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Sample records for cell surface co-receptor

  1. Functional Mimetics of the HIV-1 CCR5 Co-Receptor Displayed on the Surface of Magnetic Liposomes.

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    Kuzmina, Alona; Vaknin, Karin; Gdalevsky, Garik; Vyazmensky, Maria; Marks, Robert S; Taube, Ran; Engel, Stanislav

    2015-01-01

    Chemokine G protein coupled receptors, principally CCR5 or CXCR4, function as co-receptors for HIV-1 entry into CD4+ T cells. Initial binding of the viral envelope glycoprotein (Env) gp120 subunit to the host CD4 receptor induces a cascade of structural conformational changes that lead to the formation of a high-affinity co-receptor-binding site on gp120. Interaction between gp120 and the co-receptor leads to the exposure of epitopes on the viral gp41 that mediates fusion between viral and cell membranes. Soluble CD4 (sCD4) mimetics can act as an activation-based inhibitor of HIV-1 entry in vitro, as it induces similar structural changes in gp120, leading to increased virus infectivity in the short term but to virus Env inactivation in the long term. Despite promising clinical implications, sCD4 displays low efficiency in vivo, and in multiple HIV strains, it does not inhibit viral infection. This has been attributed to the slow kinetics of the sCD4-induced HIV Env inactivation and to the failure to obtain sufficient sCD4 mimetic levels in the serum. Here we present uniquely structured CCR5 co-receptor mimetics. We hypothesized that such mimetics will enhance sCD4-induced HIV Env inactivation and inhibition of HIV entry. Co-receptor mimetics were derived from CCR5 gp120-binding epitopes and functionalized with a palmitoyl group, which mediated their display on the surface of lipid-coated magnetic beads. CCR5-peptidoliposome mimetics bound to soluble gp120 and inhibited HIV-1 infectivity in a sCD4-dependent manner. We concluded that CCR5-peptidoliposomes increase the efficiency of sCD4 to inhibit HIV infection by acting as bait for sCD4-primed virus, catalyzing the premature discharge of its fusion potential.

  2. Characterization of a novel single-chain bispecific antibody for retargeting of T cells to tumor cells via the TCR co-receptor CD8.

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    Irene Michalk

    Full Text Available There is currently growing interest in retargeting of effector T cells to tumor cells via bispecific antibodies (bsAbs. Usually, bsAbs are directed on the one hand to the CD3 complex of T cells and on the other hand to a molecule expressed on the surface of the target cell. A bsAb-mediated cross-linkage via CD3 leads to an activation of CD8+ T cells and consequently to killing of the target cells. In parallel, CD4+ T cells including TH1, TH2, TH17 cells and even regulatory T cells (Tregs will be activated as well. Cytokines produced by CD4+ T cells can contribute to severe side effects e. g. life-threatening cytokine storms and, thinking of the immunosupressive function of Tregs, can even be counterproductive. Therefore, we asked whether or not it is feasible to limit retargeting to CD8+ T cells e. g. via targeting of the co-receptor CD8 instead of CD3. In order to test for proof of concept, a novel bsAb with specificity for CD8 and a tumor-associated surface antigen was constructed. Interestingly, we found that pre-activated (but not freshly isolated CD8+ T cells can be retargeted via CD8-engaging bsAbs leading to an efficient lysis of target cells.

  3. Genetic interactions between the hedgehog co-receptors Gas1 and Boc regulate cell proliferation during murine palatogenesis.

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    Xavier, Guilherme M; Seppala, Maisa; Papageorgiou, Spyridon N; Fan, Chen-Ming; Cobourne, Martyn T

    2016-11-29

    Abnormal regulation of Sonic hedgehog (Shh) signaling has been described in a variety of human cancers and developmental anomalies, which highlights the essential role of this signaling molecule in cell cycle regulation and embryonic development. Gas1 and Boc are membrane co-receptors for Shh, which demonstrate overlapping domains of expression in the early face. This study aims to investigate potential interactions between these co-receptors during formation of the secondary palate. Mice with targeted mutation in Gas1 and Boc were used to generate Gas1; Boc compound mutants. The expression of key Hedgehog signaling family members was examined in detail during palatogenesis via radioactive in situ hybridization. Morphometric analysis involved computational quantification of BrdU-labeling and cell packing; whilst TUNEL staining was used to assay cell death. Ablation of Boc in a Gas1 mutant background leads to reduced Shh activity in the palatal shelves and an increase in the penetrance and severity of cleft palate, associated with failed elevation, increased proliferation and reduced cell death. Our findings suggest a dual requirement for Boc and Gas1 during early development of the palate, mediating cell cycle regulation during growth and subsequent fusion of the palatal shelves.

  4. The Wnt Co-Receptor Lrp5 Is Required for Cranial Neural Crest Cell Migration in Zebrafish.

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    Bernd Willems

    Full Text Available During vertebrate neurulation, cranial neural crest cells (CNCCs undergo epithelial to mesenchymal transition (EMT, delaminate from the neural plate border, and migrate as separate streams into different cranial regions. There, they differentiate into distinct parts of the craniofacial skeleton. Canonical Wnt signaling has been shown to be essential for this process at different levels but the involved receptors remained unclear. Here we show that the frizzled co-receptor low-density-lipoprotein (LDL receptor-related protein 5 (Lrp5 plays a crucial role in CNCC migration and morphogenesis of the cranial skeleton. Early during induction and migration of CNCCs, lrp5 is expressed ubiquitously but later gets restricted to CNCC derivatives in the ventral head region besides different regions in the CNS. A knock-down of lrp5 does not interfere with induction of CNCCs but leads to reduced proliferation of premigratory CNCCs. In addition, cell migration is disrupted as CNCCs are found in clusters at ectopic positions in the dorsomedial neuroepithelium after lrp5 knock-down and transient CRISPR/Cas9 gene editing. These migratory defects consequently result in malformations of the craniofacial skeleton. To date, Lrp5 has mainly been associated with bone homeostasis in mammals. Here we show that in zebrafish, lrp5 also controls cell migration during early morphogenetic processes and contributes to shaping the craniofacial skeleton.

  5. Gene number determination and genetic polymorphism of the gamma delta T cell co-receptor WC1 genes

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    Chen Chuang

    2012-10-01

    Full Text Available Abstract Background WC1 co-receptors belong to the scavenger receptor cysteine-rich (SRCR superfamily and are encoded by a multi-gene family. Expression of particular WC1 genes defines functional subpopulations of WC1+ γδ T cells. We have previously identified partial or complete genomic sequences for thirteen different WC1 genes through annotation of the bovine genome Btau_3.1 build. We also identified two WC1 cDNA sequences from other cattle that did not correspond to sequences in the Btau_3.1 build. Their absence in the Btau_3.1 build may have reflected gaps in the genome assembly or polymorphisms among animals. Since the response of γδ T cells to bacterial challenge is determined by WC1 gene expression, it was critical to understand whether individual cattle or breeds differ in the number of WC1 genes or display polymorphisms. Results Real-time quantitative PCR using DNA from the animal whose genome was sequenced (“Dominette” and sixteen other animals representing ten breeds of cattle, showed that the number of genes coding for WC1 co-receptors is thirteen. The complete coding sequences of those thirteen WC1 genes is presented, including the correction of an error in the WC1-2 gene due to mis-assembly in the Btau_3.1 build. All other cDNA sequences were found to agree with the previous annotation of complete or partial WC1 genes. PCR amplification and sequencing of the most variable N-terminal SRCR domain (domain 1 which has the SRCR “a” pattern of each of the thirteen WC1 genes showed that the sequences are highly conserved among individuals and breeds. Of 160 sequences of domain 1 from three breeds of cattle, no additional sequences beyond the thirteen described WC1 genes were found. Analysis of the complete WC1 cDNA sequences indicated that the thirteen WC1 genes code for three distinct WC1 molecular forms. Conclusion The bovine WC1 multi-gene family is composed of thirteen genes coding for three structural forms whose

  6. Toll-like receptor 3 signalling up-regulates expression of the HIV co-receptor G-protein coupled receptor 15 on human CD4+ T cells.

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    Miriam Kiene

    Full Text Available BACKGROUND: Many HIV-2 and SIV isolates, as well as some HIV-1 strains, can use the orphan 7-transmembrane receptor GPR15 as co-receptor for efficient entry into host cells. GPR15 is expressed on central memory and effector memory CD4(+ T cells in healthy individuals and a subset of these cells is susceptible to HIV-1 and SIV infection. However, it has not been determined whether GPR15 expression is altered in the context of HIV-1 infection. RESULTS: Here, we show that GPR15 expression in CD4(+ T cells is markedly up-regulated in some HIV-1 infected individuals compared to the rest of the infected patients and to healthy controls. Infection of the PM1 T cell line with primary HIV-1 isolates was found to up-regulate GPR15 expression on the infected cells, indicating that viral components can induce GPR15 expression. Up-regulation of GPR15 expression on CD4(+ T cells was induced by activation of Toll-like receptor 3 signalling via TIR-domain-containing adapter-inducing interferon-β (TRIF and was more prominent on gut-homing compared to lymph node-homing CD4(+ T cells. CONCLUSION: These results suggest that infection-induced up-regulation of GPR15 expression could increase susceptibility of CD4(+ T cells to HIV infection and target cell availability in the gut in some infected individuals.

  7. HIV-1 replication in CD4+ T cell lines: the effects of adaptation on co-receptor use, tropism, and accessory gene function.

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    Dejucq, N

    2000-09-01

    We studied the replication of HIV-1 macrophage-tropic CCR5-using strains (R5) in CD4+ T cell lines to better understand the switch in co-receptor use of such strains during disease progression and to assess resulting changes in cell tropism. We found that the majority of R5 strains cannot replicate in CD4+ T cell lines without adaptation by serial passage. A small minority of primary R5 isolates, however, were able to infect two T cell lines, Molt4 and SupT1. This expanded tropism was due to the use of undetectable levels of CCR5 rather than CXCR4 or alternative receptors. In contrast, HIV-1sF162 adaptation for replication in the C8166 T cell line was due to the emergence of variant strains that could use CXCR4. Of two variants, one was dual-tropic and one T-tropic, although both could use CCR5 as well as CXCR4. A single mutation in the start codon of the accessory gene vpu accounted for the T-tropic phenotype of the second variant, indicating that a non-functional vpu impairs macrophage tropism. Thus, in vitro and in the absence of an immune response, R5 strains naturally adapt to infect CXCR4+ T cell lines. Such adaptation resembles the rare R5 to X4 switch that occurs in vivo. Mutations in accessory genes (e.g., vpu) not required for replication in rapidly dividing cell lines may also occur in vitro, abrogating replication in primary cell types such as macrophages. Such mutations, however, are normally selected against in vivo.

  8. Genome editing of the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4+T cells from HIV-1 infection.

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    Liu, Zhepeng; Chen, Shuliang; Jin, Xu; Wang, Qiankun; Yang, Kongxiang; Li, Chenlin; Xiao, Qiaoqiao; Hou, Panpan; Liu, Shuai; Wu, Shaoshuai; Hou, Wei; Xiong, Yong; Kong, Chunyan; Zhao, Xixian; Wu, Li; Li, Chunmei; Sun, Guihong; Guo, Deyin

    2017-01-01

    The main approach to treat HIV-1 infection is combination antiretroviral therapy (cART). Although cART is effective in reducing HIV-1 viral load and controlling disease progression, it has many side effects, and is expensive for HIV-1 infected patients who must remain on lifetime treatment. HIV-1 gene therapy has drawn much attention as studies of genome editing tools have progressed. For example, zinc finger nucleases (ZFN), transcription activator like effector nucleases (TALEN) and clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 have been utilized to successfully disrupt the HIV-1 co-receptors CCR5 or CXCR4, thereby restricting HIV-1 infection. However, the effects of simultaneous genome editing of CXCR4 and CCR5 by CRISPR-Cas9 in blocking HIV-1 infection in primary CD4 + T cells has been rarely reported. Furthermore, combination of different target sites of CXCR4 and CCR5 for disruption also need investigation. In this report, we designed two different gRNA combinations targeting both CXCR4 and CCR5, in a single vector. The CRISPR-sgRNAs-Cas9 could successfully induce editing of CXCR4 and CCR5 genes in various cell lines and primary CD4 + T cells. Using HIV-1 challenge assays, we demonstrated that CXCR4-tropic or CCR5-tropic HIV-1 infections were significantly reduced in CXCR4 - and CCR5 -modified cells, and the modified cells exhibited a selective advantage over unmodified cells during HIV-1 infection. The off-target analysis showed that no non-specific editing was identified in all predicted sites. In addition, apoptosis assays indicated that simultaneous disruption of CXCR4 and CCR5 in primary CD4 + T cells by CRISPR-Cas9 had no obvious cytotoxic effects on cell viability. Our results suggest that simultaneous genome editing of CXCR4 and CCR5 by CRISPR-Cas9 can potentially provide an effective and safe strategy towards a functional cure for HIV-1 infection.

  9. A Cinnamon-Derived Procyanidin Compound Displays Anti-HIV-1 Activity by Blocking Heparan Sulfate- and Co-Receptor- Binding Sites on gp120 and Reverses T Cell Exhaustion via Impeding Tim-3 and PD-1 Upregulation.

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    Bridgette Janine Connell

    Full Text Available Amongst the many strategies aiming at inhibiting HIV-1 infection, blocking viral entry has been recently recognized as a very promising approach. Using diverse in vitro models and a broad range of HIV-1 primary patient isolates, we report here that IND02, a type A procyanidin polyphenol extracted from cinnamon, that features trimeric and pentameric forms displays an anti-HIV-1 activity against CXCR4 and CCR5 viruses with 1-7 μM ED50 for the trimer. Competition experiments, using a surface plasmon resonance-based binding assay, revealed that IND02 inhibited envelope binding to CD4 and heparan sulphate (HS as well as to an antibody (mAb 17b directed against the gp120 co-receptor binding site with an IC50 in the low μM range. IND02 has thus the remarkable property of simultaneously blocking gp120 binding to its major host cell surface counterparts. Additionally, the IND02-trimer impeded up-regulation of the inhibitory receptors Tim-3 and PD-1 on CD4+ and CD8+ cells, thereby demonstrating its beneficial effect by limiting T cell exhaustion. Among naturally derived products significantly inhibiting HIV-1, the IND02-trimer is the first component demonstrating an entry inhibition property through binding to the viral envelope glycoprotein. These data suggest that cinnamon, a widely consumed spice, could represent a novel and promising candidate for a cost-effective, natural entry inhibitor for HIV-1 which can also down-modulate T cell exhaustion markers Tim-3 and PD-1.

  10. Normalized Synergy Predicts That CD8 Co-Receptor Contribution to T Cell Receptor (TCR and pMHC Binding Decreases As TCR Affinity Increases in Human Viral-Specific T Cells

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    Chad M. Williams

    2017-07-01

    Full Text Available The discovery of naturally occurring T cell receptors (TCRs that confer specific, high-affinity recognition of pathogen and cancer-associated antigens remains a major goal in cellular immunotherapies. The contribution of the CD8 co-receptor to the interaction between the TCR and peptide-bound major histocompatibility complex (pMHC has previously been correlated with the activation and responsiveness of CD8+ T cells. However, these studies have been limited to model systems of genetically engineered hybridoma TCRs or transgenic mouse TCRs against either a single epitope or an array of altered peptide ligands. CD8 contribution in a native human antigen-specific T cell response remains elusive. Here, using Hepatitis C Virus-specific precursor CTLs spanning a large range of TCR affinities, we discovered that the functional responsiveness of any given TCR correlated with the contribution of CD8 to TCR/pMHC binding. Furthermore, we found that CD8 contribution to TCR/pMHC binding in the two-dimensional (2D system was more accurately reflected by normalized synergy (CD8 cooperation normalized by total TCR/pMHC bonds rather than synergy (total CD8 cooperation alone. While synergy showed an increasing trend with TCR affinity, normalized synergy was demonstrated to decrease with the increase of TCR affinity. Critically, normalized synergy was shown to correlate with CTL functionality and peptide sensitivity, corroborating three-dimensional (3D analysis of CD8 contribution with respect to TCR affinity. In addition, we identified TCRs that were independent of CD8 for TCR/pMHC binding. Our results resolve the current discrepancy between 2D and 3D analysis on CD8 contribution to TCR/pMHC binding, and demonstrate that naturally occurring high-affinity TCRs are more capable of CD8-independent interactions that yield greater functional responsiveness even with CD8 blocking. Taken together, our data suggest that addition of the normalized synergy parameter to our

  11. Normalized Synergy Predicts That CD8 Co-Receptor Contribution to T Cell Receptor (TCR) and pMHC Binding Decreases As TCR Affinity Increases in Human Viral-Specific T Cells.

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    Williams, Chad M; Schonnesen, Alexandra A; Zhang, Shu-Qi; Ma, Ke-Yue; He, Chenfeng; Yamamoto, Tori; Eckhardt, S Gail; Klebanoff, Christopher A; Jiang, Ning

    2017-01-01

    The discovery of naturally occurring T cell receptors (TCRs) that confer specific, high-affinity recognition of pathogen and cancer-associated antigens remains a major goal in cellular immunotherapies. The contribution of the CD8 co-receptor to the interaction between the TCR and peptide-bound major histocompatibility complex (pMHC) has previously been correlated with the activation and responsiveness of CD8 + T cells. However, these studies have been limited to model systems of genetically engineered hybridoma TCRs or transgenic mouse TCRs against either a single epitope or an array of altered peptide ligands. CD8 contribution in a native human antigen-specific T cell response remains elusive. Here, using Hepatitis C Virus-specific precursor CTLs spanning a large range of TCR affinities, we discovered that the functional responsiveness of any given TCR correlated with the contribution of CD8 to TCR/pMHC binding. Furthermore, we found that CD8 contribution to TCR/pMHC binding in the two-dimensional (2D) system was more accurately reflected by normalized synergy (CD8 cooperation normalized by total TCR/pMHC bonds) rather than synergy (total CD8 cooperation) alone. While synergy showed an increasing trend with TCR affinity, normalized synergy was demonstrated to decrease with the increase of TCR affinity. Critically, normalized synergy was shown to correlate with CTL functionality and peptide sensitivity, corroborating three-dimensional (3D) analysis of CD8 contribution with respect to TCR affinity. In addition, we identified TCRs that were independent of CD8 for TCR/pMHC binding. Our results resolve the current discrepancy between 2D and 3D analysis on CD8 contribution to TCR/pMHC binding, and demonstrate that naturally occurring high-affinity TCRs are more capable of CD8-independent interactions that yield greater functional responsiveness even with CD8 blocking. Taken together, our data suggest that addition of the normalized synergy parameter to our previously

  12. Overlapping roles and collective requirement for the co-receptors Gas1, Cdo and Boc in Shh pathway function

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    Allen, Benjamin L.; Song, Jane Y.; Izzi, Luisa; Althaus, Irene W.; Kang, Jong-Sun; Charron, Frédéric; Krauss, Robert S.; McMahon, Andrew P.

    2011-01-01

    Summary Secreted Hedgehog (Hh) ligands signal through the canonical receptor Patched (Ptch1). However, recent studies implicate three additional Hh-binding, cell surface proteins, Gas1, Cdo and Boc, as putative co-receptors for Hh ligands. A central question is to what degree these co-receptors function similarly and their collective requirement in Hh signal transduction. Here we provide evidence that Gas1, Cdo, and Boc, play overlapping and essential roles during Hh-mediated ventral neural patterning of the mammalian neural tube. Specifically, we demonstrate two important roles for these molecules: an early role in cell fate specification of multiple neural progenitors, and a later role in motor neuron progenitor maintenance. Most strikingly, genetic loss-of-function experiments indicate an obligatory requirement for Gas1, Cdo and Boc in Hh pathway activity in multiple tissues. PMID:21664576

  13. Co-receptor choice by V alpha14i NKT cells is driven by Th-POK expression rather than avoidance of CD8-mediated negative selection.

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    Engel, Isaac; Hammond, Kirsten; Sullivan, Barbara A; He, Xi; Taniuchi, Ichiro; Kappes, Dietmar; Kronenberg, Mitchell

    2010-05-10

    Mouse natural killer T (NKT) cells with an invariant V alpha14-J alpha18 rearrangement (V alpha14 invariant [V alpha14i] NKT cells) are either CD4(+)CD8(-) or CD4(-)CD8(-). Because transgenic mice with forced CD8 expression in all T cells exhibited a profound NKT cell deficit, the absence of CD8 has been attributed to negative selection. We now present evidence that CD8 does not serve as a coreceptor for CD1d recognition and that the defect in development in CD8 transgene homozygous mice is the result of a reduction in secondary T cell receptor alpha rearrangements. Thymocytes from mice hemizygous for the CD8 transgene have a less severe rearrangement defect and have functional CD8(+) V alpha14i NKT cells. Furthermore, we demonstrate that the transcription factor Th, Poxviruses and Zinc finger, and Krüppel family (Th-POK) is expressed by V alpha14i NKT cells throughout their differentiation and is necessary both to silence CD8 expression and for the functional maturity of V alpha14i NKT cells. We therefore suggest that Th-POK expression is required for the normal development of V alpha14i NKT cells and that the absence of CD8 expression by these cells is a by-product of such expression, as opposed to the result of negative selection of CD8-expressing V alpha14i NKT cells.

  14. Evolution of the CD163 family and its relationship to the bovine gamma delta T cell co-receptor WC1

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    Baldwin Cynthia L

    2010-06-01

    Full Text Available Abstract Background The scavenger receptor cysteine rich (SRCR domain is an ancient and conserved protein domain. CD163 and WC1 molecules are classed together as group B SRCR superfamily members, along with Spα, CD5 and CD6, all of which are expressed by immune system cells. There are three known types of CD163 molecules in mammals, CD163A (M130, coded for by CD163, CD163b (M160, coded for by CD163L1 and CD163c-α (CD163L1 or SCART, while their nearest relative, WC1, is encoded by a multigene family so far identified in the artiodactyl species of cattle, sheep, and pigs. Results We annotated the bovine genome and identified genes coding for bovine CD163A and CD163c-α but found no evidence for CD163b. Bovine CD163A is widely expressed in immune cells, whereas CD163c-α transcripts are enriched in the WC1+ γδ T cell population. Phylogenetic analyses of the CD163 family genes and WC1 showed that CD163c-α is most closely related to WC1 and that chicken and platypus have WC1 orthologous genes, previously classified as among their CD163 genes. Conclusion Since it has been shown that WC1 plays an important role in the regulation of γδ T cell responses in cattle, which, like chickens, have a high percentage of γδ T cells in their peripheral blood, CD163c-α may play a similar role, especially in species lacking WC1 genes. Our results suggest that gene duplications resulted in the expansion of CD163c-α-like and WC1-like molecules. This expanded repertoire was retained by species known as "γδ T cell high", but homologous SRCR molecules were maintained by all mammals.

  15. Glycoprotein on cell surfaces

    International Nuclear Information System (INIS)

    Muramatsu, T.

    1975-01-01

    There are conjugated polysaccharides in cell membranes and outside of animal cells, and they play important role in the control of cell behavior. In this paper, the studies on the glycoprotein on cell surfaces are reported. It was found that the glycoprotein on cell surfaces have both N-glycoside type and O-glycoside type saccharic chains. Therefore it can be concluded that the basic structure of the saccharic chains in the glycoprotein on cell surfaces is similar to that of blood serum and body fluid. The main glycoprotein in the membranes of red blood corpuscles has been studied most in detail, and it also has both types of saccharic chains. The glycoprotein in liver cell membranes was found to have only the saccharic chains of acid type and to be in different pattern from that in endoplasmic reticula and nuclear membranes, which also has the saccharic chains of neutral type. The structure of the saccharic chains of H-2 antigen, i.e. the peculiar glycoprotein on the surfaces of lymph system cells, has been studied, and it is similar to the saccharic chains of glycoprotein in blood serum. The saccharic chain structures of H-2 antigen and TL antigen are different. TL, H-2 (D), Lna and H-2 (K) are the glycoprotein on cell surfaces, and are independent molecules. The analysis of the saccharic chain patterns on cell surfaces was carried out, and it was shown that the acid type saccharic chains were similar to those of ordinary glycoprotein, because the enzyme of pneumococci hydrolyzed most of the acid type saccharic chains. The change of the saccharic chain patterns of glycoprotein on cell surfaces owing to canceration and multiplication is complex matter. (Kako, I.)

  16. "Cytoplasmic domain effects on exposure of co-receptor-binding sites of HIV-1 Env".

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    Vzorov, Andrei N; Compans, Richard W

    2016-11-01

    We defined the effects of the cytoplasmic domain (CT) of the Env glycoprotein on co-receptor usage of HIV-1 by reciprocal exchanges of regions containing V3-V5 loops between CD4-dependent and CD4-independent isolates. Primary HIV-1 isolate Env clones CD8 CXCR4-tropic 92UG046 CT84 with an 84-aa truncated CT domain, CD4 CXCR4-tropic 92UG046, and CD4 CCR5-tropic SF162 with full-length (FL) CT domains were used for comparison. The parental 92UG046 Env with CT84 was not fusogenic, but a chimeric SF162 V3-V5-CT84 with an 84-aa truncated CT domain, which demonstrated a switched co-receptor specificity, exhibited syncytium-formation activity with 3T3T4X4 cells. The wild-type (WT) SF162 Env with CT84 or full-length CT was fusogenic in 3T3T4R5 cells. By exchange of V3-V5 loops, we were able to alter WT SF162 to switch its co-receptor preference, which was not dependent on CT domain length. These results provide evidence that CT domains can induce conformational changes in functional regions of gp120 and determine receptor tropism but do not modulate HIV-1 co-receptor specificity.

  17. The isolation of novel phage display-derived human recombinant antibodies against CCR5, the major co-receptor of HIV.

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    Shimoni, Moria; Herschhorn, Alon; Britan-Rosich, Yelena; Kotler, Moshe; Benhar, Itai; Hizi, Amnon

    2013-08-01

    Selecting for antibodies against specific cell-surface proteins is a difficult task due to many unrelated proteins that are expressed on the cell surface. Here, we describe a method to screen antibody-presenting phage libraries against native cell-surface proteins. We applied this method to isolate antibodies that selectively recognize CCR5, which is the major co-receptor for HIV entry (consequently, playing a pivotal role in HIV transmission and pathogenesis). We employed a phage screening strategy by using cells that co-express GFP and CCR5, along with an excess of control cells that do not express these proteins (and are otherwise identical to the CCR5-expressing cells). These control cells are intended to remove most of the phages that bind the cells nonspecifically; thus leading to an enrichment of the phages presenting anti-CCR5-specific antibodies. Subsequently, the CCR5-presenting cells were quantitatively sorted by flow cytometry, and the bound phages were eluted, amplified, and used for further successive selection rounds. Several different clones of human single-chain Fv antibodies that interact with CCR5-expressing cells were identified. The most specific monoclonal antibody was converted to a full-length IgG and bound the second extracellular loop of CCR5. The experimental approach presented herein for screening for CCR5-specific antibodies can be applicable to screen antibody-presenting phage libraries against any cell-surface expressed protein of interest.

  18. The Isolation of Novel Phage Display-Derived Human Recombinant Antibodies Against CCR5, the Major Co-Receptor of HIV

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    Shimoni, Moria; Herschhorn, Alon; Britan-Rosich, Yelena; Kotler, Moshe; Benhar, Itai

    2013-01-01

    Abstract Selecting for antibodies against specific cell-surface proteins is a difficult task due to many unrelated proteins that are expressed on the cell surface. Here, we describe a method to screen antibody-presenting phage libraries against native cell-surface proteins. We applied this method to isolate antibodies that selectively recognize CCR5, which is the major co-receptor for HIV entry (consequently, playing a pivotal role in HIV transmission and pathogenesis). We employed a phage screening strategy by using cells that co-express GFP and CCR5, along with an excess of control cells that do not express these proteins (and are otherwise identical to the CCR5-expressing cells). These control cells are intended to remove most of the phages that bind the cells nonspecifically; thus leading to an enrichment of the phages presenting anti-CCR5-specific antibodies. Subsequently, the CCR5-presenting cells were quantitatively sorted by flow cytometry, and the bound phages were eluted, amplified, and used for further successive selection rounds. Several different clones of human single-chain Fv antibodies that interact with CCR5-expressing cells were identified. The most specific monoclonal antibody was converted to a full-length IgG and bound the second extracellular loop of CCR5. The experimental approach presented herein for screening for CCR5-specific antibodies can be applicable to screen antibody-presenting phage libraries against any cell-surface expressed protein of interest. PMID:23941674

  19. HIV-1 envelope, integrins and co-receptor use in mucosal transmission of HIV

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    Cicala Claudia

    2010-01-01

    Full Text Available Abstract It is well established that HIV-1 infection typically involves an interaction between the viral envelope protein gp120/41 and the CD4 molecule followed by a second interaction with a chemokine receptor, usually CCR5 or CXCR4. In the early stages of an HIV-1 infection CCR5 using viruses (R5 viruses predominate. In some viral subtypes there is a propensity to switch to CXCR4 usage (X4 viruses. The receptor switch occurs in ~ 40% of the infected individuals and is associated with faster disease progression. This holds for subtypes B and D, but occurs less frequently in subtypes A and C. There are several hypotheses to explain the preferential transmission of R5 viruses and the mechanisms that lead to switching of co-receptor usage; however, there is no definitive explanation for either. One important consideration regarding transmission is that signaling by R5 gp120 may facilitate transmission of R5 viruses by inducing a permissive environment for HIV replication. In the case of sexual transmission, infection by HIV requires the virus to breach the mucosal barrier to gain access to the immune cell targets that it infects; however, the immediate events that follow HIV exposure at genital mucosal sites are not well understood. Upon transmission, the HIV quasispecies that is replicating in an infected donor contracts through a “genetic bottleneck”, and often infection results from a single infectious event. Many details surrounding this initial infection remain unresolved. In mucosal tissues, CD4+ T cells express high levels of CCR5, and a subset of these CD4+/CCR5high cells express the integrin α4β7, the gut homing receptor. CD4+/CCR5high/ α4β7high T cells are highly susceptible to infection by HIV-1 and are ideal targets for an efficient productive infection at the point of transmission. In this context we have demonstrated that the HIV-1 envelope protein gp120 binds to α4β7 on CD4+ T cells. On CD4+/CCR5high/ α4β7high T cells,

  20. Cell surface engineering to control cellular interactions

    OpenAIRE

    Custódio, Catarina A.; Mano, João F.

    2016-01-01

    Cell surface composition determines all interactions of the cell with its environment, thus cell functions such as adhesion, migration and cell–cell interactions can potentially be controlled by engineering and manipulating the cell membrane. Cell membranes present a rich repertoire of molecules, therefore a versatile ground for modification. However the complex and dynamic nature of the cell surface is also a major challenge for cell surface engineering that should also involve strategies co...

  1. Analysis of cell surface antigens by Surface Plasmon Resonance imaging

    NARCIS (Netherlands)

    Stojanovic, Ivan; Schasfoort, Richardus B.M.; Terstappen, Leonardus Wendelinus Mathias Marie

    2013-01-01

    Surface Plasmon Resonance (SPR) is most commonly used to measure bio-molecular interactions. SPR is used significantly less frequent for measuring whole cell interactions. Here we introduce a method to measure whole cells label free using the specific binding of cell surface antigens expressed on

  2. Metastasis-associated cell surface oncoproteomics

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    Piia-Riitta eKarhemo

    2012-11-01

    Full Text Available Oncoproteomics aims to the discovery of molecular markers, drug targets and pathways by studying cancer specific protein expression, localization, modification and interaction. Cell surface proteins play a central role in several pathological conditions, including cancer and its metastatic spread. However, cell surface proteins are underrepresented in proteomics analyses performed from the whole cell extracts due to their hydrophobicity and low abundance. Different methods have been developed to enrich and isolate the cell surface proteins to reduce sample complexity. Despite the method selected, the primary difficulty encountered is the solubilization of the hydrophobic transmembrane proteins from the lipid bilayer. This review focuses on proteomic analyses of metastasis-associated proteins identified using the cell surface biotinylation method. Interestingly, also certain intracellular proteins were identified from the cell surface samples. The function of these proteins at the cell surface might well differ from their function inside the cell.

  3. Cells behaviors and genotoxicity on topological surface

    International Nuclear Information System (INIS)

    Yang, N.; Yang, M.K.; Bi, S.X.; Chen, L.; Zhu, Z.Y.; Gao, Y.T.; Du, Z.

    2013-01-01

    To investigate different cells behaviors and genotoxicity, which were driven by specific microenvironments, three patterned surfaces (pillars, wide grooves and narrow grooves) and one smooth surface were prepared by template-based technique. Vinculin is a membrane-cytoskeletal protein in focal adhesion plaques and associates with cell–cell and cell–matrix junctions, which can promote cell adhesion and spreading. The immunofluorescence staining of vinculin revealed that the narrow grooves patterned substrate was favorable for L929 cell adhesion. For cell multiplication, the narrow grooves surface was fitted for the proliferation of L929, L02 and MSC cells, the pillars surface was only in favor of L929 cells to proliferate during 7 days of cell cultivation. Cell genetic toxicity was evaluated by cellular micronuclei test (MNT). The results indicated that topological surfaces were more suitable for L929 cells to proliferate and maintain the stability of genome. On the contrary, the narrow grooves surface induced higher micronuclei ratio of L02 and MSC cells than other surfaces. With the comprehensive results of cell multiplication and MNT, it was concluded that the wide grooves surface was best fitted for L02 cells to proliferate and have less DNA damages, and the smooth surface was optimum for the research of MSC cells in vitro. - Highlights: • Different cells behaviors on microstructure surfaces were discussed in this paper. • The expression of cell protein of Vinculin was studied in this research. • Cellular micronuclei test was applied to evaluate cells' genotoxicity. • Cell genotoxicity was first studied in the research field of topological surfaces

  4. Functions of proteoglycans at the cell surface

    DEFF Research Database (Denmark)

    Höök, M; Woods, A; Johansson, S

    1986-01-01

    Proteoglycans (primarily heparan sulphate proteoglycans) are found at the surface of most adherent eukaryotic cells. Earlier studies suggest that these molecules can be associated with the cell surface principally by two different mechanisms. Proteoglycans may occur as membrane......-intercalated glycoproteins, where the core protein of the proteoglycan is anchored in the lipid interior of the plasma membrane, or they may be bound via the polysaccharide components of the molecule to specific anchoring proteins present at the cell surface. A number of functions have been proposed for cell surface......-associated proteoglycans, including: regulation of cell-substrate adhesion; regulation of cell proliferation; participation in the binding and uptake of extracellular components; and participation in the regulation of extracellular matrix formation. Evidence is discussed suggesting that the cell-associated heparan...

  5. Radioimmunoassay to quantitatively measure cell surface immunoglobulins

    International Nuclear Information System (INIS)

    Krishman, E.C.; Jewell, W.R.

    1975-01-01

    A radioimmunoassay techniques developed to quantitatively measure the presence of immunoglobulins on the surface of cells, is described. The amount of immunoglobulins found on different tumor cells varied from 200 to 1140 ng/10 6 cells. Determination of immunoglobulins on the peripheral lymphocytes obtained from different cancer patients varied between 340 to 1040 ng/10 6 cells. Cultured tumor cells, on the other hand, were found to contain negligible quantities of human IgG [pt

  6. Functions of proteoglycans at the cell surface

    DEFF Research Database (Denmark)

    Höök, M; Woods, A; Johansson, S

    1986-01-01

    sulphate helps to connect the intracellular cytoskeleton to the extracellular matrix in focal adhesions. This evidence includes: the co-localization of actin and heparan sulphate proteoglycan during the process of cell spreading, and in isolated focal adhesions; biochemical analyses of a hydrophobic......Proteoglycans (primarily heparan sulphate proteoglycans) are found at the surface of most adherent eukaryotic cells. Earlier studies suggest that these molecules can be associated with the cell surface principally by two different mechanisms. Proteoglycans may occur as membrane......-intercalated glycoproteins, where the core protein of the proteoglycan is anchored in the lipid interior of the plasma membrane, or they may be bound via the polysaccharide components of the molecule to specific anchoring proteins present at the cell surface. A number of functions have been proposed for cell surface...

  7. A nucleation theory of cell surface capping

    International Nuclear Information System (INIS)

    Coutsias, E.A.; Wester, M.J.; Perelson, A.S.

    1997-01-01

    We propose a new theory of cell surface capping based on the principles of nucleation. When antibody interacts with cell surface molecules, the molecules initially form small aggregates called patches that later coalesce into a large aggregate called a cap. While a cap can form by patches being pulled together by action of the cell''s cytoskeleton, in the case of some molecules, disruption of the cytoskeleton does not prevent cap formation. Diffusion of large aggregates on a cell surface is slow, and thus we propose that a cap can form solely through the diffusion of small aggregates containing just one or a few cell surface molecules. Here we consider the extreme case in which single molecules are mobile, but aggregates of all larger sizes are immobile. We show that a set of patches in equilibrium with a open-quotes seaclose quotes of free cell surface molecules can undergo a nucleation-type phase transition in which the largest patch will bind free cell surface molecules, deplete the concentration of such molecules in the open-quotes seaclose quotes and thus cause the other patches to shrink in size. We therefore show that a cap can form without patches having to move, collide with each other, and aggregate

  8. Structure-based stabilization of HIV-1 gp120 enhances humoral immune responses to the induced co-receptor binding site.

    Directory of Open Access Journals (Sweden)

    Barna Dey

    2009-05-01

    Full Text Available The human immunodeficiency virus type 1 (HIV-1 exterior envelope glycoprotein, gp120, possesses conserved binding sites for interaction with the primary virus receptor, CD4, and also for the co-receptor, generally CCR5. Although gp120 is a major target for virus-specific neutralizing antibodies, the gp120 variable elements and its malleable nature contribute to evasion of effective host-neutralizing antibodies. To understand the conformational character and immunogenicity of the gp120 receptor binding sites as potential vaccine targets, we introduced structure-based modifications to stabilize gp120 core proteins (deleted of the gp120 major variable regions into the conformation recognized by both receptors. Thermodynamic analysis of the re-engineered core with selected ligands revealed significant stabilization of the receptor-binding regions. Stabilization of the co-receptor-binding region was associated with a marked increase in on-rate of ligand binding to this site as determined by surface plasmon resonance. Rabbit immunization studies showed that the conformational stabilization of core proteins, along with increased ligand affinity, was associated with strikingly enhanced humoral immune responses against the co-receptor-binding site. These results demonstrate that structure-based approaches can be exploited to stabilize a conformational site in a large functional protein to enhance immunogenic responses specific for that region.

  9. Nanostructuring of Solar Cell Surfaces

    DEFF Research Database (Denmark)

    Davidsen, Rasmus Schmidt; Schmidt, Michael Stenbæk

    Solar energy is by far the most abundant renewable energy source available, but the levelized cost of solar energy is still not competitive with that of fossil fuels. Therefore there is a need to improve the power conversion effciency of solar cells without adding to the production cost. The main...... objective of this PhD thesis is to develop nanostructured silicon (Si) solar cells with higher power conversion efficiency using only scalable and cost-efficient production methods. The nanostructures, known as 'black silicon', are fabricated by single-step, maskless reactive ion etching and used as front...... texturing of different Si solar cells. Theoretically the nanostructure topology may be described as a graded refractive index in a mean-field approximation between air and Si. The optical properties of the developed black Si were simulated and experimentally measured. Total AM1.5G-weighted average...

  10. Probes for anionic cell surface detection

    Science.gov (United States)

    Smith, Bradley D.

    2013-03-05

    Embodiments of the present invention are generally directed to compositions comprising a class of molecular probes for detecting the presence of anionic cell surfaces. Embodiments include compositions that are enriched for these compositions and preparations, particularly preparations suitable for use as laboratory/clinical reagents and diagnostic indicators, either alone or as part of a kit. An embodiment of the invention provides for a highly selective agent useful in the discernment and identification of dead or dying cells, such as apoptotic cells, in a relatively calcium-free environment. An embodiment of the invention provides a selective agent for the identification of bacteria in a mixed population of bacterial cells and nonbacterial cells.

  11. Nanotomography of Cell Surfaces with Evanescent Fields

    Directory of Open Access Journals (Sweden)

    Michael Wagner

    2008-01-01

    Full Text Available The technique of variable-angle total internal reflection fluorescence microscopy (TIRFM and its application to nanotomography of cell surfaces are described. Present applications include (1 3D imaging of chromosomes in their metaphase to demonstrate axial resolution in the nanometre range, (2 measurements of cell-substrate topology, which upon cholesterol depletion shows some loosening of cell-substrate contacts, and (3 measurements of cell topology upon photodynamic therapy (PDT, which demonstrate cell swelling and maintenance of focal contacts. The potential of the method for in vitro diagnostics, but also some requirements and limitations are discussed.

  12. Osteoblastic cell behaviour on modified titanium surfaces.

    Science.gov (United States)

    Lukaszewska-Kuska, Magdalena; Wirstlein, Przemysław; Majchrowski, Radomir; Dorocka-Bobkowska, Barbara

    2018-02-01

    The surfaces of endoosseous dental implants have been subjected to numerous modifications in order to create a surface which can provide rapid bone healing and fast implant loading. Each modification has involved changes to the chemical composition and topography of the surfaces which have resulted in various biological reactions to the implanted material. The aim of this study was to evaluate the surface topography and chemistry of various modified titanium surfaces: (1) machined surface (MA), (2) alumina-blasted (Al2O3), (3) alumina-blasted and acid-etched (Al2O3 DE), (4) hydroxyapatite/tricalcium phosphate grit-blasted (HA/TCP) and (5) hydroxyapatite/tricalcium phosphate grit-blasted and acid-etched (HA/TCP DE) and to analyse the effects of surface roughness, and chemical composition on human osteoblast vitality, differentiation, morphology and orientation. The modified surfaces were subjected to topographic analysis using Scanning Electron Microscopy (SEM), optical profilometry, roughness analysis and chemical composition evaluation using Energy Dispersion Spectroscopy (EDS) analysis. The biological effects of the titanium modifications was analysed using human osteoblasts cell culture where the cell morphology, vitality (MTS assay) and differentiation (ALP activity) was analysed. The machined surfaces were classified as anisotropic, smooth and composed of titanium and oxygen. The blasted surface samples along with the blasted and etched samples were found to be isotropic and rough. The grit-blasting procedure resulted in the incorporation of components from the blasting material. In the case of the blasted and etched samples, etching decreased the surface development as indicated by the Sdr and also reduced the amount of chemical compounds incorporated into the surfaces during the blasting procedure. The attached NHOst cells, proliferated the surfaces. With regard to the MA samples, the cells spread close to the titanium surface, with expanded cytoplasmic

  13. Determination of co-receptor usage of HIV-1

    NARCIS (Netherlands)

    Schuitemaker, Hanneke; Kootstra, Neeltje A.

    2005-01-01

    In addition to CD4, HIV-1 uses chemokine receptors for entry in their target cells. The most important chemokine receptors in this respect are beta-chemokine receptor 5 (CCR5) and alpha-chemokine receptor 4 (CXCR4). Coreceptor usage is an important feature of the biological phenotype of HIV-1

  14. Surface acoustic wave actuated cell sorting (SAWACS).

    Science.gov (United States)

    Franke, T; Braunmüller, S; Schmid, L; Wixforth, A; Weitz, D A

    2010-03-21

    We describe a novel microfluidic cell sorter which operates in continuous flow at high sorting rates. The device is based on a surface acoustic wave cell-sorting scheme and combines many advantages of fluorescence activated cell sorting (FACS) and fluorescence activated droplet sorting (FADS) in microfluidic channels. It is fully integrated on a PDMS device, and allows fast electronic control of cell diversion. We direct cells by acoustic streaming excited by a surface acoustic wave which deflects the fluid independently of the contrast in material properties of deflected objects and the continuous phase; thus the device underlying principle works without additional enhancement of the sorting by prior labelling of the cells with responsive markers such as magnetic or polarizable beads. Single cells are sorted directly from bulk media at rates as fast as several kHz without prior encapsulation into liquid droplet compartments as in traditional FACS. We have successfully directed HaCaT cells (human keratinocytes), fibroblasts from mice and MV3 melanoma cells. The low shear forces of this sorting method ensure that cells survive after sorting.

  15. Surface cell immobilization within perfluoroalkoxy microchannels

    Energy Technology Data Exchange (ETDEWEB)

    Stojkovič, Gorazd; Krivec, Matic [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia); Vesel, Alenka [Jožef Stefan Institute, Jamova cesta 39, 1000 Ljubljana (Slovenia); Marinšek, Marjan [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia); Žnidaršič-Plazl, Polona, E-mail: polona.znidarsic@fkkt.uni-lj.si [Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva 5, SI-1000 Ljubljana (Slovenia)

    2014-11-30

    Graphical abstract: - Highlights: • A very efficient approach for immobilization of cells into microreactors is presented. • It is applicable to various materials, including PFA and cyclic olefin (co)polymers. • It was used to immobilize different prokaryotic and eukaryotic microbes. • Cells were immobilized on the surface in high density and showed good stability. • Mechanisms of APTES interactions with target materials are proposed. - Abstract: Perfluoroalkoxy (PFA) is one of the most promising materials for the fabrication of cheap, solvent resistant and reusable microfluidic chips, which have been recently recognized as effective tools for biocatalytic process development. The application of biocatalysts significantly depends on efficient immobilization of enzymes or cells within the reactor enabling long-term biocatalyst use. Functionalization of PFA microchannels by 3-aminopropyltriethoxysilane (ATPES) and glutaraldehyde was used for rapid preparation of microbioreactors with surface-immobilized cells. X-ray photoelectron spectroscopy and scanning electron microscopy were used to accurately monitor individual treatment steps and to select conditions for cell immobilization. The optimized protocol for Saccharomyces cerevisiae immobilization on PFA microchannel walls comprised ethanol surface pretreatment, 4 h contacting with 10% APTES aqueous solution, 10 min treatment with 1% glutaraldehyde and 20 min contacting with cells in deionized water. The same protocol enabled also immobilization of Escherichia coli, Pseudomonas putida and Bacillus subtilis cells on PFA surface in high densities. Furthermore, the developed procedure has been proved to be very efficient also for surface immobilization of tested cells on other materials that are used for microreactor fabrication, including glass, polystyrene, poly (methyl methacrylate), polycarbonate, and two olefin-based polymers, namely Zeonor{sup ®} and Topas{sup ®}.

  16. Metabolic behavior of cell surface biotinylated proteins

    International Nuclear Information System (INIS)

    Hare, J.F.; Lee, E.

    1989-01-01

    The turnover of proteins on the surface of cultured mammalian cells was measured by a new approach. Reactive free amino or sulfhydryl groups on surface-accessible proteins were derivatized with biotinyl reagents and the proteins solubilized from culture dishes with detergent. Solubilized, biotinylated proteins were then adsorbed onto streptavidin-agarose, released with sodium dodecyl sulfate and mercaptoethanol, and separated on polyacrylamide gels. Biotin-epsilon-aminocaproic acid N-hydroxysuccinimide ester (BNHS) or N-biotinoyl-N'-(maleimidohexanoyl)hydrazine (BM) were the derivatizing agents. Only 10-12 bands were adsorbed onto streptavidin-agarose from undervatized cells or from derivatized cells treated with free avidin at 4 degrees C. Two-dimensional isoelectric focusing-sodium dodecyl sulfate gel electrophoresis resolved greater than 100 BNHS-derivatized proteins and greater than 40 BM-derivatized proteins. There appeared to be little overlap between the two groups of derivatized proteins. Short-term pulse-chase studies showed an accumulation of label into both groups of biotinylated proteins up until 1-2 h of chase and a rapid decrease over the next 1-5 h. Delayed appearance of labeled protein at the cell surface was attributed to transit time from site of synthesis. The unexpected and unexplained rapid disappearance of pulse-labeled proteins from the cell surface was invariant for all two-dimensionally resolved proteins and was sensitive to temperature reduction to 18 degrees C. Long-term pulse-chase experiments beginning 4-8 h after the initiation of chase showed the disappearance of derivatized proteins to be a simple first-order process having a half-life of 115 h in the case of BNHS-derivatized proteins and 30 h in the case of BM-derivatized proteins

  17. Zebrafish GDNF and its co-receptor GFRα1 activate the human RET receptor and promote the survival of dopaminergic neurons in vitro.

    Directory of Open Access Journals (Sweden)

    Tuulia Saarenpää

    Full Text Available Glial cell line-derived neurotrophic factor (GDNF is a ligand that activates, through co-receptor GDNF family receptor alpha-1 (GFRα1 and receptor tyrosine kinase "RET", several signaling pathways crucial in the development and sustainment of multiple neuronal populations. We decided to study whether non-mammalian orthologs of these three proteins have conserved their function: can they activate the human counterparts? Using the baculovirus expression system, we expressed and purified Danio rerio RET, and its binding partners GFRα1 and GDNF, and Drosophila melanogaster RET and two isoforms of co-receptor GDNF receptor-like. Our results report high-level insect cell expression of post-translationally modified and dimerized zebrafish RET and its binding partners. We also found that zebrafish GFRα1 and GDNF are comparably active as mammalian cell-produced ones. We also report the first measurements of the affinity of the complex to RET in solution: at least for zebrafish, the Kd for GFRα1-GDNF binding RET is 5.9 μM. Surprisingly, we also found that zebrafish GDNF as well as zebrafish GFRα1 robustly activated human RET signaling and promoted the survival of cultured mouse dopaminergic neurons with comparable efficiency to mammalian GDNF, unlike E. coli-produced human proteins. These results contradict previous studies suggesting that mammalian GFRα1 and GDNF cannot bind and activate non-mammalian RET and vice versa.

  18. Accurate and efficient gp120 V3 loop structure based models for the determination of HIV-1 co-receptor usage

    Directory of Open Access Journals (Sweden)

    Vaisman Iosif I

    2010-10-01

    Full Text Available Abstract Background HIV-1 targets human cells expressing both the CD4 receptor, which binds the viral envelope glycoprotein gp120, as well as either the CCR5 (R5 or CXCR4 (X4 co-receptors, which interact primarily with the third hypervariable loop (V3 loop of gp120. Determination of HIV-1 affinity for either the R5 or X4 co-receptor on host cells facilitates the inclusion of co-receptor antagonists as a part of patient treatment strategies. A dataset of 1193 distinct gp120 V3 loop peptide sequences (989 R5-utilizing, 204 X4-capable is utilized to train predictive classifiers based on implementations of random forest, support vector machine, boosted decision tree, and neural network machine learning algorithms. An in silico mutagenesis procedure employing multibody statistical potentials, computational geometry, and threading of variant V3 sequences onto an experimental structure, is used to generate a feature vector representation for each variant whose components measure environmental perturbations at corresponding structural positions. Results Classifier performance is evaluated based on stratified 10-fold cross-validation, stratified dataset splits (2/3 training, 1/3 validation, and leave-one-out cross-validation. Best reported values of sensitivity (85%, specificity (100%, and precision (98% for predicting X4-capable HIV-1 virus, overall accuracy (97%, Matthew's correlation coefficient (89%, balanced error rate (0.08, and ROC area (0.97 all reach critical thresholds, suggesting that the models outperform six other state-of-the-art methods and come closer to competing with phenotype assays. Conclusions The trained classifiers provide instantaneous and reliable predictions regarding HIV-1 co-receptor usage, requiring only translated V3 loop genotypes as input. Furthermore, the novelty of these computational mutagenesis based predictor attributes distinguishes the models as orthogonal and complementary to previous methods that utilize sequence

  19. First report of a healthy Indian heterozygous for delta 32 mutant of HIV-1 co-receptor-CCR5 gene.

    Science.gov (United States)

    Husain, S; Goila, R; Shahi, S; Banerjea, A

    1998-01-30

    The beta-chemokine receptor, CCR5, is a major co-receptor for macrophage tropic non-syncytia-inducing isolates of HIV-1. Recently a 32 bp homozygous deletion in the coding region of CCR5 has been reported in a very small percentage (heritage with varying frequencies (13-20%). However, when a large number of the non-Caucasian population (261 Africans and 423 Asians) were screened for the presence of this deleted allele, not a single case of either homozygous or heterozygous mutant for delta 32 allele of CCR5 was detected. We screened 100 normal individuals and found a single heterozygous case with an identical 32 bp deletion in CCR5 gene reported earlier, the rest possessed wild-type alleles. This deleted gene was inherited in Mendelian fashion among the family members of this individual. Thus, the frequency of this deleted allele in India among unrelated normal individuals is likely to be very low (< 1%). We observed a moderate transdominant effect of this mutant allele in a fusion assay. Finally, we show a significant inhibition of fusion of cell membranes when the 176-bp region of CCR5 was used as an antisense.

  20. Lipid raft localization of TLR2 and its co-receptors is independent of membrane lipid composition

    Directory of Open Access Journals (Sweden)

    Christine Hellwing

    2018-01-01

    Full Text Available Background Toll like receptors (TLRs are an important and evolutionary conserved class of pattern recognition receptors associated with innate immunity. The recognition of Gram-positive cell wall constituents strongly depends on TLR2. In order to be functional, TLR2 predominantly forms a heterodimer with TLR1 or TLR6 within specialized membrane microdomains, the lipid rafts. The membrane lipid composition and the physicochemical properties of lipid rafts are subject to modification by exogenous fatty acids. Previous investigations of our group provide evidence that macrophage enrichment with polyunsaturated fatty acids (PUFA induces a reordering of lipid rafts and non-rafts based on the incorporation of supplemented PUFA as well as their elongation and desaturation products. Methods In the present study we investigated potential constraining effects of membrane microdomain reorganization on the clustering of TLR2 with its co-receptors TLR1 and TLR6 within lipid rafts. To this end, RAW264.7 macrophages were supplemented with either docosahexaenoic acid (DHA or arachidonic acid (AA and analyzed for receptor expression and microdomain localization in context of TLR stimulation. Results and Conclusions Our analyses showed that receptor levels and microdomain localization were unchanged by PUFA supplementation. The TLR2 pathway, in contrast to the TLR4 signaling cascade, is not affected by exogenous PUFA at the membrane level.

  1. Frequency of polymorphisms of genes coding for HIV-1 co-receptors CCR5 and CCR2 in a Brazilian population

    Directory of Open Access Journals (Sweden)

    Munerato Patrícia

    2003-01-01

    Full Text Available Entry of human immunodeficiency type 1 virus (HIV-1 into target cells requires both CD4and one of the chemokine receptors. Viruses predominantly use one, or occasionally both, of the major co-receptors CCR5 and CXCR4, although other receptors, including CCR2B and CCR3, function as minor co-receptors. A 32-nucleotide deletion (delta32 within the beta-chemokine receptor 5 gene (CCR5 has been described in subjects who remain uninfected despite extensive exposition to HIV-1. The heterozygous genotype delays disease progression. This allele is common among Caucasians, but has not been found in people of African or Asian ancestry. A more common transition involving a valine to isoleucine switch in transmembrane domain I of CCR2B (64I, with unknown functional consequences, was found to delay disease progression but not to reduce infection risk. As the Brazilian population consists of a mixture of several ethnic groups, we decided to examine the genotype frequency of these polymorphisms in this country. There were 11.5% CCR5 heterozygotes among the HIV-1 infected population and 12.5% among uninfected individuals, similar to data from North America and Western Europe. The prevalence of CCR2-64I homozygotes and heterozygotes was 0.06 and 15.2%, respectively, also similar to what is known for North America and Western Europe.

  2. The chaperone and potential mannan-binding lectin (MBL) co-receptor calreticulin interacts with MBL through the binding site for MBL-associated serine proteases

    DEFF Research Database (Denmark)

    Pagh, Rasmus; Duus, Karen; Laursen, Inga

    2008-01-01

    was immobilized on a solid surface or bound to mannan on a surface. The binding was non-covalent and biphasic with an initial salt-sensitive phase followed by a more stable salt-insensitive interaction. For plasma-derived MBL, known to be complexed with MBL-associated serine proteases (MASPs), no binding...... with calreticulin. Comparative analysis of MBL with complement component C1q, its counterpart of the classical pathway, revealed that they display similar binding characteristics for calreticulin, providing further indication that calreticulin is a common co-receptor/chaperone for both proteins. In conclusion......The chaperone calreticulin has been suggested to function as a C1q and collectin receptor. The interaction of calreticulin with mannan-binding lectin (MBL) was investigated by solid-phase binding assays. Calreticulin showed saturable and time-dependent binding to recombinant MBL, provided that MBL...

  3. Cancer cell proliferation controlled by surface chemistry in its microenvironment

    Science.gov (United States)

    Yu, Xiao-Long; Zhang, Bin; Wang, Xiu-Mei; Wang, Ying; Qiao, Lin; He, Jin; Wang, Juan; Chen, Shuang-Feng; Lee, In-Seop; Cui, Fu-Zhai

    2011-12-01

    Hepatoma cells (Hepg2s) as typical cancer cells cultured on hydroxyl (-OH) and methyl (-CH3) group surfaces were shown to exhibit different proliferation and morphological changes. Hepg2s cells on -OH surfaces grew much more rapidly than those on -CH3 surfaces. Hepg2s cells on -OH surfaces had the larger contact area and the more flattened morphology, while those on -CH3 surfaces exhibited the smaller contact area and the more rounded morphology. After 7 days of culture, the migration of Hepg2s cells into clusters on the -CH3 surfaces behaved significantly slower than that on the -OH surfaces. These chemically modified surfaces exhibited regulation of Hepg2s cells on proliferation, adhesion, and migration, providing a potential treatment of liver cancer.

  4. Cell surface control of differentiation in Acanthamoeba.

    Science.gov (United States)

    Yang, S; Villemez, C

    1994-12-01

    Acanthamoeba castellanii (Neff) is a free-living soil amoeba with close relatives that are opportunistic pathogens. Trophozoites differentiate into cysts when deprived of nutrients; cysts convert into trophozoites, leaving the wall behind, in the presence of nutrients. The data presented here, which includes immunoaffinity purification of the receptor, indicate that cell surface molecular signals also control Acanthamoeba differentiation in both directions. Monoclonal antibodies that bind specifically to a 40 kD trophozoite protein initiate the encystment of trophozoites. When bound to cysts the same monoclonal antibodies prevent excystment. Washing away the antibody allows both trophozoites and cysts to resume normal activity. One of these monoclonal antibodies inhibits pinocytosis, while another has no effect on pinocytosis.

  5. Dictyostelium cells migrate similarly on surfaces of varying chemical composition.

    Science.gov (United States)

    McCann, Colin P; Rericha, Erin C; Wang, Chenlu; Losert, Wolfgang; Parent, Carole A

    2014-01-01

    During cell migration, cell-substrate binding is required for pseudopod anchoring to move the cell forward, yet the interactions with the substrate must be sufficiently weak to allow parts of the cell to de-adhere in a controlled manner during typical protrusion/retraction cycles. Mammalian cells actively control cell-substrate binding and respond to extracellular conditions with localized integrin-containing focal adhesions mediating mechanotransduction. We asked whether mechanotransduction also occurs during non-integrin mediated migration by examining the motion of the social amoeba Dictyostelium discoideum, which is thought to bind non-specifically to surfaces. We discovered that Dictyostelium cells are able to regulate forces generated by the actomyosin cortex to maintain optimal cell-surface contact area and adhesion on surfaces of various chemical composition and that individual cells migrate with similar speed and contact area on the different surfaces. In contrast, during collective migration, as observed in wound healing and metastasis, the balance between surface forces and protrusive forces is altered. We found that Dictyostelium collective migration dynamics are strongly affected when cells are plated on different surfaces. These results suggest that the presence of cell-cell contacts, which appear as Dictyostelium cells enter development, alter the mechanism cells use to migrate on surfaces of varying composition.

  6. HIV Co-receptor usage in HIV-related non-hodgkin's lymphoma

    Directory of Open Access Journals (Sweden)

    Reid Erin

    2012-03-01

    Full Text Available Abstract In this study 15 banked samples of HIV-related Non-Hodgkin's Lymphoma (NHL cases were tested for HIV co-receptor usage and SDF1 3'A polymorphism. Reportable tropism from 9 plasma samples had 1 (11.1% HIV case with CXCR4 and 8 (88.9% with CCR5 usage, even though most of the cases occurred at a late stage of HIV (2/3 had CD4 counts below 200, where expected CXCR4 usage would be 60%. Based on the expected proportion of less than 50% CCR5 in chronically infected individuals, this would suggest that in NHL may be associated with CCR5 usage (P = 0.04.

  7. Engineered cell surfaces: fertile ground for molecular landscaping.

    Science.gov (United States)

    Mahal, L K; Bertozzi, C R

    1997-06-01

    The cell surface contains a wealth of information that determines how cells interact with their environment. Methods for directing the cell surface expression of novel protein-based and oligosaccharide-based epitopes are stimulating new directions in biotechnology and biomedical research.

  8. Nanofabrication of Nonfouling Surfaces for Micropatterning of Cell and Microtissue

    Directory of Open Access Journals (Sweden)

    Hidenori Otsuka

    2010-08-01

    Full Text Available Surface engineering techniques for cellular micropatterning are emerging as important tools to clarify the effects of the microenvironment on cellular behavior, as cells usually integrate and respond the microscale environment, such as chemical and mechanical properties of the surrounding fluid and extracellular matrix, soluble protein factors, small signal molecules, and contacts with neighboring cells. Furthermore, recent progress in cellular micropatterning has contributed to the development of cell-based biosensors for the functional characterization and detection of drugs, pathogens, toxicants, and odorants. In this regards, the ability to control shape and spreading of attached cells and cell-cell contacts through the form and dimension of the cell-adhesive patches with high precision is important. Commitment of stem cells to different specific lineages depends strongly on cell shape, implying that controlled microenvironments through engineered surfaces may not only be a valuable approach towards fundamental cell-biological studies, but also of great importance for the design of cell culture substrates for tissue engineering. To develop this kind of cellular microarray composed of a cell-resistant surface and cell attachment region, micropatterning a protein-repellent surface is important because cellular adhesion and proliferation are regulated by protein adsorption. The focus of this review is on the surface engineering aspects of biologically motivated micropatterning of two-dimensional surfaces with the aim to provide an introductory overview described in the literature. In particular, the importance of non-fouling surface chemistries is discussed.

  9. Osteoblast cell response to surface-modified carbon nanotubes

    International Nuclear Information System (INIS)

    Zhang Faming; Weidmann, Arne; Nebe, J. Barbara; Burkel, Eberhard

    2012-01-01

    In order to investigate the interaction of cells with modified multi-walled carbon nanotubes (MWCNTs) for their potential biomedical applications, the MWCNTs were chemically modified with carboxylic acid groups (–COOH), polyvinyl alcohol (PVA) polymer and biomimetic apatite on their surfaces. Additionally, human osteoblast MG-63 cells were cultured in the presence of the surface-modified MWCNTs. The metabolic activities of osteoblastic cells, cell proliferation properties, as well as cell morphology were studied. The surface modification of MWCNTs with biomimetic apatite exhibited a significant increase in the cell viability of osteoblasts, up to 67.23%. In the proliferation phases, there were many more cells in the biomimetic apatite-modified MWCNT samples than in the MWCNTs–COOH. There were no obvious changes in cell morphology in osteoblastic MG-63 cells cultured in the presence of these chemically-modified MWCNTs. The surface modification of MWCNTs with apatite achieves an effective enhancement of their biocompatibility.

  10. Growth of fibroblasts and endothelial cells on wettability gradient surfaces

    NARCIS (Netherlands)

    Ruardy, TG; Moorlag, HE; Schakenraad, JM; VanderMei, HC; Busscher, HJ

    1997-01-01

    The growth, spreading, and shape of human skin fibroblasts (PK 84) and human umbilical cord endothelial cells on dichlorodimethylsilane (DDS) and dimethyloctadecylchlorosilane (DOGS) gradient surfaces were investigated in the presence of serum proteins. Gradient surfaces were prepared on glass using

  11. Defining Binding Efficiency and Specificity of Auxins for SCFTIR1/AFB-Aux/IAA Co-receptor Complex Formation

    Science.gov (United States)

    2013-01-01

    Structure–activity profiles for the phytohormone auxin have been collected for over 70 years, and a number of synthetic auxins are used in agriculture. Auxin classification schemes and binding models followed from understanding auxin structures. However, all of the data came from whole plant bioassays, meaning the output was the integral of many different processes. The discovery of Transport Inhibitor-Response 1 (TIR1) and the Auxin F-Box (AFB) proteins as sites of auxin perception and the role of auxin as molecular glue in the assembly of co-receptor complexes has allowed the development of a definitive quantitative structure–activity relationship for TIR1 and AFB5. Factorial analysis of binding activities offered two uncorrelated factors associated with binding efficiency and binding selectivity. The six maximum-likelihood estimators of Efficiency are changes in the overlap matrixes, inferring that Efficiency is related to the volume of the electronic system. Using the subset of compounds that bound strongly, chemometric analyses based on quantum chemical calculations and similarity and self-similarity indices yielded three classes of Specificity that relate to differential binding. Specificity may not be defined by any one specific atom or position and is influenced by coulomb matrixes, suggesting that it is driven by electrostatic forces. These analyses give the first receptor-specific classification of auxins and indicate that AFB5 is the preferred site for a number of auxinic herbicides by allowing interactions with analogues having van der Waals surfaces larger than that of indole-3-acetic acid. The quality factors are also examined in terms of long-standing models for the mechanism of auxin binding. PMID:24313839

  12. Cell behavior on microparticles with different surface morphology

    Energy Technology Data Exchange (ETDEWEB)

    Huang Sha [Wound Healing and Cell Biology Laboratory, Institute of Basic Medical Sciences, General Hospital of PLA, Beijing 100853 (China); Fu Xiaobing, E-mail: fuxiaobing@vip.sina.co [Wound Healing and Cell Biology Laboratory, Institute of Basic Medical Sciences, General Hospital of PLA, Beijing 100853 (China); Burns Institute, The First Affiliated Hospital, General Hospital of PLA, Trauma Center of Postgraduate Medical College, Beijing 100037 (China)

    2010-03-18

    Microparticles can serve as substrates for cell amplification and deliver the cell aggregation to the site of the defect for tissue regeneration. To develop favorable microparticles for cell delivery application, we fabricated and evaluated three types of microparticles that differ in surface properties. The microparticles with varied surface morphology (smooth, pitted and multicavity) were created from chemically crosslinked gelatin particles that underwent various drying treatments. Three types of microparticles were characterized and assessed in terms of the cell behavior of human keratinocytes and fibroblasts seeded on them. The cells could attach, spread and proliferate on all types of microparticles but spread and populated more slowly on the microparticles with smooth surfaces than on those with pitted or multicavity surfaces. Microparticles with a multicavity surface demonstrated the highest cell attachment and growth rate. Furthermore, cells tested on microparticles with a multicavity surface exhibited better morphology and induced the earlier formation of extracellular-based cell-microparticle aggregation than those on microparticles with other surface morphology (smooth and pitted). Thus, microparticles with a multicavity surface show promise for attachment and proliferation of cells in tissue engineering.

  13. Surface Passivation for Silicon Heterojunction Solar Cells

    NARCIS (Netherlands)

    Deligiannis, D.

    2017-01-01

    Silicon heterojunction solar cells (SHJ) are currently one of the most promising solar cell technologies in the world. The SHJ solar cell is based on a crystalline silicon (c-Si) wafer, passivated on both sides with a thin intrinsic hydrogenated amorphous silicon (a-Si:H) layer. Subsequently, p-type

  14. Touching Textured Surfaces: Cells in Somatosensory Cortex Respond Both to Finger Movement and to Surface Features

    Science.gov (United States)

    Darian-Smith, Ian; Sugitani, Michio; Heywood, John; Karita, Keishiro; Goodwin, Antony

    1982-11-01

    Single neurons in Brodmann's areas 3b and 1 of the macaque postcentral gyrus discharge when the monkey rubs the contralateral finger pads across a textured surface. Both the finger movement and the spatial pattern of the surface determine this discharge in each cell. The spatial features of the surface are represented unambiguously only in the responses of populations of these neurons, and not in the responses of the constituent cells.

  15. Cell Surface Proteome of Dental Pulp Stem Cells Identified by Label-Free Mass Spectrometry.

    Directory of Open Access Journals (Sweden)

    Christian Niehage

    Full Text Available Multipotent mesenchymal stromal cells (MSCs are promising tools for regenerative medicine. They can be isolated from different sources based on their plastic-adherence property. The identification of reliable cell surface markers thus becomes the Holy Grail for their prospective isolation. Here, we determine the cell surface proteomes of human dental pulp-derived MSCs isolated from single donors after culture expansion in low (2% or high (10% serum-containing media. Cell surface proteins were tagged on intact cells using cell impermeable, cleavable sulfo-NHS-SS-biotin, which allows their enrichment by streptavidin pull-down. For the proteomic analyses, we first compared label-free methods to analyze cell surface proteomes i.e. composition, enrichment and proteomic differences, and we developed a new mathematical model to determine cell surface protein enrichment using a combinatorial gene ontology query. Using this workflow, we identified 101 cluster of differentiation (CD markers and 286 non-CD cell surface proteins. Based on this proteome profiling, we identified 14 cell surface proteins, which varied consistently in abundance when cells were cultured under low or high serum conditions. Collectively, our analytical methods provide a basis for identifying the cell surface proteome of dental pulp stem cells isolated from single donors and its evolution during culture or differentiation. Our data provide a comprehensive cell surface proteome for the precise identification of dental pulp-derived MSC populations and their isolation for potential therapeutic intervention.

  16. Pheochromocytoma (PC12 Cell Response on Mechanobactericidal Titanium Surfaces

    Directory of Open Access Journals (Sweden)

    Jason V. Wandiyanto

    2018-04-01

    Full Text Available Titanium is a biocompatible material that is frequently used for making implantable medical devices. Nanoengineering of the surface is the common method for increasing material biocompatibility, and while the nanostructured materials are well-known to represent attractive substrata for eukaryotic cells, very little information has been documented about the interaction between mammalian cells and bactericidal nanostructured surfaces. In this study, we investigated the effect of bactericidal titanium nanostructures on PC12 cell attachment and differentiation—a cell line which has become a widely used in vitro model to study neuronal differentiation. The effects of the nanostructures on the cells were then compared to effects observed when the cells were placed in contact with non-structured titanium. It was found that bactericidal nanostructured surfaces enhanced the attachment of neuron-like cells. In addition, the PC12 cells were able to differentiate on nanostructured surfaces, while the cells on non-structured surfaces were not able to do so. These promising results demonstrate the potential application of bactericidal nanostructured surfaces in biomedical applications such as cochlear and neuronal implants.

  17. Biomaterial surface proteomic signature determines interaction with epithelial cells.

    Science.gov (United States)

    Abdallah, Mohamed-Nur; Tran, Simon D; Abughanam, Ghada; Laurenti, Marco; Zuanazzi, David; Mezour, Mohamed A; Xiao, Yizhi; Cerruti, Marta; Siqueira, Walter L; Tamimi, Faleh

    2017-05-01

    Cells interact with biomaterials indirectly through extracellular matrix (ECM) proteins adsorbed onto their surface. Accordingly, it could be hypothesized that the surface proteomic signature of a biomaterial might determine its interaction with cells. Here, we present a surface proteomic approach to test this hypothesis in the specific case of biomaterial-epithelial cell interactions. In particular, we determined the surface proteomic signature of different biomaterials exposed to the ECM of epithelial cells (basal lamina). We revealed that the biomaterial surface chemistry determines the surface proteomic profile, and subsequently the interaction with epithelial cells. In addition, we found that biomaterials with surface chemistries closer to that of percutaneous tissues, such as aminated PMMA and aminated PDLLA, promoted higher selective adsorption of key basal lamina proteins (laminins, nidogen-1) and subsequently improved their interactions with epithelial cells. These findings suggest that mimicking the surface chemistry of natural percutaneous tissues can improve biomaterial-epithelial integration, and thus provide a rationale for the design of improved biomaterial surfaces for skin regeneration and percutaneous medical devices. Failure of most biomaterials originates from the inability to predict and control the influence of their surface properties on biological phenomena, particularly protein adsorption, and cellular behaviour, which subsequently results in unfavourable host response. Here, we introduce a surface-proteomic screening approach using a label-free mass spectrometry technique to decipher the adsorption profile of extracellular matrix (ECM) proteins on different biomaterials, and correlate it with cellular behaviour. We demonstrated that the way a biomaterial selectively interacts with specific ECM proteins of a given tissue seems to determine the interactions between the cells of that tissue and biomaterials. Accordingly, this approach can

  18. Effect of laser modified surface microtopochemistry on endothelial cell growth.

    Science.gov (United States)

    Duncan, A C; Rouais, F; Lazare, S; Bordenave, L; Baquey, Ch

    2007-02-15

    The introduction of microelectronics technology in the area of biological sciences has brought forth previously unforeseeable applications such as DNA or protein biochips, miniaturized, multiparametric biosensors for high performance multianalyte assays, DNA sequencing, biocomputers, and substrates for controlled cell growth (i.e. tissue engineering). We developed and investigated a new method using "cold" excimer laser beam technology combined with microlithographical techniques to create surfaces with well defined 3D microdomains in order to delineate critical microscopic surface features governing cell-material interactions. Microfabricated surfaces with microgrooves 30-3 microm deep, 10 - 1 microm wide spaced 30 microm apart were obtained with micron resolution, by "microsculpturing" polymer model surfaces using a computer controlled laser KrF excimer beam coupled with a microlithographic projection technique. The laser beam after exiting a mask was focused onto the polymer target surface via an optical setup allowing for a 10-fold reduction of the mask pattern. Various 3D micropatterned features were obtained at the micron level. Reproducible submicron features could also be obtained using this method. Subsequently, model human umbilical endothelial cells (HUVEC) were cultured on the laser microfabricated surfaces in order to study the effects of specific microscopic surface features on cell deposition and orientation. Cell deposition patterns were found to be microstructure dependant, and showed cell orientation dependency for features in the cell range dimension, a behaviour significantly different from that of a previously studied cell model (osteoprogenitor cell). This model may be a promising in so far as it is very rapid (a time frame less than a second per square centimeter of micropatterned surface) and provides further insights into the effects of surface microtopography on cell response with possible applications in the field of biosensors

  19. Interactions between endothelial progenitor cells (EPC) and titanium implant surfaces.

    Science.gov (United States)

    Ziebart, Thomas; Schnell, Anne; Walter, Christian; Kämmerer, Peer W; Pabst, Andreas; Lehmann, Karl M; Ziebart, Johanna; Klein, Marc O; Al-Nawas, Bilal

    2013-01-01

    Endothelial cells play an important role in peri-implant angiogenesis during early bone formation. Therefore, interactions between endothelial progenitor cells (EPCs) and titanium dental implant surfaces are of crucial interest. The aim of our in vitro study was to investigate the reactions of EPCs in contact with different commercially available implant surfaces. EPCs from buffy coats were isolated by Ficoll density gradient separation. After cell differentiation, EPC were cultured for a period of 7 days on different titanium surfaces. The test surfaces varied in roughness and hydrophilicity: acid-etched (A), sand-blasted-blasted and acid-etched (SLA), hydrophilic A (modA), and hydrophilic SLA (modSLA). Plastic and fibronectin-coated plastic surfaces served as controls. Cell numbers and morphology were analyzed by confocal laser scanning microscopy. Secretion of vascular endothelial growth factor (VEGF)-A was measured by enzyme-linked immunosorbent assay and expressions of iNOS and eNOS were investigated by real-time polymerase chain reaction. Cell numbers were higher in the control groups compared to the cells of titanium surfaces. Initially, hydrophilic titanium surfaces (modA and modSLA) showed lower cell numbers than hydrophobic surfaces (A and SLA). After 7 days smoother surfaces (A and modA) showed increased cell numbers compared to rougher surfaces (SLA and modSLA). Cell morphology of A, modA, and control surfaces was characterized by a multitude of pseudopodia and planar cell soma architecture. SLA and modSLA promoted small and plump cell soma with little quantity of pseudopodia. The lowest VEGF level was measured on A, the highest on modSLA. The highest eNOS and iNOS expressions were found on modA surfaces. The results of this study demonstrate that biological behaviors of EPCs can be influenced by different surfaces. The modSLA surface promotes an undifferentiated phenotype of EPCs that has the ability to secrete growth factors in great quantities. In

  20. Cell response to hydroxyapatite surface topography modulated by sintering temperature.

    Science.gov (United States)

    Mealy, Jacob; O'Kelly, Kevin

    2015-11-01

    Increased mesenchymal stem cell (MSC) activity on hydroxyapatite (HA) bone tissue engineering scaffolds will improve their viability in diffusion-based in vivo environments and is therefore highly desirable. This work focused on modulating the sintered HA surface topography with a view to increasing cell activity; this was achieved by varying the sintering temperature of the HA substrates. Cells were cultured on the substrates for periods of up to 19 days and displayed a huge variation in viability. MSC metabolic activity was measured using a resazurin sodium salt assay and revealed that surfaces sintered from 1250 to 1350°C significantly outperformed their lower temperature counterparts from day one (p ≤ 0.05). Surfaces sintered at 1300°C induced 57% more cell activity than the control at day 16. No significant activity was observed on surfaces sintered below 1200°C. It is suggested that this is due to the granular morphology produced at these temperatures providing insufficient contact area for cell attachment. In addition, we propose the average surface wavelength as a more quantitative surface descriptor than those readily found in the literature. The wavelengths of the substrates presented here were highly correlated with cell activity (R(2)  = 0.9019); with a wavelength of 2.675 µm on the 1300°C surface inducing the highest cell response. © 2015 Wiley Periodicals, Inc.

  1. 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.

  2. Surface deformation during an action potential in pearled cells

    Science.gov (United States)

    Mussel, Matan; Fillafer, Christian; Ben-Porath, Gal; Schneider, Matthias F.

    2017-11-01

    Electric pulses in biological cells (action potentials) have been reported to be accompanied by a propagating cell-surface deformation with a nanoscale amplitude. Typically, this cell surface is covered by external layers of polymer material (extracellular matrix, cell wall material, etc.). It was recently demonstrated in excitable plant cells (Chara braunii) that the rigid external layer (cell wall) hinders the underlying deformation. When the cell membrane was separated from the cell wall by osmosis, a mechanical deformation, in the micrometer range, was observed upon excitation of the cell. The underlying mechanism of this mechanical pulse has, to date, remained elusive. Herein we report that Chara cells can undergo a pearling instability, and when the pearled fragments were excited even larger and more regular cell shape changes were observed (˜10 -100 μ m in amplitude). These transient cellular deformations were captured by a curvature model that is based on three parameters: surface tension, bending rigidity, and pressure difference across the surface. In this paper these parameters are extracted by curve-fitting to the experimental cellular shapes at rest and during excitation. This is a necessary step to identify the mechanical parameters that change during an action potential.

  3. Modelling cell motility and chemotaxis with evolving surface finite elements.

    Science.gov (United States)

    Elliott, Charles M; Stinner, Björn; Venkataraman, Chandrasekhar

    2012-11-07

    We present a mathematical and a computational framework for the modelling of cell motility. The cell membrane is represented by an evolving surface, with the movement of the cell determined by the interaction of various forces that act normal to the surface. We consider external forces such as those that may arise owing to inhomogeneities in the medium and a pressure that constrains the enclosed volume, as well as internal forces that arise from the reaction of the cells' surface to stretching and bending. We also consider a protrusive force associated with a reaction-diffusion system (RDS) posed on the cell membrane, with cell polarization modelled by this surface RDS. The computational method is based on an evolving surface finite-element method. The general method can account for the large deformations that arise in cell motility and allows the simulation of cell migration in three dimensions. We illustrate applications of the proposed modelling framework and numerical method by reporting on numerical simulations of a model for eukaryotic chemotaxis and a model for the persistent movement of keratocytes in two and three space dimensions. Movies of the simulated cells can be obtained from http://homepages.warwick.ac.uk/∼maskae/CV_Warwick/Chemotaxis.html.

  4. Implant Surface Design Regulates Mesenchymal Stem Cell Differentiation and Maturation.

    Science.gov (United States)

    Boyan, B D; Cheng, A; Olivares-Navarrete, R; Schwartz, Z

    2016-03-01

    Changes in dental implant materials, structural design, and surface properties can all affect biological response. While bulk properties are important for mechanical stability of the implant, surface design ultimately contributes to osseointegration. This article reviews the surface parameters of dental implant materials that contribute to improved cell response and osseointegration. In particular, we focus on how surface design affects mesenchymal cell response and differentiation into the osteoblast lineage. Surface roughness has been largely studied at the microscale, but recent studies have highlighted the importance of hierarchical micron/submicron/nanosurface roughness, as well as surface roughness in combination with surface wettability. Integrins are transmembrane receptors that recognize changes in the surface and mediate downstream signaling pathways. Specifically, the noncanonical Wnt5a pathway has been implicated in osteoblastic differentiation of cells on titanium implant surfaces. However, much remains to be elucidated. Only recently have studies been conducted on the differences in biological response to implants based on sex, age, and clinical factors; these all point toward differences that advocate for patient-specific implant design. Finally, challenges in implant surface characterization must be addressed to optimize and compare data across studies. An understanding of both the science and the biology of the materials is crucial for developing novel dental implant materials and surface modifications for improved osseointegration. © International & American Associations for Dental Research 2016.

  5. Robotic Patterning a Superhydrophobic Surface for Collective Cell Migration Screening.

    Science.gov (United States)

    Pang, Yonggang; Yang, Jing; Hui, Zhixin; Grottkau, Brian E

    2018-04-01

    Collective cell migration, in which cells migrate as a group, is fundamental in many biological and pathological processes. There is increasing interest in studying the collective cell migration in high throughput. Cell scratching, insertion blocker, and gel-dissolving techniques are some methodologies used previously. However, these methods have the drawbacks of cell damage, substrate surface alteration, limitation in medium exchange, and solvent interference. The superhydrophobic surface, on which the water contact angle is greater than 150 degrees, has been recently utilized to generate patterned arrays. Independent cell culture areas can be generated on a substrate that functions the same as a conventional multiple well plate. However, so far there has been no report on superhydrophobic patterning for the study of cell migration. In this study, we report on the successful development of a robotically patterned superhydrophobic array for studying collective cell migration in high throughput. The array was developed on a rectangular single-well cell culture plate consisting of hydrophilic flat microwells separated by the superhydrophobic surface. The manufacturing process is robotic and includes patterning discrete protective masks to the substrate using 3D printing, robotic spray coating of silica nanoparticles, robotic mask removal, robotic mini silicone blocker patterning, automatic cell seeding, and liquid handling. Compared with a standard 96-well plate, our system increases the throughput by 2.25-fold and generates a cell-free area in each well non-destructively. Our system also demonstrates higher efficiency than conventional way of liquid handling using microwell plates, and shorter processing time than manual operating in migration assays. The superhydrophobic surface had no negative impact on cell viability. Using our system, we studied the collective migration of human umbilical vein endothelial cells and cancer cells using assays of endpoint

  6. Cell surface heparan sulfate proteoglycans control adhesion and invasion of breast carcinoma cells

    DEFF Research Database (Denmark)

    Lim, Hooi Ching; Multhaupt, Hinke A. B.; Couchman, John R.

    2015-01-01

    Background: Cell surface proteoglycans interact with numerous regulators of cell behavior through their glycosaminoglycan chains. The syndecan family of transmembrane proteoglycans are virtually ubiquitous cell surface receptors that are implicated in the progression of some tumors, including bre...... syndecan-2, may be important regulators of breast carcinoma progression through regulation of cytoskeleton, cell adhesion and invasion.......Background: Cell surface proteoglycans interact with numerous regulators of cell behavior through their glycosaminoglycan chains. The syndecan family of transmembrane proteoglycans are virtually ubiquitous cell surface receptors that are implicated in the progression of some tumors, including...... breast carcinoma. This may derive from their regulation of cell adhesion, but roles for specific syndecans are unresolved. Methods: The MDA-MB231 human breast carcinoma cell line was exposed to exogenous glycosaminoglycans and changes in cell behavior monitored by western blotting, immunocytochemistry...

  7. Co-receptor tropism prediction among 1045 Indian HIV-1 subtype C sequences: Therapeutic implications for India

    Directory of Open Access Journals (Sweden)

    Kuttiatt Vijesh S

    2010-07-01

    Full Text Available Abstract Background Understanding co-receptor tropism of HIV-1 strains circulating in India will provide key analytical leverage for assessing the potential usefulness of newer antiretroviral drugs such as chemokine co-receptor antagonists among Indian HIV-infected populations. The objective of this study was to determine using in silico methods, HIV-1 tropism among a large number of Indian isolates both from primary clinical isolates as well as from database-derived sequences. Results R5-tropism was seen in 96.8% of a total of 1045 HIV-1 subtype C Indian sequences. Co-receptor prediction of 15 primary clinical isolates detected two X4-tropic strains using the C-PSSM matrix. R5-tropic HIV-1 subtype C V3 sequences were conserved to a greater extent than X4-tropic strains. X4-tropic strains were obtained from subjects who had a significantly longer time since HIV diagnosis (96.5 months compared to R5-tropic strains (20.5 months. Conclusions High prevalence of R5 tropism and greater homogeneity of the V3 sequence among HIV-1 subtype C strains in India suggests the potential benefit of CCR5 antagonists as a therapeutic option in India.

  8. Nanoscale crystallinity modulates cell proliferation on plasma sprayed surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Alan M. [School of Applied Sciences, University of Huddersfield, Huddersfield HD1 3DH (United Kingdom); Paxton, Jennifer Z.; Hung, Yi-Pei; Hadley, Martin J.; Bowen, James; Williams, Richard L. [School of Chemical Engineering, University of Birmingham, Edgbaston, B15 2TT (United Kingdom); Grover, Liam M., E-mail: l.m.grover@bham.ac.uk [School of Chemical Engineering, University of Birmingham, Edgbaston, B15 2TT (United Kingdom)

    2015-03-01

    Calcium phosphate coatings have been applied to the surface of metallic prostheses to mediate hard and soft tissue attachment for more than 40 years. Most coatings are formed of high purity hydroxyapatite, and coating methods are often designed to produce highly crystalline surfaces. It is likely however, that coatings of lower crystallinity can facilitate more rapid tissue attachment since the surface will exhibit a higher specific surface area and will be considerably more reactive than a comparable highly crystalline surface. Here we test this hypothesis by growing a population of MC3T3 osteoblast-like cells on the surface of two types of hip prosthesis with similar composition, but with differing crystallinity. The surfaces with lower crystallinity facilitated more rapid cell attachment and increased proliferation rate, despite having a less heterogeneous surface topography. This work highlights that the influence of the crystallinity of HA at the nano-scale is dominant over macro-scale topography for cell adhesion and growth. Furthermore, crystallinity could be easily adjusted by without compromising coating purity. These findings could facilitate designing novel coated calcium phosphate surfaces that more rapidly bond tissue following implantation. - Highlights: • Crystallinity of HA at the nano-scale was dominant over macro-scale topography. • Lower crystallinity caused rapid cell attachment and proliferation rate. • Crystallinity could be easily adjusted by without compromising coating purity.

  9. A case-based reasoning system for genotypic prediction of HIV-1 co-receptor tropism.

    Science.gov (United States)

    Evans, Mark C; Paquet, Agnes C; Huang, Wei; Napolitano, Laura; Frantzell, Arne; Toma, Jonathan; Stawiski, Eric W; Goetz, Matthew Bidwell; Petropoulos, Christos J; Whitcomb, Jeannette; Coakley, Eoin; Haddad, Mojgan

    2013-08-01

    Accurate co-receptor tropism (CRT) determination is critical for making treatment decisions in HIV management. We created a genotypic tropism prediction tool by utilizing the case-based reasoning (CBR) technique that attempts to solve new problems through applying the solution from similar past problems. V3 loop sequences from 732 clinical samples with diverse characteristics were used to build a case library. Additional sequence and molecular properties of the V3 loop were examined and used for similarity assessment. A similarity metric was defined based on each attribute's frequency in the CXCR4-using viruses. We implemented three other genotype-based tropism predictors, support vector machines (SVM), position specific scoring matrices (PSSM), and the 11/25 rule, and evaluated their performance as the ability to predict CRT compared to Monogram's enhanced sensitivity Trofile(®) assay (ESTA). Overall concordance of the CBR based tropism prediction algorithm was 81%, as compared to ESTA. Sensitivity to detect CXCR4 usage was 90% and specificity was at 73%. In comparison, sensitivity of the SVM, PSSM, and the 11/25 rule were 85%, 81%, and 36% respectively while achieving a specificity of 90% by SVM, 75% by PSSM, and 97% by the 11/25 rule. When we evaluated these predictors in an unseen dataset, higher sensitivity was achieved by the CBR algorithm (87%), compared to SVM (82%), PSSM (76%), and the 11/25 rule (33%), while maintaining similar level of specificity. Overall this study suggests that CBR can be utilized as a genotypic tropism prediction tool, and can achieve improved performance in independent datasets compared to model or rule based methods.

  10. Phenotypic Co-receptor Tropism in Perinatally HIV-Infected Youth Failing Antiretroviral Therapy

    Science.gov (United States)

    Agwu, Allison L.; Yao, Tzy-Jyun; Eshleman, Susan H.; Patel, Kunjal; Huang, Wei; Burchett, Sandra K.; Siberry, George K.; Van Dyke, Russell B.

    2016-01-01

    Background Perinatally HIV-infected (PHIV) children and youth are often heavily treatment-experienced, with resultant antiretroviral (ARV) resistance and limited treatment options. For those with virologic failure (VF), new agents such as CCR5 (R5) antagonists may be useful; however, reports of R5 antagonist susceptibility in children have mostly relied on genotypic testing, which may not accurately reflect the phenotypic tropism of the viral populations. We characterized phenotypic co-receptor usage among PHIV children and youth with VF on ARV treatment (ART) to identify predictors of CXCR4 (X4) tropism which preclude R5 antagonist use. Methods Plasma samples with >1,000 HIV RNA copies/mL were obtained from 73 PHIV-infected ART-treated children and youth (age 9–21 years) enrolled in the multi-center Pediatric HIV/AIDS Cohort Study. Samples were analyzed using the Trofile™ phenotypic assay. Multiple logistic regression was performed to identify factors associated with detectable X4-tropism. Results Tropism results were obtained for 59 (81%) of the 73 children and youth; 32 (54%) had X4-tropism. Persistent viremia (≥80% of HIV RNA measurements >400 copies/mL) was associated with detectable X4-tropism (adjusted odds ratio (aOR) 6.6, 95% CI 1.4, 31.4), while longer cumulative nucleoside reverse transcriptase inhibitor (NRTI) use was associated with lower risk of X4-tropism (aOR 0.6, 95% CI 0.5, 0.9). Conclusions Using a phenotypic assay, >50% of PHIV children and youth with VF had X4-tropism, similar to that in treatment-experienced adults, and higher than the 30% reported for children using genotypic assays. Persistent viremia and shorter NRTI exposure are associated with X4-tropism in children and youth and may help target phenotypic testing to those most likely to benefit from R5 antagonist. PMID:27078121

  11. Cell surface carbohydrates as prognostic markers in human carcinomas

    DEFF Research Database (Denmark)

    Dabelsteen, Erik

    1996-01-01

    Tumour development is usually associated with changes in cell surface carbohydrates. These are often divided into changes related to terminal carbohydrate structures, which include incomplete synthesis and modification of normally existing carbohydrates, and changes in the carbohydrate core struc...

  12. Cell/surface interactions on laser micro-textured titanium-coated silicon surfaces.

    Science.gov (United States)

    Mwenifumbo, Steven; Li, Mingwei; Chen, Jianbo; Beye, Aboubaker; Soboyejo, Wolé

    2007-01-01

    This paper examines the effects of nano-scale titanium coatings, and micro-groove/micro-grid patterns on cell/surface interactions on silicon surfaces. The nature of the cellular attachment and adhesion to the coated/uncoated micro-textured surfaces was elucidated by the visualization of the cells and relevant cytoskeletal & focal adhesion proteins through scanning electron microscopy and immunofluorescence staining. Increased cell spreading and proliferation rates are observed on surfaces with 50 nm thick Ti coatings. The micro-groove geometries have been shown to promote contact guidance, which leads to reduced scar tissue formation. In contrast, smooth surfaces result in random cell orientations and the increased possibility of scar tissue formation. Immunofluorescence cell staining experiments also reveal that the actin stress fibers are aligned along the groove dimensions, with discrete focal adhesions occurring along the ridges, within the grooves and at the ends of the cell extensions. The implications of the observed cell/surface interactions are discussed for possible applications of silicon in implantable biomedical systems.

  13. Directing neuronal cell growth on implant material surfaces by microstructuring.

    OpenAIRE

    Reich, Uta; Fadeeva, Elena; Warnecke, Athanasia; Paasche, Gerrit; Müller, Peter; Chichkov, Boris; Stöver, Timo; Lenarz, Thomas; Reuter, Günter

    2012-01-01

    For best hearing sensation, electrodes of auditory prosthesis must have an optimal electrical contact to the respective neuronal cells. To improve the electrode-nerve interface, microstructuring of implant surfaces could guide neuronal cells toward the electrode contact. To this end, femtosecond laser ablation was used to generate linear microgrooves on the two currently relevant cochlear implant materials, silicone elastomer and platinum. Silicone surfaces were structured by two different me...

  14. Electrochemical characterization of the bacterial cell surface

    NARCIS (Netherlands)

    Wal, van der A.

    1996-01-01


    Bacterial cells are ubiquitous in natural environments and also play important roles in domestic and industrial processes. They are found either suspended in the aqueous phase or attached to solid particles. The adhesion behaviour of bacteria is influenced by the physico-chemical

  15. Recent Insights into Cell Surface Heparan Sulphate Proteoglycans and Cancer

    DEFF Research Database (Denmark)

    Couchman, John R; Multhaupt, Hinke; Sanderson, Ralph D

    2016-01-01

    A small group of cell surface receptors are proteoglycans, possessing a core protein with one or more covalently attached glycosaminoglycan chains. They are virtually ubiquitous and their chains are major sites at which protein ligands of many types interact. These proteoglycans can signal...... and regulate important cell processes, such as adhesion, migration, proliferation, and differentiation. Since many protein ligands, such as growth factors, morphogens, and cytokines, are also implicated in tumour progression, it is increasingly apparent that cell surface proteoglycans impact tumour cell...... behaviour. Here, we review some recent advances, emphasising that many tumour-related functions of proteoglycans are revealed only after their modification in processes subsequent to synthesis and export to the cell surface. These include enzymes that modify heparan sulphate structure, recycling of whole...

  16. Multi-scale cell/surface interaction on modified titanium aluminum vanadium surfaces

    Science.gov (United States)

    Chen, Jianbo

    This dissertation presents a series of experimental studies of the effects of multi-scale cell/surface interactions on modified Ti-6Al-4V surfaces. These include laser-grooved surfaces; porous structures and RGD-coated laser-grooved surfaces. A nano-second DPSS UV lasers with a Gaussian pulse energy profile was used to introduce the desired micro-groove geometries onto Ti-6Al-4V surfaces. This was done without inducing micro-cracks or significant changes in surface chemistry within the heat affected zones. The desired 8-12 mum groove depths and widths were achieved by the control of pulse frequency, scan speed, and the lens focal length that controls spot size. The interactions between human osteosarcoma (HOS) cells and laser-grooved Ti-6Al-4V surfaces were investigated after 48 hours of cell culture. The cell behavior, including cell spreading, alignment and adhesion, was elucidated using scanning electronic microscopy (SEM), immuno-fluorescence staining and enzymatic detachment. Contact guidance was shown to increase as grooved spacing decreased. For the range of micro-groove geometries studied, micro-grooves with groove spacings of 20 mum provided the best combination of cell orientation and adhesion. Short-term adhesion experiments (15 mins to 1 day) also revealed that there is a positive correlation between cell orientation and cell adhesion. Contact guidance on the micro-grooved surfaces is shown to be enhanced by nano- and micro-scale asperities that provide sites for the attachment of lamellopodia during cell locomotion and spreading. Contact guidance is also promoted by the geometrical confinement provided by laser grooves. An experimental study of initial cell spreading and ingrowth into Ti-6Al-4V porous structures was also carried out on porous structures with different pore sizes and geometries. A combination of SEM, the tetrazolium salt (MTT) colorimetric assay and enzymatic detachment were used to study cell spreading and adhesion. The extent of cell

  17. Molecular clutch drives cell response to surface viscosity.

    Science.gov (United States)

    Bennett, Mark; Cantini, Marco; Reboud, Julien; Cooper, Jonathan M; Roca-Cusachs, Pere; Salmeron-Sanchez, Manuel

    2018-02-06

    Cell response to matrix rigidity has been explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. Here the molecular clutch model is extended to account for cell interactions with purely viscous surfaces (i.e., without an elastic component). Supported lipid bilayers present an idealized and controllable system through which to study this concept. Using lipids of different diffusion coefficients, the mobility (i.e., surface viscosity) of the presented ligands (in this case RGD) was altered by an order of magnitude. Cell size and cytoskeletal organization were proportional to viscosity. Furthermore, there was a higher number of focal adhesions and a higher phosphorylation of FAK on less-mobile (more-viscous) surfaces. Actin retrograde flow, an indicator of the force exerted on surfaces, was also seen to be faster on more mobile surfaces. This has consequential effects on downstream molecules; the mechanosensitive YAP protein localized to the nucleus more on less-mobile (more-viscous) surfaces and differentiation of myoblast cells was enhanced on higher viscosity. This behavior was explained within the framework of the molecular clutch model, with lower viscosity leading to a low force loading rate, preventing the exposure of mechanosensitive proteins, and with a higher viscosity causing a higher force loading rate exposing these sites, activating downstream pathways. Consequently, the understanding of how viscosity (regardless of matrix stiffness) influences cell response adds a further tool to engineer materials that control cell behavior. Copyright © 2018 the Author(s). Published by PNAS.

  18. Sperm cell surface dynamics during activation and fertilization

    NARCIS (Netherlands)

    Boerke, A.|info:eu-repo/dai/nl/304822922

    2013-01-01

    Before the sperm cell can reach the oocyte it needs to be activated and to undergo a series of preparative steps. The sperm surface dynamics was studied in relation to this activation process and the modifications and removal of sperm surface components havebeen investigated. Bicarbonate-induced

  19. Shape of red blood cells in contact with artificial surfaces.

    Science.gov (United States)

    Grzhibovskis, Richards; Krämer, Elisabeth; Bernhardt, Ingolf; Kemper, Björn; Zanden, Carl; Repin, Nikolay V; Tkachuk, Bogdan V; Voinova, Marina V

    2017-03-01

    The phenomenon of physical contact between red blood cells and artificial surfaces is considered. A fully three-dimensional mathematical model of a bilayer membrane in contact with an artificial surface is presented. Numerical results for the different geometries and adhesion intensities are found to be in agreement with experimentally observed geometries obtained by means of digital holographic microscopy.

  20. Nanometer polymer surface features: the influence on surface energy, protein adsorption and endothelial cell adhesion

    Science.gov (United States)

    Carpenter, Joseph; Khang, Dongwoo; Webster, Thomas J.

    2008-12-01

    Current small diameter (lactic-co-glycolic acid) (PLGA) surfaces elevated endothelial cell adhesion, proliferation, and extracellular matrix synthesis when compared to nanosmooth surfaces. Nonetheless, these studies failed to address the importance of lateral and vertical surface feature dimensionality coupled with surface free energy; nor did such studies elicit an optimum specific surface feature size for promoting endothelial cell adhesion. In this study, a series of highly ordered nanometer to submicron structured PLGA surfaces of identical chemistry were created using a technique employing polystyrene nanobeads and poly(dimethylsiloxane) (PDMS) molds. Results demonstrated increased endothelial cell adhesion on PLGA surfaces with vertical surface features of size less than 18.87 nm but greater than 0 nm due to increased surface energy and subsequently protein (fibronectin and collagen type IV) adsorption. Furthermore, this study provided evidence that the vertical dimension of nanometer surface features, rather than the lateral dimension, is largely responsible for these increases. In this manner, this study provides key design parameters that may promote vascular graft efficacy.

  1. Expanding the diversity of unnatural cell surface sialic acids

    Energy Technology Data Exchange (ETDEWEB)

    Luchansky, Sarah J.; Goon, Scarlett; Bertozzi, Carolyn R.

    2003-10-30

    Novel chemical reactivity can be introduced onto cell surfaces through metabolic oligosaccharide engineering. This technique exploits the substrate promiscuity of cellular biosynthetic enzymes to deliver unnatural monosaccharides bearing bioorthogonal functional groups into cellular glycans. For example, derivatives of N-acetylmannosamine (ManNAc) are converted by the cellular biosynthetic machinery into the corresponding sialic acids and subsequently delivered to the cell surface in the form of sialoglycoconjugates. Analogs of N-acetylglucosamine (GlcNAc) and N-acetylgalactosamine (GalNAc) are also metabolized and incorporated into cell surface glycans, likely through the sialic acid and GalNAc salvage pathways, respectively. Furthermore, GlcNAc analogs can be incorporated into nucleocytoplasmic proteins in place of {beta}-O-GlcNAc residues. These pathways have been exploited to integrate unique electrophiles such as ketones and azides into the target glycoconjugate class. These functional groups can be further elaborated in a chemoselective fashion by condensation with hydrazides and by Staudinger ligation, respectively, thereby introducing detectable probes onto the cell. In conclusion, sialic acid derivatives are efficient vehicles for delivery of bulky functional groups to cell surfaces and masking of their hydroxyl groups improves their cellular uptake and utilization. Furthermore, the successful introduction of photoactivatable aryl azides into cell surface glycans opens up new avenues for studying sialic acid-binding proteins and elucidating the role of sialic acid in essential processes such as signaling and cell adhesion.

  2. Responses of fibroblasts and glial cells to nanostructured platinum surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Pennisi, C P; Sevcencu, C; Yoshida, K [Center for Sensory-Motor Interaction (SMI), Aalborg University, Aalborg (Denmark); Dolatshahi-Pirouz, A; Foss, M; Larsen, A Nylandsted; Besenbacher, F [Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Aarhus (Denmark); Hansen, J Lundsgaard [Department of Physics and Astronomy, Aarhus University, Aarhus (Denmark); Zachar, V, E-mail: cpennisi@hst.aau.d [Laboratory for Stem Cell Research, Aalborg University (Denmark)

    2009-09-23

    The chronic performance of implantable neural prostheses is affected by the growth of encapsulation tissue onto the stimulation electrodes. Encapsulation is associated with activation of connective tissue cells at the electrode's metallic contacts, usually made of platinum. Since surface nanotopography can modulate the cellular responses to materials, the aim of the present work was to evaluate the 'in vitro' responses of connective tissue cells to platinum strictly by modulating its surface nanoroughness. Using molecular beam epitaxy combined with sputtering, we produced platinum nanostructured substrates consisting of irregularly distributed nanopyramids and investigated their effect on the proliferation, cytoskeletal organization and cellular morphology of primary fibroblasts and transformed glial cells. Cells were cultured on these substrates and their responses to surface roughness were studied. After one day in culture, the fibroblasts were more elongated and their cytoskeleton less mature when cultured on rough substrates. This effect increased as the roughness of the surface increased and was associated with reduced cell proliferation throughout the observation period (4 days). Morphological changes also occurred in glial cells, but they were triggered by a different roughness scale and did not affect cellular proliferation. In conclusion, surface nanotopography modulates the responses of fibroblasts and glial cells to platinum, which may be an important factor in optimizing the tissue response to implanted neural electrodes.

  3. Responses of fibroblasts and glial cells to nanostructured platinum surfaces

    Science.gov (United States)

    Pennisi, C. P.; Sevcencu, C.; Dolatshahi-Pirouz, A.; Foss, M.; Lundsgaard Hansen, J.; Nylandsted Larsen, A.; Zachar, V.; Besenbacher, F.; Yoshida, K.

    2009-09-01

    The chronic performance of implantable neural prostheses is affected by the growth of encapsulation tissue onto the stimulation electrodes. Encapsulation is associated with activation of connective tissue cells at the electrode's metallic contacts, usually made of platinum. Since surface nanotopography can modulate the cellular responses to materials, the aim of the present work was to evaluate the 'in vitro' responses of connective tissue cells to platinum strictly by modulating its surface nanoroughness. Using molecular beam epitaxy combined with sputtering, we produced platinum nanostructured substrates consisting of irregularly distributed nanopyramids and investigated their effect on the proliferation, cytoskeletal organization and cellular morphology of primary fibroblasts and transformed glial cells. Cells were cultured on these substrates and their responses to surface roughness were studied. After one day in culture, the fibroblasts were more elongated and their cytoskeleton less mature when cultured on rough substrates. This effect increased as the roughness of the surface increased and was associated with reduced cell proliferation throughout the observation period (4 days). Morphological changes also occurred in glial cells, but they were triggered by a different roughness scale and did not affect cellular proliferation. In conclusion, surface nanotopography modulates the responses of fibroblasts and glial cells to platinum, which may be an important factor in optimizing the tissue response to implanted neural electrodes.

  4. Directing neuronal cell growth on implant material surfaces by microstructuring.

    Science.gov (United States)

    Reich, Uta; Fadeeva, Elena; Warnecke, Athanasia; Paasche, Gerrit; Müller, Peter; Chichkov, Boris; Stöver, Timo; Lenarz, Thomas; Reuter, Günter

    2012-05-01

    For best hearing sensation, electrodes of auditory prosthesis must have an optimal electrical contact to the respective neuronal cells. To improve the electrode-nerve interface, microstructuring of implant surfaces could guide neuronal cells toward the electrode contact. To this end, femtosecond laser ablation was used to generate linear microgrooves on the two currently relevant cochlear implant materials, silicone elastomer and platinum. Silicone surfaces were structured by two different methods, either directly, by laser ablation or indirectly, by imprinting using laser-microstructured molds. The influence of surface structuring on neurite outgrowth was investigated utilizing a neuronal-like cell line and primary auditory neurons. The pheochromocytoma cell line PC-12 and primary spiral ganglion cells were cultured on microstructured auditory implant materials. The orientation of neurite outgrowth relative to the microgrooves was determined. Both cell types showed a preferred orientation in parallel to the microstructures on both, platinum and on molded silicone elastomer. Interestingly, microstructures generated by direct laser ablation of silicone did not influence the orientation of either cell type. This shows that differences in the manufacturing procedures can affect the ability of microstructured implant surfaces to guide the growth of neurites. This is of particular importance for clinical applications, since the molding technique represents a reproducible, economic, and commercially feasible manufacturing procedure for the microstructured silicone surfaces of medical implants. Copyright © 2012 Wiley Periodicals, Inc.

  5. Lactoperoxidase catalyzed radioiodination of cell surface immunoglobulin: incorporated radioactivity may not reflect relative cell surface Ig density

    International Nuclear Information System (INIS)

    Wilder, R.L.; Yuen, C.C.; Mage, R.G.

    1979-01-01

    Rabbit and mouse splenic lymphocytes were radioiodinated by the lactoperoxidase technique, extracted with non-ionic detergent, immunoprecipitated with high titered rabbit anti-kappa antisera, and compared by SDS-PAGE. Mouse sIg peaks were reproducibly larger in size than rabbit sIg peaks (often greater than 10 times). Neither differences in incorporation of label into the rabbit cell surface, nor differences in average sIg density explain this result. Total TCA-precipitable radioactivity was similar in each species. Estimation of the relative amounts of sIg in the mouse and rabbit showed similar average sIg densities. Differences in detergent solubility, proteolytic lability, or antisera used also do not adequately account for this difference. Thus, these data indicate that radioactivity incorporated after lactoperoxidase catalyzed cell surface radioiodination may not reflect cell surface Ig density. Conclusions about cell surface density based upon relative incorporation of radioactivity should be confirmed by other approaches

  6. Cell Surface Heparan Sulfate Released by Heparanase Promotes Melanoma Cell Migration and Angiogenesis

    Science.gov (United States)

    Roy, Madhuchhanda; Marchetti, Dario

    2009-01-01

    Heparan sulfate proteoglycans are essential components of the cell-surface and extracellular matrix which provide structural integrity and act as storage depots for growth factors and chemokines, through their heparan sulfate (HS) side chains. Heparanase is the only mammalian endoglycosidase known that cleaves HS, thus contributing to matrix degradation and cell invasion. The enzyme acts as an endo-β-D-glucuronidase resulting in HS fragments of discrete molecular weight size. Cell-surface HS is known to inhibit or stimulate tumorigenesis depending upon size and composition. We hypothesized that heparanase contributes to melanoma metastasis by generating bioactive HS from the cell-surface to facilitate biological activities of tumor cells as well as tumor microenvironment. We removed cell-surface HS from melanoma (B16B15b) by HPSE treatment and resulting fragments were isolated. Purified cell-surface HS stimulated in vitro B16B15b cell migration but not proliferation, and importantly, enhanced in vivo angiogenesis. Furthermore, melanoma cell-surface HS did not affect in vitro endothelioma cell (b.End3) migration. Our results provide direct evidence that, in addition to remodeling extracellular matrix and releasing growth factors and chemokines, HPSE contributes to aggressive phenotype of melanoma by releasing bioactive cell-surface HS fragments which can stimulate melanoma cell migration in vitro and angiogenesis in vivo. PMID:19115257

  7. Standing surface acoustic wave (SSAW) based multichannel cell sorting.

    Science.gov (United States)

    Ding, Xiaoyun; Lin, Sz-Chin Steven; Lapsley, Michael Ian; Li, Sixing; Guo, Xiang; Chan, Chung Yu; Chiang, I-Kao; Wang, Lin; McCoy, J Philip; Huang, Tony Jun

    2012-11-07

    We introduce a novel microfluidic device for cell sorting in continuous flow using tunable standing surface acoustic waves. This method allows individual cells to be precisely directed into five different outlet channels in a single step. It is versatile, simple, label-free, non-invasive, and highly controllable.

  8. Biomimetic surface modification of titanium surfaces for early cell capture by advanced electrospinning

    International Nuclear Information System (INIS)

    Ravichandran, Rajeswari; Raghunath, Michael; Chan, Casey K; Ng, Clarisse CH; Liao, Susan; Pliszka, Damian; Ramakrishna, S

    2012-01-01

    The time required for osseointegration with a metal implant having a smooth surface ranges from three to six months. We hypothesized that biomimetic coating surfaces with poly(lactic-co-glycolic acid) (PLGA)/collagen fibers and nano-hydroxyapatite (n-HA) on the implant would enhance the adhesion of mesenchymal stem cells. Therefore, this surface modification of dental and bone implants might enhance the process of osseointegration. In this study, we coated PLGA or PLGA/collagen (50:50 w/w ratio) fiber on Ti disks by modified electrospinning for 5 s to 2 min; after that, we further deposited n-HA on the fibers. PLGA fibers of fiber diameter 0.957 ± 0.357 µm had a contact angle of 9.9 ± 0.3° and PLGA/collagen fibers of fiber diameter 0.378 ± 0.068 µm had a contact angle of 0°. Upon n-HA incorporation, all the fibers had a contact angle of 0° owing to the hydrophilic nature of n-HA biomolecule. The cell attachment efficiency was tested on all the scaffolds for different intervals of time (10, 20, 30 and 60 min). The alkaline phosphatase activity, cell proliferation and mineralization were analyzed on all the implant surfaces on days 7, 14 and 21. Results of the cell adhesion study indicated that the cell adhesion was maximum on the implant surface coated with PLGA/collagen fibers deposited with n-HA compared to the other scaffolds. Within a short span of 60 min, 75% of the cells adhered onto the mineralized PLGA/collagen fibers. Similarly by day 21, the rate of cell proliferation was significantly higher (p ≤ 0.05) on the mineralized PLGA/collagen fibers owing to enhanced cell adhesion on these fibers. This enhanced initial cell adhesion favored higher cell proliferation, differentiation and mineralization on the implant surface coated with mineralized PLGA/collagen fibers.

  9. Surface strategies for control of neuronal cell adhesion: A review

    Science.gov (United States)

    Roach, P.; Parker, T.; Gadegaard, N.; Alexander, M. R.

    2010-06-01

    Material engineering methods have been used for many years to develop biomedical devices for use within the body to augment, repair or replace damaged tissues ranging from contact lenses to heart valves. Here we review the findings gathered from the wide and varied surface analytical approaches applied to study the interaction between biology and man-made materials. The key material characteristics identified to be important for biological recognition are surface chemistry, topography and compliance. Model surfaces with controlled chemistry and topography have provided insight into biological response to various types of topographical features over a wide range of length scales from nano to micrometres, along with 3D matrices that have been used as scaffolds to support cells for tissue formation. The cellular response to surfaces with localised areas of patterned chemistry and to those presenting gradually changing chemistry are discussed. Where previous reviews have been structured around specific classes of surface modification, e.g. self-assembly, or have broadly examined the response of various cells to numerous surfaces, we aim in this article to focus in particular on the tissues involved in the nervous system whilst providing a broad overview of key issues from the field of cell and protein surface interactions with surfaces. The goal of repair and treatment of diseases related to the central and peripheral nervous systems rely on understanding the local interfacial environment and controlling responses at the cellular level. The role of the protein layer deposited from serum containing media onto man-made surfaces is discussed. We highlight the particular problems associated with the repair of the nervous system, and review how neuronal attachment and axon guidance can be accomplished using various surface cues when cultured with single and multiple cell types. We include a brief glossary of techniques discussed in the body of this article aimed at the

  10. Influence of engineered surface on cell directionality and motility

    International Nuclear Information System (INIS)

    Tang, Qing Yuan; Pang, Stella W; Tong, Wing Yin; Shi, Peng; Lam, Yun Wah; Shi, Jue

    2014-01-01

    Control of cell migration is important in numerous key biological processes, and is implicated in pathological conditions such as cancer metastasis and inflammatory diseases. Many previous studies indicated that cell migration could be guided by micropatterns fabricated on cell culture surfaces. In this study, we designed a polydimethylsiloxane cell culture substrate with gratings punctuated by corners and ends, and studied its effects on the behavior of MC3T3-E1 osteoblast cells. MC3T3-E1 cells elongated and aligned with the gratings, and the migration paths of the cells appeared to be guided by the grating pattern. Interestingly, more than 88% of the cells cultured on these patterns were observed to reverse their migration directions at least once during the 16 h examination period. Most of the reversal events occurred at the corners and the ends of the pattern, suggesting these localized topographical features induce an abrupt loss in directional persistence. Moreover, the cell speed was observed to increase temporarily right after each directional reversal. Focal adhesion complexes were more well-established in cells on the angular gratings than on flat surfaces, but the formation of filipodia appeared to be imbalanced at the corners and the ends, possibly leading to the loss of directional persistence. This study describes the first engineered cell culture surface that consistently induces changes in the directional persistence of adherent cells. This will provide an experimental model for the study of this phenomenon and a valuable platform to control the cell motility and directionality, which can be used for cell screening and selection. (paper)

  11. Acetylation of the Cd8 Locus by KAT6A Determines Memory T Cell Diversity

    Directory of Open Access Journals (Sweden)

    Dane M. Newman

    2016-09-01

    Full Text Available How functionally diverse populations of pathogen-specific killer T cells are generated during an immune response remains unclear. Here, we propose that fine-tuning of CD8αβ co-receptor levels via histone acetylation plays a role in lineage fate. We show that lysine acetyltransferase 6A (KAT6A is responsible for maintaining permissive Cd8 gene transcription and enabling robust effector responses during infection. KAT6A-deficient CD8+ T cells downregulated surface CD8 co-receptor expression during clonal expansion, a finding linked to reduced Cd8α transcripts and histone-H3 lysine 9 acetylation of the Cd8 locus. Loss of CD8 expression in KAT6A-deficient T cells correlated with reduced TCR signaling intensity and accelerated contraction of the effector-like memory compartment, whereas the long-lived memory compartment appeared unaffected, a result phenocopied by the removal of the Cd8 E8I enhancer element. These findings suggest a direct role of CD8αβ co-receptor expression and histone acetylation in shaping functional diversity within the cytotoxic T cell pool.

  12. Cell surface topology creates high Ca2+ signalling microdomains

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Olsen, Lars Folke; Hallett, Maurice B

    2010-01-01

    20 microM Ca2+ to be activated. In this paper, we have resolved this apparent paradox by showing that the surface topology of cells represents an important and hitherto unrecognized feature for generating microdomains of high Ca2+ in cells. We show that whereas the standard modeling assumption...... of a smooth cell surface predicts only moderate localized effects, the more realistic "wrinkled" surface topology predicts that Ca2+ concentrations up to 80 microM can persist within the folds of membranes for significant times. This intra-wrinkle location may account for 5% of the total cell volume. Using......-wrinkle location is also a strategic location at which Ca2+ acts as a regulator of the cortical cytoskeleton and plasma membrane expansion....

  13. Cell adhesion and growth on ion-implanted polymer surface

    International Nuclear Information System (INIS)

    Lee, Jae-Suk; Kaibara, M.; Iwaki, M.; Sasabe, H.; Suzuki, Y.; Kusakabe, M.

    1992-01-01

    The adhesion and growth of endothelial cells on ion-implanted polystyrene and segmented polyurethane surface were investigated. Ions of Na + , N 2 + , O 2 + , Ar + and Kr + were implanted to the polymer surface with ion fluences between 1 x 10 15 and 3 x 10 17 ions/cm 2 at energy of 150 KeV at room temperature. Ion-implanted polymers were characterized by FT-IR-ATR an Raman spectroscopies. The adhesion and proliferation of bovine aorta endothelial cells on ion-implanted polymer surface were observed by an optical microscope. The rate of growth of BAECs on ion-implanted PSt was faster than that on non-implanted PSt. Complete cell adhesion and growth were observed on ion-implanted SPU, whereas the adhesion and growth of BAECs on the non-implanted SPU was not observed. It was attempted to control the cell culture on the ion-implanted domain fabricated using a mask. (author)

  14. Full Length Amelogenin Binds to Cell Surface LAMP-1 on Tooth Root/Periodontium Associated Cells

    Science.gov (United States)

    Zhang, Hai; Tompkins, Kevin; Garrigues, Jacques; Snead, Malcolm L.; Gibson, Carolyn W.; Somerman, Martha J.

    2010-01-01

    Objectives Lysosome-associated membrane protein-1 (LAMP-1) has been suggested to be a cell surface receptor for a specific amelogenin isoform, leucine-rich amelogenin peptide or LRAP. However, it is unclear if LAMP-1 is an amelogenin receptor for dental mesenchymal cells. The goal of this study was to determine if LAMP-1 serves as a cell surface binding site for full length amelogenin on tooth root/periodontium associated mesenchymal cells. Design Murine dental follicle cells and cementoblasts (OCCM-30) were cultured for 2 days followed by addition of full length recombinant mouse amelogenin, rp(H)M180. Dose-response (0 to 100 μg/ml) and time course (0 to 120 minutes) assays were performed to determine the optimal conditions for live cell surface binding using immuno-fluorescent microscopy. A competitive binding assay was performed to determine binding specificity by adding Emdogain (1 mg/ml) to the media. An antibody against LAMP-1 was used to detect the location of LAMP-1 on the cell surface and the pattern was compared to cell surface bound amelogenin. Both amelogenin and cell surface LAMP-1 were immuno-co-localized to compare the amount and distribution pattern. Results Maximum surface binding was achieved with 50 μg/ml of rp(H)M180 for 120 minutes. This binding was specific as demonstrated by competitive inhibition (79% lower) with the addition of Emdogain. The binding pattern for rp(H)M180 was similar to the distribution of surface LAMP-1 on dental follicle cells and cementoblasts. The high co-localization coefficient (0.92) for rp(H)M180 and LAMP-1 supports rp(H)M180 binding to cell surface LAMP-1. Conclusions The data from this study suggest that LAMP-1 can serve as a cell surface binding site for amelogenin on dental follicle cells and cementoblasts. PMID:20382373

  15. Surface Passivation Studies on n+pp+ Bifacial Solar Cell

    Directory of Open Access Journals (Sweden)

    Suhaila Sepeai

    2012-01-01

    Full Text Available Bifacial solar cell is a specially designed solar cell for the production of electricity from both sides of the solar cell. It is an active field of research to make photovoltaics (PV more competitive by increasing its efficiency and lowering its costs. We developed an n+pp+ structure for the bifacial solar cell. The fabrication used phosphorus-oxy-trichloride (POCl3 diffusion to form the emitter and Al diffusion using conventional screen printing to produce the back surface field (BSF. The n+pp+ bifacial solar cell was a sandwiched structure of antireflective coatings on both sides, Argentum (Ag as a front contact and Argentum/Aluminum (Ag/Al as a back contact. This paper reports the solar cell performance with different surface passivation or antireflecting coatings (ARC. Silicon nitride (SiN deposited by Plasma-Enhanced Chemical Vapor Deposition (PECVD, thermally grown silicon dioxide (SiO2, PECVD-SiO2, and SiO2/SiN stack were used as ARC. The efficiency obtained for the best bifacial solar cell having SiN as the ARC is 8.32% for front surface illumination and 3.21% for back surface illumination.

  16. Cell-Surface Proteomics Identifies Lineage-Specific Markers of Embryo-Derived Stem Cells

    OpenAIRE

    Rugg-Gunn, Peter J.; Cox, Brian J.; Lanner, Fredrik; Sharma, Parveen; Ignatchenko, Vladimir; McDonald, Angela C.H.; Garner, Jodi; Gramolini, Anthony O.; Rossant, Janet; Kislinger, Thomas

    2012-01-01

    Summary The advent of reprogramming and its impact on stem cell biology has renewed interest in lineage restriction in mammalian embryos, the source of embryonic (ES), epiblast (EpiSC), trophoblast (TS), and extraembryonic endoderm (XEN) stem cell lineages. Isolation of specific cell types during stem cell differentiation and reprogramming, and also directly from embryos, is a major technical challenge because few cell-surface proteins are known that can distinguish each cell type. We provide...

  17. Anisotropic cell growth-regulated surface micropatterns in flower petals

    Directory of Open Access Journals (Sweden)

    Xiao Huang

    2017-05-01

    Full Text Available Flower petals have not only diverse macroscopic morphologies but are rich in microscopic surface patterns, which are crucial to their biological functions. Both experimental measurements and theoretical analysis are conducted to reveal the physical mechanisms underlying the formation of minute wrinkles on flower petals. Three representative flowers, daisy, kalanchoe blossfeldiana, and Eustoma grandiflorum, are investigated as examples. A surface wrinkling model, incorporating the measured mechanical properties and growth ratio, is used to elucidate the difference in their surface morphologies. The mismatch between the anisotropic epidermal cell growth and the isotropic secretion of surficial wax is found to dictate the surface patterns.

  18. Cell surface differences of Naegleria fowleri and Naegleria lovaniensis exposed with surface markers.

    Science.gov (United States)

    González-Robles, Arturo; Castañón, Guadalupe; Cristóbal-Ramos, Ana Ruth; Hernández-Ramírez, Verónica Ivonne; Omaña-Molina, Maritza; Martínez-Palomo, Adolfo

    2007-12-01

    Differences in the distribution of diverse cell surface coat markers were found between Naegleria fowleri and Naegleria lovaniensis. The presence of carbohydrate-containing components in the cell coat of the two species was detected by selective staining with ruthenium red and alcian blue. Using both markers, N. fowleri presented a thicker deposit than N. lovaniensis. The existence of exposed mannose or glucose residues was revealed by discriminatory agglutination with the plant lectin Concanavalin A. These sugar residues were also visualized at the cell surface of these parasites either by transmission electron microscopy or by fluorescein-tagged Concanavalin A. Using this lectin cap formation was induced only in N. fowleri. The anionic sites on the cell surface detected by means of cationized ferritin were more apparent in N. fowleri. Biotinylation assays confirmed that even though the two amoebae species have some analogous plasma membrane proteins, there is a clear difference in their composition.

  19. A conserved aspartic acid is important for agonist (VUAA1 and odorant/tuning receptor-dependent activation of the insect odorant co-receptor (Orco.

    Directory of Open Access Journals (Sweden)

    Brijesh N Kumar

    Full Text Available Insect odorant receptors function as heteromeric odorant-gated cation channels comprising a conventional odorant-sensitive tuning receptor, and a conserved co-receptor (Orco. An Orco agonist, VUAA1, is able to activate both heteromeric and homomeric Orco-containing channels. Very little is known about specific residues in Orco that contribute to cation permeability and gating. We investigated the importance of two conserved Asp residues, one in each of transmembrane domains 5 and 7, for channel function by mutagenesis. Drosophila melanogaster Orco and its substitution mutants were expressed in HEK cells and VUAA1-stimulated channel activity was determined by Ca(2+ influx and whole-cell patch clamp electrophysiology. Substitution of D466 in transmembrane 7 with amino acids other than glutamic acid resulted in a substantial reduction in channel activity. The D466E Orco substitution mutant was ~2 times more sensitive to VUAA1. The permeability of the D466E Orco mutant to cations was unchanged relative to wild-type Orco. When D466E Orco is co-expressed with a conventional tuning odorant receptor, the heteromeric complex also shows increased sensitivity to an odorant. Thus, the effect of the D466E mutation is not specific to VUAA1 agonism or dependent on homomeric Orco assembly. We suggest the gain-of-activation characteristic of the D466E mutant identifies an amino acid that is likely to be important for activation of both heteromeric and homomeric insect odorant receptor channels.

  20. Micropatterned Azopolymer Surfaces Modulate Cell Mechanics and Cytoskeleton Structure.

    Science.gov (United States)

    Rianna, Carmela; Ventre, Maurizio; Cavalli, Silvia; Radmacher, Manfred; Netti, Paolo A

    2015-09-30

    Physical and chemical characteristics of materials are important regulators of cell behavior. In particular, cell elasticity is a fundamental parameter that reflects the state of a cell. Surface topography finely modulates cell fate and function via adhesion mediated signaling and cytoskeleton generated forces. However, how topographies alter cell mechanics is still unclear. In this work we have analyzed the mechanical properties of peripheral and nuclear regions of NIH-3T3 cells on azopolymer substrates with different topographic patterns. Micrometer scale patterns in the form of parallel ridges or square lattices of surface elevations were encoded on light responsive azopolymer films by means of contactless optical methods. Cell mechanics was investigated by atomic force microscopy (AFM). Cells and consequently the cell cytoskeleton were oriented along the linear patterns affecting cytoskeletal structures, e.g., formation of actin stress fibers. Our data demonstrate that topographic substrate patterns are recognized by cells and mechanical information is transferred by the cytoskeleton. Furthermore, cytoskeleton generated forces deform the nucleus, changing its morphology that appears to be related to different mechanical properties in the nuclear region.

  1. Forces due to surface water measured by force microscopy. Consequences for anchoring biological cells to surfaces

    International Nuclear Information System (INIS)

    Schilcher, K.

    1997-05-01

    Interaction forces in 'Scanning Force Microscopy' (SFM). Force curves revealed exponentially decaying, attractive forces between silicon tip and silicon sample in aqueous media. Replacing the silicon sample by a sheet of mica, the interaction forces had both, an attractive and a repulsive component. Addition of salts generally reduced the forces. At 500 mM salt concentration, the attractive force became quantized with a residual force value of 23 pN. The attractive force is attributed to the gain in energy of water molecules which are released from surface water into free water during tip-sample approach. This conclusion is supported by a statistical model. The repulsive force contribution in the case of mica, is caused by hydration forces due to the spatial organization of crystalline water on the mica surface. Anchoring of biological cells. Molecular resolution of cell surfaces by SFM requires cell anchoring without interference with cell physiology. For this a novel strategy, 'hydrophobic anchoring' was designed. It avoids strong attractive forces between cell and by using a flexible spacer molecule. It establishes anchoring by a lipid (bound to the spacer), which weakly interacts with the hydrophobic core of the cell membrane. The method was subjected to tests using RBL-2H3, CH0 αβ and HEK-293 cells. The strength of cell anchoring was assayed by shear forces. In all cases 'hydrophobic anchoring' via a spacer caused elective anchoring much beyond controls. Such cell anchoring was employed for the imaging of RBL-2H3 cells by SFM. Images showed considerable finer details than images of loosely adsorbed cells. With about 50 rim resolution, SFM succeeded in imaging microvilli, filopodia, single cytoskeletal fibers (microtubules, microfilaments) and vesicles. In addition, as a consequence of cell stimulation upon ionomycin treatment, lamellae formation and the appearance of secretory granules on top of them were observed which indicates the viability of anchored

  2. Application of various surface passivation layers in solar cells

    International Nuclear Information System (INIS)

    Lee, Ji Youn; Lee, Soo Hong

    2004-01-01

    In this work, we have used different techniques for surface passivation: conventional thermal oxidation (CTO), rapid thermal oxidation (RTO), and plasma-enhanced chemical vapour deposition (PECVD). The surface passivation qualities of eight different single and combined double layers have been investigated both on phosphorus non-diffused p-type Float Zone (FZ) silicon wafers and on diffused emitters (100 Ω/□ and 40 Ω/□). CTO/SiN 1 passivates very well not only on a non-diffused surface (τ eff = 1361 μs) but also on an emitter (τ eff = 414 μs). However, we concluded that RTO/SiN 1 and RTO/SiN 2 stacks were more suitable than CTO/SiN stacks for surface passivation in solar cells since those stacks had relatively good passivation qualities and suitable optical reflections. RTO/SiN 1 for rear-surface passivation and RTO/SiN 2 for front-surface passivation were applied to the fabrication of solar cells. We achieved efficiencies of 18.5 % and 18.8 % on 0.5 Ω-cm (FZ) silicon with planar and textured front surfaces, respectively. An excellent open circuit voltage (V oc ) of 675.6 mV was obtained for the planar cell.

  3. Development of exosome surface display technology in living human cells

    Energy Technology Data Exchange (ETDEWEB)

    Stickney, Zachary, E-mail: zstickney@scu.edu; Losacco, Joseph, E-mail: jlosacco@scu.edu; McDevitt, Sophie, E-mail: smmcdevitt@scu.edu; Zhang, Zhiwen, E-mail: zzhang@scu.edu; Lu, Biao, E-mail: blu2@scu.edu

    2016-03-25

    Surface display technology is an emerging key player in presenting functional proteins for targeted drug delivery and therapy. Although a number of technologies exist, a desirable mammalian surface display system is lacking. Exosomes are extracellular vesicles that facilitate cell–cell communication and can be engineered as nano-shuttles for cell-specific delivery. In this study, we report the development of a novel exosome surface display technology by exploiting mammalian cell secreted nano-vesicles and their trans-membrane protein tetraspanins. By constructing a set of fluorescent reporters for both the inner and outer surface display on exosomes at two selected sites of tetraspanins, we demonstrated the successful exosomal display via gene transfection and monitoring fluorescence in vivo. We subsequently validated our system by demonstrating the expected intracellular partitioning of reporter protein into sub-cellular compartments and secretion of exosomes from human HEK293 cells. Lastly, we established the stable engineered cells to harness the ability of this robust system for continuous production, secretion, and uptake of displayed exosomes with minimal impact on human cell biology. In sum, our work paved the way for potential applications of exosome, including exosome tracking and imaging, targeted drug delivery, as well as exosome-mediated vaccine and therapy.

  4. Investigation of the Cell Surface Proteome of Human Periodontal Ligament Stem Cells.

    Science.gov (United States)

    Xiong, Jimin; Menicanin, Danijela; Zilm, Peter S; Marino, Victor; Bartold, P Mark; Gronthos, Stan

    2016-01-01

    The present study examined the cell surface proteome of human periodontal ligament stem cells (PDLSC) compared to human fibroblasts. Cell surface proteins were prelabelled with CyDye before processing to extract the membrane lysates, which were separated using 2D electrophoresis. Selected differentially expressed protein "spots" were identified using Mass spectrometry. Four proteins were selected for validation: CD73, CD90, Annexin A2, and sphingosine kinase 1 previously associated with mesenchymal stem cells. Flow cytometric analysis found that CD73 and CD90 were highly expressed by human PDLSC and gingival fibroblasts but not by keratinocytes, indicating that these antigens could be used as potential markers for distinguishing between mesenchymal cells and epithelial cell populations. Annexin A2 was also found to be expressed at low copy number on the cell surface of human PDLSC and gingival fibroblasts, while human keratinocytes lacked any cell surface expression of Annexin A2. In contrast, sphingosine kinase 1 expression was detected in all the cell types examined using immunocytochemical analysis. These proteomic studies form the foundation to further define the cell surface protein expression profile of PDLSC in order to better characterise this cell population and help develop novel strategies for the purification of this stem cell population.

  5. Investigation of the Cell Surface Proteome of Human Periodontal Ligament Stem Cells

    Directory of Open Access Journals (Sweden)

    Jimin Xiong

    2016-01-01

    Full Text Available The present study examined the cell surface proteome of human periodontal ligament stem cells (PDLSC compared to human fibroblasts. Cell surface proteins were prelabelled with CyDye before processing to extract the membrane lysates, which were separated using 2D electrophoresis. Selected differentially expressed protein “spots” were identified using Mass spectrometry. Four proteins were selected for validation: CD73, CD90, Annexin A2, and sphingosine kinase 1 previously associated with mesenchymal stem cells. Flow cytometric analysis found that CD73 and CD90 were highly expressed by human PDLSC and gingival fibroblasts but not by keratinocytes, indicating that these antigens could be used as potential markers for distinguishing between mesenchymal cells and epithelial cell populations. Annexin A2 was also found to be expressed at low copy number on the cell surface of human PDLSC and gingival fibroblasts, while human keratinocytes lacked any cell surface expression of Annexin A2. In contrast, sphingosine kinase 1 expression was detected in all the cell types examined using immunocytochemical analysis. These proteomic studies form the foundation to further define the cell surface protein expression profile of PDLSC in order to better characterise this cell population and help develop novel strategies for the purification of this stem cell population.

  6. Adhesion defective BHK cell mutant has cell surface heparan sulfate proteoglycan of altered properties

    DEFF Research Database (Denmark)

    Couchman, J R; Austria, R; Woods, A

    1988-01-01

    In the light of accumulating data that implicate cell surface heparan sulfate proteoglycans (HSPGs) with a role in cell interactions with extracellular matrix molecules such as fibronectin, we have compared the properties of these molecules in wild-type BHK cells and an adhesion-defective ricin......-resistant mutant (RicR14). Our results showed that the mutant, unlike BHK cells, cannot form focal adhesions when adherent to planar substrates in the presence of serum. Furthermore, while both cell lines possess similar amounts of cell surface HSPG with hydrophobic properties, that of RicR14 cells had decreased...... sulfation, reduced affinity for fibronectin and decreased half-life on the cell surface when compared to the normal counterpart. Our conclusions based on this data are that these altered properties may, in part, account for the adhesion defect in the ricin-resistant mutant. Whether this results from...

  7. Relationships between cell surface insulin binding and endocytosis in adipocytes

    International Nuclear Information System (INIS)

    Jochen, A.L.

    1988-01-01

    Chymotrypsin substrate analogues, such as N-acetyl-Tyr ethyl ester, have recently been demonstrated to inhibit the endocytic uptake of insulin in isolated rat adipocytes. In this study, the effect of N-acetyl-Tyr ethyl ester on cell surface insulin binding and dissociation were examined. Surface-bound 125 I-insulin was distinguished from intracellular 125 I-insulin by the sensitivity of the former to rapid dissociation with an acidic buffer. Plateau levels of surface-bound insulin at 37 degree C were increased 70% by inhibiting the internalization pathway. This increase was temperature and insulin concentration dependent. Thus differences in surface binding were small at 12 degree C and also at high insulin concentrations. Inhibition of internalization with N-acetyl-Tyr ethyl ester markedly slowed the loss of surface-bound insulin observed during dissociation the loss of surface-bound insulin observed during dissociation studies. After 20-30 min of dissociation, the remaining levels of surface-bound insulin were three- to fourfold higher in treated adipocytes compared with control adipocytes. Added unlabeled insulin retained its ability to accelerate the dissociation of insulin in N-acetyl-Tyr ethyl ester-treated cells. These observations indicate that the internalization pathway is a quantitatively important factor in determining levels of surface binding at 37 degree C and in determining the rat of deactivation of insulin binding

  8. Role of lactobacillus cell surface hydrophobicity as probed by AFM in adhesion to surfaces at low and high ionic strength

    NARCIS (Netherlands)

    Vadillo Rodriguez, Virginia; Busscher, Hendrik; van der Mei, Henderina; Norde, Willem; de Vries, Jacob

    2005-01-01

    The S-layer present at the outermost cell surface of some lactobacillus species is known to convey hydrophobicity to the lactobacillus cell surface. Yet, it is commonly found that adhesion of lactobacilli to solid substrata does not proceed according to expectations based on cell surface

  9. Role of lactobacillus cell surface hydrophobicity as probed by AMF in adhesion to surfaces at low and high ionic strength

    NARCIS (Netherlands)

    Vadillo-Rodriguez, V.; Busscher, H.J.; Meij, van der H.C.; Vries, de J.; Norde, W.

    2005-01-01

    The S-layer present at the outermost cell surface of some lactobacillus species is known to convey hydrophobicity to the lactobacillus cell surface. Yet, it is commonly found that adhesion of lactobacilli to solid substrata does not proceed according to expectations based on cell surface

  10. Surface-Enhanced Raman Scattering Nanoparticles as Optical Labels for Imaging Cell Surface Proteins

    Science.gov (United States)

    MacLaughlin, Christina M.

    Assaying the expression of cell surface proteins has widespread application for characterizing cell type, developmental stage, and monitoring disease transformation. Immunophenotyping is conducted by treating cells with labelled targeting moieties that have high affinity for relevant surface protein(s). The sensitivity and specificity of immunophenotyping is defined by the choice of contrast agent and therefore, the number of resolvable signals that can be used to simultaneously label cells. Narrow band width surface-enhanced Raman scattering (SERS) nanoparticles are proposed as optical labels for multiplexed immunophenotying. Two types of surface coatings were investigated to passivate the gold nanoparticles, incorporate SERS functionality, and to facilitate attachment of targeting antibodies. Thiolated poly(ethylene glycol) forms dative bonds with the gold surface and is compatible with multiple physisorbed Raman-active reporter molecules. Ternary lipid bilayers are used to encapsulate the gold nanoparticles particles, and incorporate three different classes of Raman reporters. TEM, UV-Visible absorbance spectroscopy, DLS, and electrophoretic light scattering were used characterize the particle coating. Colourimetric protein assay, and secondary antibody labelling were used to quantify the antibody conjugation. Three different in vitromodels were used to investigate the binding efficacy and specificity of SERS labels for their biomarker targets. Primary human CLL cells, LY10 B lymphoma, and A549 adenocarcinoma lines were targeted. Dark field imaging was used to visualize the colocalization of SERS labels with cells, and evidence of receptor clustering was obtained based on colour shifts of the particles' Rayleigh scattering. Widefield, and spatially-resolved Raman spectra were used to detect labels singly, and in combination from labelled cells. Fluorescence flow cytometry was used to test the particles' binding specificity, and SERS from labelled cells was also

  11. Mapping Cellular Hierarchy by Single-Cell Analysis of the Cell Surface Repertoire

    OpenAIRE

    Guo, Guoji; Luc, Sidinh; Marco, Eugenio; Lin, Ta-Wei; Peng, Cong; Kerenyi, Marc A.; Beyaz, Semir; Kim, Woojin; Xu, Jian; Das, Partha Pratim; Neff, Tobias; Zou, Keyong; Yuan, Guo-Cheng; Orkin, Stuart H.

    2013-01-01

    Stem cell differentiation pathways are most often studied at the population level, whereas critical decisions are executed at the level of single cells. We have established a highly multiplexed, quantitative PCR assay to profile in an unbiased manner a panel of all commonly used cell surface markers (280 genes) from individual cells. With this method we analyzed over 1500 single cells throughout the mouse hematopoietic system, and illustrate its utility for revealing important biological insi...

  12. Inefficient entry of vicriviroc-resistant HIV-1 via the inhibitor-CCR5 complex at low cell surface CCR5 densities

    OpenAIRE

    Pugach, Pavel; Ray, Neelanjana; Klasse, Per Johan; Ketas, Thomas J.; Michael, Elizabeth; Doms, Robert W.; Lee, Benhur; Moore, John P.

    2009-01-01

    HIV-1 variants resistant to small molecule CCR5 inhibitors such as vicriviroc (VVC) have modified Env complexes that can use both the inhibitor-bound and -free forms of the CCR5 co-receptor to enter target cells. However, entry via the inhibitor-CCR5 complex is inefficient in some, but not all, cell types, particularly cell lines engineered to express CCR5. We investigated the effect of increasing CCR5 expression, and hence the density of the inhibitor-CCR5 complex when a saturating inhibitor...

  13. Dynamic and reversible surface topography influences cell morphology.

    Science.gov (United States)

    Kiang, Jennifer D; Wen, Jessica H; del Álamo, Juan C; Engler, Adam J

    2013-08-01

    Microscale and nanoscale surface topography changes can influence cell functions, including morphology. Although in vitro responses to static topography are novel, cells in vivo constantly remodel topography. To better understand how cells respond to changes in topography over time, we developed a soft polyacrylamide hydrogel with magnetic nickel microwires randomly oriented in the surface of the material. Varying the magnetic field around the microwires reversibly induced their alignment with the direction of the field, causing the smooth hydrogel surface to develop small wrinkles; changes in surface roughness, ΔRRMS , ranged from 0.05 to 0.70 μm and could be oscillated without hydrogel creep. Vascular smooth muscle cell morphology was assessed when exposed to acute and dynamic topography changes. Area and shape changes occurred when an acute topographical change was imposed for substrates exceeding roughness of 0.2 μm, but longer-term oscillating topography did not produce significant changes in morphology irrespective of wire stiffness. These data imply that cells may be able to use topography changes to transmit signals as they respond immediately to changes in roughness. Copyright © 2013 Wiley Periodicals, Inc.

  14. Investigation of the Cell Surface Proteome of Human Periodontal Ligament Stem Cells

    OpenAIRE

    Xiong, Jimin; Menicanin, Danijela; Zilm, Peter S.; Marino, Victor; Bartold, P. Mark; Gronthos, Stan

    2016-01-01

    The present study examined the cell surface proteome of human periodontal ligament stem cells (PDLSC) compared to human fibroblasts. Cell surface proteins were prelabelled with CyDye before processing to extract the membrane lysates, which were separated using 2D electrophoresis. Selected differentially expressed protein ?spots? were identified using Mass spectrometry. Four proteins were selected for validation: CD73, CD90, Annexin A2, and sphingosine kinase 1 previously associated with mesen...

  15. Lining cells on normal human vertebral bone surfaces

    International Nuclear Information System (INIS)

    Henning, C.B.; Lloyd, E.L.

    1982-01-01

    Thoracic vertebrae from two individuals with no bone disease were studied with the electron microscope to determine cell morphology in relation to bone mineral. The work was undertaken to determine if cell morphology or spatial relationships between the bone lining cells and bone mineral could account for the relative infrequency of bone tumors which arise at this site following radium intake, when compared with other sites, such as the head of the femur. Cells lining the vertebral mineral were found to be generally rounded in appearance with varied numbers of cytoplasmic granules, and they appeared to have a high density per unit of surface area. These features contrasted with the single layer of flattened cells characteristic of the bone lining cells of the femur. A tentative discussion of the reasons for the relative infrequency of tumors in the vertebrae following radium acquisition is presented

  16. Mesenchymal Stem Cells, Nanofiber Scaffolds and Ocular Surface Reconstruction

    Czech Academy of Sciences Publication Activity Database

    Holáň, Vladimír; Javorková, Eliška

    2013-01-01

    Roč. 9, č. 5 (2013), s. 609-619 ISSN 1550-8943 R&D Projects: GA ČR GAP304/11/0653; GA ČR(CZ) GAP301/11/1568 Grant - others:GA MŠk(CZ) UK668012; GA MŠk(CZ) SVV 265211 Institutional support: RVO:68378041 Keywords : mesenchymal stem cell s * limbal stem cell s * ocular surface injuries Subject RIV: EC - Immunology Impact factor: 3.214, year: 2013

  17. HIV co-receptor tropism prediction remains stable over time in treatment-naïve patients.

    Science.gov (United States)

    Philip, Keir Ej; Macartney, Malcolm J; Conibear, Tim Cr; Smith, Colette J; Marshall, Neal; Johnson, Margaret A; Haque, Tanzina; Webster, Daniel P

    2016-06-01

    HIV co-receptor tropism determination is essential before prescribing the CCR5 antagonist maraviroc. British HIV Association guidelines suggest tropism testing may remain valid for only 90 days in antiretroviral-naïve patients. We aimed to determine the accuracy of this figure. Tropism was assessed in 26 antiretroviral-naïve patients with ongoing viral replication, sampled yearly from first clinic visit. The V3 region of HIV-1 was sequenced in triplicate, then tropism predicted using the Geno2Pheno system. Baseline tropism prediction remained valid for a median of 52 months (range 7-81). For 19/26 individuals baseline tropism remained unchanged throughout a median of 54 months follow-up; 18 R5 tropic and 1 X4 tropic. In seven patients (27%) baseline tropism switched at least once (range 1-4 switches) during follow-up; however, their baseline tropism prediction remained valid for a median of 45 months. Co-receptor tropism in treatment-naïve patients with ongoing viral replication appears highly stable over time, suggesting that baseline genotypic tropism prediction may be valid for a longer duration in patients delaying ART initiation. In this study, baseline tropism prediction remained valid for a median of 52 months, suggesting current guidelines recommending repeat testing after 90 days may be excessively conservative in their assessment of tropism stability. © The Author(s) 2015.

  18. Micromechanical and surface adhesive properties of single saccharomyces cerevisiae cells

    Science.gov (United States)

    Farzi, Bahman; Cetinkaya, Cetin

    2017-09-01

    The adhesion and mechanical properties of a biological cell (e.g. cell membrane elasticity and adhesiveness) are often strong indicators for the state of its health. Many existing techniques for determining mechanical properties of cells require direct physical contact with a single cell or a group of cells. Physical contact with the cell can trigger complex mechanotransduction mechanisms, leading to cellular responses, and consequently interfering with measurement accuracy. In the current work, based on ultrasonic excitation and interferometric (optical) motion detection, a non-contact method for characterizing the adhesion and mechanical properties of single cells is presented. It is experimentally demonstrated that the rocking (rigid body) motion and internal vibrational resonance frequencies of a single saccharomyces cerevisiae (SC) (baker’s yeast) cell can be acquired with the current approach, and the Young’s modulus and surface tension of the cell membrane as well as surface adhesion energy can be extracted from the values of these acquired resonance frequencies. The detected resonance frequency ranges for single SC cells include a rocking (rigid body) frequency of 330  ±  70 kHz and two breathing resonance frequencies of 1.53  ±  0.12 and 2.02  ±  0.31 MHz. Based on these values, the average work-of-adhesion of SC cells on a silicon substrate in aqueous medium is extracted, for the first time, as WASC-Si=16.2+/- 3.8 mJ {{m}-2} . Similarly, the surface tension and the Young’s modulus of the SC cell wall are predicted as {{σ }SC}=0.16+/- 0.02 N {{m}-1} and {{E}SC}= 9.20  ±  2.80 MPa, respectively. These results are compared to those reported in the literature by utilizing various methods, and good agreements are found. The current approach eliminates the measurement inaccuracies associated with the physical contact. Exciting and detecting cell dynamics at micro-second time-scales is significantly faster than the

  19. Surface-modified magnetic nanoparticles for cell labeling

    Czech Academy of Sciences Publication Activity Database

    Zasońska, Beata Anna; Patsula, Vitalii; Stoika, R.; Horák, Daniel

    2014-01-01

    Roč. 13, č. 4 (2014), s. 63-73 ISSN 2305-7815 R&D Projects: GA MŠk(CZ) LH14318 Institutional support: RVO:61389013 Keywords : magnetic nanoparticles * surface-modified * cell labeling Subject RIV: CD - Macromolecular Chemistry

  20. LANTHANUM STAINING OF THE SURFACE COAT OF CELLS

    Science.gov (United States)

    Shea, Stephen M.

    1971-01-01

    Among the techniques which have been reported to stain the surface coat of cells, for electron microscopy, is lanthanum staining en bloc. Similarly, the presence of the cationic dye, Alcian blue 8GX, in a primary glutaraldehyde fixative has been reported to improve the preservation of the surface coat of cells of many types; however, the preserved coat is not very electron opaque unless thin sections are counterstained. The present paper shows that for several rat tissues lanthanum staining en bloc is an effective electron stain for the cell surface, giving excellent contrast, if combined sequentially with prefixation in an aldehyde fixative containing Alcian blue. The cationic substance cetylpyridinium chloride was found to have a similar effect to that of Alcian blue in enhancing the lanthanum staining of the surface coat material of the brush border of intestinal epithelial cells. The patterns of lanthanum staining obtained for the tissues studied strikingly resemble those reported in the literature where tissues are stained by several standard methods for demonstrating mucosubstances at the ultrastructural level. This fact and the reproduction of the effect of Alcian blue by cetylpyridinium chloride constitute a persuasive empirical argument that the material visualized is a mucopolysaccharide or mucopolysaccharide-protein complex. PMID:4108476

  1. Bacterial Cell Surface Damage Due to Centrifugal Compaction

    NARCIS (Netherlands)

    Peterson, Brandon W.; Sharma, Prashant K.; van der Mei, Henny C.; Busscher, Henk J.

    Centrifugal damage has been known to alter bacterial cell surface properties and interior structures, including DNA. Very few studies exist on bacterial damage caused by centrifugation because of the difficulty in relating centrifugation speed and container geometry to the damage caused. Here, we

  2. Multijunction Solar Cells Optimized for the Mars Surface Solar Spectrum

    Science.gov (United States)

    Edmondson, Kenneth M.; Fetzer, Chris; Karam, Nasser H.; Stella, Paul; Mardesich, Nick; Mueller, Robert

    2007-01-01

    This paper gives an update on the performance of the Mars Exploration Rovers (MER) which have been continually performing for more than 3 years beyond their original 90-day missions. The paper also gives the latest results on the optimization of a multijunction solar cell that is optimized to give more power on the surface of Mars.

  3. Surface determinants of low density lipoprotein uptake by endothelial cells

    International Nuclear Information System (INIS)

    Goeroeg, P.; Pearson, J.D.

    1984-01-01

    The surface sialic acid content of aortic endothelial cells in vitro was substantially lower in sparse cultures than at confluence. Binding of LDL to endothelial cells did not change at different culture densities and was unaffected by brief pretreatment with neuraminidase to partially remove surface sialic acid residues. In contrast, internalisation of LDL declined by a factor of 3 between low density cell cultures and confluent monolayers; neuraminidase pretreatment increased LDL uptake and the effect was most marked (>10-fold) at confluence. Pretreatment with cationised ferritin, which removed most of the surface sialic acid residues as well as glycosaminoglycans, increased LDL internalisation by up to 20-fold, again with most effect on confluent monolayers. Thus LDL uptake is inversely correlated with sialic acid content. We conclude that changes in the surface density of sialic acid (and possibly other charged) residues significantly modulate endothelial LDL uptake, and suggest that focal increases in LDL accumulation during atherogenesis may be related to alterations in endothelial endocytic properties at sites of increased cell turnover or damage. (author)

  4. Cell adhesion on Ti surface with controlled roughness

    Energy Technology Data Exchange (ETDEWEB)

    Burgos-Asperilla, L.; Garcia-Alonso, M. C.; Escudero, M. L.; Alonso, C.

    2015-07-01

    In this report, the in situ interaction between Saos-2 osteoblast cells and a smooth Ti surface was examined over time. The adhesion kinetics and mechanisms of cellular proliferation were monitored by quartz crystal microbalance (QCM) and electrochemical impedance spectroscopy (EIS). The rate of Saos-2 attachment on Ti surfaces, obtained from the measurements performed with the QCM, is a first-order reaction, with k=2.10{sup -}3 min{sup -}1. The impedance measurements indicate that in the absence of cells, the Ti resistance diminishes over time (7 days), due to the presence of amino acids and proteins from the culture medium that have been adsorbed, while in the presence of osteoblasts, this decrease is much greater because of the compounds generated by the cells that accelerate the dissolution of Ti. (Author)

  5. Surface modification of hydrophobic polymers for improvement of endothelial cell-surface interactions

    NARCIS (Netherlands)

    Dekker, A.; Dekker, A.; Reitsma, K.; Beugeling, T.; Beugeling, T.; Bantjes, A.; Bantjes, A.; Feijen, Jan; Kirkpatrick, C.J.; van Aken, W.G.

    1992-01-01

    The aim of this study is to improve the interaction of endothelial cells with polymers used in vascular prostheses. Polytetrafluoroethylene (PTFE; Teflon) films were treated by means of nitrogen and oxygen plasmas. Depending on the plasma exposure time, modified PTFE surfaces showed water-contact

  6. Established cell surface markers efficiently isolate highly overlapping populations of skeletal muscle satellite cells by fluorescence-activated cell sorting.

    Science.gov (United States)

    Maesner, Claire C; Almada, Albert E; Wagers, Amy J

    2016-01-01

    Fluorescent-activated cell sorting (FACS) has enabled the direct isolation of highly enriched skeletal muscle stem cell, or satellite cell, populations from postnatal tissue. Several distinct surface marker panels containing different positively selecting surface antigens have been used to distinguish muscle satellite cells from other non-myogenic cell types. Because functional and transcriptional heterogeneity is known to exist within the satellite cell population, a direct comparison of results obtained in different laboratories has been complicated by a lack of clarity as to whether commonly utilized surface marker combinations select for distinct or overlapping subsets of the satellite cell pool. This study therefore sought to evaluate phenotypic and functional overlap among popular satellite cell sorting paradigms. Utilizing a transgenic Pax7 -zsGreen reporter mouse, we compared the overlap between the fluorescent signal of canonical paired homeobox protein 7 ( Pax7 ) expressing satellite cells to cells identified by combinations of surface markers previously published for satellite cells isolation. We designed two panels for mouse skeletal muscle analysis, each composed of markers that exclude hematopoietic and stromal cells (CD45, CD11b, Ter119, CD31, and Sca1), combined with previously published antibody clones recognizing surface markers present on satellite cells (β1-integrin/CXCR4, α7-integrin/CD34, and Vcam1). Cell populations were comparatively analyzed by flow cytometry and FACS sorted for functional assessment of myogenic activity. Consistent with prior reports, each of the commonly used surface marker schemes evaluated here identified a highly enriched satellite cell population, with 89-90 % positivity for Pax7 expression based on zsGreen fluorescence. Distinct surface marker panels were also equivalent in their ability to identify the majority of the satellite cell pool, with 90-93 % of all Pax7-zsGreen positive cells marked by each of the

  7. MEMS-based dynamic cell-to-cell culture platforms using electrochemical surface modifications

    International Nuclear Information System (INIS)

    Chang, Jiyoung; Lin, Liwei; Yoon, Sang-Hee; Mofrad, Mohammad R K

    2011-01-01

    MEMS-based biological platforms with the capability of both spatial placements and time releases of living cells for cell-to-cell culture experiments have been designed and demonstrated utilizing electrochemical surface modification effects. The spatial placement is accomplished by electrochemical surface modification of substrate surfaces to be either adhesive or non-adhesive for living cells. The time control is achieved by the electrical activation of the selective indium tin oxide co-culture electrode to allow the migration of living cells onto the electrode to start the cell-to-cell culture studies. Prototype devices have a three-electrode design with an electrode size of 50 × 50 µm 2 and the separation gaps of 2 µm between them. An electrical voltage of −1.5 V has been used to activate the electrodes independently and sequentially to demonstrate the dynamic cell-to-cell culture experiments of NIH 3T3 fibroblast and Madin Darby canine kidney cells. As such, this MEMS platform could be a basic yet versatile tool to characterize transient cell-to-cell interactions

  8. Molecular Tension Probes for Imaging Forces at the Cell Surface.

    Science.gov (United States)

    Liu, Yang; Galior, Kornelia; Ma, Victor Pui-Yan; Salaita, Khalid

    2017-12-19

    Mechanical forces are essential for a variety of biological processes ranging from transcription and translation to cell adhesion, migration, and differentiation. Through the activation of mechanosensitive signaling pathways, cells sense and respond to physical stimuli from the surrounding environment, a process widely known as mechanotransduction. At the cell membrane, many signaling receptors, such as integrins, cadherins and T- or B-cell receptors, bind to their ligands on the surface of adjacent cells or the extracellular matrix (ECM) to mediate mechanotransduction. Upon ligation, these receptor-ligand bonds transmit piconewton (pN) mechanical forces that are generated, in part, by the cytoskeleton. Importantly, these forces expose cryptic sites within mechanosensitive proteins and modulate the binding kinetics (on/off rate) of receptor-ligand complexes to further fine-tune mechanotransduction and the corresponding cell behavior. Over the past three decades, two categories of methods have been developed to measure cell receptor forces. The first class is traction force microscopy (TFM) and micropost array detectors (mPADs). In these methods, cells are cultured on elastic polymers or microstructures that deform under mechanical forces. The second category of techniques is single molecule force spectroscopy (SMFS) including atomic force microscopy (AFM), optical or magnetic tweezers, and biomembrane force probe (BFP). In SMFS, the experimenter applies external forces to probe the mechanics of individual cells or single receptor-ligand complexes, serially, one bond at a time. Although these techniques are powerful, the limited throughput of SMFS and the nN force sensitivity of TFM have hindered further elucidation of the molecular mechanisms of mechanotransduction. In this Account, we introduce the recent advent of molecular tension fluorescence microscopy (MTFM) as an emerging tool for molecular imaging of receptor mechanics in living cells. MTFM probes are

  9. Surface code—biophysical signals for apoptotic cell clearance

    International Nuclear Information System (INIS)

    Biermann, Mona; Maueröder, Christian; Brauner, Jan M; Chaurio, Ricardo; Herrmann, Martin; Muñoz, Luis E; Janko, Christina

    2013-01-01

    Apoptotic cell death and the clearance of dying cells play an important and physiological role in embryonic development and normal tissue turnover. In contrast to necrosis, apoptosis proceeds in an anti-inflammatory manner. It is orchestrated by the timed release and/or exposure of so-called ‘find-me’, ‘eat me’ and ‘tolerate me’ signals. Mononuclear phagocytes are attracted by various ‘find-me’ signals, including proteins, nucleotides, and phospholipids released by the dying cell, whereas the involvement of granulocytes is prevented via ‘stay away’ signals. The exposure of anionic phospholipids like phosphatidylserine (PS) by apoptotic cells on the outer leaflet of the plasma membrane is one of the main ‘eat me’ signals. PS is recognized by a number of innate receptors as well as by soluble bridging molecules on the surface of phagocytes. Importantly, phagocytes are able to discriminate between viable and apoptotic cells both exposing PS. Due to cytoskeleton remodeling PS has a higher lateral mobility on the surfaces of apoptotic cells thereby promoting receptor clustering on the phagocyte. PS not only plays an important role in the engulfment process, but also acts as ‘tolerate me’ signal inducing the release of anti-inflammatory cytokines by phagocytes. An efficient and fast clearance of apoptotic cells is required to prevent secondary necrosis and leakage of intracellular danger signals into the surrounding tissue. Failure or prolongation of the clearance process leads to the release of intracellular antigens into the periphery provoking inflammation and development of systemic inflammatory autoimmune disease like systemic lupus erythematosus. Here we review the current findings concerning apoptosis-inducing pathways, important players of apoptotic cell recognition and clearance as well as the role of membrane remodeling in the engulfment of apoptotic cells by phagocytes. (paper)

  10. Encapsulant Adhesion to Surface Metallization on Photovoltaic Cells

    Energy Technology Data Exchange (ETDEWEB)

    Tracy, Jared; Bosco, Nick; Dauskardt, Reinhold

    2017-11-01

    Delamination of encapsulant materials from PV cell surfaces often appears to originate at regions with metallization. Using a fracture mechanics based metrology, the adhesion of ethylene vinyl acetate (EVA) encapsulant to screen-printed silver metallization was evaluated. At room temperature, the fracture energy Gc [J/m2] of the EVA/silver interface (952 J/m2) was ~70% lower than that of the EVA/antireflective (AR) coating (>2900 J/m2) and ~60% lower than that of the EVA to the surface of cell (2265 J/m2). After only 300 h of damp heat aging, the adhesion energy of the silver interface dropped to and plateaued at ~50-60 J/m2 while that of the EVA/AR coating and EVA/cell remained mostly unchanged. Elemental surface analysis showed that the EVA separates from the silver in a purely adhesive manner, indicating that bonds at the interface were likely displaced in the presence of humidity and chemical byproducts at elevated temperature, which in part accounts for the propensity of metalized surfaces to delaminate in the field.

  11. Surface science studies of model fuel cell electrocatalysts

    Science.gov (United States)

    Marković, N. M.; Ross, P. N.

    2002-04-01

    The purpose of this review is to discuss progress in the understanding of electrocatalytic reactions through the study of model systems with surface spectroscopies. Pure metal single crystals and well-characterized bulk alloys have been used quite successfully as models for real (commercial) electrocatalysts. Given the sheer volume of all work in electrocatalysis that is on fuel cell reactions, we will focus on electrocatalysts for fuel cells. Since Pt is the model fuel cell electrocatalyst, we will focus entirely on studies of pure Pt and Pt bimetallic alloys. The electrode reactions discussed include hydrogen oxidation/evolution, oxygen reduction, and the electrooxidation of carbon monoxide, formic acid, and methanol. Surface spectroscopies emphasized are FTIR, STM/AFM and surface X-ray scattering (SXS). The discussion focuses on the relation between the energetics of adsorption of intermediates and the reaction pathway and kinetics, and how the energetics and kinetics relate to the extrinsic properties of the model system, e.g. surface structure and/or composition. Finally, we conclude by discussing the limitations that are reached by using pure metal single crystals and well-characterized bulk alloys as models for real catalysts, and suggest some directions for developing more realistic systems.

  12. Surface-modified low-temperature solid oxide fuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young Beom; Holme, Timothy P. [Department of Mechanical Engineering, Stanford University, Stanford, CA 94305 (United States); Guer, Turgut M. [Department of Materials Science and Engineering, Stanford University, Stanford, CA 94305 (United States); Prinz, Fritz B. [Department of Mechanical Engineering, Stanford University, Stanford, CA 94305 (United States); Department of Materials Science and Engineering, Stanford University, Stanford, CA 94305 (United States)

    2011-12-20

    This paper reports both experimental and theoretical results of the role of surface modification on the oxygen reduction reaction in low-temperature solid oxide fuel cells (LT-SOFC). Epitaxial ultrathin films of yttria-doped ceria (YDC) cathode interlayers (<10-130 nm) are grown by pulsed laser deposition (PLD) on single-crystalline YSZ(100). Fuel cell current-voltage measurements and electrochemical impedance spectroscopy are performed in the temperature range of 350 C {approx} 450 C. Quantum mechanical simulations of oxygen incorporation energetics support the experimental results and indicate a low activation energy of only 0.07 eV for YDC, while the incorporation reaction on YSZ is activated by a significantly higher energy barrier of 0.38 eV. Due to enhanced oxygen incorporation at the modified Pt/YDC interface, the cathodic interface resistance is reduced by two-fold, while fuel cell performance shows more than a two-fold enhancement with the addition of an ultrathin YDC interlayer at the cathode side of an SOFC element. The results of this study open up opportunities for improving cell performance, particularly of LT-SOFCs by adopting surface modification of YSZ surface with catalytically superior, ultrathin cathodic interlayers. (Copyright copyright 2011 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  13. Surface modified alginate microcapsules for 3D cell culture

    Science.gov (United States)

    Chen, Yi-Wen; Kuo, Chiung Wen; Chueh, Di-Yen; Chen, Peilin

    2016-06-01

    Culture as three dimensional cell aggregates or spheroids can offer an ideal platform for tissue engineering applications and for pharmaceutical screening. Such 3D culture models, however, may suffer from the problems such as immune response and ineffective and cumbersome culture. This paper describes a simple method for producing microcapsules with alginate cores and a thin shell of poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) to encapsulate mouse induced pluripotent stem (miPS) cells, generating a non-fouling surface as an effective immunoisolation barrier. We demonstrated the trapping of the alginate microcapsules in a microwell array for the continuous observation and culture of a large number of encapsulated miPS cells in parallel. miPS cells cultured in the microcapsules survived well and proliferated to form a single cell aggregate. Droplet formation of monodisperse microcapsules with controlled size combined with flow cytometry provided an efficient way to quantitatively analyze the growth of encapsulated cells in a high-throughput manner. The simple and cost-effective coating technique employed to produce the core-shell microcapsules could be used in the emerging field of cell therapy. The microwell array would provide a convenient, user friendly and high-throughput platform for long-term cell culture and monitoring.

  14. Glucocorticoids and the cell surface of human glioma cells: relationship to cytostasis.

    Science.gov (United States)

    Mackie, A E; Freshney, R I; Akturk, F; Hunt, G

    1988-12-01

    The glucocorticoid hormones methyl prednisolone and dexamethasone were shown to be cytostatic, but not cytotoxic, at high cell densities for early passage and continuous cell lines from human glioma at 0.25 microM and above, in the presence or absence of serum. In the absence of serum both steroids at 2.5 nM increased the saturation density close to the level reached in serum. Examination of the iodinated glycoproteins of the cell surface by gel electrophoresis did not reveal any consistent change. However, gel exclusion chromatography of protease digests of the cell surface and of material released into the medium showed an increase in incorporation of 3H-glucosamine in pronase digests after treatment with methyl prednisolone. Ion exchange chromatography showed that sulphated glycosaminoglycans, particularly heparan sulphate, increased and hyaluronic acid decreased in response to steroids, and there was increased retention of GAGs on the cell surface relative to the released fraction. It was concluded that glucocorticoid hormones modify the cell surface of human glioma cells and that this may contribute to enhanced cell intraction and lead to increased density limitation of cell proliferation.

  15. Stable isotope labeling of oligosaccharide cell surface antigens

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, C.J.; Silks, L.A. III; Martinez, R.A. [and others

    1998-12-31

    The overall goal of this Laboratory Directed Research and Development (LDRD) project was to develop new methods for synthesis of {sup 13}C-labeled oligosaccharides that are required for nuclear magnetic resonance (NMR) studies of their solution conformation. Oligosaccharides are components of the cell`s outer surface and are involved in important processes such as cell-cell recognition and adhesion. Recently, Danishefsky and coworkers at Slone-Kettering Cancer Center developed a method for the solid-phase chemical synthesis of oligosaccharides. The specific goal of this LDRD project was to prepare uniform {sup 13}C-labeled aldohexose precursors required for the solid-phase synthesis of the Lewis blood-group antigenic determinants. We report the synthesis of {sup 13}C-labeled D-glucal, D-galactal and Fucosyl precursors. We have been collaborating with the Danishefsky group on the synthesis of the Lewis oligosaccharides and the NMR analysis of their solution conformation.

  16. Characterization of atrial natriuretic peptide degradation by cell-surface peptidase activity on endothelial cells

    Science.gov (United States)

    Frost, S. J.; Whitson, P. A.

    1993-01-01

    Atrial natriuretic peptide (ANP) is a fluid-regulating peptide hormone that promotes vasorelaxation, natriuresis, and diuresis. The mechanisms for the release of ANP and for its clearance from the circulation play important roles in modulating its biological effects. Recently, we have reported that the cell surface of an endothelial cell line, CPA47, could degrade 125I-ANP in the presence of EDTA. In this study, we have characterized this degradation of 125I-ANP. The kinetics of ANP degradation by the surface of CPA47 cells were first order, with a Km of 320 +/- 60 nM and Vmax of 35 +/- 14 pmol of ANP degraded/10 min/10(5) cells at pH 7.4. ANP is degraded by the surface of CPA47 cells over a broad pH range from 7.0-8.5. Potato carboxypeptidase inhibitor and bestatin inhibited 125I-ANP degradation, suggesting that this degradative activity on the surface of CPA47 cells has exopeptidase characteristics. The selectivity of CPA47 cell-surface degradation of ANP was demonstrated when 125I-ANP degradation was inhibited in the presence of neuropeptide Y and angiotensin I and II but not bradykinin, bombesin, endothelin-1, or substance P. The C-terminal amino acids phe26 and tyr28 were deduced to be important for ANP interaction with the cell-surface peptidase(s) based on comparison of the IC50 of various ANP analogues and other natriuretic peptides for the inhibition of ANP degradation. These data suggest that a newly characterized divalent cation-independent exopeptidase(s) that selectively recognizes ANP and some other vasoactive peptides exists on the surface of endothelial cells.

  17. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

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

    clinically applied reagents, such as alkyl-lysophospholipides, chemotherapeutic agents, and anti-inflammatory reagents, have been found to enhance Hsp70 surface expression on cancer cells. We have found that inhibition of histone deacetylase (HDAC) activity leads to surface expression of Hsp70 on various...

  18. Probing and Mapping Electrode Surfaces in Solid Oxide Fuel Cells

    Science.gov (United States)

    Blinn, Kevin S.; Li, Xiaxi; Liu, Mingfei; Bottomley, Lawrence A.; Liu, Meilin

    2012-01-01

    Solid oxide fuel cells (SOFCs) are potentially the most efficient and cost-effective solution to utilization of a wide variety of fuels beyond hydrogen 1-7. The performance of SOFCs and the rates of many chemical and energy transformation processes in energy storage and conversion devices in general are limited primarily by charge and mass transfer along electrode surfaces and across interfaces. Unfortunately, the mechanistic understanding of these processes is still lacking, due largely to the difficulty of characterizing these processes under in situ conditions. This knowledge gap is a chief obstacle to SOFC commercialization. The development of tools for probing and mapping surface chemistries relevant to electrode reactions is vital to unraveling the mechanisms of surface processes and to achieving rational design of new electrode materials for more efficient energy storage and conversion2. Among the relatively few in situ surface analysis methods, Raman spectroscopy can be performed even with high temperatures and harsh atmospheres, making it ideal for characterizing chemical processes relevant to SOFC anode performance and degradation8-12. It can also be used alongside electrochemical measurements, potentially allowing direct correlation of electrochemistry to surface chemistry in an operating cell. Proper in situ Raman mapping measurements would be useful for pin-pointing important anode reaction mechanisms because of its sensitivity to the relevant species, including anode performance degradation through carbon deposition8, 10, 13, 14 ("coking") and sulfur poisoning11, 15 and the manner in which surface modifications stave off this degradation16. The current work demonstrates significant progress towards this capability. In addition, the family of scanning probe microscopy (SPM) techniques provides a special approach to interrogate the electrode surface with nanoscale resolution. Besides the surface topography that is routinely collected by AFM and STM

  19. Exogenous cathepsin G upregulates cell surface MHC class I molecules on immune and glioblastoma cells.

    Science.gov (United States)

    Giese, Madleen; Turiello, Nadine; Molenda, Nicole; Palesch, David; Meid, Annika; Schroeder, Roman; Basilico, Paola; Benarafa, Charaf; Halatsch, Marc-Eric; Zimecki, Michal; Westhoff, Mike-Andrew; Wirtz, Christian Rainer; Burster, Timo

    2016-11-15

    Major histocompatibility complex (MHC) class I molecules present antigenic peptides to cytotoxic T cells. During an adaptive immune response, MHC molecules are regulated by several mechanisms including lipopolysaccharide (LPS) and interferon gamma (IFN-g). However, it is unclear whether the serine protease cathepsin G (CatG), which is generally secreted by neutrophils at the site of inflammation, might regulate MHC I molecules. We identified CatG, and to a higher extend CatG and lactoferrin (LF), as an exogenous regulator of cell surface MHC I expression of immune cells and glioblastoma stem cells. In addition, levels of MHC I molecules are reduced on dendritic cells from CatG deficient mice compared to their wild type counterparts. Furthermore, cell surface CatG on immune cells, including T cells, B cells, and NK cells triggers MHC I on THP-1 monocytes suggesting a novel mechanism for CatG to facilitate intercellular communication between infiltrating cells and the respective target cell. Subsequently, our findings highlight the pivotal role of CatG as a checkpoint protease which might force target cells to display their intracellular MHC I:antigen repertoire.

  20. Cdon, a cell surface protein, mediates oligodendrocyte differentiation and myelination.

    Science.gov (United States)

    Wang, Li-Chun; Almazan, Guillermina

    2016-06-01

    During central nervous system development, oligodendrocyte progenitors (OLPs) establish multiple branched processes and axonal contacts to initiate myelination. A complete understanding of the molecular signals implicated in cell surface interaction to initiate myelination/remyelination is currently lacking. The objective of our study was to assess whether Cdon, a cell surface protein that was shown to participate in muscle and neuron cell development, is involved in oligodendrocyte (OLG) differentiation and myelination. Here, we demonstrate that endogenous Cdon protein is expressed in OLPs, increasing in the early differentiation stages and decreasing in mature OLGs. Immunocytochemistry of endogenous Cdon showed localization on both OLG cell membranes and cellular processes exhibiting puncta- or varicosity-like structures. Cdon knockdown with siRNA decreased protein levels by 62% as well as two myelin-specific proteins, MBP and MAG. Conversely, overexpression of full-length rat Cdon increased myelin proteins in OLGs. The complexity of OLGs branching and contact point numbers with axons were also increased in Cdon overexpressing cells growing alone or in coculture with dorsal root ganglion neurons (DRGNs). Furthermore, myelination of DRGNs was decreased when OLPs were transfected with Cdon siRNA. Altogether, our results suggest that Cdon participates in OLG differentiation and myelination, most likely in the initial stages of development. © 2016 Wiley Periodicals, Inc.

  1. Bacterial Cell Surface Adsorption of Rare Earth Elements

    Science.gov (United States)

    Jiao, Y.; Park, D.; Reed, D.; Fujita, Y.; Yung, M.; Anderko, A.; Eslamimanesh, A.

    2015-12-01

    Rare earth elements (REE) play a critical role in many emerging clean energy technologies, including high-power magnets, wind turbines, solar panels, hybrid/electric vehicle batteries and lamp phosphors. In order to sustain demand for such technologies given current domestic REE shortages, there is a need to develop new approaches for ore processing/refining and recycling of REE-containing materials. To this end, we have developed a microbially-mediated bioadsorption strategy with application towards enrichment of REE from complex mixtures. Specifically, the bacterium Caulobacter crescentus was genetically engineered to display lanthanide binding tags (LBTs), short peptides that possess high affinity and specificity for rare earth elements, on its cell surface S-layer protein. Under optimal conditions, LBT-displayed cells adsorbed greater than 5-fold more REE than control cells lacking LBTs. Competition binding experiments with a selection of REEs demonstrated that our engineered cells could facilitate separation of light- from heavy- REE. Importantly, binding of REE onto our engineered strains was much more favorable compared to non-REE metals. Finally, REE bound to the cell surface could be stripped off using citrate, providing an effective and non-toxic REE recovery method. Together, this data highlights the potential of our approach for selective REE enrichment from REE containing mixtures.

  2. Interfacing biomembrane mimetic polymer surfaces with living cells - Surface modification for reliable bioartificial liver

    International Nuclear Information System (INIS)

    Iwasaki, Yasuhiko; Takami, Utae; Sawada, Shin-ichi; Akiyoshi, Kazunari

    2008-01-01

    The surface design used for reducing nonspecific biofouling is one of the most important issues for the fabrication of medical devices. We present here a newly synthesized a carbohydrate-immobilized phosphorylcholine polymer for surface modification of medical devices to control the interface with living cells. A random copolymer composed of 2-methacryloyloxyethyl phosphorylcholine (MPC), n-butyl methacrylate (BMA), and 2-lactobionamidoethyl methacrylate (LAMA) was synthesized by conventional radical polymerization. The monomer feeding ratio in the copolymer was adjusted to 24/75/1 (MPC/BMA/LAMA). The copolymer (PMBL1.0) could be coated by solvent evaporation from an ethanol solution. Cells of the human hepatocellular liver carcinoma cell line (HepG2) having asialoglycoprotein receptors (ASGPRs) were seeded on PMBL1.0 or poly(BMA) (PBMA)-coated PET plates. On PBMA, many adherent cells were observed and were well spread with monolayer adhesion. HepG2 adhesion was observed on PMBL1.0 because the cell has ASGPRs. Furthermore, some of the cells adhering to PMBL1.0 had a spheroid formation and similarly shaped spheroids were scattered on the surface. According to confocal laser microscopic observation after 96 h cultivation, it was found that albumin production preferentially occurred in the center of the spheroid. The albumin production of the cells that adhered to PBMA was sparse. The amount of albumin production per unit cell that adhered to PMBL1.0 was determined by ELISA and was significantly higher than that which adhered to PBMA. Long-term cultivation of HepG2 was also performed using hollow fiber mini-modules coated with PMBL1.0. The concentration of albumin produced from HepG2 increased continuously for one month. In the mini-module, the function of HepG2 was effectively preserved for that period. On the hollow fiber membrane, spheroid formation of HepG2 cells was also observed. In conclusion, PMBL1.0 can provide a suitable surface for the cultivation of

  3. Surface proteome analysis and characterization of surface cell antigen (Sca or autotransporter family of Rickettsia typhi.

    Directory of Open Access Journals (Sweden)

    Khandra T Sears

    Full Text Available Surface proteins of the obligate intracellular bacterium Rickettsia typhi, the agent of murine or endemic typhus fever, comprise an important interface for host-pathogen interactions including adherence, invasion and survival in the host cytoplasm. In this report, we present analyses of the surface exposed proteins of R. typhi based on a suite of predictive algorithms complemented by experimental surface-labeling with thiol-cleavable sulfo-NHS-SS-biotin and identification of labeled peptides by LC MS/MS. Further, we focus on proteins belonging to the surface cell antigen (Sca autotransporter (AT family which are known to be involved in rickettsial infection of mammalian cells. Each species of Rickettsia has a different complement of sca genes in various states; R. typhi, has genes sca1 thru sca5. In silico analyses indicate divergence of the Sca paralogs across the four Rickettsia groups and concur with previous evidence of positive selection. Transcripts for each sca were detected during infection of L929 cells and four of the five Sca proteins were detected in the surface proteome analysis. We observed that each R. typhi Sca protein is expressed during in vitro infections and selected Sca proteins were expressed during in vivo infections. Using biotin-affinity pull down assays, negative staining electron microscopy, and flow cytometry, we demonstrate that the Sca proteins in R. typhi are localized to the surface of the bacteria. All Scas were detected during infection of L929 cells by immunogold electron microscopy. Immunofluorescence assays demonstrate that Scas 1-3 and 5 are expressed in the spleens of infected Sprague-Dawley rats and Scas 3, 4 and 5 are expressed in cat fleas (Ctenocephalides felis. Sca proteins may be crucial in the recognition and invasion of different host cell types. In short, continuous expression of all Scas may ensure that rickettsiae are primed i to infect mammalian cells should the flea bite a host, ii to remain

  4. Active screen plasma nitriding enhances cell attachment to polymer surfaces

    International Nuclear Information System (INIS)

    Kaklamani, Georgia; Bowen, James; Mehrban, Nazia; Dong, Hanshan; Grover, Liam M.; Stamboulis, Artemis

    2013-01-01

    Active screen plasma nitriding (ASPN) is a well-established technique used for the surface modification of materials, the result of which is often a product with enhanced functional performance. Here we report the modification of the chemical and mechanical properties of ultra-high molecular weight poly(ethylene) (UHMWPE) using 80:20 (v/v) N 2 /H 2 ASPN, followed by growth of 3T3 fibroblasts on the treated and untreated polymer surfaces. ASPN-treated UHMWPE showed extensive fibroblast attachment within 3 h of seeding, whereas fibroblasts did not successfully attach to untreated UHMWPE. Fibroblast-coated surfaces were maintained for up to 28 days, monitoring their metabolic activity and morphology throughout. The chemical properties of the ASPN-treated UHMWPE surface were studied using X-ray photoelectron spectroscopy, revealing the presence of C-N, C=N, and C≡N chemical bonds. The elastic modulus, surface topography, and adhesion properties of the ASPN-treated UHMWPE surface were studied over 28 days during sample storage under ambient conditions and during immersion in two commonly used cell culture media.

  5. Cell-surface phosphatidylserine regulates osteoclast precursor fusion.

    Science.gov (United States)

    Verma, Santosh K; Leikina, Evgenia; Melikov, Kamran; Gebert, Claudia; Kram, Vardit; Young, Marian F; Uygur, Berna; Chernomordik, Leonid V

    2018-01-05

    Bone-resorbing multinucleated osteoclasts that play a central role in the maintenance and repair of our bones are formed from bone marrow myeloid progenitor cells by a complex differentiation process that culminates in fusion of mononuclear osteoclast precursors. In this study, we uncoupled the cell fusion step from both pre-fusion stages of osteoclastogenic differentiation and the post-fusion expansion of the nascent fusion connections. We accumulated ready-to-fuse cells in the presence of the fusion inhibitor lysophosphatidylcholine and then removed the inhibitor to study synchronized cell fusion. We found that osteoclast fusion required the dendrocyte-expressed seven transmembrane protein (DC-STAMP)-dependent non-apoptotic exposure of phosphatidylserine at the surface of fusion-committed cells. Fusion also depended on extracellular annexins, phosphatidylserine-binding proteins, which, along with annexin-binding protein S100A4, regulated fusogenic activity of syncytin 1. Thus, in contrast to fusion processes mediated by a single protein, such as epithelial cell fusion in Caenorhabditis elegans , the cell fusion step in osteoclastogenesis is controlled by phosphatidylserine-regulated activity of several proteins.

  6. Vaccines based on the cell surface carbohydrates of pathogenic bacteria

    Directory of Open Access Journals (Sweden)

    Jones Christopher

    2005-01-01

    Full Text Available Glycoconjugate vaccines, in which a cell surface carbohydrate from a micro-organism is covalently attached to an appropriate carrier protein are proving to be the most effective means to generate protective immune responses to prevent a wide range of diseases. The technology appears to be generic and applicable to a wide range of pathogens, as long as antibodies against surface carbohydrates help protect against infection. Three such vaccines, against Haemophilus influenzae type b, Neisseria meningitidis Group C and seven serotypes of Streptococcus pneumoniae, have already been licensed and many others are in development. This article discusses the rationale for the development and use of glycoconjugate vaccines, the mechanisms by which they elicit T cell-dependent immune responses and the implications of this for vaccine development, the role of physicochemical methods in the characterisation and quality control of these vaccines, and the novel products which are under development.

  7. Isolation of Pancreatic Progenitor Cells with the Surface Marker of Hematopoietic Stem Cells

    Directory of Open Access Journals (Sweden)

    Fengxia Ma

    2012-01-01

    Full Text Available To isolate pancreatic progenitor cells with the surface markers of hematopoietic stem cells, the expression of stem cell antigen (Sca-1 and c-Kit and the coexpression of them with pancreatic duodenal homeobox-1 (PDX-1, neurogenin 3 (Ngn3, and insulin were examined in murine embryonic pancreas. Then different pancreatic cell subpopulations were isolated by magnet-activated cell sorting. Isolated cells were cultured overnight in hanging drops. When cells formed spheres, they were laid on floating filters at the air/medium interface. With this new culture system, pancreatic progenitor cells were induced to differentiate to endocrine and exocrine cells. It was shown that c-Kit and Sca-1 were expressed differently in embryonic pancreas at 12.5, 15.5, and 17.5 days of gestation. The expression of c-Kit and Sca-1 was the highest at 15.5 days of gestation. c-Kit rather than Sca-1 coexpressed with PDX-1, Ngn3, and insulin. Cells differentiated from c-Kit-positive cells contained more insulin-producing cells and secreted more insulin in response to glucose stimulation than that from c-Kit-negative cells. These results suggested that c-Kit could be used to isolate pancreatic progenitor cells and our new culture system permitted pancreatic progenitor cells to differentiate to mature endocrine cells.

  8. The Mesenchymal Precursor Cell Marker Antibody STRO-1 Binds to Cell Surface Heat Shock Cognate 70.

    Science.gov (United States)

    Fitter, Stephen; Gronthos, Stan; Ooi, Soo Siang; Zannettino, Andrew C W

    2017-04-01

    Since its discovery more than 25 years ago, the STRO-1 antibody has played a fundamental role in defining the hierarchical nature of mesenchymal precursor cells (MPC) and their progeny. STRO-1 antibody binding remains a hallmark of immature pluripotent MPC. Despite the significance of STRO-1 in the MPC field, the identity of the antigen has remained elusive. Using a combination of two-dimensional gel electrophoresis, coupled with Western blotting and Tandem mass spectroscopy, we have identified the STRO-1 antigen as heat shock cognate 70 (HSC70;HSPA8). STRO-1 binds to immune-precipitated HSC70 and siRNA-mediated knock down of HSPA8 reduced STRO-1 binding. STRO-1 surface binding does not correlate with HSC70 expression and sequestration of cholesterol reduces STRO-1 surface binding, suggesting that the plasma membrane lipid composition may be an important determinant in the presentation of HSC70 on the cell surface. HSC70 is present on the surface of STRO-1 + but not STRO-1 - cell lines as assessed by cell surface biotinylation and recombinant HSC70 blocks STRO-1 binding to the cell surface. The STRO-1 epitope on HSC70 was mapped to the ATPase domain using a series of deletion mutants in combination with peptide arrays. Deletion of the first four amino acids of the consensus epitope negated STRO-1 binding. Notably, in addition to HSC70, STRO-1 cross-reacts with heat shock protein 70 (HSP70), however all the clonogenic cell activity is restricted to the STRO-1 BRIGHT /HSP70 - fraction. These results provide important insight into the properties that define multipotent MPC and provide the impetus to explore the role of cell surface HSC70 in MPC biology. Stem Cells 2017;35:940-951. © 2016 AlphaMed Press.

  9. Simulation and Optimization of Silicon Solar Cell Back Surface Field

    Directory of Open Access Journals (Sweden)

    Souad TOBBECHE

    2015-11-01

    Full Text Available In this paper, TCAD Silvaco (Technology Computer Aided Design software has been used to study the Back Surface Field (BSF effect of a p+ silicon layer for a n+pp+ silicon solar cell. To study this effect, the J-V characteristics and the external quantum efficiency (EQE are simulated under AM 1.5 illumination for two types of cells. The first solar cell is without BSF (n+p structure while the second one is with BSF (n+pp+ structure. The creation of the BSF on the rear face of the cell results in efficiency h of up to 16.06% with a short-circuit current density Jsc = 30.54 mA/cm2, an open-circuit voltage Voc = 0.631 V, a fill factor FF = 0.832 and a clear improvement of the spectral response obtained in the long wavelengths range. An electric field and a barrier of potential are created by the BSF and located at the junction p+/p with a maximum of 5800 V/cm and 0.15 V, respectively. The optimization of the BSF layer shows that the cell performance improves with the p+ thickness between 0.35 – 0.39 µm, the p+ doping dose is about 2 × 1014 cm-2, the maximum efficiency up to 16.19 %. The cell efficiency is more sensitive to the value of the back surface recombination velocity above a value of 103 cm/s in n+p than n+pp+ solar cell.DOI: http://dx.doi.org/10.5755/j01.ms.21.4.9565

  10. Interaction of Human Tumor Viruses with Host Cell Surface Receptors and Cell Entry

    Directory of Open Access Journals (Sweden)

    Georgia Schäfer

    2015-05-01

    Full Text Available Currently, seven viruses, namely Epstein-Barr virus (EBV, Kaposi’s sarcoma-associated herpes virus (KSHV, high-risk human papillomaviruses (HPVs, Merkel cell polyomavirus (MCPyV, hepatitis B virus (HBV, hepatitis C virus (HCV and human T cell lymphotropic virus type 1 (HTLV-1, have been described to be consistently associated with different types of human cancer. These oncogenic viruses belong to distinct viral families, display diverse cell tropism and cause different malignancies. A key to their pathogenicity is attachment to the host cell and entry in order to replicate and complete their life cycle. Interaction with the host cell during viral entry is characterized by a sequence of events, involving viral envelope and/or capsid molecules as well as cellular entry factors that are critical in target cell recognition, thereby determining cell tropism. Most oncogenic viruses initially attach to cell surface heparan sulfate proteoglycans, followed by conformational change and transfer of the viral particle to secondary high-affinity cell- and virus-specific receptors. This review summarizes the current knowledge of the host cell surface factors and molecular mechanisms underlying oncogenic virus binding and uptake by their cognate host cell(s with the aim to provide a concise overview of potential target molecules for prevention and/or treatment of oncogenic virus infection.

  11. Surface recombination analysis in silicon-heterojunction solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Barrio, R.; Gandia, J.J.; Carabe, J.; Gonzalez, N.; Torres, I. [CIEMAT, Madrid (Spain); Munoz, D.; Voz, C. [Universitat Politecnica de Catalunya, Barcelona (Spain)

    2010-02-15

    The origin of this work is the understanding of the correlation observed between efficiency and emitter-deposition temperature in single silicon-heterojunction solar cells prepared by depositing an n-doped hydrogenated-amorphous-silicon thin film onto a p-type crystalline-silicon wafer. In order to interpret these results, surface-recombination velocities have been determined by two methods, i.e. by fitting the current-voltage characteristics to a theoretical model and by means of the Quasi-Steady-State Photoconductance Technique (QSSPC). In addition, effective diffusion lengths have been estimated from internal quantum efficiencies. The analysis of these data has led to conclude that the performance of the cells studied is limited by back-surface recombination rather than by front-heterojunction quality. A 12%-efficient cell has been prepared by combining optimum emitter-deposition conditions with back-surface-field (BSF) formation by vacuum annealing of the back aluminium contact. This result has been achieved without using any transparent conductive oxide. (author)

  12. Systemic alteration of cell-surface and secreted glycoprotein expression in malignant breast cancer cell lines

    OpenAIRE

    Timpe, Leslie C; Yen, Roger; Haste, Nicole V; Litsakos-Cheung, Christina; Yen, Ten-Yang; Macher, Bruce A

    2013-01-01

    Breast cancer cell lines express fewer transmembrane and secreted glycoproteins than nonmalignant ones. The objective of these experiments was to characterize the changes in the expression of several hundred glycoproteins quantitatively. Secreted and cell-surface glycoproteins were isolated using a glycoprotein capture protocol and then identified by tandem mass spectrometry. Glycoproteins expressed by a group of cell lines originating from malignant tumors of the breast were compared with th...

  13. Paired Expression Analysis of Tumor Cell Surface Antigens

    Directory of Open Access Journals (Sweden)

    Rimas J. Orentas

    2017-08-01

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

  14. RPE cell surface proteins in normal and dystrophic rats

    International Nuclear Information System (INIS)

    Clark, V.M.; Hall, M.O.

    1986-01-01

    Membrane-bound proteins in plasma membrane enriched fractions from cultured rat RPE were analyzed by two-dimensional gel electrophoresis. Membrane proteins were characterized on three increasingly specific levels. Total protein was visualized by silver staining. A maximum of 102 separate proteins were counted in silver-stained gels. Glycoproteins were labeled with 3H-glucosamine or 3H-fucose and detected by autoradiography. Thirty-eight fucose-labeled and 61-71 glucosamine-labeled proteins were identified. All of the fucose-labeled proteins were labeled with glucosamine-derived radioactivity. Proteins exposed at the cell surface were labeled by lactoperoxidase-catalyzed radioiodination prior to preparation of membranes for two-dimensional analysis. Forty separate 125I-labeled surface proteins were resolved by two-dimensional electrophoresis/autoradiography. Comparison with the glycoprotein map showed that a number of these surface labeled proteins were glycoproteins. Two-dimensional maps of total protein, fucose-labeled, and glucosamine-labeled glycoproteins, and 125I-labeled surface proteins of membranes from dystrophic (RCS rdy-p+) and normal (Long Evans or RCS rdy+p+) RPE were compared. No differences in the total protein or surface-labeled proteins were observed. However, the results suggest that a 183K glycoprotein is more heavily glycosylated with glucosamine and fucose in normal RPE membranes as compared to membranes from dystrophic RPE

  15. Defining binding efficiency and specificity of auxins for SCF(TIR1/AFB)-Aux/IAA co-receptor complex formation.

    Science.gov (United States)

    Lee, Sarah; Sundaram, Shanthy; Armitage, Lynne; Evans, John P; Hawkes, Tim; Kepinski, Stefan; Ferro, Noel; Napier, Richard M

    2014-03-21

    Structure-activity profiles for the phytohormone auxin have been collected for over 70 years, and a number of synthetic auxins are used in agriculture. Auxin classification schemes and binding models followed from understanding auxin structures. However, all of the data came from whole plant bioassays, meaning the output was the integral of many different processes. The discovery of Transport Inhibitor-Response 1 (TIR1) and the Auxin F-Box (AFB) proteins as sites of auxin perception and the role of auxin as molecular glue in the assembly of co-receptor complexes has allowed the development of a definitive quantitative structure-activity relationship for TIR1 and AFB5. Factorial analysis of binding activities offered two uncorrelated factors associated with binding efficiency and binding selectivity. The six maximum-likelihood estimators of Efficiency are changes in the overlap matrixes, inferring that Efficiency is related to the volume of the electronic system. Using the subset of compounds that bound strongly, chemometric analyses based on quantum chemical calculations and similarity and self-similarity indices yielded three classes of Specificity that relate to differential binding. Specificity may not be defined by any one specific atom or position and is influenced by coulomb matrixes, suggesting that it is driven by electrostatic forces. These analyses give the first receptor-specific classification of auxins and indicate that AFB5 is the preferred site for a number of auxinic herbicides by allowing interactions with analogues having van der Waals surfaces larger than that of indole-3-acetic acid. The quality factors are also examined in terms of long-standing models for the mechanism of auxin binding.

  16. Surface modification for interaction study with bacteria and preosteoblast cells

    Science.gov (United States)

    Song, Qing

    Surface modification plays a pivotal role in bioengineering. Polymer coatings can provide biocompatibility and biofunctionalities to biomaterials through surface modification. In this dissertation, initiated chemical vapor deposition (iCVD) was utilized to coat two-dimensional (2D) and three-dimensional (3D) substrates with differently charged polyelectrolytes in order to generate antimicrobial and osteocompatible biomaterials. ICVD is a modified CVD technique that enables surface modification in an all-dry condition without substrate damage and solvent contamination. The free-radical polymerization allows the vinyl polymers to conformally coat on various micro- and nano-structured substrates and maintains the delicate structure of the functional groups. The vapor deposition of polycations provided antimicrobial activity to planar and porous substrates through destroying the negatively charged bacterial membrane and brought about high contact-killing efficiency (99.99%) against Gram-positive Bacillus subtilis and Gram-negative Escherichia coli. Additionally, the polyampholytes synthesized by iCVD exhibited excellent antifouling performance against the adhesion of Gram-positive Listeria innocua and Gram-negative E. coli in phosphate buffered saline (PBS). Their antifouling activities were attributed to the electrostatic interaction and hydration layers that served as physical and energetic barriers to prevent bacterial adhesion. The contact-killing and antifouling polymers synthesized by iCVD can be applied to surface modification of food processing equipment and medical devices with the aim of reducing foodborne diseases and medical infections. Moreover, the charged polyelectrolyte modified 2D polystyrene surfaces displayed good osteocompatibility and enhanced osteogenesis of preosteoblast cells than the un-modified polystyrene surface. In order to promote osteoinduction of hydroxyapatite (HA) scaffolds, bioinspired polymer-controlled mineralization was conducted

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

    DEFF Research Database (Denmark)

    Dolatshahi-Pirouz, Alireza; Jensen, T H L; Kolind, K

    2011-01-01

    . In subsequent cell studies with hMSC's we studied the cell spreading, cytoskeletal organization and cell morphology on the respective surfaces. When the cells were adsorbed on the uncoated substrates, a diffuse cell actin cytoskeleton was revealed, and the cells had a highly elongated shape. On the fibronectin...... coated surfaces the cells adapted to a more polygonal shape with a well-defined actin cytoskeleton, while a larger cell area and roundness values were observed for cells cultured on the coated surfaces. Among the coated surfaces a slightly larger cell area and roundness values was observed on HA......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...

  18. Functional mapping of cell surface proteins with localized stimulation of single cells

    Science.gov (United States)

    Sun, Bingyun; Chiu, Daniel T.

    2003-11-01

    This paper describes the development of using individual micro and nano meter-sized vesicles as delivery vessels to functionally map the distribution of cell surface proteins at the level of single cells. The formation of different sizes of vesicles from tens of nanometers to a few micrometers in diameter that contain the desired molecules is addressed. An optical trap is used to manipulate the loaded vesicle to specific cell morphology of interest, and a pulsed UV laser is used to photo-release the stimuli onto the cell membrane. Carbachol activated cellular calcium flux, upon binding to muscarinic acetylcholine receptors, is studied by this method, and the potential of using this method for the functional mapping of localized proteins on the cell surface membrane is discussed.

  19. The outer membrane protein TolC of Vibrio cholerae serves as a second cell-surface receptor for the VP3 phage.

    Science.gov (United States)

    Fan, Fenxia; Li, Xu; Pang, Bo; Zhang, Cheng; Li, Zhe; Zhang, Lijuan; Li, Jie; Zhang, Jingyun; Yan, Meiying; Liang, Weili; Kan, Biao

    2017-12-19

    Receptor recognition is a key step in the initiation of phage infection. Previously, we found that VP3, the T7 family phage of the Vibrio cholerae serogroup O1 biotype El Tor, can adsorb to the core oligosaccharide (OS) of lipopolysaccharides of V. cholerae. However, some wild-type strains of V. cholerae possessing the intact OS gene cluster still have VP3 binding but are resistant to VP3 infection. Moreover, an OS gene deletion mutant still exhibits weak VP3 binding, suggesting multiple factors are possibly involved in VP3 binding to V. cholerae. Here, we report that the outer-membrane protein TolC of V. cholerae is involved in the host adsorption of VP3. We observed that TolC directly interacts with the VP3 tail fiber protein gp44 and its C-terminal domains, and also found that three amino acid residues in the outside loops of TolC, at positions 78, 290 and 291, are critical for binding to gp44. Among VP3-resistant wild type V. cholerae strains, frequent amino acid residue mutations were observed in the loops around the sites 78, 290 and 291 which were predicted to be exposed to the cell surface. These findings reveal a co-receptor-binding mechanism for VP3 infection of V. cholerae and that both outer membrane TolC and OS are necessary for successful VP3 infection of V. cholerae. We conclude that mutations on the outside loops of the receptor may confer V. cholerae strains with VP3 phage-resistance, enabling these strains to survive in environments containing VP3 or related phages. Copyright © 2017, The American Society for Biochemistry and Molecular Biology.

  20. Enhanced cell surface expression, immunogenicity and genetic stability resulting from a spontaneous truncation of HIV Env expressed by a recombinant MVA

    International Nuclear Information System (INIS)

    Wyatt, Linda S.; Belyakov, Igor M.; Earl, Patricia L.; Berzofsky, Jay A.; Moss, Bernard

    2008-01-01

    During propagation of modified vaccinia virus Ankara (MVA) encoding HIV 89.6 Env, a few viral foci stained very prominently. Virus cloned from such foci replicated to higher titers than the parent and displayed enhanced genetic stability on passage. Sequence analysis showed a single nucleotide deletion in the 89.6 env gene of the mutant that caused a frame shift and truncation of 115 amino acids from the cytoplasmic domain. The truncated Env was more highly expressed on the cell surface, induced higher antibody responses than the full-length Env, reacted with HIV neutralizing monoclonal antibodies and mediated CD4/co-receptor-dependent fusion. Intramuscular (IM), intradermal (ID) needleless, and intrarectal (IR) catheter inoculations gave comparable serum IgG responses. However, intraoral (IO) needleless injector route gave the highest IgA in lung washings and IR gave the highest IgA and IgG responses in fecal extracts. Induction of CTL responses in the spleens of individual mice as assayed by intracellular cytokine staining was similar with both the full-length and truncated Env constructs. Induction of acute and memory CTL in the spleens of mice immunized with the truncated Env construct by ID, IO, and IR routes was comparable and higher than by the IM route, but only the IR route induced CTL in the gut-associated lymphoid tissue. Thus, truncation of Env enhanced genetic stability as well as serum and mucosal antibody responses, suggesting the desirability of a similar modification in MVA-based candidate HIV vaccines

  1. Interaction of KSHV with Host Cell Surface Receptors and Cell Entry

    Directory of Open Access Journals (Sweden)

    Mohanan Valiya Veettil

    2014-10-01

    Full Text Available Virus entry is a complex process characterized by a sequence of events. Since the discovery of KSHV in 1994, tremendous progress has been made in our understanding of KSHV entry into its in vitro target cells. KSHV entry is a complex multistep process involving viral envelope glycoproteins and several cell surface molecules that is utilized by KSHV for its attachment and entry. KSHV has a broad cell tropism and the attachment and receptor engagement on target cells have an important role in determining the cell type-specific mode of entry. KSHV utilizes heparan sulfate, integrins and EphrinA2 molecules as receptors which results in the activation of host cell pre-existing signal pathways that facilitate the subsequent cascade of events resulting in the rapid entry of virus particles, trafficking towards the nucleus followed by viral and host gene expression. KSHV enters human fibroblast cells by dynamin dependant clathrin mediated endocytosis and by dynamin independent macropinocytosis in dermal endothelial cells. Once internalized into endosomes, fusion of the viral envelope with the endosomal membranes in an acidification dependent manner results in the release of capsids which subsequently reaches the nuclear pore vicinity leading to the delivery of viral DNA into the nucleus. In this review, we discuss the principal mechanisms that enable KSHV to interact with the host cell surface receptors as well as the mechanisms that are required to modulate cell signaling machinery for a successful entry.

  2. Interaction of KSHV with Host Cell Surface Receptors and Cell Entry

    Science.gov (United States)

    Veettil, Mohanan Valiya; Bandyopadhyay, Chirosree; Dutta, Dipanjan; Chandran, Bala

    2014-01-01

    Virus entry is a complex process characterized by a sequence of events. Since the discovery of KSHV in 1994, tremendous progress has been made in our understanding of KSHV entry into its in vitro target cells. KSHV entry is a complex multistep process involving viral envelope glycoproteins and several cell surface molecules that is utilized by KSHV for its attachment and entry. KSHV has a broad cell tropism and the attachment and receptor engagement on target cells have an important role in determining the cell type-specific mode of entry. KSHV utilizes heparan sulfate, integrins and EphrinA2 molecules as receptors which results in the activation of host cell pre-existing signal pathways that facilitate the subsequent cascade of events resulting in the rapid entry of virus particles, trafficking towards the nucleus followed by viral and host gene expression. KSHV enters human fibroblast cells by dynamin dependant clathrin mediated endocytosis and by dynamin independent macropinocytosis in dermal endothelial cells. Once internalized into endosomes, fusion of the viral envelope with the endosomal membranes in an acidification dependent manner results in the release of capsids which subsequently reaches the nuclear pore vicinity leading to the delivery of viral DNA into the nucleus. In this review, we discuss the principal mechanisms that enable KSHV to interact with the host cell surface receptors as well as the mechanisms that are required to modulate cell signaling machinery for a successful entry. PMID:25341665

  3. Cancer cell uptake behavior of Au nanoring and its localized surface plasmon resonance induced cell inactivation

    Science.gov (United States)

    Chu, Che-Kuan; Tu, Yi-Chou; Chang, Yu-Wei; Chu, Chih-Ken; Chen, Shih-Yang; Chi, Ting-Ta; Kiang, Yean-Woei; Yang, Chih-Chung

    2015-02-01

    Au nanorings (NRIs), which have the localized surface plasmon resonance (LSPR) wavelength around 1058 nm, either with or without linked antibodies, are applied to SAS oral cancer cells for cell inactivation through the LSPR-induced photothermal effect when they are illuminated by a laser of 1065 nm in wavelength. Different incubation times of cells with Au NRIs are considered for observing the variations of cell uptake efficiency of Au NRI and the threshold laser intensity for cell inactivation. In each case of incubation time, the cell sample is washed for evaluating the total Au NRI number per cell adsorbed and internalized by the cells based on inductively coupled plasma mass spectrometry measurement. Also, the Au NRIs remaining on cell membrane are etched with KI/I2 solution to evaluate the internalized Au NRI number per cell. The threshold laser intensities for cell inactivation before washout, after washout, and after KI/I2 etching are calibrated from the circular area sizes of inactivated cells around the illuminated laser spot center with various laser power levels. By using Au NRIs with antibodies, the internalized Au NRI number per cell increases monotonically with incubation time up to 24 h. However, the number of Au NRI remaining on cell membrane reaches a maximum at 12 h in incubation time. The cell uptake behavior of an Au NRI without antibodies is similar to that with antibodies except that the uptake NRI number is significantly smaller and the incubation time for the maximum NRI number remaining on cell membrane is delayed to 20 h. By comparing the threshold laser intensities before and after KI/I2 etching, it is found that the Au NRIs remaining on cell membrane cause more effective cancer cell inactivation, when compared with the internalized Au NRIs.

  4. Cancer cell uptake behavior of Au nanoring and its localized surface plasmon resonance induced cell inactivation

    International Nuclear Information System (INIS)

    Chu, Che-Kuan; Tu, Yi-Chou; Chang, Yu-Wei; Chu, Chih-Ken; Chen, Shih-Yang; Chi, Ting-Ta; Kiang, Yean-Woei; Yang, Chih-Chung

    2015-01-01

    Au nanorings (NRIs), which have the localized surface plasmon resonance (LSPR) wavelength around 1058 nm, either with or without linked antibodies, are applied to SAS oral cancer cells for cell inactivation through the LSPR-induced photothermal effect when they are illuminated by a laser of 1065 nm in wavelength. Different incubation times of cells with Au NRIs are considered for observing the variations of cell uptake efficiency of Au NRI and the threshold laser intensity for cell inactivation. In each case of incubation time, the cell sample is washed for evaluating the total Au NRI number per cell adsorbed and internalized by the cells based on inductively coupled plasma mass spectrometry measurement. Also, the Au NRIs remaining on cell membrane are etched with KI/I 2 solution to evaluate the internalized Au NRI number per cell. The threshold laser intensities for cell inactivation before washout, after washout, and after KI/I 2 etching are calibrated from the circular area sizes of inactivated cells around the illuminated laser spot center with various laser power levels. By using Au NRIs with antibodies, the internalized Au NRI number per cell increases monotonically with incubation time up to 24 h. However, the number of Au NRI remaining on cell membrane reaches a maximum at 12 h in incubation time. The cell uptake behavior of an Au NRI without antibodies is similar to that with antibodies except that the uptake NRI number is significantly smaller and the incubation time for the maximum NRI number remaining on cell membrane is delayed to 20 h. By comparing the threshold laser intensities before and after KI/I 2 etching, it is found that the Au NRIs remaining on cell membrane cause more effective cancer cell inactivation, when compared with the internalized Au NRIs. (paper)

  5. Evaluation of Relative Yeast Cell Surface Hydrophobicity Measured by Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Lisa Colling

    2005-01-01

    Full Text Available Objective: To develop an efficient method for evaluating cell surface hydrophobicity and to apply the method to demonstrate the effects of fungal growth conditions on cell surface properties.

  6. Imaging and reconstruction of cell cortex structures near the cell surface

    Science.gov (United States)

    Jin, Luhong; Zhou, Xiaoxu; Xiu, Peng; Luo, Wei; Huang, Yujia; Yu, Feng; Kuang, Cuifang; Sun, Yonghong; Liu, Xu; Xu, Yingke

    2017-11-01

    Total internal reflection fluorescence microscopy (TIRFM) provides high optical sectioning capability and superb signal-to-noise ratio for imaging of cell cortex structures. The development of multi-angle (MA)-TIRFM permits high axial resolution imaging and reconstruction of cellular structures near the cell surface. Cytoskeleton is composed of a network of filaments, which are important for maintenance of cell function. The high-resolution imaging and quantitative analysis of filament organization would contribute to our understanding of cytoskeleton regulation in cell. Here, we used a custom-developed MA-TIRFM setup, together with stochastic photobleaching and single molecule localization method, to enhance the lateral resolution of TIRFM imaging to about 100 nm. In addition, we proposed novel methods to perform filament segmentation and 3D reconstruction from MA-TIRFM images. Furthermore, we applied these methods to study the 3D localization of cortical actin and microtubule structures in U373 cancer cells. Our results showed that cortical actins localize ∼ 27 nm closer to the plasma membrane when compared with microtubules. We found that treatment of cells with chemotherapy drugs nocodazole and cytochalasin B disassembles cytoskeletal network and induces the reorganization of filaments towards the cell periphery. In summary, this study provides feasible approaches for 3D imaging and analyzing cell surface distribution of cytoskeletal network. Our established microscopy platform and image analysis toolkits would facilitate the study of cytoskeletal network in cells.

  7. Surface receptor Toso controls B cell-mediated regulation of T cell immunity.

    Science.gov (United States)

    Yu, Jinbo; Duong, Vu Huy Hoang; Westphal, Katrin; Westphal, Andreas; Suwandi, Abdulhadi; Grassl, Guntram A; Brand, Korbinian; Chan, Andrew C; Föger, Niko; Lee, Kyeong-Hee

    2018-04-03

    The immune system is tightly controlled by regulatory processes that allow for the elimination of invading pathogens, while limiting immunopathological damage to the host. In the present study, we found that conditional deletion of the cell surface receptor Toso on B cells unexpectedly resulted in impaired proinflammatory T cell responses, which led to impaired immune protection in an acute viral infection model and was associated with reduced immunopathological tissue damage in a chronic inflammatory context. Toso exhibited its B cell-inherent immunoregulatory function by negatively controlling the pool of IL-10-competent B1 and B2 B cells, which were characterized by a high degree of self-reactivity and were shown to mediate immunosuppressive activity on inflammatory T cell responses in vivo. Our results indicate that Toso is involved in the differentiation/maintenance of regulatory B cells by fine-tuning B cell receptor activation thresholds. Furthermore, we showed that during influenza A-induced pulmonary inflammation, the application of Toso-specific antibodies selectively induced IL-10-competent B cells at the site of inflammation and resulted in decreased proinflammatory cytokine production by lung T cells. These findings suggest that Toso may serve as a novel therapeutic target to dampen pathogenic T cell responses via the modulation of IL-10-competent regulatory B cells.

  8. Mechanotransduction across the cell surface and through the cytoskeleton

    Science.gov (United States)

    Wang, N.; Butler, J. P.; Ingber, D. E.

    1993-01-01

    Mechanical stresses were applied directly to cell surface receptors with a magnetic twisting device. The extracellular matrix receptor, integrin beta 1, induced focal adhesion formation and supported a force-dependent stiffening response, whereas nonadhesion receptors did not. The cytoskeletal stiffness (ratio of stress to strain) increased in direct proportion to the applied stress and required intact microtubules and intermediate filaments as well as microfilaments. Tensegrity models that incorporate mechanically interdependent struts and strings that reorient globally in response to a localized stress mimicked this response. These results suggest that integrins act as mechanoreceptors and transmit mechanical signals to the cytoskeleton. Mechanotransduction, in turn, may be mediated simultaneously at multiple locations inside the cell through force-induced rearrangements within a tensionally integrated cytoskeleton.

  9. Surface Passivation of CIGS Solar Cells Using Gallium Oxide

    KAUST Repository

    Garud, Siddhartha

    2018-02-27

    This work proposes gallium oxide grown by plasma-enhanced atomic layer deposition, as a surface passivation material at the CdS buffer interface of Cu(In,Ga)Se2 (CIGS) solar cells. In preliminary experiments, a metal-insulator-semiconductor (MIS) structure is used to compare aluminium oxide, gallium oxide, and hafnium oxide as passivation layers at the CIGS-CdS interface. The findings suggest that gallium oxide on CIGS may show a density of positive charges and qualitatively, the least interface trap density. Subsequent solar cell results with an estimated 0.5 nm passivation layer show an substantial absolute improvement of 56 mV in open-circuit voltage (VOC), 1 mA cm−2 in short-circuit current density (JSC), and 2.6% in overall efficiency as compared to a reference (with the reference showing 8.5% under AM 1.5G).

  10. Cell-surface display of enzymes by the yeast Saccharomyces cerevisiae for synthetic biology.

    Science.gov (United States)

    Tanaka, Tsutomu; Kondo, Akihiko

    2015-02-01

    In yeast cell-surface displays, functional proteins, such as cellulases, are genetically fused to an anchor protein and expressed on the cell surface. Saccharomyces cerevisiae, which is often utilized as a cell factory for the production of fuels, chemicals, and proteins, is the most commonly used yeast for cell-surface display. To construct yeast cells with a desired function, such as the ability to utilize cellulose as a substrate for bioethanol production, cell-surface display techniques for the efficient expression of enzymes on the cell membrane need to be combined with metabolic engineering approaches for manipulating target pathways within cells. In this Minireview, we summarize the recent progress of biorefinery fields in the development and application of yeast cell-surface displays from a synthetic biology perspective and discuss approaches for further enhancing cell-surface display efficiency. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  11. Control of cell behavior on PTFE surface using ion beam irradiation

    International Nuclear Information System (INIS)

    Kitamura, Akane; Kobayashi, Tomohiro; Meguro, Takashi; Suzuki, Akihiro; Terai, Takayuki

    2009-01-01

    A polytetrafluoroethylene (PTFE) surface is smooth and biologically inert, so that cells cannot attach to it. Ion beam irradiation of the PTFE surface forms micropores and a melted layer, and the surface is finally covered with a large number of small protrusions. Recently, we found that cells could adhere to this irradiated PTFE surface and spread over the surface. Because of their peculiar attachment behavior, these surfaces can be used as biological tools. However, the factors regulating cell adhesion are still unclear, although some new functional groups formed by irradiation seem to contribute to this adhesion. To control cell behavior on PTFE surfaces, we must determine the effects of the outermost irradiated surface on cell adhesion. In this study, we removed the thin melted surface layer by postirradiation annealing and investigated cell behavior on the surface. On the surface irradiated with 3 x 10 16 ions/cm 2 , cells spread only on the remaining parts of the melted layer. From these results, it is clear that the melted layer had a capacity for cell attachment. When the surface covered with protrusions was irradiated with a fluence of 1 x 10 17 ions/cm 2 , the distribution of cells changed after the annealing process from 'sheet shaped' into multicellular aggregates with diameters of around 50 μm. These results indicate that we can control cell behavior on PTFE surfaces covered with protrusions using irradiation and subsequent annealing. Multicellular spheroids can be fabricated for tissue engineering using this surface.

  12. Ultraviolet light depletes surface markers of Langerhans cells

    International Nuclear Information System (INIS)

    Aberer, W.; Schuler, G.; Stingl, G.; Hoenigsmann, H.; Wolff, K.

    1981-01-01

    This report defines the influence of ultraviolet light (UV) on Langerhans cells (LC). Human volunteers and hairless mice (Swiss ha/ha) were exposed to various single and/or cumulative doses of either UV-A, UV-B, or UV-A plus small amounts of UV-B (UV-A (+B)). 24 hr after the last irradiation, morphology of the entire epidermis was evaluated by both light and electron microscopy while LC, in addition, were tested for expression of specific histochemical (ATPase) and functional immunological markers (Ia antigens). In both men and mice, cumulative doses of either 80-120 J/cm2 UV-A (+B) or 1-2 X 100 J/cm2 UV-A resulted in a dramatic reduction of cells exhibiting ATPase and Ia-reactivity. In the UV-B spectrum, single doses of 60-80 mJ/cm2 produced a virtually complete elimination of LC membrane markers. By contrast, pemphigus antigens of keratinocytes were unaffected by these energy doses. Electron microscopy revealed cellular damage of some LC after UV-doses which produce a virtually complete abolition of LC membrane markers. At certain dose ranges (15-30 mJ/cm2 UV-B and 1 x 40 to 2 x 100 J/cm2 UV-A) LC were the only epidermal cells to display morphological damage at the ultrastructural level whereas higher doses affected all epidermal cells. The finding that LC surface markers and to a lesser extent the cells themselves are particularly susceptible to UV irradiation has important implications in view of previous findings that LC are potent stimulators of antigen-specific and allogeneic T cell activation. UV-induced alteration of LC plasma membrane integrity may represent a tool to manipulate adverse immune reactions involving the epidermis

  13. Endothelialization of artificial surfaces: does surface tension determine in vitro growth of human saphenous vein endothelial cells?

    Science.gov (United States)

    Fasol, R; Zilla, P; Deutsch, M; Fischlein, T; Kadletz, M; Griesmacher, A; Müller, M M

    1987-06-01

    To evaluate the possibility of providing, in vitro, an endothelial lining for artificial hearts, we cultivated adult autologous endothelial cells on two polyurethane and two silicone rubber surfaces. Over the ensuing 11-day period, we investigated the resulting cell proliferation and morphology by means of scanning electron and light microscopy. On the silicone rubber surfaces, seeding of 200,000 human saphenous vein endothelial cells per cm(2) produced an ideal cobblestone monolayer within a single day. In contrast, the polyurethane surfaces displayed an uneven, patchy distribution of endothelial cells. Scanning electron microscopy revealed microvilli and marginal overlapping in both groups. After the first day, the cell count on the polyurethane surfaces increased, whereas the count on the silicone rubber surfaces decreased. Morphologic investigations revealed that the ideally shaped cells initially on the silicone rubber had begun to overspread and subsequently to become detached, leaving denuded spheroid areas. Moreover, cultivation for 11 days on the polyurethane surfaces resulted in an unevenness of cell distribution that far exceeded the unevenness seen on the first day. Thus, despite the fact that materials with a high surface tension (such as silicone rubbers) seem to be ideal for initial cell spreading, subsequent cultivation results in cell detachment and death. On materials with a lower surface tension (such as polyurethanes), the less differentiated monolayers do at least proliferate, although their morphology remains unsatisfactory. Even if adult human endothelial cells should prove shear-stress-resistant, a minimum of 6 to 8 weeks would be required to establish autologous endothelial cell monolayers on the inner surface. Therefore, the endothelialization of artificial hearts is not possible when such hearts are used for urgent "bridging" before cardiac transplantation.

  14. Surface-Engineering of Red Blood Cells as Artificial Antigen Presenting Cells Promising for Cancer Immunotherapy.

    Science.gov (United States)

    Sun, Xiaoqi; Han, Xiao; Xu, Ligeng; Gao, Min; Xu, Jun; Yang, Rong; Liu, Zhuang

    2017-10-01

    The development of artificial antigen presenting cells (aAPCs) to mimic the functions of APCs such as dendritic cells (DCs) to stimulate T cells and induce antitumor immune responses has attracted substantial interests in cancer immunotherapy. In this work, a unique red blood cell (RBC)-based aAPC system is designed by engineering antigen peptide-loaded major histocompatibility complex-I and CD28 activation antibody on RBC surface, which are further tethered with interleukin-2 (IL2) as a proliferation and differentiation signal. Such RBC-based aAPC-IL2 (R-aAPC-IL2) can not only provide a flexible cell surface with appropriate biophysical parameters, but also mimic the cytokine paracrine delivery. Similar to the functions of matured DCs, the R-aAPC-IL2 cells can facilitate the proliferation of antigen-specific CD8+ T cells and increase the secretion of inflammatory cytokines. As a proof-of-concept, we treated splenocytes from C57 mice with R-aAPC-IL2 and discovered those splenocytes induced significant cancer-cell-specific lysis, implying that the R-aAPC-IL2 were able to re-educate T cells and induce adoptive immune response. This work thus presents a novel RBC-based aAPC system which can mimic the functions of antigen presenting DCs to activate T cells, promising for applications in adoptive T cell transfer or even in direct activation of circulating T cells for cancer immunotherapy. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Endothelialization of Artificial Surfaces: Does Surface Tension Determine in vitro Growth of Human Saphenous Vein Endothelial Cells?

    OpenAIRE

    Fasol, Roland; Zilla, Peter; Deutsch, Manfred; Fischlein, Teddy; Kadletz, Margit; Griesmacher, Andrea; Müller, Mathias M.

    1987-01-01

    To evaluate the possibility of providing, in vitro, an endothelial lining for artificial hearts, we cultivated adult autologous endothelial cells on two polyurethane and two silicone rubber surfaces. Over the ensuing 11-day period, we investigated the resulting cell proliferation and morphology by means of scanning electron and light microscopy. On the silicone rubber surfaces, seeding of 200,000 human saphenous vein endothelial cells per cm2 produced an ideal cobblestone monolayer within a s...

  16. A monoclonal antibody recognizes undifferentiation-specific carbohydrate moieties expressed on cell surface of the human dental pulp cells.

    Science.gov (United States)

    Kang, Kyung-Jung; Ko, Seon-Yle; Ryu, Chun-Jeih; Jang, Young-Joo

    2017-05-01

    Human dental pulp cells are obtained from dental pulp tissue, and have the ability to form dentin and a pulp-like complex. Although adult stem cells have been identified from the primary culture by using specific cell surface markers, the identity of surface markers for the purification of stem cells within the dental pulp population are still unclear. Previously, we had constructed monoclonal antibodies against the undifferentiated cell-specific surface markers of human dental pulp cells (hDPCs) by performing decoy immunization. Among them, a monoclonal antibody against the cell surface antigen of the undifferentiated hDPCs (named UPSA-1) was purified and its heavy and light chain consensus regions were analyzed. The cell surface binding affinity of UPSA-1 mAb on the undifferentiated hDPCs was stronger than that on the differentiated cells. When tunicamycin was applied to hDPSCs during culture, the cell surface binding affinity of the antibody was dramatically decreased, and dentinogenic differentiation was reduced. The purified UPSA-1 antigen band resulting from immunoprecipitation disappeared or shifted down on the SDS-PAGE by deglycosylation. These data suggested that glycosylation on the cell surface might be a marker of an undifferentiated state, and that UPSA-1 mAb might be useful for identifying the carbohydrate moiety on the cell surface of undifferentiated pulp cells. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  17. Substance P Increases Cell-Surface Expression of CD74 (Receptor for Macrophage Migration Inhibitory Factor: In Vivo Biotinylation of Urothelial Cell-Surface Proteins

    Directory of Open Access Journals (Sweden)

    Katherine L. Meyer-Siegler

    2009-01-01

    N-hydroxysulfosuccinimide biotin ester-labeled surface urothelial proteins in rats treated either with saline or substance P (SP, 40 μg/kg. The bladder was examined by histology and confocal microscopy. Biotinylated proteins were purified by avidin agarose, immunoprecipitated with anti-MIF or anti-CD74 antibodies, and detected with strepavidin-HRP. Only superficial urothelial cells were biotinylated. These cells contained a biotinylated MIF/CD74 cell-surface complex that was increased in SP-treated animals. SP treatment increased MIF and CD74 mRNA in urothelial cells. Our data indicate that intraluminal MIF, released from urothelial cells as a consequence of SP treatment, interacts with urothelial cell-surface CD74. These results document that our previously described MIF-CD74 interaction occurs at the urothelial cell surface.

  18. Odorant receptor co-receptor Orco is upregulated by methyl eugenol in male Bactrocera dorsalis (Diptera: Tephritidae).

    Science.gov (United States)

    Zheng, Weiwei; Zhu, Chipan; Peng, Tao; Zhang, Hongyu

    2012-08-01

    Bactrocera dorsalis is a destructive fruit-eating pest that causes severe economic damage to the fruit and vegetable industry. Methyl eugenol (ME) has been widely used as an effective sexual attractant for male fruit flies through olfactory perception. However, the molecular mechanism underlying the olfactory perception of ME remains unknown. Here, we report the characterization and functional analysis of a newly discovered cDNA that encodes a Drosophila melanogaster odorant receptor co-receptor Orco ortholog in B. dorsalis. qRT-PCR analysis revealed that it was abundantly expressed in the antenna of adult B. dorsalis. Notably, Orco was upregulated by ME in the antenna of male flies. Mature males of B. dorsalis showed significant taxis toward ME within 0.5h, and Orco was significantly upregulated in the attracted adults within the same period. Silencing Orco through the ingestion of dsRNA reduced the attractive effects of ME. These data suggest that Orco may play an essential role in ME attraction in the olfactory signal transduction pathway. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Genetic predisposition of variants in TLR2 and its co-receptors to severe malaria in Odisha, India.

    Science.gov (United States)

    Panigrahi, Subhendu; Kar, Avishek; Tripathy, Sagnika; Mohapatra, Manoj K; Dhangadamajhi, Gunanidhi

    2016-02-01

    Although the role of TLRs signalling in malaria pathogenesis is well established, contribution of individual TLR to clinical outcome of malaria still remains inconclusive. Given the importance of TLR2 and its co-receptors in recognising distinct structural forms of key malaria toxins and mediating innate immune response, it is essential to delineate their genetic contribution. Variants in TLR1 (I602S) and TLR6 (P249S) were genotyped by PCR-RFLP methods, and TLR2 (I/D) was genotyped by PCR in 200 samples each from uncomplicated malaria (UM) and severe malaria (SM). Further, SM was categorised into its sub-clinical groups (CM and NCSM or SOD and MODS) and analysed. The results showed the PP genotype of TLR6 (P249S) to be significantly more common in UM (P genetic predisposition to SM and that its association with either TLR2 'D' or TLR1 '602S' modulates for CM development. The present study opens up several new avenues for their exploration and validation in future studies in different global settings for malaria.

  20. Bio-Orthogonal Mediated Nucleic Acid Transfection of Cells via Cell Surface Engineering

    Science.gov (United States)

    2017-01-01

    The efficient delivery of foreign nucleic acids (transfection) into cells is a critical tool for fundamental biomedical research and a pillar of several biotechnology industries. There are currently three main strategies for transfection including reagent, instrument, and viral based methods. Each technology has significantly advanced cell transfection; however, reagent based methods have captured the majority of the transfection market due to their relatively low cost and ease of use. This general method relies on the efficient packaging of a reagent with nucleic acids to form a stable complex that is subsequently associated and delivered to cells via nonspecific electrostatic targeting. Reagent transfection methods generally use various polyamine cationic type molecules to condense with negatively charged nucleic acids into a highly positively charged complex, which is subsequently delivered to negatively charged cells in culture for association, internalization, release, and expression. Although this appears to be a straightforward procedure, there are several major issues including toxicity, low efficiency, sorting of viable transfected from nontransfected cells, and limited scope of transfectable cell types. Herein, we report a new strategy (SnapFect) for nucleic acid transfection to cells that does not rely on electrostatic interactions but instead uses an integrated approach combining bio-orthogonal liposome fusion, click chemistry, and cell surface engineering. We show that a target cell population is rapidly and efficiently engineered to present a bio-orthogonal functional group on its cell surface through nanoparticle liposome delivery and fusion. A complementary bio-orthogonal nucleic acid complex is then formed and delivered to which chemoselective click chemistry induced transfection occurs to the primed cell. This new strategy requires minimal time, steps, and reagents and leads to superior transfection results for a broad range of cell types

  1. Distinct roles for dystroglycan, beta1 integrin and perlecan in cell surface laminin organization

    DEFF Research Database (Denmark)

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

    2001-01-01

    integrin-deficient ES cells, laminin-1 binds to the cell surface, but fails to organize into more morphologically complex structures. This result indicates that beta1 integrin function is required after DG function in the cell surface-mediated laminin assembly process. In perlecan-deficient ES cells......Dystroglycan (DG) is a cell surface receptor for several extracellular matrix (ECM) molecules including laminins, agrin and perlecan. Recent data indicate that DG function is required for the formation of basement membranes in early development and the organization of laminin on the cell surface...

  2. Enteroendocrine cells are specifically marked by cell surface expression of claudin-4 in mouse small intestine.

    Directory of Open Access Journals (Sweden)

    Takahiro Nagatake

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

  3. Interaction of progenitor bone cells with different surface modifications of titanium implant

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Wen-Cheng, E-mail: wencchen@fcu.edu.tw [Advanced Medical Devices and Composites Laboratory, Department of Fiber and Composite Materials, College of Engineering, Feng Chia University, Taichung 40724, Taiwan (China); Chen, Ya-Shun [Advanced Medical Devices and Composites Laboratory, Department of Fiber and Composite Materials, College of Engineering, Feng Chia University, Taichung 40724, Taiwan (China); Ko, Chia-Ling [Advanced Medical Devices and Composites Laboratory, Department of Fiber and Composite Materials, College of Engineering, Feng Chia University, Taichung 40724, Taiwan (China); Dental Medical Devices and Materials Research Center, College of Dental Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan (China); Lin, Yi; Kuo, Tzu-Huang; Kuo, Hsien-Nan [Medical Device Development Division, Metal Industries Research and Development Centre, Kaohsiung 82151, Taiwan (China)

    2014-04-01

    Changes in the physical and chemical properties of Ti surfaces can be attributed to cell performance, which improves surface biocompatibility. The cell proliferation, mineralization ability, and gene expression of progenitor bone cells (D1 cell) were compared on five different Ti surfaces, namely, mechanical grinding (M), electrochemical modification through potentiostatic anodization (ECH), sandblasting and acid etching (SLA), sandblasting, hydrogen peroxide treatment, and heating (SAOH), and sandblasting, alkali heating, and etching (SMART). SAOH treatment produced the most hydrophilic surface, whereas SLA produced the most hydrophobic surface. Cell activity indicated that SLA and SMART produced significantly rougher surfaces and promoted D1 cell attachment within 1 day of culturing, whereas SAOH treatment produced moderate roughness (Ra = 1.26 μm) and accelerated the D1 cell proliferation up to 7 days after culturing. The ECH surface significantly promoted alkaline phosphatase (ALP) expression and osteocalcin (OCN) secretion in the D1 cells compared with the other surface groups. The ECH and SMART-treated Ti surfaces resulted in maximum ALP and OCN expressions during the D1 cell culture. SLA, SAOH, and SMART substrate surfaces were rougher and exhibited better cell metabolic responses during the early stage of cell attachment, proliferation, and morphologic expressions within 1 day of D1 cell culture. The D1 cells cultured on the ECH and SMART substrates exhibited higher differentiation, and higher ALP and OCN expressions after 10 days of culture. Thus, the ECH and SMART treatments promote better ability of cell mineralization in vitro, which demonstrate their great potential for clinical use. - Highlights: • Progenitor bone cells onto Ti with different modifications are characterized. • Surface roughness and hydrophilicity encourage early stage cell attachment. • Composition and surface treatments are more vital in bone cell mineralization.

  4. The Cell-Surface N-Glycome of Human Embryonic Stem Cells and Differentiated Hepatic Cells thereof.

    Science.gov (United States)

    Montacir, Houda; Freyer, Nora; Knöspel, Fanny; Urbaniak, Thomas; Dedova, Tereza; Berger, Markus; Damm, Georg; Tauber, Rudolf; Zeilinger, Katrin; Blanchard, Véronique

    2017-07-04

    Human embryonic stem cells (hESCs) are pluripotent stem cells that offer a wide range of applications in regenerative medicine. In addition, they have been proposed as an appropriate alternative source of hepatocytes. In this work, hESCs were differentiated into definitive endodermal cells (DECs), followed by maturation into hepatocyte-like cells (HLCs). Their cell-surface N-glycome was profiled and also compared with that of primary human hepatocytes (PHHs). Undifferentiated hESCs contained large amounts of high-mannose N-glycans. In contrast, complex-type N-glycans such as asialylated or monosialylated biantennary and triantennary N-glycans were dominant in HLCs, and fully galactosylated structures were significantly more abundant than in undifferentiated hESCs. The cell-surface N-glycosylation of PHHs was more biologically processed than that of HLCs, with bisialylated biantennary and trisialylated triantennary structures predominant. This is the first report of the cell surface N-glycome of PHHs and of HLCs being directly generated from hESCs without embryoid body formation. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Continuous signaling of CD79b and CD19 is required for the fitness of Burkitt lymphoma B cells.

    Science.gov (United States)

    He, Xiaocui; Kläsener, Kathrin; Iype, Joseena M; Becker, Martin; Maity, Palash C; Cavallari, Marco; Nielsen, Peter J; Yang, Jianying; Reth, Michael

    2018-04-18

    Expression of the B-cell antigen receptor (BCR) is essential not only for the development but also for the maintenance of mature B cells. Similarly, many B-cell lymphomas, including Burkitt lymphoma (BL), require continuous BCR signaling for their tumor growth. This growth is driven by immunoreceptor tyrosine-based activation motif (ITAM) and PI3 kinase (PI3K) signaling. Here, we employ CRISPR/Cas9 to delete BCR and B-cell co-receptor genes in the human BL cell line Ramos. We find that Ramos B cells require the expression of the BCR signaling component Igβ (CD79b), and the co-receptor CD19, for their fitness and competitive growth in culture. Furthermore, we show that in the absence of any other BCR component, Igβ can be expressed on the B-cell surface, where it is found in close proximity to CD19 and signals in an ITAM-dependent manner. These data suggest that Igβ and CD19 are part of an alternative B-cell signaling module that use continuous ITAM/PI3K signaling to promote the survival of B lymphoma and normal B cells. © 2018 The Authors. Published under the terms of the CC BY 4.0 license.

  6. Primordial germ cell differentiation of nuclear transfer embryonic stem cells using surface modified electroconductive scaffolds.

    Science.gov (United States)

    Eslami-Arshaghi, Tarlan; Vakilian, Saeid; Seyedjafari, Ehsan; Ardeshirylajimi, Abdolreza; Soleimani, Masoud; Salehi, Mohammad

    2017-04-01

    A combination of nanotopographical cues and surface modification of collagen and fibronectin is a potential platform in primordial germ cells (PGCs) differentiation. In the present study, the synergistic effect of nanotopography and surface modification on differentiation of nuclear transfer embryonic stem cells (nt-ESCs) toward PGC lineage was investigated. In order to achieve this goal, poly-anyline (PANi) was mix within poly-L-lactic acid (PLLA). Afterward, the random composite mats were fabricated using PLLA and PANi mix solution. The nanofiber topography notably upregulated the expressions of prdm14, mvh and c-kit compared with tissue culture polystyrene (TCP). Moreover, the combination of nanofiber topography and surface modification resulted in more enhancement of PGCs differentiation compared with non-modified nanofibrous scaffold. Additionally, gene expression results showed that mvh and c-kit were expressed at higher intensity in cells exposed to collagen and fibronectin rather than collagen or fibronectin solitary. These results demonstrated the importance of combined effect of collagen and fibronectin in order to develop a functional extracellular matrix (ECM) mimic in directing stem cell fate and the potential of such biofunctional scaffolds for treatment of infertility.

  7. Impaired cell surface expression of HLA-B antigens on mesenchymal stem cells and muscle cell progenitors

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  8. Flow cytometry detection of planktonic cells with polycyclic aromatic hydrocarbons sorbed to cell surfaces

    KAUST Repository

    Cerezo, Maria I.

    2017-02-17

    Polycyclic aromatic hydrocarbons are very important components of oil pollution. These pollutants tend to sorb to cell surfaces, exerting toxic effects on organisms. Our study developed a flow cytometric method for the detection of PAHs sorbed to phytoplankton by exploiting their spectral characteristics. We discriminated between cells with PAHs from cells free of PAHs. Clear discrimination was observed with flow cytometer provided with 375 or 405nm lasers in addition to the standard 488nm laser necessary to identify phytoplankton. Using this method, we measured the relationship between the percentages of phytoplankton organisms with PAHs, with the decrease in the growth rate. Moreover, the development of this method could be extended to facilitate the study of PAHs impact on cell cultures from a large variety of organisms.

  9. Surface topography during neural stem cell differentiation regulates cell migration and cell morphology.

    Science.gov (United States)

    Czeisler, Catherine; Short, Aaron; Nelson, Tyler; Gygli, Patrick; Ortiz, Cristina; Catacutan, Fay Patsy; Stocker, Ben; Cronin, James; Lannutti, John; Winter, Jessica; Otero, José Javier

    2016-12-01

    We sought to determine the contribution of scaffold topography to the migration and morphology of neural stem cells by mimicking anatomical features of scaffolds found in vivo. We mimicked two types of central nervous system scaffolds encountered by neural stem cells during development in vitro by constructing different diameter electrospun polycaprolactone (PCL) fiber mats, a substrate that we have shown to be topographically similar to brain scaffolds. We compared the effects of large fibers (made to mimic blood vessel topography) with those of small-diameter fibers (made to mimic radial glial process topography) on the migration and differentiation of neural stem cells. Neural stem cells showed differential migratory and morphological reactions with laminin in different topographical contexts. We demonstrate, for the first time, that neural stem cell biological responses to laminin are dependent on topographical context. Large-fiber topography without laminin prevented cell migration, which was partially reversed by treatment with rock inhibitor. Cell morphology complexity assayed by fractal dimension was inhibited in nocodazole- and cytochalasin-D-treated neural precursor cells in large-fiber topography, but was not changed in small-fiber topography with these inhibitors. These data indicate that cell morphology has different requirements on cytoskeletal proteins dependent on the topographical environment encountered by the cell. We propose that the physical structure of distinct scaffolds induces unique signaling cascades that regulate migration and morphology in embryonic neural precursor cells. J. Comp. Neurol. 524:3485-3502, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  10. Involvement of cell surface TG2 in the aggregation of K562 cells triggered by gluten.

    Science.gov (United States)

    Feriotto, G; Calza, R; Bergamini, C M; Griffin, M; Wang, Z; Beninati, S; Ferretti, V; Marzola, E; Guerrini, R; Pagnoni, A; Cavazzini, A; Casciano, F; Mischiati, C

    2017-03-01

    Gluten-induced aggregation of K562 cells represents an in vitro model reproducing the early steps occurring in the small bowel of celiac patients exposed to gliadin. Despite the clear involvement of TG2 in the activation of the antigen-presenting cells, it is not yet clear in which compartment it occurs. Herein we study the calcium-dependent aggregation of these cells, using either cell-permeable or cell-impermeable TG2 inhibitors. Gluten induces efficient aggregation when calcium is absent in the extracellular environment, while TG2 inhibitors do not restore the full aggregating potential of gluten in the presence of calcium. These findings suggest that TG2 activity is not essential in the cellular aggregation mechanism. We demonstrate that gluten contacts the cells and provokes their aggregation through a mechanism involving the A-gliadin peptide 31-43. This peptide also activates the cell surface associated extracellular TG2 in the absence of calcium. Using a bioinformatics approach, we identify the possible docking sites of this peptide on the open and closed TG2 structures. Peptide docks with the closed TG2 structure near to the GTP/GDP site, by establishing molecular interactions with the same amino acids involved in stabilization of GTP binding. We suggest that it may occur through the displacement of GTP, switching the TG2 structure from the closed to the active open conformation. Furthermore, docking analysis shows peptide binding with the β-sandwich domain of the closed TG2 structure, suggesting that this region could be responsible for the different aggregating effects of gluten shown in the presence or absence of calcium. We deduce from these data a possible mechanism of action by which gluten makes contact with the cell surface, which could have possible implications in the celiac disease onset.

  11. Cell surface alteration in Epstein-Barr virus-transformed cells from patients with extreme insulin resistance

    International Nuclear Information System (INIS)

    Gorden, D.L.; Robert, A.; Moncada, V.Y.; Taylor, S.I.; Muehlhauser, J.C.; Carpentier, J.L.

    1990-01-01

    An abnormality was detected in the morphology of the cell surface of Epstein-Barr virus-transformed lymphocytes of patients with genetic forms of insulin resistance. In cells from two patients with leprechaunism and two patients with type A extreme insulin resistance, scanning electron microscopy demonstrated a decrease in the percentage of the cell surface occupied by microvilli in cells from the patients with leprechaunism and type A insulin resistance compared with control cells. When cells from a healthy control subject and one of the patients with leprechaunism (Lep/Ark-1) were incubated with 125 I-labeled insulin, there was a decrease in the percentage of 125 I-insulin associated with microvilli on the cell surface. Thus, the decreased localization of insulin receptors with the microvillous region of the cell surface was in proportion to the decrease in microvilli

  12. Cell adhesion on Ti surface with controlled roughness

    Directory of Open Access Journals (Sweden)

    Burgos-Asperilla, Laura

    2015-06-01

    Full Text Available In this report, the in situ interaction between Saos-2 osteoblast cells and a smooth Ti surface was examined over time. The adhesion kinetics and mechanisms of cellular proliferation were monitored by quartz crystal microbalance (QCM and electrochemical impedance spectroscopy (EIS. The rate of Saos-2 attachment on Ti surfaces, obtained from the measurements performed with the QCM, is a first-order reaction, with k=2.10−3 min−1. The impedance measurements indicate that in the absence of cells, the Ti resistance diminishes over time (7 days, due to the presence of amino acids and proteins from the culture medium that have been a dsorbed, while in the presence of osteoblasts, this decrease is much greater because of the compounds generated by the cells that accelerate the dissolution of Ti.En este trabajo, se ha estudiado la interacción in situ entre células osteoblásticas Saos-2 y una superficie de Ti de rugosidad controlada a lo largo del tiempo. El estudio de la cinética y los mecanismos de proliferación celular de adhesión se ha realizado a través de la microbalanza de cristal de cuarzo (QCM y espectroscopía de impedancia electroquímica (EIS. La velocidad de adhesión de los osteoblastos sobre la superficie de Ti obtenida a través de medidas con la QCM, sigue una reacción de primer orden, con k=2×10−3 min−1. Los ensayos de impedancia indican que, en ausencia de las células, la resistencia del Ti disminuye con el tiempo (7 días, debido a la presencia de aminoácidos y proteínas del medio de cultivo que se han adsorbido, mientras que en presencia de células, esta disminución es mucho mayor debido a los productos metabólicos generados por las células que aceleran la disolución del Ti.

  13. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Borodina, Irina; Jensen, B. M.; Søndergaard, Ib

    2010-01-01

    Background: Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast...

  14. The role of cell walls and pectins in cation exchange and surface area of plant roots.

    Science.gov (United States)

    Szatanik-Kloc, A; Szerement, J; Józefaciuk, G

    2017-08-01

    We aimed to assess role of cell walls in formation of cation exchange capacity, surface charge, surface acidity, specific surface, water adsorption energy and surface charge density of plant roots, and to find the input of the cell wall pectins to the above properties. Whole roots, isolated cell walls and the residue after the extraction of pectins from the cell walls of two Apiaceae L. species (celeriac and parsnip) were studied using potentiometric titration curves and water vapor adsorption - desorption isotherms. Total amount of surface charge, as well as the cation exchange capacity were markedly higher in roots than in their cell walls, suggesting large contribution of other cell organelles to the binding of cations by the whole root cells. Significantly lower charge of the residues after removal of pectins was noted indicating that pectins play the most important role in surface charge formation of cell walls. The specific surface was similar for all of the studied materials. For the separated cell walls it was around 10% smaller than of the whole roots, and it increased slightly after the removal of pectins. The surface charge density and water vapor adsorption energy were the highest for the whole roots and the lowest for the cell walls residues after removal of pectins. The results indicate that the cell walls and plasma membranes are jointly involved in root ion exchange and surface characteristics and their contribution depends upon the plant species. Copyright © 2017 Elsevier GmbH. All rights reserved.

  15. Characterization and Comparative Analysis of Olfactory Receptor Co-Receptor Orco Orthologs Among Five Mirid Bug Species

    Directory of Open Access Journals (Sweden)

    Qi Wang

    2018-03-01

    Full Text Available The phytophagous mirid bugs of Apolygus lucorum, Lygus pratensis as well as three Adelphocoris spp., including Adelphocoris lineolatus, A. suturalis, and A. fasciaticollis are major pests of multiple agricultural crops in China, which have distinct geographical distribution and occurrence ranges. Like many insect species, these bugs heavily rely on olfactory cues to search preferred host plants, thereby investigation on functional co-evolution and divergence of olfactory genes seems to be necessary and is of great interest. In the odorant detection pathway, olfactory receptor co-receptor (Orco plays critical role in the perception of odors. In this study, we identified the full-length cDNA sequences encoding three putative Orcos (AsutOrco, AfasOrco, and LpraOrco in bug species of A. suturalis, A. fasciaticollis, and L. pratensis based on homology cloning method. Next, sequence alignment, membrane topology and gene structure analysis showed that these three Orco orthologs together with previously reported AlinOrco and AlucOrco shared high amino acid identities and similar topology structure, but had different gene structure especially at the length and insertion sites of introns. Furthermore, the evolutional estimation on the ratios of non-synonymous to synonymous (Ka/Ks revealed that Orco genes were under strong purifying selection, but the degrees of variation were significant different between genera. The results of quantitative real-time PCR experiments showed that these five Orco genes had a similar antennae-biased tissue expression pattern. Taking these data together, it is thought that Orco genes in the mirid species could share conserved olfaction roles but had different evolution rates. These findings would lay a foundation to further investigate the molecular mechanisms of evolutionary interactions between mirid bugs and their host plants, which might in turn contribute to the development of pest management strategy for mirid bugs.

  16. Mapping cellular hierarchy by single-cell analysis of the cell surface repertoire.

    Science.gov (United States)

    Guo, Guoji; Luc, Sidinh; Marco, Eugenio; Lin, Ta-Wei; Peng, Cong; Kerenyi, Marc A; Beyaz, Semir; Kim, Woojin; Xu, Jian; Das, Partha Pratim; Neff, Tobias; Zou, Keyong; Yuan, Guo-Cheng; Orkin, Stuart H

    2013-10-03

    Stem cell differentiation pathways are most often studied at the population level, whereas critical decisions are executed at the level of single cells. We have established a highly multiplexed, quantitative PCR assay to profile in an unbiased manner a panel of all commonly used cell surface markers (280 genes) from individual cells. With this method, we analyzed over 1,500 single cells throughout the mouse hematopoietic system and illustrate its utility for revealing important biological insights. The comprehensive single cell data set permits mapping of the mouse hematopoietic stem cell differentiation hierarchy by computational lineage progression analysis. Further profiling of 180 intracellular regulators enabled construction of a genetic network to assign the earliest differentiation event during hematopoietic lineage specification. Analysis of acute myeloid leukemia elicited by MLL-AF9 uncovered a distinct cellular hierarchy containing two independent self-renewing lineages with different clonal activities. The strategy has broad applicability in other cellular systems. Copyright © 2013 Elsevier Inc. All rights reserved.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2008-01-01

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

  18. Poly(vinyl alcohol)-micropatterned surfaces for manipulation of mesenchymal stem cell functions.

    Science.gov (United States)

    Chen, Guoping

    2014-01-01

    Micropatterning is a useful method to study the effects of biological and physical cues on cell functions. Various micropatterning methods have been developed for investigation of cell-cell interaction and cell-material interaction. As one of the potent methods, poly(vinyl alcohol) (PVA)-based micropatterning has been used to array cells in a pre-designed manner for a long-term cell culture. Cell population and single cell arrays can be formed in the micropatterned surfaces. The micropatterned surfaces have used to generate a gradient cell density, different degree of cell spreading, protrusion and cell-cell interaction and different geometry to investigate their effects on the differentiation of bone marrow-derived mesenchymal stem cells. This chapter highlights the latest development of PVA-based micropatterning and its application for manipulation of stem cell functions. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Quantitative comparison of a human cancer cell surface proteome between interphase and mitosis.

    Science.gov (United States)

    Özlü, Nurhan; Qureshi, Mohammad H; Toyoda, Yusuke; Renard, Bernhard Y; Mollaoglu, Gürkan; Özkan, Nazlı E; Bulbul, Selda; Poser, Ina; Timm, Wiebke; Hyman, Anthony A; Mitchison, Timothy J; Steen, Judith A

    2015-01-13

    The cell surface is the cellular compartment responsible for communication with the environment. The interior of mammalian cells undergoes dramatic reorganization when cells enter mitosis. These changes are triggered by activation of the CDK1 kinase and have been studied extensively. In contrast, very little is known of the cell surface changes during cell division. We undertook a quantitative proteomic comparison of cell surface-exposed proteins in human cancer cells that were tightly synchronized in mitosis or interphase. Six hundred and twenty-eight surface and surface-associated proteins in HeLa cells were identified; of these, 27 were significantly enriched at the cell surface in mitosis and 37 in interphase. Using imaging techniques, we confirmed the mitosis-selective cell surface localization of protocadherin PCDH7, a member of a family with anti-adhesive roles in embryos. We show that PCDH7 is required for development of full mitotic rounding pressure at the onset of mitosis. Our analysis provided basic information on how cell cycle progression affects the cell surface. It also provides potential pharmacodynamic biomarkers for anti-mitotic cancer chemotherapy. © 2014 The Authors.

  20. Increased cell surface metallopeptidase activity in cells undergoing UV-induced apoptosis

    International Nuclear Information System (INIS)

    Piva, T.J.; Davern, C.M.; Ellem, K.A.O.

    1999-01-01

    Full text: We have previously shown that UVC irradiation activated a range of cell surface peptidases (CSP) in HeLa cell monolayer cultures 20 h post-irradiation (1). In cells undergoing apoptosis there is an increase in CSP activity compared to control viable cells in cultures which have been treated by a wide range of agents including UV-irradiation (2). In order to further understand the mechanism involved in this process, we induced apoptosis in HeLa cells using 500 Jm -2 UVB. The separation of viable, apoptotic and necrotic cells of irradiated HeLa cell cultures was made by FACS analysis and sorting. The three populations were distinguished by their staining with PI and Hoechst 33342 dyes. CSP activity was measured using the P9 assay developed in this laboratory (1-3). The viable fraction of the irradiated cells had a higher level of CSP activity compared to unirradiated controls. The level of CSP activity in the apoptotic fraction was higher than that of the viable fraction, however that of the necrotic fraction was significantly lower. This finding agreed with that seen in UVC-irradiated (50 Jm -2 ) cultures (2). In order to elucidate the mechanism by which CSP activity was increased in UVB-irradiated cells undergoing apoptosis, the cultures were treated with the following agents: bestatin, aminopeptidase inhibitor, DEVD, caspase 3 inhibitor, and 3-aminobenzamide (3AB), PARP activation inhibitor. Bestatin and DEVD did not affect the level of CSP activity in the different cell subpopulations following UVB-irradiation. Treatment with 3AB abolished the increased CSP activity seen in the viable and apoptotic fraction following UVB-irradiation. All treated cells had the same morphology as observed under EM. The degree of phosphatidylserine eversion on the cell membrane was similar as were the cleavage profiles of PARP and actin. Only DEVD-treated cells had reduced caspase 3 activity which confirmed that the activation of CSP activity in apoptotic cells is

  1. The effect of cerium valence states at cerium oxide nanoparticle surfaces on cell proliferation

    KAUST Repository

    Naganuma, Tamaki

    2014-05-01

    Understanding and controlling cell proliferation on biomaterial surfaces is critical for scaffold/artificial-niche design in tissue engineering. The mechanism by which underlying integrin ligates with functionalized biomaterials to induce cell proliferation is still not completely understood. In this study, poly-l-lactide (PL) scaffold surfaces were functionalized using layers of cerium oxide nanoparticles (CNPs), which have recently attracted attention for use in therapeutic application due to their catalytic ability of Ce4+ and Ce3+ sites. To isolate the influence of Ce valance states of CNPs on cell proliferation, human mesenchymal stem cells (hMSCs) and osteoblast-like cells (MG63) were cultured on the PL/CNP surfaces with dominant Ce4+ and Ce3+ regions. Despite cell type (hMSCs and MG63 cells), different surface features of Ce4+ and Ce3+ regions clearly promoted and inhibited cell spreading, migration and adhesion behavior, resulting in rapid and slow cell proliferation, respectively. Cell proliferation results of various modified CNPs with different surface charge and hydrophobicity/hydrophilicity, indicate that Ce valence states closely correlated with the specific cell morphologies and cell-material interactions that trigger cell proliferation. This finding suggests that the cell-material interactions, which influence cell proliferation, may be controlled by introduction of metal elements with different valence states onto the biomaterial surface. © 2014 Elsevier Ltd.

  2. Cell-surface proteoglycan in sea urchin primary mesenchyme cell migration

    Energy Technology Data Exchange (ETDEWEB)

    Lane, M.C.

    1989-01-01

    Early in the development of the sea urchin embryo, the primary mesenchyme cells (PMC) migrate along the basal lamina of the blastocoel. Migration is inhibited in L. pictus embryos cultured in sulfate-free seawater and in S. purpuratus embryos exposed to exogenous {beta}-D-xylosides. An in vitro assay was developed to test the migratory capacity of normal PMC on normal and treated blastocoelic matrix. Sulfate deprivation and exposure to exogenous xyloside render PMC nonmotile on either matrix. Materials removed from the surface of normal PMC by treatment with 1 M urea restored migratory ability to defective cells, whereas a similar preparation isolated from the surface of epithelial cells at the same stage did not. Migration also resumed when cells were removed from the xyloside or returned to normal seawater. The urea extract was partially purified and characterized by radiolabeling, gel electrophoresis, fluorography, ion exchange chromatography, and western blotting. The PMC synthesize a large chondroitin sulfate/dermatan sulfate proteoglycan that is present in an active fraction isolated by chromatography. Chondroitinase ABC digestion of live cells blocked migration reversibly, further supporting the identification of the chondroitin sulfate/dermatan sulfate proteoglycan as the active component in the urea extract. Much of the incorporated sulfate was distributed along the filopodia in {sup 35}SO{sub 4}-labelled PMC by autoradiography. The morphology of normal and treated S. purpuratus PMC was examined by scanning electron microscopy, and differences in spreading, particularly of the extensive filopodia present on the cells, was observed. A model for the role of the chondroitin sulfate/dermatan sulfate proteoglycan in cell detachment during migration is proposed.

  3. Microarray of neuroblastoma cells on the selectively functionalized nanocrystalline diamond thin film surface

    Energy Technology Data Exchange (ETDEWEB)

    Park, Young-Sang; Son, Hyeong-Guk; Kim, Dae-Hoon; Oh, Hong-Gi; Lee, Da-Som; Kim, Min-Hye; Lim, Ki-Moo; Song, Kwang-Soup, E-mail: kssong10@kumoh.ac.kr

    2016-01-15

    Graphical abstract: - Highlights: • The nanocrystalline diamond (NCD) surface is functionalized with F or O. • The cell adhesion and growth are evaluated on the functionalized NCD surface. • The cell adhesion and growth depend on the wettability of the surface. • Cell patterning was achieved by using of hydrophilic and hydrophobic surfaces. • Neuroblastoma cells were arrayed on the micro-patterned NCD surface. - Abstract: Nanocrystalline diamond (NCD) film surfaces were modified with fluorine or oxygen by plasma treatment in an O{sub 2} or C{sub 3}F{sub 8} gas environment in order to induce wettability. The oxygenated-NCD (O-NCD) film surface was hydrophilic and the fluorinated-NCD (F-NCD) surface was hydrophobic. The efficiency of early cell adhesion, which is dependent on the wettability of the cell culture plate and necessary for the growth and proliferation of cells, was 89.62 ± 3.92% on the O-NCD film and 7.78 ± 0.77% on the F-NCD film surface after 3 h of cell culture. The wettability of the NCD film surface was artificially modified using a metal mask and plasma treatment to fabricate a micro-pattern. Four types of micro-patterns were fabricated (line, circle, mesh, and word) on the NCD film surface. We precisely arrayed the neuroblastoma cells on the micro-patterned NCD film surfaces by controlling the surface wettability and cell seeding density. The neuroblastoma cells adhered and proliferated along the O-NCD film surface.

  4. Cell surface estrogen receptor alpha is upregulated during subchronic metabolic stress and inhibits neuronal cell degeneration.

    Directory of Open Access Journals (Sweden)

    Cristiana Barbati

    Full Text Available In addition to the classical nuclear estrogen receptor, the expression of non-nuclear estrogen receptors localized to the cell surface membrane (mER has recently been demonstrated. Estrogen and its receptors have been implicated in the development or progression of numerous neurodegenerative disorders. Furthermore, the pathogenesis of these diseases has been associated with disturbances of two key cellular programs: apoptosis and autophagy. An excess of apoptosis or a defect in autophagy has been implicated in neurodegeneration. The aim of this study was to clarify the role of ER in determining neuronal cell fate and the possible implication of these receptors in regulating either apoptosis or autophagy. The human neuronal cell line SH-SY5Y and mouse neuronal cells in primary culture were thus exposed to chronic minimal peroxide treatment (CMP, a form of subcytotoxic minimal chronic stress previously that mimics multiple aspects of long-term cell stress and represents a limited molecular proxy for neurodegenerative processes. We actually found that either E2 or E2-bovine serum albumin construct (E2BSA, i.e. a non-permeant form of E2 was capable of modulating intracellular cell signals and regulating cell survival and death. In particular, under CMP, the up-regulation of mERα, but not mERβ, was associated with functional signals (ERK phosphorylation and p38 dephosphorylation compatible with autophagic cytoprotection triggering and leading to cell survival. The mERα trafficking appeared to be independent of the microfilament system cytoskeletal network but was seemingly associated with microtubular apparatus network, i.e., to MAP2 molecular chaperone. Importantly, antioxidant treatments, administration of siRNA to ERα, or the presence of antagonist of ERα hindered these events. These results support that the surface expression of mERα plays a pivotal role in determining cell fate, and that ligand-induced activation of mER signalling exerts a

  5. Nanoscale surfaces for the long-term maintenance of mesenchymal stem cell phenotype and multipotency

    Science.gov (United States)

    McMurray, Rebecca J.; Gadegaard, Nikolaj; Tsimbouri, P. Monica; Burgess, Karl V.; McNamara, Laura E.; Tare, Rahul; Murawski, Kate; Kingham, Emmajayne; Oreffo, Richard O. C.; Dalby, Matthew J.

    2011-08-01

    There is currently an unmet need for the supply of autologous, patient-specific stem cells for regenerative therapies in the clinic. Mesenchymal stem cell differentiation can be driven by the material/cell interface suggesting a unique strategy to manipulate stem cells in the absence of complex soluble chemistries or cellular reprogramming. However, so far the derivation and identification of surfaces that allow retention of multipotency of this key regenerative cell type have remained elusive. Adult stem cells spontaneously differentiate in culture, resulting in a rapid diminution of the multipotent cell population and their regenerative capacity. Here we identify a nanostructured surface that retains stem-cell phenotype and maintains stem-cell growth over eight weeks. Furthermore, the study implicates a role for small RNAs in repressing key cell signalling and metabolomic pathways, demonstrating the potential of surfaces as non-invasive tools with which to address the stem cell niche.

  6. Tuning cell adhesion on polymeric and nanocomposite surfaces: Role of topography versus superhydrophobicity

    Energy Technology Data Exchange (ETDEWEB)

    Zangi, Sepideh [Department of Chemical Engineering, Shahrood Branch, Islamic Azad University, P.O. Box 36155-163, Shahrood (Iran, Islamic Republic of); Hejazi, Iman [Department of Polymer Engineering & Color Technology, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Seyfi, Javad, E-mail: Jseyfi@gmail.com [Department of Chemical Engineering, Shahrood Branch, Islamic Azad University, P.O. Box 36155-163, Shahrood (Iran, Islamic Republic of); Hejazi, Ehsan [Department of Clinical Nutrition and Dietetics, Faculty of Nutrition Sciences and Food Technology, Shahid Beheshti University of Medical Sciences, Tehran (Iran, Islamic Republic of); Khonakdar, Hossein Ali [Department of Polymer Engineering, Faculty of Engineering, South Tehran Branch, Islamic Azad University, P.O. Box 19585-466, Tehran (Iran, Islamic Republic of); Davachi, Seyed Mohammad [School of Chemical Engineering, University of Tehran, P.O. Box 11155-4563, Tehran (Iran, Islamic Republic of)

    2016-06-01

    Development of surface modification procedures which allow tuning the cell adhesion on the surface of biomaterials and devices is of great importance. In this study, the effects of different topographies and wettabilities on cell adhesion behavior of polymeric surfaces are investigated. To this end, an improved phase separation method was proposed to impart various wettabilities (hydrophobic and superhydrophobic) on polypropylene surfaces. Surface morphologies and compositions were characterized by scanning electron microscopy and X-ray photoelectron spectroscopy, respectively. Cell culture was conducted to evaluate the adhesion of 4T1 mouse mammary tumor cells. It was found that processing conditions such as drying temperature is highly influential in cell adhesion behavior due to the formation of an utterly different surface topography. It was concluded that surface topography plays a more significant role in cell adhesion behavior rather than superhydrophobicity since the nano-scale topography highly inhibited the cell adhesion as compared to the micro-scale topography. Such cell repellent behavior could be very useful in many biomedical devices such as those in drug delivery and blood contacting applications as well as biosensors. - Highlights: • A novel method is presented for fabrication of superhydrophobic surfaces. • The presence of nanoparticles in non-solvent bath notably promoted phase separation. • Topography had a more notable impact on cell adhesion than superhydrophobicity. • Nano-scale topographical features highly impeded cell adhesion on polymer surfaces.

  7. An efficient delivery of DAMPs on the cell surface by the unconventional secretion pathway

    International Nuclear Information System (INIS)

    Zhu, Haiyan; Wang, Lan; Ruan, Yuanyuan; Zhou, Lei; Zhang, Dongmei; Min, Zhihui; Xie, Jianhui; Yu, Min; Gu, Jianxin

    2011-01-01

    Research highlights: → Hsp60 transported to cell surface through the classical secretory pathway was modified with N-glycosylation. → HSAPB-N18 could efficiently deliver Hsp60 to the cell surface via the unconventional secretory pathway. → Cell surface Hsp60 delivered by HASPB-N18 has a proper conformation. → HASPB-N18 is an efficient delivery signal for other DAMP molecules such as Hsp70 and HMGB1. -- Abstract: Damage-associated molecular patterns (DAMPs) are signals released from dying cells evoking the immune system response in several inflammatory disorders. In normal situations, many of DAMPs are nuclear or cytosolic proteins with defined intracellular function, but they could be found on the cell surface following tissue injury. The biological function of the translocated DAMPs is still not well known and an efficient delivery of these molecules on the cell surface is required to clarify their biological effects. In this study, we demonstrated that an unclassical secretory signal peptide, N-terminal 18 amino acids of HASPB (HASPB-N18), could efficiently deliver Hsp60, Hsp70, and HMGB1 on the cell surface. Furthermore, the delivery of these molecules on the cell surface by HASPB-N18 is not limited to a special cell line because several cell lines could use this delivery signal to deliver these molecules on the cell surface. Moreover, we demonstrated that Hsp60 on the cell surface delivered by HASPB-N18 could be recognized by a soluble form of LOX-1, which implies that DAMPs on the cell surface delivered by HASPB-N18 have a proper conformation during transport. Therefore, delivery of DAMPs by HASPB-N18 is a reliable model to further understand the biological significance of DAMPs on the cell surface.

  8. LapF and Its Regulation by Fis Affect the Cell Surface Hydrophobicity of Pseudomonas putida.

    Science.gov (United States)

    Lahesaare, Andrio; Ainelo, Hanna; Teppo, Annika; Kivisaar, Maia; Heipieper, Hermann J; Teras, Riho

    2016-01-01

    The ability of bacteria to regulate cell surface hydrophobicity is important for the adaptation to different environmental conditions. The hydrophobicity of cell surface can be determined by several factors, including outer membrane and surface proteins. In this study, we report that an adhesin LapF influences cell surface hydrophobicity of Pseudomonas putida. Cells lacking LapF are less hydrophobic than wild-type cells in stationary growth phase. Moreover, the overexpression of the global regulator Fis decreases surface hydrophobicity by repressing the expression of lapF. Flow cytometry analysis revealed that bacteria producing LapF are more viable when confronted with methanol (a hydrophilic compound) but are more susceptible to 1-octanol (a hydrophobic compound). Thus, these results revealed that LapF is the hydrophobicity factor for the cell surface of P. putida.

  9. Adherence of Helicobacter pylori cells and their surface components to HeLa cell membranes.

    Science.gov (United States)

    Fauchère, J L; Blaser, M J

    1990-12-01

    Four Helicobacter pylori strains were used to develop in vitro methods to assess adherence to HeLa cells. Using direct detection by microscopy, adhesion scores increased with the initial bacteria-to-cell ratio. The urease method assessed H. pylori bound to HeLa cells by their urease activity. The percentage of the original inoculum adhering to HeLa cells remained constant for initial ratios from 10(2) to 10(5) bacteria per cell. An ELISA using anti-H. pylori serum assessed whole bacteria or components bound to HeLa cell fractions. By all three methods, the four H. pylori strains were adherent to HeLa cells or membranes whereas Campylobacter fetus and Providencia control strains were not. The adherence of H. pylori whole cells decreased following extraction with saline, water, or glycine buffer and most of the superficial adhering material (SAM) was present in the saline or water extracts. SAM bound better to HeLa membranes than to calf fetuin or bovine serum albumin (BSA); binding was inhibited by preincubation of SAM with HeLa membranes but not with fetuin or BSA or by pretreatment of HeLa membranes with neuraminidase. These data indicate that SAM has a specific receptor on the HeLa cell membranes. By gel exclusion chromatography of bacterial extracts, the most adherent components were found in the fractions which also contained the highest urease activity; these fractions included urease subunit antigens. We conclude that adherence of H. pylori can be assessed by microtiter assays and involves bacterial surface material which co-purifies with urease and is different from the N-acetyl-neuraminyl-lactose binding hemagglutinin.

  10. Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration

    Science.gov (United States)

    Wu, Tao; Kooi, Craig Vander; Shah, Pritom; Charnigo, Richard; Huang, Cai; Smyth, Susan S.; Morris, Andrew J.

    2014-01-01

    Autotaxin (ATX) is a secreted lysophospholipase D (lysoPLD) that binds to integrin adhesion receptors. We dissected the roles of integrin binding and lysoPLD activity in stimulation of human breast cancer and mouse aortic vascular smooth muscle cell migration by ATX. We compared effects of wild-type human ATX, catalytically inactive ATX, an integrin binding-defective ATX variant with wild-type lysoPLD activity, the isolated ATX integrin binding N-terminal domain, and a potent ATX selective lysoPLD inhibitor on cell migration using transwell and single-cell tracking assays. Stimulation of transwell migration was reduced (18 or 27% of control, respectively) but not ablated by inactivation of integrin binding or inhibition of lysoPLD activity. The N-terminal domain increased transwell migration (30% of control). ATX lysoPLD activity and integrin binding were necessary for a 3.8-fold increase in the fraction of migrating breast cancer cell step velocities >0.7 μm/min. ATX increased the persistent directionality of single-cell migration 2-fold. This effect was lysoPLD activity independent and recapitulated by the integrin binding N-terminal domain. Integrin binding enables uptake and intracellular sequestration of ATX, which redistributes to the front of migrating cells. ATX binding to integrins and lysoPLD activity therefore cooperate to promote rapid persistent directional cell migration.—Wu, T., Kooi, C. V., Shah, P., Charnigo, R., Huang, C., Smyth, S. S., Morris, A. J. Integrin-mediated cell surface recruitment of autotaxin promotes persistent directional cell migration. PMID:24277575

  11. Proteomic inventory of "anchorless" proteins on the colon adenocarcinoma cell surface.

    NARCIS (Netherlands)

    Tjalsma, H.; Pluk, W.J.G.; Heuvel, L.P.W.J. van den; Peters, W.H.M.; Roelofs, R.H.W.M.; Swinkels, D.W.

    2006-01-01

    Surface proteins play important pathophysiological roles in health and disease, and accumulating proteomics-based studies suggest that several "non-membrane" proteins are sorted to the cell surface by unconventional mechanisms. Importantly, these proteins may comprise attractive therapeutic targets

  12. 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.

  13. Cell-surface galactosyltransferase acts as a modulator of rat and human acinar cell proliferation.

    Science.gov (United States)

    Humphreys-Beher, M G; Zelles, T; Maeda, N; Purushotham, K R; Cassisi, N; Schneyer, C A

    1990-06-01

    Several physiological parameters were examined for inducing acinar cell proliferation and corresponding increased expression of beta 1-4 galactosyltransferase. In this study, dietary changes causing acinar cell proliferation included the following: the introduction of animals to a liquid diet (causing gland atrophy) followed by re-introduction of solid chow, gustatory stimulation provided by the introduction of 0.5% citric acid to animal drinking water, and removal of the submandibular gland with subsequent reliance on the parotid gland for saliva protein and fluid. Alterations in growth factor levels were produced by injecting animals with a chronic (three-day) regimen of either nerve growth factor (NGF) or epidermal growth factor (EGF). In all cases of acinar cell proliferation in vivo, generated by the above treatments, cell-surface galactosyltransferase was detected along with the unique expression of a 4.5-kb proliferation-associated mRNA. Parotid gland proliferation could be blocked in all cases by the injection of the galactosyltransferase specific modifier protein, alpha-lactalbumin. Propranolol, a beta-adrenergic receptor antagonist, blocked proliferation in all cases except EGF treatment. EGF-induced proliferation could, however, be prevented if the animals were treated with monoclonal antibody to EGF receptor or with the galactosyltransferase modifier alpha-lactalbumin. As a comparison, human parotid tissue samples obtained from neoplastic pleomorphic adenomas, muco-epidermoid carcinoma, adenoid cystic carcinoma, and a bulimia patient were analyzed for galactosyltransferase expression by Northern blot of mRNA and plasma membrane isolation. Elevated levels of galactosyltransferase were found in all neoplastic tissue preparations as well as in the bulimia sample. Amylase synthesis was reduced in samples compared with surrounding normal tissue from the same patient. In vitro cell culturing of pleomorphic adenoma cells in the presence of

  14. Cell surface GRP78 facilitates hepatoma cells proliferation and migration by activating IGF-IR.

    Science.gov (United States)

    Yin, Yancun; Chen, Chen; Chen, Jinliang; Zhan, Renhui; Zhang, Qiang; Xu, Xiaoyan; Li, Defang; Li, Minjing

    2017-07-01

    The 78kDa glucose regulated protein (GRP78) is a multifunctional chaperone that is involved in a variety of cellular processes. Insulin like growth factor I receptor (IGF-IR) often aberrant expresses in many types of tumor cells. The IGF-IR signaling plays key roles in carcinogenesis and maintenance of the malignant phenotype. The crosstalk between GRP78 and IGF-IR molecules has not well been illuminated. Here, we demonstrated a reciprocal regulation of GRP78 expression and IGF-IR pathway activation. IGF-I induced GRP78 expression in hepatoma cells. IGF-IR knockdown or IGF-IR inhibitor repressed GRP78 expression. Both phosphatidylinositol 3-kianase (PI3K) and mitogen-activated protein kinase (MAPK) pathways involved in IGF-I induction of GRP78 expression. Interestingly, treatment of hepatoma cells with IGF-I re-distributes GRP78 from endoplasmic reticulum (ER) to cell surface and promotes its physical interaction with IGF-IR. Also, GRP78 promotes IGF-IR phosphorylation and activation. Blocked of GRP78 by small interfering RNA or inhibition of GRP78 function by (-)-epigallocatechin gallate (EGCG) blocks IGF-I induced IGF-IR phosphorylation and its downstream signaling. Further, blocked cell surface GRP78 with antibody inhibits IGF-I stimulated cellular proliferation and migration. These data reveal an essential role for the molecular chaperone GRP78 in IGF-IR signaling and implicate the use of GRP78 inhibitors in blocking IGF-IR signaling in hepatoma cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

    Jensen, Helle

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

  16. Enhanced cell disruption strategy in the release of recombinant hepatitis B surface antigen from Pichia pastoris using response surface methodology

    Directory of Open Access Journals (Sweden)

    Tam Yew

    2012-10-01

    Full Text Available Abstract Background Cell disruption strategies by high pressure homogenizer for the release of recombinant Hepatitis B surface antigen (HBsAg from Pichia pastoris expression cells were optimized using response surface methodology (RSM based on the central composite design (CCD. The factors studied include number of passes, biomass concentration and pulse pressure. Polynomial models were used to correlate the above mentioned factors to project the cell disruption capability and specific protein release of HBsAg from P. pastoris cells. Results The proposed cell disruption strategy consisted of a number of passes set at 20 times, biomass concentration of 7.70 g/L of dry cell weight (DCW and pulse pressure at 1,029 bar. The optimized cell disruption strategy was shown to increase cell disruption efficiency by 2-fold and 4-fold for specific protein release of HBsAg when compared to glass bead method yielding 75.68% cell disruption rate (CDR and HBsAg concentration of 29.20 mg/L respectively. Conclusions The model equation generated from RSM on cell disruption of P. pastoris was found adequate to determine the significant factors and its interactions among the process variables and the optimum conditions in releasing HBsAg when validated against a glass bead cell disruption method. The findings from the study can open up a promising strategy for better recovery of HBsAg recombinant protein during downstream processing.

  17. Enhanced cell disruption strategy in the release of recombinant hepatitis B surface antigen from Pichia pastoris using response surface methodology

    Science.gov (United States)

    2012-01-01

    Background Cell disruption strategies by high pressure homogenizer for the release of recombinant Hepatitis B surface antigen (HBsAg) from Pichia pastoris expression cells were optimized using response surface methodology (RSM) based on the central composite design (CCD). The factors studied include number of passes, biomass concentration and pulse pressure. Polynomial models were used to correlate the above mentioned factors to project the cell disruption capability and specific protein release of HBsAg from P. pastoris cells. Results The proposed cell disruption strategy consisted of a number of passes set at 20 times, biomass concentration of 7.70 g/L of dry cell weight (DCW) and pulse pressure at 1,029 bar. The optimized cell disruption strategy was shown to increase cell disruption efficiency by 2-fold and 4-fold for specific protein release of HBsAg when compared to glass bead method yielding 75.68% cell disruption rate (CDR) and HBsAg concentration of 29.20 mg/L respectively. Conclusions The model equation generated from RSM on cell disruption of P. pastoris was found adequate to determine the significant factors and its interactions among the process variables and the optimum conditions in releasing HBsAg when validated against a glass bead cell disruption method. The findings from the study can open up a promising strategy for better recovery of HBsAg recombinant protein during downstream processing. PMID:23039947

  18. Dictyostelium Cells Migrate Similarly on Surfaces of Varying Chemical Composition

    OpenAIRE

    McCann, Colin P.; Rericha, Erin C.; Wang, Chenlu; Losert, Wolfgang; Parent, Carole A.

    2014-01-01

    During cell migration, cell-substrate binding is required for pseudopod anchoring to move the cell forward, yet the interactions with the substrate must be sufficiently weak to allow parts of the cell to de-adhere in a controlled manner during typical protrusion/retraction cycles. Mammalian cells actively control cell-substrate binding and respond to extracellular conditions with localized integrin-containing focal adhesions mediating mechanotransduction. We asked whether mechanotransduction ...

  19. Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Makarova, Olga V. [Creatv MicroTech, Inc., 2242 West Harrison St., Chicago 60612, IL (United States); Adams, Daniel L., E-mail: dan@creatvmicrotech.com [Creatv MicroTech, Inc., 1 Deer Park Drive, Monmouth Junction, NJ 08852 (United States); Divan, Ralu; Rosenmann, Daniel [Center for Nanoscale Materials, Argonne National Laboratory, 9700 South Cass Ave., Argonne 60439, IL (United States); Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei [Creatv MicroTech, Inc., 11609 Lake Potomac Drive, Potomac 20854, MD (United States)

    2016-09-01

    There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. - Highlights: • Surface topographical effects the growth patterns and cell function of cancer cells • Nanoscale surface topography on polymer filters for circulating tumor cell culture • Membrane fabricated directly on polymer surfaces utilized for polymer etching • Nanotopography alters cell shape, phenotype and growth patterns of cancer cells • Nanoscale surface topography dictates monolayering or 3D structured cell culture.

  20. Hydrophobic fractal surface from glycerol tripalmitate and the effects on C6 glioma cell growth.

    Science.gov (United States)

    Zhang, Shanshan; Chen, Xuerui; Yu, Jing; Hong, Biyuan; Lei, Qunfang; Fang, Wenjun

    2016-06-01

    To provide a biomimic environment for glial cell culture, glycerol tripalmitate (PPP) has been used as a raw material to prepare fractal surfaces with different degrees of hydrophobicity. The spontaneous formation of the hydrophobic fractal surfaces was monitored by differential scanning calorimetry (DSC) and X-ray diffraction (XRD). The surface morphologies were observed by a scanning electron microscope (SEM), and then the fractal dimension (FD) values of the surfaces were determined with the box-counting method. C6 glioma cells were cultured and compared on different hydrophobic PPP surfaces and poly-L-lysine (PLL)-coated surface. The cell numbers as a function of incubation time on different surfaces during the cell proliferation process were measured, and the cell morphologies were observed under a fluorescence microscope. Influences of hydrophobic fractal surfaces on the cell number and morphology were analyzed. The experimental results show that the cell proliferation rates decrease while the cell morphology complexities increase with the growth of the fractal dimensions of the PPP surfaces. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Various domains of the B-cell regulatory molecule CD72 has diverged at different rates in mammals

    DEFF Research Database (Denmark)

    Petersen, Cathrine Bie; Hillig, Ann-Britt Nygaard; Fredholm, Merete

    2007-01-01

    We report the cloning of the porcine B-cell co-receptor CD72, as well as genomic mapping and examination of transcription. The B-cell receptor (BCR) complex mediates signalling upon antigen recognition by the membrane bound BCR. Several co-receptors modulate this signal positively or negatively. CD......72 has been shown to be a negatively regulating BCR co-receptor. We isolated and sequenced three porcine CD72 transcript variants. Using a pig radiation hybrid panel we found the porcien CD72 gene to be located on chromosome 1q21-28 in a region syntenic to human chromosome 9. The porcine CD72 gene...

  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. Tracking Traction Force Changes of Single Cells on the Liquid Crystal Surface

    Directory of Open Access Journals (Sweden)

    Chin Fhong Soon

    2015-01-01

    Full Text Available Cell migration is a key contributor to wound repair. This study presents findings indicating that the liquid crystal based cell traction force transducer (LCTFT system can be used in conjunction with a bespoke cell traction force mapping (CTFM software to monitor cell/surface traction forces from quiescent state in real time. In this study, time-lapse photo microscopy allowed cell induced deformations in liquid crystal coated substrates to be monitored and analyzed. The results indicated that the system could be used to monitor the generation of cell/surface forces in an initially quiescent cell, as it migrated over the culture substrate, via multiple points of contact between the cell and the surface. Future application of this system is the real-time assaying of the pharmacological effects of cytokines on the mechanics of cell migration.

  5. Regulation of ADAM12 cell-surface expression by protein kinase C epsilon

    DEFF Research Database (Denmark)

    Sundberg, Christina; Thodeti, Charles Kumar; Kveiborg, Marie

    2004-01-01

    The ADAM (a disintegrin and metalloprotease) family consists of multidomain cell-surface proteins that have a major impact on cell behavior. These transmembrane-anchored proteins are synthesized as proforms that have (from the N terminus): a prodomain; a metalloprotease-, disintegrin......-like-, cysteine-rich, epidermal growth factor-like, and transmembrane domain; and a cytoplasmic tail. The 90-kDa mature form of human ADAM12 is generated in the trans-Golgi through cleavage of the prodomain by a furin-peptidase and is stored intracellularly until translocation to the cell surface...... as a constitutively active protein. However, little is known about the regulation of ADAM12 cell-surface translocation. Here, we used human RD rhabdomyosarcoma cells, which express ADAM12 at the cell surface, in a temporal pattern. We report that protein kinase C (PKC) epsilon induces ADAM12 translocation to the cell...

  6. Enhancing the stability of xylanase from Cellulomonas fimi by cell-surface display on Escherichia coli.

    Science.gov (United States)

    Chen, Y-P; Hwang, I-E; Lin, C-J; Wang, H-J; Tseng, C-P

    2012-03-01

    The cell-surface display of Cex, which encodes xylanase and exoglucanase from Cellulomonas fimi, was constructed on Escherichia coli using PgsA as the anchor protein. Characterization of the cell-surface display of Cex was performed. PgsA was fused to the N-terminus of Cex and six histidines were utilized as spacers between the targeting and anchor proteins. Successful cell-surface display of Cex was demonstrated by Western blot and immunofluorescence analyses on E. coli C41 (DE3). According to the time-course analysis, the xylanase activity of Cex was achieved at 49Ug(-1) dry cell weight after 12 h culture at 37°C. The optimal temperature and pH ranges of the cell-surface displayed protein with whole-cell were broader than the corresponding ranges of the purified form. Further determination of thermostability indicated that the half-life of cell-surface displayed Cex was 1·6 times longer than that of purified Cex at 60°C. We have successfully developed the cell-surface display of xylanase on E. coli. The cell-surface display can enhance the stability of xylanase against changes in temperature and has the potential of becoming a whole-cell biocatalyst for industrial applications, such as biobleaching of paper and production of renewable energy. The results demonstrated that the cell-surface display of xylanase embedded in the cell membrane is more stable than that of the purified enzyme. Thus, to improve the stability of heterologous proteins production, cell-surface display using the PgsA anchor protein as a tool can be considered in E. coli. © 2012 The Authors. Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

  7. Characterization of rat basophilic leukemia cell surface proteins using monoclonal antibodies

    International Nuclear Information System (INIS)

    Buonocore-Buzzelli, L.M.

    1988-01-01

    Rat basophilic leukemia (RBL) cells express both immunoglobulin E (IgE) and immunoglobulin G (IgG) receptors. In this study, mouse monoclonal antibodies were produced against the RBL cell and screened for their ability to precipitate specific bands from 125 I surface labeled cells. Fourteen hybridomas were selected and divided into five groups since many of the hybridomas precipitated bands of identical molecular weight. One or more of the hybridomas from each group, and the cell surface antigens they identified, were further characterized. Binding of all the monoclonal antibodies to the RBL-2H3 cell surface was saturable and of high affinity. In cross inhibition studies, two of the antibodies were found to bind to identical or neighboring epitopes, presumably on the same cell surface molecule. Binding studies using other cell populations demonstrated that the monoclonal antibodies react not only with commonly expressed rat cell surface molecules but also with molecules specifically expressed on rat mast cells and basophils. None of the antibodies were found to induce or inhibit serotonin release from the RBL cells. Western blotting showed most of the antibodies to react with bands whose molecular weights resembled those seen by immuno-precipitation. Antibodies number sign 8 and number sign 12, although from the same group, were found to react with different subunits of the same cell surface protein. Sequential immunoprecipitation and peptide mapping confirmed that the antigens defined by these antibodies were structurally related

  8. Survey of surface proteins from the pathogenic Mycoplasma hyopneumoniae strain 7448 using a biotin cell surface labeling approach.

    Science.gov (United States)

    Reolon, Luciano Antonio; Martello, Carolina Lumertz; Schrank, Irene Silveira; Ferreira, Henrique Bunselmeyer

    2014-01-01

    The characterization of the repertoire of proteins exposed on the cell surface by Mycoplasma hyopneumoniae (M. hyopneumoniae), the etiological agent of enzootic pneumonia in pigs, is critical to understand physiological processes associated with bacterial infection capacity, survival and pathogenesis. Previous in silico studies predicted that about a third of the genes in the M. hyopneumoniae genome code for surface proteins, but so far, just a few of them have experimental confirmation of their expression and surface localization. In this work, M. hyopneumoniae surface proteins were labeled in intact cells with biotin, and affinity-captured biotin-labeled proteins were identified by a gel-based liquid chromatography-tandem mass spectrometry approach. A total of 20 gel slices were separately analyzed by mass spectrometry, resulting in 165 protein identifications corresponding to 59 different protein species. The identified surface exposed proteins better defined the set of M. hyopneumoniae proteins exposed to the host and added confidence to in silico predictions. Several proteins potentially related to pathogenesis, were identified, including known adhesins and also hypothetical proteins with adhesin-like topologies, consisting of a transmembrane helix and a large tail exposed at the cell surface. The results provided a better picture of the M. hyopneumoniae cell surface that will help in the understanding of processes important for bacterial pathogenesis. Considering the experimental demonstration of surface exposure, adhesion-like topology predictions and absence of orthologs in the closely related, non-pathogenic species Mycoplasma flocculare, several proteins could be proposed as potential targets for the development of drugs, vaccines and/or immunodiagnostic tests for enzootic pneumonia.

  9. Survey of surface proteins from the pathogenic Mycoplasma hyopneumoniae strain 7448 using a biotin cell surface labeling approach.

    Directory of Open Access Journals (Sweden)

    Luciano Antonio Reolon

    Full Text Available The characterization of the repertoire of proteins exposed on the cell surface by Mycoplasma hyopneumoniae (M. hyopneumoniae, the etiological agent of enzootic pneumonia in pigs, is critical to understand physiological processes associated with bacterial infection capacity, survival and pathogenesis. Previous in silico studies predicted that about a third of the genes in the M. hyopneumoniae genome code for surface proteins, but so far, just a few of them have experimental confirmation of their expression and surface localization. In this work, M. hyopneumoniae surface proteins were labeled in intact cells with biotin, and affinity-captured biotin-labeled proteins were identified by a gel-based liquid chromatography-tandem mass spectrometry approach. A total of 20 gel slices were separately analyzed by mass spectrometry, resulting in 165 protein identifications corresponding to 59 different protein species. The identified surface exposed proteins better defined the set of M. hyopneumoniae proteins exposed to the host and added confidence to in silico predictions. Several proteins potentially related to pathogenesis, were identified, including known adhesins and also hypothetical proteins with adhesin-like topologies, consisting of a transmembrane helix and a large tail exposed at the cell surface. The results provided a better picture of the M. hyopneumoniae cell surface that will help in the understanding of processes important for bacterial pathogenesis. Considering the experimental demonstration of surface exposure, adhesion-like topology predictions and absence of orthologs in the closely related, non-pathogenic species Mycoplasma flocculare, several proteins could be proposed as potential targets for the development of drugs, vaccines and/or immunodiagnostic tests for enzootic pneumonia.

  10. Cell behavior related to implant surfaces with different microstructure and chemical composition: an in vitro analysis.

    Science.gov (United States)

    Conserva, Enrico; Lanuti, Anna; Menini, Maria

    2010-01-01

    This paper reports on an in vitro comparison of osteoblast and mesenchymal stem cell (MSC) adhesion, proliferation, and differentiation related to two different surface treatments applied to the same implant design to determine whether the interaction between cells and implants is influenced by surface structure and chemical composition of the implants. Thirty-nine implants with a sandblasted (SB) surface and 39 implants with a grit-blasted and high-temperature acid-etched (GBAE) surface were used. The implant macrostructures and microstructures were analyzed by high- and low-voltage scanning electron microscopy (SEM) and by stereo-SEM. The surface chemical composition was investigated by energy dispersive analysis and x-ray photoemission spectroscopy. SaOS-2 osteoblasts and human MSCs were used for the evaluation of cell proliferation and alkaline phosphatase enzymatic activity in contact with the two surfaces. The GBAE surface showed fewer contaminants and a very high percentage of titanium (19.7%) compared to the SB surface (14.2%). The two surfaces showed similar mean roughness (Ra), but the depth (Rz) and density (RSm) of the porosity were significantly increased in the GBAE surface. The GBAE surface presented more osteoblast and MSC proliferation than the SB surface. No statistically significant differences in alkaline phosphatase activity were found between surfaces for either cellular line. The GBAE surface showed less surface contaminants and a higher percentage of titanium (19.7%) than the SB surface. The macro/micropore structured design and chemical composition of the GBAE surface allowed greater cell adhesion and proliferation and an earlier cell spreading but did not play an obvious role in in vitro cellular differentiation.

  11. Microarray of neuroblastoma cells on the selectively functionalized nanocrystalline diamond thin film surface

    Science.gov (United States)

    Park, Young-Sang; Son, Hyeong-Guk; Kim, Dae-Hoon; Oh, Hong-Gi; Lee, Da-Som; Kim, Min-Hye; Lim, Ki-Moo; Song, Kwang-Soup

    2016-01-01

    Nanocrystalline diamond (NCD) film surfaces were modified with fluorine or oxygen by plasma treatment in an O2 or C3F8 gas environment in order to induce wettability. The oxygenated-NCD (O-NCD) film surface was hydrophilic and the fluorinated-NCD (F-NCD) surface was hydrophobic. The efficiency of early cell adhesion, which is dependent on the wettability of the cell culture plate and necessary for the growth and proliferation of cells, was 89.62 ± 3.92% on the O-NCD film and 7.78 ± 0.77% on the F-NCD film surface after 3 h of cell culture. The wettability of the NCD film surface was artificially modified using a metal mask and plasma treatment to fabricate a micro-pattern. Four types of micro-patterns were fabricated (line, circle, mesh, and word) on the NCD film surface. We precisely arrayed the neuroblastoma cells on the micro-patterned NCD film surfaces by controlling the surface wettability and cell seeding density. The neuroblastoma cells adhered and proliferated along the O-NCD film surface.

  12. Improved adherence and spreading of Saos-2 cells on polypropylene surfaces achieved by surface texturing and carbon nitride coating.

    Science.gov (United States)

    Myllymaa, Katja; Myllymaa, Sami; Korhonen, Hannu; Lammi, Mikko J; Saarenpää, Hanna; Suvanto, Mika; Pakkanen, Tapani A; Tiitu, Virpi; Lappalainen, Reijo

    2009-11-01

    The adhesion and contact guidance of human primary osteogenic sarcoma cells (Saos-2) were characterized on smooth, microstructured (MST) and micro- and nano-structured (MNST) polypropylene (PP) and on the same samples with a silicon-doped carbon nitride (C(3)N(4)-Si) coating. Injection molding was used to pattern the PP surfaces and the coating was obtained by using ultra-short pulsed laser deposition (USPLD). Surfaces were characterized using atomic force microscopy and surface energy components were calculated according to the Owens-Wendt model. The results showed C(3)N(4)-Si coated surfaces to be significantly more hydrophilic than uncoated ones. In addition, there were 86% more cells in the smooth C(3)N(4)-Si coated PP compared to smooth uncoated PP and 551%/476% more cells with MST/MNST C(3)N(4)-Si coated PP than could be obtained with MST/MNST uncoated PP. Thus the adhesion, spreading and contact guidance of osteoblast-like cells was effectively improved by combining surface texturing and deposition of osteocompatible C(3)N(4)-Si coating.

  13. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

    NARCIS (Netherlands)

    Tarazanova, Mariya; Huppertz, Thom; Beerthuyzen, Marke; van Schalkwijk, Saskia; Janssen, Patrick; Wels, Michiel; Kok, Jan; Bachmann, Herwig

    2017-01-01

    Surface properties of bacteria are determined by the molecular composition of the cell wall and they are important for interactions of cells with their environment. Well-known examples of bacterial interactions with surfaces are biofilm formation and the fermentation of solid materials like food and

  14. Spatial and temporal changes in the morphology of preosteoblastic cells seeded on microstructured tantalum surfaces

    DEFF Research Database (Denmark)

    Justesen, Jørn; Lorentzen, M.; Andersen, L. K.

    2009-01-01

    It has been widely reported that surface morphology on the micrometer scale affects cell function as well as cell shape. In this study, we have systematically compared the influence of 13 topographically micropatterned tantalum surfaces on the temporal development of morphology, including spreadi...

  15. Cell Surface Enzymatic Engineering-Based Approaches to Improve Cellular Therapies

    KAUST Repository

    AbuElela, Ayman

    2014-06-06

    The cell surface represents the interface between the cell and its environment. As such, the cell surface controls cell–cell interactions and functions such as adhesion and migration, and will transfer external cues to regulate processes such as survival, death, and differentiation. Redefining the cell surface by temporarily (or permanently) modifying the molecular landscape of the plasma membrane affects the way in which the cell interacts with its environment and influences the information that is relayed into the cell along downstream signaling pathways. This chapter outlines the role of key enzymes, the glycosyltransferases, in posttranslationally modifying proteins and lipids to fine-tune cells, ability to migrate. These enzymes are critical in controlling the formation of a platform structure, sialyl Lewis x (sLex), on circulating cells that plays a central role in the recognition and recruitment by selectin counter receptors on endothelial cells that line blood vessels of tissues throughout the body. By developing methods to manipulate the activity of these enzymes and hence the cell surface structures that result, treatments can be envisioned that direct the migration of therapeutic cells to specific locations throughout the body and also to inhibit metastasis of detrimental cells such as circulating tumor cells.

  16. Nanotextured titanium surfaces stimulate spreading, migration, and growth of rat mast cells.

    Science.gov (United States)

    Marcatti Amarú Maximiano, William; Marino Mazucato, Vivian; Tambasco de Oliveira, Paulo; Célia Jamur, Maria; Oliver, Constance

    2017-08-01

    Titanium is a biomaterial widely used in dental and orthopedic implants. Since tissue-implant interactions occur at the nanoscale level, nanotextured titanium surfaces may affect cellular activity and modulate the tissue response that occurs at the tissue-implant interface. Therefore, the characterization of diverse cell types in response to titanium surfaces with nanotopography is important for the rational design of implants. Mast cells are multifunctional cells of the immune system that release a range of chemical mediators involved in the inflammatory response that occurs at the tissue-implant interface. Therefore, the aim of this study was to investigate the effects of the nanotopography of titanium surfaces on the physiology of mast cells. The results show that the nanotopography of titanium surfaces promoted the spreading of mast cells, which was accompanied by the reorganization of the cytoskeleton. Also, the nanotopography of titanium surfaces enhanced cell migration and cell growth, but did not alter the number of adherent cells in first hours of culture or affect focal adhesions and mediator release. Thus, the results show that nanotopography of titanium surfaces can affect mast cell physiology, and represents an improved strategy for the rational production of surfaces that stimulate tissue integration with the titanium implants. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2150-2161, 2017. © 2017 Wiley Periodicals, Inc.

  17. Multidimensional profiling of cell surface proteins and nuclear markers

    Energy Technology Data Exchange (ETDEWEB)

    Han, Ju; Chang, Hang; Andarawewa, Kumari; Yaswen, Paul; Helen Barcellos-Hoff, Mary; Parvin, Bahram

    2009-01-30

    Cell membrane proteins play an important role in tissue architecture and cell-cell communication. We hypothesize that segmentation and multidimensional characterization of the distribution of cell membrane proteins, on a cell-by-cell basis, enable improved classification of treatment groups and identify important characteristics that can otherwise be hidden. We have developed a series of computational steps to (i) delineate cell membrane protein signals and associate them with a specific nucleus; (ii) compute a coupled representation of the multiplexed DNA content with membrane proteins; (iii) rank computed features associated with such a multidimensional representation; (iv) visualize selected features for comparative evaluation through heatmaps; and (v) discriminate between treatment groups in an optimal fashion. The novelty of our method is in the segmentation of the membrane signal and the multidimensional representation of phenotypic signature on a cell-by-cell basis. To test the utility of this method, the proposed computational steps were applied to images of cells that have been irradiated with different radiation qualities in the presence and absence of other small molecules. These samples are labeled for their DNA content and E-cadherin membrane proteins. We demonstrate that multidimensional representations of cell-by-cell phenotypes improve predictive and visualization capabilities among different treatment groups, and identify hidden variables.

  18. Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

    Energy Technology Data Exchange (ETDEWEB)

    Kado, T.; Hidaka, T. [Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Aita, H. [Division of Occlusion and Removable Prosthodontics, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Endo, K. [Division of Biomaterials and Bioengineering, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Furuichi, Y., E-mail: furuichi@hoku-iryo-u.ac.jp [Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan)

    2012-12-01

    Highlights: Black-Right-Pointing-Pointer Cell-adhesive molecules were covalently immobilized on a Ti surface. Black-Right-Pointing-Pointer Immobilized cell-adhesive molecules maintained native function on the Ti surface. Black-Right-Pointing-Pointer Immobilized collagen enhanced adhesion of periodontal ligament cells to the Ti. - Abstract: A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully

  19. CXCR3 surface expression in human airway epithelial cells: cell cycle dependence and effect on cell proliferation.

    Science.gov (United States)

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

    2006-05-01

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

  20. Adsorption of Amorphous Silica Nanoparticles onto Hydroxyapatite Surfaces Differentially Alters Surfaces Properties and Adhesion of Human Osteoblast Cells.

    Directory of Open Access Journals (Sweden)

    Priya Kalia

    Full Text Available Silicon (Si is suggested to be an important/essential nutrient for bone and connective tissue health. Silicon-substituted hydroxyapatite (Si-HA has silicate ions incorporated into its lattice structure and was developed to improve attachment to bone and increase new bone formation. Here we investigated the direct adsorption of silicate species onto an HA coated surface as a cost effective method of incorporating silicon on to HA surfaces for improved implant osseointegration, and determined changes in surface characteristics and osteoblast cell adhesion. Plasma-sprayed HA-coated stainless steel discs were incubated in silica dispersions of different concentrations (0-42 mM Si, at neutral pH for 12 h. Adsorbed Si was confirmed by XPS analysis and quantified by ICP-OES analysis following release from the HA surface. Changes in surface characteristics were determined by AFM and measurement of surface wettability. Osteoblast cell adhesion was determined by vinculin plaque staining. Maximum Si adsorption to the HA coated disc occurred after incubation in the 6 mM silica dispersion and decreased progressively with higher silica concentrations, while no adsorption was observed with dispersions below 6 mM Si. Comparison of the Si dispersions that produced the highest and lowest Si adsorption to the HA surface, by TEM-based analysis, revealed an abundance of small amorphous nanosilica species (NSP of ~1.5 nm in diameter in the 6 mM Si dispersion, with much fewer and larger NSP in the 42 mM Si dispersions. 29Si-NMR confirmed that the NSPs in the 6 mM silica dispersion were polymeric and similar in composition to the larger NSPs in the 42 mM Si dispersion, suggesting that the latter were aggregates of the former. Amorphous NSP adsorbed from the 6 mM dispersion on to a HA-coated disc surface increased the surface's water contact angle by 53°, whereas that adsorbed from the 42 mM dispersion decreased the contact angle by 18°, indicating increased and

  1. A probabilistic approach to measure the strength of bone cell adhesion to chemically modified surfaces.

    Science.gov (United States)

    Rezania, A; Thomas, C H; Healy, K E

    1997-01-01

    Patterned surfaces with alternating regions of amino silanes [N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane (EDS)] and alkyl silanes [dimethyldichlorosilane (DMS)] have been used to alter the kinetics of spatial distribution of cells in vitro. In particular, we have previously observed the preferential spatial distribution of bone cells on the EDS regions of EDS/ DMS patterned surfaces (10). In this study, we examined whether the mechanism of spatial distribution of cells on the EDS regions was adhesion mediated. Homogeneous layers of EDS and DMS were immobilized on quartz substrates and characterized by contact angle. X-ray photoelectron spectroscopy, and spectroscopic ellipsometry. The strength of bone cell attachment to the modified substrates was examined using a radial flow apparatus, within either 20 min or 2 hr of cell incubation in the presence of serum. A Weibull distribution was chosen to characterize the strength of cell-substratum adhesion. Within 20 min of cell exposure, the strength of adhesion was significantly larger on EDS and clean surfaces, compared with DMS surfaces (p < 0.001). Within 2 hr of cell incubation, there was no statistical difference between the strength of cell adhesion to EDS, DMS, and clean surfaces. The results of this study suggest that the surface chemistry mediates adhesion-based spatial cell arrangement through a layer of adsorbed serum proteins.

  2. Surface effects and electrochemical cell capacitance in desorption electrospray ionization.

    Science.gov (United States)

    Volný, Michael; Venter, Andre; Smith, Scott A; Pazzi, Marco; Cooks, R Graham

    2008-04-01

    Time resolved measurements show that during a desorption electrospray ionization (DESI) experiment, the current initially rises sharply, followed by an exponential decrease to a relatively steady current. When the high voltage on the spray emitter is switched off, the current drops to negative values, suggesting that the direction of current flow in the equivalent DESI circuit is reversed. These data demonstrate that the DESI source behaves as a dc capacitor and that the addition of a surface between the sprayer and the counter electrode in DESI introduces a new electrically active element into the system. The charging and discharging behavior was observed using different surfaces and it could be seen both by making current measurements on a plate at the entrance to the mass spectrometer as well as by measuring ion current in the linear ion trap within the vacuum system of the mass spectrometer. The magnitude of the steady state current obtained without analyte present on the surface is different for different surface materials, and different capacitor time constants of the equivalent RC circuits were calculated for different DESI surfaces. The PTFE surface has by far the greatest time constant and is also able to produce the highest DESI currents. Surface properties play a crucial role in charge transfer during DESI in addition to the effects of the chemical properties of the analyte. It is suggested that surface energy (wettability) is an important factor controlling droplet behavior on the surface. The experimental data are correlated with critical surface tension values of different materials. It is proposed, based on the results presented, that super-hydrophobic materials with extremely high contact angles have the potential to be excellent DESI substrates. It is also demonstrated, using the example of the neurotransmitter dopamine, that the surface charge that develops during a DESI-MS experiment can cause electrochemical oxidation of the analyte.

  3. Poly-lactic-glycolic-acid surface nanotopographies selectively decrease breast adenocarcinoma cell functions

    Science.gov (United States)

    Zhang, Lijuan; Webster, Thomas J.

    2012-04-01

    The ability of poly(lactic-co-glycolic acid) (PLGA, 50:50 PLG/PGA, wt%) nanotopographies to decrease lung epithelial carcinoma cell functions (including adhesion, proliferation, apoptosis and vascular endothelial growth factor (VEGF) secretion) has been previously reported. Specifically, results demonstrated decreased lung epithelial carcinoma cell VEGF synthesis on 23 nm surface-featured PLGA compared to traditional nanosmooth PLGA. However, clearly, different cell lines could have different behaviors on similar biomaterials. Thus, to investigate the universality of nanopatterned PLGA substrates to inhibit numerous cancer cell functions, here, breast epithelial adenocarcinoma cell (MCF-7) adhesion, proliferation, apoptosis and VEGF secretion were determined on different PLGA nanometer surface topographies. To isolate surface nanotopographical effects from all other surface properties, PLGA surfaces with various nanotopographies but similar chemistry and hydrophobicity were fabricated here. Atomic force microscopy (AFM) verified the varied nanotopographies on the PLGA surfaces prepared in this study. Importantly, results demonstrated for the first time significantly decreased breast adenocarcinoma cell functions (including decreased proliferation rate, increased apoptosis and decreased VEGF synthesis) on 23 nm featured PLGA surfaces compared to all other PLGA surface topographies fabricated (specifically, nanosmooth, 300 and 400 nm surface-featured PLGA surfaces). In contrast, healthy breast epithelial cells proliferated more (24%) on the 23 nm featured PLGA surfaces compared to all other PLGA samples. In summary, these results provided further insights into understanding the role PLGA surface nanotopographies can have on cancer cell functions and, more importantly, open the possibility of using polymer nanotopographies for a wide range of anticancer regenerative medicine applications (without resorting to the use of chemotherapeutics).

  4. Surface nanotopography guides kidney-derived stem cell differentiation into podocytes.

    Science.gov (United States)

    MacGregor-Ramiasa, Melanie; Hopp, Isabel; Bachhuka, Akash; Murray, Patricia; Vasilev, Krasimir

    2017-07-01

    Stem cells have enormous potential for developing novel therapies for kidney disease but our current inability to direct their differentiation to specialised renal cells presents a barrier to their use in renal bioengineering and drug development programmes. Here, a plasma-based technology was used to produce a range of biocompatible substrates comprising controlled surface nanotopography and tailored outermost chemical functionalities. These novel substrata were used to investigate the response of mouse kidney-derived stem cells to changes in both substrate nanotopography and surface chemistry. The stem cells proliferated to a similar extent on all substrates, but specific combinations of nanotopography and surface chemistry promoted differentiation into either podocyte or proximal tubule-like cells. The data reveal that high density of surface nanodefects in association with amine rich chemistry primarily lead to differentiation into podocytes while surfaces with low amine content constituted better substrates for differentiation into proximal tubule cells regardless of the surface nanotopographic profile. Thus plasma coated nanorough substrate may provide useful platform for guiding the fate kidney stem cell in vitro. Adult kidney-derived stem cells have been identified as a promising way to regenerate damaged nephrons. Artificial growth platforms capable to guide the stem cells differentiation into useful cell lineages are needed to expand regenerative cell therapies for chronic kidney diseases. Chemically homogeneous growth substrates endowed with nanotopography gradients were generated via plasma assisted methods in order to investigate the effect of physical cues on the proliferation and differentiation of kidney-derived stem cells. For the first time it is shown that the surface density of the nano-structures had a greater impact on fate of the stem cells than their size. Careful design of the growth substrate nanotopography may help directing the

  5. Enterococcus faecalis strains show culture heterogeneity in cell surface charge

    NARCIS (Netherlands)

    van Merode, Annet; van der Mei, HC; Busscher, HJ; Waar, K; Krom, BP

    Adhesion of micro-organisms to biotic and abiotic surfaces is an important virulence factor and involves different types of interactions. Enterococcus faecalis, a human commensal and an important opportunistic pathogen, has the ability to adhere to surfaces. Biliary stents frequently become clogged

  6. Surface-engineered substrates for improved human pluripotent stem cell culture under fully defined conditions.

    Science.gov (United States)

    Saha, Krishanu; Mei, Ying; Reisterer, Colin M; Pyzocha, Neena Kenton; Yang, Jing; Muffat, Julien; Davies, Martyn C; Alexander, Morgan R; Langer, Robert; Anderson, Daniel G; Jaenisch, Rudolf

    2011-11-15

    The current gold standard for the culture of human pluripotent stem cells requires the use of a feeder layer of cells. Here, we develop a spatially defined culture system based on UV/ozone radiation modification of typical cell culture plastics to define a favorable surface environment for human pluripotent stem cell culture. Chemical and geometrical optimization of the surfaces enables control of early cell aggregation from fully dissociated cells, as predicted from a numerical model of cell migration, and results in significant increases in cell growth of undifferentiated cells. These chemically defined xeno-free substrates generate more than three times the number of cells than feeder-containing substrates per surface area. Further, reprogramming and typical gene-targeting protocols can be readily performed on these engineered surfaces. These substrates provide an attractive cell culture platform for the production of clinically relevant factor-free reprogrammed cells from patient tissue samples and facilitate the definition of standardized scale-up friendly methods for disease modeling and cell therapeutic applications.

  7. Bioadsorption of cadmium ion by cell surface-engineered yeasts displaying metallothionein and hexa-His

    Energy Technology Data Exchange (ETDEWEB)

    Kuroda, K.; Ueda, M. [Lab. of Applied Biological Chemistry, Kyoto Univ., Yoshida, Kyoto (Japan)

    2004-07-01

    The Cd{sup 2+}-chelating abilities of yeast metallothionein (YMT) and hexa-His displayed on the yeast-cell surface were compared. Display of YMT and hexa-His by {alpha}-agglutinin-based cell-surface engineering was confirmed by immunofluorescent labeling. Surface-engineered yeast cells with YMT and hexa-His fused in tandem showed superior cell-surface adsorption and recovery of Cd{sup 2+} under EDTA treatment on the cell surface than hexa-His-displaying cells. YMT was demonstrated to be more effective than hexa-His for the adsorption of Cd{sup 2+}. Yeast cells displaying YMT and/or hexa-His exhibited a higher potential for the adsorption of Cd{sup 2+} than Escherichia coli cells displaying these molecules. In order to investigate the effect of the displayed YMT and hexa-His on sensitivity to toxic Cd{sup 2+}, growth in Cd{sup 2+}-containing liquid medium was monitored. Unlike hexa-His-displaying cells, cells displaying YMT and hexa-His fused in tandem induced resistance to Cd{sup 2+} through active and enhanced adsorption of toxic Cd{sup 2+}. These results indicate that YMT-displaying yeast cells are a unique bioadsorbent with a functional chelating ability superior to that of E. coli. (orig.)

  8. Aquatic flower-inspired cell culture platform with simplified medium exchange process for facilitating cell-surface interaction studies.

    Science.gov (United States)

    Hong, Hyeonjun; Park, Sung Jea; Han, Seon Jin; Lim, Jiwon; Kim, Dong Sung

    2016-02-01

    Establishing fundamentals for regulating cell behavior with engineered physical environments, such as topography and stiffness, requires a large number of cell culture experiments. However, cell culture experiments in cell-surface interaction studies are generally labor-intensive and time-consuming due to many experimental tasks, such as multiple fabrication processes in sample preparation and repetitive medium exchange in cell culture. In this work, a novel aquatic flower-inspired cell culture platform (AFIP) is presented. AFIP aims to facilitate the experiments on the cell-surface interaction studies, especially the medium exchange process. AFIP was devised to capture and dispense cell culture medium based on interactions between an elastic polymer substrate and a liquid medium. Thus, the medium exchange can be performed easily and without the need of other instruments, such as a vacuum suction and pipette. An appropriate design window of AFIP, based on scaling analysis, was identified to provide a criterion for achieving stability in medium exchange as well as various surface characteristics of the petal substrates. The developed AFIP, with physically engineered petal substrates, was also verified to exchange medium reliably and repeatedly. A closed structure capturing the medium was sustained stably during cell culture experiments. NIH3T3 proliferation results also demonstrated that AFIP can be applied to the cell-surface interaction studies as an alternative to the conventional method.

  9. Systemic alteration of cell-surface and secreted glycoprotein expression in malignant breast cancer cell lines.

    Science.gov (United States)

    Timpe, Leslie C; Yen, Roger; Haste, Nicole V; Litsakos-Cheung, Christina; Yen, Ten-Yang; Macher, Bruce A

    2013-11-01

    Breast cancer cell lines express fewer transmembrane and secreted glycoproteins than nonmalignant ones. The objective of these experiments was to characterize the changes in the expression of several hundred glycoproteins quantitatively. Secreted and cell-surface glycoproteins were isolated using a glycoprotein capture protocol and then identified by tandem mass spectrometry. Glycoproteins expressed by a group of cell lines originating from malignant tumors of the breast were compared with those expressed by a nonmalignant set. The average number of spectral counts (proportional to relative protein abundance) and the total number of glycopeptides in the malignant samples were reduced to about two-thirds of the level in the nonmalignant samples. Most glycoproteins were expressed at a different level in the malignant samples, with nearly as many increasing as decreasing. The glycoproteins with reduced expression accounted for a larger change in spectral counts, and hence for the net loss of spectral counts in the malignant lines. Similar results were found when the glycoproteins were studied via identified glycosylation sites only, or through identified sites together with non-glycopeptides. The overall reduction is largely due to the loss of integrins, laminins and other proteins that form or interact with the basement membrane.

  10. Cell-surface acceleration of urokinase-catalyzed receptor cleavage

    DEFF Research Database (Denmark)

    Høyer-Hansen, G; Ploug, M; Behrendt, N

    1997-01-01

    relative to the reaction in solution. The time course of uPA-catalyzed cleavage of cell-bound uPAR was studied using U937 cells stimulated with phorbol 12-myristate 13-acetate. Only 30 min was required for 10 nM uPA to cleave 50% of the cell-bound uPAR. This uPA-catalyzed cleavage reaction was inhibited...

  11. Modeling the Excess Cell Surface Stored in a Complex Morphology of Bleb-Like Protrusions.

    Directory of Open Access Journals (Sweden)

    Maryna Kapustina

    2016-03-01

    Full Text Available Cells transition from spread to rounded morphologies in diverse physiological contexts including mitosis and mesenchymal-to-amoeboid transitions. When these drastic shape changes occur rapidly, cell volume and surface area are approximately conserved. Consequently, the rounded cells are suddenly presented with a several-fold excess of cell surface whose area far exceeds that of a smooth sphere enclosing the cell volume. This excess is stored in a population of bleb-like protrusions (BLiPs, whose size distribution is shown by electron micrographs to be skewed. We introduce three complementary models of rounded cell morphologies with a prescribed excess surface area. A 2D Hamiltonian model provides a mechanistic description of how discrete attachment points between the cell surface and cortex together with surface bending energy can generate a morphology that satisfies a prescribed excess area and BLiP number density. A 3D random seed-and-growth model simulates efficient packing of BLiPs over a primary rounded shape, demonstrating a pathway for skewed BLiP size distributions that recapitulate 3D morphologies. Finally, a phase field model (2D and 3D posits energy-based constitutive laws for the cell membrane, nematic F-actin cortex, interior cytosol, and external aqueous medium. The cell surface is equipped with a spontaneous curvature function, a proxy for the cell surface-cortex couple, that is a priori unknown, which the model "learns" from the thin section transmission electron micrograph image (2D or the "seed and growth" model image (3D. Converged phase field simulations predict self-consistent amplitudes and spatial localization of pressure and stress throughout the cell for any posited stationary morphology target and cell compartment constitutive properties. The models form a general framework for future studies of cell morphological dynamics in a variety of biological contexts.

  12. Nanostructured Surfaces to Target and Kill Circulating Tumor Cells While Repelling Leukocytes

    Directory of Open Access Journals (Sweden)

    Michael J. Mitchell

    2012-01-01

    Full Text Available Hematogenous metastasis, the process of cancer cell migration from a primary to distal location via the bloodstream, typically leads to a poor patient prognosis. Selectin proteins hold promise in delivering drug-containing nanocarriers to circulating tumor cells (CTCs in the bloodstream, due to their rapid, force-dependent binding kinetics. However, it is challenging to deliver such nanocarriers while avoiding toxic effects on healthy blood cells, as many possess ligands that adhesively interact with selectins. Herein, we describe a nanostructured surface to capture flowing cancer cells, while preventing human neutrophil adhesion. Microtube surfaces with immobilized halloysite nanotubes (HNTs and E-selectin functionalized liposomal doxorubicin (ES-PEG L-DXR significantly increased the number of breast adenocarcinoma MCF7 cells captured from flow, yet also significantly reduced the number of captured neutrophils. Neutrophils firmly adhered and projected pseudopods on surfaces coated only with liposomes, while neutrophils adherent to HNT-liposome surfaces maintained a round morphology. Perfusion of both MCF7 cells and neutrophils resulted in primarily cancer cell adhesion to the HNT-liposome surface, and induced significant cancer cell death. This work demonstrates that nanostructured surfaces consisting of HNTs and ES-PEG L-DXR can increase CTC recruitment for chemotherapeutic delivery, while also preventing healthy cell adhesion and uptake of therapeutic intended for CTCs.

  13. Cell surface of sea urchin micromeres and primary mesenchyme. [Arbacia punctulata; Strongylocentrotus drobachiensis; Strongylocentrotus purpuratus

    Energy Technology Data Exchange (ETDEWEB)

    DeSimone, D.W.

    1985-01-01

    The cell surface and extracellular matrix (ECM) of the sea urchin embryo were studied during the early morphogenetic events involved in the differentiation of the micromere cell lineage. Sixteen-cell and early cleavage stage blastomeres were isolated and the protein composition of their cell surfaces examined by /sup 125/I-labelling followed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Micromere-specific cell surface proteins are reported for Arbacia punctulata, Strongylocentrotus droebachiensis, and Strongylocentrotus purpuratus. Cell surface glycoproteins were characterized on the basis of lectin binding specificity with a novel lectin affinity transfer technique. Using this procedure, cell-type specific surface proteins, which are also lectin-binding specific, can be detected. In addition, fluorescein conjugated lectins were microinjected into the blastocoels of living S. drobachiensis and Lytechinus pictus embryos and the patterns of lectin bindings observed by fluorescence microscopy. The evidence presented in this thesis suggests that the differentiation of the primary mesenchyme cells is correlated with changes in the molecular composition of the cell-surface and the ECM.

  14. Cell-surface display of the active mannanase in Yarrowia lipolytica with a novel surface-display system.

    Science.gov (United States)

    Yang, Xiao-Song; Jiang, Zheng-Bing; Song, Hui-Ting; Jiang, Si-Jing; Madzak, Catherine; Ma, Li-Xin

    2009-10-13

    A novel surface-display system was constructed using the cell-wall anchor protein Flo1p from Saccharomyces cerevisiae, the mannanase (man1) from Bacillus subtilis fused with the C-terminus of Flo1p and the 6xHis tag was inserted between Flo1p and man1. The fusion protein was displayed on the cell surface of Yarrowia lipolytica successfully, and it was confirmed by immunofluorescence. In succession, the surface-displayed mannanase was characterized. The optimum catalytic conditions for the recombinant mannanase were 55 degrees C at pH 6.0, and it exhibited high stability against pH variation. The highest activity of the recombinant mannanase reached 62.3 IU/g (dry cell weight) after the recombinant was cultivated for 96 h in YPD medium [1% (w/v) yeast extract/2% (w/v) peptone/2% (w/v) glucose]. To our knowledge, the present paper is the first to report that high-activity mannanase is displayed on the cell surface of Y. lipolytica with Flo1p.

  15. Surface-modified yeast cells: A novel eukaryotic carrier for oral application.

    Science.gov (United States)

    Kenngott, Elisabeth E; Kiefer, Ruth; Schneider-Daum, Nicole; Hamann, Alf; Schneider, Marc; Schmitt, Manfred J; Breinig, Frank

    2016-02-28

    The effective targeting and subsequent binding of particulate carriers to M cells in Peyer's patches of the gut is a prerequisite for the development of oral delivery systems. We have established a novel carrier system based on cell surface expression of the β1-integrin binding domain of invasins derived from Yersinia enterocolitica and Yersinia pseudotuberculosis on the yeast Saccharomyces cerevisiae. All invasin derivatives were shown to be effectively expressed on the cell surface and recombinant yeast cells showed improved binding to both human HEp-2 cells and M-like cells in vitro. Among the different derivatives tested, the integrin-binding domain of Y. enterocolitica invasin proved to be the most effective and was able to target Peyer's patches in vivo. In conclusion, cell surface-modified yeasts might provide a novel bioadhesive, eukaryotic carrier system for efficient and targeted delivery of either antigens or drugs via the oral route. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Distinct roles for dystroglycan, beta1 integrin and perlecan in cell surface laminin organization

    DEFF Research Database (Denmark)

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

    2001-01-01

    . Here we show that DG-mediated laminin clustering on mouse embryonic stem (ES) cells is a dynamic process in which clusters are consolidated over time into increasingly more complex structures. Utilizing various null-mutant ES cell lines, we define roles for other molecules in this process. In beta1...... integrin-deficient ES cells, laminin-1 binds to the cell surface, but fails to organize into more morphologically complex structures. This result indicates that beta1 integrin function is required after DG function in the cell surface-mediated laminin assembly process. In perlecan-deficient ES cells......, the formation of complex laminin-1 structures is defective, implicating perlecan in the laminin matrix assembly process. Moreover, laminin and perlecan reciprocally modulate the organization of the other on the cell surface. Taken together, the data support a model whereby DG serves as a receptor essential...

  17. Field Measurements of PCB emissions from Building Surfaces Using a New Portable Emission Test Cell

    DEFF Research Database (Denmark)

    Lyng, Nadja; Haven, Rune; Gunnarsen, Lars Bo

    2016-01-01

    Danish elementary school. The emission test cell was capable of measuring widely varying specific emission rates of PCBtotal (8-3357 ng/(m2·h)). Remediated measures were found to reduce the emission rates by more than 96% compared with similar untreated surfaces. Emission rates may be affected...... by the conditions in the test cell (such as clean air and increased air velocity) and thereby potentially be different without the test cell attached to the surface. Still the measured emission rates obtained by using the test cell are valuable for determination of mitigation strategies. Additionally the test cell...

  18. Proximity Hybridization-Regulated Immunoassay for Cell Surface Protein and Protein-Overexpressing Cancer Cells via Electrochemiluminescence.

    Science.gov (United States)

    Wang, Xiaofei; Gao, Hongfang; Qi, Honglan; Gao, Qiang; Zhang, Chengxiao

    2018-03-06

    A simple electrochemiluminescence (ECL) immunoassay based on a proximity hybridization-regulated strategy was developed for highly sensitive and specific detection of cell surface protein and protein-overexpressing cancer cells. A biosensor was fabricated by self-assembling a thiolated capture ss-DNA3 (partially hybridize with ss-DNA1 and ss-DNA2) and blocking with 6-mercapto-1-hexanol on a gold electrode surface. Target protein was simultaneously bound by two ss-DNA-tagged antibody probes (DNA1-Ab1 and DNA2-Ab2), while DNA1 and DNA2 were brought in sufficient proximity and hybridized with capture DNA3 on the surface of the biosensor. After ECL signal reagent Ru(phen) 3 2+ was intercalated into the hybridized ds-DNAs, ECL measurement was performed in the coreactant solution. A "signal on" proximity hybridization-regulated ECL immunoassay for alpha-fetoprotein (AFP) was developed. The ECL intensity increased with the increase of AFP concentration in the range of 0.05-20.0 ng/mL with a detection limit of 6.2 pg/mL. Moreover, the developed ECL method was successfully used to detect AFP-overexpressing cancer cells (MCF-7 cancer cells as model) with a detection limit of 620 cells/mL (∼60 MCF-7 cells in 100 μL of cell suspension) and discriminate AFP expression on different types of the living cell surface. This work for the first time reports a proximity hybridization-regulated ECL immunoassay for the detection of the cell surface protein on a living cell surface with good specificity and sensitivity. This simple, specific, and sensitive strategy is greatly promising for the detection of proteins and specific cells.

  19. Plasma surface modification of chitosan membranes : characterization and preliminary cell response studies

    OpenAIRE

    Silva, Simone Santos; Luna, Sandra M.; Gomes, Manuela E.; Benesch, Johan; Pashkuleva, I.; Mano, J. F.; Reis, R. L.

    2008-01-01

    Surface modification of biomaterials is a way to tailor cell responses whilst retaining the bulk properties. In this work, chitosan membranes were prepared by solvent casting and treated with nitrogen or argon plasma at 20Wfor 10–40 min. AFM indicated an increase in the surface roughness as a result of the ongoing etching process. XPS and contact angle measurements showed different surface elemental compositions and higher surface free energy. The MTS test and direct contact...

  20. CARbodies: Human Antibodies Against Cell Surface Tumor Antigens Selected From Repertoires Displayed on T Cell Chimeric Antigen Receptors

    Directory of Open Access Journals (Sweden)

    Vanesa Alonso-Camino

    2013-01-01

    Full Text Available A human single-chain variable fragment (scFv antibody library was expressed on the surface of human T cells after transduction with lentiviral vectors (LVs. The repertoire was fused to a first-generation T cell receptor ζ (TCRζ-based chimeric antigen receptor (CAR. We used this library to isolate antibodies termed CARbodies that recognize antigens expressed on the tumor cell surface in a proof-of-principle system. After three rounds of activation-selection there was a clear repertoire restriction, with the emergence dominant clones. The CARbodies were purified from bacterial cultures as soluble and active proteins. Furthermore, to validate its potential application for adoptive cell therapy, human T cells were transduced with a LV encoding a second-generation costimulatory CAR (CARv2 bearing the selected CARbodies. Transduced human primary T cells expressed significant levels of the CARbodies-based CARv2 fusion protein on the cell surface, and importantly could be specifically activated, after stimulation with tumor cells. This approach is a promising tool for the generation of antibodies fully adapted to the display format (CAR and the selection context (cell synapse, which could extend the scope of current adoptive cell therapy strategies with CAR-redirected T cells.

  1. Alzheimer's amyloid precursor protein is expressed on the surface of hematopoietic cells upon activation.

    Science.gov (United States)

    Bullido, M J; Muñoz-Fernandez, M A; Recuero, M; Fresno, M; Valdivieso, F

    1996-08-21

    A4-amyloid is the major component of senile plaques and neurofibrillary tangles found in the brain of patients suffering Alzheimer's disease. This 39-42 amino acid peptide is derived from a larger precursor protein (APP). Since APP gene encodes for a putative membrane protein, the study of APP expression at the cell surface may provide useful data for understanding its physiological function. In this report, we present data on APP expression, that was detected by APP specific mAbs in cells of the hematopoietic system. APP was weakly expressed on the cell surface of resting human lymphocytes and monocytes, but it could be induced to the surface of those cells upon stimulation. The cell activators capable of inducing APP membrane expression comprehended mitogenic lectins, calcium ionophores, phosphatase inhibitors, and anti mu-chain or anti-CD3 antibodies in B and T cells, respectively. Interestingly, phorbol esters were able to induce APP membrane expression in monocytic, but not in lymphoid cells. In contrast to lymphocytes and monocytes, granulocytes never expressed cell surface or cytoplasmic APP, even after the activation. The induction of membrane APP in response to lymphocyte activation signals, including antibodies to the antigen receptor of B and T cells, raises the possibility that APP might play the role of a cell surface receptor in the immune system.

  2. Surface modification of Chlorella vulgaris cells using magnetite particles

    Czech Academy of Sciences Publication Activity Database

    Procházková, G.; Šafařík, Ivo; Brányik, T.

    2012-01-01

    Roč. 42, č. 2012 (2012), s. 1778-1787 E-ISSN 1877-7058 Institutional support: RVO:67179843 Keywords : microalgae * physicochemical approaches * surface interactions * magnetite * XDLVO theory * harvesting Subject RIV: EI - Biotechnology ; Bionics

  3. Macromolecular cell surface engineering for accelerated and reversible cellular aggregation.

    OpenAIRE

    Amaral, A. J.; Pasparakis, G.

    2015-01-01

    We report the synthesis of two simple copolymers that induce rapid cell aggregation within minutes in a fully reversible manner. The polymers can act as self-supporting "cellular glues" or as "drivers" of 3D cell spheroids/aggregates formation at minute concentrations.

  4. Macromolecular cell surface engineering for accelerated and reversible cellular aggregation.

    Science.gov (United States)

    Amaral, Adérito J R; Pasparakis, George

    2015-12-25

    We report the synthesis of two simple copolymers that induce rapid cell aggregation within minutes in a fully reversible manner. The polymers can act as self-supporting "cellular glues" or as "drivers" of 3D cell spheroids/aggregates formation at minute concentrations.

  5. Antimicrobial design of titanium surface that kill sessile bacteria but support stem cells adhesion

    Science.gov (United States)

    Zhu, Chen; Bao, Ni-Rong; Chen, Shuo; Zhao, Jian-Ning

    2016-12-01

    Implant-related bacterial infection is one of the most severe postoperative complications in orthopedic or dental surgery. In this context, from the perspective of surface modification, increasing efforts have been made to enhance the antibacterial capability of titanium surface. In this work, a hierarchical hybrid surface architecture was firstly constructed on titanium surface by two-step strategy of acid etching and H2O2 aging. Then silver nanoparticles were firmly immobilized on the hierarchical surface by ion implantation, showing no detectable release of silver ions from surface. The designed titanium surface showed good bioactivity. More importantly, this elaborately designed titanium surface can effectively inactivate the adherent S. aureus on surface by virtue of a contact-killing mode. Meanwhile, the designed titanium surface can significantly facilitate the initial adhesion and spreading behaviors of bone marrow mesenchymal stem cells (MSCs) on titanium. The results suggested that, the elaborately designed titanium surface might own a cell-favoring ability that can help mammalian cells win the initial adhesion race against bacteria. We hope the present study can provide a new insight for the better understanding and designing of antimicrobial titanium surface, and pave the way to satisfying clinical requirements.

  6. Skeletal (stromal) stem cells

    DEFF Research Database (Denmark)

    Abdallah, Basem M; Kermani, Abbas Jafari; Zaher, Walid

    2015-01-01

    Skeletal (marrow stromal) stem cells (BMSCs) are a group of multipotent cells that reside in the bone marrow stroma and can differentiate into osteoblasts, chondrocytes and adipocytes. Studying signaling pathways that regulate BMSC differentiation into osteoblastic cells is a strategy....../preadipocyte factor 1 (Dlk1/Pref-1), the Wnt co-receptor Lrp5 and intracellular kinases. This article is part of a Special Issue entitled: Stem Cells and Bone....

  7. Tumor suppressor protein SMAR1 modulates the roughness of cell surface: combined AFM and SEM study

    Directory of Open Access Journals (Sweden)

    Mamgain Hitesh

    2009-10-01

    Full Text Available Abstract Background Imaging tools such as scanning electron microscope (SEM and atomic force microscope (AFM can be used to produce high-resolution topographic images of biomedical specimens and hence are well suited for imaging alterations in cell morphology. We have studied the correlation of SMAR1 expression with cell surface smoothness in cell lines as well as in different grades of human breast cancer and mouse tumor sections. Methods We validated knockdown and overexpression of SMAR1 using RT-PCR as well as Western blotting in human embryonic kidney (HEK 293, human breast cancer (MCF-7 and mouse melanoma (B16F1 cell lines. The samples were then processed for cell surface roughness studies using atomic force microscopy (AFM and scanning electron microscopy (SEM. The same samples were used for microarray analysis as well. Tumors sections from control and SMAR1 treated mice as well as tissues sections from different grades of human breast cancer on poly L-lysine coated slides were used for AFM and SEM studies. Results Tumor sections from mice injected with melanoma cells showed pronounced surface roughness. In contrast, tumor sections obtained from nude mice that were first injected with melanoma cells followed by repeated injections of SMAR1-P44 peptide, exhibited relatively smoother surface profile. Interestingly, human breast cancer tissue sections that showed reduced SMAR1 expression exhibited increased surface roughness compared to the adjacent normal breast tissue. Our AFM data establishes that treatment of cells with SMAR1-P44 results into increase in cytoskeletal volume that is supported by comparative gene expression data showing an increase in the expression of specific cytoskeletal proteins compared to the control cells. Altogether, these findings indicate that tumor suppressor function of SMAR1 might be exhibited through smoothening of cell surface by regulating expression of cell surface proteins. Conclusion Tumor suppressor

  8. Surface characteristics determining the cell compatibility of ionically cross-linked alginate gels

    International Nuclear Information System (INIS)

    Machida-Sano, Ikuko; Hirakawa, Makoto; Matsumoto, Hiroki; Kamada, Mitsuki; Ogawa, Sakito; Satoh, Nao; Namiki, Hideo

    2014-01-01

    In this study we investigated differences in the characteristics determining the suitability of five types of ion (Fe 3+ , Al 3+ , Ca 2+ , Ba 2+ and Sr 2+ )-cross-linked alginate films as culture substrates for cells. Human dermal fibroblasts were cultured on each alginate film to examine the cell affinity of the alginates. Since cell behavior on the surface of a material is dependent on the proteins adsorbed to it, we investigated the protein adsorption ability and surface features (wettability, morphology and charge) related to the protein adsorption abilities of alginate films. We observed that ferric, aluminum and barium ion-cross-linked alginate films supported better cell growth and adsorbed higher amounts of serum proteins than other types. Surface wettability analysis demonstrated that ferric and aluminum ion-cross-linked alginates had moderate hydrophilic surfaces, while other types showed highly hydrophilic surfaces. The roughness was exhibited only on barium ion-cross-linked alginate surface. Surface charge measurements revealed that alginate films had negatively charged surfaces, and showed little difference among the five types of gel. These results indicate that the critical factors of ionically cross-linked alginate films determining the protein adsorption ability required for their cell compatibility may be surface wettability and morphology. (paper)

  9. Cell-surface serglycin promotes adhesion of myeloma cells to collagen type I and affects the expression of matrix metalloproteinases.

    Science.gov (United States)

    Skliris, Antonis; Labropoulou, Vassiliki T; Papachristou, Dionysios J; Aletras, Alexios; Karamanos, Nikos K; Theocharis, Achilleas D

    2013-05-01

    Serglycin (SG) is mainly expressed by hematopoetic cells as an intracellular proteoglycan. Multiple myeloma cells constitutively secrete SG, which is also localized on the cell surface in some cell lines. In this study, SG isolated from myeloma cells was found to interact with collagen type I (Col I), which is a major bone matrix component. Notably, myeloma cells positive for cell-surface SG (csSG) adhered significantly to Col I, compared to cells lacking csSG. Removal of csSG by treatment of the cells with chondroitinase ABC or blocking of csSG by an SG-specific polyclonal antibody significantly reduced the adhesion of myeloma cells to Col I. Significant up-regulation of expression of the matrix metalloproteinases MMP-2 and MMP-9 at both the mRNA and protein levels was observed when culturing csSG-positive myeloma cells on Col I-coated dishes or in the presence of soluble Col I. MMP-9 and MMP-2 were also expressed in increased amounts by myeloma cells in the bone marrow of patients with multiple myeloma. Our data indicate that csSG of myeloma cells affects key functional properties, such as adhesion to Col I and the expression of MMPs, and imply that csSG may serve as a potential prognostic factor and/or target for pharmacological interventions in multiple myeloma. © 2013 The Authors Journal compilation © 2013 FEBS.

  10. Evolvement of cell-substrate interaction over time for cells cultivated on a 3-aminopropyltriethoxysilane (γ-APTES) modified silicon dioxide (SiO2) surface

    Science.gov (United States)

    Hsu, Chung-Ping; Hsu, Po-Yen; Wu, You-Lin; Hsu, Wan-Yun; Lin, Jing-Jenn

    2012-09-01

    Since cell-substrate interaction is directly related to the traction force of the cell, the cell property can be judged from the imprint it leaves on the soft substrate surface onto which the cell is cultured. In this letter, the evolvement of the cell-substrate interaction over time was observed by cultivating cells on a 3-aminopropyltriethoxysilane (γ-APTES) modified silicon dioxide (SiO2) surface for different periods of time. The cell-substrate interaction property as a function of time can then be found from the post-cell-removal surface morphology profiles determined by atomic force microscopy (AFM). Different surface morphology profiles were found between normal cells and cancer cells. It was found that the cancer cells tend to form deeper trenches along the circumference of the imprints, while the normal cells do not. In addition, our results indicated that normal cells involve cell-substrate interaction mechanisms that are different from those for cancer cells.

  11. Vector vortex beam generation with dolphin-shaped cell meta-surface.

    Science.gov (United States)

    Yang, Zhuo; Kuang, Deng-Feng; Cheng, Fang

    2017-09-18

    We present a dolphin-shaped cell meta-surface, which is a combination of dolphin-shaped metallic cells and dielectric substrate, for vector vortex beam generation with the illumination of linearly polarized light. Surface plasmon polaritons are excited at the boundary of the metallic cells, then guided by the metallic structures, and finally squeezed to the tips to form highly localized strong electromagnetic fields, which generate the intensity of vector vortex beams at z component. Synchronously, the abrupt phase change produced by the meta-surface is utilized to explain the vortex phase generated by elements. The new kind of structure can be utilized for communication, bioscience, and materiality.

  12. A radioimmunoassay for antibodies against surface membrane antigens using adhering cells

    International Nuclear Information System (INIS)

    Tax, A.; Manson, L.A.

    1976-01-01

    A radioimmunoassay using cells adhering to plastic is described. In this assay, A-10 mammary carcinoma attached to the surface of plastic in microtiter plates were permitted to bind antibody and the bound antibody was detected with purified rabbit 125 I-antimouse-Fab. The bound radioactive material was eluted with glycine-HCl buffer (pH 2.5), and the acid eluates were counted in a gamma counter. This assay can be used to detect cytolic or noncytolic antibody to cell surface antigens in studies with any tumor or normal cell that will adhere to a solid surface

  13. Genetic and proteomic evidences support the localization of yeast enolase in the cell surface

    DEFF Research Database (Denmark)

    López-Villar, Elena; Monteoliva, Lucía; Larsen, Martin Røssel

    2006-01-01

    protein (GFP) as reporter proteins, proved that the 169 N-terminal amino acids are sufficient to target the protein to the cell surface. Furthermore, the enolase-GFP fusion co-localized with a plasma membrane marker. Enolase was also identified among membrane proteins obtained by a purification protocol...... that different experimental approaches (genetics, cellular biology and proteomics) show that yeast enolase can reach the cell surface and describe the protein regions involved in its cell surface targeting. Hybrid enolase truncates, fused at their C terminus with the yeast internal invertase or green fluorescent...

  14. Cellular Cancer Vaccines: an Update on the Development of Vaccines Generated from Cell Surface Antigens

    Directory of Open Access Journals (Sweden)

    Petr G. Lokhov, Elena E. Balashova

    2010-01-01

    Full Text Available A recent advance in anti-cancer therapies has been the use of cancer cells to develop vaccines. However, immunization with cancer cell-based vaccines has not resulted in significant long-term therapeutic benefits. A possible reason for this is that while cancer cells provide surface antigens that are targets for a desired immune response, they also contain a high abundance of housekeeping proteins, carbohydrates, nucleic acids, lipids, and other intracellular contents that are ubiquitous in all mammalian cells. These ubiquitous molecules are not the intended targets of this therapy approach, and thus, the immune response generated is not sufficient to eliminate the cancer cells present. In this review, a discussion of the cell surface of cancer cells is presented in relation to the goals of improving antigen composition of cancer cell-based vaccines. Strategies to enrich vaccines for cancer-specific antigens are also discussed.

  15. Polymer microfilters with nanostructured surfaces for the culture of circulating cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Makarova, Olga V.; Adams, Daniel L.; Divan, Ralu; Rosenmann, Daniel; Zhu, Peixuan; Li, Shuhong; Amstutz, Platte; Tang, Cha-Mei

    2016-09-01

    There is a critical need to improve the accuracy of drug screening and testing through the development of in vitro culture systems that more effectively mimic the in vivo environment. Surface topographical features on the nanoscale level, in short nanotopography, effect the cell growth patterns, and hence affect cell function in culture. We report the preliminary results on the fabrication, and subsequent cellular growth, of nanoscale surface topography on polymer microfilters using cell lines as a precursor to circulating tumor cells (CTCs). To create various nanoscale features on the microfilter surface, we used reactive ion etching (RIE) with and without an etching mask. An anodized aluminum oxide (AAO) membrane fabricated directly on the polymer surface served as an etching mask. Polymer filters with a variety of modified surfaces were used to compare the effects on the culture of cancer cell lines in blank culture wells, with untreated microfilters or with RIE-treated microfilters. We then report the differences of cell shape, phenotype and growth patterns of bladder and glioblastoma cancer cell lines after isolation on the various types of material modifications. Our data suggest that RIE modified polymer filters can isolate model cell lines while retaining ell viability, and that the RIE filter modification allows T24 monolayering cells to proliferate as a structured cluster. Copyright 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Remote control of tissue interactions via engineered photo-switchable cell surfaces.

    Science.gov (United States)

    Luo, Wei; Pulsipher, Abigail; Dutta, Debjit; Lamb, Brian M; Yousaf, Muhammad N

    2014-09-10

    We report a general cell surface molecular engineering strategy via liposome fusion delivery to create a dual photo-active and bio-orthogonal cell surface for remote controlled spatial and temporal manipulation of microtissue assembly and disassembly. Cell surface tailoring of chemoselective functional groups was achieved by a liposome fusion delivery method and quantified by flow cytometry and characterized by a new cell surface lipid pull down mass spectrometry strategy. Dynamic co-culture spheroid tissue assembly in solution and co-culture tissue multilayer assembly on materials was demonstrated by an intercellular photo-oxime ligation that could be remotely cleaved and disassembled on demand. Spatial and temporal control of microtissue structures containing multiple cell types was demonstrated by the generation of patterned multilayers for controlling stem cell differentiation. Remote control of cell interactions via cell surface engineering that allows for real-time manipulation of tissue dynamics may provide tools with the scope to answer fundamental questions of cell communication and initiate new biotechnologies ranging from imaging probes to drug delivery vehicles to regenerative medicine, inexpensive bioreactor technology and tissue engineering therapies.

  17. Remote Control of Tissue Interactions via Engineered Photo-switchable Cell Surfaces

    Science.gov (United States)

    Luo, Wei; Pulsipher, Abigail; Dutta, Debjit; Lamb, Brian M.; Yousaf, Muhammad N.

    2014-01-01

    We report a general cell surface molecular engineering strategy via liposome fusion delivery to create a dual photo-active and bio-orthogonal cell surface for remote controlled spatial and temporal manipulation of microtissue assembly and disassembly. Cell surface tailoring of chemoselective functional groups was achieved by a liposome fusion delivery method and quantified by flow cytometry and characterized by a new cell surface lipid pull down mass spectrometry strategy. Dynamic co-culture spheroid tissue assembly in solution and co-culture tissue multilayer assembly on materials was demonstrated by an intercellular photo-oxime ligation that could be remotely cleaved and disassembled on demand. Spatial and temporal control of microtissue structures containing multiple cell types was demonstrated by the generation of patterned multilayers for controlling stem cell differentiation. Remote control of cell interactions via cell surface engineering that allows for real-time manipulation of tissue dynamics may provide tools with the scope to answer fundamental questions of cell communication and initiate new biotechnologies ranging from imaging probes to drug delivery vehicles to regenerative medicine, inexpensive bioreactor technology and tissue engineering therapies. PMID:25204325

  18. Characterization of fucosyltransferase activity during mouse spermatogenesis: Evidence for a cell surface fucosyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Cardullo, R.A.; Armant, D.R.; Millette, C.F. (Harvard Medical School, Boston, MA (USA))

    1989-02-21

    Fucosyltransferase activity was quantified in mouse germ cells at different stages of spermatogenesis. Specifically, fucosyltransferase activities of pachytene spermatocytes, round spermatids, and cauda epididymal sperm were compared. Fucosyltranferase activity of mixed germ cells displayed an apparent V{sub max} of 17 pmol (mg of protein){sup {minus}1} min{sup {minus}1} and an apparent K{sub m} of approximately 13 {mu}M for GDP-L-({sup 14}C)fucose in the presence of saturating amounts of asialofetuin at 33{degree}C. Under these conditions, cellular fucosyltransferase activity was found to increase during spermatogenesis. In agreement with assays of intact cells, examination of subcellular fractions indicated that a large fraction of fucosyltransferase activity was associated with the cell surface. The fraction of fucosyltransferase activity that was associated with the cell surface progressively increased throughout spermatogenesis and epididymal maturation so that nearly all of the fucosyltransferase in epididymal sperm was on the cell surface. Specifically, by comparison of activities in the presence and absence of the detergent NP-40, the fraction of fucosyltransferase activity that was associated with the cell surface in pachytene spermatocytes, round spermatids, and epididymal sperm was 0.36, 0.5, and 0.85, respectively. These results suggest that a cell surface fucosyltransferase may be important during differentiation of spermatogenic cells in the testis as well as during epididymal maturation and fertilization.

  19. In vitro osteoinduction of human mesenchymal stem cells in biomimetic surface modified titanium alloy implants.

    Science.gov (United States)

    Santander, Sonia; Alcaine, Clara; Lyahyai, Jaber; Pérez, Maria Angeles; Rodellar, Clementina; Doblaré, Manuel; Ochoa, Ignacio

    2014-01-01

    Interaction between cells and implant surface is crucial for clinical success. This interaction and the associated surface treatment are essential for achieving a fast osseointegration process. Several studies of different topographical or chemical surface modifications have been proposed previously in literature. The Biomimetic Advanced Surface (BAS) topography is a combination of a shot blasting and anodizing procedure. Macroroughness, microporosity of titanium oxide and Calcium/Phosphate ion deposition is obtained. Human mesenchymal stem cells (hMCSs) response in vitro to this treatment has been evaluated. The results obtained show an improved adhesion capacity and a higher proliferation rate when hMSCs are cultured on treated surfaces. This biomimetic modification of the titanium surface induces the expression of osteblastic differentiation markers (RUNX2 and Osteopontin) in the absence of any externally provided differentiation factor. As a main conclusion, our biomimetic surface modification could lead to a substantial improvement in osteoinduction in titanium alloy implants.

  20. Dynamic expression of cell surface hydrophobicity during initial yeast cell growth and before germ tube formation of Candida albicans.

    OpenAIRE

    Hazen, B W; Hazen, K C

    1988-01-01

    Expression of cell surface hydrophobicity (CSH) during initial growth of Candida albicans was monitored. CSH of hydrophobic and hydrophilic yeast cells changed within 30 min upon subculture into fresh medium. Morphologic evidence of germination was preceded by expression of CSH. These results indicate that CSH expression is important in C. albicans growth.

  1. Microassay for measurement of binding of radiolabelled ligands to cell surface molecules

    International Nuclear Information System (INIS)

    Woof, J.M.; Burton, D.R.

    1988-01-01

    An improved technique for measuring the binding of radiolabelled ligands to cell surface molecules has been developed by modification of a procedure using centrifugation through a water-immiscible oil to separate free and cell-bound ligand. It maximises the percentage of ligand bound since cell-bound and free ligand can be separated easily and reproducibly even when very small reaction volumes are used. This permits low levels of ligand radiolabelling and relatively low numbers of cells to be used

  2. IN VITRO TRANSPLANTATION OF GENETICALLY MODIFIED CELLS TO THE TENDON SURFACE

    OpenAIRE

    Couvreur, Paulus J. J.; Zhao, Chunfeng; Murphy, Stephen; Amadio, Peter C.

    2008-01-01

    The objective of this paper was to study in vitro transfection of tendon cells and adherence of transfected cells to different tendon surfaces. Achilles tendon fibroblasts from 2-month-old New Zealand white rabbits were cultured to confluence, after which the cells were transfected by an adenovirus carrying either the β-galactosidase reporter gene or the green fluorescent protein (GFP) gene at multiplicities of infection (MOIs) of 50, 100, or 500. Two days later, the cells were transplanted o...

  3. The control of mesenchymal stem cell differentiation using dynamically tunable surface microgrooves.

    Science.gov (United States)

    Gong, Tao; Zhao, Kun; Yang, Guang; Li, Jinrong; Chen, Hongmei; Chen, Yuping; Zhou, Shaobing

    2014-10-01

    Many studies have demonstrated the potential to modulate stem cell differentiation by using static material substrate surfaces. However, cells actually grow in a dynamically diverse microenvironment in vivo. The regulated signals to the differentiation provided by these materials should not be passive or static but be active and dynamic. To mimic the endogenous cell culture microenvironment, a novel system is designed to realize the dynamic change of the surface geometries as well as a resultant mechanical force using a thermally activated four-stage shape memory polymer. The parallel microgroove surface patterns are fabricated via thermal embossing lithography on the polymer substrate surface. The dynamic microgroove surfaces accompanying with the mechanical force can effectively and significantly regulate the shape and the cytoskeletal arrangement of rBMSC compared with the static patterned and non-patterned surfaces. Cellular and molecular analyses reveal that the spatiotemporally programmed regulation of cell shape is more viable to coax lineage-specific differentiation of stem cell in contrast to the general reports with the static surfaces. Therefore, this study provides a facile strategy in designing and manufacturing an artificial substrate with a mimic natural cellular environment to precisely direct the cell differentiation. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Polycarbonate surface cell's adhesion examination after Nd:YAG laser irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Ramazani, S.A. Ahmad, E-mail: Ramazani@sharif.ir [Polymer Group, Department of Chemical and Petroleum Engineering, Sharif University of Technology, Tehran (Iran, Islamic Republic of); Mousavi, Seyyed Abbas, E-mail: Musavi@che.sharif.ir [Department of Chemistry, Sharif University of Technology, Tehran (Iran, Islamic Republic of); Seyedjafari, Ehsan [Department of Biotechnology, University College of Science, University of Tehran (Iran, Islamic Republic of); Poursalehi, Reza [Department of Physics, University of Shahed, Tehran (Iran, Islamic Republic of); Sareh, Shohreh [Research Center of Iranian Blood Transfusion Organization, Tehran (Iran, Islamic Republic of); Silakhori, Kaveh [Laser Research Center, Atomic Energy Organization, Tehran (Iran, Islamic Republic of); Poorfatollah, Ali Akbar [Research Center of Iranian Blood Transfusion Organization, Tehran (Iran, Islamic Republic of); Shamkhali, Amir Nasser [Department of Chemistry, Sharif University of Technology, Tehran (Iran, Islamic Republic of)

    2009-05-05

    Nd:YAG laser treatment was used in order to increase surface cell adhesion aspects of polycarbonate (PC) films prepared via melt process. The treatment was carried out under different wavelengths and beam diameters. ATR-FTIR and UV spectra obtained from different samples before and after laser treatment in air showed that laser irradiation has induced some chemical and physical changes in surface properties. The irradiated films were also characterized using scanning electron microscopy (SEM) and contact angle measurements. Effect of pulse numbers on the surface properties was also investigated. Cell culture test was used to evaluate cell adhesion property on the PC films before and after treatment. The results obtained from this test showed that after laser treatment, the cells were attached and proliferated extensively on the Nd:YAG laser treated films in comparison with the unmodified PC. Moreover, it was revealed that a decrease in the laser beam diameter and an increase in the irradiated pulse numbers increased surface wettability and caused a better cell attachment on the polymer surface. The obtained results also showed that a decrease in the laser beam diameter and an increase in the irradiated pulse numbers increased surface wettability and caused a better cell attachment on the polymer surface.

  5. Yeast cell surface display: An efficient strategy for improvement of bioethanol fermentation performance.

    Science.gov (United States)

    Chen, Xianzhong

    2017-03-04

    The cell surface serves as a functional interface between the inside and the outside of the cell. Within the past 20 y the ability of yeast (Saccharomyces cerevisiae) to display heterologous proteins on the cell surface has been demonstrated. Furthermore, S. cerevisiae has been both developed and applied in expression of various proteins on the cell surface. Using this novel and useful strategy, proteins and peptides of various kinds can be displayed on the yeast cell surface by fusing the protein of interest with the glycosylphosphatidylinositol (GPI)-anchoring system. Consolidated bioprocessing (CBP) using S. cerevisiae represents a promising technology for bioethanol production. However, further work is needed to improve the fermentation performance. There is some excellent previous research regarding construction of yeast biocatalyst using the surface display system to decrease cost, increase efficiency of ethanol production and directly utilize starch or biomass for fuel production. In this commentary, we reviewed the yeast surface display system and highlighted recent work. Additionally, the strategy for decrease of phytate phosphate content in dried distillers grains with solubles (DDGS) by display of phytase on the yeast cell surface is discussed.

  6. FAP-1 Association with Fas (Apo-1) Inhibits Fas Expression on the Cell Surface

    Science.gov (United States)

    Ivanov, Vladimir N.; Bergami, Pablo Lopez; Maulit, Gabriel; Sato, Taka-Aki; Sassoon, David; Ronai, Ze'ev

    2003-01-01

    As revealed by intracellular pools of nonactive Fas (Apo-1), export of Fas to the cell surface is often impaired in human tumors, thereby inactivating Fas ligand-mediated apoptosis. Here, we demonstrate that association with Fas-associated phosphatase 1 (FAP-1) attenuates Fas export to the cell surface. Forced expression of FAP-1 reduces cell surface Fas levels and increases the intracellular pool of Fas within the cytoskeleton network. Conversely, expression of dominant-negative forms of FAP-1, or inhibition of FAP-1 expression by short interfering RNA, efficiently up-regulates surface expression of Fas. Inhibition of Fas surface expression by FAP-1 depends on its association with the C terminus of Fas. Mutation within amino acid 275 results in decreased association with FAP-1 and greater export of Fas to the cell surface in melanomas, normal fibroblasts, or Fas null cells. Identifying the role of FAP-1 in binding to, and consequently inhibition of, Fas export to the cell surface provides novel insight into the mechanism underlying the regulation of Fas trafficking, which is commonly impaired in advanced tumors with FAP-1 overexpression. PMID:12724420

  7. Titanium Surface Coating with a Laminin-Derived Functional Peptide Promotes Bone Cell Adhesion

    Directory of Open Access Journals (Sweden)

    Seung-Ki Min

    2013-01-01

    Full Text Available Laminin-derived peptide coatings can enhance epithelial cell adhesion to implants, and the positive effect of these peptides on bone cell adhesion has been anticipated. The purpose of this study was to evaluate the improvement in bone cell attachment to and activity on titanium (Ti scaffolds coated with a laminin-derived functional peptide, Ln2-P3 (the DLTIDDSYWYRI motif. Four Ti disc surfaces were prepared, and a human osteosarcoma (HOS cell attachment test was performed to select two candidate surfaces for peptide coating. These two candidates were then coated with Ln2-P3 peptide, a scrambled peptide, or left uncoated to measure cell attachment to each surface, following which one surface was chosen to assess alkaline phosphatase (ALP activity and osteogenic marker gene expression with quantitative real-time PCR. On the commercially pure Ti surface, the Ln2-P3 coating significantly increased cellular ALP activity and the expression levels of ALP and bone sialoprotein mRNA as compared with the scrambled peptide-coated and uncoated surfaces. In conclusion, although further in vivo studies are needed, the findings of this in vitro study indicate that the Ln2-P3-coated implant surface promotes bone cell adhesion, which has clinical implications for reducing the overall treatment time of dental implant therapy.

  8. Adhesion, Proliferation and Migration of NIH/3T3 Cells on Modified Polyaniline Surfaces.

    Science.gov (United States)

    Rejmontová, Petra; Capáková, Zdenka; Mikušová, Nikola; Maráková, Nela; Kašpárková, Věra; Lehocký, Marián; Humpolíček, Petr

    2016-09-15

    Polyaniline shows great potential and promises wide application in the biomedical field thanks to its intrinsic conductivity and material properties, which closely resemble natural tissues. Surface properties are crucial, as these predetermine any interaction with biological fluids, proteins and cells. An advantage of polyaniline is the simple modification of its surface, e.g., by using various dopant acids. An investigation was made into the adhesion, proliferation and migration of mouse embryonic fibroblasts on pristine polyaniline films and films doped with sulfamic and phosphotungstic acids. In addition, polyaniline films supplemented with poly (2-acrylamido-2-methyl-1-propanesulfonic) acid at various ratios were tested. Results showed that the NIH/3T3 cell line was able to adhere, proliferate and migrate on the pristine polyaniline films as well as those films doped with sulfamic and phosphotungstic acids; thus, utilization of said forms in biomedicine appears promising. Nevertheless, incorporating poly (2-acrylamido-2-methyl-1-propanesulfonic) acid altered the surface properties of the polyaniline films and significantly affected cell behavior. In order to reveal the crucial factor influencing the surface/cell interaction, cell behavior is discussed in the context of the surface energy of individual samples. It was clearly demonstrated that the lesser the difference between the surface energy of the sample and cell, the more cyto-compatible the surface is.

  9. Culture of neural cells on silicon wafers with nano-scale surface topograph.

    Science.gov (United States)

    Fan, Y W; Cui, F Z; Hou, S P; Xu, Q Y; Chen, L N; Lee, I-S

    2002-10-15

    The adherence and viability of central neural cells (substantia nigra) on a thin layer of SiO(2) on Si wafers with different surface roughness were investigated. Variable roughness of the Si wafer surface was achieved by etching. The nano-scale surface topography was evaluated by atomic force microscopy. The adherence and subsequent viability of the cells on the wafer were examined by scanning electron microscopy (SEM) and fluorescence immunostaining of tyrosine hydroxylase (TH). It is found that the surface roughness significantly affected cell adhesion and viability. Cells survived for over 5 days with normal morphology and expressed neuronal TH when grown on surfaces with an average roughness (Ra) ranging from 20 to 50 nm. However, cell adherence was adversely affected when surfaces with Ra less than 10 nm and rough surfaces with Ra above 70 nm were used as the substrate. Such a simple preparation procedure may provide a suitable interface surface for silicon-based devices and neurones or other living tissues.

  10. Visible tumor surface response to physical plasma and apoptotic cell kill in head and neck cancer.

    Science.gov (United States)

    Schuster, Matthias; Seebauer, Christian; Rutkowski, Rico; Hauschild, Anna; Podmelle, Fred; Metelmann, Camilla; Metelmann, Bibiana; von Woedtke, Thomas; Hasse, Sybille; Weltmann, Klaus-Dieter; Metelmann, Hans-Robert

    2016-09-01

    The aim of the study was to learn, whether clinical application of cold atmospheric pressure plasma (CAP) is able to cause (i) visible tumor surface effects and (ii) apoptotic cell kill in squamous cell carcinoma and (iii) whether CAP-induced visible tumor surface response occurs as often as CAP-induced apoptotic cell kill. Twelve patients with advanced head and neck cancer and infected ulcerations received locally CAP followed by palliative treatment. Four of them revealed tumor surface response appearing 2 weeks after intervention. The tumor surface response expressed as a flat area with vascular stimulation (type 1) or a contraction of tumor ulceration rims forming recesses covered with scabs, in each case surrounded by tumor tissue in visible progress (type 2). In parallel, 9 patients with the same kind of cancer received CAP before radical tumor resection. Tissue specimens were analyzed for apoptotic cells. Apoptotic cells were detectable and occurred more frequently in tissue areas previously treated with CAP than in untreated areas. Bringing together both findings and placing side by side the frequency of clinical tumor surface response and the frequency of analytically proven apoptotic cell kill, detection of apoptotic cells is as common as clinical tumor surface response. There was no patient showing signs of an enhanced or stimulated tumor growth under influence of CAP. CAP was made applicable by a plasma jet, kINPen(®) MED (neoplas tools GmbH, Greifswald, Germany). Copyright © 2016. Published by Elsevier Ltd.

  11. Near-surface alloys for hydrogen fuel cell applications

    DEFF Research Database (Denmark)

    Greeley, Jeffrey Philip; Mavrikakis, Manos

    2006-01-01

    Near-surface alloys (NSAs) possess a variety of unusual catalytic properties that could make them useful candidates for improved catalysts in a variety of chemical processes. It is known from previous work, for example, that some NSAs bind hydrogen very weakly while, at the same time, permitting ...

  12. Hydrophobic and Electrostatic Cell Surface Properties of Thermophilic Dairy Streptococci

    NARCIS (Netherlands)

    Van der Mei, HC; de Vries, Jacob; Busscher, HJ

    1993-01-01

    Microbial adhesion to hydrocarbons (MATH) and microelectrophoresis were done in 10 mM potassium phosphate solutions to characterize the surfaces of thermophilic dairy streptococci, isolated from pasteurizers. Regardless of whether they were grown (in M17 broth) with lactose, sucrose, or glucose

  13. Surface Characterization and Cell Adhesion of Different Zirconia Treatments: An in vitro Study.

    Science.gov (United States)

    Nassif, Wadih; Rifai, Mohamad

    2018-02-01

    The aim of this study was to characterize the surface of zirconia subjected to different treatments and evaluate its effect on cell adhesion and proliferation. A total of 80 zirconia disks were divided into four groups (n = 20) according to the surface treatments used: group I: as-sintered (AS), no surface treatment applied; group II: abrasion treatment applied using Rocatec (ROC; 3M ESPE) system with silica-coated alumina powder of grit size 110 μm; group III: erbium, chromium:yttrium, scandium, gallium, garnet (Er, Cr:YSGG) laser (LAS; BIOLASE) was used at a frequency of 20 Hz and output power of 3 W; and group IV: specimens were subjected to the selective infiltration etching (SIE) technique. Surface characterization was evaluated for the different groups (roughness, hardness, and morphology), and cell behavior (adhesion and proliferation) was tested (a = 0.05). The ROC group reported a significant increase in surface roughness (2.201 ± 0.352) and Vickers hardness (1758 ± 16.6) compared with the other surface treatments. The SIE surface-treated group reported a significantly higher number of cells (64.5 ± 2.6 and 53.5 ± 2.2 respectively) compared with the other surface-treated groups. The SIE is a promising surface treatment for zirconia that significantly enhances cell adhesion and osseointegration. The SIE treatment of zirconia implants may help in a faster and better osseointegration.

  14. Functionalization of CoCr surfaces with cell adhesive peptides to promote HUVECs adhesion and proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Castellanos, Maria Isabel, E-mail: maria.isabel.castellanos@upc.edu [Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgical Engineering, Technical University of Catalonia (UPC), ETSEIB, 08028 Barcelona (Spain); Centre for Research in Nanoengineering (CRNE), UPC, 08028 Barcelona (Spain); Mas-Moruno, Carlos, E-mail: carles.mas.moruno@upc.edu [Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgical Engineering, Technical University of Catalonia (UPC), ETSEIB, 08028 Barcelona (Spain); Centre for Research in Nanoengineering (CRNE), UPC, 08028 Barcelona (Spain); Grau, Anna, E-mail: agraugar@gmail.com [Biomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgical Engineering, Technical University of Catalonia (UPC), ETSEIB, 08028 Barcelona (Spain); Centre for Research in Nanoengineering (CRNE), UPC, 08028 Barcelona (Spain); Serra-Picamal, Xavier, E-mail: xserrapicamal@gmail.com [Institute for Bioengineering of Catalonia (IBEC), 08028 Barcelona (Spain); University of Barcelona and CIBER-BBN, 08036 Barcelona (Spain); Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona (Spain); Trepat, Xavier, E-mail: xtrepat@ub.edu [Institute for Bioengineering of Catalonia (IBEC), 08028 Barcelona (Spain); University of Barcelona and CIBER-BBN, 08036 Barcelona (Spain); Institució Catalana de Recerca i Estudis Avançats (ICREA), 08010 Barcelona (Spain); Albericio, Fernando, E-mail: fernando.albericio@irbbarcelona.org [Department of Chemistry, University of Barcelona, CIBER-BBN, 08028 Barcelona (Spain); Joner, Michael, E-mail: michaeljoner@me.com [Department of Cardiology, Deutsches Herzzentrum München, 80636 Munich (Germany); CVPath Institute, Gaithersburg, MD 20878 (United States); and others

    2017-01-30

    Highlights: • We immobilized peptides on CoCr alloy through physisorption and covalent bonding. • Surface activation is an essential step prior to silanization to enhance peptide attachment. • Biofunctionalized surface characteristics were discussed. • RGDS, YIGSR and combination peptides display an improved HUVECs adhesion and proliferation. - Abstract: Biomimetic surface modification with peptides that have specific cell-binding moieties is a promising approach to improve endothelialization of metal-based stents. In this study, we functionalized CoCr surfaces with RGDS, REDV, YIGSR peptides and their combinations to promote endothelial cells (ECs) adhesion and proliferation. An extensive characterization of the functionalized surfaces was performed by XPS analysis, surface charge and quartz crystal microbalance with dissipation monitoring (QCM-D), which demonstrated the successful immobilization of the peptides to the surface. Cell studies demonstrated that the covalent functionalization of CoCr surfaces with an equimolar combination of RGDS and YIGSR represents the most powerful strategy to enhance the early stages of ECs adhesion and proliferation, indicating a positive synergistic effect between the two peptide motifs. Although these peptide sequences slightly increased smooth muscle cells (SMCs) adhesion, these values were ten times lower than those observed for ECs. The combination of RGDS with the REDV sequence did not show synergistic effects in promoting the adhesion or proliferation of ECs. The strategy presented in this study holds great potential to overcome clinical limitations of current metal stents by enhancing their capacity to support surface endothelialization.

  15. Cell-to-cell heterogeneity in cortical tension specifies curvature of contact surfaces in Caenorhabditis elegans embryos.

    Directory of Open Access Journals (Sweden)

    Masashi Fujita

    Full Text Available In the two-cell stage embryos of Caenorhabditis elegans, the contact surface of the two blastomeres forms a curve that bulges from the AB blastomere to the P₁ blastomere. This curve is a consequence of the high intracellular hydrostatic pressure of AB compared with that of P₁. However, the higher pressure in AB is intriguing because AB has a larger volume than P₁. In soap bubbles, which are a widely used model of cell shape, a larger bubble has lower pressure than a smaller bubble. Here, we reveal that the higher pressure in AB is mediated by its higher cortical tension. The cell fusion experiments confirmed that the curvature of the contact surface is related to the pressure difference between the cells. Chemical and genetic interferences showed that the pressure difference is mediated by actomyosin. Fluorescence imaging indicated that non-muscle myosin is enriched in the AB cortex. The cell killing experiments provided evidence that AB but not P₁ is responsible for the pressure difference. Computer simulation clarified that the cell-to-cell heterogeneity of cortical tensions is indispensable for explaining the pressure difference. This study demonstrates that heterogeneity in surface tension results in significant deviations of cell behavior compared to simple soap bubble models, and thus must be taken into consideration in understanding cell shape within embryos.

  16. Transfer plate radioassay using cell monolayers to detect anti-cell surface antibodies synthesized by lymphocyte hybridomas

    International Nuclear Information System (INIS)

    Schneider, M.D.; Eisenbarth, G.S.

    1979-01-01

    A solid phase [ 125 I] Protein A radioassay for anti-cell surface antibodies is described, which employs target cell monolayers cultured on fenestrated polyvinyl chloride 96-well plates ('transfer plates'). The calibrated aperture in the bottom of each well is small enough to retain fluid contents by surface tension during monolayer growth, but also permits fluid to enter the wells when transfer plate are lowered into receptacles containing washing buffer on test sera. To assay for antibodies directed against target cell surface antigens, transfer plates bearing monolayers are inserted into microculture plates with corresponding 96-well geometry, thereby simultaneously sampling 96 wells. This assay allows rapid screening of hundreds of hybrid cell colonies for production of antibodies with desired tissue specificity. (Auth.)

  17. Isthmin targets cell-surface GRP78 and triggers apoptosis via induction of mitochondrial dysfunction

    Science.gov (United States)

    Chen, M; Zhang, Y; Yu, V C; Chong, Y-S; Yoshioka, T; Ge, R

    2014-01-01

    Isthmin (ISM) is a secreted 60-kDa protein that potently induces endothelial cell (EC) apoptosis. It suppresses tumor growth and angiogenesis in mice when stably overexpressed in cancer cells. Although αvβ5 integrin serves as a low-affinity receptor for ISM, the mechanism by which ISM mediates antiangiogenesis and apoptosis in ECs remain to be fully resolved. In this work, we report the identification of cell-surface glucose-regulated protein 78 kDa (GRP78) as a high-affinity receptor for ISM (Kd=8.6 nM). We demonstrated that ISM-GRP78 interaction triggers apoptosis not only in activated ECs but also in cancer cells expressing high level of cell-surface GRP78. Normal cells and benign tumor cells tend to express low level of cell-surface GRP78 and are resistant to ISM-induced apoptosis. Upon binding to GRP78, ISM is internalized into ECs through clathrin-dependent endocytosis that is essential for its proapoptotic activity. Once inside the cell, ISM co-targets with GRP78 to mitochondria where it interacts with ADP/ATP carriers on the inner membrane and blocks ATP transport from mitochondria to cytosol, thereby causing apoptosis. Hence, ISM is a novel proapoptotic ligand that targets cell-surface GRP78 to trigger apoptosis by inducing mitochondrial dysfunction. The restricted and high-level expression of cell-surface GRP78 on cancer cells and cancer ECs make them uniquely susceptible to ISM-targeted apoptosis. Indeed, systemic delivery of recombinant ISM potently suppressed subcutaneous 4T1 breast carcinoma and B16 melanoma growth in mice by eliciting apoptosis selectively in the cancer cells and cancer ECs. Together, this work reveals a novel ISM-GRP78 apoptosis pathway and demonstrates the potential of ISM as a cancer-specific and dual-targeting anticancer agent. PMID:24464222

  18. Cell wall trapping of autocrine peptides for human G-protein-coupled receptors on the yeast cell surface.

    Directory of Open Access Journals (Sweden)

    Jun Ishii

    Full Text Available G-protein-coupled receptors (GPCRs regulate a wide variety of physiological processes and are important pharmaceutical targets for drug discovery. Here, we describe a unique concept based on yeast cell-surface display technology to selectively track eligible peptides with agonistic activity for human GPCRs (Cell Wall Trapping of Autocrine Peptides (CWTrAP strategy. In our strategy, individual recombinant yeast cells are able to report autocrine-positive activity for human GPCRs by expressing a candidate peptide fused to an anchoring motif. Following expression and activation, yeast cells trap autocrine peptides onto their cell walls. Because captured peptides are incapable of diffusion, they have no impact on surrounding yeast cells that express the target human GPCR and non-signaling peptides. Therefore, individual yeast cells can assemble the autonomous signaling complex and allow single-cell screening of a yeast population. Our strategy may be applied to identify eligible peptides with agonistic activity for target human GPCRs.

  19. Determining Cell-surface Expression and Endocytic Rate of Proteins in Primary Astrocyte Cultures Using Biotinylation.

    Science.gov (United States)

    Tham, Daniel Kai Long; Moukhles, Hakima

    2017-07-03

    Cell-surface proteins mediate a wide array of functions. In many cases, their activity is regulated by endocytic processes that modulate their levels at the plasma membrane. Here, we present detailed protocols for 2 methods that facilitate the study of such processes, both of which are based on the principle of the biotinylation of cell-surface proteins. The first is designed to allow for the semi-quantitative determination of the relative levels of a particular protein at the cell-surface. In it, the lysine residues of the plasma membrane proteins of cells are first labeled with a biotin moiety. Once the cells are lysed, these proteins may then be specifically precipitated via the use of agarose-immobilized streptavidin by exploiting the natural affinity of the latter for biotin. The proteins isolated in such a manner may then be analyzed via a standard western blotting approach. The second method provides a means of determining the endocytic rate of a particular cell-surface target over a period of time. Cell-surface proteins are first modified with a biotin derivative containing a cleavable disulfide bond. The cells are then shifted back to normal culture conditions, which causes the endocytic uptake of a proportion of biotinylated proteins. Next, the disulfide bonds of non-internalized biotin groups are reduced using the membrane-impermeable reducing agent glutathione. Via this approach, endocytosed proteins may thus be isolated and quantified with a high degree of specificity.

  20. Improved accuracy of cell surface shaving proteomics in Staphylococcus aureus using a false-positive control

    DEFF Research Database (Denmark)

    Solis, Nestor; Larsen, Martin Røssel; Cordwell, Stuart J

    2010-01-01

    Proteolytic treatment of intact bacterial cells is an ideal means for identifying surface-exposed peptide epitopes and has potential for the discovery of novel vaccine targets. Cell stability during such treatment, however, may become compromised and result in the release of intracellular proteins...... that complicate the final analysis. Staphylococcus aureus is a major human pathogen, causing community and hospital-acquired infections, and is a serious healthcare concern due to the increasing prevalence of multiple antibiotic resistances amongst clinical isolates. We employed a cell surface "shaving" technique...... to trypsin and three identified in the control. The use of a subtracted false-positive strategy improved enrichment of surface-exposed peptides in the trypsin data set to approximately 80% (124/155 peptides). Predominant surface proteins were those associated with methicillin resistance-surface protein SACOL...

  1. Time-kill profiles and cell-surface morphological effects of crude ...

    African Journals Online (AJOL)

    MK1201 mycelial extract on the viability and cell surface morphology of methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA). Methods: Time-kill assays were conducted by incubating test ...

  2. Role of the retinoblastoma protein in cell cycle arrest mediated by a novel cell surface proliferation inhibitor

    Science.gov (United States)

    Enebo, D. J.; Fattaey, H. K.; Moos, P. J.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    A novel cell regulatory sialoglycopeptide (CeReS-18), purified from the cell surface of bovine cerebral cortex cells has been shown to be a potent and reversible inhibitor of proliferation of a wide array of fibroblasts as well as epithelial-like cells and nontransformed and transformed cells. To investigate the possible mechanisms by which CeReS-18 exerts its inhibitory action, the effect of the inhibitor on the posttranslational regulation of the retinoblastoma susceptibility gene product (RB), a tumor suppressor gene, has been examined. It is shown that CeReS-18 mediated cell cycle arrest of both human diploid fibroblasts (HSBP) and mouse fibroblasts (Swiss 3T3) results in the maintenance of the RB protein in the hypophosphorylated state, consistent with a late G1 arrest site. Although their normal nontransformed counterparts are sensitive to cell cycle arrest mediated by CeReS-18, cell lines lacking a functional RB protein, through either genetic mutation or DNA tumor virus oncoprotein interaction, are less sensitive. The refractory nature of these cells is shown to be independent of specific surface receptors for the inhibitor, and another tumor suppressor gene (p53) does not appear to be involved in the CeReS-18 inhibition of cell proliferation. The requirement for a functional RB protein product, in order for CeReS-18 to mediate cell cycle arrest, is discussed in light of regulatory events associated with density-dependent growth inhibition.

  3. Near-surface alloys for hydrogen fuel cell applications

    DEFF Research Database (Denmark)

    Greeley, Jeffrey Philip; Mavrikakis, Manos

    2006-01-01

    facile H-2 activation. These NSAs could, potentially, facilitate highly selective hydrogenation reactions at low temperatures. In the present work, the suitability of NSAs for use as hydrogen fuel cell anodes has been evaluated: the combination of properties, possessed by selected NSAs, of weak binding...... of such materials for use in fuel cells and in an ever. increasing range of catalytic applications. Furthermore, we introduce a new concept for NSA-defect sites, which could be responsible for the promotional catalytic effects of a second metal added. even in minute quantities, to a host metal catalyst....

  4. Surface grafting of carboxylic groups onto thermoplastic polyurethanes to reduce cell adhesion

    Energy Technology Data Exchange (ETDEWEB)

    Alves, P., E-mail: palves@eq.uc.pt [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal); Ferreira, P. [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal); Kaiser, Jean-Pierre [EMPA, St. Gallen, Lerchenfeldstrasse 5, CH-9014 St. Gallen (Switzerland); Salk, Natalie [Mikrofertigung – Micro Engineering, Fraunhofer IFAM, Wiener Strasse 12, D-288359 Bremen (Germany); Bruinink, Arie [EMPA, St. Gallen, Lerchenfeldstrasse 5, CH-9014 St. Gallen (Switzerland); Sousa, Hermínio C. de; Gil, M.H. [CIEPQPF, Departamento de Engenharia Química, Universidade de Coimbra, Polo II, Pinhal de Marrocos, 3030-790 Coimbra (Portugal)

    2013-10-15

    The interaction of polymers with other materials is an important issue, being their surface properties clearly crucial. For some important polymer applications, their surfaces have to be modified. Surface modification aims to tailor the surface characteristics of a material for a specific application without affecting its bulk properties. Materials can be surface modified by using biological, chemical or physical methods. The aim of this work was to improve the reactivity of the thermoplastic polyurethane (TPU) material (Elastollan{sup ®}) surface and to make its surface cell repellent by grafting carboxylic groups onto its surface. Two TPU materials were studied: a polyether-based TPU and a polyester-based TPU. The grafting efficiency was evaluated by contact angle measurements and by analytical determination of the COOH groups. Scanning electron microscopy (SEM) of the membranes surface was performed as well as cell adhesion tests. It was proved that the surfaces of the TPUs membranes were successfully modified and that cell adhesion was remarkably reduced.

  5. Cell surface carbohydrate changes during embryonic and fetal skin development

    DEFF Research Database (Denmark)

    Dabelsteen, Erik; Holbrook, K; Clausen, H

    1986-01-01

    Monoclonal antibodies to four type 2 chain carbohydrate antigens were used for immunohistochemical studies of embryonic and fetal skin. The antibodies detected N-acetyllactosamine and 3 fucosyl substitutes of this, blood group antigen H, Lex, and Ley. Periderm consistently stained for N-acetyllac......Monoclonal antibodies to four type 2 chain carbohydrate antigens were used for immunohistochemical studies of embryonic and fetal skin. The antibodies detected N-acetyllactosamine and 3 fucosyl substitutes of this, blood group antigen H, Lex, and Ley. Periderm consistently stained for N......-acetyllactosamine, Lex and Ley. The H antigen showed a variable and weak expression on peridermal cells from day 57 to day 84 estimated gestation age (EGA). After this period the H antigen was no longer expressed at peridermal cells. In the epidermis, N-acetyllactosamine was present on all cells until the age of 15...... weeks EGA. After this period N-acetyllactosamine could only be demonstrated on basal cells after treatment with neuraminidase, indicating a masking of N-acetyllactosamine by sialic acid. The H antigen could not be demonstrated in the epithelium before 14 weeks EGA. At this time it appeared on spinous...

  6. DMPD: Innate immune sensing of pathogens and danger signals by cell surface Toll-likereceptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17275324 Innate immune sensing of pathogens and danger signals by cell surface Toll... Show Innate immune sensing of pathogens and danger signals by cell surface Toll-likereceptors. PubmedID 172...75324 Title Innate immune sensing of pathogens and danger signals by cell surface

  7. Recent Insights into Cell Surface Heparan Sulphate Proteoglycans and Cancer [version 1; referees: 3 approved

    Directory of Open Access Journals (Sweden)

    John R Couchman

    2016-06-01

    Full Text Available A small group of cell surface receptors are proteoglycans, possessing a core protein with one or more covalently attached glycosaminoglycan chains. They are virtually ubiquitous and their chains are major sites at which protein ligands of many types interact. These proteoglycans can signal and regulate important cell processes, such as adhesion, migration, proliferation, and differentiation. Since many protein ligands, such as growth factors, morphogens, and cytokines, are also implicated in tumour progression, it is increasingly apparent that cell surface proteoglycans impact tumour cell behaviour. Here, we review some recent advances, emphasising that many tumour-related functions of proteoglycans are revealed only after their modification in processes subsequent to synthesis and export to the cell surface. These include enzymes that modify heparan sulphate structure, recycling of whole or fragmented proteoglycans into exosomes that can be paracrine effectors or biomarkers, and lateral interactions between some proteoglycans and calcium channels that impact the actin cytoskeleton.

  8. Exploring the Leishmania Hydrophilic Acylated Surface Protein B (HASPB) Export Pathway by Live Cell Imaging Methods.

    Science.gov (United States)

    MacLean, Lorna; Price, Helen; O'Toole, Peter

    2016-01-01

    Leishmania major is a human-infective protozoan parasite transmitted by the bite of the female phlebotomine sand fly. The L. major hydrophilic acylated surface protein B (HASPB) is only expressed in infective parasite stages suggesting a role in parasite virulence. HASPB is a "nonclassically" secreted protein that lacks a conventional signal peptide, reaching the cell surface by an alternative route to the classical ER-Golgi pathway. Instead HASPB trafficking to and exposure on the parasite plasma membrane requires dual N-terminal acylation. Here, we use live cell imaging methods to further explore this pathway allowing visualization of key events in real time at the individual cell level. These methods include live cell imaging using fluorescent reporters to determine the subcellular localization of wild type and acylation site mutation HASPB18-GFP fusion proteins, fluorescence recovery after photobleaching (FRAP) to analyze the dynamics of HASPB in live cells, and live antibody staining to detect surface exposure of HASPB by confocal microscopy.

  9. Prognostic significance of cell surface phenotype in acute lymphoblastic leukemia

    Directory of Open Access Journals (Sweden)

    Shiek Aejaz Aziz

    2015-01-01

    Full Text Available Context: To find out the phenotypic character of lymphoblasts of acute lymphoblastic leukemia (ALL patients in our study cohort and their possible effect on the prognosis. Aims: To investigate the phenotype in ALL in our demographic population and to prognosticate various upfront current protocols employed in our hospital. Settings and Design: The study spanned over a period of 4 years with retrospective and prospective data of January 2008 through December 2011. Materials and Methods: 159 patients of all age groups were enrolled for the study, of which flow cytometry was done in 144 patients. Statistical Analysis Used: Analysis was done using the variables on SPSS (statistical package for social sciences software on computer. Survival curves were estimated by method of Kaplan-Meir. Results: Majority of the patients were of B-cell (68.1% and 30.6% patients were of T-cell lineage. Of these, 80.6% patients were having cALLa positivity. Complete remission (CR was achieved in 59.1%, 16.4% relapsed, and 20.1% patients died. Conclusions: Phenotyping has become an important and integral part of diagnosis, classification, management and prognosticating in ALL. B-cell has been found to have a better survival over T-cell lymphoblastic leukemia. cALLa antigen positivity has good impact in achieving CR in only B-cell lineage, myeloid coexpression has no significant effect on the outcome. BFM (Berlin-Frankfurt-Münster based protocols though showed a higher CR and survival vis-a-vis UKALL-XII. However, patients enrolled in former group being of low risk category and lesser in numbers cannot be compared statistically with a fair degree of confidence.

  10. Effect of micro- and nanoscale topography on the adhesion of bacterial cells to solid surfaces.

    Science.gov (United States)

    Hsu, Lillian C; Fang, Jean; Borca-Tasciuc, Diana A; Worobo, Randy W; Moraru, Carmen I

    2013-04-01

    Attachment and biofilm formation by bacterial pathogens on surfaces in natural, industrial, and hospital settings lead to infections and illnesses and even death. Minimizing bacterial attachment to surfaces using controlled topography could reduce the spreading of pathogens and, thus, the incidence of illnesses and subsequent human and financial losses. In this context, the attachment of key microorganisms, including Escherichia coli, Listeria innocua, and Pseudomonas fluorescens, to silica and alumina surfaces with micron and nanoscale topography was investigated. The results suggest that orientation of the attached cells occurs preferentially such as to maximize their contact area with the surface. Moreover, the bacterial cells exhibited different morphologies, including different number and size of cellular appendages, depending on the topographical details of the surface to which they attached. This suggests that bacteria may utilize different mechanisms of attachment in response to surface topography. These results are important for the design of novel microbe-repellant materials.

  11. Efficient Isolation and Quantitative Proteomic Analysis of Cancer Cell Plasma Membrane Proteins for Identification of Metastasis-Associated Cell Surface Markers

    DEFF Research Database (Denmark)

    Lund, Rikke; Leth-Larsen, Rikke; Jensen, Ole N

    2009-01-01

    Cell surface membrane proteins are involved in central processes such as cell signaling, cell-cell interactions, ion and solute transport, and they seem to play a pivotal role in several steps of the metastatic process of cancer cells. The low abundance and hydrophobic nature of cell surface...... membrane proteins complicate their purification and identification by MS. We used two isogenic cell lines with opposite metastatic capabilities in nude mice to optimize cell surface membrane protein purification and to identify potential novel markers of metastatic cancer. The cell surface membrane...... peptides of which 622 (300 at SL80) were membrane proteins. The quantitative proteomic analysis identified 16 cell surface proteins as potential markers of the ability of breast cancer cells to form distant metastases....

  12. Surface chemical functionalities affect the behavior of human adipose-derived stem cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Xujie [State key laboratory of new ceramics and fine processing, School of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng, Qingling, E-mail: biomater@mail.tsinghua.edu.cn [State key laboratory of new ceramics and fine processing, School of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Bachhuka, Akash [Mawson Institute, University of South Australia, Mawson Lakes 5095 (Australia); Vasilev, Krasimir [Mawson Institute, University of South Australia, Mawson Lakes 5095 (Australia); School of Advanced Manufacturing, University of South Australia, Mawson Lakes 5095 (Australia)

    2013-04-01

    This study examines the effect of surface chemical functionalities on the behavior of human adipose-derived stem cells (hASCs) in vitro. Plasma polymerized films rich in amine (-NH{sub 2}), carboxyl (-COOH) and methyl (-CH{sub 3}), were generated on hydroxyapatite (HAp) substrates. The surface chemical functionalities were characterized by X-ray photoelectron spectroscopy (XPS). The ability of different substrates to absorb proteins was evaluated. The results showed that substrates modified with hydrophilic functional group (-COOH and -NH{sub 2}) can absorb more proteins than these modified with more hydrophobic functional group (-CH{sub 3}). The behavior of human adipose-derived stem cells (hASCs) cultured on different substrates was investigated in vitro: cell counting kit-8 (CCK-8) analysis was used to characterize cell proliferation, scanning electronic microscopy (SEM) analysis was used to characterize cell morphology and alkaline phosphatase (ALP) activity analysis was used to account for differentiation. The results of this study demonstrated that the -NH{sub 2} modified surfaces encourage osteogenic differentiation; the -COOH modified surfaces promote cell adhesion and spreading and the -CH{sub 3} modified surfaces have the lowest ability to induce osteogenic differentiation. These findings confirmed that the surface chemical states of biomaterials can affect the behavior of hASCs in vitro.

  13. Effect of surface organic coatings of cellulose nanocrystals on the viability of mammalian cell lines.

    Science.gov (United States)

    Jimenez, Ambar S; Jaramillo, Francesca; Hemraz, Usha D; Boluk, Yaman; Ckless, Karina; Sunasee, Rajesh

    2017-01-01

    Cellulose nanocrystals (CNCs) have emerged as promising candidates for a number of bio-applications. Surface modification of CNCs continues to gain significant research interest as it imparts new properties to the surface of the nanocrystals for the design of multifunctional CNCs-based materials. A small chemical surface modification can potentially lead to drastic behavioral changes of cell-material interactions thereby affecting the intended bio-application. In this work, unmodified CNCs were covalently decorated with four different organic moieties such as a diaminobutane fragment, a cyclic oligosaccharide ( β -cyclodextrin), a thermoresponsive polymer (poly[ N -isopropylacrylamide]), and a cationic aminomethacrylamide-based polymer using different synthetic covalent methods. The effect of surface coatings of CNCs and the respective dose-response of the above organic moieties on the cell viability were evaluated on mammalian cell cultures (J774A.1 and MFC-7), using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphe-nyltetrazolium bromide and lactate dehydrogenase assays. Overall, the results indicated that cells exposed to surface-coated CNCs for 24 h did not display major changes in cell viability, membrane permeability as well as cell morphology. However, with longer exposure, all these parameters were somewhat affected, which appears not to be correlated with either anionic or cationic surface coatings of CNCs used in this study.

  14. Focal Adhesion of Osteoblastic Cells on Titanium Surface with Amine Functionalities Formed by Plasma Polymerization

    Science.gov (United States)

    Song, Heesang; Jung, Sang Chul; Kim, Byung Hoon

    2012-08-01

    To enhance the focal adhesion of osteoblastic cells on a titanium surface, plasma polymerized allyl amine (AAm) thin films were deposited by plasma polymerization. This plasma polymer functionalization of titanium is advantageous for osteoblastic focal adhesion formation. Such Ti surfaces are useful for the fabrication of titanium-based dental implants for enhancement of osseointegration.

  15. Topochip: technology for instructing cell fate and morphology via designed surface topography

    NARCIS (Netherlands)

    Hulshof, G.F.B.

    2016-01-01

    The control of biomaterial surface topography is emerging as a tool to influence cells and tissues. Due to a lack a theoretical framework of the underlying molecular mechanisms, high-throughput screening (HTS) technology is valuable to identify and study bioactive surface topographies. To identify

  16. Pressure effects on interfacial surface contacts and performance of organic solar cells

    NARCIS (Netherlands)

    Agyei-Tuffour, B.; Doumon, Nutifafa Y.; Rwenyagila, E. R.; Asare, J.; Oyewole, O. K.; Shen, Z.; Petoukhoff, C. E.; Zebaze Kana, M. G.; Ocarroll, D. M.; Soboyejo, W. O.

    2017-01-01

    This paper explores the effects of pressure on the interfacial surface contacts and the performance of organic solar cells. A combination of experimental techniques and analytical/computational models is used to study the evolving surface contacts profiles that occur when compliant, semi-rigid and

  17. Genotoxicity of copper oxide nanoparticles with different surface chemistry on rat bone marrow mesenchymal stem cells

    DEFF Research Database (Denmark)

    Zhang, Wenjing; Jiang, Pengfei; Chen, Wei

    2016-01-01

    The surface chemistry of nanoparticles (NPs) is one of the critical factors determining their cellular responses. In this study, the cytotoxicity and genotoxicity of copper oxide (CuO) NPs with a similar size but different surface chemistry to rat bone marrow mesenchymal stem cells (MSCs) were in...

  18. A Synthetic Peptide-Acrylate Surface for Production of Insulin-Producing Cells from Human Embryonic Stem Cells

    Science.gov (United States)

    Lin, Pei-Yi; Hung, Shih-Han; Yang, Yao-Chen; Liao, Li-Chuan; Hsieh, Yi-Cheng; Yen, Hsan-Jan; Lu, Huai-En; Lee, Maw-Sheng

    2014-01-01

    Human embryonic stem cells (hESCs), due to their self-renewal capacity and pluripotency, have become a potential source of transplantable β-cells for the treatment of diabetes. However, it is imperative that the derived cells fulfill the criteria for clinical treatment. In this study, we replaced common Matrigel with a synthetic peptide-acrylate surface (Synthemax) to expand undifferentiated hESCs and direct their differentiation in a defined and serum-free medium. We confirmed that the cells still expressed pluripotent markers, had the ability to differentiate into three germ layers, and maintained a normal karyotype after 10 passages of subculture. Next, we reported an efficient protocol for deriving nearly 86% definitive endoderm cells from hESCs under serum-free conditions. Moreover, we were able to obtain insulin-producing cells within 21 days following a simple three-step protocol. The results of immunocytochemical and quantitative gene expression analysis showed that the efficiency of induction was not significantly different between the Synthemax surface and the Matrigel-coated surface. Thus, we provided a totally defined condition from hESC culture to insulin-producing cell differentiation, and the derived cells could be a therapeutic resource for diabetic patients in the future. PMID:24083371

  19. Application of Environmental Scanning Electron Microscope-Nanomanipulation System on Spheroplast Yeast Cells Surface Observation.

    Science.gov (United States)

    Rad, Maryam Alsadat; Ahmad, Mohd Ridzuan; Nakajima, Masahiro; Kojima, Seiji; Homma, Michio; Fukuda, Toshio

    2017-01-01

    The preparation and observations of spheroplast W303 cells are described with Environmental Scanning Electron Microscope (ESEM). The spheroplasting conversion was successfully confirmed qualitatively, by the evaluation of the morphological change between the normal W303 cells and the spheroplast W303 cells, and quantitatively, by determining the spheroplast conversion percentage based on the OD 800 absorbance data. From the optical microscope observations as expected, the normal cells had an oval shape whereas spheroplast cells resemble a spherical shape. This was also confirmed under four different mediums, that is, yeast peptone-dextrose (YPD), sterile water, sorbitol-EDTA-sodium citrate buffer (SCE), and sorbitol-Tris-Hcl-CaCl 2 (CaS). It was also observed that the SCE and CaS mediums had a higher number of spheroplast cells as compared to the YPD and sterile water mediums. The OD 800 absorbance data also showed that the whole W303 cells were fully converted to the spheroplast cells after about 15 minutes. The observations of the normal and the spheroplast W303 cells were then performed under an environmental scanning electron microscope (ESEM). The normal cells showed a smooth cell surface whereas the spheroplast cells had a bleb-like surface after the loss of its integrity when removing the cell wall.

  20. Immunophenotypic characterization of human T cells after in vitro exposure to different silicone breast implant surfaces.

    Directory of Open Access Journals (Sweden)

    Giuseppe Cappellano

    Full Text Available The most common complication of silicone breast implants is capsular contracture (massive scar formation around the implant. We postulate that capsular contracture is always a sequel to inflammatory processes, with both innate and adaptive immune mechanisms participating. In general, fibroblasts and macrophages have been used as cell types to evaluate in vitro the biocompatibility of breast implant surfaces. Moreover, also T cells have been found at the implant site at the initial stage of fibrous capsule formation. However, only few studies have addressed the influence of surfaces with different textures on T-cell responses. The aim of the present study was to investigate the immune response of human peripheral blood mononuclear cells (PBMC to commercially available silicone breast implants in vitro. PBMC from healthy female blood donors were cultured on each silicone surface for 4 days. Proliferation and phenotype of cultured cells were assessed by flow cytometry. Cytokine levels were determined by multiplex and real-time assay. We found that silicone surfaces do not induce T-cell proliferation, nor do they extensively alter the proportion of T cell subsets (CD4, CD8, naïve, effector memory. Interestingly, cytokine profiling identified matrix specific differences, especially for IL-6 and TNF-α on certain surface topographies that could lead to increased fibrosis.

  1. Cells responding to surface structure of calcium phosphate ceramics for bone regeneration.

    Science.gov (United States)

    Zhang, Jingwei; Sun, Lanying; Luo, Xiaoman; Barbieri, Davide; de Bruijn, Joost D; van Blitterswijk, Clemens A; Moroni, Lorenzo; Yuan, Huipin

    2017-11-01

    Surface structure largely affects the inductive bone-forming potential of calcium phosphate (CaP) ceramics in ectopic sites and bone regeneration in critical-sized bone defects. Surface-dependent osteogenic differentiation of bone marrow stromal cells (BMSCs) partially explained the improved bone-forming ability of submicron surface structured CaP ceramics. In this study, we investigated the possible influence of surface structure on different bone-related cells, which may potentially participate in the process of improved bone formation in CaP ceramics. Besides BMSCs, the response of human brain vascular pericytes (HBVP), C2C12 (osteogenic inducible cells), MC3T3-E1 (osteogenic precursors), SV-HFO (pre-osteoblasts), MG63 (osteoblasts) and SAOS-2 (mature osteoblasts) to the surface structure was evaluated in terms of cell proliferation, osteogenic differentiation and gene expression. The cells were cultured on tricalcium phosphate (TCP) ceramics with either micron-scaled surface structure (TCP-B) or submicron-scaled surface structure (TCP-S) for up to 14 days, followed by DNA, alkaline phosphatase (ALP) and quantitative polymerase chain reaction gene assays. HBVP were not sensitive to surface structure with respect to cell proliferation and osteogenic differentiation, but had downregulated angiogenesis-related gene expression (i.e. vascular endothelial growth factor) on TCP-S. Without additional osteogenic inducing factors, submicron-scaled surface structure enhanced ALP activity and osteocalcin gene expression of human (h)BMSCs and C2C12 cells, favoured the proliferation of MC3T3-E1, MG63 and SAOS-2, and increased ALP activity of MC3T3-E1 and SV-HFO. The results herein indicate that cells with osteogenic potency (either osteogenic inducible cells or osteogenic cells) could be sensitive to surface structure and responded to osteoinductive submicron-structured CaP ceramics in cell proliferation, ALP production or osteogenic gene expression, which favour bone

  2. Modified Titanium Surface-Mediated Effects on Human Bone Marrow Stromal Cell Response

    Science.gov (United States)

    Chaudhari, Amol; Duyck, Joke; Braem, Annabel; Vleugels, Jozef; Petite, Hervé; Logeart-Avramoglou, Delphine; Naert, Ignace; Martens, Johan A.; Vandamme, Katleen

    2013-01-01

    Surface modification of titanium implants is used to enhance osseointegration. The study objective was to evaluate five modified titanium surfaces in terms of cytocompatibility and pro-osteogenic/pro-angiogenic properties for human mesenchymal stromal cells: amorphous microporous silica (AMS), bone morphogenetic protein-2 immobilized on AMS (AMS + BMP), bio-active glass (BAG) and two titanium coatings with different porosity (T1; T2). Four surfaces served as controls: uncoated Ti (Ti), Ti functionalized with BMP-2 (Ti + BMP), Ti surface with a thickened titanium oxide layer (TiO2) and a tissue culture polystyrene surface (TCPS). The proliferation of eGFP-fLuc (enhanced green fluorescence protein-firefly luciferase) transfected cells was tracked non-invasively by fluorescence microscopy and bio-luminescence imaging. The implant surface-mediated effects on cell differentiation potential was tracked by determination of osteogenic and angiogenic parameters [alkaline phosphatase (ALP); osteocalcin (OC); osteoprotegerin (OPG); vascular endothelial growth factor-A (VEGF-A)]. Unrestrained cell proliferation was observed on (un)functionalized Ti and AMS surfaces, whereas BAG and porous titanium coatings T1 and T2 did not support cell proliferation. An important pro-osteogenic and pro-angiogenic potential of the AMS + BMP surface was observed. In contrast, coating the Ti surface with BMP did not affect the osteogenic differentiation of the progenitor cells. A significantly slower BMP-2 release from AMS compared to Ti supports these findings. In the unfunctionalized state, Ti was found to be superior to AMS in terms of OPG and VEGF-A production. AMS is suggested to be a promising implant coating material for bioactive agents delivery. PMID:28788407

  3. Feedback Regulation of Intracellular Hydrostatic Pressure in Surface Cells of the Lens.

    Science.gov (United States)

    Gao, Junyuan; Sun, Xiurong; White, Thomas W; Delamere, Nicholas A; Mathias, Richard T

    2015-11-03

    In wild-type lenses from various species, an intracellular hydrostatic pressure gradient goes from ∼340 mmHg in central fiber cells to 0 mmHg in surface cells. This gradient drives a center-to-surface flow of intracellular fluid. In lenses in which gap-junction coupling is increased, the central pressure is lower, whereas if gap-junction coupling is reduced, the central pressure is higher but surface pressure is always zero. Recently, we found that surface cell pressure was elevated in PTEN null lenses. This suggested disruption of a feedback control system that normally maintained zero surface cell pressure. Our purpose in this study was to investigate and characterize this feedback control system. We measured intracellular hydrostatic pressures in mouse lenses using a microelectrode/manometer-based system. We found that all feedback went through transport by the Na/K ATPase, which adjusted surface cell osmolarity such that pressure was maintained at zero. We traced the regulation of Na/K ATPase activity back to either TRPV4, which sensed positive pressure and stimulated activity, or TRPV1, which sensed negative pressure and inhibited activity. The inhibitory effect of TRPV1 on Na/K pumps was shown to signal through activation of the PI3K/AKT axis. The stimulatory effect of TRPV4 was shown in previous studies to go through a different signal transduction path. Thus, there is a local two-legged feedback control system for pressure in lens surface cells. The surface pressure provides a pedestal on which the pressure gradient sits, so surface pressure determines the absolute value of pressure at each radial location. We speculate that the absolute value of intracellular pressure may set the radial gradient in the refractive index, which is essential for visual acuity. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  4. Modified Titanium Surface-Mediated Effects on Human Bone Marrow Stromal Cell Response

    Directory of Open Access Journals (Sweden)

    Amol Chaudhari

    2013-11-01

    Full Text Available Surface modification of titanium implants is used to enhance osseointegration. The study objective was to evaluate five modified titanium surfaces in terms of cytocompatibility and pro-osteogenic/pro-angiogenic properties for human mesenchymal stromal cells: amorphous microporous silica (AMS, bone morphogenetic protein-2 immobilized on AMS (AMS + BMP, bio-active glass (BAG and two titanium coatings with different porosity (T1; T2. Four surfaces served as controls: uncoated Ti (Ti, Ti functionalized with BMP-2 (Ti + BMP, Ti surface with a thickened titanium oxide layer (TiO2 and a tissue culture polystyrene surface (TCPS. The proliferation of eGFP-fLuc (enhanced green fluorescence protein-firefly luciferase transfected cells was tracked non-invasively by fluorescence microscopy and bio-luminescence imaging. The implant surface-mediated effects on cell differentiation potential was tracked by determination of osteogenic and angiogenic parameters [alkaline phosphatase (ALP; osteocalcin (OC; osteoprotegerin (OPG; vascular endothelial growth factor-A (VEGF-A]. Unrestrained cell proliferation was observed on (unfunctionalized Ti and AMS surfaces, whereas BAG and porous titanium coatings T1 and T2 did not support cell proliferation. An important pro-osteogenic and pro-angiogenic potential of the AMS + BMP surface was observed. In contrast, coating the Ti surface with BMP did not affect the osteogenic differentiation of the progenitor cells. A significantly slower BMP-2 release from AMS compared to Ti supports these findings. In the unfunctionalized state, Ti was found to be superior to AMS in terms of OPG and VEGF-A production. AMS is suggested to be a promising implant coating material for bioactive agents delivery.

  5. Cell surface expression and function of the macromolecular C1 complex on the surface of human monocytes

    Directory of Open Access Journals (Sweden)

    Kinga K Hosszu

    2012-03-01

    Full Text Available The synthesis of the subunits of the C1 complex (C1q, C1s, C1r, and its regulator C1 inhibitor (C1-Inh by human monocytes has been previously established. However, surface expression of these molecules by monocytes has not been shown. Using flow cytometry and antigen-capture ELISA, we show here for the first time that, in addition to C1q, PB monocytes and the monocyte-derived U937 cells express C1s and C1r, as well as Factor B and C1-Inh on their surface. C1s and C1r immunoprecipitated with C1q, suggesting that at least some of the C1q on these cells is part of the C1 complex. Furthermore, the C1 complex on U937 cells was able to trigger complement activation via the classical pathway. The presence of C1-Inh may ensure that an unwarranted autoactivation of the C1 complex does not take place. Since C1-Inh closely monitors the activation of the C1 complex in a sterile or infectious inflammatory environment, further elucidation of the role of C1 complex is crucial to dissect its function in monocyte, DC and T cell activities, and its implications in host defense and tolerance.

  6. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Poulsen Lars K

    2010-09-01

    Full Text Available Abstract Background Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results The proteins were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained their enzymatic activities. Phospholipase A1 severely inhibited the growth of the yeast cells. Antigen 5 - expressing yeast cells bound IgE antibodies from wasp venom allergic patient sera but not from control sera as demonstrated by FACS. Moreover, antigen 5 - expressing yeast cells were capable of mediating allergen-specific histamine release from human basophils. Conclusions All the three major wasp venom allergens were expressed on the yeast surface. A high-level expression, which was observed only for antigen 5, was needed for detection of IgE binding by FACS and for induction of histamine release. The non-modified S. cerevisiae cells did not cause any unspecific reaction in FACS or histamine release assay despite the expression of high-mannose oligosaccharides. In perspective the yeast surface display may be used for allergen discovery from cDNA libraries and possibly for sublingual immunotherapy as the cells can serve as good adjuvant and can be produced in large amounts at a low price.

  7. Effects of Streptococcus sanguinis Bacteriocin on Cell Surface Hydrophobicity, Membrane Permeability, and Ultrastructure of Candida Thallus

    Directory of Open Access Journals (Sweden)

    Shengli Ma

    2015-01-01

    Full Text Available Candida albicans (C.a and Candida tropicalis (C.t were treated with Streptococcus sanguinis bacteriocin (S.s bacteriocin, respectively; the bacteriostatic dynamics of S.s bacteriocin, their effects on cell surface hydrophobicity, leakage of inorganic phosphorus and macromolecular substance, cytosolic calcium concentration, and ultrastructure changes of Candida thallus were detected and analyzed. The results showed that inhibitory effect of S.s bacteriocin on C.a and C.t reached peak level at 24 h, the cell-surface hydrophobicity decreased significantly (P < 0.05 after S.s bacteriocin treatment, and there was leakage of cytoplasmic inorganic phosphorus and macromolecular substance from C.a and C.t; cytosolic calcium concentration decreased greatly. After 24 h treatment by S.s bacteriocin, depressive deformity and defect could be found in the cell surface of C.a and C.t; the thallus displayed irregular forms: C.a was shrunken, there was unclear margins abutting upon cell wall and cell membrane, nucleus disappeared, and cytoplasm was inhomogeneous; likewise, C.t was first plasmolysis, and then the cytoplasm was shrunk, the ultrastructure of cell wall and cell membrane was continuously damaged, and the nucleus was karyolysis. It was illustrated that S.s bacteriocin had similar antifungal effect on C.a and C.t; their cell surface hydrophobicity, membrane permeability, and ultrastructure were changed significantly on exposure to S.s bacteriocin.

  8. Morphology, proliferation, and osteogenic differentiation of mesenchymal stem cells cultured on titanium, tantalum, and chromium surfaces

    DEFF Research Database (Denmark)

    Stiehler, Maik; Lind, M.; Mygind, Tina

    2007-01-01

    the interactions between human mesenchymal stem cells (MSCs) and smooth surfaces of titanium (Ti), tantalum (Ta), and chromium (Cr). Mean cellular area was quantified using fluorescence microscopy (4 h). Cellular proliferation was assessed by (3)H-thymidine incorporation and methylene blue cell counting assays (4...

  9. Protein secretion in the Archaea : multiple paths towards a unique cell surface

    NARCIS (Netherlands)

    Albers, Sonja-Verena; Szabo, Zalan; Driessen, Arnold J. M.

    Archaea are similar to other prokaryotes in most aspects of cell structure but are unique with respect to the lipid composition of the cytoplasmic membrane and the structure of the cell surface. Membranes of archaea are composed of glycerol-ether lipids instead of glycerol-ester lipids and are based

  10. Acrylic acid surface-modified contact lens for the culture of limbal stem cells.

    Science.gov (United States)

    Zhang, Hong; Brown, Karl David; Lowe, Sue Peng; Liu, Guei-Sheung; Steele, David; Abberton, Keren; Daniell, Mark

    2014-06-01

    Surface treatment to a biomaterial surface has been shown to modify and help cell growth. Our aim was to determine the best surface-modified system for the treatment of limbal stem cell deficiency (LSCD), which would facilitate expansion of autologous limbal epithelial cells, while maintaining cultivated epithelial cells in a less differentiated state. Commercially available contact lenses (CLs) were variously surface modified by plasma polymerization with ratios of acrylic acid to octadiene tested at 100% acrylic acid, 50:50% acrylic acid:octadiene, and 100% octadiene to produce high-, mid-, and no-acid. X-ray photoelectron spectroscopy was used to analyze the chemical composition of the plasma polymer deposited layer. Limbal explants cultured on high acid-modified CLs outgrew more cells. Immunofluorescence and RT2-PCR array results indicated that a higher acrylic acid content can also help maintain progenitor cells during ex vivo expansion of epithelial cells. This study provides the first evidence for the ability of high acid-modified CLs to preserve the stemness and to be used as substrates for the culture of limbal cells in the treatment of LSCD.

  11. Detection of apoptosis in cancer cell lines using Surface Plasmon Resonance imaging

    NARCIS (Netherlands)

    Stojanović, I.; van Hal, Y.; van der Velden, T.J.G.; Schasfoort, Richardus B.M.; Terstappen, Leonardus Wendelinus Mathias Marie

    2016-01-01

    Induction of apoptosis in cancer cells by therapeutic agents is an important event to detect the potential effectiveness of therapies. Here we explore the potential of Surface Plasmon Resonance imaging (SPRi) to assess apoptosis in cancer cells exposed to therapeutic agents by measuring the

  12. The modulation of cell surface cAMP receptors from Dictyostelium disscoideum by ammonium sulfate

    NARCIS (Netherlands)

    Haastert, Peter J.M. van

    1985-01-01

    Dictyostelium discoideum cells contain a heterogeneous population of cell surface cAMP receptors with components possessing different affinities (Kd between 15 and 450 nM) and different off-rates of the cAMP-receptor complex (t½ between 0.7 and 150 s). The association of cAMP to the receptor and the

  13. Surface modifications by gas plasma control osteogenic differentiation of MC3T3-E1 cells

    NARCIS (Netherlands)

    Barradas, A.M.C.; Lachmann, K.; Hlawacek, G.; Frielink, C.; Truckenmüller, R.K.; Boerman, O.C.; van Gastel, Raoul; Garritsen, H.S.P.; Garritsen, H.S.P.; Thomas, M.; Moroni, Lorenzo; van Blitterswijk, Clemens; de Boer, Jan

    2012-01-01

    Numerous studies have shown that the physicochemical properties of biomaterials can control cell activity. Cell adhesion, proliferation, differentiation as well as tissue formation in vivo can be tuned by properties such as the porosity, surface micro- and nanoscale topography and chemical

  14. Surface modifications by gas plasma control osteogenic differentiation of MC3T3-E1 cells.

    NARCIS (Netherlands)

    Barradas, A.M.; Lachmann, K.; Hlawacek, G.; Frielink, C.; Truckenmoller, R.; Boerman, O.C.; Gastel, R. van; Garritsen, H.; Thomas, M.; Moroni, L.; Blitterswijk, C. Van; Boer, J. den

    2012-01-01

    Numerous studies have shown that the physicochemical properties of biomaterials can control cell activity. Cell adhesion, proliferation, differentiation as well as tissue formation in vivo can be tuned by properties such as the porosity, surface micro- and nanoscale topography and chemical

  15. Cell surface glycan engineering of neural stem cells augments neurotropism and improves recovery in a murine model of multiple sclerosis

    KAUST Repository

    Merzaban, Jasmeen S.

    2015-09-13

    Neural stem cell (NSC)-based therapies offer potential for neural repair in central nervous system (CNS) inflammatory and degenerative disorders. Typically, these conditions present with multifocal CNS lesions making it impractical to inject NSCs locally, thus mandating optimization of vascular delivery of the cells to involved sites. Here, we analyzed NSCs for expression of molecular effectors of cell migration and found that these cells are natively devoid of E-selectin ligands. Using glycosyltransferase-programmed stereosubstitution (GPS), we glycan engineered the cell surface of NSCs ("GPS-NSCs") with resultant enforced expression of the potent E-selectin ligand HCELL (hematopoietic cell E-/L-selectin ligand) and of an E-selectin-binding glycoform of neural cell adhesion molecule ("NCAM-E"). Following intravenous (i.v.) injection, short-term homing studies demonstrated that, compared with buffer-treated (control) NSCs, GPS-NSCs showed greater neurotropism. Administration of GPS-NSC significantly attenuated the clinical course of experimental autoimmune encephalomyelitis (EAE), with markedly decreased inflammation and improved oligodendroglial and axonal integrity, but without evidence of long-term stem cell engraftment. Notably, this effect of NSC is not a universal property of adult stem cells, as administration of GPS-engineered mouse hematopoietic stem/progenitor cells did not improve EAE clinical course. These findings highlight the utility of cell surface glycan engineering to boost stem cell delivery in neuroinflammatory conditions and indicate that, despite the use of a neural tissue-specific progenitor cell population, neural repair in EAE results from endogenous repair and not from direct, NSC-derived cell replacement.

  16. Targeting and molecular imaging of HepG2 cells using surface-functionalized gold nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Rathinaraj, Pierson [Auckland University of Technology, Institute of Biomedical Technologies (New Zealand); Lee, Kyubae; Choi, Yuri; Park, Soo-Young [Kyungpook National University, School of Applied Chemical Engineering, Graduate School (Korea, Republic of); Kwon, Oh Hyeong [Kumoh National Institute of Technology, Department of Polymer Science and Engineering (Korea, Republic of); Kang, Inn-Kyu, E-mail: ikkang@knu.ac.kr [Kyungpook National University, School of Applied Chemical Engineering, Graduate School (Korea, Republic of)

    2015-07-15

    Mercaptosuccinic acid (M)-conjugated gold nanoparticles (GM) were prepared and characterized by transmission electron microscope and dynamic light scattering. M was used to improve the monodispersity and non-specific intracellular uptake of nanoparticles. Lactobionic acid (L) was subsequently conjugated to the GM to target preferentially HepG2 cells (liver cancer cells) that express asialoglycoprotein receptors (ASGPR) on their membrane surfaces and facilitate the transit of nanoparticles across the cell membrane. The mean size of lactobionic acid-conjugated gold nanoparticle (GL) was approximately 10 ± 0.2 nm. Finally, the Atto 680 dye (A6) was coupled to the nanoparticles to visualize their internalization into HepG2 cells. The interaction of surface-modified gold nanoparticles with HepG2 cells was studied after culturing cells in media containing the GM or L-conjugated GM (GL)

  17. Cell Surface Glycosylation Is Required for Efficient Mating of Haloferax volcanii

    Directory of Open Access Journals (Sweden)

    Yarden Shalev

    2017-07-01

    Full Text Available Halophilic archaea use a fusion-based mating system for lateral gene transfer across cells, yet the molecular mechanisms involved remain unknown. Previous work implied that cell fusion involves cell–cell recognition since fusion occurs more efficiently between cells from the same species. Long believed to be restricted only to Eukarya, it is now known that cells of all three domains of life perform N-glycosylation, the covalent attachment of glycans to select target asparagine residues in proteins, and that this post-translational modification is common for archaeal cell surface proteins. Here, we show that differences in glycosylation of the Haloferax volcanii surface-layer glycoprotein, brought about either by changing medium salinity or by knocking out key glycosylation genes, reduced mating success. Thus, different glycosylation patterns are likely to underlie mating preference in halophilic archaea, contributing to speciation processes.

  18. Adjustment of surface chemical and physical properties with functionalized polymers to control cell adhesion

    Science.gov (United States)

    Zhou, Zhaoli

    Cell-surface interaction is crucial in many cellular functions such as movement, growth, differentiation, proliferation and survival. In the present work, we have developed several strategies to design and prepare synthetic polymeric materials with selected cues to control cell attachment. To promote neuronal cell adhesion on the surfaces, biocompatible, non-adhesive PEG-based materials were modified with neurotransmitter acetylcholine functionalities to produce hydrogels with a range of porous structures, swollen states, and mechanical strengths. Mice hippocampal cells cultured on the hydrogels showed differences in number, length of processes and exhibited different survival rates, thereby highlighting the importance of chemical composition and structure in biomaterials. Similar strategies were used to prepare polymer brushes to assess how topographical cues influence neuronal cell behaviors. The brushes were prepared using the "grown from" method through surface-initiated atom transfer radical polymerization (SI-ATRP) reactions and further patterned via UV photolithography. Protein absorption tests and hippocampal neuronal cell culture of the brush patterns showed that both protein and neuronal cells can adhere to the patterns and therefore can be guided by the patterns at certain length scales. We also prepared functional polymers to discourage attachment of undesirable cells on the surfaces. For example, we synthesized PEG-perfluorinated alkyl amphiphilic surfactants to modify polystyrene-block-poly(ethylene-ran-butylene)- block-polyisoprene (SEBI or K3) triblock copolymers for marine antifouling/fouling release surface coatings. Initial results showed that the polymer coated surfaces can facilitate removal of Ulva sporelings on the surfaces. In addition, we prepared both bioactive and dual functional biopassive/bioactive antimicrobial coatings based on SEBI polymers. Incubating the polymer coated surfaces with gram-positive bacteria (S. aureus), gram

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

    Science.gov (United States)

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

    2018-04-05

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

  20. Selective labelling of cell-surface proteins using CyDye DIGE Fluor minimal dyes.

    Science.gov (United States)

    Hagner-McWhirter, Asa; Winkvist, Maria; Bourin, Stephanie; Marouga, Rita

    2008-11-26

    Surface proteins are central to the cell's ability to react to its environment and to interact with neighboring cells. They are known to be inducers of almost all intracellular signaling. Moreover, they play an important role in environmental adaptation and drug treatment, and are often involved in disease pathogenesis and pathology (1). Protein-protein interactions are intrinsic to signaling pathways, and to gain more insight in these complex biological processes, sensitive and reliable methods are needed for studying cell surface proteins. Two-dimensional (2-D) electrophoresis is used extensively for detection of biomarkers and other targets in complex protein samples to study differential changes. Cell surface proteins, partly due to their low abundance (1 2% of cellular proteins), are difficult to detect in a 2-D gel without fractionation or some other type of enrichment. They are also often poorly represented in 2-D gels due to their hydrophobic nature and high molecular weight (2). In this study, we present a new protocol for intact cells using CyDye DIGE Fluor minimal dyes for specific labeling and detection of this important group of proteins. The results showed specific labeling of a large number of cell surface proteins with minimal labeling of intracellular proteins. This protocol is rapid, simple to use, and all three CyDye DIGE Fluor minimal dyes (Cy 2, Cy 3 and Cy 5) can be used to label cell-surface proteins. These features allow for multiplexing using the 2-D Fluorescence Difference Gel Electrophoresis (2-D DIGE) with Ettan DIGE technology and analysis of protein expression changes using DeCyder 2-D Differential Analysis Software. The level of cell-surface proteins was followed during serum starvation of CHO cells for various lengths of time (see Table 1). Small changes in abundance were detected with high accuracy, and results are supported by defined statistical methods.

  1. Folic acid functionalized surface highlights 5-methylcytosine-genomic content within circulating tumor cells

    KAUST Repository

    Malara, Natalia

    2014-07-01

    Although the detection of methylated cell free DNA represents one of the most promising approaches for relapse risk assessment in cancer patients, the low concentration of cell-free circulating DNA constitutes the biggest obstacle in the development of DNA methylation-based biomarkers from blood. This paper describes a method for the measurement of genomic methylation content directly on circulating tumor cells (CTC), which could be used to deceive the aforementioned problem. Since CTC are disease related blood-based biomarkers, they result essential to monitor tumor\\'s stadiation, therapy, and early relapsing lesions. Within surface\\'s bio-functionalization and cell\\'s isolation procedure standardization, the presented approach reveals a singular ability to detect high 5-methylcytosine CTC-subset content in the whole CTC compound, by choosing folic acid (FA) as transducer molecule. Sensitivity and specificity, calculated for FA functionalized surface (FA-surface), result respectively on about 83% and 60%. FA-surface, allowing the detection and characterization of early metastatic dissemination, provides a unique advance in the comprehension of tumors progression and dissemination confirming the presence of CTC and its association with high risk of relapse. This functionalized surface identifying and quantifying high 5-methylcytosine CTC-subset content into the patient\\'s blood lead significant progress in cancer risk assessment, also providing a novel therapeutic strategy.© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Comparison of biological characteristics of mesenchymal stem cells grown on two different titanium implant surfaces

    International Nuclear Information System (INIS)

    Wang Chengyue; Zhao Baohong; Ai Hongjun; Wang Yiwei

    2008-01-01

    This study examined the biological characteristics of mesenchymal stem cells (MSCs) grown on sand-blasted, large-grit, acid-etched (SLA) surface and hydroxyapatite (HA) coating on the SLA (HA/SLA) surface of titanium dental implants. The HA/SLA surfaces of titanium dental implants were formed by the ion beam assisted deposition (IBAD) method. Rabbit bone marrow derived mesenchymal stem cells cultured in vitro were seeded onto the surface of SLA and HA/SLA; the growth states of MSCs on the two samples were observed by a scanning electron microscope; the proliferation index, alkaline phosphatase (ALP) activity, osteocalcin (OCN) content of MSCs and mRNA relative expression level of osteopontin (opn) were compared between two groups. MSCs were found to be easier to adhere to the HA/SLA surface compared to the SLA surface. At the same time, the ALP activity and the OCN content of MSCs grown on the HA/SLA surface were obviously higher, and the relative expression level of opn mRNA was 4.78 times higher than that on the SLA surface. The HA coating formed by the IBAD method on the SLA surface of titanium dental implants significantly improves proliferation and well-differentiated osteoblastic phenotype of MSCs, which indicates a promising method for the surface modification of titanium dental implants

  3. Surface localization of the nuclear receptor CAR in influenza A virus-infected cells

    International Nuclear Information System (INIS)

    Takahashi, Tadanobu; Moriyama, Yusuke; Ikari, Akira; Sugatani, Junko; Suzuki, Takashi; Miwa, Masao

    2008-01-01

    Constitutive active/androstane receptor CAR is a member of the nuclear receptors which regulate transcription of xenobiotic metabolism enzymes. CAR is usually localized in the cytosol and nucleus. Here, we found that CAR was localized at the cell surface of influenza A virus (IAV)-infected cells. Additionally, we demonstrated that expression of a viral envelope glycoprotein, either hemagglutinin (HA) or neuraminidase (NA), but not viral nucleoprotein (NP), was responsible for this localization. This report is the first demonstration of CAR at the surface of tissue culture cells, and suggests that CAR may exert the IAV infection mechanism

  4. Cell-surface metalloprotease ADAM12 is internalized by a clathrin- and Grb2-dependent mechanism

    DEFF Research Database (Denmark)

    Hansen, Dorte Stautz; Leyme, Anthony; Grandal, Michael Vibo

    2012-01-01

    ADAM12 (A Disintegrin And Metalloprotease 12), a member of the ADAMs family of transmembrane proteins, is involved in ectodomain shedding, cell-adhesion and signaling, with important implications in cancer. Therefore, mechanisms that regulate the levels and activity of ADAM12 at the cell...... that regulates ADAM cell surface levels and show that ADAM12 internalization involves the clathrin-dependent pathway and Grb2.......-surface are possibly crucial in these contexts. We here investigated internalization and subsequent recycling or degradation of ADAM12 as a potentially important regulatory mechanism. Our results show that ADAM12 is constitutively internalized primarily via the clathrin-dependent pathway and is subsequently detected...

  5. Femtosecond laser-induced periodic nanostructure creation on PET surface for controlling of cell spreading

    Science.gov (United States)

    Sato, Yuji; Tsukamoto, Masahiro; Shinonaga, Togo; Kawa, Takuya

    2016-03-01

    A new method of periodic nanostructure formation on a polyethylene terephthalate (PET) surface has been developed, employing a femtosecond laser with a wavelength of 1045 nm. To generate structured films, the PET was placed in contact with a silicon (Si) wafer, followed by irradiation with the laser focused on the Si wafer, passing through the PET film. In order to evaluate the surface morphology, atomic force microscopy analysis was conducted on both treated and untreated PET surfaces. From the results, nanostructures with a period of 600 nm and height of 100 nm were formed on the PET film surface by laser treatment. A cell cultivation test was carried out on PET films with and without periodic nanostructures, showing that for nanostructured films, the cells (MG-63) were spread along the periodic grooves; in contrast, random cell spreading was observed for cultures grown on the untreated PET film.

  6. Pathogenic Acinetobacter: from the Cell Surface to Infinity and Beyond

    Science.gov (United States)

    Weber, Brent S.; Harding, Christian M.

    2015-01-01

    The genus Acinetobacter encompasses multiple nosocomial opportunistic pathogens that are of increasing worldwide relevance because of their ability to survive exposure to various antimicrobial and sterilization agents. Among these, Acinetobacter baumannii, Acinetobacter nosocomialis, and Acinetobacter pittii are the most frequently isolated in hospitals around the world. Despite the growing incidence of multidrug-resistant Acinetobacter spp., little is known about the factors that contribute to pathogenesis. New strategies for treating and managing infections caused by multidrug-resistant Acinetobacter strains are urgently needed, and this requires a detailed understanding of the pathobiology of these organisms. In recent years, some virulence factors important for Acinetobacter colonization have started to emerge. In this review, we focus on several recently described virulence factors that act at the bacterial surface level, such as the capsule, O-linked protein glycosylation, and adhesins. Furthermore, we describe the current knowledge regarding the type II and type VI secretion systems present in these strains. PMID:26712938

  7. Novel AgoshRNA molecules for silencing of the CCR5 co-receptor for HIV-1 infection.

    Science.gov (United States)

    Herrera-Carrillo, Elena; Berkhout, Ben

    2017-01-01

    Allogeneic transplantation of blood stem cells from a CCR5-Δ32 homozygous donor to an HIV-infected individual, the "Berlin patient", led to a cure. Since then there has been a search for approaches that mimic this intervention in a gene therapy setting. RNA interference (RNAi) has evolved as a powerful tool to regulate gene expression in a sequence-specific manner and can be used to inactivate the CCR5 mRNA. Short hairpin RNA (shRNA) molecules can impair CCR5 expression, but these molecules may cause unintended side effects and they will not be processed in cells that lack Dicer, such as monocytes. Dicer-independent RNAi pathways have opened opportunities for new AgoshRNA designs that rely exclusively on Ago2 for maturation. Furthermore, AgoshRNA processing yields a single active guide RNA, thus reducing off-target effects. In this study, we tested different AgoshRNA designs against CCR5. We selected AgoshRNAs that potently downregulated CCR5 expression on human T cells and peripheral blood mononuclear cells (PBMC) and that had no apparent adverse effect on T cell development as assessed in a competitive cell growth assay. CCR5 knockdown significantly protected T cells from CCR5 tropic HIV-1 infection.

  8. The influence of the surface chemistry of silver nanoparticles on cell death

    International Nuclear Information System (INIS)

    Sur, Ilknur; Altunbek, Mine; Kahraman, Mehmet; Culha, Mustafa

    2012-01-01

    The influence of the surface chemistry of silver nanoparticles (AgNPs) on p53 mediated cell death was evaluated using human dermal fibroblast (HDF) and lung cancer (A549) cells. The citrate reduced AgNPs (C-AgNPs) were modified with either lactose (L-AgNPs) or a 12-base long oligonucleotide (O-AgNPs). Both unmodified and modified AgNPs showed increased concentration and time dependent cytotoxicity and genotoxicity causing an increased p53 up-regulation within 6 h and led to apoptotic or necrotic cell deaths. The C-AgNPs induced more cytotoxicity and cellular DNA damage than the surface modified AgNPs. Modifying the C-AgNPs with lactose or the oligonucleotide reduced both necrotic and apoptotic cell deaths in the HDF cells. The C-AgNPs caused an insignificant necrosis in A549 cells whereas the modified AgNPs caused necrosis and apoptosis in both cell types. Compared to the O-AgNPs, the L-AgNPs triggered more cellular DNA damage, which led to up-regulation of p53 gene inducing apoptosis in A549 cells compared to HDF cells. This suggests that the different surface chemistries of the AgNPs cause different cellular responses that may be important not only for their use in medicine but also for reducing their toxicity. (paper)

  9. Facile method to stain the bacterial cell surface for super-resolution fluorescence microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Gunsolus, Ian L.; Hu, Dehong; Mihai, Cosmin; Lohse, Samuel E.; Lee, Chang-Soo; Torelli, Marco; Hamers, Robert J.; Murphy, Catherine; Orr, Galya; Haynes, Christy L.

    2014-01-01

    A method to fluorescently stain the surfaces of both Gram-negative and Gram-positive bacterial cells compatible with super-resolution fluorescence microscopy is presented. This method utilizes a commercially-available fluorescent probe to label primary amines at the surface of the cell. We demonstrate efficient staining of two bacterial strains, the Gram-negative Shewanella oneidensis MR-1 and the Gram-positive Bacillus subtilis 168. Using structured illumination microscopy and stochastic optical reconstruction microscopy, which require high quantum yield or specialized dyes, we show that this staining method may be used to resolve the bacterial cell surface with sub-diffraction-limited resolution. We further use this method to identify localization patterns of nanomaterials, specifically cadmium selenide quantum dots, following interaction with bacterial cells.

  10. AFM study of hippocampal cells cultured on silicon wafers with nano-scale surface topograph.

    Science.gov (United States)

    Ma, J; Liu, B F; Xu, Q Y; Cui, F Z

    2005-08-01

    The rat hippocampal cells were selected as model to study the interaction between the neural cells and silicon substrates using atomic force microscopy (AFM). The hippocampal cells show tight adherence on silicon wafers with nano-scale surface topograph. The lateral friction force investigated by AFM shows significant increase on the boundary around the cellular body. It is considered to relate to the cytoskeleton and cellular secretions. After ultrasonic wash in ethanol and acetone step by step, the surface of silicon wafers was observed by AFM sequentially. We have found that the culture leftovers form tight porous networks and a monolayer on the silicon wafers. It is concluded that the leftovers overspreading on the silicon substrates are the base of cell adherence on such smooth inert surfaces.

  11. Adhesion of neural cells on silicon wafer with nano-topographic surface

    Science.gov (United States)

    Fan, Y. W.; Cui, F. Z.; Chen, L. N.; Zhai, Y.; Xu, Q. Y.; Lee, I.-S.

    2002-02-01

    The adherence and subsequent viability of central neural cells (substantia nigra) on silicon wafers with different surface roughness conditions were investigated. Various roughness conditions of the silicon wafer were achieved by etching at different times. The topography was evaluated by AFM. Primary neurons were obtained from Wistar rats. The adherence and subsequent viability of the cells on the wafer were examined by scanning electronic microscopy and fluorescence immunostaining of tyrosine hydroxylase. It is found that the surface roughness affects significantly cell adhesion and viability. Cells can survive for over 5 days on the surface with average roughness in the range 20-70 nm. Such a treatment may provide a new method to make a mild interface of silicon-based electronic devices and neurons as well as other living tissues.

  12. Localized Surface Plasmons Enhanced Light Transmission into c-Silicon Solar Cells

    Directory of Open Access Journals (Sweden)

    Y. Premkumar Singh

    2013-01-01

    Full Text Available The paper investigates the light incoupling into c-Si solar cells due to the excitation of localized surface plasmon resonances in periodic metallic nanoparticles by finite-difference time-domain (FDTD technique. A significant enhancement of AM1.5G solar radiation transmission has been demonstrated by depositing nanoparticles of various metals on the upper surface of a semi-infinite Si substrate. Plasmonic nanostructures located close to the cell surface can scatter incident light efficiently into the cell. Al nanoparticles were found to be superior to Ag, Cu, and Au nanoparticles due to the improved transmission of light over almost the entire solar spectrum and, thus, can be a potential low-cost plasmonic metal for large-scale implementation of solar cells.

  13. Emergence of fractal geometry on the surface of human cervical epithelial cells during progression towards cancer

    International Nuclear Information System (INIS)

    Dokukin, M E; Sokolov, I; Guz, N V; Woodworth, C D

    2015-01-01

    Despite considerable advances in understanding the molecular nature of cancer, many biophysical aspects of malignant development are still unclear. Here we study physical alterations of the surface of human cervical epithelial cells during stepwise in vitro development of cancer (from normal to immortal (premalignant), to malignant). We use atomic force microscopy to demonstrate that development of cancer is associated with emergence of simple fractal geometry on the cell surface. Contrary to the previously expected correlation between cancer and fractals, we find that fractal geometry occurs only at a limited period of development when immortal cells become cancerous; further cancer progression demonstrates deviation from fractal. Because of the connection between fractal behaviour and chaos (or far from equilibrium behaviour), these results suggest that chaotic behaviour coincides with the cancer transformation of the immortalization stage of cancer development, whereas further cancer progression recovers determinism of processes responsible for cell surface formation. (paper)

  14. Comparative surface energetic study of Matrigel®and collagen I interactions with endothelial cells.

    Science.gov (United States)

    Hill, Michael J; Sarkar, Debanjan

    2017-07-01

    Understanding of the surface energetic aspects of the spontaneously deposited proteins on biomaterial surfaces and how this influences cell adhesion and differentiation is an area of regenerative medicine that has not received adequate attention. Current controversies surround the role of the biomaterial substratum surface chemistry, the range of influence of said substratum, and the effects of different surface energy components of the protein interface. Endothelial cells (ECs) are a highly important cell type for regenerative medicine applications, such as tissue engineering, and In-vivo they interact with collagen I based stromal tissue and basement membranes producing different behavioral outcomes. The surface energetic properties of these tissue types and how they control EC behavior is not well known. In this work we studied the surface energetic properties of collagen I and Matrigel ® on various previously characterized substratum polyurethanes (PU) via contact angle analysis and examined the subsequent EC network forming characteristics. A combinatorial surface energy approach was utilized that compared Zisman's critical surface tension, Kaelble's numerical method, and van Oss-Good-Chaudhury theory (vOGCT). We found that the unique, rapid network forming characteristics of ECs on Matrigel ® could be attributed to the apolar or monopolar basic interfacial characteristics according to Zisman/Kaelble or vOGCT, respectively. We also found a lack of significant substratum influence on EC network forming characteristics for Matrigel ® but collagen I showed a distinct influence where more apolar PU substrata tended to produce higher Lewis acid character collagen I interfaces which led to a greater interaction with ECs. Collagen I interfaces on more polar PU substrata lacked Lewis acid character and led to similar EC network characteristics as Matrigel ® . We hypothesized that bipolar character of the protein film favored cell-substratum over cell-cell adhesive

  15. Bong-Han Corpuscles as Possible Stem Cell Niches on the Organ-Surfaces

    Directory of Open Access Journals (Sweden)

    Min Su Kim

    2008-03-01

    Full Text Available Objectives : Showing that Bong-Han corpuscles(BHC are suppliers of the stem cells in adulthood, and the Bong-Han ducts(BHD are transportation routes of stem cells. Methods : BHC and BHD were obtained from the internal organ-surfaces of rats. The sliced BHC and BHD were immunostained with various stem cell markers. Extracellular matrices were also analyzed by immunohistochemistry. Result : The presence of mesenchymal stem cells was confirmed by the expression of Integrin beta 1, Collagen type 1 and Fibronectin. But CD54 was not expressed. The hematopoietic stem cell marker, Thy 1 was strongly expressed. BHDs showed Collagen type 1, Fibronectin, and vWF expression. Conclusion : Both hematopoietic and mesenchymal stem cell markers were expressed strongly in BHC similarly as in bone marrow. An endothelial cell marker(vWF demonstrated the possibility of the stem cell transportation routes of BHD.

  16. Cell and fiber attachment to demineralized dentin from normal root surfaces.

    Science.gov (United States)

    Hanes, P J; Polson, A M; Ladenheim, S

    1985-12-01

    The study assessed connective tissue and epithelial responses to dentin specimens (obtained from normal roots of human teeth) after surface demineralization. Rectangular dental specimens with opposite faces of root and pulpal dentin were prepared from beneath root surfaces covered by periodontal ligament. One-half of the specimens were treated with citric acid, pH 1, for 3 minutes, while the remainder served as untreated control specimens. Specimens were implanted vertically into incisional wounds on the dorsal surface of rats with one end of the implant protruding through the skin. Four specimens in each group were available 1, 3, 5 and 10 days after implantation. Histologic and histometric analyses included counts of adhering cells, evaluation of connective tissue fiber relationships and assessment of epithelial migration. Analyses within each group comparing root and pulpal surfaces showed no differences between any of the parameters. Comparisons between experimental and control groups showed that demineralized surfaces had a greater number of cells attached, fiber attachment occurred and epithelial downgrowth was inhibited. The fiber attachment to experimental specimens differed morphologically from fiber attachment to normal root surfaces: the number of fibers attached per unit length and the diameter of attached fibers were significantly less on experimental specimens. Demineralized specimens at 10 days had a distinct eosinophilic surface zone. Surface demineralization of dentin predisposed toward a cell and fiber attachment system which inhibited migration of epithelium.

  17. The surface nanostructures of titanium alloy regulate the proliferation of endothelial cells

    Directory of Open Access Journals (Sweden)

    Min Lai

    2014-02-01

    Full Text Available To investigate the effect of surface nanostructures on the behaviors of human umbilical vein endothelial cells (HUVECs, surface nanostructured titanium alloy (Ti-3Zr2Sn-3Mo-25Nb, TLM was fabricated by surface mechanical attrition treatment (SMAT technique. Field emission scanning electron microscopy (FE-SEM, atomic force microscopy (AFM, transmission electron microscopy (TEM and X-ray diffraction (XRD were employed to characterize the surface nanostructures of the TLM, respectively. The results demonstrated that nano-crystalline structures with several tens of nanometers were formed on the surface of TLM substrates. The HUVECs grown onto the surface nanostructured TLM spread well and expressed more vinculin around the edges of cells. More importantly, HUVECs grown onto the surface nanostructured TLM displayed significantly higher (p < 0.01 or p < 0.05 cell adhesion and viabilities than those of native titanium alloy. HUVECs cultured on the surface nanostructured titanium alloy displayed significantly higher (p < 0.01 or p < 0.05 productions of nitric oxide (NO and prostacyclin (PGI2 than those of native titanium alloy, respectively. This study provides an alternative for the development of titanium alloy based vascular stents.

  18. Effect of surface organic coatings of cellulose nanocrystals on the viability of mammalian cell lines

    Directory of Open Access Journals (Sweden)

    Jimenez AS

    2017-09-01

    Full Text Available Ambar S Jimenez,1 Francesca Jaramillo,1 Usha D Hemraz,2 Yaman Boluk,3 Karina Ckless,1 Rajesh Sunasee1 1Department of Chemistry, State University of New York at Plattsburgh, Plattsburgh, NY, USA; 2National Research Council, Montreal, QC, Canada, 3Department of Civil & Environmental Engineering, University of Alberta and National Institute for Nanotechnology, National Research Council, Edmonton, AB, Canada Abstract: Cellulose nanocrystals (CNCs have emerged as promising candidates for a number of bio-applications. Surface modification of CNCs continues to gain significant research interest as it imparts new properties to the surface of the nanocrystals for the design of multifunctional CNCs-based materials. A small chemical surface modification can potentially lead to drastic behavioral changes of cell-material interactions thereby affecting the intended bio-application. In this work, unmodified CNCs were covalently decorated with four different organic moieties such as a diaminobutane fragment, a cyclic oligosaccharide (β-cyclodextrin, a thermoresponsive polymer (poly[N-isopropylacrylamide], and a cationic aminomethacrylamide-based polymer using different synthetic covalent methods. The effect of surface coatings of CNCs and the respective dose-response of the above organic moieties on the cell viability were evaluated on mammalian cell cultures (J774A.1 and MFC-7, using 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assays. Overall, the results indicated that cells exposed to surface-coated CNCs for 24 h did not display major changes in cell viability, membrane permeability as well as cell morphology. However, with longer exposure, all these parameters were somewhat affected, which appears not to be correlated with either anionic or cationic surface coatings of CNCs used in this study. Keywords: cellulose nanocrystals, surface coating, cell viability, MTT, LDH

  19. 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.

  20. Efficient Capture of Cancer Cells by Their Replicated Surfaces Reveals Multiscale Topographic Interactions Coupled with Molecular Recognition.

    Science.gov (United States)

    Wang, Wenshuo; Cui, Haijun; Zhang, Pengchao; Meng, Jingxin; Zhang, Feilong; Wang, Shutao

    2017-03-29

    Cell-surface topographic interactions can direct the design of biointerfaces, which have been widely used in isolation of circulating tumor cells or fundamental cell biological research. By using three kinds of cancer cell-replicated surfaces with differentiated structures, we uncover that multiscale-cooperative topographic interactions (at both nanoscale and microscale) coupled with molecular recognition enable efficient and specific isolation of cancer cells. The cell replicas precisely inherit the structural features from the original cancer cells, providing not only preferable structures for matching with cancer cells but also a unique platform to interrogate whether certain cancer cells can optimally match with their own replicated surfaces. The results reveal that cancer cells do not show preferential recognitions to their respective replicas, while the capture agent-modified surfaces with hierarchical structures exhibit improved cancer cell capture efficiencies. Two levels of topographic interactions between cancer cells and cell replica surfaces exist. Nanoscale filopodia on cancer cells can topographically interact with different nanostructures on replica surfaces. In addition, microscale concave/convex on surfaces provide suitable sites for trapping cancer cells. This study may promote smart design of multiscale biofunctional materials that can specifically recognize cancer cells.

  1. Surface Markers for Chondrogenic Determination: A Highlight of Synovium-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Douglas D. Campbell

    2012-11-01

    Full Text Available Cartilage tissue engineering is a promising field in regenerative medicine that can provide substantial relief to people suffering from degenerative cartilage disease. Current research shows the greatest chondrogenic potential for healthy articular cartilage growth with minimal hypertrophic differentiation to be from mesenchymal stem cells (MSCs of synovial origin. These stem cells have the capacity for differentiation into multiple cell lineages related to mesenchymal tissue; however, evidence exists for cell surface markers that specify a greater potential for chondrogenesis than other differentiation fates. This review will examine relevant literature to summarize the chondrogenic differentiation capacities of tested synovium-derived stem cell (SDSC surface markers, along with a discussion about various other markers that may hold potential, yet require further investigation. With this information, a potential clinical benefit exists to develop a screening system for SDSCs that will produce the healthiest articular cartilage possible.

  2. Cell-surface interactions involving immobilized magnetite nanoparticles on flat magnetic substrates.

    Science.gov (United States)

    Loichen, Juliane; Hartmann, Uwe

    2009-09-01

    A new method to affect cells by cell-surface interaction is introduced. Biocompatible magnetic nanobeads are deposited onto a biocompatible magnetic thin layer. The particles are composed of small magnetite crystals embedded in a matrix which can be functionalized by different molecules, proteins or growth factors. The magnetic interaction between surface and beads prevents endocytosis if the setup is utilized for cell culturing. The force acting between particles and magnetic layer is calculated by a magnetostatic approach. Biocompatibility is ensured by using garnet layers which turned out to be nontoxic and stable under culturing conditions. The garnet thin films exhibit spatially and temporally variable magnetic domain configurations in changing external magnetic fields and depending on their thermal pretreatment. Several patterns and bead deposition methods as well as the cell-surface interactions were analyzed. In some cases the cells show directed growth. Theoretical considerations explaining particular cell behavior on this magnetic material involve calculations of cell growth on elastic substrates and bending of cell membranes.

  3. Longitudinal microarray analysis of cell surface antigens on peripheral blood mononuclear cells from HIV+ individuals on highly active antiretroviral therapy

    Directory of Open Access Journals (Sweden)

    Wang Bin

    2008-03-01

    Full Text Available Abstract Background The efficacy of highly active antiretroviral therapy (HAART determined by simultaneous monitoring over 100 cell-surface antigens overtime has not been attempted. We used an antibody microarray to analyze changes in the expression of 135 different cell-surface antigens overtime on PBMC from HIV+ patients on HAART. Two groups were chosen, one (n = 6 achieved sustainable response by maintaining below detectable plasma viremia and the other (n = 6 responded intermittently. Blood samples were collected over an average of 3 years and 5–8 time points were selected for microarray assay and statistical analysis. Results Significant trends over time were observed for the expression of 7 cell surface antigens (CD2, CD3epsilon, CD5, CD95, CD36, CD27 and CD28 for combined patient groups. Between groups, expression levels of 10 cell surface antigens (CD11a, CD29, CD38, CD45RO, CD52, CD56, CD57, CD62E, CD64 and CD33 were found to be differential. Expression levels of CD9, CD11a, CD27, CD28 and CD52, CD44, CD49d, CD49e, CD11c strongly correlated with CD4+ and CD8+ T cell counts, respectively. Conclusion Our findings not only detected markers that may have potential prognostic/diagnostic values in evaluating HAART efficacy, but also showed how density of cell surface antigens could be efficiently exploited in an array-like manner in relation to HAART and HIV-infection. The antigens identified in this study should be further investigated by other methods such as flow cytometry for confirmation as biological analysis of these antigens may help further clarify their role during HAART and HIV infection.

  4. Modified titanium surface with gelatin nano gold composite increases osteoblast cell biocompatibility

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Young-Hee; Bhattarai, Govinda [Department of Oral Biochemistry, School of Dentistry and Institute of Oral Bioscience, BK21 program, Chonbuk National University, Jeonju (Korea, Republic of); Aryal, Santosh [Department of Bionanosystem Engineering, Chonbuk National University, Jeonju (Korea, Republic of); Lee, Nan-Hee [Department of Oral Biochemistry, School of Dentistry and Institute of Oral Bioscience, BK21 program, Chonbuk National University, Jeonju (Korea, Republic of); Lee, Min-Ho [Department of Dental Biomaterials, School of Dentistry and Institute of Oral Bioscience, BK21 program, Chonbuk National University, Jeonju (Korea, Republic of); Kim, Tae-Gun [Department of Conservative Dentistry, School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Jhee, Eun-Chung [Department of Oral Biochemistry, School of Dentistry and Institute of Oral Bioscience, BK21 program, Chonbuk National University, Jeonju (Korea, Republic of); Kim, Hak-Yong [Department of Bionanosystem Engineering, Chonbuk National University, Jeonju (Korea, Republic of); Yi, Ho-Keun, E-mail: yihokn@chonbuk.ac.kr [Department of Oral Biochemistry, School of Dentistry and Institute of Oral Bioscience, BK21 program, Chonbuk National University, Jeonju (Korea, Republic of)

    2010-08-01

    This study examined the gelatin nano gold (GnG) composite for surface modification of titanium in addition to insure biocompatibility on dental implants or biomaterials. The GnG composite was constructed by gelatin and hydrogen tetrachloroaurate in presence of reducing agent, sodium borohydrate (NabH{sub 4}). The GnG composite was confirmed by UV-VIS spectroscopy and transmission electron microscopy (TEM). A dipping method was used to modify the titanium surface by GnG composite. Surface was characterized by scanning electron microscopy (SEM) and energy dispersive X-ray (EDX). The MC-3T3 E1 cell viability was assessed by trypan blue and the expression of proteins to biocompatibility were analyzed by Western blotting. The GnG composite showed well dispersed character, the strong absorption at 530 nm, roughness, regular crystal and clear C, Na, Cl, P, and Au signals onto titanium. Further, this composite allowed MC-3T3 E1 growth and viability compared to gelatin and pure titanium. It induced ERK activation and the expression of cell adherent molecules, FAK and SPARC, and growth factor, VEGF. However, GnG decreased the level of SAPK/JNK. This shows that GnG composite coated titanium surfaces have a good biocompatibility for osteoblast growth and attachment than in intact by simple and versatile dipping method. Furthermore, it offers good communication between cell and implant surfaces by regulating cell signaling and adherent molecules, which are useful to enhance the biocompatibility of titanium surfaces.

  5. Modified titanium surface with gelatin nano gold composite increases osteoblast cell biocompatibility

    Science.gov (United States)

    Lee, Young-Hee; Bhattarai, Govinda; Aryal, Santosh; Lee, Nan-Hee; Lee, Min-Ho; Kim, Tae-Gun; Jhee, Eun-Chung; Kim, Hak-Yong; Yi, Ho-Keun

    2010-08-01

    This study examined the gelatin nano gold (GnG) composite for surface modification of titanium in addition to insure biocompatibility on dental implants or biomaterials. The GnG composite was constructed by gelatin and hydrogen tetrachloroaurate in presence of reducing agent, sodium borohydrate (NabH 4). The GnG composite was confirmed by UV-VIS spectroscopy and transmission electron microscopy (TEM). A dipping method was used to modify the titanium surface by GnG composite. Surface was characterized by scanning electron microscopy (SEM) and energy dispersive X-ray (EDX). The MC-3T3 E1 cell viability was assessed by trypan blue and the expression of proteins to biocompatibility were analyzed by Western blotting. The GnG composite showed well dispersed character, the strong absorption at 530 nm, roughness, regular crystal and clear C, Na, Cl, P, and Au signals onto titanium. Further, this composite allowed MC-3T3 E1 growth and viability compared to gelatin and pure titanium. It induced ERK activation and the expression of cell adherent molecules, FAK and SPARC, and growth factor, VEGF. However, GnG decreased the level of SAPK/JNK. This shows that GnG composite coated titanium surfaces have a good biocompatibility for osteoblast growth and attachment than in intact by simple and versatile dipping method. Furthermore, it offers good communication between cell and implant surfaces by regulating cell signaling and adherent molecules, which are useful to enhance the biocompatibility of titanium surfaces.

  6. Enhanced adhesion of osteoblastic cells on polystyrene films by independent control of surface topography and wettability

    International Nuclear Information System (INIS)

    Yang, Seung Yun; Kim, Eung-Sam; Jeon, Gumhye; Choi, Kwan Yong; Kim, Jin Kon

    2013-01-01

    We independently controlled surface topography and wettability of polystyrene (PS) films by CF 4 and oxygen plasma treatments, respectively, to evaluate the adhesion and proliferation of human fetal osteoblastic (hFOB) cells on the films. Among the CF 4 plasma-treated PS films with the average surface roughness ranging from 0.9 to 70 nm, the highest adhesion of hFOB cells was observed on a PS film with roughness of ∼ 11 nm. When this film was additionally treated by oxygen plasma to provide a hydrophilic surface with a contact angle less than 10°, the proliferation of bone-forming cell was further enhanced. Thus, the plasma-based independent modification of PS film into an optimum nanotexture for human osteoblast cells could be appplied to materials used in bone tissue engineering. Highlights: ► New approach based on plasma treatment to independently control the surface topography and wettability ► The adhesion of human fetal osteoblast (hFOB) was enhanced on a surface with an average roughness of ∼ 11 nm. ► The adhesion and proliferation of hFOB was maximized when nanotextured surface became highly hydrophilic

  7. Cell Surface and Membrane Engineering: Emerging Technologies and Applications

    Directory of Open Access Journals (Sweden)

    Christopher T. Saeui

    2015-06-01

    Full Text Available Membranes constitute the interface between the basic unit of life—a single cell—and the outside environment and thus in many ways comprise the ultimate “functional biomaterial”. To perform the many and often conflicting functions required in this role, for example to partition intracellular contents from the outside environment while maintaining rapid intake of nutrients and efflux of waste products, biological membranes have evolved tremendous complexity and versatility. This article describes how membranes, mainly in the context of living cells, are increasingly being manipulated for practical purposes with drug discovery, biofuels, and biosensors providing specific, illustrative examples. Attention is also given to biology-inspired, but completely synthetic, membrane-based technologies that are being enabled by emerging methods such as bio-3D printers. The diverse set of applications covered in this article are intended to illustrate how these versatile technologies—as they rapidly mature—hold tremendous promise to benefit human health in numerous ways ranging from the development of new medicines to sensitive and cost-effective environmental monitoring for pathogens and pollutants to replacing hydrocarbon-based fossil fuels.

  8. Cells transplanted onto the surface of the glial scar reveal hidden potential for functional neural regeneration.

    Science.gov (United States)

    Sekiya, Tetsuji; Holley, Matthew C; Hashido, Kento; Ono, Kazuya; Shimomura, Koichiro; Horie, Rie T; Hamaguchi, Kiyomi; Yoshida, Atsuhiro; Sakamoto, Tatsunori; Ito, Juichi

    2015-06-30

    Cell transplantation therapy has long been investigated as a therapeutic intervention for neurodegenerative disorders, including spinal cord injury, Parkinson's disease, and amyotrophic lateral sclerosis. Indeed, patients have high hopes for a cell-based therapy. However, there are numerous practical challenges for clinical translation. One major problem is that only very low numbers of donor cells survive and achieve functional integration into the host. Glial scar tissue in chronic neurodegenerative disorders strongly inhibits regeneration, and this inhibition must be overcome to accomplish successful cell transplantation. Intraneural cell transplantation is considered to be the best way to deliver cells to the host. We questioned this view with experiments in vivo on a rat glial scar model of the auditory system. Our results show that intraneural transplantation to the auditory nerve, preceded by chondroitinase ABC (ChABC)-treatment, is ineffective. There is no functional recovery, and almost all transplanted cells die within a few weeks. However, when donor cells are placed on the surface of a ChABC-treated gliotic auditory nerve, they autonomously migrate into it and recapitulate glia- and neuron-guided cell migration modes to repair the auditory pathway and recover auditory function. Surface transplantation may thus pave the way for improved functional integration of donor cells into host tissue, providing a less invasive approach to rescue clinically important neural tracts.

  9. Altered expression of epithelial cell surface glycoconjugates and intermediate filaments at the margins of mucosal wounds

    DEFF Research Database (Denmark)

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

    1998-01-01

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

  10. Automatic analysis of image of surface structure of cell wall-deficient EVC.

    Science.gov (United States)

    Li, S; Hu, K; Cai, N; Su, W; Xiong, H; Lou, Z; Lin, T; Hu, Y

    2001-01-01

    Some computer applications for cell characterization in medicine and biology, such as analysis of surface structure of cell wall-deficient EVC (El Tor Vibrio of Cholera), operate with cell samples taken from very small areas of interest. In order to perform texture characterization in such an application, only a few texture operators can be employed: the operators should be insensitive to noise and image distortion and be reliable in order to estimate texture quality from images. Therefore, we introduce wavelet theory and mathematical morphology to analyse the cellular surface micro-area image obtained by SEM (Scanning Electron Microscope). In order to describe the quality of surface structure of cell wall-deficient EVC, we propose a fully automatic computerized method. The image analysis process is carried out in two steps. In the first, we decompose the given image by dyadic wavelet transform and form an image approximation with higher resolution, by doing so, we perform edge detection of given images efficiently. In the second, we introduce many operations of mathematical morphology to obtain morphological quantitative parameters of surface structure of cell wall-deficient EVC. The obtained results prove that the method can eliminate noise, detect the edge and extract the feature parameters validly. In this work, we have built automatic analytic software named "EVC.CELL".

  11. A Comparative Study of the Therapeutic Potential of Mesenchymal Stem Cells and Limbal Epithelial Stem Cells for Ocular Surface Reconstruction

    Czech Academy of Sciences Publication Activity Database

    Holáň, Vladimír; Trošan, Peter; Čejka, Čestmír; Javorková, Eliška; Zajícová, Alena; Heřmánková, Barbora; Chudíčková, Milada; Čejková, Jitka

    2015-01-01

    Roč. 4, č. 9 (2015), s. 1052-1063 ISSN 2157-6564 R&D Projects: GA ČR(CZ) GA14-12580S; GA MZd NT14102; GA MŠk(CZ) LO1309; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:68378041 Keywords : limbal stem cells * mesenchymal stem cells * alkali-injured ocular surface * corneal regeneration * stem cell-based therapy Subject RIV: FF - HEENT, Dentistry Impact factor: 4.247, year: 2015

  12. Immobilization of microbial cells on cellulose-polymer surfaces by radiation polymerization

    International Nuclear Information System (INIS)

    Kumakura, M.; Kaetsu, I.

    1983-01-01

    Streptomyces phaeochromogens cells were immobilized on cellulose-polymer surfaces by radiation polymerization using hydrophilic monomers and paper. The enzyme activity of immobilized cell sheets was higher than that of immobilized cell composites obtained by the usual radiation polymerization technique. The enzyme activity of the sheets was affected by monomer concentration, the thickness of paper, and the degree of polymerization of paper. The copolymerization of hydroxyethyl methacrylate and methoxytetraethyleneglycol methacrylate in the sheets led to a further increase of the enzyme activity due to the increase of the hydrophilicity of the polymer matrix. The Michaelis constant of the sheets from low monomer concentration was close to that of intact cells

  13. Surface chemistry and acid-base activity of Shewanella putrefaciens: Cell wall charging and metal binding to bacterial cell walls

    NARCIS (Netherlands)

    Claessens, Jacqueline Wilhelmien

    2006-01-01

    To gain insight into the surface chemistry of live microorganisms, pH stat experiments are combined with analyses of the time-dependent changes in solution chemistry using suspensions of live cells of Shewanella putrefaciens. The results of this study illustrate the complex response of the live

  14. Surface chemistry and acid-base activity of Shewanella putrefaciens : Cell wall charging and metal binding to bacterial cell walls

    NARCIS (Netherlands)

    Claessens, J.W.

    2006-01-01

    To gain insight into the surface chemistry of live microorganisms, pH stat experiments are combined with analyses of the time-dependent changes in solution chemistry using suspensions of live cells of Shewanella putrefaciens. The results of this study illustrate the complex response of the live

  15. Bleb Expansion in Migrating Cells Depends on Supply of Membrane from Cell Surface Invaginations.

    Science.gov (United States)

    Goudarzi, Mohammad; Tarbashevich, Katsiaryna; Mildner, Karina; Begemann, Isabell; Garcia, Jamie; Paksa, Azadeh; Reichman-Fried, Michal; Mahabaleshwar, Harsha; Blaser, Heiko; Hartwig, Johannes; Zeuschner, Dagmar; Galic, Milos; Bagnat, Michel; Betz, Timo; Raz, Erez

    2017-12-04

    Cell migration is essential for morphogenesis, organ formation, and homeostasis, with relevance for clinical conditions. The migration of primordial germ cells (PGCs) is a useful model for studying this process in the context of the developing embryo. Zebrafish PGC migration depends on the formation of cellular protrusions in form of blebs, a type of protrusion found in various cell types. Here we report on the mechanisms allowing the inflation of the membrane during bleb formation. We show that the rapid expansion of the protrusion depends on membrane invaginations that are localized preferentially at the cell front. The formation of these invaginations requires the function of Cdc42, and their unfolding allows bleb inflation and dynamic cell-shape changes performed by migrating cells. Inhibiting the formation and release of the invaginations strongly interfered with bleb formation, cell motility, and the ability of the cells to reach their target. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Novel AgoshRNA molecules for silencing of the CCR5 co-receptor for HIV-1 infection

    NARCIS (Netherlands)

    Herrera-Carrillo, Elena; Berkhout, Ben

    2017-01-01

    Allogeneic transplantation of blood stem cells from a CCR5-Δ32 homozygous donor to an HIV-infected individual, the "Berlin patient", led to a cure. Since then there has been a search for approaches that mimic this intervention in a gene therapy setting. RNA interference (RNAi) has evolved as a

  17. Behavior of osteoblastic cells cultured on titanium and structured zirconia surfaces

    Science.gov (United States)

    Depprich, Rita; Ommerborn, Michelle; Zipprich, Holger; Naujoks, Christian; Handschel, Jörg; Wiesmann, Hans-Peter; Kübler, Norbert R; Meyer, Ulrich

    2008-01-01

    Background Osseointegration is crucial for the long-term success of dental implants and depends on the tissue reaction at the tissue-implant interface. Mechanical properties and biocompatibility make zirconia a suitable material for dental implants, although surface processings are still problematic. The aim of the present study was to compare osteoblast behavior on structured zirconia and titanium surfaces under standardized conditions. Methods The surface characteristics were determined by scanning electron microscopy (SEM). In primary bovine osteoblasts attachment kinetics, proliferation rate and synthesis of bone-associated proteins were tested on different surfaces. Results The results demonstrated that the proliferation rate of cells was significantly higher on zirconia surfaces than on titanium surfaces (p zirconia and titanium surfaces. Conclusion The study demonstrates distinct effects of the surface composition on osteoblasts in culture. Zirconia improves cell proliferation significantly during the first days of culture, but it does not improve attachment and adhesion strength. Both materials do not differ with respect to protein synthesis or ultrastructural appearance of osteoblasts. Zirconium oxide may therefore be a suitable material for dental implants. PMID:19063728

  18. Ocular surface reconstruction with a tissue-engineered nasal mucosal epithelial cell sheet for the treatment of severe ocular surface diseases.

    Science.gov (United States)

    Kobayashi, Masakazu; Nakamura, Takahiro; Yasuda, Makoto; Hata, Yuiko; Okura, Shoki; Iwamoto, Miyu; Nagata, Maho; Fullwood, Nigel J; Koizumi, Noriko; Hisa, Yasuo; Kinoshita, Shigeru

    2015-01-01

    Severe ocular surface diseases (OSDs) with severe dry eye can be devastating and are currently some of the most challenging eye disorders to treat. To investigate the feasibility of using an autologous tissue-engineered cultivated nasal mucosal epithelial cell sheet (CNMES) for ocular surface reconstruction, we developed a novel technique for the culture of nasal mucosal epithelial cells expanded ex vivo from biopsy-derived human nasal mucosal tissues. After the protocol, the CNMESs had 4-5 layers of stratified, well-differentiated cells, and we successfully generated cultured epithelial sheets, including numerous goblet cells. Immunohistochemistry confirmed the presence of keratins 3, 4, and 13; mucins 1, 16, and 5AC; cell junction and basement membrane assembly proteins; and stem/progenitor cell marker p75 in the CNMESs. We then transplanted the CNMESs onto the ocular surfaces of rabbits and confirmed the survival of this tissue, including the goblet cells, up to 2 weeks. The present report describes an attempt to overcome the problems of treating severe OSDs with the most severe dry eye by treating them using tissue-engineered CNMESs to supply functional goblet cells and to stabilize and reconstruct the ocular surface. The present study is a first step toward assessing the use of tissue-engineered goblet-cell transplantation of nonocular surface origin for ocular surface reconstruction. ©AlphaMed Press.

  19. Measurement of surface recombination velocity for silicon solar cells using a scanning electron microscope with pulsed beam

    Science.gov (United States)

    Daud, T.; Cheng, L. J.

    1981-01-01

    The role of surface recombination velocity in the design and fabrication of silicon solar cells is discussed. A scanning electron microscope with pulsed electron beam was used to measure this parameter of silicon surfaces. It is shown that the surface recombination velocity, s, increases by an order of magnitude when an etched surface degrades, probably as a result of environmental reaction. A textured front-surface-field cell with a high-low junction near the surface shows the effect of minority carrier reflection and an apparent reduction of s, whereas a tandem-junction cell shows an increasing s value. Electric fields at junction interfaces in front-surface-field and tandem-junction cells acting as minority carrier reflectors or sinks tend to alter the value of effective surface recombination velocity for different beam penetration depths. A range of values of s was calculated for different surfaces.

  20. The Control of Mesenchymal Stromal Cell Osteogenic Differentiation through Modified Surfaces

    Directory of Open Access Journals (Sweden)

    Niall Logan

    2013-01-01

    Full Text Available Stem cells continue to receive widespread attention due to their potential to revolutionise treatments in the fields of both tissue engineering and regenerative medicine. Adult stem cells, specifically mesenchymal stromal cells (MSCs, play a vital role in the natural events surrounding bone healing and osseointegration through being stimulated to differentiate along their osteogenic lineage and in doing so, they form new cortical and trabecular bone tissue. Understanding how to control, manipulate, and enhance the intrinsic healing events modulated through osteogenic differentiation of MSCs by the use of modified surfaces and biomaterials could potentially advance the fields of both orthopaedics and dentistry. This could be by either using surface modification to generate greater implant stability and more rapid healing following implantation or the stimulation of MSCs ex vivo for reimplantation. This review aims to gather publications targeted at promoting, enhancing, and controlling the osteogenic differentiation of MSCs through biomaterials, nanotopographies, and modified surfaces for use in implant procedures.

  1. Studying cell-surface interactions in vitro: a survey of experimental approaches and techniques.

    Science.gov (United States)

    Michaelis, Stefanie; Robelek, Rudolf; Wegener, Joachim

    2012-01-01

    A better understanding of the interactions of animal (or human) cells with in vitro surfaces is the key to the successful development, improvement and optimization of biomaterials for biomedical or biotechnological purposes. State-of-the-art experimental approaches and techniques are a prerequisite for further and deeper insights into the mechanisms and processes involved in cell-surface adhesion. This chapter provides a brief but not complete survey of optical, mechanical, electrochemical and acoustic devices that are currently used to study the structural and functional properties of the cell-surface junction. Each technique is introduced with respect to the underlying principles before example data are discussed. At the end of the chapter all techniques are compared in terms of their strengths, limitations and technical requirements.

  2. Annexin A8 controls leukocyte recruitment to activated endothelial cells via cell surface delivery of CD63

    Science.gov (United States)

    Poeter, Michaela; Brandherm, Ines; Rossaint, Jan; Rosso, Gonzalo; Shahin, Victor; Skryabin, Boris V.; Zarbock, Alexander; Gerke, Volker; Rescher, Ursula

    2014-04-01

    To enable leukocyte adhesion to activated endothelium, the leukocyte receptor P-selectin is released from Weibel-Palade bodies (WPB) to the endothelial cell surface where it is stabilized by CD63. Here we report that loss of annexin A8 (anxA8) in human umbilical vein endothelial cells (HUVEC) strongly decreases cell surface presentation of CD63 and P-selectin, with a concomitant reduction in leukocyte rolling and adhesion. We confirm the compromised leukocyte adhesiveness in inflammatory-activated endothelial venules of anxA8-deficient mice. We find that WPB of anxA8-deficient HUVEC contain less CD63, and that this is caused by improper transport of CD63 from late multivesicular endosomes to WPB, with CD63 being retained in intraluminal vesicles. Consequently, reduced CD63 cell surface levels are seen following WPB exocytosis, resulting in enhanced P-selectin re-internalization. Our data support a model in which anxA8 affects leukocyte recruitment to activated endothelial cells by supplying WPB with sufficient amounts of the P-selectin regulator CD63.

  3. Regulating Immunogenicity and Tolerogenicity of Bone Marrow-Derived Dendritic Cells through Modulation of Cell Surface Glycosylation by Dexamethasone Treatment

    Directory of Open Access Journals (Sweden)

    Kevin Lynch

    2017-10-01

    Full Text Available Dendritic cellular therapies and dendritic cell vaccines show promise for the treatment of autoimmune diseases, the prolongation of graft survival in transplantation, and in educating the immune system to fight cancers. Cell surface glycosylation plays a crucial role in the cell–cell interaction, uptake of antigens, migration, and homing of DCs. Glycosylation is known to change with environment and the functional state of DCs. Tolerogenic DCs (tDCs are commonly generated using corticosteroids including dexamethasone, however, to date, little is known on how corticosteroid treatment alters glycosylation and what functional consequences this may have. Here, we present a comprehensive profile of rat bone marrow-derived dendritic cells, examining their cell surface glycosylation profile before and after Dexa treatment as resolved by both lectin microarrays and lectin-coupled flow cytometry. We further examine the functional consequences of altering cell surface glycosylation on immunogenicity and tolerogenicity of DCs. Dexa treatment of rat DCs leads to profoundly reduced expression of markers of immunogenicity (MHC I/II, CD80, CD86 and pro-inflammatory molecules (IL-6, IL-12p40, inducible nitric oxide synthase indicating a tolerogenic phenotype. Moreover, by comprehensive lectin microarray profiling and flow cytometry analysis, we show that sialic acid (Sia is significantly upregulated on tDCs after Dexa treatment, and that this may play a vital role in the therapeutic attributes of these cells. Interestingly, removal of Sia by neuraminidase treatment increases the immunogenicity of immature DCs and also leads to increased expression of pro-inflammatory cytokines while tDCs are moderately protected from this increase in immunogenicity. These findings may have important implications in strategies aimed at increasing tolerogenicity where it is advantageous to reduce immune activation over prolonged periods. These findings are also relevant in

  4. Cell fitting to adhesive surfaces: A prerequisite to firm attachment and subsequent events

    Directory of Open Access Journals (Sweden)

    Pierres A.

    2002-06-01

    Full Text Available Cell adhesion usually involves extensive shape reorganization. This process is important because i it is required for efficient cross-linking of interacting surfaces by adhesion receptors the length of which does not exceed several tens of nanometers and ii it influences subsequent cell differentiation and activation. This review focuses on the initial phase of cell deformation, preceding the extensive reorganization process known as spreading. This first phase includes local flattening at the micrometer scale and membrane alignment at the nanometer level, resulting in fitting of the cell to an adhesive surface. Three main points are considered. First, experimental methods available to study cell apposition to a surface are described, with an emphasis on interference reflection microscopy. Second, selected experimental evidence is presented to show that there is a quantitative relationship between "adhesiveness" and "contact extension", and some theoretical models aimed at relating these parameters are briefly sketched. Third, experimental data on the kinetics of initial contact extension are described and possible mechanisms for driving this extension are discussed, including nonspecific forces, receptor-mediated interactions, active cell movements or passive membrane fluctuations. It is concluded that both passive physical phenomena and random active cell movements are possible candidates for the initial triggering of contact extension.

  5. Identification of polymer surface adsorbed proteins implicated in pluripotent human embryonic stem cell expansion.

    Science.gov (United States)

    Hammad, Moamen; Rao, Wei; Smith, James G W; Anderson, Daniel G; Langer, Robert; Young, Lorraine E; Barrett, David A; Davies, Martyn C; Denning, Chris; Alexander, Morgan R

    2016-08-16

    Improved biomaterials are required for application in regenerative medicine, biosensing, and as medical devices. The response of cells to the chemistry of polymers cultured in media is generally regarded as being dominated by proteins adsorbed to the surface. Here we use mass spectrometry to identify proteins adsorbed from a complex mouse embryonic fibroblast (MEF) conditioned medium found to support pluripotent human embryonic stem cell (hESC) expansion on a plasma etched tissue culture polystyrene surface. A total of 71 proteins were identified, of which 14 uniquely correlated with the surface on which pluripotent stem cell expansion was achieved. We have developed a microarray combinatorial protein spotting approach to test the potential of these 14 proteins to support expansion of a hESC cell line (HUES-7) and a human induced pluripotent stem cell line (ReBl-PAT) on a novel polymer (N-(4-Hydroxyphenyl) methacrylamide). These proteins were spotted to form a primary array yielding several protein mixture 'hits' that enhanced cell attachment to the polymer. A second array was generated to test the function of a refined set of protein mixtures. We found that a combination of heat shock protein 90 and heat shock protein-1 encourage elevated adherence of pluripotent stem cells at a level comparable to fibronectin pre-treatment.

  6. Cell surface protein disulfide isomerase regulates natriuretic peptide generation of cyclic guanosine monophosphate.

    Directory of Open Access Journals (Sweden)

    Shuchong Pan

    Full Text Available The family of natriuretic peptides (NPs, including atrial natriuretic peptide (ANP, B-type natriuretic peptide (BNP, and C-type natriuretic peptide (CNP, exert important and diverse actions for cardiovascular and renal homeostasis. The autocrine and paracrine functions of the NPs are primarily mediated through the cellular membrane bound guanylyl cyclase-linked receptors GC-A (NPR-A and GC-B (NPR-B. As the ligands and receptors each contain disulfide bonds, a regulatory role for the cell surface protein disulfide isomerase (PDI was investigated.We utilized complementary in vitro and in vivo models to determine the potential role of PDI in regulating the ability of the NPs to generate its second messenger, cyclic guanosine monophosphate.Inhibition of PDI attenuated the ability of ANP, BNP and CNP to generate cGMP in human mesangial cells (HMCs, human umbilical vein endothelial cells (HUVECs, and human aortic smooth muscle cells (HASMCs, each of which were shown to express PDI. In LLC-PK1 cells, where PDI expression was undetectable by immunoblotting, PDI inhibition had a minimal effect on cGMP generation. Addition of PDI to cultured LLC-PK1 cells increased intracellular cGMP generation mediated by ANP. Inhibition of PDI in vivo attenuated NP-mediated generation of cGMP by ANP. Surface Plasmon Resonance demonstrated modest and differential binding of the natriuretic peptides with immobilized PDI in a cell free system. However, PDI was shown to co-localize on the surface of cells with GC-A and GC-B by co-immunoprecpitation and immunohistochemistry.These data demonstrate for the first time that cell surface PDI expression and function regulate the capacity of natriuretic peptides to generate cGMP through interaction with their receptors.

  7. Surface enhanced Raman spectroscopy measurements of MCF7 cells adhesion in confined micro-environments

    KAUST Repository

    De Vitis, Stefania

    2015-05-01

    Undoubtedly cells can perceive the external environment, not only from a biochemical point of view with the related signalling pathways, but also from a physical and topographical perspective. In this sense controlled three dimensional micro-structures as well as patterns at the nano-scale can affect and guide the cell evolution and proliferation, due to the fact that the surrounding environment is no longer isotropic (like the flat surfaces of standard cell culturing) but possesses well defined symmetries and anisotropies. In this work regular arrays of silicon micro-pillars with hexagonal arrangement are used as culturing substrates for MCF-7 breast cancer cells. The characteristic size and spacing of the pillars are tens of microns, comparable with MCF-7 cell dimensions and then well suited to induce acceptable external stimuli. It is shown that these cells strongly modify their morphology for adapting themselves to the micro-structured landscape, by means of protrusions from the main body of the cell. Scanning electron microscopy along with both Raman micro-spectroscopy and surface enhanced Raman spectroscopy are used for topographical and biochemical studies of the new cell arrangement. We have revealed that single MCF-7 cells exploit their capability to produce invadopodia, usually generated to invade the neighboring tissue in metastatic activity, for spanning and growing across separate pillars. © 2015 Elsevier Ltd.

  8. Olopatadine Inhibits Exocytosis in Rat Peritoneal Mast Cells by Counteracting Membrane Surface Deformation

    Directory of Open Access Journals (Sweden)

    Asuka Baba

    2015-01-01

    Full Text Available Backgroud/Aims: Besides its anti-allergic properties as a histamine receptor antagonist, olopatadine stabilizes mast cells by inhibiting the release of chemokines. Since olopatadine bears amphiphilic features and is preferentially partitioned into the lipid bilayers of the plasma membrane, it would induce some morphological changes in mast cells and thus affect the process of exocytosis. Methods: Employing the standard patch-clamp whole-cell recording technique, we examined the effects of olopatadine and other anti-allergic drugs on the membrane capacitance (Cm in rat peritoneal mast cells during exocytosis. Using confocal imaging of a water-soluble fluorescent dye, lucifer yellow, we also examined their effects on the deformation of the plasma membrane. Results: Low concentrations of olopatadine (1 or 10 µM did not significantly affect the GTP-γ-S-induced increase in the Cm. However, 100 µM and 1 mM olopatadine almost totally suppressed the increase in the Cm. Additionally, these doses completely washed out the trapping of the dye on the cell surface, indicating that olopatadine counteracted the membrane surface deformation induced by exocytosis. As shown by electron microscopy, olopatadine generated inward membrane bending in mast cells. Conclusion: This study provides electrophysiological evidence for the first time that olopatadine dose-dependently inhibits the process of exocytosis in rat peritoneal mast cells. Such mast cell stabilizing properties of olopatadine may be attributed to its counteracting effects on the plasma membrane deformation in degranulating mast cells.

  9. Surface enhanced Raman spectroscopy measurements of MCF7 cells adhesion in confined micro-environments

    Science.gov (United States)

    De Vitis, Stefania; Coluccio, Maria Laura; Gentile, Francesco; Malara, Natalia; Perozziello, Gerardo; Dattola, Elisabetta; Candeloro, Patrizio; Di Fabrizio, Enzo

    2016-01-01

    Undoubtedly cells can perceive the external environment, not only from a biochemical point of view with the related signalling pathways, but also from a physical and topographical perspective. In this sense controlled three dimensional micro-structures as well as patterns at the nano-scale can affect and guide the cell evolution and proliferation, due to the fact that the surrounding environment is no longer isotropic (like the flat surfaces of standard cell culturing) but possesses well defined symmetries and anisotropies. In this work regular arrays of silicon micro-pillars with hexagonal arrangement are used as culturing substrates for MCF-7 breast cancer cells. The characteristic size and spacing of the pillars are tens of microns, comparable with MCF-7 cell dimensions and then well suited to induce acceptable external stimuli. It is shown that these cells strongly modify their morphology for adapting themselves to the micro-structured landscape, by means of protrusions from the main body of the cell. Scanning electron microscopy along with both Raman micro-spectroscopy and surface enhanced Raman spectroscopy are used for topographical and biochemical studies of the new cell arrangement. We have revealed that single MCF-7 cells exploit their capability to produce invadopodia, usually generated to invade the neighboring tissue in metastatic activity, for spanning and growing across separate pillars.

  10. A cell surface clicked navigation system to direct specific bone targeting.

    Science.gov (United States)

    Kim, Young; Zhang, Zhe; Shim, Jae-Hyuck; Lee, Tae Sup; Tung, Ching-Hsuan

    2018-02-01

    Cell therapies are promising up-and-coming therapeutic strategies for many diseases. For maximal therapeutic benefits, injected cells have to navigate their way to a designated area, including organ and tissue; unfortunately, the majority of therapeutic cells are currently administered without a guide or homing device. To improve this serious shortcoming, a functionalization method was developed to equip cells with a homing signal. Its application was demonstrated by applying an Azadibenzocyclooctyne-bisphosphonate (ADIBO-BP) and azide paired bioorthogonal chemistry on cells for bone specific homing. Jurkat T cells and bone marrow derived stromal cells (BMSCs) were cultured with tetraacetylated N-azidoacetyl-d-mannosamine (Ac 4 ManNAz) to place unnatural azido groups onto the cell's surface; these azido groups were then reacted with ADIBO-BP. The tethered bisphosphonates were able to bring Jurkat cells to hydroxyapatite, the major component of bone, and mineralized SAOS-2 cells. The incorporated BP groups also enhanced the specific affinity of BMSCs to mouse femur bone fragments in vitro. The introduced navigation strategy is expected to have a broad application in cell therapy, because through the biocompatible ADIBO and azide reactive pair, various homing signals could be efficiently anchored onto therapeutic cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Plasma of Argon Increases Cell Attachment and Bacterial Decontamination on Different Implant Surfaces.

    Science.gov (United States)

    Canullo, Luigi; Genova, Tullio; Wang, Hom-Lay; Carossa, Stefano; Mussano, Federico

    This in vitro study tested the effects of argon atmospheric pressure dielectric barrier discharge (APDBD) on different implant surfaces with regard to physical changes, bacterial decontamination, and osteoblast adhesion. Seven hundred twenty disks with three different surface topographies-machined (MAC), titanium plasma-sprayed (TPS), and zirconia-blasted and acid-etched (ZRT)-were tested in this experiment. Bacterial adhesion tests were performed repeatedly on a simplified biofilm of Streptococcus mitis. Bacteria were incubated in the presence of the samples, which were subsequently either left untreated as controls or treated with APDBD for 30, 60, and 120 seconds. Samples were then metalized, prior to the recurring acquisition of images using a scanning electronic microscope (SEM). Protein adsorption, surface wettability, and early biologic response were determined for both treated (120 seconds) and untreated implant surfaces. For depicting the eukaryotic cell behavior, preosteoblastic murine cells were used. Cells were conveniently stained, and nuclei were counted. Cell viability was assessed by a chemiluminescent assay at 1, 2, and 3 days. On all treated samples, values of the contact angle measurements were lower than 10 degrees. The untreated samples showed values of contact angle of 80, 100, and 110 degrees, respectively, for MAC, TPS, and ZRT. The protein adsorption on TPS and ZRT was significantly increased after the plasma of argon treatment. However, no significant effect was noted on the MAC disks. The number and the cell spreading area of adherent osteoblasts significantly increased in all treated surfaces. Nonetheless, argon treatment did not influence the osteoblast proliferation and viability at different time points. Bacteria adhesion was significantly reduced, even after 60 seconds of argon treatment. Preliminary data showed that argon atmospheric pressure dielectric barrier discharge disinfected the implant surface, with potential to promote

  12. Bacterial cell surface properties: role of loosely bound extracellular polymeric substances (LB-EPS).

    Science.gov (United States)

    Zhao, Wenqiang; Yang, Shanshan; Huang, Qiaoyun; Cai, Peng

    2015-04-01

    This study investigated the effect of loosely bound extracellular polymeric substances (LB-EPS) on the comprehensive surface properties of four bacteria (Bacillus subtilis, Streptococcus suis, Escherichia coli and Pseudomonas putida). The removal of LB-EPS from bacterial surfaces by high-speed centrifugation (12,000×g) was confirmed by SEM images. Viability tests showed that the percentages of viable cells ranged from 95.9% to 98.0%, and no significant difference was found after treatment (P>0.05). FTIR spectra revealed the presence of phosphodiester, carboxylic, phosphate, and amino functional groups on bacteria surfaces, and the removal of LB-EPS did not alter the types of cell surface functional groups. Potentiometric titration results suggested the total site concentrations on the intact bacteria were higher than those on LB-EPS free bacteria. Most of the acidity constants (pKa) were almost identical, except the increased pKa values of phosphodiester groups on LB-EPS free S. suis and E. coli surfaces. The electrophoretic mobilities and hydrodynamic diameters of the intact and LB-EPS free bacteria were statistically unchanged (P>0.05), indicating LB-EPS had no influence on the net surface charges and size distribution of bacteria. However, LB-ESP could enhance cell aggregation processes. The four LB-EPS free bacteria all exhibited fewer hydrophobicity values (26.1-65.0%) as compared to the intact cells (47.4-69.3%), suggesting the removal of uncharged nonpolar compounds (e.g., carbohydrates) in LB-EPS. These findings improve our understanding of the changes in cell surface characterizations induced by LB-EPS, and have important implications for assessing the role of LB-EPS in bacterial adhesion and transport behaviors. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Proteomic analysis of cell surface-associated proteins from probiotic Lactobacillus plantarum

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Madsen, Søren M; Glenting, Jacob

    2009-01-01

    In the present study, we used a proteomic approach to identify surface-associated proteins from the probiotic bacterium Lactobacillus plantarum 299v. Proteins were extracted from the cell surface using a mild wash in phosphate buffer and analysed by sodium dodecyl sulphate-polyacrylamide gel...... of probiotics in the gastrointestinal tract. The results provide the basis for future studies on the molecular mechanisms of probiotics....

  14. Efficiency improvement of multicrystalline silicon solar cells after surface and grain boundaries passivation using vanadium oxide

    International Nuclear Information System (INIS)

    Derbali, L.; Ezzaouia, H.

    2012-01-01

    Highlights: ► Evaporation of vanadium pentoxide onto the front surface leads to reduce the surface reflectivity considerably. ► An efficient surface passivation can be obtained after thermal treatment of obtained films. ► Efficiency of the obtained solar cells has been improved noticeably after thermal treatment of deposited thin films. - Abstract: The aim of this work is to investigate the effect of vanadium oxide deposition onto the front surface of multicrystalline silicon (mc-Si) substrat, without any additional cost in the fabrication process and leading to an efficient surface and grain boundaries (GBs) passivation that have not been reported before. The lowest reflectance of mc-Si coated with vanadium oxide film of 9% was achieved by annealing the deposited film at 600 °C. Vanadium pentoxide (V 2 O 5 ) were thermally evaporated onto the surface of mc-Si substrates, followed by a short annealing duration at a temperature ranging between 600 °C and 800 °C, under O 2 atmosphere. The chemical composition of the films was analyzed by means of Fourier transform infrared spectroscopy (FTIR). Surface and cross-section morphology were determined by atomic force microscope (AFM) and a scanning electron microscope (SEM), respectively. The deposited vanadium oxide thin films make the possibility of combining in one processing step an antireflection coating deposition along with efficient surface state passivation, as compared to a reference wafer. Silicon solar cells based on untreated and treated mc-Si wafers were achieved. We showed that mc-silicon solar cells, subjected to the above treatment, have better short circuit currents and open-circuit voltages than those made from untreated wafers. Thus, the efficiency of obtained solar cells has been improved.

  15. Pancreatic hormones are expressed on the surfaces of human and rat islet cells through exocytotic sites

    DEFF Research Database (Denmark)

    Larsson, L I; Hutton, J C; Madsen, O D

    1989-01-01

    . Electron microscopy reveals the labeling to occur at sites of exocytotic granule release, involving the surfaces of extruded granule cores. The surfaces of islet cells were labeled both by polyclonal and monoclonal antibodies, excluding that receptor-interacting, anti-idiotypic hormone antibodies were...... for these results. It is concluded that the staining reflects interactions between the appropriate antibodies and exocytotic sites of hormone release....

  16. Radiation Response of Cancer Stem-Like Cells From Established Human Cell Lines After Sorting for Surface Markers

    International Nuclear Information System (INIS)

    Al-Assar, Osama; Muschel, Ruth J.; Mantoni, Tine S.; McKenna, W. Gillies; Brunner, Thomas B.

    2009-01-01

    Purpose: A subpopulation of cancer stem-like cells (CSLC) is hypothesized to exist in different cancer cell lines and to mediate radioresistance in solid tumors. Methods and Materials: Cells were stained for CSLC markers and sorted (fluorescence-activated cell sorter/magnetic beads) to compare foci and radiosensitivity of phosphorylated histone H2AX at Ser 139 (γ-H2AX) in sorted vs. unsorted populations in eight cell lines from different organs. CSLC properties were examined using anchorage-independent growth and levels of activated Notch1. Validation consisted of testing tumorigenicity and postirradiation enrichment of CSLC in xenograft tumors. Results: The quantity of CSLC was generally in good agreement with primary tumors. CSLC from MDA-MB-231 (breast) and Panc-1 and PSN-1 (both pancreatic) cells had fewer residual γ-H2AX foci than unsorted cells, pointing to radioresistance of CSLC. However, only MDA-MB-231 CSLC were more radioresistant than unsorted cells. Furthermore, MDA-MB-231 CSLC showed enhanced anchorage-independent growth and overexpression of activated Notch1 protein. The expression of cancer stem cell surface markers in the MDA-MB-231 xenograft model was increased after exposure to fractionated radiation. In contrast to PSN-1 cells, a growth advantage for MDA-MB-231 CSLC xenograft tumors was found compared to tumors arising from unsorted cells. Conclusions: CSLC subpopulations showed no general radioresistant phenotype, despite the quantities of CSLC subpopulations shown to correspond relatively well in other reports. Likewise, CSLC characteristics were found in some but not all of the tested cell lines. The reported problems in testing for CSLC in cell lines may be overcome by additional techniques, beyond sorting for markers.

  17. The lid wiper contains goblet cells and goblet cell crypts for ocular surface lubrication during the blink.

    Science.gov (United States)

    Knop, Nadja; Korb, Donald R; Blackie, Caroline A; Knop, Erich

    2012-06-01

    The conjunctival side of the upper and lower inner eyelid borders, termed the lid wiper, has a thickened epithelial lip for apposition to the globe, assumed to distribute the preocular tear film. The human lid wiper structure and its goblet cells are investigated. Conjunctival whole mounts, including lid margins from 17 eyes of human body donors, were investigated by routine histology and semithin plastic sections, using histology, histochemistry, and immunohistochemistry. In routine histology, the conjunctival lid wiper epithelium regularly showed goblet cells, single and in clusters, at the luminal surface and also deep within the epithelium without apparent surface contact. Semithin sections revealed that the deep goblet cells were often connected to cryptal epithelial infoldings that opened to the surface, hence making their mucins available at the surface. The goblet cells produced mucins of neutral (periodic acid-Schiff) and acidic (Alcian blue) type and stained positive for the gel-forming mucin MUC5AC. Surprisingly, MUC5AC-negative goblet cells were also observed in the lid wiper. Contrary to conventional assumptions, the lid wiper is part of the conjunctiva. It contains previously undescribed goblet cell crypts deep in the epithelium, suitable as an internal lubrication system for reduction of friction between the lid margin and the globe. This provides the first evidence of the morphological basis for the hydrodynamic type of lubrication and a more conclusive understanding of lid-margin lubrication and tear film distribution. It is another strong indication that the lid wiper is that area in apposition with the globe for distributing the thin preocular tear film during the blink.

  18. Antibody-protein A conjugated quantum dots for multiplexed imaging of surface receptors in living cells.

    Science.gov (United States)

    Jin, Takashi; Tiwari, Dhermendra K; Tanaka, Shin-Ichi; Inouye, Yasushi; Yoshizawa, Keiko; Watanabe, Tomonobu M

    2010-11-01

    To use quantum dots (QDs) as fluorescent probes for receptor imaging, QD surface should be modified with biomolecules such as antibodies, peptides, carbohydrates, and small-molecule ligands for receptors. Among these QDs, antibody conjugated QDs are the most promising fluorescent probes. There are many kinds of coupling reactions that can be used for preparing antibody conjugated QDs. Most of the antibody coupling reactions, however, are non-selective and time-consuming. In this paper, we report a facile method for preparing antibody conjugated QDs for surface receptor imaging. We used ProteinA as an adaptor protein for binding of antibody to QDs. By using ProteinA conjugated QDs, various types of antibodies are easily attached to the surface of the QDs via non-covalent binding between the F(c) (fragment crystallization) region of antibody and ProteinA. To show the utility of ProteinA conjugated QDs, HER2 (anti-human epidermal growth factor receptor 2) in KPL-4 human breast cancer cells were stained by using anti-HER2 antibody conjugated ProteinA-QDs. In addition, multiplexed imaging of HER2 and CXCR4 (chemokine receptor) in the KPL-4 cells was performed. The result showed that CXCR4 receptors coexist with HER2 receptors in the membrane surface of KPL-4 cells. ProteinA mediated antibody conjugation to QDs is very useful to prepare fluorescent probes for multiplexed imaging of surface receptors in living cells.

  19. Construction and characterization of a thermostable whole-cell chitinolytic enzyme using yeast surface display.

    Science.gov (United States)

    Li, Xiaobo; Jin, Xiaobao; Lu, Xuemei; Chu, Fujiang; Shen, Juan; Ma, Yan; Liu, Manyu; Zhu, Jiayong

    2014-10-01

    To develop a novel yeast whole-cell biocatalyst by yeast surface display technology that can hydrolyze chitin, the chitinaseC gene from Serratia marcescens AS1.1652 strain was cloned and subcloned into the yeast surface display plasmid pYD1, and the recombinant plasmid pYD1/SmchiC was electroporated into Saccharomyces cerevisiae EBY100 cell. Aga2p-SmChiC fusion protein was expressed and anchored on the yeast cell surface by induction with galactose, which was verified by indirect immunofluorescence and Western blotting. The chitinolytic activity of the yeast whole-cell biocatalyst or partially purified enzyme was detected by agar plate clear zone test, SDS-PAGE zymography and dinitrosalicylic acid method. The results showed that the chitinaseC gene from S. marcescens AS1.1652 strain was successfully cloned and expressed on the yeast cell surface, Aga2p-SmChiC fusion protein with molecular weight (67 kDa) was determined. Tests on the effect of temperature and pH on enzyme activity and stability revealed that the yeast whole-cell biocatalyst and partially purified enzyme possessed both thermal stability and activity, and even maintained some activity under acidic and weakly alkaline conditions. The optimum reaction temperature and pH value were set at 52 °C and 5.0, respectively. Yeast surface display technology succeeded in preparing a yeast whole-cell biocatalyst with chitinolytic activity, and the utilization of chitin could benefit from this process of enzyme preparation.

  20. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

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

    2009-01-01

    surface-negative despite effective induction of apoptosis. Interestingly, inhibition of endolysosomes or normal ER/Golgi transport did not affect Hsp70 surface expression. Intracellular calcium and the transcription factor Sp1, which has been shown previously to be important for the intracellular stress...

  1. Cell-surface expression of Hsp70 on hematopoietic cancer cells after inhibition of HDAC activity

    DEFF Research Database (Denmark)

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

    or normal ER/Golgi transport did not affect Hsp70 surface expression. Intracellular Calcium and the transcription factor Sp1, that has previously been shown to be important for the intracellular stress mediated by HDAC-inhibitors, were not involved in Hsp70 surface expression. We also found that HDAC...

  2. Actin dynamics in cells on nanotopographical surfaces in competition with chemotaxis and electrotaxis

    Science.gov (United States)

    Schmidt, Sebastian

    Directed cell migration can be guided by different types of gradients, for example chemotaxis. We use surfaces with nanotopographical ridges to examine a type of guidance called esotaxis on migration in the well-studied amoeba Dictyostelium Discoideum. In this work we compare chemotaxis with esotaxis on ridges as well as the influence of electrotaxis on the formation of the actin cytoskeleton on these nanotopographies. These esotactic surfaces have more guidance cues for cells than planar 2D cultures and can disrupt other guidance types like chemotaxis.

  3. Development and Application of a Flow Reactor Cell for Studies of Surface Chemistry

    Science.gov (United States)

    Algrim, L. B.; Pagonis, D.; Price, D.; Day, D. A.; De Gouw, J. A.; Jimenez, J. L.; Ziemann, P. J.

    2017-12-01

    We have designed, constructed, characterized, and employed a flow reactor cell that can be used to investigate the interaction of gaseous species such as volatile organic compounds (VOCs), oxidants, acids, and water vapor with authentic and model surfaces that are present in indoor and outdoor environments. The 3.9 L rectangular cell is made of FEP-coated aluminum and has one open face that can be sealed to the surface of interest. An internal plunger is raised (lowered) to expose (cover) the surface while various probe chemicals are added to the flow. To date we have exposed painted surfaces to O3, OH radicals (made from reaction of O3 with tetramethylethene and from photolysis of methyl nitrate/NO mixtures), and NO3 radicals (made from thermal decomposition N2O5) and analyzed the emitted oxidation products with a proton transfer reaction mass spectrometer (PTR-MS) and chemical ionization mass spectrometer (CIMS) equipped with an iodide reagent ion source. Further studies have included the reaction of oxidants with surfaces coated with organic films such as squalene and polyethylene glycol, as well as uptake of ketones and acids from the gas-phase to painted surfaces. The cell was also recently deployed at the University of Colorado-Boulder Art Museum during spring of 2017 to investigate the oxidation products released from the museum walls and floors. Results from all of these studies will be presented.

  4. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

    Science.gov (United States)

    Tarazanova, Mariya; Huppertz, Thom; Beerthuyzen, Marke; van Schalkwijk, Saskia; Janssen, Patrick; Wels, Michiel; Kok, Jan; Bachmann, Herwig

    2017-01-01

    Surface properties of bacteria are determined by the molecular composition of the cell wall and they are important for interactions of cells with their environment. Well-known examples of bacterial interactions with surfaces are biofilm formation and the fermentation of solid materials like food and feed. Lactococcus lactis is broadly used for the fermentation of cheese and buttermilk and it is primarily isolated from either plant material or the dairy environment. In this study, we characterized surface hydrophobicity, charge, emulsification properties, and the attachment to milk proteins of 55 L. lactis strains in stationary and exponential growth phases. The attachment to milk protein was assessed through a newly developed flow cytometry-based protocol. Besides finding a high degree of biodiversity, phenotype-genotype matching allowed the identification of candidate genes involved in the modification of the cell surface. Overexpression and gene deletion analysis allowed to verify the predictions for three identified proteins that altered surface hydrophobicity and attachment of milk proteins. The data also showed that lactococci isolated from a dairy environment bind higher amounts of milk proteins when compared to plant isolates. It remains to be determined whether the alteration of surface properties also has potential to alter starter culture functionalities. PMID:28936202

  5. Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

    Directory of Open Access Journals (Sweden)

    Mariya Tarazanova

    2017-09-01

    Full Text Available Surface properties of bacteria are determined by the molecular composition of the cell wall and they are important for interactions of cells with their environment. Well-known examples of bacterial interactions with surfaces are biofilm formation and the fermentation of solid materials like food and feed. Lactococcus lactis is broadly used for the fermentation of cheese and buttermilk and it is primarily isolated from either plant material or the dairy environment. In this study, we characterized surface hydrophobicity, charge, emulsification properties, and the attachment to milk proteins of 55 L. lactis strains in stationary and exponential growth phases. The attachment to milk protein was assessed through a newly developed flow cytometry-based protocol. Besides finding a high degree of biodiversity, phenotype-genotype matching allowed the identification of candidate genes involved in the modification of the cell surface. Overexpression and gene deletion analysis allowed to verify the predictions for three identified proteins that altered surface hydrophobicity and attachment of milk proteins. The data also showed that lactococci isolated from a dairy environment bind higher amounts of milk proteins when compared to plant isolates. It remains to be determined whether the alteration of surface properties also has potential to alter starter culture functionalities.

  6. Bacterial whole-cell biocatalysts by surface display of enzymes: toward industrial application.

    Science.gov (United States)

    Schüürmann, Jan; Quehl, Paul; Festel, Gunter; Jose, Joachim

    2014-10-01

    Despite the first report on the bacterial display of a recombinant peptide appeared almost 30 years ago, industrial application of cells with surface-displayed enzymes is still limited. To display an enzyme on the surface of a living cell bears several advantages. First of all, neither the substrate nor the product of the enzymatic reaction needs to cross a membrane barrier. Second, the enzyme being linked to the cell can be separated from the reaction mixture and hence the product by simple centrifugation. Transfer to a new substrate preparation results in multiple cycles of enzymatic conversion. Finally, the anchoring in a matrix, in this case, the cell envelope stabilizes the enzyme and makes it less accessible to proteolytic degradation and material adsorption resulting in continuous higher activities. These advantages in common need to balance some disadvantages before this application can be taken into account for industrial processes, e.g., the exclusion of the enzyme from the cellular metabolome and hence from redox factors or other co-factors that need to be supplied. Therefore, this digest describes the different systems in Gram-positive and Gram-negative bacteria that have been used for the surface display of enzymes so far and focuses on examples among these which are suitable for industrial purposes or for the production of valuable resources, not least in order to encourage a broader application of whole-cell biocatalysts with surface-displayed enzymes.

  7. Surface and protein analyses of normal human cell attachment on PIII-modified chitosan membranes

    Energy Technology Data Exchange (ETDEWEB)

    Saranwong, N. [Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Inthanon, K. [Human and Animal Cell Technology Research Laboratory, Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Wongkham, W., E-mail: weerah@chiangmai.ac.th [Human and Animal Cell Technology Research Laboratory, Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Wanichapichart, P. [Nanotechnology Center of Excellence and Membrane Science and Technology Research Center, Department of Physics, Faculty of Science, Prince of Songkla University, Hat Yai, Songkla 90110 (Thailand); Suwannakachorn, D. [Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Yu, L.D., E-mail: yuld@fnrf.science.cmu.ac.th [Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand)

    2012-02-01

    Surface of chitosan membrane was modified with argon (Ar) and nitrogen (N) plasma immersion ion implantation (PIII) for human skin fibroblasts F1544 cell attachment. The modified surfaces were characterized by Fourier transform infrared spectroscopy (FTIR) and atomic force microscopy (AFM). Cell attachment patterns were evaluated by scanning electron microscopy (SEM). The enzyme-linked immunosorbent assay (ELISA) was used to quantify levels of focal adhesion kinase (FAK). The results showed that Ar PIII had an enhancement effect on the cell attachment while N-PIII had an inhibition effect. Filopodial analysis revealed more microfilament cytoplasmic spreading on the edge of cells attached on the Ar-treated membranes than N-treated membranes. Higher level FAK was found in Ar-treated membranes than that in N-treated membranes.

  8. Bottom-up engineering of the surface roughness of nanostructured cubic zirconia to control cell adhesion

    International Nuclear Information System (INIS)

    Singh, A V; Ferri, M; Tamplenizza, M; Borghi, F; Lenardi, C; Piazzoni, C; Podestà, A; Milani, P; Divitini, G; Ducati, C; Merlini, M

    2012-01-01

    Nanostructured cubic zirconia is a strategic material for biomedical applications since it combines superior structural and optical properties with a nanoscale morphology able to control cell adhesion and proliferation. We produced nanostructured cubic zirconia thin films at room temperature by supersonic cluster beam deposition of nanoparticles produced in the gas phase. Precise control of film roughness at the nanoscale is obtained by operating in a ballistic deposition regime. This allows one to study the influence of nanoroughness on cell adhesion, while keeping the surface chemistry constant. We evaluated cell adhesion on nanostructured zirconia with an osteoblast-like cell line using confocal laser scanning microscopy for detailed morphological and cytoskeleton studies. We demonstrated that the organization of cytoskeleton and focal adhesion formation can be controlled by varying the evolution of surface nanoroughness. (paper)

  9. Surface Engineering of ZnO Thin Film for High Efficiency Planar Perovskite Solar Cells

    Science.gov (United States)

    Tseng, Zong-Liang; Chiang, Chien-Hung; Wu, Chun-Guey

    2015-09-01

    Sputtering made ZnO thin film was used as an electron-transport layer in a regular planar perovskite solar cell based on high quality CH3NH3PbI3 absorber prepared with a two-step spin-coating. An efficiency up to 15.9% under AM 1.5G irradiation is achieved for the cell based on ZnO film fabricated under Ar working gas. The atmosphere of the sputtering chamber can tune the surface electronic properties (band structure) of the resulting ZnO thin film and therefore the photovoltaic performance of the corresponding perovskite solar cell. Precise surface engineering of ZnO thin film was found to be one of the key steps to fabricate ZnO based regular planar perovskite solar cell with high power conversion efficiency. Sputtering method is proved to be one of the excellent techniques to prepare ZnO thin film with controllable properties.

  10. Selection and syntheses of tentacle type peptides as 'artificial' lectins against various cell-surface carbohydrates.

    Science.gov (United States)

    Hyun, Soonsil; Kim, Jiyoung; Kwon, Miyun; Yu, Jaehoon

    2007-01-01

    Sialyl Lewis X and its derivatives are cell-surface carbohydrates that are involved in cell-cell recognition by carbohydrate-mediated interactions. Unfortunately, owing to the similarities between carbohydrates only a limited number of tools are available for their differentiation. In this study, we prepared a selected phage-displayed peptide library against LeX (2), SLN (3), or LN (4), which compared to sLeX (1) lack sialic acid, fucose, and both sialic acid and fucose from constituents, respectively. Sequences of the selected peptides, prepared as tentacle type dimeric peptides, were prepared and shown to have micromolar affinities for the cognate carbohydrates. The specificities displayed by these 'artificial' lectins overwhelm those of natural lectins. These results suggest that they can serve as useful tools to detect changes in the terminal monosaccharide of cell-surface carbohydrates.

  11. Selective cell culture on UV transparent polymer by F2 laser surface modification

    International Nuclear Information System (INIS)

    Hanada, Yasutaka; Sugioka, Koji; Kawano, Hiroyuki; Tsuchimoto, Takayoshi; Miyamoto, Iwao; Miyawaki, Atsushi; Midorikawa, Katsumi

    2009-01-01

    A microchip made of UV transparent polymer (CYTOP) that can perform selective cell culture has been fabricated by F 2 laser surface modification. The refractive index of CYTOP is almost the same as that of culture medium, which is essential for three-dimensional (3D) observation of cells. The F 2 laser modification of CYTOP achieves hydrophilicity only on the laser irradiated area with little deterioration of the optical properties and surface smoothness. After the laser modification, HeLa cells were successfully cultured and strongly adhered only on the modified area of CYTOP. The cells patterned on CYTOP were applied for clear 3D observation using an optical microscope in phase contrast mode.

  12. Development of genetically modified eliminable human dermal fibroblast feeder cells for ocular surface regeneration medicine.

    Science.gov (United States)

    Li, Yingli; Inoue, Tomoyuki; Takamatsu, Fumihiko; Maeda, Naoyuki; Ohashi, Yuichi; Nishida, Kohji

    2013-11-15

    Cultured human corneal limbal stem/progenitor cells are usually established and maintained on feeder layers. However, animal feeder cells are associated with viral infection, pathogen transmission, and xenogenic contamination. All feeder cells also can be mixed easily into cell-sheet production, causing self-contamination. We developed a line of labeled, immortalized, eliminable human dermal fibroblast cells to eliminate these problems. The enhanced green fluorescent protein gene, human-derived telomerase reverse transcriptase gene, and herpes simplex virus thymidine kinase gene were transfected into human dermal fibroblast cells to establish labeled, immortalized, eliminable feeder cells. Established eliminable dermal fibroblasts (TERT+TK-D) were treated with mitomycin, cocultured with human limbal stem/progenitor cells to regenerate epithelium sheets, and compared with 3T3 feeder cells. Established TERT+TK-D feeder cells maintained immortalization, visualization, and eliminable characteristics during 6 months of continuous passages. The colony-forming efficiency of limbal stem/progenitor cells was similar in the TERT+TK-D group (11.77 ± 0.21%) and the 3T3 group (12.8 ± 1.61%) (P = 0.332). All cell sheets were well stratified into 4 to 5 layers. The TERT+TK-D group colonies and epithelial cell sheets showed weaker staining of corneal epithelium differentiation marker K3 than the 3T3 group and quantitative analysis of mRNA transcripts. Moreover, PCR analysis against the long terminal repeat sequence of the lentiviral vector integrated into the genetically modified feeder cells showed no contamination of ganciclovir-treated regeneration epithelial sheets. Genetically modified, labeled, immortalized, eliminable human dermal feeder cells are promising substitutes for 3T3 feeder cells for xenogeny-free ocular surface regeneration.

  13. Study of double porous silicon surfaces for enhancement of silicon solar cell performance

    Science.gov (United States)

    Razali, N. S. M.; Rahim, A. F. A.; Radzali, R.; Mahmood, A.

    2017-09-01

    In this work, design and simulation of double porous silicon surfaces for enhancement of silicon solar cell is carried out. Both single and double porous structures are constructed by using TCAD ATHENA and TCAD DEVEDIT tools of the SILVACO software respectively. After the structures were created, I-V characteristics and spectral response of the solar cell were extracted using ATLAS device simulator. Finally, the performance of the simulated double porous solar cell is compared with the performance of both single porous and bulk-Si solar cell. The results showed that double porous silicon solar cell exhibited 1.8% efficiency compared to 1.3% and 1.2% for single porous silicon and bulk-Si solar cell.

  14. Micro patterned surfaces: an effective tool for long term digital holographic microscopy cell imaging

    Science.gov (United States)

    Mues, Sarah; Lilge, Inga; Schönherr, Holger; Kemper, Björn; Schnekenburger, Jürgen

    2017-02-01

    The major problem of Digital Holographic Microscopy (DHM) long term live cell imaging is that over time most of the tracked cells move out of the image area and other ones move in. Therefore, most of the cells are lost for the evaluation of individual cellular processes. Here, we present an effective solution for this crucial problem of long-term microscopic live cell analysis. We have generated functionalized slides containing areas of 250 μm per 200 μm. These micropatterned biointerfaces consist of passivating polyaclrylamide brushes (PAAm). Inner areas are backfilled with octadecanthiol (ODT), which allows cell attachment. The fouling properties of these surfaces are highly controllable and therefore the defined areas designed for the size our microscopic image areas were effective in keeping all cells inside the rectangles over the selected imaging period.

  15. Reconstruction of limbal stem cell deficient corneal surface with induced human bone marrow mesenchymal stem cells on amniotic membrane.

    Science.gov (United States)

    Rohaina, Che Man; Then, Kong Yong; Ng, Angela Min Hwei; Wan Abdul Halim, Wan Haslina; Zahidin, Aida Zairani Mohd; Saim, Aminuddin; Idrus, Ruszymah B H

    2014-03-01

    The cornea can be damaged by a variety of clinical disorders or chemical, mechanical, and thermal injuries. The objectives of this study were to induce bone marrow mesenchymal stem cells (BMSCs) to corneal lineage, to form a tissue engineered corneal substitute (TEC) using BMSCs, and to treat corneal surface defects in a limbal stem cell deficiency model. BMSCs were induced to corneal lineage using limbal medium for 10 days. Induced BMSCs demonstrated upregulation of corneal stem cell markers; β1-integrin, C/EBPδ, ABCG2, and p63, increased protein expression of CK3 and p63 significantly compared with the uninduced ones. For TEC formation, passage 1 BMSCs were trypsinized and seeded on amniotic membrane in a transwell co-culture system and were grown in limbal medium. Limbal stem cell deficiency models were induced by alkaline injury, and the TEC was implanted for 8 weeks. Serial slit lamp evaluation revealed remarkable improvement in corneal regeneration in terms of corneal clarity and reduced vascularization. Histologic and optical coherence tomography analyses demonstrated comparable corneal thickness and achieved stratified epithelium with a compact stromal layer resembling that of normal cornea. CK3 and p63 were expressed in the newly regenerated cornea. In conclusion, BMSCs can be induced into corneal epithelial lineage, and these cells are viable for the formation of TEC, to be used for the reconstruction of the corneal surface in the limbal stem cell deficient model. Copyright © 2014 Mosby, Inc. All rights reserved.

  16. Data of continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surfaces

    Directory of Open Access Journals (Sweden)

    Chin-Chen Yeh

    2016-03-01

    Full Text Available This data article contains two figures and one table supporting the research article entitled: “Continuous harvest of stem cells via partial detachment from thermoresponsive nanobrush surface” [1]. The table shows coating conditions of three copolymers, poly(styrene-co-acrylic acid grafted with oligovitronectin, poly(styrene-co-N-isopropylacrylamide and poly(styrene-co-polyethylene glycol methacrylate to prepare thermoresponsive surface. XPS spectra show the nitrogen peak of the polystyrene surface coated with poly(styrene-co-acrylic acid grafted with oligovitronectin. The surface coating density analyzed from sorption of poly(styrene-co-acrylic acid grafted with oligovitronectin by UV–vis spectroscopy is also presented.

  17. The ultrastructural surface morphology of oral cancer cells and keratinocytes after exposure to chitosan

    Science.gov (United States)

    Fatimah; Sarsito, A. S.; Wimardhani, Y. S.

    2017-08-01

    Low-molecular-weight chitosan (LMWC) has the same selective cytotoxic effects on oral cancer cells as cisplatin. The cell deaths caused by the anticancer characteristics of chitosan show that apoptosis is not the death pathway of the primary cells involved. The interactions between LMWC and the cells need to be explored. The objective of this study was to compare the ultrastructural morphology of oral Squamous Cell Carcinoma (SCC Ca)-922 and noncancer keratinocyte HaCaT cell lines after exposure to LMWC and cisplatin. The cells were treated with LMWC and cisplatin, and their ultrastructural morphology was analyzed using scanning electron micrographs. Features of early apoptosis, seen as the loss of microvilli, were detected in the LMWC-exposed Ca9-22 cells, and there was a material surrounding the cells. In contrast, the LMWC-exposed HaCaT cells showed no changes related to apoptosis. The results were the opposite when cisplatin was used. This study confirms that there are differences in the ultrastructural surface morphology of LMWC-exposed and cisplatin-exposed oral cancer cells and keratinocytes that could be correlated with their biological activity.

  18. A cell-surface superoxide dismutase is a binding protein for peroxinectin, a cell-adhesive peroxidase in crayfish.

    Science.gov (United States)

    Johansson, M W; Holmblad, T; Thörnqvist, P O; Cammarata, M; Parrinello, N; Söderhäll, K

    1999-03-01

    Peroxinectin, a cell-adhesive peroxidase (homologous to human myeloperoxidase), from the crayfish Pacifastacus leniusculus, was shown by immuno-fluorescence to bind to the surface of crayfish blood cells (haemocytes). In order to identify a cell surface receptor for peroxinectin, labelled peroxinectin was incubated with a blot of haemocyte membrane proteins. It was found to specifically bind two bands of 230 and 90 kDa; this binding was decreased in the presence of unlabelled peroxinectin. Purified 230/90 kDa complex also bound peroxinectin in the same assay. In addition, the 230 kDa band binds the crayfish beta-1,3-glucan-binding protein. The 230 kDa band could be reduced to 90 kDa, thus showing that the 230 kDa is a multimer of 90 kDa units. The peroxinectin-binding protein was cloned from a haemocyte cDNA library, using immuno-screening or polymerase chain reaction based on partial amino acid sequence of the purified protein. It has a signal sequence, a domain homologous to CuZn-containing superoxide dismutases, and a basic, proline-rich, C-terminal tail, but no membrane-spanning segment. In accordance, the 90 and 230 kDa bands had superoxide dismutase activity. Immuno-fluorescence of non-permeabilized haemocytes with affinity-purified antibodies confirmed that the crayfish CuZn-superoxide dismutase is localized at the cell surface; it could be released from the membrane with high salt. It was thus concluded that the peroxinectin-binding protein is an extracellular SOD (EC-SOD) and a peripheral membrane protein, presumably kept at the cell surface via ionic interaction with its C-terminal region. This interaction with a peroxidase seems to be a novel function for an SOD. The binding of the cell surface SOD to the cell-adhesive/opsonic peroxinectin may mediate, or regulate, cell adhesion and phagocytosis; it may also be important for efficient localized production of microbicidal substances.

  19. A molecular smart surface for spatio-temporal studies of cell mobility.

    Science.gov (United States)

    Lee, Eun-ju; Luo, Wei; Chan, Eugene W L; Yousaf, Muhammad N

    2015-01-01

    Active migration in both healthy and malignant cells requires the integration of information derived from soluble signaling molecules with positional information gained from interactions with the extracellular matrix and with other cells. How a cell responds and moves involves complex signaling cascades that guide the directional functions of the cytoskeleton as well as the synthesis and release of proteases that facilitate movement through tissues. The biochemical events of the signaling cascades occur in a spatially and temporally coordinated manner then dynamically shape the cytoskeleton in specific subcellular regions. Therefore, cell migration and invasion involve a precise but constantly changing subcellular nano-architecture. A multidisciplinary effort that combines new surface chemistry and cell biological tools is required to understand the reorganization of cytoskeleton triggered by complex signaling during migration. Here we generate a class of model substrates that modulate the dynamic environment for a variety of cell adhesion and migration experiments. In particular, we use these dynamic substrates to probe in real-time how the interplay between the population of cells, the initial pattern geometry, ligand density, ligand affinity and integrin composition affects cell migration and growth. Whole genome microarray analysis indicates that several classes of genes ranging from signal transduction to cytoskeletal reorganization are differentially regulated depending on the nature of the surface conditions.

  20. Cell treatment and surface functionalization using a miniature atmospheric pressure glow discharge plasma torch

    International Nuclear Information System (INIS)

    Yonson, S; Coulombe, S; Leveille, V; Leask, R L

    2006-01-01

    A miniature atmospheric pressure glow discharge plasma torch was used to detach cells from a polystyrene Petri dish. The detached cells were successfully transplanted to a second dish and a proliferation assay showed the transplanted cells continued to grow. Propidium iodide diffused into the cells, suggesting that the cell membrane had been permeabilized, yet the cells remained viable 24 h after treatment. In separate experiments, hydrophobic, bacteriological grade polystyrene Petri dishes were functionalized. The plasma treatment reduced the contact angle from 93 0 to 35 0 , and promoted cell adhesion. Two different torch nozzles, 500 μm and 150 μm in internal diameter, were used in the surface functionalization experiments. The width of the tracks functionalized by the torch, as visualized by cell adhesion, was approximately twice the inside diameter of the nozzle. These results indicate that the miniature plasma torch could be used in biological micropatterning, as it does not use chemicals like the present photolithographic techniques. Due to its small size and manouvrability, the torch also has the ability to pattern complex 3D surfaces

  1. Proteomic Profiling of Neuroblastoma Cells Adhesion on Hyaluronic Acid-Based Surface for Neural Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Ming-Hui Yang

    2016-01-01

    Full Text Available The microenvironment of neuron cells plays a crucial role in regulating neural development and regeneration. Hyaluronic acid (HA biomaterial has been applied in a wide range of medical and biological fields and plays important roles in neural regeneration. PC12 cells have been reported to be capable of endogenous NGF synthesis and secretion. The purpose of this research was to assess the effect of HA biomaterial combining with PC12 cells conditioned media (PC12 CM in neural regeneration. Using SH-SY5Y cells as an experimental model, we found that supporting with PC12 CM enhanced HA function in SH-SY5Y cell proliferation and adhesion. Through RP-nano-UPLC-ESI-MS/MS analyses, we identified increased expression of HSP60 and RanBP2 in SH-SY5Y cells grown on HA-modified surface with cotreatment of PC12 CM. Moreover, we also identified factors that were secreted from PC12 cells and may promote SH-SY5Y cell proliferation and adhesion. Here, we proposed a biomaterial surface enriched with neurotrophic factors for nerve regeneration application.

  2. Cellular Homeostasis and Antioxidant Response in Epithelial HT29 Cells on Titania Nanotube Arrays Surface

    Directory of Open Access Journals (Sweden)

    Rabiatul Basria SMN Mydin

    2017-01-01

    Full Text Available Cell growth and proliferative activities on titania nanotube arrays (TNA have raised alerts on genotoxicity risk. Present toxicogenomic approach focused on epithelial HT29 cells with TNA surface. Fledgling cell-TNA interaction has triggered G0/G1 cell cycle arrests and initiates DNA damage surveillance checkpoint, which possibly indicated the cellular stress stimuli. A profound gene regulation was observed to be involved in cellular growth and survival signals such as p53 and AKT expressions. Interestingly, the activation of redox regulator pathways (antioxidant defense was observed through the cascade interactions of GADD45, MYC, CHECK1, and ATR genes. These mechanisms furnish to protect DNA during cellular division from an oxidative challenge, set in motion with XRRC5 and RAD50 genes for DNA damage and repair activities. The cell fate decision on TNA-nanoenvironment has been reported to possibly regulate proliferative activities via expression of p27 and BCL2 tumor suppressor proteins, cogent with SKP2 and BCL2 oncogenic proteins suppression. Findings suggested that epithelial HT29 cells on the surface of TNA may have a positive regulation via cell-homeostasis mechanisms: a careful circadian orchestration between cell proliferation, survival, and death. This nanomolecular knowledge could be beneficial for advanced medical applications such as in nanomedicine and nanotherapeutics.

  3. Antimicrobial efficacy of copper surfaces against spores and vegetative cells of Clostridium difficile: the germination theory.

    Science.gov (United States)

    Wheeldon, L J; Worthington, T; Lambert, P A; Hilton, A C; Lowden, C J; Elliott, T S J

    2008-09-01

    Persistent contamination of surfaces by spores of Clostridium difficile is a major factor influencing the spread of C. difficile-associated diarrhoea (CDAD) in the clinical setting. In recent years, the antimicrobial efficacy of metal surfaces has been investigated against microorganisms including methicillin-resistant Staphylococcus aureus. This study compared the survival of C. difficile on stainless steel, a metal contact surface widely used in hospitals, and copper surfaces. Antimicrobial efficacy was assessed using a carrier test method against dormant spores, germinating spores and vegetative cells of C. difficile (NCTC 11204 and ribotype 027) over a 3 h period in the presence and absence of organic matter. Copper metal eliminated all vegetative cells of C. difficile within 30 min, compared with stainless steel which demonstrated no antimicrobial activity (P or=2.5 log reduction (99.8% reduction) at 3 h. Organic material did not reduce the antimicrobial efficacy of the copper surface (P > 0.05). The use of copper surfaces within the clinical environment and application of a germination solution in infection control procedures may offer a novel way forward in eliminating C. difficile from contaminated surfaces and reducing CDAD.

  4. Surface enhanced imaging and IR spectroscopy of the biological cells on the nanostructured gold film

    Directory of Open Access Journals (Sweden)

    G.I. Dovbeshko

    2017-07-01

    Full Text Available New approach for optical imaging, structural study and cell cultivation based on the effect of the enhancement of optical signals from biomolecules and biological cells near nanostructured rough gold surface is proposed. The surface enhanced IR absorption (SEIRA spectroscopy and confocal microscopy experiments were made using the culture of SPEV (porcine embryonic kidney epithelium transplantable line and fibroblast cells, cultivated and/or adsorbed on the gold substrate. The SEIRA spectra registered from monolayer of the SPEV cells cultivated on the rough gold showed a low frequency shift of about 2 to 7 cm 1 for the most characteristic IR vibrations, compared with those adsorbed from suspension on the same substrate. An enhancement factor of 15…30 was obtained for different molecular vibrations. The confocal microscopy contrast images of the SPEV cells on rough gold substrate were obtained in laser fluorescence mode. This approach opens new possibilities for visualization of the living cells in vivo without staining. The fluorescence of the rough gold surfaces and effects responsible for our findings have been discussed.

  5. SEPARATION OF CELL POPULATIONS BY SUPER-PARAMAGNETIC PARTICLES WITH CONTROLLED SURFACE FUNCTIONALITY

    Directory of Open Access Journals (Sweden)

    Lootsik M. D.

    2014-02-01

    Full Text Available The recognition and isolation of specific mammalian cells by the biocompatible polymer coated super-paramagnetic particles with determined surface functionality were studied. The method of synthesis of nanoscaled particles on a core of iron III oxide (Fe2O3, magemit coated with a polymer shell containing reactive oligoperoxide groups for attachment of ligands is described. By using the developed superparamagnetic particles functionalized with peanut agglutinin (PNA we have separated the sub-populations of PNA+ and PNA– cells from ascites of murine Nemeth-Kellner lymphoma. In another type of experiment, the particles were opsonized with proteins of the fetal calf serum that improved biocompatibility of the particles and their ingestion by cultivated murine macrophages J774.2. Macrophages loaded with the particles were effeciently separated from the particles free cells by using the magnet. Thus, the developed surface functionalized superparamagnetic particles showed to be a versatile tool for cell separation independent on the mode of particles’ binding with cell surface or their engulfment by the targeted cells.

  6. Determining the fate of fluorescent quantum dots on surface of engineered budding S. cerevisiae cell molecular landscape.

    Science.gov (United States)

    Chouhan, Raghuraj S; Qureshi, Anjum; Niazi, Javed H

    2015-07-15

    In this study, we surface engineered living S. cerevisiae cells by decorating quantum dots (QDs) and traced the fate of QDs on molecular landscape of single mother cell through several generation times (progeny cells). The fate of QDs on cell-surface was tracked through the cellular division events using confocal microscopy and fluorescence emission profiles. The extent of cell-surface QDs distribution among the offspring was determined as the mother cell divides into daughter cells. Fluorescence emission from QDs on progeny cells was persistent through the second-generation time (~240min) until all of the progeny cells lost their cell-bound QDs during the third generation time (~360min). The surface engineered yeast cells were unaffected by the QDs present on their molecular landscapes and retained their normal cellular growth, architecture and metabolic activities as confirmed by their viability, scanning electron microscopy (SEM) examinations and cytotoxicity tests, respectively. Our results demonstrated that QDs on mother cell landscape tend to distribute among its progeny cells that accompanied with concomitant reduction in QDs' fluorescence, which can be quantified. We suggest that surface engineered cells with QDs will enable investigating the cellular behavior and monitoring cell growth patterns as nanobiosensors for screening of drugs/chemicals at single cell level with fewer side effects. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Additive manufactured polymeric 3D scaffolds with tailored surface topography influence mesenchymal stromal cells activity.

    Science.gov (United States)

    Neves, Sara C; Mota, Carlos; Longoni, Alessia; Barrias, Cristina C; Granja, Pedro L; Moroni, Lorenzo

    2016-05-24

    Additive manufactured three-dimensional (3D) scaffolds with tailored surface topography constitute a clear advantage in tissue regeneration strategies to steer cell behavior. 3D fibrous scaffolds of poly(ethylene oxide terephthalate)/poly(butylene terephthalate) block copolymer presenting different fiber surface features were successfully fabricated by additive manufacturing combined with wet-spinning, in a single step, without any post-processing. The optimization of the processing parameters, mainly driven by different solvent/non-solvent combinations, led to four distinct scaffold types, with average surface roughness values ranging from 0.071 ± 0.012 μm to 1.950 ± 0.553 μm, average pore sizes in the x- and y-axis between 351.1 ± 33.6 μm and 396.1 ± 32.3 μm, in the z-axis between 36.5 ± 5.3 μm and 70.7 ± 8.8 μm, average fiber diameters between 69.4 ± 6.1 μm and 99.0 ± 9.4 μm, and porosity values ranging from 60.2 ± 0.8% to 71.7 ± 2.6%. Human mesenchymal stromal cells (hMSCs) cultured on these scaffolds adhered, proliferated, and produced endogenous extracellular matrix. The effect of surface roughness and topography on hMSCs differentiation was more evident for cells seeded at lower density, where the percentage of cells in direct contact with the surface was higher compared to more densely seeded scaffolds. Under osteogenic conditions, lower surface roughness values (0.227 ± 0.035 μm) had a synergistic effect on hMSCs behavior, while chondrogenesis was favored on rougher surfaces (1.950 ± 0.553 μm).

  8. Directing and Potentiating Stem Cell-Mediated Angiogenesis and Tissue Repair by Cell Surface E-Selectin Coating.

    Science.gov (United States)

    Liu, Zhao-Jun; Daftarian, Pirouz; Kovalski, Letícia; Wang, Bo; Tian, Runxia; Castilla, Diego M; Dikici, Emre; Perez, Victor L; Deo, Sapna; Daunert, Sylvia; Velazquez, Omaida C

    2016-01-01

    Stem cell therapy has emerged as a promising approach for treatment of a number of diseases, including delayed and non-healing wounds. However, targeted systemic delivery of therapeutic cells to the dysfunctional tissues remains one formidable challenge. Herein, we present a targeted nanocarrier-mediated cell delivery method by coating the surface of the cell to be delivered with dendrimer nanocarriers modified with adhesion molecules. Infused nanocarrier-coated cells reach to destination via recognition and association with the counterpart adhesion molecules highly or selectively expressed on the activated endothelium in diseased tissues. Once anchored on the activated endothelium, nanocarriers-coated transporting cells undergo transendothelial migration, extravasation and homing to the targeted tissues to execute their therapeutic role. We now demonstrate feasibility, efficacy and safety of our targeted nanocarrier for delivery of bone marrow cells (BMC) to cutaneous wound tissues and grafted corneas and its advantages over conventional BMC transplantation in mouse models for wound healing and neovascularization. This versatile platform is suited for targeted systemic delivery of virtually any type of therapeutic cell.

  9. Anticoagulation and endothelial cell behaviors of heparin-loaded graphene oxide coating on titanium surface

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Chang-Jiang, E-mail: panchangjiang@hyit.edu.cn [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an 223003 (China); Pang, Li-Qun [Department of General Surgery, Huai' an First People' s Hospital, Nanjing Medical University, Huai' an 223300 (China); Gao, Fei [Zhejiang Zylox Medical Devices Co., Ltd., Hangzhou 310000 (China); Wang, Ya-Nan; Liu, Tao; Ye, Wei; Hou, Yan-Hua [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an 223003 (China)

    2016-06-01

    Owing to its unique physical and chemical properties, graphene oxide (GO) has attracted tremendous interest in many fields including biomaterials and biomedicine. The purpose of the present study is to investigate the endothelial cell behaviors and anticoagulation of heparin-loaded GO coating on the titanium surface. To this end, the titanium surface was firstly covered by the polydopamine coating followed by the deposition of the GO coating. Heparin was finally loaded on the GO coating to improve the blood compatibility. The results of attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR), Raman spectroscopy and X-ray photoelectron spectroscopy (XPS) indicated that the heparin-loaded GO coating was successfully created on the titanium surface. The scanning electron microscopy (SEM) images indicated that a relative uniform GO coating consisting of multilayer GO sheets was formed on the substrate. The hydrophilicity of the titanium surface was enhanced after the deposition of GO and further improved significantly by the loading heparin. The GO coating can enhance the endothelial cell adhesion and proliferation as compared with polydopamine coating and the blank titanium. Loading heparin on the GO coating can significantly reduce the platelet adhesion and prolong the activated partial thromboplastin time (APTT) while not influence the endothelial cell adhesion and proliferation. Therefore, the heparin-loaded GO coating can simultaneously enhance the cytocompatibility to endothelial cells and blood compatibility of biomaterials. Because the polydopamine coating can be easily prepared on most of biomaterials including polymer, ceramics and metal, thus the approach of the present study may open up a new window of promising an effective and efficient way to promote endothelialization and improve the blood compatibility of blood-contact biomedical devices such as intravascular stents. - Highlights: • Heparin-loaded graphene oxide coating was

  10. Motion of cells sedimenting on a solid surface in a laminar shear flow.

    Science.gov (United States)

    Tissot, O; Pierres, A; Foa, C; Delaage, M; Bongrand, P

    1992-01-01

    Cell adhesion often occurs under dynamic conditions, as in flowing blood. A quantitative understanding of this process requires accurate knowledge of the topographical relationships between the cell membrane and potentially adhesive surfaces. This report describes an experimental study made on both the translational and rotational velocities of leukocytes sedimenting of a flat surface under laminar shear flow. The main conclusions are as follows: (a) Cells move close to the wall with constant velocity for several tens of seconds. (b) The numerical values of translational and rotational velocities are inconsistent with Goldman's model of a neutrally buoyant sphere in a laminar shear flow, unless a drag force corresponding to contact friction between cells and the chamber floor is added. The phenomenological friction coefficient was 7.4 millinewton.s/m. (c) Using a modified Goldman's theory, the width of the gap separating cells (6 microns radius) from the chamber floor was estimated at 1.4 micron. (d) It is shown that a high value of the cell-to-substrate gap may be accounted for by the presence of cell surface protrusions of a few micrometer length, in accordance with electron microscope observations performed on the same cell population. (e) In association with previously reported data (Tissot, O., C. Foa, C. Capo, H. Brailly, M. Delaage, and P. Bongrand. 1991. Biocolloids and Biosurfaces. In press), these results are consistent with the possibility that cell-substrate attachment be initiated by the formation of a single molecular bond, which might be considered as the rate limiting step.

  11. An AFM-based pit-measuring method for indirect measurements of cell-surface membrane vesicles

    International Nuclear Information System (INIS)

    Zhang, Xiaojun; Chen, Yuan; Chen, Yong

    2014-01-01

    Highlights: • Air drying induced the transformation of cell-surface membrane vesicles into pits. • An AFM-based pit-measuring method was developed to measure cell-surface vesicles. • Our method detected at least two populations of cell-surface membrane vesicles. - Abstract: Circulating membrane vesicles, which are shed from many cell types, have multiple functions and have been correlated with many diseases. Although circulating membrane vesicles have been extensively characterized, the status of cell-surface membrane vesicles prior to their release is less understood due to the lack of effective measurement methods. Recently, as a powerful, micro- or nano-scale imaging tool, atomic force microscopy (AFM) has been applied in measuring circulating membrane vesicles. However, it seems very difficult for AFM to directly image/identify and measure cell-bound membrane vesicles due to the similarity of surface morphology between membrane vesicles and cell surfaces. Therefore, until now no AFM studies on cell-surface membrane vesicles have been reported. In this study, we found that air drying can induce the transformation of most cell-surface membrane vesicles into pits that are more readily detectable by AFM. Based on this, we developed an AFM-based pit-measuring method and, for the first time, used AFM to indirectly measure cell-surface membrane vesicles on cultured endothelial cells. Using this approach, we observed and quantitatively measured at least two populations of cell-surface membrane vesicles, a nanoscale population (<500 nm in diameter peaking at ∼250 nm) and a microscale population (from 500 nm to ∼2 μm peaking at ∼0.8 μm), whereas confocal microscopy only detected the microscale population. The AFM-based pit-measuring method is potentially useful for studying cell-surface membrane vesicles and for investigating the mechanisms of membrane vesicle formation/release

  12. Droplet Microarray Based on Patterned Superhydrophobic Surfaces Prevents Stem Cell Differentiation and Enables High-Throughput Stem Cell Screening.

    Science.gov (United States)

    Tronser, Tina; Popova, Anna A; Jaggy, Mona; Bastmeyer, Martin; Levkin, Pavel A

    2017-12-01

    Over the past decades, stem cells have attracted growing interest in fundamental biological and biomedical research as well as in regenerative medicine, due to their unique ability to self-renew and differentiate into various cell types. Long-term maintenance of the self-renewal ability and inhibition of spontaneous differentiation, however, still remain challenging and are not fully understood. Uncontrolled spontaneous differentiation of stem cells makes high-throughput screening of stem cells also difficult. This further hinders investigation of the underlying mechanisms of stem cell differentiation and the factors that might affect it. In this work, a dual functionality of nanoporous superhydrophobic-hydrophilic micropatterns is demonstrated in their ability to inhibit differentiation of mouse embryonic stem cells (mESCs) and at the same time enable formation of arrays of microdroplets (droplet microarray) via the effect of discontinuous dewetting. Such combination makes high-throughput screening of undifferentiated mouse embryonic stem cells possible. The droplet microarray is used to investigate the development, differentiation, and maintenance of stemness of mESC, revealing the dependence of stem cell behavior on droplet volume in nano- and microliter scale. The inhibition of spontaneous differentiation of mESCs cultured on the droplet microarray for up to 72 h is observed. In addition, up to fourfold increased cell growth rate of mESCs cultured on our platform has been observed. The difference in the behavior of mESCs is attributed to the porosity and roughness of the polymer surface. This work demonstrates that the droplet microarray possesses the potential for the screening of mESCs under conditions of prolonged inhibition of stem cells' spontaneous differentiation. Such a platform can be useful for applications in the field of stem cell research, pharmacological testing of drug efficacy and toxicity, biomedical research as well as in the field of

  13. Glucocorticoid-regulated and constitutive trafficking of proteolytically processed cell surface-associated glycoproteins in wild type and variant rat hepatoma cells

    International Nuclear Information System (INIS)

    Amacher, S.L.; Goodman, L.J.; Bravo, D.A.; Wong, K.Y.; Goldfine, I.D.; Hawley, D.M.; Firestone, G.L.

    1989-01-01

    Glucocorticoids regulate the trafficking of mouse mammary tumor virus (MMTV) glycoproteins to the cell surface in the rat hepatoma cell line M1.54, but not in the immunoselected sorting variant CR4. To compare the localization of MMTV glycoproteins to another proteolytically processed glycoprotein, both wild type M1.54 cells and variant CR4 cells were transfected with a human insulin receptor (hIR) expression vector, pRSVhIR. The production of cell surface hIR was monitored in dexamethasone-treated and -untreated wild type M1.54 and variant CR4 cells by indirect immunofluorescence, direct plasma membrane immunoprecipitation, and by [125I] insulin binding. In both wild type and variant rat hepatoma cells, hIR were localized at the cell surface in the presence or in the absence of 1 microM dexamethasone. In contrast, the glucocorticoid-regulated trafficking of cell surface MMTV glycoproteins occurred only in wild type M1.54 cells. We conclude that the hIR, which undergoes posttranslational processing reactions similar to MMTV glycoproteins, does not require glucocorticoids to be transported to the plasma membrane and is representative of a subset of cell surface glycoproteins whose trafficking is constitutive in rat hepatoma cells. Thus, MMTV glycoproteins and hIR provide specific cell surface markers to characterize the glucocorticoid-regulated and constitutive sorting pathways

  14. Germline cells in ovarian surface epithelium of mammalians: a promising notion

    Directory of Open Access Journals (Sweden)

    Celik Onder

    2012-12-01

    Full Text Available Abstract It is a long held doctrine in reproductive biology that women are born with a finite number of oocytes and there is no oogenesis during the postnatal period. However, recent evidence challenges this by showing the presence of germ line stem cells in the human ovarian surface epithelium (OSE, which can serve as a source of germ cells, and differentiate into oocyte like structures. Postnatal renewal of oocytes may have enormous therapeutic potential especially in women facing the risk of premature ovarian failure idiopathically or iatrogenically after exposure to gonadotoxic chemotherapy and radiation for cancer therapy. This article reviews current knowledge on germ line stem cells in human OSE.

  15. Surface-enhanced Raman spectroscopy of cell lysates mixed with silver nanoparticles for tumor classification

    Directory of Open Access Journals (Sweden)

    Mohamed Hassoun

    2017-06-01

    Full Text Available The throughput of spontaneous Raman spectroscopy for cell identification applications is limited to the range of one cell per second because of the relatively low sensitivity. Surface-enhanced Raman scattering (SERS is a widespread way to amplify the intensity of Raman signals by several orders of magnitude and, consequently, to improve the sensitivity and throughput. SERS protocols using immuno-functionalized nanoparticles turned out to be challenging for cell identification because they require complex preparation procedures. Here, a new SERS strategy is presented for cell classification using non-functionalized silver nanoparticles and potassium chloride to induce aggregation. To demonstrate the principle, cell lysates were prepared by ultrasonication that disrupts the cell membrane and enables interaction of released cellular biomolecules to nanoparticles. This approach was applied to distinguish four cell lines – Capan-1, HepG2, Sk-Hep1 and MCF-7 – using SERS at 785 nm excitation. Six independent batches were prepared per cell line to check the reproducibility. Principal component analysis was applied for data reduction and assessment of spectral variations that were assigned to proteins, nucleotides and carbohydrates. Four principal components were selected as input for classification models based on support vector machines. Leave-three-batches-out cross validation recognized four cell lines with sensitivities, specificities and accuracies above 96%. We conclude that this reproducible and specific SERS approach offers prospects for cell identification using easily preparable silver nanoparticles.

  16. Evaluation of single-cell force spectroscopy and fluorescence microscopy to determine cell interactions with femtosecond-laser microstructured titanium surfaces.

    Science.gov (United States)

    Aliuos, Pooyan; Fadeeva, Elena; Badar, Muhammad; Winkel, Andreas; Mueller, Peter P; Warnecke, Athanasia; Chichkov, Boris; Lenarz, Thomas; Reich, Uta; Reuter, Guenter

    2013-04-01

    One goal in biomaterials research is to limit the formation of connective tissue around the implant. Antiwetting surfaces are known to reduce ability of cells to adhere. Such surfaces can be achieved by special surface structures (lotus effect). Aim of the study was to investigate the feasibility for creating antiwetting surface structures on titanium and to characterize their effect on initial cell adhesion and proliferation. Titanium microstructures were generated using femtosecond- (fs-) laser pulses. Murine fibroblasts served as a model for connective tissue cells. Quantitative investigation of initial cell adhesion was performed using atomic force microscopy. Fluorescence microscopy was used for the characterization of cell-adhesion pattern, cell morphology, and proliferation. Water contact angle (WCA) measurements evinced antiwetting properties of laser-structured surfaces. However, the WCA was decreased in serum-containing medium. Initial cell adhesion to microstructured titanium was significantly promoted when compared with polished titanium. Microstructures did not influence cell proliferation on titanium surfaces. However, on titanium microstructures, cells showed a flattened morphology, and the cell orientation was biased according to the surface topography. In conclusion, antiwetting properties of surfaces were absent in the presence of serum and did not hinder adhesion and proliferation of NIH 3T3 fibroblasts. Copyright © 2012 Wiley Periodicals, Inc.

  17. 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.

  18. Roles for Cell Wall Glycopeptidolipid in Surface Adherence and Planktonic Dispersal of Mycobacterium avium

    Science.gov (United States)

    The opportunistic pathogen Mycobacterium avium is a significant inhabitant of biofilms in drinking water distribution systems. M. avium expresses on its cell surface serovar-specific glycopeptidolipids (ssGPLs). Studies have implicated the core GPL in biofilm formation by M. aviu...

  19. Microarray analysis of genes associated with cell surface NIS protein levels in breast cancer.

    Science.gov (United States)

    Beyer, Sasha J; Zhang, Xiaoli; Jimenez, Rafael E; Lee, Mei-Ling T; Richardson, Andrea L; Huang, Kun; Jhiang, Sissy M

    2011-10-11

    Na+/I- symporter (NIS)-mediated iodide uptake allows radioiodine therapy for thyroid cancer. NIS is also expressed in breast tumors, raising potential for radionuclide therapy of breast cancer. However, NIS expression in most breast cancers is low and may not be sufficient for radionuclide therapy. We aimed to identify biomarkers associated with NIS expression such that mechanisms underlying NIS modulation in human breast tumors may be elucidated. Published oligonucleotide microarray data within the National Center for Biotechnology Information Gene Expression Omnibus database were analyzed to identify gene expression tightly correlated with NIS mRNA level among human breast tumors. NIS immunostaining was performed in a tissue microarray composed of 28 human breast tumors which had corresponding oligonucleotide microarray data available for each tumor such that gene expression associated with cell surface NIS protein level could be identified. NIS mRNA levels do not vary among breast tumors or when compared to normal breast tissues when detected by Affymetrix oligonucleotide microarray platforms. Cell surface NIS protein levels are much more variable than their corresponding NIS mRNA levels. Despite a limited number of breast tumors examined, our analysis identified cysteinyl-tRNA synthetase as a biomarker that is highly associated with cell surface NIS protein levels in the ER-positive breast cancer subtype. Further investigation on genes associated with cell surface NIS protein levels within each breast cancer molecular subtype may lead to novel targets for selectively increasing NIS expression/function in a subset of breast cancers patients.

  20. Effect of Growth Conditions on Flocculation and Cell Surface Hydrophobicity of Brewing Yeast

    Czech Academy of Sciences Publication Activity Database

    Kopecká, J.; Němec, M.; Matoulková, D.; Čejka, P.; Jelínková, Markéta; Felsberg, Jürgen; Sigler, Karel

    2015-01-01

    Roč. 73, č. 2 (2015), s. 143-150 ISSN 0361-0470 Institutional support: RVO:61388971 Keywords : Ale and lager yeast * Cell surface hydrophobicity * FLO genes Subject RIV: EI - Biotechnology ; Bionics Impact factor: 0.492, year: 2015

  1. Bioinspired superhydrophobic poly(L-lactic acid) surfaces control bone marrow derived cells adhesion and proliferation.

    Science.gov (United States)

    Alves, Natália M; Shi, Jun; Oramas, Elena; Santos, José L; Tomás, Helena; Mano, João F

    2009-11-01

    The aptitude of a cell to adhere, migrate, and differentiate on a compact substrate or scaffold is important in the field of tissue engineering and biomaterials. It is well known that cell behavior can be controlled and guided through the change in micro- and nano-scale topographic features. In this work, we intend to demonstrate that special topographic features that control wettability may also have an important role in the biological performance of biodegradable substrates. Poly(L-lactic acid) surfaces with superhydrophobic characteristics were produced, based on the so-called Lotus effect, exhibiting dual micro- and nano-scale roughness. The water contact angle could be higher than 150 degrees and a value of that order could be kept even upon immersion in a simulated body fluid solution for more than 20 days. Such water repellent surfaces were found to prevent adhesion and proliferation of bone marrow derived cells previously isolated from the femurs of 6-week-old male Wistar rats, when compared with smoother surfaces prepared by simple solvent casting. Such results demonstrate that these superhydrophobic surfaces may be used to control cell behavior onto biodegradable substrates. (c) 2008 Wiley Periodicals, Inc.

  2. Combined Contamination and Space Environmental Effects on Solar Cells and Thermal Control Surfaces

    Science.gov (United States)

    Dever, Joyce A.; Bruckner, Eric J.; Scheiman, David A.; Stidham, Curtis R.

    1994-01-01

    For spacecraft in low Earth orbit (LEO), contamination can occur from thruster fuel, sputter contamination products and from products of silicone degradation. This paper describes laboratory testing in which solar cell materials and thermal control surfaces were exposed to simulated spacecraft environmental effects including contamination, atomic oxygen, ultraviolet radiation and thermal cycling. The objective of these experiments was to determine how the interaction of the natural LEO environmental effects with contaminated spacecraft surfaces impacts the performance of these materials. Optical properties of samples were measured and solar cell performance data was obtained. In general, exposure to contamination by thruster fuel resulted in degradation of solar absorptance for fused silica and various thermal control surfaces and degradation of solar cell performance. Fused silica samples which were subsequently exposed to an atomic oxygen/vacuum ultraviolet radiation environment showed reversal of this degradation. These results imply that solar cells and thermal control surfaces which are susceptible to thruster fuel contamination and which also receive atomic oxygen exposure may not undergo significant performance degradation. Materials which were exposed to only vacuum ultraviolet radiation subsequent to contamination showed slight additional degradation in solar absorptance.

  3. Interaction of human mesenchymal stem cells with osteopontin coated hydroxyapatite surfaces

    DEFF Research Database (Denmark)

    Jensen, Thomas; Dolatshahi-Pirouz, Alireza; Foss, Morten

    2010-01-01

    In vitro studies of the initial attachment, spreading and motility of human bone mesenchymal stem cells have been carried out on bovine osteopontin (OPN) coated hydroxyapatite (HA) and gold (Au) model surfaces. The adsorption of OPN extracted from bovine milk was monitored by the quartz crystal...

  4. Reduced PDZ Interactions of Rescued ΔF508CFTR Increases Its Cell Surface Mobility*

    Science.gov (United States)

    Valentine, Cathleen D.; Lukacs, Gergely L.; Verkman, Alan S.; Haggie, Peter M.

    2012-01-01

    Deletion of phenylalanine 508 (ΔF508) in the cystic fibrosis transmembrane conductance regulator (CFTR) plasma membrane chloride channel is the most common cause of cystic fibrosis (CF). Though several maneuvers can rescue endoplasmic reticulum-retained ΔF508CFTR and promote its trafficking to the plasma membrane, rescued ΔF508CFTR remains susceptible to quality control mechanisms that lead to accelerated endocytosis, ubiquitination, and lysosomal degradation. To investigate the role of scaffold protein interactions in rescued ΔF508CFTR surface instability, the plasma membrane mobility of ΔF508CFTR was measured in live cells by quantum dot single particle tracking. Following rescue by low temperature, chemical correctors, thapsigargin, or overexpression of GRASP55, ΔF508CFTR diffusion was more rapid than that of wild-type CFTR because of reduced interactions with PDZ domain-containing scaffold proteins. Knock-down of the plasma membrane quality control proteins CHIP and Hsc70 partially restored ΔF508CFTR-scaffold association. Quantitative comparisons of CFTR cell surface diffusion and endocytosis kinetics suggested an association between reduced scaffold binding and CFTR internalization. Our surface diffusion measurements in live cells indicate defective scaffold interactions of rescued ΔF508CFTR at the cell surface, which may contribute to its defective peripheral processing. PMID:23115232

  5. Simultaneous interaction of bacteria and tissue cells with photocatalytically activated, anodized titanium surfaces

    NARCIS (Netherlands)

    Yue, Chongxia; Kuijer, Roelof; Kaper, H. J.; van der Mei, Henderina; Busscher, Hendrik; Kuijer, Roelof

    Photocatalytic-activation of anodized TiO2-surfaces has been demonstrated to yield antibacterial and tissue integrating effects, but effects on simultaneous growth of tissue cells and bacteria in co-culture have never been studied. Moreover, it is unknown how human-bone-marrow-mesenchymal-stem

  6. Growth condition-dependent cell surface proteome analysis of Enterococcus faecium

    NARCIS (Netherlands)

    Sinnige, Jan C; de Been, Mark; Zhou, Miaomiao; Bonten, Marc J M; Willems, Rob J L; Top, Janetta

    2015-01-01

    The last 30 years Enterococcus faecium has become an important nosocomial pathogen in hospitals worldwide. The aim of this study was to obtain insight in the cell surface proteome of E. faecium when grown in laboratory and clinically relevant conditions. Enterococcus faecium E1162, a clinical blood

  7. Illuminated up close: near-field optical microscopy of cell surfaces.

    Science.gov (United States)

    Czajkowsky, Daniel M; Sun, Jielin; Shao, Zhifeng

    2015-01-01

    Invented in the 1990s, near-field optical microscopy (NSOM) was the first optical microscopy method to hold the promise of finally breaking the diffraction barrier in studies of biological samples. This promise, though, failed to materialize at that time, largely owing to the inability to image soft samples, such as cell surfaces, without damage. However, steady technical improvements have now produced NSOM devices that can routinely achieve images of cell surfaces with sub-100nm resolution in aqueous solution. Further, beyond just optical information, these instruments can also provide simultaneous topographic, mechanical, and/or chemical details of the sample, an ability not yet matched by any other optics-based methodology. With the long recognized important roles of many biological processes at cell surfaces in human health and disease, near-field probing of cell surfaces is indeed now well poised to directly illume in biomedicine what has, until recently, been unknowable with classic light microscopy. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Comprehensive proteomic analysis of Trypanosoma cruzi epimastigote cell surface proteins by two complementary methods

    DEFF Research Database (Denmark)

    Queiroz, Rayner M L; Charneau, Sébastien; Motta, Flávia N

    2013-01-01

    Trypanosoma cruzi is a protozoan that causes Chagas' disease, a neglected infectious illness that affects millions of people, mostly in Latin America. Here, the cell surface subproteome of the T. cruzi epimastigote life form was characterized. In order to prepare samples enriched in epimastigote...

  9. Cell Wall-anchored Proteins of Enterococcus faecium: Exploring a Novel Surface

    NARCIS (Netherlands)

    Hendrickx, A.P.A.|info:eu-repo/dai/nl/304820741

    2009-01-01

    The past 4 years my research focussed on the identification, expression and function of surface-exposed LPXTG proteins and filamentous structures (also called pili or fimbriae) at the Enterococcus faecium cell wall. E. faecium is a commensal organism of the mammalian gastrointestinal tract, but the

  10. IMPLICATIONS OF MICROBIAL ADHESION TO HYDROCARBONS FOR EVALUATING CELL-SURFACE HYDROPHOBICITY .2. ADHESION MECHANISMS

    NARCIS (Netherlands)

    VANDERMEI, HC; VANDEBELTGRITTER, B; BUSSCHER, HJ

    1995-01-01

    Microbial adhesion to hydrocarbons (MATH) is generally considered to be a measure of the organisms cell surface hydrophobicity. Recent observations that the zeta potentials of hydrocarbons can be highly negative in the various solutions commonly used in MATH, have suggested that MATH may measure a

  11. Stem Cell Surface Marker Expression Defines Late Stages of Reprogramming to Pluripotency in Human Fibroblasts.

    Science.gov (United States)

    Pomeroy, Jordan E; Hough, Shelley R; Davidson, Kathryn C; Quaas, Alex M; Rees, Jordan A; Pera, Martin F

    2016-07-01

    Our current understanding of the induction of pluripotency by defined factors indicates that this process occurs in discrete stages characterized by specific alterations in the cellular transcriptome and epigenome. However, the final phase of the reprogramming process is incompletely understood. We sought to generate tools to characterize the transition to a fully reprogramed state. We used combinations of stem cell surface markers to isolate colonies emerging after transfection of human fibroblasts with reprogramming factors and then analyzed their expression of genes associated with pluripotency and early germ lineage specification. We found that expression of a subset of these genes, including the cell-cell adhesion molecule CDH3, characterized a late stage in the reprogramming process. Combined live-cell staining with the antibody GCTM-2 and anti-CDH3 during reprogramming identified colonies of cells that showed gene expression patterns very similar to those of embryonic stem cell or established induced pluripotent stem cell lines, and gave rise to stable induced pluripotent stem cell lines at high frequency. Our findings will facilitate studies of the final stages of reprogramming of human cells to pluripotency and will provide a simple means for prospective identification of fully reprogrammed cells. Reprogramming of differentiated cells back to an embryonic pluripotent state has wide ranging applications in understanding and treating human disease. However, how cells traverse the barriers on the journey to pluripotency still is not fully understood. This report describes tools to study the late stages of cellular reprogramming. The findings enable a more precise approach to dissecting the final phases of conversion to pluripotency, a process that is particularly poorly defined. The results of this study also provide a simple new method for the selection of fully reprogrammed cells, which could enhance the efficiency of derivation of cell lines for research

  12. Superhydrophilic-Superhydrophobic Patterned Surfaces as High-Density Cell Microarrays: Optimization of Reverse Transfection.

    Science.gov (United States)

    Ueda, Erica; Feng, Wenqian; Levkin, Pavel A

    2016-10-01

    High-density microarrays can screen thousands of genetic and chemical probes at once in a miniaturized and parallelized manner, and thus are a cost-effective alternative to microwell plates. Here, high-density cell microarrays are fabricated by creating superhydrophilic-superhydrophobic micropatterns in thin, nanoporous polymer substrates such that the superhydrophobic barriers confine both aqueous solutions and adherent cells within each superhydrophilic microspot. The superhydrophobic barriers confine and prevent the mixing of larger droplet volumes, and also control the spreading of droplets independent of the volume, minimizing the variability that arises due to different liquid and surface properties. Using a novel liposomal transfection reagent, ScreenFect A, the method of reverse cell transfection is optimized on the patterned substrates and several factors that affect transfection efficiency and cytotoxicity are identified. Higher levels of transfection are achieved on HOOC- versus NH 2 -functionalized superhydrophilic spots, as well as when gelatin and fibronectin are added to the transfection mixture, while minimizing the amount of transfection reagent improves cell viability. Almost no diffusion of the printed transfection mixtures to the neighboring microspots is detected. Thus, superhydrophilic-superhydrophobic patterned surfaces can be used as cell microarrays and for optimizing reverse cell transfection conditions before performing further cell screenings. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Cell adhesion, morphology and bioche