan, James D; Yang, Yandan; Webster, Daniel E; Rui, Lixin; Kohlhammer, Holger; Nakagawa, Masao; Waldmann, Thomas A; Staudt, Louis M

    2016-04-05

    The requirement for the B-cell transcription factor OCT2 (octamer-binding protein 2, encoded by Pou2f2) in germinal center B cells has proved controversial. Here, we report that germinal center B cells are formed normally after depletion of OCT2 in a conditional knockout mouse, but their proliferation is reduced and in vivo differentiation to antibody-secreting plasma cells is blocked. This finding led us to examine the role of OCT2 in germinal center-derived lymphomas. shRNA knockdown showed that almost all diffuse large B-cell lymphoma (DLBCL) cell lines are addicted to the expression of OCT2 and its coactivator OCA-B. Genome-wide chromatin immunoprecipitation (ChIP) analysis and gene-expression profiling revealed the broad transcriptional program regulated by OCT2 that includes the expression of STAT3, IL-10, ELL2, XBP1, MYC, TERT, and ADA. Importantly, genetic alteration of OCT2 is not a requirement for cellular addiction in DLBCL. However, we detected amplifications of the POU2F2 locus in DLBCL tumor biopsies and a recurrent mutation of threonine 223 in the DNA-binding domain of OCT2. This neomorphic mutation subtly alters the DNA-binding preference of OCT2, leading to the transactivation of noncanonical target genes including HIF1a and FCRL3 Finally, by introducing mutations designed to disrupt the OCT2-OCA-B interface, we reveal a requirement for this protein-protein interface that ultimately might be exploited therapeutically. Our findings, combined with the predominantly B-cell-restricted expression of OCT2 and the absence of a systemic phenotype in our knockout mice, suggest that an OCT2-targeted therapeutic strategy would be efficacious in both major subtypes of DLBCL while avoiding systemic toxicity.

  3. A balance of FGF and BMP signals regulates cell cycle exit and Equarin expression in lens cells

    Science.gov (United States)

    Jarrin, Miguel; Pandit, Tanushree; Gunhaga, Lena

    2012-01-01

    In embryonic and adult lenses, a balance of cell proliferation, cell cycle exit, and differentiation is necessary to maintain physical function. The molecular mechanisms regulating the transition of proliferating lens epithelial cells to differentiated primary lens fiber cells are poorly characterized. To investigate this question, we used gain- and loss-of-function analyses to modulate fibroblast growth factor (FGF) and/or bone morphogenetic protein (BMP) signals in chick lens/retina explants. Here we show that FGF activity plays a key role for proliferation independent of BMP signals. Moreover, a balance of FGF and BMP signals regulates cell cycle exit and the expression of Ccdc80 (also called Equarin), which is expressed at sites where differentiation of lens fiber cells occurs. BMP activity promotes cell cycle exit and induces Equarin expression in an FGF-dependent manner. In contrast, FGF activity is required but not sufficient to induce cell cycle exit or Equarin expression. Furthermore, our results show that in the absence of BMP activity, lens cells have increased cell cycle length or are arrested in the cell cycle, which leads to decreased cell cycle exit. Taken together, these findings suggest that proliferation, cell cycle exit, and early differentiation of primary lens fiber cells are regulated by counterbalancing BMP and FGF signals. PMID:22718906

  4. Protein kinase C signaling and cell cycle regulation

    OpenAIRE

    Black, Adrian R.; Black, Jennifer D.

    2013-01-01

    A link between T cell proliferation and the protein kinase C (PKC) family of serine/threonine kinases has been recognized for about thirty years. However, despite the wealth of information on PKC-mediated control of T cell activation, understanding of the effects of PKCs on the cell cycle machinery in this cell type remains limited. Studies in other systems have revealed important cell cycle-specific effects of PKC signaling that can either positively or negatively impact proliferation. Th...

  5. HDAC up-regulation in early colon field carcinogenesis is involved in cell tumorigenicity through regulation of chromatin structure.

    Directory of Open Access Journals (Sweden)

    Yolanda Stypula-Cyrus

    Full Text Available Normal cell function is dependent on the proper maintenance of chromatin structure. Regulation of chromatin structure is controlled by histone modifications that directly influence chromatin architecture and genome function. Specifically, the histone deacetylase (HDAC family of proteins modulate chromatin compaction and are commonly dysregulated in many tumors, including colorectal cancer (CRC. However, the role of HDAC proteins in early colorectal carcinogenesis has not been previously reported. We found HDAC1, HDAC2, HDAC3, HDAC5, and HDAC7 all to be up-regulated in the field of human CRC. Furthermore, we observed that HDAC2 up-regulation is one of the earliest events in CRC carcinogenesis and observed this in human field carcinogenesis, the azoxymethane-treated rat model, and in more aggressive colon cancer cell lines. The universality of HDAC2 up-regulation suggests that HDAC2 up-regulation is a novel and important early event in CRC, which may serve as a biomarker. HDAC inhibitors (HDACIs interfere with tumorigenic HDAC activity; however, the precise mechanisms involved in this process remain to be elucidated. We confirmed that HDAC inhibition by valproic acid (VPA targeted the more aggressive cell line. Using nuclease digestion assays and transmission electron microscopy imaging, we observed that VPA treatment induced greater changes in chromatin structure in the more aggressive cell line. Furthermore, we used the novel imaging technique partial wave spectroscopy (PWS to quantify nanoscale alterations in chromatin. We noted that the PWS results are consistent with the biological assays, indicating a greater effect of VPA treatment in the more aggressive cell type. Together, these results demonstrate the importance of HDAC activity in early carcinogenic events and the unique role of higher-order chromatin structure in determining cell tumorigenicity.

  6. Analysis of cell-cycle regulation following exposure of lung-derived cells to γ-rays

    Science.gov (United States)

    Trani, D.; Lucchetti, C.; Cassone, M.; D'Agostino, L.; Caputi, M.; Giordano, A.

    Acute exposure of mammalian cells to ionizing radiation results in a delay of cell-cycle progression and/or augmentation of apoptosis. Following ionizing radiation-induced DNA damage, cell-cycle arrest in the G1- or G2-phase of the cell-cycle prevents or delays DNA replication or mitosis, providing time for the DNA repair machinery to exert its function. Deregulation or failing of cell-cycle checkpoints and/or DNA repair mechanisms may lead normal cells bearing chromosome mutations to acquire neoplastic autonomy, which in turn can trigger the onset of cancer. Existing studies have focused on the impact of p53 status on the radiation response of lung cancer (LC) cell lines in terms of both cell-cycle regulation and apoptosis, while no comparative studies have been performed on the radiation response of lung derived normal and cancerous epithelial cells. To investigate the radiation response in normal and cancerous phenotypes, along with the role and impact of p53 status, and possible correlations with pRb/p105 or other proteins involved in carcinogenesis and cell-cycle regulation, we selected two lung-derived epithelial cell lines, one normal (NL20, p53 wild-type) and one non-small cell lung cancer (NSCLC), H358 (known to be p53-deficient). We compared the levels of γ-induced cell proliferation ability, cell-cycle arrest, apoptotic index, and expression levels of cell-cycle regulating and regulated proteins. The different cell sensitivity, apoptotic response and protein expression profiles resulting from our study for NL20 and H358 cells suggest that still unknown mechanisms involving p53, pRb/p105 and their target molecules might play a pivotal role in determining cell sensitivity and resistance upon exposure to ionizing radiation.

  7. Spread of Canine Flu

    Centers for Disease Control (CDC) Podcasts

    2018-04-05

    Dr. Colin Parrish, a Professor of Virology at the College of Veterinary Medicine, Cornell University, discusses the spread of influenza among dogs.  Created: 4/5/2018 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 4/5/2018.

  8. Engineering amount of cell-cell contact demonstrates biphasic proliferative regulation through RhoA and the actin cytoskeleton

    International Nuclear Information System (INIS)

    Gray, Darren S.; Liu, Wendy F.; Shen, Colette J.; Bhadriraju, Kiran; Nelson, Celeste M.; Chen, Christopher S.

    2008-01-01

    Endothelial cell-cell contact via VE-cadherin plays an important role in regulating numerous cell functions, including proliferation. However, using different experimental approaches to manipulate cell-cell contact, investigators have observed both inhibition and stimulation of proliferation depending on the adhesive context. In this study, we used micropatterned wells combined with active positioning of cells by dielectrophoresis in order to investigate whether the number of contacting neighbors affected the proliferative response. Varying cell-cell contact resulted in a biphasic effect on proliferation; one contacting neighbor increased proliferation, while two or more neighboring cells partially inhibited this increase. We also observed that cell-cell contact increased the formation of actin stress fibers, and that expression of dominant negative RhoA (RhoN19) blocked the contact-mediated increase in stress fibers and proliferation. Furthermore, examination of heterotypic pairs of untreated cells in contact with RhoN19-expressing cells revealed that intracellular, but not intercellular, tension is required for the contact-mediated stimulation of proliferation. Moreover, engagement of VE-cadherin with cadherin-coated beads was sufficient to stimulate proliferation in the absence of actual cell-cell contact. In all, these results demonstrate that cell-cell contact signals through VE-cadherin, RhoA, and intracellular tension in the actin cytoskeleton to regulate proliferation

  9. Mitochondria-Associated Membranes As Networking Platforms and Regulators of Cancer Cell Fate

    Directory of Open Access Journals (Sweden)

    Maria Livia Sassano

    2017-08-01

    Full Text Available The tight cross talk between two essential organelles of the cell, the endoplasmic reticulum (ER and mitochondria, is spatially and functionally regulated by specific microdomains known as the mitochondria-associated membranes (MAMs. MAMs are hot spots of Ca2+ transfer between the ER and mitochondria, and emerging data indicate their vital role in the regulation of fundamental physiological processes, chief among them mitochondria bioenergetics, proteostasis, cell death, and autophagy. Moreover, and perhaps not surprisingly, it has become clear that signaling events regulated at the ER–mitochondria intersection regulate key processes in oncogenesis and in the response of cancer cells to therapeutics. ER–mitochondria appositions have been shown to dynamically recruit oncogenes and tumor suppressors, modulating their activity and protein complex formation, adapt the bioenergetic demand of cancer cells and to regulate cell death pathways and redox signaling in cancer cells. In this review, we discuss some emerging players of the ER–mitochondria contact sites in mammalian cells, the key processes they regulate and recent evidence highlighting the role of MAMs in shaping cell-autonomous and non-autonomous signals that regulate cancer growth.

  10. New insights into redox regulation of stem cell self-renewal and differentiation.

    Science.gov (United States)

    Ren, Fenglian; Wang, Kui; Zhang, Tao; Jiang, Jingwen; Nice, Edouard Collins; Huang, Canhua

    2015-08-01

    Reactive oxygen species (ROS), the natural byproducts of aerobic metabolism, are precisely orchestrated to evoke diverse signaling pathways. To date, studies have focused mainly on the detrimental effects of ROS in stem cells. Recently, accumulating evidence has suggested that ROS also function as second messengers that modulate stem cell self-renewal and differentiation by regulating intricate signaling networks. Although many efforts have been made to clarify the general effects of ROS on signal transduction in stem cells, less is known about the initial and direct executors of ROS signaling, which are known as 'redox sensors'. Modifications of cysteine residues in redox sensors are of significant importance in the modulation of protein function in response to different redox conditions. Intriguingly, most key molecules in ROS signaling and cell cycle regulation (including transcriptional factors and kinases) that are crucial in the regulation of stem cell self-renewal and differentiation have the potential to be redox sensors. We highlight herein the importance of redox regulation of these key regulators in stem cell self-renewal and differentiation. Understanding the mechanisms of redox regulation in stem cell self-renewal and differentiation will open exciting new perspectives for stem cell biology. This article is part of a Special Issue entitled Redox regulation of differentiation and de-differentiation. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. [Astragalus polysaccharide may increase sensitivity of cervical cancer HeLa cells to cisplatin by regulating cell autophagy].

    Science.gov (United States)

    Zhai, Qiu-Li; Hu, Xiang-Dan; Xiao, Jing; Yu, Dong-Qing

    2018-02-01

    This study aimed to investigate the possible sensitivity of Astragalus polysaccharides, in order to improve the chemosensitivity of cervical cancer HeLa cells to cisplatin by regulating the cell autophagy, and explore its possible mechanism. In this study, HeLa cells were divided into control group, cisplatin group, Astragalus polysaccharide group, and Astragalus polysaccharide combined with cisplatin group. MTT assay was used to detect the proliferation of cervical cancer HeLa cells. Flow cytometry was used to detect the apoptosis and cycle of HeLa cells in each experimental group. RT-PCR was used to detect the mRNA expression of autophagy-related proteins beclin1, LC3Ⅱ and p62. The expression levels of autophagy-related proteins beclin1, LC3Ⅱ, LC3Ⅰ and p62 were detected by WB method. MTT results showed that compared with the control group, the proliferation of HeLa cells was significantly inhibited in each administration group( P HeLa cells was significantly increased( P HeLa cells to cisplatin by regulating the cell autophagy. Its possible mechanism of action is correlated with the up-regulation of autophagy-related proteins beclin1, the promote the conversion from LC3Ⅰ to LC3Ⅱ, the down-regulation of labeled protein p62, and the enhancement of HeLa cell autophagic activity, thereby increasing the sensitivity of HeLa cells to cisplatin chemotherapy. Copyright© by the Chinese Pharmaceutical Association.

  12. Molecular machinery of signal transduction and cell cycle regulation in Plasmodium

    OpenAIRE

    Koyama, Fernanda C.; Chakrabarti, Debopam; Garcia, Célia R.S.

    2009-01-01

    The regulation of the Plasmodium cell cycle is not understood. Although the Plasmodium falciparum genome is completely sequenced, about 60% of the predicted proteins share little or no sequence similarity with other eukaryotes. This feature impairs the identification of important proteins participating in the regulation of the cell cycle. There are several open questions that concern cell cycle progression in malaria parasites, including the mechanism by which multiple nuclear divisions is co...

  13. RCAN1.4 regulates VEGFR-2 internalisation, cell polarity and migration in human microvascular endothelial cells.

    Science.gov (United States)

    Alghanem, Ahmad F; Wilkinson, Emma L; Emmett, Maxine S; Aljasir, Mohammad A; Holmes, Katherine; Rothermel, Beverley A; Simms, Victoria A; Heath, Victoria L; Cross, Michael J

    2017-08-01

    Regulator of calcineurin 1 (RCAN1) is an endogenous inhibitor of the calcineurin pathway in cells. It is expressed as two isoforms in vertebrates: RCAN1.1 is constitutively expressed in most tissues, whereas transcription of RCAN1.4 is induced by several stimuli that activate the calcineurin-NFAT pathway. RCAN1.4 is highly upregulated in response to VEGF in human endothelial cells in contrast to RCAN1.1 and is essential for efficient endothelial cell migration and tubular morphogenesis. Here, we show that RCAN1.4 has a role in the regulation of agonist-stimulated VEGFR-2 internalisation and establishment of endothelial cell polarity. siRNA-mediated gene silencing revealed that RCAN1 plays a vital role in regulating VEGF-mediated cytoskeletal reorganisation and directed cell migration and sprouting angiogenesis. Adenoviral-mediated overexpression of RCAN1.4 resulted in increased endothelial cell migration. Antisense-mediated morpholino silencing of the zebrafish RCAN1.4 orthologue revealed a disrupted vascular development further confirming a role for the RCAN1.4 isoform in regulating vascular endothelial cell physiology. Our data suggest that RCAN1.4 plays a novel role in regulating endothelial cell migration by establishing endothelial cell polarity in response to VEGF.

  14. Hydrogels with tunable stress relaxation regulate stem cell fate and activity

    Science.gov (United States)

    Chaudhuri, Ovijit; Gu, Luo; Klumpers, Darinka; Darnell, Max; Bencherif, Sidi A.; Weaver, James C.; Huebsch, Nathaniel; Lee, Hong-Pyo; Lippens, Evi; Duda, Georg N.; Mooney, David J.

    2016-03-01

    Natural extracellular matrices (ECMs) are viscoelastic and exhibit stress relaxation. However, hydrogels used as synthetic ECMs for three-dimensional (3D) culture are typically elastic. Here, we report a materials approach to tune the rate of stress relaxation of hydrogels for 3D culture, independently of the hydrogel's initial elastic modulus, degradation, and cell-adhesion-ligand density. We find that cell spreading, proliferation, and osteogenic differentiation of mesenchymal stem cells (MSCs) are all enhanced in cells cultured in gels with faster relaxation. Strikingly, MSCs form a mineralized, collagen-1-rich matrix similar to bone in rapidly relaxing hydrogels with an initial elastic modulus of 17 kPa. We also show that the effects of stress relaxation are mediated by adhesion-ligand binding, actomyosin contractility and mechanical clustering of adhesion ligands. Our findings highlight stress relaxation as a key characteristic of cell-ECM interactions and as an important design parameter of biomaterials for cell culture.

  15. Ursodeoxycholic acid inhibits the proliferation of colon cancer cells by regulating oxidative stress and cancer stem-like cell growth

    Science.gov (United States)

    Kim, EuiJoo

    2017-01-01

    Introduction The regulation of reactive oxygen species (ROS) exists as a therapeutic target for cancer treatments. Previous studies have shown that ursodeoxycholic acid (UDCA) suppresses the proliferation of colon cancer cells. The aim of this study was to evaluate the effect of UDCA upon the proliferation of colon cancer cells as a direct result of the regulation of ROS. Method Colon cancer cell lines (HT29 and HCT116) were treated with UDCA. The total number of cells and the number of dead cells were determined using cell counters. A fluorescein isothiocyanate-bromodeoxyuridine flow kit was used to analyze cell cycle variations. Upon exposure to UDCA, the protein levels of p27, p21, CDK2, CDK4 and CDK6 were determined using western blotting, and qRT-PCR was used to determine levels of mRNA. We preformed dichlorofluorescindiacetate (DCF-DA) staining to detect alteration of intracellular ROS using fluorescence activated cell sorting (FACS). Colon cancer stem-like cell lines were generated by tumorsphere culture and treated with UDCA for seven days. The total number of tumorspheres was determined using microscopy. Results We found that UDCA reduced the total number of colon cancer cells, but did not increase the number of dead cells. UDCA inhibited the G1/S and G2/M transition phases in colon cancer cells. UDCA induced expression of cell cycle inhibitors such as p27 and p21. However, it was determined that UDCA suppressed levels of CDK2, CDK4, and CDK6. UDCA regulated intracellular ROS generation in colon cancer cells, and induced activation of Erk1/2. Finally, UDCA inhibited formation of colon cancer stem-like cells. Conclusion Our results indicate that UDCA suppresses proliferation through regulation of oxidative stress in colon cancer cells, as well as colon cancer stem-like cells. PMID:28708871

  16. Sensors and signal transduction pathways in vertebrate cell volume regulation

    DEFF Research Database (Denmark)

    Hoffmann, Else K; Pedersen, Stine F

    2006-01-01

    The ability to control cell volume is fundamental for proper cell function. This review highlights recent advances in the understanding of the complex sequences of events by which acute cell volume perturbation alters the activity of osmolyte transport proteins in cells from vertebrate organisms...... will be discussed. In contrast to the simple pathway of osmosensing in yeast, cells from vertebrate organisms appear to exhibit multiple volume sensing systems, the specific mechanism(s) activated being cell type- and stimulus-dependent. Candidate sensors include integrins and growth factor receptors, while other...

  17. Cyclic Mechanical Stretch Up-regulates Hepatoma-Derived Growth Factor Expression in Cultured Rat Aortic Smooth Muscle Cells.

    Science.gov (United States)

    Kao, Ying-Hsien; Chen, Po-Han; Sun, Cheuk-Kwan; Chang, Yo-Chen; Lin, Yu-Chun; Tsai, Ming-Shian; Lee, Po-Huang; Cheng, Cheng-I

    2018-02-21

    Hepatoma-derived growth factor (HDGF) is a potent mitogen for vascular smooth muscle cells (SMCs) during embryogenesis and injury repair of vessel walls. Whether mechanical stimuli modulate HDGF expression remains unknown. This study aimed at investigating whether cyclic mechanical stretch plays a regulatory role in HDGF expression and regenerative cytokine production in aortic SMCs. A SMC cell line was grown on a silicone-based elastomer chamber with extracellular matrix coatings (either type I collagen or fibronectin) and received cyclic and uni-axial mechanical stretches with 10% deformation at frequency 1 Hz. Morphological observation showed that fibronectin coating provided better cell adhesion and spreading and that consecutive 6 hours of cyclic mechanical stretch remarkably induced reorientation and realignment of SMCs. Western blotting detection demonstrated that continuous mechanical stimuli elicited up-regulation of HDGF and PCNA, a cell proliferative marker. Signal kinetic profiling study indicated that cyclic mechanical stretch induced signaling activity in RhoA/ROCK and PI3K/Akt cascades. Kinase inhibition study further showed that blockade of PI3K activity suppressed the stretch-induced TNF-a, whereas RhoA/ROCK inhibition significantly blunted the IL-6 production and HDGF over-expression. Moreover, siRNA-mediated HDGF gene silencing significantly suppressed constitutive expression of IL-6, but not TNF-α, in SMCs. These findings support the role of HDGF in maintaining vascular expression of IL-6, which has been regarded a crucial regenerative factor for acute vascular injury. In conclusion, cyclic mechanical stretch may maintain constitutive expression of HDGF in vascular walls and be regarded an important biophysical regulator in vascular regeneration. ©2018 The Author(s).

  18. Benzyl isothiocyanate alters the gene expression with cell cycle regulation and cell death in human brain glioblastoma GBM 8401 cells.

    Science.gov (United States)

    Tang, Nou-Ying; Chueh, Fu-Shin; Yu, Chien-Chih; Liao, Ching-Lung; Lin, Jen-Jyh; Hsia, Te-Chun; Wu, King-Chuen; Liu, Hsin-Chung; Lu, Kung-Wen; Chung, Jing-Gung

    2016-04-01

    Glioblastoma multiforme (GBM) is a highly malignant devastating brain tumor in adults. Benzyl isothiocyanate (BITC) is one of the isothiocyanates that have been shown to induce human cancer cell apoptosis and cell cycle arrest. Herein, the effect of BITC on cell viability and apoptotic cell death and the genetic levels of human brain glioblastoma GBM 8401 cells in vitro were investigated. We found that BITC induced cell morphological changes, decreased cell viability and the induction of cell apoptosis in GBM 8401 cells was time-dependent. cDNA microarray was used to examine the effects of BITC on GBM 8401 cells and we found that numerous genes associated with cell death and cell cycle regulation in GBM 8401 cells were altered after BITC treatment. The results show that expression of 317 genes was upregulated, and two genes were associated with DNA damage, the DNA-damage-inducible transcript 3 (DDIT3) was increased 3.66-fold and the growth arrest and DNA-damage-inducible α (GADD45A) was increased 2.34-fold. We also found that expression of 182 genes was downregulated and two genes were associated with receptor for cell responses to stimuli, the EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1) was inhibited 2.01-fold and the TNF receptor-associated protein 1 (TRAP1) was inhibited 2.08-fold. BITC inhibited seven mitochondria ribosomal genes, the mitochondrial ribosomal protein; tumor protein D52 (MRPS28) was inhibited 2.06-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein L23 (MRPL23) decreased 2.08-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein S12 (MRPS12) decreased 2.08-fold, the mitochondria ribosomal protein L12 (MRPL12) decreased 2.25-fold and the mitochondria ribosomal protein S34 (MRPS34) was decreased 2.30-fold in GBM 8401 cells. These changes of gene expression can provide the effects of BITC on the genetic level and are

  19. Cellular volume regulation and substrate stiffness modulate the detachment dynamics of adherent cells

    Science.gov (United States)

    Yang, Yuehua; Jiang, Hongyuan

    2018-03-01

    Quantitative characterizations of cell detachment are vital for understanding the fundamental mechanisms of cell adhesion. Experiments have found that cell detachment shows strong rate dependence, which is mostly attributed to the binding-unbinding kinetics of receptor-ligand bond. However, our recent study showed that the cellular volume regulation can significantly regulate the dynamics of adherent cell and cell detachment. How this cellular volume regulation contributes to the rate dependence of cell detachment remains elusive. Here, we systematically study the role of cellular volume regulation in the rate dependence of cell detachment by investigating the cell detachments of nonspecific adhesion and specific adhesion. We find that the cellular volume regulation and the bond kinetics dominate the rate dependence of cell detachment at different time scales. We further test the validity of the traditional Johnson-Kendall-Roberts (JKR) contact model and the detachment model developed by Wyart and Gennes et al (W-G model). When the cell volume is changeable, the JKR model is not appropriate for both the detachments of convex cells and concave cells. The W-G model is valid for the detachment of convex cells but is no longer applicable for the detachment of concave cells. Finally, we show that the rupture force of adherent cells is also highly sensitive to substrate stiffness, since an increase in substrate stiffness will lead to more associated bonds. These findings can provide insight into the critical role of cell volume in cell detachment and might have profound implications for other adhesion-related physiological processes.

  20. Bovine lactoferrin regulates cell survival, apoptosis and inflammation in intestinal epithelial cells and preterm pig intestine.

    Science.gov (United States)

    Nguyen, Duc Ninh; Jiang, Pingping; Stensballe, Allan; Bendixen, Emøke; Sangild, Per T; Chatterton, Dereck E W

    2016-04-29

    Bovine lactoferrin (bLF) may modulate neonatal intestinal inflammation. Previous studies in intestinal epithelial cells (IECs) indicated that moderate bLF doses enhance proliferation whereas high doses trigger inflammation. To further elucidate cellular mechanisms, we profiled the porcine IEC proteome after stimulation with bLF at 0, 0.1, 1 and 10g/L by LC-MS-based proteomics. Key pathways were analyzed in the intestine of formula-fed preterm pigs with and without supplementation of 10g/L bLF. Levels of 123 IEC proteins were altered by bLF. Low bLF doses (0.1-1g/L) up-regulated 11 proteins associated with glycolysis, energy metabolism and protein synthesis, indicating support of cell survival. In contrast, a high bLF dose (10g/L) up-regulated three apoptosis-inducing proteins, down-regulated five anti-apoptotic and proliferation-inducing proteins and 15 proteins related to energy and amino acid metabolism, and altered three proteins enhancing the hypoxia inducible factor-1 (HIF-1) pathway. In the preterm pig intestine, bLF at 10g/L decreased villus height/crypt depth ratio and up-regulated the Bax/Bcl-2 ratio and HIF-1α, indicating elevated intestinal apoptosis and inflammation. In conclusion, bLF dose-dependently affects IECs via metabolic, apoptotic and inflammatory pathways. It is important to select an appropriate dose when feeding neonates with bLF to avoid detrimental effects exerted by excessive doses. The present work elucidates dose-dependent effects of bLF on the proteomic changes of IECs in vitro supplemented with data from a preterm pig study confirming detrimental effects of enteral feeding with the highest dose of bLF (10g/L). The study contributes to further understanding on mechanisms that bLF, as an important milk protein, can regulate the homeostasis of the immature intestine. Results from this study urge neonatologists to carefully consider the dose of bLF to supplement into infant formula used for preterm neonates. Copyright © 2016 Elsevier B

  1. Cholinergic regulation of VIP gene expression in human neuroblastoma cells

    DEFF Research Database (Denmark)

    Kristensen, Bo; Georg, Birgitte; Fahrenkrug, Jan

    1997-01-01

    Vasoactive intestinal polypeptide, muscarinic receptor, neuroblastoma cell, mRNA, gene expression, peptide processing......Vasoactive intestinal polypeptide, muscarinic receptor, neuroblastoma cell, mRNA, gene expression, peptide processing...

  2. Epigenetic Regulation of Epidermal Stem Cell Biomarkers and Their Role in Wound Healing

    Directory of Open Access Journals (Sweden)

    Sabita N. Saldanha

    2015-12-01

    Full Text Available As an actively renewable tissue, changes in skin architecture are subjected to the regulation of stem cells that maintain the population of cells responsible for the formation of epidermal layers. Stems cells retain their self-renewal property and express biomarkers that are unique to this population. However, differential regulation of the biomarkers can initiate the pathway of terminal cell differentiation. Although, pockets of non-clarity in stem cell maintenance and differentiation in skin still exist, the influence of epigenetics in epidermal stem cell functions and differentiation in skin homeostasis and wound healing is clearly evident. The focus of this review is to discuss the epigenetic regulation of confirmed and probable epidermal stem cell biomarkers in epidermal stratification of normal skin and in diseased states. The role of epigenetics in wound healing, especially in diseased states of diabetes and cancer, will also be conveyed.

  3. Classification of Hydrogels Based on Their Source: A Review and Application in Stem Cell Regulation

    Science.gov (United States)

    Khansari, Maziyar M.; Sorokina, Lioudmila V.; Mukherjee, Prithviraj; Mukhtar, Farrukh; Shirdar, Mostafa Rezazadeh; Shahidi, Mahnaz; Shokuhfar, Tolou

    2017-08-01

    Stem cells are recognized by their self-renewal ability and can give rise to specialized progeny. Hydrogels are an established class of biomaterials with the ability to control stem cell fate via mechanotransduction. They can mimic various physiological conditions to influence the fate of stem cells and are an ideal platform to support stem cell regulation. This review article provides a summary of recent advances in the application of different classes of hydrogels based on their source (e.g., natural, synthetic, or hybrid). This classification is important because the chemistry of substrate affects stem cell differentiation and proliferation. Natural and synthetic hydrogels have been widely used in stem cell regulation. Nevertheless, they have limitations that necessitate a new class of material. Hybrid hydrogels obtained by manipulation of the natural and synthetic ones can potentially overcome these limitations and shape the future of research in application of hydrogels in stem cell regulation.

  4. Down-regulation of mTOR leads to up-regulation of osteoprotegerin in bone marrow cells

    International Nuclear Information System (INIS)

    Mogi, Makio; Kondo, Ayami

    2009-01-01

    Osteoprotegerin (OPG)/osteoclastogenesis inhibitory factor regulates bone mass by inhibiting osteoclastic bone resorption. mTOR, which is the mammalian target of rapamycin, is a kinase and central regulator of cell growth, proliferation, and survival. By using Rapamycin, we studied whether mTOR pathway is associated with OPG protein production in the mouse bone marrow-derived stromal cell line ST2. Rapamycin markedly increased the level of soluble OPG in ST2 cells. This antibiotic treatment resulted in the suppression of phosphorylation of mTOR. Rapamycin had no effects on the proliferation, differentiation, or apoptosis of the cells. Treatment with bone morphogenetic protein-4, which can induce OPG protein in ST2 cells, also resulted in a decrease in the density of the phospho-mTOR-band, suggesting that the suppression of the phospho-mTOR pathway is necessary for OPG production in ST2 cells. Thus, suitable suppression of mTOR phosphorylation is a necessary requirement for OPG production in bone marrow stromal cells.

  5. Lycopene inhibits regulator of calcineurin 1-mediated apoptosis by reducing oxidative stress and down-regulating Nucling in neuronal cells.

    Science.gov (United States)

    Lim, Seiyoung; Hwang, Sinwoo; Yu, Ji Hoon; Lim, Joo Weon; Kim, Hyeyoung

    2017-05-01

    Regulator of calcineurin 1 (RCAN1) is located on the Down syndrome critical region (DSCR) locus in human chromosome 21. Oxidative stress and overexpression of RCAN1 are implicated in neuronal impairment in Down's syndrome (DS) and Alzheimer's disease (AD). Serum level of lycopene, an antioxidant pigment, is low in DS and AD patients, which may be related to neuronal damage. The present study is to investigate whether lycopene inhibits apoptosis by reducing ROS levels, NF-κB activation, expression of the apoptosis regulator Nucling, cell viability, and indices of apoptosis (cytochrome c release, caspase-3 activation) in RCAN1-overexpressing neuronal cells. Cells transfected with either pcDNA or RCAN1 were treated with or without lycopene. Lycopene decreased intracellular and mitochondrial ROS levels, NF-κB activity, and Nucling expression while it reversed decrease in mitochondrial membrane potential, mitochondrial respiration, and glycolytic function in RCAN1-overexpressing cells. Lycopene inhibited cell death, DNA fragmentation, caspase-3 activation, and cytochrome c release in RCAN1-overexpressing cells. Lycopene inhibits RCAN1-mediated apoptosis by reducing ROS levels and by inhibiting NF-κB activation, Nucling induction, and the increase in apoptotic indices in neuronal cells. Consumption of lycopene-rich foods may prevent oxidative stress-associated neuronal damage in some pathologic conditions such as DS or AD. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Genome Binding and Gene Regulation by Stem Cell Transcription Factors

    NARCIS (Netherlands)

    J.H. Brandsma (Johan)

    2016-01-01

    markdownabstractNearly all cells of an individual organism contain the same genome. However, each cell type transcribes a different set of genes due to the presence of di