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Sample records for adhesion cell proliferation

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

    NASA Astrophysics Data System (ADS)

    Castellanos, Maria Isabel; Mas-Moruno, Carlos; Grau, Anna; Serra-Picamal, Xavier; Trepat, Xavier; Albericio, Fernando; Joner, Michael; Gil, Francisco Javier; Ginebra, Maria Pau; Manero, Jose María; Pegueroles, Marta

    2017-01-01

    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.

  2. Cell adhesion and proliferation on polyethylene grafted with Au nanoparticles

    NASA Astrophysics Data System (ADS)

    Kasálková, N. Slepičková; Slepička, P.; Kolská, Z.; Sajdl, P.; Bačáková, L.; Rimpelová, S.; Švorčík, V.

    2012-02-01

    Plasma treatment and subsequent Au nano-particles grafting of polyethylene (PE) lead to changes in surface morphology, roughness and wettability, significantly increasing the attractiveness of the material for cells. The PE samples were exposed to argon plasma. Plasma modified PE was chemically grafted by immersion to biphenyldithiol and consequently into solution of Au nano-particles. Changes in chemical structure of the modified PE were studied using X-ray Photoelectron Spectroscopy (XPS) and electrokinetic analysis ( ζ-potential). The surface wettability of the modified PE samples was examined by measurement of the contact angle by standard goniometry. The surface morphology of the plasma modified PE and that grafted with Au nano-particles was studied by Atomic Force Microscopy (AFM). The modified PE samples were seeded with rat vascular smooth muscle cells (VSMCs) and their adhesion and proliferation were studied. Chemically bounded biphenyldithiol increases the number of the incorporated gold nano-particles and changes sample surface properties. The presence of the biphenyldithiol and the gold nano-particles on the PE surface influences dramatically adhesion and proliferation of VSMCs.

  3. Endothelial cell responses in terms of adhesion, proliferation, and morphology to stiffness of polydimethylsiloxane elastomer substrates.

    PubMed

    Ataollahi, Forough; Pramanik, Sumit; Moradi, Ali; Dalilottojari, Adel; Pingguan-Murphy, Belinda; Wan Abas, Wan Abu Bakar; Abu Osman, Noor Azuan

    2015-07-01

    Extracellular environments can regulate cell behavior because cells can actively sense their mechanical environments. This study evaluated the adhesion, proliferation and morphology of endothelial cells on polydimethylsiloxane (PDMS)/alumina (Al2 O3 ) composites and pure PDMS. The substrates were prepared from pure PDMS and its composites with 2.5, 5, 7.5, and 10 wt % Al2 O3 at a curing temperature of 50°C for 4 h. The substrates were then characterized by mechanical, structural, and morphological analyses. The cell adhesion, proliferation, and morphology of cultured bovine aortic endothelial (BAEC) cells on substrate materials were evaluated by using resazurin assay and 1,1'-dioctadecyl-1,3,3,3',3'-tetramethylindocarbocyanine perchlorate-acetylated LDL (Dil-Ac-LDL) cell staining, respectively. The composites (PDMS/2.5, 5, 7.5, and 10 wt % Al2 O3 ) exhibited higher stiffness than the pure PDMS substrate. The results also revealed that stiffer substrates promoted endothelial cell adhesion and proliferation and also induced spread morphology in the endothelial cells compared with lesser stiff substrates. Statistical analysis showed that the effect of time on cell proliferation depended on stiffness. Therefore, this study concludes that the addition of different Al2 O3 percentages to PDMS elevated substrate stiffness which in turn increased endothelial cell adhesion and proliferation significantly and induced spindle shape morphology in endothelial cells.

  4. Effects of SOX2 on Proliferation, Migration and Adhesion of Human Dental Pulp Stem Cells.

    PubMed

    Liu, Pengfei; Cai, Jinglei; Dong, Delu; Chen, Yaoyu; Liu, Xiaobo; Wang, Yi; Zhou, Yulai

    2015-01-01

    As a key factor for cell pluripotent and self-renewing phenotypes, SOX2 has attracted scientists' attention gradually in recent years. However, its exact effects in dental pulp stem cells (DPSCs) are still unclear. In this study, we mainly investigated whether SOX2 could affect some biological functions of DPSCs. DPSCs were isolated from the dental pulp of human impacted third molar. SOX2 overexpressing DPSCs (DPSCs-SOX2) were established through retroviral infection. The effect of SOX2 on cell proliferation, migration and adhesion ability was evaluated with CCK-8, trans-well system and fibronectin-induced cell attachment experiment respectively. Whole genome expression of DPSCs-SOX2 was analyzed with RNA microarray. Furthermore, a rescue experiment was performed with SOX2-siRNA in DPSC-SOX2 to confirm the effect of SOX2 overexpression in DPSCs. We found that SOX2 overexpression could result in the enhancement of cell proliferation, migration, and adhesion in DPSCs obviously. RNA microarray analysis indicated that some key genes in the signal pathways associated with cell cycle, migration and adhesion were upregulated in different degree, and the results were further confirmed with qPCR and western-blot. Finally, DPSC-SOX2 transfected with SOX2-siRNA showed a decrease of cell proliferation, migration and adhesion ability, which further confirmed the biological effect of SOX2 in human DPSCs. This study indicated that SOX2 could improve the cell proliferation, migration and adhesion ability of DPSCs through regulating gene expression about cell cycle, migration and adhesion, and provided a novel strategy to develop seed cells with strong proliferation, migration and adhesion ability for tissue engineering.

  5. Adhesion and proliferation of human mesenchymal stem cells from dental pulp on porous silicon scaffolds.

    PubMed

    Collart-Dutilleul, Pierre-Yves; Secret, Emilie; Panayotov, Ivan; Deville de Périère, Dominique; Martín-Palma, Raúl J; Torres-Costa, Vicente; Martin, Marta; Gergely, Csilla; Durand, Jean-Olivier; Cunin, Frédérique; Cuisinier, Frédéric J

    2014-02-12

    In regenerative medicine, stem-cell-based therapy often requires a scaffold to deliver cells and/or growth factors to the injured site. Porous silicon (pSi) is a promising biomaterial for tissue engineering as it is both nontoxic and bioresorbable. Moreover, surface modification can offer control over the degradation rate of pSi and can also promote cell adhesion. Dental pulp stem cells (DPSC) are pluripotent mesenchymal stem cells found within the teeth and constitute a readily source of stem cells. Thus, coupling the good proliferation and differentiation capacities of DPSC with the textural and chemical properties of the pSi substrates provides an interesting approach for therapeutic use. In this study, the behavior of human DPSC is analyzed on pSi substrates presenting pores of various sizes, 10 ± 2 nm, 36 ± 4 nm, and 1.0 ± 0.1 μm, and undergoing different chemical treatments, thermal oxidation, silanization with aminopropyltriethoxysilane (APTES), and hydrosilylation with undecenoic acid or semicarbazide. DPSC adhesion and proliferation were followed for up to 72 h by fluorescence microscopy, scanning electron microscopy (SEM), enzymatic activity assay, and BrdU assay for mitotic activity. Porous silicon with 36 nm pore size was found to offer the best adhesion and the fastest growth rate for DPSC compared to pSi comporting smaller pore size (10 nm) or larger pore size (1 μm), especially after silanization with APTES. Hydrosilylation with semicarbazide favored cell adhesion and proliferation, especially mitosis after cell adhesion, but such chemical modification has been found to led to a scaffold that is stable for only 24-48 h in culture medium. Thus, semicarbazide-treated pSi appeared to be an appropriate scaffold for stem cell adhesion and immediate in vivo transplantation, whereas APTES-treated pSi was found to be more suitable for long-term in vitro culture, for stem cell proliferation and differentiation.

  6. Surface modifications of photocrosslinked biodegradable elastomers and their influence on smooth muscle cell adhesion and proliferation.

    PubMed

    Ilagan, Bernadette G; Amsden, Brian G

    2009-09-01

    Photocrosslinked, biodegradable elastomers based on aliphatic polyesters have many desirable features as scaffolds for smooth muscle tissue engineering. However, they lack cell adhesion motifs. To address this shortcoming, two different modification procedures were studied utilizing a high and a low crosslink density elastomer: base etching and the incorporation of acryloyl-poly(ethylene glycol) (PEG)-Gly-Arg-Gly-Asp-Ser (GRGDS) into the elastomer network during photocrosslinking. Base etching improved surface hydrophilicity without altering surface topography, but did not improve bovine aortic smooth muscle cell adhesion. Incorporation of PEG-GRGDS into the elastomer network significantly improved cell adhesion for both high and low crosslink density elastomers, with a greater effect with the higher crosslink density elastomer. Incorporation of GRGDS into the high crosslink density elastomer also enhanced smooth muscle cell proliferation, while proliferation on the low crosslink density unmodified, base etched, and PEG-GRGDS incorporated elastomers was significantly greater than on the high crosslink density unmodified and base etched elastomer.

  7. Hyaluronan synthase 3 (HAS3) overexpression downregulates MV3 melanoma cell proliferation, migration and adhesion

    SciTech Connect

    Takabe, Piia; Bart, Geneviève; Ropponen, Antti; Rilla, Kirsi; Tammi, Markku; Tammi, Raija; Pasonen-Seppänen, Sanna

    2015-09-10

    Malignant skin melanoma is one of the most deadly human cancers. Extracellular matrix (ECM) influences the growth of malignant tumors by modulating tumor cells adhesion and migration. Hyaluronan is an essential component of the ECM, and its amount is altered in many tumors, suggesting an important role for hyaluronan in tumorigenesis. Nonetheless its role in melanomagenesis is not understood. In this study we produced a MV3 melanoma cell line with inducible expression of the hyaluronan synthase 3 (HAS3) and studied its effect on the behavior of the melanoma cells. HAS3 overexpression expanded the cell surface hyaluronan coat and decreased melanoma cell adhesion, migration and proliferation by cell cycle arrest at G1/G0. Melanoma cell migration was restored by removal of cell surface hyaluronan by Streptomyces hyaluronidase and by receptor blocking with hyaluronan oligosaccharides, while the effect on cell proliferation was receptor independent. Overexpression of HAS3 decreased ERK1/2 phosphorylation suggesting that inhibition of MAP-kinase signaling was responsible for these suppressive effects on the malignant phenotype of MV3 melanoma cells. - Highlights: • Inducible HAS3-MV3 melanoma cell line was generated using Lentiviral transduction. • HAS3 overexpression inhibits MV3 cell migration via hyaluronan–receptor interaction. • HAS3 overexpression decreases MV3 melanoma cell proliferation and adhesion. • ERK1/2 phosphorylation is downregulated by 50% in HAS3 overexpressing cells. • The results suggest that hyaluronan has anti-cancer like effects in melanoma.

  8. Cell Adhesion and Proliferation on Sulfonated and Non-Modified Chitosan Films.

    PubMed

    Martínez-Campos, Enrique; Civantos, Ana; Redondo, Juan Alfonso; Guzmán, Rodrigo; Pérez-Perrino, Mónica; Gallardo, Alberto; Ramos, Viviana; Aranaz, Inmaculada

    2016-09-15

    Three types of chitosan-based films have been prepared and evaluated: a non-modified chitosan film bearing cationizable aliphatic amines and two films made of N-sulfopropyl chitosan derivatives bearing both aliphatic amines and negative sulfonate groups at different ratios. Cell adhesion and proliferation on chitosan films of C2C12 pre-myoblastic cells and B16 cells as tumoral model have been tested. A differential cell behavior has been observed on chitosan films due to their different surface modification. B16 cells have shown lower vinculin expression when cultured on sulfonated chitosan films. This study shows how the interaction among cells and material surface can be modulated by physicochemical characteristics of the biomaterial surface, altering tumoral cell adhesion and proliferation processes.

  9. Receptor FGFRL1 does not promote cell proliferation but induces cell adhesion

    PubMed Central

    YANG, XIAOCHEN; STEINBERG, FLORIAN; ZHUANG, LEI; BESSEY, RALPH; TRUEB, BEAT

    2016-01-01

    Fibroblast growth factor receptor (FGFR)-like protein 1 (FGFRL1) is the most recently discovered member of the FGFR family. Owing to the fact that it interacts with FGF ligands, but lacks the intracellular tyrosine kinase domain, several researchers have speculated that it may function as a decoy receptor and exert a negative effect on cell proliferation. In this study, we performed overexpression experiments with TetOn-inducible cell clones and downregulation experiments with siRNA oligonucleotides, and found that FGFRL1 had absolutely no effect on cell growth and proliferation. Likewise, we did not observe any influence of FGFRL1 on ERK1/2 activation and on the phosphorylation of 250 other signaling proteins analyzed by the Kinexus antibody microarray. On the other hand, with bacterial petri dishes, we observed a clear effect of FGFRL1 on cell adhesion during the initial hours after cell seeding. Our results suggest that FGFRL1 is a cell adhesion protein similar to the nectins rather than a signaling receptor similar to FGFR1-FGFR4. PMID:27220341

  10. Bio-active molecules modified surfaces enhanced mesenchymal stem cell adhesion and proliferation.

    PubMed

    Mobasseri, Rezvan; Tian, Lingling; Soleimani, Masoud; Ramakrishna, Seeram; Naderi-Manesh, Hossein

    2017-01-29

    Surface modification of the substrate as a component of in vitro cell culture and tissue engineering, using bio-active molecules including extracellular matrix (ECM) proteins or peptides derived ECM proteins can modulate the surface properties and thereby induce the desired signaling pathways in cells. The aim of this study was to evaluate the behavior of human bone marrow mesenchymal stem cells (hBM-MSCs) on glass substrates modified with fibronectin (Fn), collagen (Coll), RGD peptides (RGD) and designed peptide (R-pept) as bio-active molecules. The glass coverslips were coated with fibronectin, collagen, RGD peptide and R-peptide. Bone marrow mesenchymal stem cells were cultured on different substrates and the adhesion behavior in early incubation times was investigated using scanning electron microscopy (SEM) and confocal microscopy. The MTT assay was performed to evaluate the effect of different bio-active molecules on MSCs proliferation rate during 24 and 72 h. Formation of filopodia and focal adhesion (FA) complexes, two steps of cell adhesion process, were observed in MSCs cultured on bio-active molecules modified coverslips, specifically in Fn coated and R-pept coated groups. SEM image showed well adhesion pattern for MSCs cultured on Fn and R-pept after 2 h incubation, while the shape of cells cultured on Coll and RGD substrates indicated that they might experience stress condition in early hours of culture. Investigation of adhesion behavior, as well as proliferation pattern, suggests R-peptide as a promising bio-active molecule to be used for surface modification of substrate in supporting and inducing cell adhesion and proliferation.

  11. Cholic acid functionalized star poly(DL-lactide) for promoting cell adhesion and proliferation.

    PubMed

    Fu, Hui-Li; Zou, Tao; Cheng, Si-Xue; Zhang, Xian-Zheng; Zhuo, Ren-Xi

    2007-01-01

    Cholic acid functionalized star poly(DL-lactide) was synthesized through the ring-opening polymerization of DL-lactide initiated by cholic acid. The properties and cell behaviour of the cholic acid functionalized star poly(DL-lactide) were investigated as compared with linear poly(DL-lactide)s with different molecular weights and a star poly(DL-lactide) initiated by glycerol. In comparison to linear poly(DL-lactide)s, the cholic acid functionalized star poly(DL-lactide) had better wettability and slightly higher surface energy. The cell adhesion and proliferation on different materials were evaluated using two types of cells, 3T3 mouse fibroblasts and ECV304 human endothelial cells. Compared with the linear poly(DL-lactide)s, the cholic acid functionalized star poly(DL-lactide) showed obviously improved property for cell adhesion. The cell proliferation on the cholic acid functionalized star poly(DL-lactide) was also enhanced. The improvement in cell proliferation was not so significant as compared with the improvement in cell adhesion. This modification strategy provides an effective and simple way to promote cell attachment and growth in tissue engineering.

  12. Effect of surface potential on epithelial cell adhesion, proliferation and morphology.

    PubMed

    Chang, Hsun-Yun; Kao, Wei-Lun; You, Yun-Wen; Chu, Yi-Hsuan; Chu, Kuo-Jui; Chen, Peng-Jen; Wu, Chen-Yi; Lee, Yu-Hsuan; Shyue, Jing-Jong

    2016-05-01

    Cell adhesion is the basis of individual cell survival, division and motility. Hence, understanding the effects that the surface properties have on cell adhesion, proliferation and morphology are crucial. In particular, surface charge/potential has been identified as an important factor that affects cell behavior. However, how cells respond to incremental changes in surface potential remains unclear. By using binary self-assembled monolayer (SAM) modified Au surfaces that are similar in mechanical/chemical properties and provide a series of surface potentials, the effect of surface potential on the behavior of cells can be studied. In this work, the effect of surface potential on epithelial cells, including human embryonic kidney (HEK293T) and human hepatocellular carcinoma (HepG2), were examined. The results showed that the adhesion density of epithelial cells increased with increasing surface potential, which is similar to but varied more significantly compared with fibroblasts. The proliferation rate is found to be independent of surface potential in both cell types. Furthermore, epithelial cells show no morphological change with respect to surface potential, whereas the morphology of the fibroblasts clearly changed with the surface potential. These differences between the cell types were rationalized by considering the difference in extracellular matrix composition. Laminin-dominant epithelial cells showed higher adhesion density and less morphological change than did fibronectin-dominant fibroblasts because the more significant adsorption of positively charged laminin on the surface enhanced the adhesion of epithelial cells. In contrast, due to the dominance of negatively charged fibronectin that adsorbed weakly on the surface, fibroblasts had to change their morphology to fit the inhomogeneous fibronectin-adsorbed area.

  13. Expression of the melanoma cell adhesion molecule in human mesenchymal stromal cells regulates proliferation, differentiation, and maintenance of hematopoietic stem and progenitor cells.

    PubMed

    Stopp, Sabine; Bornhäuser, Martin; Ugarte, Fernando; Wobus, Manja; Kuhn, Matthias; Brenner, Sebastian; Thieme, Sebastian

    2013-04-01

    The melanoma cell adhesion molecule defines mesenchymal stromal cells in the human bone marrow that regenerate bone and establish a hematopoietic microenvironment in vivo. The role of the melanoma cell adhesion molecule in primary human mesenchymal stromal cells and the maintenance of hematopoietic stem and progenitor cells during ex vivo culture has not yet been demonstrated. We applied RNA interference or ectopic overexpression of the melanoma cell adhesion molecule in human mesenchymal stromal cells to evaluate the effect of the melanoma cell adhesion molecule on their proliferation and differentiation as well as its influence on co-cultivated hematopoietic stem and progenitor cells. Knockdown and overexpression of the melanoma cell adhesion molecule affected several characteristics of human mesenchymal stromal cells related to osteogenic differentiation, proliferation, and migration. Furthermore, knockdown of the melanoma cell adhesion molecule in human mesenchymal stromal cells stimulated the proliferation of hematopoietic stem and progenitor cells, and strongly reduced the formation of long-term culture-initiating cells. In contrast, melanoma cell adhesion molecule-overexpressing human mesenchymal stromal cells provided a supportive microenvironment for hematopoietic stem and progenitor cells. Expression of the melanoma cell adhesion molecule increased the adhesion of hematopoietic stem and progenitor cells to human mesenchymal stromal cells and their migration beneath the monolayer of human mesenchymal stromal cells. Our results demonstrate that the expression of the melanoma cell adhesion molecule in human mesenchymal stromal cells determines their fate and regulates the maintenance of hematopoietic stem and progenitor cells through direct cell-cell contact.

  14. Critical role of heparin binding domains of ameloblastin for dental epithelium cell adhesion and ameloblastoma proliferation.

    PubMed

    Sonoda, Akira; Iwamoto, Tsutomu; Nakamura, Takashi; Fukumoto, Emiko; Yoshizaki, Keigo; Yamada, Aya; Arakaki, Makiko; Harada, Hidemitsu; Nonaka, Kazuaki; Nakamura, Seiji; Yamada, Yoshihiko; Fukumoto, Satoshi

    2009-10-02

    AMBN (ameloblastin) is an enamel matrix protein that regulates cell adhesion, proliferation, and differentiation of ameloblasts. In AMBN-deficient mice, ameloblasts are detached from the enamel matrix, continue to proliferate, and form a multiple cell layer; often, odontogenic tumors develop in the maxilla with age. However, the mechanism of AMBN functions in these biological processes remains unclear. By using recombinant AMBN proteins, we found that AMBN had heparin binding domains at the C-terminal half and that these domains were critical for AMBN binding to dental epithelial cells. Overexpression of full-length AMBN protein inhibited proliferation of human ameloblastoma AM-1 cells, but overexpression of heparin binding domain-deficient AMBN protein had no inhibitory effect. In full-length AMBN-overexpressing AM-1 cells, the expression of Msx2, which is involved in the dental epithelial progenitor phenotype, was decreased, whereas the expression of cell proliferation inhibitors p21 and p27 was increased. We also found that the expression of enamelin, a marker of differentiated ameloblasts, was induced, suggesting that AMBN promotes odontogenic tumor differentiation. Thus, our results suggest that AMBN promotes cell binding through the heparin binding sites and plays an important role in preventing odontogenic tumor development by suppressing cell proliferation and maintaining differentiation phenotype through Msx2, p21, and p27.

  15. Toward Cell Selective Surfaces: Cell Adhesion and Proliferation on Breath Figures with Antifouling Surface Chemistry.

    PubMed

    Martínez-Campos, Enrique; Elzein, Tamara; Bejjani, Alice; García-Granda, Maria Jesús; Santos-Coquillat, Ana; Ramos, Viviana; Muñoz-Bonilla, Alexandra; Rodríguez-Hernández, Juan

    2016-03-01

    We report the preparation of microporous functional polymer surfaces that have been proven to be selective surfaces toward eukaryotic cells while maintaining antifouling properties against bacteria. The fabrication of functional porous films has been carried out by the breath figures approach that allowed us to create porous interfaces with either poly(ethylene glycol) methyl ether methacrylate (PEGMA) or 2,3,4,5,6-pentafluorostyrene (5FS). For this purpose, blends of block copolymers in a polystyrene homopolymer matrix have been employed. In contrast to the case of single functional polymer, using blends enables us to vary the chemical distribution of the functional groups inside and outside the formed pores. In particular, fluorinated groups were positioned at the edges while the hydrophilic PEGMA groups were selectively located inside the pores, as demonstrated by TOF-SIMS. More interestingly, studies of cell adhesion, growth, and proliferation on these surfaces confirmed that PEGMA functionalized interfaces are excellent candidates to selectively allow cell growth and proliferation while maintaining antifouling properties.

  16. Fucosyltransferase 1 mediates angiogenesis, cell adhesion and rheumatoid arthritis synovial tissue fibroblast proliferation

    PubMed Central

    2014-01-01

    Introduction We previously reported that sialyl Lewisy, synthesized by fucosyltransferases, is involved in angiogenesis. Fucosyltransferase 1 (fut1) is an α(1,2)-fucosyltransferase responsible for synthesis of the H blood group and Lewisy antigens. However, the angiogenic involvement of fut 1 in the pathogenesis of rheumatoid arthritis synovial tissue (RA ST) has not been clearly defined. Methods Assay of α(1,2)-linked fucosylated proteins in RA was performed by enzyme-linked lectin assay. Fut1 expression was determined in RA ST samples by immunohistological staining. We performed angiogenic Matrigel assays using a co-culture system of human dermal microvascular endothelial cells (HMVECs) and fut1 small interfering RNA (siRNA) transfected RA synovial fibroblasts. To determine if fut1 played a role in leukocyte retention and cell proliferation in the RA synovium, myeloid THP-1 cell adhesion assays and fut1 siRNA transfected RA synovial fibroblast proliferation assays were performed. Results Total α(1,2)-linked fucosylated proteins in RA ST were significantly higher compared to normal (NL) ST. Fut1 expression on RA ST lining cells positively correlated with ST inflammation. HMVECs from a co-culture system with fut1 siRNA transfected RA synovial fibroblasts exhibited decreased endothelial cell tube formation compared to control siRNA transfected RA synovial fibroblasts. Fut1 siRNA also inhibited myeloid THP-1 adhesion to RA synovial fibroblasts and RA synovial fibroblast proliferation. Conclusions These data show that α(1,2)-linked fucosylated proteins are upregulated in RA ST compared to NL ST. We also show that fut1 in RA synovial fibroblasts is important in angiogenesis, leukocyte-synovial fibroblast adhesion, and synovial fibroblast proliferation, all key processes in the pathogenesis of RA. PMID:24467809

  17. Hyaluronan synthase 3 (HAS3) overexpression downregulates MV3 melanoma cell proliferation, migration and adhesion.

    PubMed

    Takabe, Piia; Bart, Geneviève; Ropponen, Antti; Rilla, Kirsi; Tammi, Markku; Tammi, Raija; Pasonen-Seppänen, Sanna

    2015-09-10

    Malignant skin melanoma is one of the most deadly human cancers. Extracellular matrix (ECM) influences the growth of malignant tumors by modulating tumor cells adhesion and migration. Hyaluronan is an essential component of the ECM, and its amount is altered in many tumors, suggesting an important role for hyaluronan in tumorigenesis. Nonetheless its role in melanomagenesis is not understood. In this study we produced a MV3 melanoma cell line with inducible expression of the hyaluronan synthase 3 (HAS3) and studied its effect on the behavior of the melanoma cells. HAS3 overexpression expanded the cell surface hyaluronan coat and decreased melanoma cell adhesion, migration and proliferation by cell cycle arrest at G1/G0. Melanoma cell migration was restored by removal of cell surface hyaluronan by Streptomyces hyaluronidase and by receptor blocking with hyaluronan oligosaccharides, while the effect on cell proliferation was receptor independent. Overexpression of HAS3 decreased ERK1/2 phosphorylation suggesting that inhibition of MAP-kinase signaling was responsible for these suppressive effects on the malignant phenotype of MV3 melanoma cells.

  18. Role of HLA-G1 in trophoblast cell proliferation, adhesion and invasion

    SciTech Connect

    Jiang, Feng; Zhao, Hongxi; Wang, Li; Guo, Xinyu; Wang, Xiaohong; Yin, Guowu; Hu, Yunsheng; Li, Yi; Yao, Yuanqing

    2015-02-27

    Trophoblast cells are important in embryo implantation and fetomaternal tolerance. HLA-G is specifically expressed at the maternal–fetal interface and is a regulator in pregnancy. The aim of the present study was to detect the effect of HLA-G1 on trophoblast cell proliferation, adhesion, and invasion. Human trophoblast cell lines (JAR and HTR-8/SVneo cells) were infected with HLA-G1-expressing lentivirus. After infection, HLA-G1 expression of the cells was detected by western blotting. Cell proliferation was detected by the BrdU assay. The cell cycle and apoptosis of JAR and HTR-8/SVneo cells was measured by flow cytometry (FCM). The invasion of the cells under different conditions was detected by the transwell invasion chamber assay. HLA-G1 didn't show any significant influence on the proliferation, apoptosis, adhesion, and invasion of trophocytes in normal culture conditions. However, HLA-G1 inhibited JAR and HTR-8/SVneo cells invasion induced by hepatocyte growth factor (HGF) under normal oxygen conditions. In conditions of hypoxia, HLA-G1 couldn't inhibit the induction of cell invasion by HGF. HLA-G1 is not an independent factor for regulating the trophocytes. It may play an indirect role in embryo implantation and formation of the placenta. - Highlights: • HLA-G1 could not influence trophocytes under normal conditions. • HLA-G1 inhibited cell invasion induced by HGF under normal oxygen condition. • HLA-G1 could not influence cell invasion under hypoxia conditions.

  19. The selective role of ECM components on cell adhesion, morphology, proliferation and communication in vitro.

    PubMed

    Schlie-Wolter, Sabrina; Ngezahayo, Anaclet; Chichkov, Boris N

    2013-06-10

    Cell binding to the extracellular matrix (ECM) is essential for cell and tissue functions. In this context, each tissue consists of a unique ECM composition, which may be responsible for tissue-specific cell responses. Due to the complexity of ECM-cell interactions-which depend on the interplay of inside-out and outside-in signaling cascades, cell and tissue specificity of ECM-guidance is poorly understood. In this paper, we investigate the role of different ECM components like laminin, fibronectin, and collagen type I with respect to the essential cell behaviour patterns: attachment dynamics such as adhesion kinetic and force, formation of focal adhesion complexes, morphology, proliferation, and intercellular communication. A detailed in vitro comparison of fibroblasts, endothelial cells, osteoblasts, smooth muscle cells, and chondrocytes reveals significant differences in their cell responses to the ECM: cell behaviour follows a cell specific ligand priority ranking, which was independent of the cell type origin. Fibroblasts responded best to fibronectin, chondrocytes best to collagen I, the other cell types best to laminin. This knowledge is essential for optimization of tissue-biomaterial interfaces in all tissue engineering applications and gives insight into tissue-specific cell guidance.

  20. Adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene.

    PubMed

    Gumpenberger, T; Heitz, J; Bäuerle, D; Kahr, H; Graz, I; Romanin, C; Svorcik, V; Leisch, F

    2003-12-01

    We studied the adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene samples. The polymer surfaces were modified by exposure to the ultraviolet light of a Xe(2)(*)-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere. Treatment times were between 10 and 20 min. The endothelial cell density was determined 1, 3 and 8 days after seeding by image analysis. Surface modification of the samples resulted in a significant increase in the number of adhering cells and in the formation of a confluent cell layer after 3-8 days. The results were comparable than those obtained on polystyrene Petri dishes, which are used as standard substrates in cell cultivation. Thus modified PTFE appears to be a promising material for the fabrication of artificial vascular prostheses coated with endothelial cells.

  1. Investigation of In Vitro Bone Cell Adhesion and Proliferation on Ti Using Direct Current Stimulation.

    PubMed

    Bodhak, Subhadip; Bose, Susmita; Kinsel, William C; Bandyopadhyay, Amit

    2012-12-01

    Our objective was to establish an in vitro cell culture protocol to improve bone cell attachment and proliferation on Ti substrate using direct current stimulation. For this purpose, a custom made electrical stimulator was developed and a varying range of direct currents, from 5 to 25 µA, were used to study the current stimulation effect on bone cells cultured on conducting Ti samples in vitro. Cell-materials interaction was studied for a maximum of 5 days by culturing with human fetal osteoblast cells (hFOB). The direct current was applied in every 8 h time interval and the duration of electrical stimulation was kept constant at 15 min for all cases. In vitro results showed that direct current stimulation significantly favored bone cell attachment and proliferation in comparison to nonstimulated Ti surface. Immunochemistry and confocal microscopy results confirmed that the cell adhesion was most pronounced on 25 µA direct current stimulated Ti surfaces as hFOB cells expressed higher vinculin protein with increasing amount of direct current. Furthermore, MTT assay results established that cells grew 30% higher in number under 25 µA electrical stimulation as compared to nonstimulated Ti surface after 5 days of culture period. In this work we have successfully established a simple and cost effective in vitro protocol offering easy and rapid analysis of bone cell-materials interaction which can be used in promotion of bone cell attachment and growth on Ti substrate using direct current electrical stimulation in an in vitro model.

  2. Surface chemistry and polymer film thickness effects on endothelial cell adhesion and proliferation

    PubMed Central

    Bhattacharyya, Dhiman; Xu, Hao; Deshmukh, Rajendra R.; Timmons, Richard B.; Nguyen, Kytai T.

    2010-01-01

    Adherence and growth rates of human aortic endothelial cells (HAEC) on plasma polymerized poly(vinylacetic acid) films were measured as functions of the surface density of —COOH groups and plasma deposited film thickness. Pulsed plasma polymerization was employed to produce films containing 3.6 to 9% —COOH groups, expressed as a percent of total carbon content. Endothelial cells exhibited increased cell adherence and proliferation with increasing —COOH surface densities. Additionally, and unexpectedly, cell growth was also dependent on the film thicknesses, which ranged from 25 to 200 nm. The results indicate that optimization of the functional group surface density and film thickness could produce significant enhancements in initial adhesion and subsequent growth of the HAEC cells. PMID:20213813

  3. The selective role of ECM components on cell adhesion, morphology, proliferation and communication in vitro

    SciTech Connect

    Schlie-Wolter, Sabrina; Ngezahayo, Anaclet; Chichkov, Boris N.

    2013-06-10

    Cell binding to the extracellular matrix (ECM) is essential for cell and tissue functions. In this context, each tissue consists of a unique ECM composition, which may be responsible for tissue-specific cell responses. Due to the complexity of ECM-cell interactions—which depend on the interplay of inside-out and outside-in signaling cascades, cell and tissue specificity of ECM-guidance is poorly understood. In this paper, we investigate the role of different ECM components like laminin, fibronectin, and collagen type I with respect to the essential cell behaviour patterns: attachment dynamics such as adhesion kinetic and force, formation of focal adhesion complexes, morphology, proliferation, and intercellular communication. A detailed in vitro comparison of fibroblasts, endothelial cells, osteoblasts, smooth muscle cells, and chondrocytes reveals significant differences in their cell responses to the ECM: cell behaviour follows a cell specific ligand priority ranking, which was independent of the cell type origin. Fibroblasts responded best to fibronectin, chondrocytes best to collagen I, the other cell types best to laminin. This knowledge is essential for optimization of tissue-biomaterial interfaces in all tissue engineering applications and gives insight into tissue-specific cell guidance. -- Highlights: • We analyse the impact of ECM components on cell behaviour in vitro. • We compare five different cell types, using the same culture conditions. • The ECM significantly guides all cell responses. • Cell behaviour follows a cell specific ligand-priority ranking. • This gives insight in tissue formation and is essential for biomedical applications.

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

    PubMed Central

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

    2016-01-01

    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. PMID:27649159

  5. Amphiphilic Macromolecule Nanoassemblies Suppress Smooth Muscle Cell Proliferation and Platelet Adhesion

    PubMed Central

    Chan, Jennifer W.; Lewis, Daniel R.; Petersen, Latrisha K.; Moghe, Prabhas V.; Uhrich, Kathryn E.

    2016-01-01

    While the development of second- and third-generation drug-eluting stents (DES) have significantly improved patient outcomes by reducing smooth muscle cell (SMC) proliferation, DES have also been associated with an increased risk of late-stent thrombosis due to delayed re-endothelialization and hypersensitivity reactions from the drug-polymer coating. Furthermore, DES anti-proliferative agents do not counteract the upstream oxidative stress that triggers the SMC proliferation cascade. In this study, we investigate biocompatible amphiphilic macromolecules (AMs) that address high oxidative lipoprotein microenvironments by competitively binding oxidized lipid receptors and suppressing SMC proliferation with minimal cytotoxicity. To determine the influence of nanoscale assembly on proliferation, micelles and nanoparticles were fabricated from AM unimers containing a phosphonate or carboxylate end-group, sugar-based hydrophobic domain, and a hydrophilic poly(ethylene glycol) domain. The results indicate that when SMCs are exposed to high levels of oxidized lipid stimuli, nanotherapeutics inhibit lipid uptake, downregulate scavenger receptor expression, and attenuate scavenger receptor gene transcription in SMCs, and thus significantly suppress proliferation. Although both functional end-groups were similarly efficacious, nanoparticles suppressed oxidized lipid uptake and scavenger receptor expression more effectively compared to micelles, indicating the relative importance of formulation characteristics (e.g., higher localized AM concentrations and nanotherapeutic stability) in scavenger receptor binding as compared to AM end-group functionality. Furthermore, AM coatings significantly prevented platelet adhesion to metal, demonstrating its potential as an anti-platelet therapy to treat thrombosis. Thus, AM micelles and NPs can effectively repress early stage SMC proliferation and thrombosis through non-cytotoxic mechanisms, highlighting the promise of nanomedicine for

  6. Amphiphilic macromolecule nanoassemblies suppress smooth muscle cell proliferation and platelet adhesion.

    PubMed

    Chan, Jennifer W; Lewis, Daniel R; Petersen, Latrisha K; Moghe, Prabhas V; Uhrich, Kathryn E

    2016-04-01

    While the development of second- and third-generation drug-eluting stents (DES) have significantly improved patient outcomes by reducing smooth muscle cell (SMC) proliferation, DES have also been associated with an increased risk of late-stent thrombosis due to delayed re-endothelialization and hypersensitivity reactions from the drug-polymer coating. Furthermore, DES anti-proliferative agents do not counteract the upstream oxidative stress that triggers the SMC proliferation cascade. In this study, we investigate biocompatible amphiphilic macromolecules (AMs) that address high oxidative lipoprotein microenvironments by competitively binding oxidized lipid receptors and suppressing SMC proliferation with minimal cytotoxicity. To determine the influence of nanoscale assembly on proliferation, micelles and nanoparticles were fabricated from AM unimers containing a phosphonate or carboxylate end-group, a sugar-based hydrophobic domain, and a hydrophilic poly(ethylene glycol) domain. The results indicate that when SMCs are exposed to high levels of oxidized lipid stimuli, nanotherapeutics inhibit lipid uptake, downregulate scavenger receptor expression, and attenuate scavenger receptor gene transcription in SMCs, and thus significantly suppress proliferation. Although both functional end-groups were similarly efficacious, nanoparticles suppressed oxidized lipid uptake and scavenger receptor expression more effectively compared to micelles, indicating the relative importance of formulation characteristics (e.g., higher localized AM concentrations and nanotherapeutic stability) in scavenger receptor binding as compared to AM end-group functionality. Furthermore, AM coatings significantly prevented platelet adhesion to metal, demonstrating its potential as an anti-platelet therapy to treat thrombosis. Thus, AM micelles and NPs can effectively repress early stage SMC proliferation and thrombosis through non-cytotoxic mechanisms, highlighting the promise of nanomedicine for

  7. Thymus vulgaris (thyme) inhibits proliferation, adhesion, migration, and invasion of human colorectal cancer cells.

    PubMed

    Al-Menhali, Afnan; Al-Rumaihi, Aisha; Al-Mohammed, Hana; Al-Mazrooey, Hana; Al-Shamlan, Maryam; AlJassim, Meaad; Al-Korbi, Noof; Eid, Ali Hussein

    2015-01-01

    Colorectal cancer (CRC) remains one of the most common malignancies and a leading cause of cancer-related deaths. Its prognosis remains poor for patients with several grades of this disease. This underscores the need for alternative modalities, such as herbal medicines, to treat this disease. A commonly used plant that appears to be of high medicinal value is Thymus vulgaris L. However, the effects of this plant on the malignant behavior of human CRC cells remains poorly investigated. This study was undertaken to determine the anticancer efficacy of T. vulgaris extract (TVE) in CRC cells. Our results show that TVE inhibits proliferation in a concentration- and time-dependent fashion. This decreased proliferation was concomitant with increased apoptotic cell death as evidenced by increased caspase3/7 activity. Moreover, TVE also decreased adhesion to fibronectin in a concentration-dependent manner. The migratory and invasive capacities of HCT116 cells were significantly inhibited by TVE. Taken together, these data suggest that the TVE inhibits malignant phenotype of colon cancer cells. Therefore, T. vulgaris could have an anticancer effect and that some of its bioactive compounds may prove to be effective treatment modalities for human CRC.

  8. The effect of graphene substrate on osteoblast cell adhesion and proliferation.

    PubMed

    Aryaei, Ashkan; Jayatissa, Ahalapitiya H; Jayasuriya, Ambalangodage C

    2014-09-01

    Understanding the effect of graphene substrate on graphene-cell interaction is important for considering graphene as a potential candidate for biomedical applications. In this article, biocompatibility of few layers of graphene film transferred to different substrates was evaluated using osteoblasts. The substrates were oxidized silicon wafer (SiO2/Si stack), soda lime glass, and stainless steel. Chemical vapor deposition method was employed to synthesize graphene on copper substrate using methane and hydrogen as precursors. The quality and the thickness of graphene films on different substrates were estimated by Raman spectra, whereas the thickness of graphene film was confirmed by reflectance and transmittance spectroscopy. The study was also focused on cell attachment and morphology at two time points. The results show that graphene does not have any toxic effect on osteoblasts. The cell adhesion improves with graphene coated substrate than the substrate alone. It seems that graphene substrate properties play a dominant role in cell adhesion. The result of this study suggests that a layer of graphene on bone implants will be beneficial for osteoblast attachment and proliferation.

  9. Tumor suppressor KAI1 affects integrin {alpha}v{beta}3-mediated ovarian cancer cell adhesion, motility, and proliferation

    SciTech Connect

    Ruseva, Zlatna; Geiger, Pamina Xenia Charlotte; Hutzler, Peter; Kotzsch, Matthias; Luber, Birgit; Schmitt, Manfred; Gross, Eva; Reuning, Ute

    2009-06-10

    The tetraspanin KAI1 had been described as a metastasis suppressor in many different cancer types, a function for which associations of KAI1 with adhesion and signaling receptors of the integrin superfamily likely play a role. In ovarian cancer, integrin {alpha}v{beta}3 correlates with tumor progression and its elevation in vitro provoked enhanced cell adhesion accompanied by significant increases in cell motility and proliferation in the presence of its major ligand vitronectin. In the present study, we characterized integrin {alpha}v{beta}3-mediated tumor biological effects as a function of cellular KAI1 restoration and proved for the first time that KAI1, besides its already known physical crosstalk with {beta}1-integrins, also colocalizes with integrin {alpha}v{beta}3. Functionally, elevated KAI1 levels drastically increased integrin {alpha}v{beta}3/vitronectin-dependent ovarian cancer cell adhesion. Since an intermediate level of cell adhesive strength is required for optimal cell migration, we next studied ovarian cancer cell motility as a function of KAI1 restoration. By time lapse video microscopy, we found impaired integrin {alpha}v{beta}3/vitronectin-mediated cell migration most probably due to strongly enhanced cellular immobilization onto the adhesion-supporting matrix. Moreover, KAI1 reexpression significantly diminished cell proliferation. These data strongly indicate that KAI1 may suppress ovarian cancer progression by inhibiting integrin {alpha}v{beta}3/vitronectin-provoked tumor cell motility and proliferation as important hallmarks of the oncogenic process.

  10. Virtual-tissue computer simulations define the roles of cell adhesion and proliferation in the onset of kidney cystic disease

    PubMed Central

    Belmonte, Julio M.; Clendenon, Sherry G.; Oliveira, Guilherme M.; Swat, Maciej H.; Greene, Evan V.; Jeyaraman, Srividhya; Glazier, James A.; Bacallao, Robert L.

    2016-01-01

    In autosomal dominant polycystic kidney disease (ADPKD), cysts accumulate and progressively impair renal function. Mutations in PKD1 and PKD2 genes are causally linked to ADPKD, but how these mutations drive cell behaviors that underlie ADPKD pathogenesis is unknown. Human ADPKD cysts frequently express cadherin-8 (cad8), and expression of cad8 ectopically in vitro suffices to initiate cystogenesis. To explore cell behavioral mechanisms of cad8-driven cyst initiation, we developed a virtual-tissue computer model. Our simulations predicted that either reduced cell–cell adhesion or reduced contact inhibition of proliferation triggers cyst induction. To reproduce the full range of cyst morphologies observed in vivo, changes in both cell adhesion and proliferation are required. However, only loss-of-adhesion simulations produced morphologies matching in vitro cad8-induced cysts. Conversely, the saccular cysts described by others arise predominantly by decreased contact inhibition, that is, increased proliferation. In vitro experiments confirmed that cell–cell adhesion was reduced and proliferation was increased by ectopic cad8 expression. We conclude that adhesion loss due to cadherin type switching in ADPKD suffices to drive cystogenesis. Thus, control of cadherin type switching provides a new target for therapeutic intervention. PMID:27193300

  11. Investigation of In Vitro Bone Cell Adhesion and Proliferation on Ti Using Direct Current Stimulation

    PubMed Central

    Bodhak, Subhadip; Bose, Susmita; Kinsel, William C.; Bandyopadhyay, Amit

    2012-01-01

    Our objective was to establish an in vitro cell culture protocol to improve bone cell attachment and proliferation on Ti substrate using direct current stimulation. For this purpose, a custom made electrical stimulator was developed and a varying range of direct currents, from 5 to 25 µA, were used to study the current stimulation effect on bone cells cultured on conducting Ti samples in vitro. Cell–materials interaction was studied for a maximum of 5 days by culturing with human fetal osteoblast cells (hFOB). The direct current was applied in every 8 h time interval and the duration of electrical stimulation was kept constant at 15 min for all cases. In vitro results showed that direct current stimulation significantly favored bone cell attachment and proliferation in comparison to nonstimulated Ti surface. Immunochemistry and confocal microscopy results confirmed that the cell adhesion was most pronounced on 25 µA direct current stimulated Ti surfaces as hFOB cells expressed higher vinculin protein with increasing amount of direct current. Furthermore, MTT assay results established that cells grew 30% higher in number under 25 µA electrical stimulation as compared to nonstimulated Ti surface after 5 days of culture period. In this work we have successfully established a simple and cost effective in vitro protocol offering easy and rapid analysis of bone cell-materials interaction which can be used in promotion of bone cell attachment and growth on Ti substrate using direct current electrical stimulation in an in vitro model. PMID:23144532

  12. Hydrophilic PCU scaffolds prepared by grafting PEGMA and immobilizing gelatin to enhance cell adhesion and proliferation.

    PubMed

    Shi, Changcan; Yuan, Wenjie; Khan, Musammir; Li, Qian; Feng, Yakai; Yao, Fanglian; Zhang, Wencheng

    2015-05-01

    Gelatin contains many functional motifs which can modulate cell specific adhesion, so we modified polycarbonate urethane (PCU) scaffold surface by immobilization of gelatin. PCU-g-gelatin scaffolds were prepared by direct immobilizing gelatins onto the surface of aminated PCU scaffolds. To increase the immobilization amount of gelatin, poly(ethylene glycol) methacrylate (PEGMA) was grafted onto PCU scaffolds by surface initiated atom transfer radical polymerization. Then, following amination and immobilization, PCU-g-PEGMA-g-gelatin scaffolds were obtained. Both modified scaffolds were characterized by chemical and biological methods. After immobilization of gelatin, the microfiber surface became rough, but the original morphology of scaffolds was maintained successfully. PCU-g-PEGMA-g-gelatin scaffolds were more hydrophilic than PCU-g-gelatin scaffolds. Because hydrophilic PEGMA and gelatin were grafted and immobilized onto the surface, the PCU-g-PEGMA-g-gelatin scaffolds showed low platelet adhesion, perfect anti-hemolytic activity and excellent cell growth and proliferation capacity. It could be envisioned that PCU-g-PEGMA-g-gelatin scaffolds might have potential applications in tissue engineering artificial scaffolds.

  13. Vasostatin-2 inhibits cell proliferation and adhesion in vascular smooth muscle cells, which are associated with the progression of atherosclerosis.

    PubMed

    Hou, Jianghong; Xue, Xiaolin; Li, Junnong

    2016-01-22

    Recently, the serum expression level of vasostatin-2 was found to be reduced and is being studied as an important indicator to assess the presence and severity of coronary artery disease; the functional properties of vasostatin-2 and its relationship with the development of atherosclerosis remains unclear. In this study, we attempted to detect the expression of vasostatin-2 and its impact on human vascular smooth muscle cells (VSMCs). Quantitative real-time PCR (qRT-PCR) and western blot were used to assess the expression level of vasostatin-2 in VSMCs between those from atherosclerosis and disease-free donors; we found that vasostatin-2 was significantly down-regulated in atherosclerosis patient tissues and cell lines. In addition, the over-expression of vasostatin-2 apparently inhibits cell proliferation and migration in VSMCs. Gain-of-function in vitro experiments further show that vasostatin-2 over-expression significantly inhibits inflammatory cytokines release in VSMCs. In addition, cell adhesion experimental analysis showed that soluble adhesion molecules (sICAM-1, sVCAM-1) had decreased expression when vasostatin-2 was over-expressed in VSMCs. Therefore, our results indicate that vasostatin-2 is an atherosclerosis-related factor that can inhibit cell proliferation, inflammatory response and cell adhesion in VSMCs. Taken together, our results indicate that vasostatin-2 could serve as a potential diagnostic biomarker and therapeutic option for human atherosclerosis in the near future.

  14. Decreased astroglial cell adhesion and proliferation on zinc oxide nanoparticle polyurethane composites

    PubMed Central

    Seil, Justin T; Webster, Thomas J

    2008-01-01

    Nanomaterials offer a number of properties that are of interest to the field of neural tissue engineering. Specifically, materials that exhibit nanoscale surface dimensions have been shown to promote neuron function while simultaneously minimizing the activity of cells such as astrocytes that inhibit central nervous system regeneration. Studies demonstrating enhanced neural tissue regeneration in electrical fields through the use of conductive materials have led to interest in piezoelectric materials (or those materials which generate a transient electrical potential when mechanically deformed) such as zinc oxide (ZnO). It has been speculated that ZnO nanoparticles possess increased piezoelectric properties over ZnO micron particles. Due to this promise in neural applications, the objective of the present in vitro study was, for the first time, to assess the activity of astroglial cells on ZnO nanoparticle polymer composites. ZnO nanoparticles embedded in polyurethane were analyzed via scanning electron microscopy to evaluate nanoscale surface features of the composites. The surface chemistry was characterized via X-ray photoelectron spectroscopy. Astroglial cell response was evaluated based on cell adhesion and proliferation. Astrocyte adhesion was significantly reduced on ZnO nanoparticle/polyurethane (PU) composites with a weight ratio of 50:50 (PU:ZnO) wt.%, 75:25 (PU:ZnO) wt.%, and 90:10 (PU:ZnO) wt.% in comparison to pure PU. The successful production of ZnO nanoparticle composite scaffolds suitable for decreasing astroglial cell density demonstrates their potential as a nerve guidance channel material with greater efficiency than what may be available today. PMID:19337420

  15. Videography supported adhesion, and proliferation behavior of MG-63 osteoblastic cells on 2.5D titania nanotube matrices.

    PubMed

    Manurung, Robeth Viktoria; Fu, Pei-Wen; Chu, Yeh-Shiu; Lo, Chun-Min; Chattopadhyay, Surojit

    2016-04-01

    Human osteosarcoma cells MG-63 were cultured on anodically etched titania nanotubes (TiO2 NT), with diameters ranging from 40-100 nm, to study the correlations between cell proliferation and adhesion on the 2.5 dimensional (2.5D) extracellular matrix (ECM). Unlike other reports, mostly based on mouse stem cells, and 2D cell culture, our studies indicate that the 2.5D NT promote higher proliferation and activity, but less 2D adhesion. Proliferation of the MG-63 cells was significantly higher in the NTs, the best being the 70 nm diameter sample, compared to planar titania (control). This is consistent with previous studies. However, cellular adhesion was stronger on TiO2 NT with increasing diameter, and highest on the control as obtained from shear stress measurement, paxilin imaging, and western blot measurements probing focal adhesion kinase, p130 CAS, and extracellular-regulated kinase, in addition to cell morphology imaging by fluorescence microscopy. We provide direct videography of cell migration, and cell speed data indicating faster filopodial activity on the TiO2 NT surfaces having lower adhesion. This evidence was not available previously. The NT matrices promote cells with smaller surface area, because of less 2D stretching. In contrast, on comparatively planar 2D-like surfaces uniaxial stretching of the cell body with strong anchoring of the filopodia, resulted in larger cell surface area, and demonstrated stronger adhesion. The difference in the results, with those previously published, may be generally attributed to, among others, the use of mouse stem cells (human osteosarcoma used here), and unannealed as-grown TiO2 NTs used previously (annealed ECMs used here).

  16. Bioinspired superhydrophobic poly(L-lactic acid) surfaces control bone marrow derived cells adhesion and proliferation.

    PubMed

    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.

  17. β-eudesmol, a sesquiterpene from Teucrium ramosissimum, inhibits superoxide production, proliferation, adhesion and migration of human tumor cell.

    PubMed

    Ben Sghaier, Mohamed; Mousslim, Mohamed; Pagano, Alessandra; Ammari, Youssef; Luis, José; Kovacic, Hervé

    2016-09-01

    Reactive oxygen species are well-known mediators of various biological responses. Recently, new homologues of the catalytic subunit of NADPH oxidase have been discovered in non phagocytic cells. These new homologues (Nox1-Nox5) produce low levels of superoxides compared to the phagocytic homologue Nox2/gp91phox. In this study we examined the effect of β-eudesmol, a sesquiterpenoid alcohol isolated from Teucrium ramosissimum leaves, on proliferation, superoxide anion production, adhesion and migration of human lung (A549) and colon (HT29 and Caco-2) cancer cell lines. Proliferation of tumor cells was inhibited by β-eudesmol. It also significantly inhibited superoxide production in A549 cells. Furthermore, β-eudesmol inhibited adhesion and migration of A549 and HT29 cell. These results demonstrate that β-eudesmol may be a novel anticancer agent for the treatment of lung and colon cancer by different ways: by inhibition of superoxide production or by blocking proliferation, adhesion and migration.

  18. cAMP effects in neuroendocrine tumors: The role of Epac and PKA in cell proliferation and adhesion.

    PubMed

    Vitali, E; Cambiaghi, V; Spada, A; Tresoldi, A; Zerbi, A; Peverelli, E; Carnaghi, C; Mantovani, G; Lania, A G

    2015-12-10

    cAMP effects have been initially attributed to protein kinase A (PKA) activation. Subsequently, two exchange proteins directly activated by cAMP (Epac1/2) have been identified as cAMP targets. Aim of this study was to investigate cAMP effects in pancreatic-NET (P-NET) and bronchial carcinoids and in corresponding cell lines (QGP-1 and H727) on cell proliferation and adhesion and to determine PKA and Epac role in mediating these effects. We found that cAMP increased cyclin D1 expression in P-NET and QGP-1 cells, whereas it had opposite effects on bronchial carcinoids and H727 cells and it promoted cell adhesion in QGP-1 and H727 cells. These effects are mimicked by Epac and PKA specific analogs, activating the small GTPase Rap1. In conclusion, we demonstrated that cAMP exerted divergent effects on proliferation and promoted cell adhesion of different neuroendocrine cell types, these effects being mediated by both Epac and PKA and involving the same effector GTPase Rap1.

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

    SciTech Connect

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

    2009-03-10

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

  20. Rutin inhibits proliferation, attenuates superoxide production and decreases adhesion and migration of human cancerous cells.

    PubMed

    Ben Sghaier, Mohamed; Pagano, Alessandra; Mousslim, Mohamed; Ammari, Youssef; Kovacic, Hervé; Luis, José

    2016-12-01

    Lung and colorectal cancer are the principal causes of death in the world. Rutin, an active flavonoid compound, is known for possessing a wide range of biological activities. In this study, we examined the effect of rutin on the viability, superoxide anion production, adhesion and migration of human lung (A549) and colon (HT29 and Caco-2) cancer cell lines. In order to control the harmlessness of the tested concentrations of rutin, the viability of cancer cell lines was assessed using a 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. ROS generation was measured by lucigenin chemiluminescence detecting superoxide ions. To investigate the effect of rutin on the behavior of human lung and colon cancer cell lines, we performed adhesion assays, using various purified extracellular matrix (ECM) proteins. Finally, in vitro cell migration assays were explored using modified Boyden chambers. The viability of cancerous cells was inhibited by rutin. It also significantly attenuated the superoxide production in HT29 cells. In addition, rutin affected adhesion and migration of A549 and HT29 cell. These findings indicate that rutin, a natural molecule, might have potential as anticancer agent against lung and colorectal carcinogenesis.

  1. Modulation of keratin in adhesion, proliferation, adipogenic, and osteogenic differentiation of porcine adipose-derived stem cells.

    PubMed

    Wu, Yen-Lin; Lin, Che-Wei; Cheng, Nai-Chen; Yang, Kai-Chiang; Yu, Jiashing

    2017-01-01

    Recently, keratin attracts tremendous interest because of its intrinsic ability to interact with different cells. It has the potential to serve as a controllable extracellular matrix protein that can be used to demonstrate cell mechanism and cell-matrix interaction. However, there have been relatively few studies on the effects of keratin on stem cells. In the present work, we study the effects of human keratin on porcine adipose-derived stem cells (pASCs) and a series of selective cell lines: 3T3 fibroblasts, Madin-Darby canine kidney (MDCK) cells, and MG63 osteoblasts. Relative to un-treated culture plate, our results showed that keratin coating substrates promote cell adhesion and proliferation to above cell lines. Keratin also improved pASCs adhesion, proliferation, and enhanced cell viability. Evaluation of genetic markers showed that adipogenic and osteogenic differentiations of pASCs can be successfully induced, thus demonstrating that keratin did not influence the stemness of pASCs. Furthermore, keratin improved adipogenic differentiations of pASCs in terms of up-regulations in lipoprotein lipase, peroxisome proliferator-activated receptor gamma, and CCAAT-enhancer-binding protein alpha. The osteogenic markers type I collagen, runt-related transcription factor 2, and vitamin D receptor were also upregulated when pASCs cultured on keratin substrates. Therefore, keratin can serve as a biological derived material for surface modification and scaffold fabrication for biomedical purpose. The combination of keratin with stem cells may be a potential candidate for tissue repair in the field of regenerative medicine. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 180-192, 2017.

  2. Mimicking bone extracellular matrix: integrin-binding peptidomimetics enhance osteoblast-like cells adhesion, proliferation, and differentiation on titanium.

    PubMed

    Fraioli, Roberta; Rechenmacher, Florian; Neubauer, Stefanie; Manero, José M; Gil, Javier; Kessler, Horst; Mas-Moruno, Carlos

    2015-04-01

    Interaction between the surface of implants and biological tissues is a key aspect of biomaterials research. Apart from fulfilling the non-toxicity and structural requirements, synthetic materials are asked to direct cell response, offering engineered cues that provide specific instructions to cells. This work explores the functionalization of titanium with integrin-binding peptidomimetics as a novel and powerful strategy to improve the adhesion, proliferation and differentiation of osteoblast-like cells to implant materials. Such biomimetic strategy aims at targeting integrins αvβ3 and α5β1, which are highly expressed on osteoblasts and are essential for many fundamental functions in bone tissue development. The successful grafting of the bioactive molecules on titanium is proven by contact angle measurements, X-ray photoelectron spectroscopy and fluorescent labeling. Early attachment and spreading of cells are statistically enhanced by both peptidomimetics compared to unmodified titanium, reaching values of cell adhesion comparable to those obtained with full-length extracellular matrix proteins. Moreover, an increase in alkaline phosphatase activity, and statistically higher cell proliferation and mineralization are observed on surfaces coated with the peptidomimetics. This study shows an unprecedented biological activity for low-molecular-weight ligands on titanium, and gives striking evidence of the potential of these molecules to foster bone regeneration on implant materials.

  3. Lowered Expression of Tumor Suppressor Candidate MYO1C Stimulates Cell Proliferation, Suppresses Cell Adhesion and Activates AKT

    PubMed Central

    Visuttijai, Kittichate; Pettersson, Jennifer; Mehrbani Azar, Yashar; van den Bout, Iman; Örndal, Charlotte; Marcickiewicz, Janusz; Nilsson, Staffan; Hörnquist, Michael; Olsson, Björn; Ejeskär, Katarina

    2016-01-01

    Myosin-1C (MYO1C) is a tumor suppressor candidate located in a region of recurrent losses distal to TP53. Myo1c can tightly and specifically bind to PIP2, the substrate of Phosphoinositide 3-kinase (PI3K), and to Rictor, suggesting a role for MYO1C in the PI3K pathway. This study was designed to examine MYO1C expression status in a panel of well-stratified endometrial carcinomas as well as to assess the biological significance of MYO1C as a tumor suppressor in vitro. We found a significant correlation between the tumor stage and lowered expression of MYO1C in endometrial carcinoma samples. In cell transfection experiments, we found a negative correlation between MYO1C expression and cell proliferation, and MYO1C silencing resulted in diminished cell migration and adhesion. Cells expressing excess of MYO1C had low basal level of phosphorylated protein kinase B (PKB, a.k.a. AKT) and cells with knocked down MYO1C expression showed a quicker phosphorylated AKT (pAKT) response in reaction to serum stimulation. Taken together the present study gives further evidence for tumor suppressor activity of MYO1C and suggests MYO1C mediates its tumor suppressor function through inhibition of PI3K pathway and its involvement in loss of contact inhibition. PMID:27716847

  4. ENO1 promotes tumor proliferation and cell adhesion mediated drug resistance (CAM-DR) in Non-Hodgkin's Lymphomas

    SciTech Connect

    Zhu, Xinghua; Miao, Xiaobing; Wu, Yaxun; Li, Chunsun; Guo, Yan; Liu, Yushan; Chen, Yali; Lu, Xiaoyun; Wang, Yuchan; He, Song

    2015-07-15

    Enolases are glycolytic enzymes responsible for the ATP-generated conversion of 2-phosphoglycerate to phosphoenolpyruvate. In addition to the glycolytic function, Enolase 1 (ENO1) has been reported up-regulation in several tumor tissues. In this study, we investigated the expression and biologic function of ENO1 in Non-Hodgkin's Lymphomas (NHLs). Clinically, by western blot analysis we observed that ENO1 expression was apparently higher in diffuse large B-cell lymphoma than in the reactive lymphoid tissues. Subsequently, immunohistochemical staining of 144 NHLs suggested that the expression of ENO1 was significantly lower in the indolent lymphomas compared with the progressive lymphomas. Further, we identified ENO1 as an independent prognostic factor, and it was significantly correlated with overall survival of NHL patients. In addition, we found that ENO1 could promote cell proliferation, regulate cell cycle associated gene and PI3K/AKT signaling pathway in NHLs. Finally, we verified that ENO1 participated in the process of lymphoma cell adhesion mediated drug resistance (CAM-DR). Adhesion to FN or HS5 cells significantly protected OCI-Ly8 and Daudi cells from cytotoxicity compared with those cultured in suspension, and these effects were attenuated when transfected with ENO1-siRNA. Based on the study, we propose that inhibition of ENO1 expression may be a novel strategy for therapy for NHLs patients, and it may be a target for drug resistance. - Highlights: • ENO1 expression is reversely correlated with clinical outcomes of patients with NHLs. • ENO1 promotes the proliferation of NHL cells. • ENO1 regulates cell adhesion mediated drug resistance.

  5. Insulin signaling via Akt2 switches plakophilin 1 function from stabilizing cell adhesion to promoting cell proliferation.

    PubMed

    Wolf, Annika; Rietscher, Katrin; Glaß, Markus; Hüttelmaier, Stefan; Schutkowski, Mike; Ihling, Christian; Sinz, Andrea; Wingenfeld, Aileen; Mun, Andrej; Hatzfeld, Mechthild

    2013-04-15

    Downregulation of adherens junction proteins is a frequent event in carcinogenesis. How desmosomal proteins contribute to tumor formation by regulating the balance between adhesion and proliferation is not well understood. The desmosomal protein plakophilin 1 can increase intercellular adhesion by recruiting desmosomal proteins to the plasma membrane or stimulate proliferation by enhancing translation rates. Here, we show that these dual functions of plakophilin 1 are regulated by growth factor signaling. Insulin stimulation induced the phosphorylation of plakophilin 1, which correlated with reduced intercellular adhesion and an increased activity of plakophilin 1 in the stimulation of translation. Phosphorylation was mediated by Akt2 at four motifs within the plakophilin 1 N-terminal domain. A plakophilin 1 phospho-mimetic mutant revealed reduced intercellular adhesion and accumulated in the cytoplasm, where it increased translation and proliferation rates and conferred the capacity of anchorage-independent growth. The cytoplasmic accumulation was mediated by the stabilization of phosphorylated plakophilin 1, which displayed a considerably increased half-life, whereas non-phosphorylated plakophilin 1 was more rapidly degraded. Our data indicate that upon activation of growth factor signaling, plakophilin 1 switches from a desmosome-associated growth-inhibiting to a cytoplasmic proliferation-promoting function. This supports the view that the deregulation of plakophilin 1, as observed in several tumors, directly contributes to hyperproliferation and carcinogenesis in a context-dependent manner.

  6. The influence of high intensity terahertz radiation on mammalian cell adhesion, proliferation and differentiation

    NASA Astrophysics Data System (ADS)

    Williams, Rachel; Schofield, Amy; Holder, Gareth; Downes, Joan; Edgar, David; Harrison, Paul; Siggel-King, Michele; Surman, Mark; Dunning, David; Hill, Stephen; Holder, David; Jackson, Frank; Jones, James; McKenzie, Julian; Saveliev, Yuri; Thomsen, Neil; Williams, Peter; Weightman, Peter

    2013-01-01

    Understanding the influence of exposure of biological systems to THz radiation is becoming increasingly important. There is some evidence to suggest that THz radiation can influence important activities within mammalian cells. This study evaluated the influence of the high peak power, low average power THz radiation produced by the ALICE (Daresbury Laboratory, UK) synchrotron source on human epithelial and embryonic stem cells. The cells were maintained under standard tissue culture conditions, during which the THz radiation was delivered directly into the incubator for various exposure times. The influence of the THz radiation on cell morphology, attachment, proliferation and differentiation was evaluated. The study demonstrated that there was no difference in any of these parameters between irradiated and control cell cultures. It is suggested that under these conditions the cells are capable of compensating for any effects caused by exposure to THz radiation with the peak powers levels employed in these studies.

  7. Mycobacterium ulcerans mycolactone interferes with adhesion, migration and proliferation of primary human keratinocytes and HaCaT cell line.

    PubMed

    Graziola, Francesca; Colombo, Elena; Tiberio, Rossana; Leigheb, Giorgio; Bozzo, Chiarella

    2017-04-01

    The pathogenicity of Mycobacterium ulcerans (Buruli ulcer) is closely associated with the secretion of exotoxin mycolactone. The cytotoxicity of mycolactone has been linked to its apoptogenic activity. We explored if low mycolactone concentrations, which are not able to induce apoptosis, can influence other essential activities on two primary human keratinocyte populations, keratinocyte stem cells (KSC) and transit amplifying cells (TAC), and on a human keratinocyte line, HaCaT. We demonstrated that 0.01 and 0.1 ng/ml mycolactone A/B are not able to induce apoptosis in primary human keratinocytes, but interfere with KSC wound repair. Moreover, the same toxin concentrations reduce cell proliferation of KSC and TAC and their ability to adhere to type IV collagen. HaCaT cells are more resistant to the toxin; nevertheless, they show a delayed woud repair when treated with 1 and 10 ng/ml mycolactone A/B. Moreover, these sub-apoptotic concentrations affect their ability to proliferate and adhere to collagen IV. Wound healing is a complex mechanism, which occurs "in vivo" as the outcome of many co-ordinated events. Sub-apoptotic mycolactone concentrations can affect essential mechanisms, which are required to achieve wound repair, such as adhesion, migration and proliferation of human keratinocytes.

  8. Porous titania surfaces on titanium with hierarchical macro- and mesoporosities for enhancing cell adhesion, proliferation and mineralization.

    PubMed

    Han, Guang; Müller, Werner E G; Wang, Xiaohong; Lilja, Louise; Shen, Zhijian

    2015-02-01

    Titanium received a macroporous titania surface layer by anodization, which contains open pores with average pore diameter around 5 μm. An additional mesoporous titania top layer following the contour of the macropores, of 100-200 nm thickness and with a pore diameter of 10nm, was formed by using the evaporation-induced self-assembly (EISA) method with titanium (IV) tetraethoxide as the precursor. A coherent laminar titania surface layer was thus obtained, creating a hierarchical macro- and mesoporous surface that was characterized by high-resolution electron microscopy. The interfacial bonding between the surface layers and the titanium matrix was characterized by the scratch test that confirmed a stable and strong bonding of titania surface layers on titanium. The wettability to water and the effects on the osteosarcoma cell line (SaOS-2) proliferation and mineralization of the formed titania surface layers were studied systematically by cell culture and scanning electron microscopy. The results proved that the porous titania surface with hierarchical macro- and mesoporosities was hydrophilic that significantly promoted cell attachment and spreading. A synergistic role of the hierarchical macro- and mesoporosities was revealed in terms of enhancing cell adhesion, proliferation and mineralization, compared with the titania surface with solo scale topography.

  9. Unsaturated Fatty Acids Drive Disintegrin and Metalloproteinase (ADAM)-dependent Cell Adhesion, Proliferation, and Migration by Modulating Membrane Fluidity*

    PubMed Central

    Reiss, Karina; Cornelsen, Isabell; Husmann, Matthias; Gimpl, Gerald; Bhakdi, Sucharit

    2011-01-01

    The disintegrin-metalloproteinases ADAM10 and ADAM17 mediate the release of several cell signaling molecules and cell adhesion molecules such as vascular endothelial cadherin or L-selectin affecting endothelial permeability and leukocyte transmigration. Dysregulation of ADAM activity may contribute to the pathogenesis of vascular diseases, but the mechanisms underlying the control of ADAM functions are still incompletely understood. Atherosclerosis is characterized by lipid plaque formation and local accumulation of unsaturated free fatty acids (FFA). Here, we show that unsaturated FFA increase ADAM-mediated substrate cleavage. We demonstrate that these alterations are not due to genuine changes in enzyme activity, but correlate with changes in membrane fluidity as revealed by measurement of 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy and fluorescence recovery after photobleaching analyses. ELISA and immunoblot experiments conducted with granulocytes, endothelial cells, and keratinocytes revealed rapid increase of ectodomain shedding of ADAM10 and ADAM17 substrates upon membrane fluidization. Large amounts of unsaturated FFA may be liberated from cholesteryl esters in LDL that is entrapped in atherosclerotic lesions. Incubation of cells with thus modified LDL resulted in rapid cleavage of ADAM substrates with corresponding functional consequences on cell proliferation, cell migration, and endothelial permeability, events of high significance in atherogenesis. We propose that FFA represent critical regulators of ADAM function that may assume relevance in many biological settings through their influence on mobility of enzyme and substrate in lipid bilayers. PMID:21642425

  10. Unsaturated fatty acids drive disintegrin and metalloproteinase (ADAM)-dependent cell adhesion, proliferation, and migration by modulating membrane fluidity.

    PubMed

    Reiss, Karina; Cornelsen, Isabell; Husmann, Matthias; Gimpl, Gerald; Bhakdi, Sucharit

    2011-07-29

    The disintegrin-metalloproteinases ADAM10 and ADAM17 mediate the release of several cell signaling molecules and cell adhesion molecules such as vascular endothelial cadherin or L-selectin affecting endothelial permeability and leukocyte transmigration. Dysregulation of ADAM activity may contribute to the pathogenesis of vascular diseases, but the mechanisms underlying the control of ADAM functions are still incompletely understood. Atherosclerosis is characterized by lipid plaque formation and local accumulation of unsaturated free fatty acids (FFA). Here, we show that unsaturated FFA increase ADAM-mediated substrate cleavage. We demonstrate that these alterations are not due to genuine changes in enzyme activity, but correlate with changes in membrane fluidity as revealed by measurement of 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy and fluorescence recovery after photobleaching analyses. ELISA and immunoblot experiments conducted with granulocytes, endothelial cells, and keratinocytes revealed rapid increase of ectodomain shedding of ADAM10 and ADAM17 substrates upon membrane fluidization. Large amounts of unsaturated FFA may be liberated from cholesteryl esters in LDL that is entrapped in atherosclerotic lesions. Incubation of cells with thus modified LDL resulted in rapid cleavage of ADAM substrates with corresponding functional consequences on cell proliferation, cell migration, and endothelial permeability, events of high significance in atherogenesis. We propose that FFA represent critical regulators of ADAM function that may assume relevance in many biological settings through their influence on mobility of enzyme and substrate in lipid bilayers.

  11. Adhesion and proliferation of skeletal muscle cells on single layer poly(lactic acid) ultra-thin films.

    PubMed

    Ricotti, Leonardo; Taccola, Silvia; Pensabene, Virginia; Mattoli, Virgilio; Fujie, Toshinori; Takeoka, Shinji; Menciassi, Arianna; Dario, Paolo

    2010-10-01

    An increasing interest in bio-hybrid systems and cell-material interactions is evident in the last years. This leads towards the development of new nano-structured devices and the assessment of their biocompatibility. In the present study, the development of free-standing single layer poly(lactic acid) (PLA) ultra-thin films is described, together with the analysis of topography and roughness properties. The biocompatibility of the PLA films has been tested in vitro, by seeding C2C12 skeletal muscle cells, and thus assessing cells shape, density and viability after 24, 48 and 72 h. The results show that free-standing flexible PLA nanofilms represent a good matrix for C2C12 cells adhesion, spreading and proliferation. Early differentiation into myotubes is also allowed. The biocompatibility of the novel ultra-thin films as substrates for cell growth promotes their application in the fields of regenerative medicine, muscle tissue engineering, drug delivery, and-in general-in the field of bio-hybrid devices.

  12. Nano-hydroxyapatite-coated metal-ceramic composite of iron-tricalcium phosphate: Improving the surface wettability, adhesion and proliferation of mesenchymal stem cells in vitro.

    PubMed

    Surmeneva, Maria A; Kleinhans, Claudia; Vacun, Gabriele; Kluger, Petra Juliane; Schönhaar, Veronika; Müller, Michaela; Hein, Sebastian Boris; Wittmar, Alexandra; Ulbricht, Mathias; Prymak, Oleg; Oehr, Christian; Surmenev, Roman A

    2015-11-01

    Thin radio-frequency magnetron sputter deposited nano-hydroxyapatite (HA) films were prepared on the surface of a Fe-tricalcium phosphate (Fe-TCP) bioceramic composite, which was obtained using a conventional powder injection moulding technique. The obtained nano-hydroxyapatite coated Fe-TCP biocomposites (nano-HA-Fe-TCP) were studied with respect to their chemical and phase composition, surface morphology, water contact angle, surface free energy and hysteresis. The deposition process resulted in a homogeneous, single-phase HA coating. The ability of the surface to support adhesion and the proliferation of human mesenchymal stem cells (hMSCs) was studied using biological short-term tests in vitro. The surface of the uncoated Fe-TCP bioceramic composite showed an initial cell attachment after 24h of seeding, but adhesion, proliferation and growth did not persist during 14 days of culture. However, the HA-Fe-TCP surfaces allowed cell adhesion, and proliferation during 14 days. The deposition of the nano-HA films on the Fe-TCP surface resulted in higher surface energy, improved hydrophilicity and biocompatibility compared with the surface of the uncoated Fe-TCP. Furthermore, it is suggested that an increase in the polar component of the surface energy was responsible for the enhanced cell adhesion and proliferation in the case of the nano-HA-Fe-TCP biocomposites.

  13. Repeated morphine treatment alters polysialylated neural cell adhesion molecule, glutamate decarboxylase-67 expression and cell proliferation in the adult rat hippocampus.

    PubMed

    Kahn, Laëtitia; Alonso, Gérard; Normand, Elisabeth; Manzoni, Olivier J

    2005-01-01

    Altered synaptic transmission and plasticity in brain areas involved in reward and learning are thought to underlie the long-lasting effects of addictive drugs. In support of this idea, opiates reduce neurogenesis [A.J. Eisch et al. (2000) Proceedings of the National Academy of Sciences USA, 97, 7579-7584] and enhance long-term potentiation in adult rodent hippocampus [J.M. Harrison et al. (2002) Journal of Neurophysiology, 87, 2464-2470], a key structure of learning and memory processes. Here we studied how repeated morphine treatment and withdrawal affect cell proliferation and neuronal phenotypes in the dentate gyrus-CA3 region of the adult rat hippocampus. Our data showed a strong reduction of cellular proliferation in morphine-dependent animals (54% of control) that was followed by a rebound increase after 1 week withdrawal and a return to normal after 2 weeks withdrawal. Morphine dependence was also associated with a drastic reduction in the expression levels of the polysialylated form of neural cell adhesion molecule (68% of control), an adhesion molecule expressed by newly generated neurons and involved in cell migration and structural plasticity. Polysialylated neural cell adhesion molecule levels quickly returned to normal following withdrawal. In morphine-dependent rats, we found a significant increase of glutamate decarboxylase-67 mRNA transcription (170% of control) in dentate gyrus granular cells which was followed by a marked rebound decrease after 1 week withdrawal and a return to normal after 4 weeks withdrawal. Together, the results show, for the first time, that, in addition to reducing cell proliferation and neurogenesis, chronic exposure to morphine dramatically alters neuronal phenotypes in the dentate gyrus-CA3 region of the adult rat hippocampus.

  14. Strategies to prepare TiO2 thin films, doped with transition metal ions, that exhibit specific physicochemical properties to support osteoblast cell adhesion and proliferation.

    PubMed

    Dhayal, Marshal; Kapoor, Renu; Sistla, Pavana Goury; Pandey, Ravi Ranjan; Kar, Satabisha; Saini, Krishan Kumar; Pande, Gopal

    2014-04-01

    Metal ion doped titanium oxide (TiO2) thin films, as bioactive coatings on metal or other implantable materials, can be used as surfaces for studying the cell biological properties of osteogenic and other cell types. Bulk crystallite phase distribution and surface carbon-oxygen constitution of thin films, play an important role in determining the biological responses of cells that come in their contact. Here we present a strategy to control the polarity of atomic interactions between the dopant metal and TiO2 molecules and obtain surfaces with smaller crystallite phases and optimal surface carbon-oxygen composition to support the maximum proliferation and adhesion of osteoblast cells. Our results suggest that surfaces, in which atomic interactions between the dopant metals and TiO2 were less polar, could support better adhesion, spreading and proliferation of cells.

  15. Carbonated apatites obtained by the hydrolysis of monetite: influence of carbonate content on adhesion and proliferation of MC3T3-E1 osteoblastic cells.

    PubMed

    Pieters, Ilse Y; Van den Vreken, Natasja M F; Declercq, Heidi A; Cornelissen, Maria J; Verbeeck, Ronald M H

    2010-04-01

    The influence of the carbonate content in apatites on the adhesion and the proliferation of MC3T3-E1 osteoblastic cells was investigated. B-type carbonated apatites (DCAps) were prepared by the hydrolysis of monetite (CaHPO(4), DCP) in solutions with a carbonate concentration ranging from 0.001 to 0.075 mol l(-1). Stoichiometric hydroxyapatite (DCAp0) was synthesized in carbonate-free solution. MC3T3-E1 cells were seeded on the compacted DCAps and cell adhesion and proliferation were analysed after 24h and 7 days, respectively, using a MTS assay and fluorescence microscopy. Cell adhesion tends to increase with increasing carbonate content for carbonate contents between 0 and 6.9 wt.% and levels out to an acceptable value (+ or - 50% compared to the control) for carbonate contents between 6.9 and 16.1 wt.%. Only DCAps with a carbonate content equal to or higher than 11% support high cell proliferation comparable to the control. On the latter DCAps, the cells have a spread morphology and form a near-confluent layer. A decrease in charge density and crystallinity at the apatite surface, as well as the formation of more spheroidal crystals with increasing carbonate content, might attribute to changes in composition and three-dimensional structure of the protein adsorption layer and hence to the observed cell behaviour. Consequently, only DCAps with a high carbonate content, mimicking early in vivo mineralization, are possible candidates for bone regeneration.

  16. Increasing α7β1-integrin promotes muscle cell proliferation, adhesion, and resistance to apoptosis without changing gene expression

    PubMed Central

    Liu, Jianming; Burkin, Dean J.; Kaufman, Stephen J.

    2008-01-01

    The dystrophin-glycoprotein complex maintains the integrity of skeletal muscle by associating laminin in the extracellular matrix with the actin cytoskeleton. Several human muscular dystrophies arise from defects in the components of this complex. The α7β1-integrin also binds laminin and links the extracellular matrix with the cytoskeleton. Enhancement of α7-integrin levels alleviates pathology in mdx/utrn−/− mice, a model of Duchenne muscular dystrophy, and thus the integrin may functionally compensate for the absence of dystrophin. To test whether increasing α7-integrin levels affects transcription and cellular functions, we generated α7-integrin-inducible C2C12 cells and transgenic mice that overexpress the integrin in skeletal muscle. C2C12 myoblasts with elevated levels of integrin exhibited increased adhesion to laminin, faster proliferation when serum was limited, resistance to staurosporine-induced apoptosis, and normal differentiation. Transgenic expression of eightfold more integrin in skeletal muscle did not result in notable toxic effects in vivo. Moreover, high levels of α7-integrin in both myoblasts and in skeletal muscle did not disrupt global gene expression profiles. Thus increasing integrin levels can compensate for defects in the extracellular matrix and cytoskeleton linkage caused by compromises in the dystrophin-glycoprotein complex without triggering apparent overt negative side effects. These results support the use of integrin enhancement as a therapy for muscular dystrophy. PMID:18045857

  17. Pravastatin and simvastatin inhibit the adhesion, replication and proliferation of Toxoplasma gondii (RH strain) in HeLa cells.

    PubMed

    Sanfelice, Raquel Arruda; da Silva, Suelen Santos; Bosqui, Larissa Rodrigues; Miranda-Sapla, Milena Menegazzo; Barbosa, Bellisa Freitas; Silva, Rafaela José; Ferro, Eloísa A Vieira; Panagio, Luciano Aparecido; Navarro, Italmar Teodorico; Bordignon, Juliano; Conchon-Costa, Ivete; Pavanelli, Wander Rogerio; Almeida, Ricardo Sergio; Costa, Idessania Nazareth

    2017-03-01

    The conventional treatment for toxoplasmosis with pyrimethamine and sulfadiazine shows toxic effects to the host, and it is therefore necessary to search for new drugs. Some studies suggest the use of statins, which inhibit cholesterol synthesis in humans and also the initial processes of isoprenoid biosynthesis in the parasite. Thus, the objective of this study was to evaluate the activity of the statins pravastatin and simvastatin in HeLa cells infected in vitro with the RH strain of T. gondii. HeLa cells (1×10(5)) were infected with T. gondii tachyzoites (5×10(5)) following two different treatment protocols. In the first protocol, T. gondii tachyzoites were pretreated with pravastatin (50 and 100μg/mL) and simvastatin (1.56 and 3.125μg/mL) for 30min prior to infection. In the second, HeLa cells were first infected (5×10(5)) with tachyzoites and subsequently treated with pravastatin and simvastatin for 24h at the concentrations noted above. Initially, we evaluated the cytotoxicity of drugs by the MTT assay, number of tachyzoites adhered to cells, number of infected cells, and viability of tachyzoites by trypan blue exclusion. The supernatant of the cell cultures was collected post-treatment for determination of the pattern of Th1/Th2/Th17 cytokines by cytometric bead array. There was no cytotoxicity to HeLa cells with 50 and 100μg/mL pravastatin and 1.56 and 3.125μg/mL simvastatin. There was no change in the viability of tachyzoites that received pretreatment. Regarding the pre- and post-treatment of the cells with pravastatin and simvastatin alone, there was a reduction in adhesion, invasion and proliferation of cells to T. gondii. As for the production of cytokines, we found that IL-6 and IL-17 were significantly reduced in cells infected with T. gondii and treated with pravastatin and simvastatin, when compared to control. Based on these results, we can infer that pravastatin and simvastatin alone possess antiproliferative effects on tachyzoites forms

  18. TiN x O y coatings facilitate the initial adhesion of osteoblasts to create a suitable environment for their proliferation and the recruitment of endothelial cells.

    PubMed

    Moussa, M; Banakh, O; Wehrle-Haller, B; Fontana, P; Scherrer, S; Cattani, M; Wiskott, A; Durual, S

    2017-02-28

    Titanium-nitride-oxide coatings (TiN x O y ) improve osseointegration of endosseous implants. The exact mechanisms by which these effects are mediated are poorly understood except for an increase of osteoblast proliferation while a high degree of differentiation is maintained. One hypothesis holds that TiN x O y facilitates the initial spreading and adhesion of the osteoblasts. The aim of this work was to investigate the molecular mechanisms of osteoblast adhesion on TiN x O y as compared to microrough titanium SLA. A global view of the osseointegrative process, that is, taking into account other cell groups, especially endothelial cells, is also presented. To this aim, gene expression and focal adhesion analysis, cocultures and wound assays were performed early after seeding, from 6 h to 3 days. We demonstrated that TiN x O y coatings enhance osteoblast adhesion and spreading when compared to the standard microrough titanium. The integrin β1, either in association with α1 or with α2 plays a central role in these mechanisms. TiN x O y coatings optimize the process of osseointegration by acting at several levels, especially by upregulating osteoblast adhesion and proliferation, but also by supporting neovascularization and the development of a suitable inflammatory environment.

  19. Interplay of Anionic Charge, Poly(ethylene glycol), and Iodinated Tyrosine Incorporation within Tyrosine-derived Polycarbonates: Effects on Vascular Smooth Muscle Cell Adhesion, Proliferation and Motility

    PubMed Central

    Johnson, Patrick A.; Luk, Arnold; Demtchouk, Aleksey; Patel, Hiral; Sung, Hak-Joon; Treiser, Matthew D.; Gordonov, Simon; Sheihet, Larisa; Bolikal, Das; Kohn, Joachim; Moghe, Prabhas V.

    2009-01-01

    Regulation of smooth muscle cell adhesion, proliferation, and motility on biomaterials is critical to the performance of blood-contacting implants and vascular tissue engineering scaffolds. The goal of this study was to examine the underlying substrate-smooth muscle cell response relations, using a selection of polymers representative of an expansive library of multifunctional, tyrosine-derived polycarbonates. Three chemical components within the polymer structure were selectively varied through copolymerization: 1) the content of iodinated tyrosine to achieve X-ray visibility; 2) the content of poly(ethylene glycol) (PEG) to decrease protein adsorption and cell adhesivity; and 3) the content of desaminotyrosyl-tyrosine (DT) which regulates the rate of polymer degradation. Using quartz crystal microbalance with dissipation, we quantified differential serum protein adsorption behavior due to the chemical components DT, iodinated tyrosine, and PEG: increased PEG content within the polymer structure progressively decreased protein adsorption but the simultaneous presence of both DT and iodinated tyrosine reversed the effects of PEG. The complex interplay of these components was next tested on the adhesion, proliferation, and motility behavior cultured human aortic smooth muscle cells. The incorporation of PEG into the polymer reduced cell attachment, which was reversed in the presence of iodinated tyrosine. Further, we found that as little as 10% DT content was sufficient to negate the PEG effect in polymers containing iodinated tyrosine while in non-iodinated polymers the PEG effect on cell attachment was reversed. Cross-functional analysis of motility and proliferation revealed divergent substrate chemistry related cell response regimes. For instance, within the series of polymers containing both iodinated tyrosine and 10% of DT, increasing PEG levels lowered smooth muscle cell motility without a change in the rate of cell proliferation. In contrast, for non

  20. β1 integrin adhesion enhances IL-6 mediated STAT3 signaling in Myeloma cells: Implications for Microenvironment Influence on Tumor Survival and Proliferation

    PubMed Central

    Shain, Kenneth H.; Yarde, Danielle N.; Meads, Mark B.; Huang, Mei; Jove, Richard; Hazlehurst, Lori A.; Dalton, William S.

    2009-01-01

    The bone marrow microenvironmental components interleukin (IL)-6 and fibronectin (FN) individually influence the proliferation and survival of Multiple Myeloma (MM) cells; however, in vivo these effectors most likely work together. We examined signaling events, cell cycle progression, and levels of drug response in MM cells either adhered to FN via β1 integrins, stimulated with IL-6, or treated with the two combined. While G1/S cell cycle arrest associated with FN adhesion was overcome when IL-6 as added, the cell adhesion mediated drug resistance (CAM-DR) was maintained in the presence of IL-6. Concomitant exposure of MM cells to IL-6 and FN adhesion revealed a dramatic increase in STAT3 phosphorylation, nuclear translocation and DNA-binding, as compared to either IL-6 or FN adhesion alone in four MM cell lines. Importantly, this increase in STAT3 activation correlated with a novel association between STAT3 and gp130 in cells adhered to FN prior to stimulation with IL-6, relative to non-adherent cells. Taken together, these results suggest a mechanism by which collaborative signaling by β1 integrin and gp130 confers an increased survival advantage to MM cells. PMID:19155309

  1. Effects of decellularized matrices derived from periodontal ligament stem cells and SHED on the adhesion, proliferation and osteogenic differentiation of human dental pulp stem cells in vitro.

    PubMed

    Heng, Boon Chin; Zhu, Shaoyue; Xu, Jianguang; Yuan, Changyong; Gong, Ting; Zhang, Chengfei

    2016-04-01

    A major bottleneck to the therapeutic applications of dental pulp stem cells (DPSC) are their limited proliferative capacity ex vivo and tendency to undergo senescence. This may be partly due to the sub-optimal in vitro culture milieu, which could be improved by an appropriate extracellular matrix substratum. This study therefore examined decellularized matrix (DECM) from stem cells derived from human exfoliated deciduous teeth (SHED) and periodontal ligament stem cells (PDLSC), as potential substrata for DPSC culture. Both SHED-DECM and PDLSC-DECM promoted rapid adhesion and spreading of newly-seeded DPSC compared to bare polystyrene (TCPS), with vinculin immunocytochemistry showing expression of more focal adhesions by newly-adherent DPSC cultured on DECM versus TCPS. Culture of DPSC on SHED-DECM and PDLSC-DECM yielded higher proliferation of cell numbers compared to TCPS. The qRT-PCR data showed significantly higher expression of nestin by DPSC cultured on DECM versus the TCPS control. Osteogenic differentiation of DPSC was enhanced by culturing on PDLSC-DECM and SHED-DECM versus TCPS, as demonstrated by alizarin red S staining for mineralized calcium deposition, alkaline phosphatase assay and qRT-PCR analysis of key osteogenic marker expression. Hence, both SHED-DECM and PDLSC-DECM could enhance the ex vivo culture of DPSC under both non-inducing and osteogenic-inducing conditions.

  2. The effects of caffeic, coumaric and ferulic acids on proliferation, superoxide production, adhesion and migration of human tumor cells in vitro.

    PubMed

    Nasr Bouzaiene, Nouha; Kilani Jaziri, Soumaya; Kovacic, Hervé; Chekir-Ghedira, Leila; Ghedira, Kamel; Luis, José

    2015-11-05

    Reactive oxygen species are well-known mediators of various biological responses. In this study, we examined the effect of three phenolic acids, caffeic, coumaric and ferulic acids, on superoxide anion production, adhesion and migration of human lung (A549) and colon adenocarcinoma (HT29-D4) cancer cell lines. Proliferation of both tumor cells was inhibited by phenolic acids. Caffeic, coumaric and ferulic acids also significantly inhibited superoxide production in A549 and HT29-D4 cells. Superoxide anion production decreased by 92% and 77% at the highest tested concentration (200 µM) of caffeic acid in A549 and HT29-D4 cell lines respectively. Furthermore, A549 and HT29-D4 cell adhesion was reduced by 77.9% and 79.8% respectively at the higher tested concentration of ferulic acid (200 µM). Migration assay performed towards A549 cell line, revealed that tested compounds reduced significantly cell migration. At the highest concentration tested (200 µM), the covered surface was 7.7%, 9.5% and 35% for caffeic, coumaric or ferulic acids, respectively. These results demonstrate that caffeic, coumaric and ferulic acids may participate as active ingredients in anticancer agents against lung and colon cancer development, at adhesion and migration steps of tumor progression.

  3. Influence of different modifications of a calcium phosphate bone cement on adhesion, proliferation, and osteogenic differentiation of human bone marrow stromal cells.

    PubMed

    Vater, Corina; Lode, Anja; Bernhardt, Anne; Reinstorf, Antje; Heinemann, Christiane; Gelinsky, Michael

    2010-03-15

    Collagen and noncollagenous proteins of the extracellular bone matrix are able to stimulate bone cell activities and bone healing. The modification of calcium phosphate bone cements used as temporary bone replacement materials with these proteins seems to be a promising approach to accelerate new bone formation. In this study, we investigated adhesion, proliferation, and osteogenic differentiation of human bone marrow stromal cells (hBMSC) on Biocement D/collagen composites which have been modified with osteocalcin and O-phospho-L-serine. Modification with osteocalcin was carried out by its addition to the cement precursor before setting as well as by functionalization of the cement samples after setting and sterilization. hBMSC were cultured on these samples for 28 days with and without osteogenic supplements. We found a positive impact especially of the phosphoserine-modifications but also of both osteocalcin-modifications on differentiation of hBMSC indicated by higher expression of the osteoblastic markers matrix metalloproteinase-13 and bone sialo protein II. For hBMSC cultured on phosphoserine-containing composites, an increased proliferation has been observed. However, in case of the osteocalcin-modified samples, only osteocalcin adsorbed after setting and sterilization of the cement samples was able to promote initial adhesion and proliferation of hBMSC. The addition of osteocalcin before setting results in a finer microstructure but the biological activity of osteocalcin might be impaired due to the sterilization process. Thus, our data indicate that the initial adhesion and proliferation of hBMSC is enhanced rather by the biological activity of osteocalcin than by the finer microstructure.

  4. Blood-compatible poly(2-methoxyethyl acrylate) for the adhesion and proliferation of endothelial and smooth muscle cells.

    PubMed

    Sato, Chikako; Aoki, Makiko; Tanaka, Masaru

    2016-09-01

    Thrombus formation presents a serious hindrance in the development of functional artificial blood vessels, especially those with a small diameter. Endothelialization can prevent thrombus formation; however, the adhesion of endothelial cells to existing polymer materials is generally weak. Therefore, polymers that have both anti-thrombotic and endothelialization properties do not currently exist. We previously reported that platelets do not adhere to poly(2-methoxyethyl acrylate) (PMEA) or poly(tetrahydrofurfuryl acrylate)(PTHFA). Here, we investigated whether endothelial cells and smooth muscle cells, both of which are blood vessel components, could adhere to these synthetic polymers. Polyethylene terephthalate films were coated with PMEA and PTHFA using a spin-coater. Human umbilical vein endothelial cells or aorta smooth muscle cells were seeded on the polymer surfaces, after which we analyzed the number of adherent cells, their morphologies and vinculin expression. We found that both endothelial and smooth muscle cells adhered to PMEA and PTHFA, while platelets did not. We propose that, by using PMEA and PTHFA with no modifications, it should be possible to develop artificial blood vessels with both anti-thrombotic and endothelialization properties. In addition, we discuss the mechanism of selective cell adhesion in PMEA and PTHFA.

  5. Increased Cell Proliferation and Gene Expression of Genes Related to Bone Remodeling, Cell Adhesion and Collagen Metabolism in the Periodontal Ligament of Unopposed Molars in Growing Rats

    PubMed Central

    Dorotheou, Domna; Farsadaki, Vassiliki; Bochaton-Piallat, Marie-Luce; Giannopoulou, Catherine; Halazonetis, Thanos D.; Kiliaridis, Stavros

    2017-01-01

    Tooth eruption, the process by which teeth emerge from within the alveolar bone into the oral cavity, is poorly understood. The post-emergent phase of tooth eruption continues throughout life, in particular, if teeth are not opposed by antagonists. The aim of the present study was to better understand the molecular processes underlying post-emergent tooth eruption. Toward this goal, we removed the crowns of the maxillary molars on one side of the mouth of 14 young rats and examined gene expression patterns in the periodontal ligaments (PDLs) of the ipsilateral and contralateral mandibular molars, 3 and 15 days later. Nine untreated rats served as controls. Expression of six genes, Adamts18, Ostn, P4ha3, Panx3, Pth1r, and Tnmd, was upregulated in unopposed molars relative to molars with antagonists. These genes function in osteoblast differentiation and proliferation, cell adhesion and collagen metabolism. Proliferation of PDL cells also increased following loss of the antagonist teeth. Interestingly, mutations in PTH1R have been linked to defects in the post-emergent phase of tooth eruption in humans. We conclude that post-emergent eruption of unopposed teeth is associated with gene expression patterns conducive to alveolar bone formation and PDL remodeling. PMID:28239357

  6. Tetraploid cells produced by absence of substrate adhesion during cytokinesis are limited in their proliferation and enter senescence after DNA replication.

    PubMed

    De Santis Puzzonia, Marco; Gonzalez, Laetitia; Ascenzi, Sonia; Cundari, Enrico; Degrassi, Francesca

    2016-01-01

    Tetraploidy has been proposed as an intermediate state in neoplastic transformation due to the intrinsic chromosome instability of tetraploid cells. Despite the identification of p53 as a major factor in growth arrest of tetraploid cells, it is still unclear whether the p53-dependent mechanism for proliferation restriction is intrinsic to the tetraploid status or dependent on the origin of tetraploidy. Substrate adherence is fundamental for cytokinesis completion in adherent untransformed cells. Here we show that untransformed fibroblast cells undergoing mitosis in suspension produce binucleated tetraploid cells due to defective cleavage furrow constriction that leads to incomplete cell abscission. Binucleated cells obtained after loss of substrate adhesion maintain an inactive p53 status and are able to progress into G1 and S phase. However, binucleated cells arrest in G2, accumulate p53 and are not able to enter mitosis as no tetraploid metaphases were recorded after one cell cycle time. In contrast, tetraploid metaphases were found following pharmacological inhibition of Chk1 kinase, suggesting the involvement of the ATR/Chk1 pathway in the G2 arrest of binucleated cells. Interestingly, after persistence in the G2 phase of the cell cycle, a large fraction of binucleated cells become senescent. These findings identify a new pathway of proliferation restriction for tetraploid untransformed cells that seems to be specific for loss of adhesion-dependent cytokinesis failure. This involves Chk1 and p53 activation during G2. Inhibition of growth and entrance into senescence after cytokinesis in suspension may represent an important mechanism to control tumor growth. In fact, anchorage independent growth is a hallmark of cancer and it has been demonstrated that binucleated transformed cells can enter a cycle of anchorage independent growth.

  7. Indirect coating of RGD peptides using a poly-L-lysine spacer enhances jaw periosteal cell adhesion, proliferation, and differentiation into osteogenic tissue.

    PubMed

    Ardjomandi, N; Klein, C; Kohler, K; Maurer, A; Kalbacher, H; Niederländer, J; Reinert, S; Alexander, D

    2012-08-01

    The aim of our study was to generate a biofunctionalized, three-dimensional (3D) biomaterial to enhance jaw periosteal cell (JPC) adhesion and differentiation into osteogenic tissue. Therefore, open-cell polylactic acid (OPLA) scaffolds were coated covalently with different RGD peptides (a conserved recognition sequence of the most ECM proteins--arginine-glycine-asparagine) and different coating variants. The linear and cyclic RGD peptides were either applied directly or indirectly via a poly-L-lysine (PLL) spacer. JPCs were analyzed on coated constructs in 2D and 3D cultures and showed enhanced rates for indirectly coated scaffolds using the PLL spacer. By gene expression, we detected significantly increased levels of osteogenic marker genes, such as alkaline phosphatase, RUNX2, and AMELY in JPCs seeded onto PLL/linear RGD constructs compared to the otherwise-coated constructs. An analysis of the JPC mineralization capacity revealed the highest amounts of calcium-phosphate precipitates in cells growing within the PLL/linear scaffolds. Additionally, the JPC adhesion behavior on OPLA scaffolds seems to be mediated by ITGB3, ITGB1, and ITGAV, as shown by blocking assays. We concluded that coating of OPLA constructs with linear RGD peptides via PLL represents a suitable approach for functionalizing the polymer surface and enhancing adhesion, proliferation, and mineralization of JPCs.

  8. PED/PEA-15 interacts with the 67 kD laminin receptor and regulates cell adhesion, migration, proliferation and apoptosis

    PubMed Central

    Formisano, Pietro; Ragno, Pia; Pesapane, Ada; Alfano, Daniela; Alberobello, Anna Teresa; Rea, Vincenza Elena Anna; Giusto, Raffaella; Rossi, Francesca W; Beguinot, Francesco; Rossi, Guido; Montuori, Nunzia

    2012-01-01

    Abstract Phosphoprotein enriched in diabetes/phosphoprotein enriched in astrocytes-15 kD (PED/PEA-15) is an anti-apoptotic protein whose expression is increased in several human cancers. In addition to apoptosis, PED/PEA-15 is involved in the regulation of other major cellular functions, including cell adhesion, migration, proliferation and glucose metabolism. To further understand the functions of this protein, we performed a yeast two-hybrid screening using PED/PEA-15 as a bait and identified the 67 kD high-affinity laminin receptor (67LR) as an interacting partner. 67 kD laminin receptor is a non-integrin cell-surface receptor for the extracellular matrix (ECM), derived from the dimerization of a 37 kD cytosolic precursor (37LRP). The 67LR is highly expressed in human cancers and widely recognized as a molecular marker of metastatic aggressiveness. The molecular interaction of PED/PEA-15 with 67LR was confirmed by pull-down experiments with recombinant His-tagged 37LRP on lysates of PED/PEA-15 transfected HEK-293 cells. Further, overexpressed or endogenous PED/PEA-15 was co-immunoprecipitated with 67LR in PED/PEA-15-transfected HEK-293 cells and in U-373 glioblastoma cells, respectively. PED/PEA-15 overexpression significantly increased 67LR-mediated HEK-293 cell adhesion and migration to laminin that, in turn, determined PED/PEA-15 phosphorylation both in Ser-104 and Ser-116, thus enabling cell proliferation and resistance to apoptosis. PED/PEA-15 ability to induce cell responses to ECM-derived signals through interaction with 67LR may be of crucial importance for tumour cell survival in a poor microenvironment, thus favouring the metastatic spread and colonization. PMID:21895963

  9. EGCG Inhibits Proliferation, Invasiveness and Tumor Growth by Up-Regulation of Adhesion Molecules, Suppression of Gelatinases Activity, and Induction of Apoptosis in Nasopharyngeal Carcinoma Cells

    PubMed Central

    Fang, Chih-Yeu; Wu, Chung-Chun; Hsu, Hui-Yu; Chuang, Hsin-Ying; Huang, Sheng-Yen; Tsai, Ching-Hwa; Chang, Yao; Tsao, George Sai-Wah; Chen, Chi-Long; Chen, Jen-Yang

    2015-01-01

    (−)-Epigallocatechin-3-gallate (EGCG), a major green tea polyphenol, has been shown to inhibit the proliferation of a variety of tumor cells. Epidemiological studies have shown that drinking green tea can reduce the incidence of nasopharyngeal carcinoma (NPC), yet the underlying mechanism is not well understood. In this study, the inhibitory effect of EGCG was tested on a set of Epstein Barr virus-negative and -positive NPC cell lines. Treatment with EGCG inhibited the proliferation of NPC cells but did not affect the growth of a non-malignant nasopharyngeal cell line, NP460hTert. Moreover, EGCG treated cells had reduced migration and invasive properties. The expression of the cell adhesion molecules E-cadherin and β-catenin was found to be up-regulated by EGCG treatment, while the down-regulation of matrix metalloproteinases (MMP)-2 and MMP-9 were found to be mediated by suppression of extracellular signal-regulated kinase (ERK) phosphorylation and AP-1 and Sp1 transactivation. Spheroid formation by NPC cells in suspension was significantly inhibited by EGCG. Oral administration of EGCG was capable of suppressing tumor growth in xenografted mice bearing NPC tumors. Treatment with EGCG was found to elevate the expression of p53 and p21, and eventually led to apoptosis of NPC cells via caspase 3 activation. The nuclear translocation of NF-κB and β-catenin was also suppressed by EGCG treatment. These results indicate that EGCG can inhibit the proliferation and invasiveness, and induce apoptosis, of NPC cells, making it a promising agent for chemoprevention or adjuvant therapy of NPC. PMID:25625511

  10. Involvement of small G protein RhoB in the regulation of proliferation, adhesion and migration by dexamethasone in osteoblastic cells

    PubMed Central

    Wang, Yan; Li, Yidong; Xu, Weidong; Lu, Jian

    2017-01-01

    Long-term exposure to therapeutic doses of glucocorticoids (GCs) results in bone remodeling, which frequently causes osteoporosis and fracture healing retardation because of the abnormality of osteoblastic proliferation and differentiation. The mechanisms of GCs’ effect on osteoblasts are largely unknown. In this present study, we found that dexamethasone (Dex) could induce the expression of the small G protein, RhoB, in mRNA and protein levels in the osteoblast-derived osteosarcoma cell lines MG-63. The up-regulation of RhoB mRNA by Dex mainly occurs at posttranscriptional level by increasing its mRNA stability through PI-3K/Akt and p38 mitogen-activated protein kinase signaling pathways. Over-expression of RhoB in MG-63 cells magnified while down-regulation of RhoB level by RNA interference impaired Dex-induced growth inhibition but not differentiation. What’s more, over-expression of RhoB mimicked the effect of Dex on cell adhesion and migration. And interfering RhoB expression partially suppressed Dex-induced pro-adhesion and anti-migration in MG-63 cells. In conclusion, these results indicate that RhoB plays an important role in the pathological effect of Dex on osteoblastic growth and migration, which is a part of the mechanisms of GCs’ adverse effect on bone remodeling. PMID:28323887

  11. Silicon-hydroxyapatite bioactive coatings (Si-HA) from diatomaceous earth and silica. Study of adhesion and proliferation of osteoblast-like cells.

    PubMed

    López-Alvarez, M; Solla, E L; González, P; Serra, J; León, B; Marques, A P; Reis, R L

    2009-05-01

    The aim of this study consisted on investigating the influence of silicon substituted hydroxyapatite (Si-HA) coatings over the human osteoblast-like cell line (SaOS-2) behaviour. Diatomaceous earth and silica, together with commercial hydroxyapatite were respectively the silicon and HA sources used to produce the Si-HA coatings. HA coatings with 0 wt% of silicon were used as control of the experiment. Pulsed laser deposition (PLD) was the selected technique to deposit the coatings. The Si-HA thin films were characterized by Fourier Transformed Infrared Spectroscopy (FTIR) demonstrating the efficient transfer of Si to the HA structure. The in vitro cell culture was established to assess the cell attachment, proliferation and osteoblastic activity respectively by, Scanning Electron Microscopy (SEM), DNA and alkaline phosphatase (ALP) quantification. The SEM analysis demonstrated a similar adhesion behaviour of the cells on the tested materials and the maintenance of the typical osteoblastic morphology along the time of culture. The Si-HA coatings did not evidence any type of cytotoxic behaviour when compared with HA coatings. Moreover, both the proliferation rate and osteoblastic activity results showed a slightly better performance on the Si-HA coatings from diatoms than on the Si-HA from silica.

  12. Influence of sodium hypochlorite treatment of electropolished and magnetoelectropolished nitinol surfaces on adhesion and proliferation of MC3T3 pre-osteoblast cells.

    PubMed

    Rokicki, Ryszard; Haider, Waseem; Hryniewicz, Tadeusz

    2012-09-01

    The influence of 6 % sodium hypochlorite (NaClO) treatment on adhesion and proliferation of MC3T3 pre-osteoblast cells seeded on electropolished (EP) and magnetoelectropolished (MEP) nitinol surfaces were investigated. The chemistry, topography, roughness, surface energy, wettability of EP and MEP nitinol surfaces before and after NaClO treatment were studied with X-ray photoelectron spectroscopy (XPS), profilometry, and contact angle meter. In vitro interaction of osteoblast cell and NaClO treated EP and MEP nitinol surfaces were assessed after 3 days of incubation by scanning electron microscopy. The XPS analysis shows that NaClO treatment increases oxygen content especially in subsurface oxide layer of EP and MEP nitinol. The changes of both basic components of nitinol, namely nickel and titanium in oxide layer, were negligible. The NaClO treatment did not influence physico-morphological surface properties of EP and MEP nitinol to a big extent. The osteoblast cells show remarkable adherence and proliferation improvement on NaClO treated EP and MEP nitinol surfaces. After 3 days of incubation they show almost total confluence on both NaClO treated surfaces. The present study shows that NaClO treatment of EP and MEP nitinol surfaces alters oxide layer by enriching it in oxygen and by this improves bone cell-nitinol interaction.

  13. Photo-initiated grafting of gelatin/N-maleic acyl-chitosan to enhance endothelial cell adhesion, proliferation and function on PLA surface.

    PubMed

    Zhu, Aiping; Zhao, Feng; Ma, Teng

    2009-07-01

    Vascular graft surface properties significantly affect adhesion, growth and function of endothelial cells (ECs). The bulk degradation property of poly(lactic acid) (PLA) makes it possible for it to be replaced by cellular materials and PLA is desirable as a scaffold material for vascular grafts. However, PLA has an unfavorable surface property for EC adhesion and proliferation due to the lack of a selective cell adhesion motif. Photo-initiated surface-grafting polymerization is a promising method for immobilizing certain biomacromolecules on material surfaces without compromising bulk properties. N-Maleic acyl-chitosan (NMCS) is a novel biocompatible amphiphilic derivative of chitosan with double bonds and can be initiated by ultraviolet light. In this study, gelatin was complexed with NMCS via hydrophobic interaction, and gel/NMCS complex thus formed was then grafted on the PLA surface to improve EC biocompatibility. X-ray photoelectron and Fourier transform infrared spectroscopy, and water contact angle measurement confirmed immobilization of the gel/NMCS complex on PLA surface. Moreover, the gel/NMCS modified PLA enhanced human umbilical vein endothelial cell (HUVEC) spreading and flattening, and promoted the expression of more structured CD31 and vWF compared to unmodified PLA film. Compared to the unmodified PLA surface, the HUVECs on the modified PLA surface had elevated uptake of acetylated low-density lipoprotein, and maintained the ability to modulate metabolic activity upon exposure to shear stress at 5dyncm(-2) by up-regulating nitric oxide and prostacyclin production. Cell retention was 1.6 times higher on the gel/NMCS-PLA surface, demonstrating its improved potential for hemocompatibility. These results indicate that photo-initiated surface-grafting of the biomimetic gel/NMCS complex is an effective method to modify material surfaces as vascular grafts.

  14. [Endothelial cell adhesion molecules].

    PubMed

    Ivanov, A N; Norkin, I A; Puchin'ian, D M; Shirokov, V Iu; Zhdanova, O Iu

    2014-01-01

    The review presents current data concerning the functional role of endothelial cell adhesion molecules belonging to different structural families: integrins, selectins, cadherins, and the immunoglobulin super-family. In this manuscript the regulatory mechanisms and factors of adhesion molecules expression and distribution on the surface of endothelial cells are discussed. The data presented reveal the importance of adhesion molecules in the regulation of structural and functional state of endothelial cells in normal conditions and in pathology. Particular attention is paid to the importance of these molecules in the processes of physiological and pathological angiogenesis, regulation of permeability of the endothelial barrier and cell transmigration.

  15. Bio-safe processing of polylactic-co-caprolactone and polylactic acid blends to fabricate fibrous porous scaffolds for in vitro mesenchymal stem cells adhesion and proliferation.

    PubMed

    Salerno, Aurelio; Guarino, Vincenzo; Oliviero, Olimpia; Ambrosio, Luigi; Domingo, Concepción

    2016-06-01

    In this study, the design and fabrication of porous scaffolds, made of blends of polylactic-co-caprolactone (PLC) and polylactic acid (PLA) polymers, for tissue engineering applications is reported. The scaffolds are prepared by means of a bio-safe thermally induced phase separation (TIPS) approach with or without the addition of NaCl particles used as particulate porogen. The scaffolds are characterized to assess their crystalline structure, morphology and mechanical properties, and the texture of the pores and the pore size distribution. Moreover, in vitro human mesenchymal stem cells (hMSCs) culture tests have been carried out to demonstrate the biocompatibility of the scaffolds. The results of this study demonstrate that all of the scaffold materials processed by means of TIPS process are semi-crystalline. Furthermore, the blend composition affected polymer crystallization and, in turn, the nano and macro-structural properties of the scaffolds. Indeed, neat PLC and neat PLA crystallize into globular and randomly arranged sub micro-size scale fibrous conformations, respectively. Concomitantly, the addition of NaCl particles during the fabrication route allows for the creation of an interconnected network of large pores inside the primary structure while resulted in a significant decrease of scaffolds mechanical response. Finally, the results of cell culture tests demonstrate that both the micro and macro-structure of the scaffold affect the in vitro hMSCs adhesion and proliferation.

  16. Sodium phenylacetate (NaPa) induces modifications of the proliferation, the adhesion and the cell cycle of tumoral epithelial breast cells.

    PubMed

    Thibout, D; Kraemer, M; Di Benedetto, M; Saffar, L; Gattegno, L; Derbin, C; Crépin, M

    1999-01-01

    Sodium phenylacetate (NaPa), a physiological product of phenylalanine metabolism, present in micromolar concentrations in human plasma, has been shown to induce in vivo and in vitro cytostatic antiproliferative effects at millimolar concentrations. Cadherin molecules are powerful invasion suppressor molecules and the reduction of E-cadherin expression plays an important role in the invasion and metastasis of human breast cancer. In this study, we demonstrated, on one hand, that NaPa stimulated aggregation by increasing the expression of E-cadherin at the surface of breast cancer MCF-7ras cells transformed by Ha-ras oncogene and inhibited its expression in MCF-7 cells. We demonstrated that NaPa increased the formation of MCF-7ras cell aggregates and did not alter the formation of MCF-7 cell aggregates. By Northern blot, we demonstrated that the E-cadherin expression was not regulated at the transcriptional level. On the other hand, we analyzed the cell cycle of these 2 cell lines after NaPa treatment and showed that NaPa induced arrest at the G1/S phase in both MCF-7 and MCF-7ras cells. bFGF increased the growth of MCF-7 cells, but inhibited MCF-7ras cell proliferation. NaPa treatment suppressed the stimulation of MCF-7 cell proliferation and increased MCF-7ras cell growth inhibition. We have demonstrated a new target of NaPa action in blocking the cell cycle of tumor cells in G0/G1. We suggest that the anti-proliferative effect of NaPa associated to the restoration of the cadherin function in human mammary carcinoma cells indicates that NaPa could be a novel therapeutic agent in breast cancer.

  17. MiR-9-5p, miR-675-5p and miR-138-5p Damages the Strontium and LRP5-Mediated Skeletal Cell Proliferation, Differentiation, and Adhesion.

    PubMed

    Sun, Tianhao; Leung, Frankie; Lu, William W

    2016-02-15

    This study was designed to evaluate the effects of strontium on the expression levels of microRNAs (miRNAs) and to explore their effects on skeletal cell proliferation, differentiation, adhesion, and apoptosis. The targets of these miRNAs were also studied. Molecular cloning, cell proliferation assay, cell apoptosis assay, quantitative real-time PCR, and luciferase reporter assay were used. Strontium altered the expression levels of miRNAs in vitro and in vivo. miR-9-5p, miR-675-5p, and miR-138-5p impaired skeletal cell proliferation, cell differentiation and cell adhesion. miR-9-5p and miR-675-5p induced MC3T3-E1 cell apoptosis more specifically than miR-138-5p. miR-9-5p, miR-675-5p, and miR-138-5p targeted glycogen synthase kinase 3 β (GSK3β), ATPase Aminophospholipid Transporter Class I Type 8A Member 2 (ATP8A2), and Eukaryotic Translation Initiation Factor 4E Binding Protein 1 (EIF4EBP1), respectively. Low-density lipoprotein receptor-related protein 5 (LRP5) played a positive role in skeletal development. miR-9-5p, miR-675-5p, and miR-138-5p damage strontium and LRP5-mediated skeletal cell proliferation, differentiation, and adhesion, and induce cell apoptosis by targeting GSK3β, ATP8A2, and EIF4EBP1, respectively.

  18. BAG-1 enhances cell-cell adhesion, reduces proliferation and induces chaperone-independent suppression of hepatocyte growth factor-induced epidermal keratinocyte migration

    SciTech Connect

    Hinitt, C.A.M.; Wood, J.; Lee, S.S.; Williams, A.C.; Howarth, J.L.; Glover, C.P.; Uney, J.B.; Hague, A.

    2010-08-01

    Cell motility is important in maintaining tissue homeostasis, facilitating epithelial wound repair and in tumour formation and progression. The aim of this study was to determine whether BAG-1 isoforms regulate epidermal cell migration in in vitro models of wound healing. In the human epidermal cell line HaCaT, endogenous BAG-1 is primarily nuclear and increases with confluence. Both transient and stable p36-Bag-1 overexpression resulted in increased cellular cohesion. Stable transfection of either of the three human BAG-1 isoforms p36-Bag-1 (BAG-1S), p46-Bag-1 (BAG-1M) and p50-Bag-1 (BAG-1L) inhibited growth and wound closure in serum-containing medium. However, in response to hepatocyte growth factor (HGF) in serum-free medium, BAG-1S/M reduced communal motility and colony scattering, but BAG-1L did not. In the presence of HGF, p36-Bag-1 transfectants retained proliferative response to HGF with no change in ERK1/2 activation. However, the cells retained E-cadherin localisation at cell-cell junctions and exhibited pronounced cortical actin. Point mutations in the BAG domain showed that BAG-1 inhibition of motility is independent of its function as a chaperone regulator. These findings are the first to suggest that BAG-1 plays a role in regulating cell-cell adhesion and suggest an important function in epidermal cohesion.

  19. Mapping cell surface adhesion by rotation tracking and adhesion footprinting

    NASA Astrophysics Data System (ADS)

    Li, Isaac T. S.; Ha, Taekjip; Chemla, Yann R.

    2017-03-01

    Rolling adhesion, in which cells passively roll along surfaces under shear flow, is a critical process involved in inflammatory responses and cancer metastasis. Surface adhesion properties regulated by adhesion receptors and membrane tethers are critical in understanding cell rolling behavior. Locally, adhesion molecules are distributed at the tips of membrane tethers. However, how functional adhesion properties are globally distributed on the individual cell’s surface is unknown. Here, we developed a label-free technique to determine the spatial distribution of adhesive properties on rolling cell surfaces. Using dark-field imaging and particle tracking, we extract the rotational motion of individual rolling cells. The rotational information allows us to construct an adhesion map along the contact circumference of a single cell. To complement this approach, we also developed a fluorescent adhesion footprint assay to record the molecular adhesion events from cell rolling. Applying the combination of the two methods on human promyelocytic leukemia cells, our results surprisingly reveal that adhesion is non-uniformly distributed in patches on the cell surfaces. Our label-free adhesion mapping methods are applicable to the variety of cell types that undergo rolling adhesion and provide a quantitative picture of cell surface adhesion at the functional and molecular level.

  20. Mapping cell surface adhesion by rotation tracking and adhesion footprinting

    PubMed Central

    Li, Isaac T. S.; Ha, Taekjip; Chemla, Yann R.

    2017-01-01

    Rolling adhesion, in which cells passively roll along surfaces under shear flow, is a critical process involved in inflammatory responses and cancer metastasis. Surface adhesion properties regulated by adhesion receptors and membrane tethers are critical in understanding cell rolling behavior. Locally, adhesion molecules are distributed at the tips of membrane tethers. However, how functional adhesion properties are globally distributed on the individual cell’s surface is unknown. Here, we developed a label-free technique to determine the spatial distribution of adhesive properties on rolling cell surfaces. Using dark-field imaging and particle tracking, we extract the rotational motion of individual rolling cells. The rotational information allows us to construct an adhesion map along the contact circumference of a single cell. To complement this approach, we also developed a fluorescent adhesion footprint assay to record the molecular adhesion events from cell rolling. Applying the combination of the two methods on human promyelocytic leukemia cells, our results surprisingly reveal that adhesion is non-uniformly distributed in patches on the cell surfaces. Our label-free adhesion mapping methods are applicable to the variety of cell types that undergo rolling adhesion and provide a quantitative picture of cell surface adhesion at the functional and molecular level. PMID:28290531

  1. Using real-time impedance-based assays to monitor the effects of fibroblast-derived media on the adhesion, proliferation, migration and invasion of colon cancer cells.

    PubMed

    Dowling, Catríona M; Herranz Ors, Carmen; Kiely, Patrick A

    2014-07-29

    Increasing our knowledge of the mechanisms regulating cell proliferation, migration and invasion are central to understanding tumour progression and metastasis. The local tumour microenvironment contributes to the transformed phenotype in cancer by providing specific environmental cues that alter the cells behaviour and promotes metastasis. Fibroblasts have a strong association with cancer and in recent times there has been some emphasis in designing novel therapeutic strategies that alter fibroblast behaviour in the tumour microenvironment. Fibroblasts produce growth factors, chemokines and many of the proteins laid down in the ECM (extracellular matrix) that promote angiogenesis, inflammation and tumour progression. In this study, we use a label-free RTCA (real-time cell analysis) platform (xCELLigence) to investigate how media derived from human fibroblasts alters cancer cell behaviour. We used a series of complimentary and novel experimental approaches to show HCT116 cells adhere, proliferate and migrate significantly faster in the presence of media from human fibroblasts. As well as this, we used the xCELLigence CIM-plates system to show that HCT116 cells invade matrigel layers aggressively when migrating towards media derived from human fibroblasts. These data strongly suggest that fibroblasts have the ability to increase the migratory and invasive properties of HCT116 cells. This is the first study that provides real-time data on fibroblast-mediated migration and invasion kinetics of colon cancer cells.

  2. CD44 interaction with ankyrin and IP3 receptor in lipid rafts promotes hyaluronan-mediated Ca2+ signaling leading to nitric oxide production and endothelial cell adhesion and proliferation.

    PubMed

    Singleton, Patrick A; Bourguignon, Lilly Y W

    2004-04-15

    In this study, we have showed that aortic endothelial cells (GM7372A cell line) express CD44v10 [a hyaluronan (HA) receptor], which is significantly enriched in cholesterol-containing lipid rafts (characterized as caveolin-rich plasma membrane microdomains). HA binding to CD44v10 promotes recruitment of the cytoskeletal protein, ankyrin and inositol 1,4,5-triphosphate (IP3) receptor into cholesterol-containing lipid rafts. The ankyrin repeat domain (ARD) of ankyrin is responsible for binding IP3 receptor to CD44v10 at lipid rafts and subsequently triggering HA/CD44v10-mediated intracellular calcium (Ca2+) mobilization leading to a variety of endothelial cell functions such as nitric oxide (NO) production, cell adhesion and proliferation. Further analyses indicate (i) disruption of lipid rafts by depleting cholesterol from the membranes of GM7372A cells (using methyl-beta-cyclodextrin treatment) or (ii) interference of endogenous ankyrin binding to CD44 and IP3 receptor using overexpression of ARD fragments (by transfecting cells with ARDcDNA) not only abolishes ankyrin/IP3 receptor accumulation into CD44v10/cholesterol-containing lipid rafts, but also blocks HA-mediated Ca2+ signaling and endothelial cell functions. Taken together, our findings suggest that CD44v10 interaction with ankyrin and IP3 receptor in cholesterol-containing lipid rafts plays an important role in regulating HA-mediated Ca2+ signaling and endothelial cell functions such as NO production, cell adhesion and proliferation.

  3. Notch-Mediated Cell Adhesion

    PubMed Central

    Murata, Akihiko; Hayashi, Shin-Ichi

    2016-01-01

    Notch family members are generally recognized as signaling molecules that control various cellular responses in metazoan organisms. Early fly studies and our mammalian studies demonstrated that Notch family members are also cell adhesion molecules; however, information on the physiological roles of this function and its origin is limited. In this review, we discuss the potential present and ancestral roles of Notch-mediated cell adhesion in order to explore its origin and the initial roles of Notch family members dating back to metazoan evolution. We hypothesize that Notch family members may have initially emerged as cell adhesion molecules in order to mediate multicellularity in the last common ancestor of metazoan organisms. PMID:26784245

  4. Effect of various concentrations of Ti in hydrocarbon plasma polymer films on the adhesion, proliferation and differentiation of human osteoblast-like MG-63 cells

    NASA Astrophysics Data System (ADS)

    Vandrovcova, Marta; Grinevich, Andrey; Drabik, Martin; Kylian, Ondrej; Hanus, Jan; Stankova, Lubica; Lisa, Vera; Choukourov, Andrei; Slavinska, Danka; Biederman, Hynek; Bacakova, Lucie

    2015-12-01

    Hydrocarbon polymer films (ppCH) enriched with various concentrations of titanium were deposited on microscopic glass slides by magnetron sputtering from a Ti target. The maximum concentration of Ti (about 20 at.%) was achieved in a pure argon atmosphere. The concentration of Ti decreased rapidly after n-hexane vapors were introduced into the plasma discharge, and reached zero values at n-hexane flow of 0.66 sccm. The decrease in Ti concentration was associated with decreasing oxygen and titanium carbide concentration in the films, decreasing wettability (the water drop contact angle increased from 20° to 91°) and decreasing root-mean-square roughness (from 3.3 nm to 1.0 nm). The human osteoblast-like MG-63 cells cultured on pure ppCH films and on films with 20 at.% of Ti showed relatively high concentrations of ICAM-1, a marker of cell immune activation. Lower concentrations of Ti (mainly 5 at.%) improved cell adhesion and osteogenic differentiation, as revealed by higher concentrations of talin, vinculin and osteocalcin. Higher Ti concentrations (15 at.%) supported cell growth, as indicated by the highest final cell population densities on day 7 after seeding. Thus, enrichment of ppCH films with appropriate concentrations of Ti makes these films more suitable for potential coatings of bone implants.

  5. Hydroxyapatite promotes superior keratocyte adhesion and proliferation in comparison with current keratoprosthesis skirt materials

    PubMed Central

    Mehta, J S; Futter, C E; Sandeman, S R; Faragher, R G A F; Hing, K A; Tanner, K E; Allan, B D S

    2005-01-01

    Aim: Published clinical series suggest the osteoodontokeratoprosthesis (OOKP) may have a lower extrusion rate than current synthetic keratoprostheses. The OOKP is anchored in the eye wall by autologous tooth. The authors’ aim was to compare adhesion, proliferation, and morphology for telomerase transformed keratocytes seeded on calcium hydroxyapatite (the principal mineral constituent of tooth) and materials used in the anchoring elements of commercially available synthetic keratoprostheses. Methods: Test materials were hydroxyapatite, polytetrafluoroethylene (PTFE), polyhydroxyethyl methacrylate (HEMA), and glass (control). Cell adhesion and viability were quantified at 4 hours, 24 hours, and 1 week using a calcein-AM/EthD-1 viability/cytotoxicity assay. Focal contact expression and cytoskeletal organisation were studied at 24 hours by confocal microscopy with immunoflourescent labelling. Further studies of cell morphology were performed using light and scanning electron microscopy. Results: Live cell counts were significantly greater on hydroxyapatite surfaces at each time point (p<0.04). Dead cell counts were significantly higher for PTFE at 7 days (p<0.002). ß1 integrin expression was highest on hydroxyapatite. Adhesion structures were well expressed in flat, spread out keratocytes on both HA and glass. Keratocytes tended to be thinner and spindle shaped on PTFE. The relatively few keratocytes visible on HEMA test surfaces were rounded and poorly adherent. Conclusions: Keratocyte adhesion, spreading, and viability on hydroxyapatite test surfaces is superior to that seen on PTFE and HEMA. Improving the initial cell adhesion environment in the skirt element of keratoprostheses may enhance tissue integration and reduce device failure rates. PMID:16170132

  6. Surface modification of tantalum pentoxide coatings deposited by magnetron sputtering and correlation with cell adhesion and proliferation in in vitro tests

    NASA Astrophysics Data System (ADS)

    Zykova, A.; Safonov, V.; Goltsev, A.; Dubrava, T.; Rossokha, I.; Donkov, N.; Yakovin, S.; Kolesnikov, D.; Goncharov, I.; Georgieva, V.

    2016-03-01

    The effect was analyzed of surface treatment by argon ions on the surface properties of tantalum pentoxide coatings deposited by reactive magnetron sputtering. The structural parameters of the as-deposited coatings were investigated by means of transmission electron microscopy, atomic force microscopy and scanning electron microscopy. X-ray diffraction profiles and X-ray photoelectron spectra were also acquired. The total surface free energy (SFE), the polar, dispersion parts and fractional polarities, were estimated by the Owens-Wendt-Rabel-Kaeble method. The adhesive and proliferative potentials of bone marrow cells were evaluated for both Ta2O5 coatings and Ta2O5 coatings deposited by simultaneous bombardment by argon ions in in vitro tests.

  7. Mechanics of Nascent Cell Adhesions

    NASA Astrophysics Data System (ADS)

    Mejean, Cecile O.; Schaefer, Andrew W.; Forscher, Paul; Dufresne, Eric R.

    2009-03-01

    Cells have the ability to sense and respond to mechanical and biochemical cues from their environment. In neurons, the binding and restraint of transmembrane cell adhesion molecules (CAMs) can trigger acute periods of axon growth. Preceding growth, the cell must create a stiff mechanical linkage between the CAM and the cytoskeleton. Using holographic optical tweezers, we manipulate CAM-coated beads on the membrane of the cell. We investigate the dynamics of the mechanical properties of this linkage as a function of time, applied force, and CAM density. We find that CAM-coated beads exhibit stochastic intermittent binding to the cytoskeleton. In time, we observed that the adhesions stiffen and their mechanical properties depend on the applied force. Treatment of cells with small molecules that alter cytoskeletal dynamics are used to probe the roles of actin filament assembly and myosin motor activity in adhesion formation.

  8. In vitro evaluation of poly(ethylene glycol)-block-poly(ɛ-caprolactone) methyl ether copolymer coating effects on cells adhesion and proliferation

    NASA Astrophysics Data System (ADS)

    Rusen, Laurentiu; Neacsu, Patricia; Cimpean, Anisoara; Valentin, Ion; Brajnicov, Simona; Dumitrescu, L. N.; Banita, Janina; Dinca, Valentina; Dinescu, Maria

    2016-06-01

    Understanding and controlling natural and synthetic biointerfaces is known to be the key to a wide variety of application within cell culture and tissue engineering field. As both material characteristics and methods are important in tailoring biointerfaces characteristics, in this work we explore the feasibility of using Matrix Assisted Pulsed Laser Evaporation technique for obtaining synthetic copolymeric biocoatings (i.e. poly(ethylene glycol)-block-poly(ɛ-caprolactone) methyl ether) for evaluating in vitro Vero and MC3T3-E1 pre-osteoblasts cell response. Characterization and evaluation of the coated substrates were carried out using different techniques. The Fourier transform infrared spectroscopy data demonstrated that the main functional groups in the MAPLE-deposited films remained intact. Atomic Force Microscopy images showed the coatings to be continuous, with the surface roughness depending on the deposition parameters. Moreover, the behaviour of the coatings in medium mimicking the pH and temperature of the human body was studied and corelated to degradation. Spectro-ellipsometry (SE) and AFM measurements revealed the degradation trend during immersion time by the changes in coating thickness and roughness. In vitro biocompatibility was studied by indirect contact tests on Vero cells in accordance with ISO 10993-5/2009. The results obtained in terms of cell morphology (phase contrast microscopy) and cytotoxicity (LDH and MTT assays) proved biocompatibility. Furthermore, direct contact assays on MC3T3-E1 pre-osteoblasts demonstrated the capacity of all analyzed specimens to support cell adhesion, normal cellular morphology and growth.

  9. Cell Proliferation and Cytotoxicity Assays.

    PubMed

    Adan, Aysun; Kiraz, Yağmur; Baran, Yusuf

    Cell viability is defined as the number of healthy cells in a sample and proliferation of cells is a vital indicator for understanding the mechanisms in action of certain genes, proteins and pathways involved cell survival or death after exposing to toxic agents. Generally, methods used to determine viability are also common for the detection of cell proliferation. Cell cytotoxicity and proliferation assays are generally used for drug screening to detect whether the test molecules have effects on cell proliferation or display direct cytotoxic effects. Regardless of the type of cell-based assay being used, it is important to know how many viable cells are remaining at the end of the experiment. There are a variety of assay methods based on various cell functions such as enzyme activity, cell membrane permeability, cell adherence, ATP production, co-enzyme production, and nucleotide uptake activity. These methods could be basically classified into different categories: (I) dye exclusion methods such as trypan blue dye exclusion assay, (II) methods based on metabolic activity, (III) ATP assay, (IV) sulforhodamine B assay, (V) protease viability marker assay, (VI) clonogenic cell survival assay, (VII) DNA synthesis cell proliferation assays and (V) raman micro-spectroscopy. In order to choose the optimal viability assay, the cell type, applied culture conditions, and the specific questions being asked should be considered in detail. This particular review aims to provide an overview of common cell proliferation and cytotoxicity assays together with their own advantages and disadvantages, their methodologies, comparisons and intended purposes.

  10. Cell Proliferation, Cell Death, and Size Regulation

    DTIC Science & Technology

    1998-10-01

    Cell Death , and Size Regulation PRINCIPAL INVESTIGATOR: Nicholas E. Baker, Ph.D. CONTRACTING ORGANIZATION: Albert Einstein College of Medicine of Yeshiva...SUBTITLE 5. FUNDING NUMBERS Cell Proliferation, Cell Death , and Size Regulation DAMD17-97-1-7034 6. AUTHOR(S) Nicholas E. Baker, Ph.D. 7. PERFORMING...Contains unpublished data 5 CELL PROLIFERATION, CELL DEATH , AND SIZE REGULATION INTRODUCTION Cell proliferation and cell death come to attention through

  11. Lensless imaging system to quantify cell proliferation

    NASA Astrophysics Data System (ADS)

    Vinjimore Kesavan, S.; Allier, C. P.; Navarro, F.; Mittler, F.; Chalmond, B.; Dinten, J.-M.

    2013-02-01

    Owing to its simplicity, lensless imaging system is adept at continuous monitoring of adherent cells inside the incubator. The setup consists of a CMOS sensor with pixel pitch of 2.2 μm and field of view of 24 mm2, LED with a dominating wavelength of 525 nm, along with a pinhole of 150 μm as the source of illumination. The in-line hologram obtained from cells depends on the degree of cell-substrate adhesion. Drastic difference is observed between the holographic patterns of floating and adherent cells. In addition, the well-established fact of reduction of cell-substrate contact during cell division is observed with our system based on corresponding spontaneous transition in the holographic pattern. Here, we demonstrate that by recognizing this specific holographic pattern, number of cells undergoing mitosis in a cell culture with a population of approximately 5000 cells, can be estimated in real-time. The method is assessed on comparison with Edu-based proliferation assay. The approach is straightforward and it eliminates the use of markers to estimate the proliferation rate of a given cell culture. Unlike most proliferation assays, the cells are not harvested enabling continuous monitoring of cell culture.

  12. Focal Adhesion Kinase Modulates Cell Adhesion Strengthening via Integrin Activation

    PubMed Central

    Michael, Kristin E.; Dumbauld, David W.; Burns, Kellie L.; Hanks, Steven K.

    2009-01-01

    Focal adhesion kinase (FAK) is an essential nonreceptor tyrosine kinase regulating cell migration, adhesive signaling, and mechanosensing. Using FAK-null cells expressing FAK under an inducible promoter, we demonstrate that FAK regulates the time-dependent generation of adhesive forces. During the early stages of adhesion, FAK expression in FAK-null cells enhances integrin activation to promote integrin binding and, hence, the adhesion strengthening rate. Importantly, FAK expression regulated integrin activation, and talin was required for the FAK-dependent effects. A role for FAK in integrin activation was confirmed in human fibroblasts with knocked-down FAK expression. The FAK autophosphorylation Y397 site was required for the enhancements in adhesion strengthening and integrin-binding responses. This work demonstrates a novel role for FAK in integrin activation and the time-dependent generation of cell–ECM forces. PMID:19297531

  13. Cell-Cell Adhesion and Breast Cancer.

    DTIC Science & Technology

    1998-01-01

    Staging of breast cancer. In: K.I. Bland and E.M. Copeland (eds.), The breast: Comprehensive management of benign and malignant diseases , pp. 313-330... desmosomes . The physical strength of adhesion between two cells is likely to be dependent upon a number of factors, including the number of adhesion

  14. Cellular Adhesion Promotes Prostate Cancer Cells Escape from Dormancy

    PubMed Central

    Ruppender, Nazanin; Larson, Sandy; Lakely, Bryce; Kollath, Lori; Brown, Lisha; Coleman, Ilsa; Coleman, Roger; Nguyen, Holly; Nelson, Peter S.; Corey, Eva; Snyder, Linda A.; Vessella, Robert L.; Morrissey, Colm; Lam, Hung-Ming

    2015-01-01

    Dissemination of prostate cancer (PCa) cells to the bone marrow is an early event in the disease process. In some patients, disseminated tumor cells (DTC) proliferate to form active metastases after a prolonged period of undetectable disease known as tumor dormancy. Identifying mechanisms of PCa dormancy and reactivation remain a challenge partly due to the lack of in vitro models. Here, we characterized in vitro PCa dormancy-reactivation by inducing cells from three patient-derived xenograft (PDX) lines to proliferate through tumor cell contact with each other and with bone marrow stroma. Proliferating PCa cells demonstrated tumor cell-cell contact and integrin clustering by immunofluorescence. Global gene expression analyses on proliferating cells cultured on bone marrow stroma revealed a downregulation of TGFB2 in all of the three proliferating PCa PDX lines when compared to their non-proliferating counterparts. Furthermore, constitutive activation of myosin light chain kinase (MLCK), a downstream effector of integrin-beta1 and TGF-beta2, in non-proliferating cells promoted cell proliferation. This cell proliferation was associated with an upregulation of CDK6 and a downregulation of E2F4. Taken together, our data provide the first clinically relevant in vitro model to support cellular adhesion and downregulation of TGFB2 as a potential mechanism by which PCa cells may escape from dormancy. Targeting the TGF-beta2-associated mechanism could provide novel opportunities to prevent lethal PCa metastasis. PMID:26090669

  15. Cell adhesion to cathodic arc plasma deposited CrAlSiN thin films

    NASA Astrophysics Data System (ADS)

    Kim, Sun Kyu; Pham, Vuong-Hung; Kim, Chong-Hyun

    2012-07-01

    Osteoblast cell response (cell adhesion, actin cytoskeleton and focal contact adhesion as well as cell proliferation) to CrN, CrAlSiN and Ti thin films was evaluated in vitro. Cell adhesion and actin stress fibers organization depended on the film composition significantly. Immunofluorescent staining of vinculin in osteoblast cells showed good focal contact adhesion on the CrAlSiN and Ti thin films but not on the CrN thin films. Cell proliferation was significantly greater on the CrAlSiN thin films as well as on Ti thin films than on the CrN thin films.

  16. Tuning cell adhesion by direct nanostructuring silicon into cell repulsive/adhesive patterns.

    PubMed

    Premnath, Priyatha; Tavangar, Amirhossein; Tan, Bo; Venkatakrishnan, Krishnan

    2015-09-10

    Developing platforms that allow tuning cell functionality through incorporating physical, chemical, or mechanical cues onto the material surfaces is one of the key challenges in research in the field of biomaterials. In this respect, various approaches have been proposed and numerous structures have been developed on a variety of materials. Most of these approaches, however, demand a multistep process or post-chemical treatment. Therefore, a simple approach would be desirable to develop bio-functionalized platforms for effectively modulating cell adhesion and consequently programming cell functionality without requiring any chemical or biological surface treatment. This study introduces a versatile yet simple laser approach to structure silicon (Si) chips into cytophobic/cytophilic patterns in order to modulate cell adhesion and proliferation. These patterns are fabricated on platforms through direct laser processing of Si substrates, which renders a desired computer-generated configuration into patterns. We investigate the morphology, chemistry, and wettability of the platform surfaces. Subsequently, we study the functionality of the fabricated platforms on modulating cervical cancer cells (HeLa) behaviour. The results from in vitro studies suggest that the nanostructures efficiently repel HeLa cells and drive them to migrate onto untreated sites. The study of the morphology of the cells reveals that cells evade the cytophobic area by bending and changing direction. Additionally, cell patterning, cell directionality, cell channelling, and cell trapping are achieved by developing different platforms with specific patterns. The flexibility and controllability of this approach to effectively structure Si substrates to cell-repulsive and cell-adhesive patterns offer perceptible outlook for developing bio-functionalized platforms for a variety of biomedical devices. Moreover, this approach could pave the way for developing anti-cancer platforms that selectively repel

  17. Cucurbitacin B purified from Ecballium elaterium (L.) A. Rich from Tunisia inhibits α5β1 integrin-mediated adhesion, migration, proliferation of human glioblastoma cell line and angiogenesis.

    PubMed

    Touihri-Barakati, Imen; Kallech-Ziri, Olfa; Ayadi, Wiem; Kovacic, Hervé; Hanchi, Belgacem; Hosni, Karim; Luis, José

    2017-02-15

    Integrins are essential protagonists in the complex multistep process of cancer progression and are thus attractive targets for the development of anticancer agents. Cucurbitacin B, a triterpenoid purified from the leaves of Tunisian Ecballium elaterium exhibited an anticancer effect and displayed anti-integrin activity on human glioblastoma U87 cells, without being cytotoxic at concentrations up to 500nM. Here we show that cucurbitacin B affected the adhesion and migration of U87 cells to fibronectin in a dose-dependent manner with IC50 values of 86.2nM and 84.6nM, respectively. Time-lapse videomicroscopy showed that cucurbitacin B significantly reduced U87 cells motility and affected directional persistence. Cucurbitacin B also inhibited proliferation with IC50 value of 70.1nM using Crystal Violet assay. Moreover, cucurbitacin B efficiently inhibited in vitro human microvascular endothelial cells (HMEC) angiogenesis with concentration up to 10nM. Interestingly, we demonstrate for the first time that this effect was specifically mediated by α5β1 integrins. These findings reveal a novel mechanism of action for cucurbitacin B, which displays a potential interest as a specific anti-integrin drug.

  18. On the relation between surface roughness of metallic substrates and adhesion of human primary bone cells.

    PubMed

    Anselme, K; Bigerelle, M

    2014-01-01

    Surface characteristics of materials, whether their topography, chemistry, or surface energy, play an essential part in osteoblast adhesion on biomaterials. Thus, the quality of cell adhesion will influence the cell's capacity to proliferate and differentiate in contact with a biomaterial. We have developed for more than ten years numerous studies on the influence of topography and chemistry of metallic substrates on the response of primary human bone cells. The originality of our approach is that contrary to most of other authors, we quantified the adhesion of primary human bone cells on metallic substrates with perfectly characterized surface topography after some hours but also over 21 days. Moreover, we have developed original statistical approaches for characterizing the relation between surface roughness and cell-adhesion parameters. In this article, we will illustrate different studies we did these last ten years concerning the development of a new adhesion parameter, the adhesion power; the correlation between short-term adhesion, long-term adhesion, and proliferation; the influence of roughness organization on cell adhesion and the development of the order parameter; our modeling approach of cell adhesion on surface topography; the relative influence of surface chemistry and topography on cell adhesion and contact angle; the relation between surface features dimensions and cell adhesion. Further, some considerations will be given on the methods for scanning surface topography for cell-adhesion studies. Finally, perspectives will be given to elucidate these intracellular mechanotransduction mechanisms induced by the deformation of cells on model sinusoidal peaks-or-valleys surfaces.

  19. Cell-Substrate Adhesion by Amoeboid Cells

    NASA Astrophysics Data System (ADS)

    Flanders, Bret; Panta, Krishna

    Amoeboid migration is a rapid (10 μm min-1) mode of migration that some tumor cells exhibit. To permit such rapid movement, the adhesive contacts between the cell and the substrate must be relatively short-lived and weak. In this study, we investigate the basic adhesive character of amoeboid cells (D. discoideum) in contact with silanized glass substrates. We observe the initiation and spreading of the adhesive contacts that these cells establish as they settle under gravity onto the substrate and relax towards mechanical equilibrium. The use of interference reflection microscopy and cellular tethering measurements have allowed us to determine the basic adhesive properties of the cell: the membrane-medium interfacial energy; the bending modulus; the equilibrium contact angle; and the work of adhesion. We find the time scale on which settling occurs to be longer than expected. Implications of these results on adhesion and migration will be discussed. The authors are grateful for support from NSF (CBET-1451903) and NIH (1R21EY026392).

  20. Microfluidic devices for cell cultivation and proliferation

    PubMed Central

    Tehranirokh, Masoomeh; Kouzani, Abbas Z.; Francis, Paul S.; Kanwar, Jagat R.

    2013-01-01

    Microfluidic technology provides precise, controlled-environment, cost-effective, compact, integrated, and high-throughput microsystems that are promising substitutes for conventional biological laboratory methods. In recent years, microfluidic cell culture devices have been used for applications such as tissue engineering, diagnostics, drug screening, immunology, cancer studies, stem cell proliferation and differentiation, and neurite guidance. Microfluidic technology allows dynamic cell culture in microperfusion systems to deliver continuous nutrient supplies for long term cell culture. It offers many opportunities to mimic the cell-cell and cell-extracellular matrix interactions of tissues by creating gradient concentrations of biochemical signals such as growth factors, chemokines, and hormones. Other applications of cell cultivation in microfluidic systems include high resolution cell patterning on a modified substrate with adhesive patterns and the reconstruction of complicated tissue architectures. In this review, recent advances in microfluidic platforms for cell culturing and proliferation, for both simple monolayer (2D) cell seeding processes and 3D configurations as accurate models of in vivo conditions, are examined. PMID:24273628

  1. Tuning cell adhesion by direct nanostructuring silicon into cell repulsive/adhesive patterns

    SciTech Connect

    Premnath, Priyatha; Venkatakrishnan, Krishnan

    2015-09-10

    Developing platforms that allow tuning cell functionality through incorporating physical, chemical, or mechanical cues onto the material surfaces is one of the key challenges in research in the field of biomaterials. In this respect, various approaches have been proposed and numerous structures have been developed on a variety of materials. Most of these approaches, however, demand a multistep process or post-chemical treatment. Therefore, a simple approach would be desirable to develop bio-functionalized platforms for effectively modulating cell adhesion and consequently programming cell functionality without requiring any chemical or biological surface treatment. This study introduces a versatile yet simple laser approach to structure silicon (Si) chips into cytophobic/cytophilic patterns in order to modulate cell adhesion and proliferation. These patterns are fabricated on platforms through direct laser processing of Si substrates, which renders a desired computer-generated configuration into patterns. We investigate the morphology, chemistry, and wettability of the platform surfaces. Subsequently, we study the functionality of the fabricated platforms on modulating cervical cancer cells (HeLa) behaviour. The results from in vitro studies suggest that the nanostructures efficiently repel HeLa cells and drive them to migrate onto untreated sites. The study of the morphology of the cells reveals that cells evade the cytophobic area by bending and changing direction. Additionally, cell patterning, cell directionality, cell channelling, and cell trapping are achieved by developing different platforms with specific patterns. The flexibility and controllability of this approach to effectively structure Si substrates to cell-repulsive and cell-adhesive patterns offer perceptible outlook for developing bio-functionalized platforms for a variety of biomedical devices. Moreover, this approach could pave the way for developing anti-cancer platforms that selectively repel

  2. High-Frequency Mechanostimulation of Cell Adhesion.

    PubMed

    Kadem, Laith F; Suana, K Grace; Holz, Michelle; Wang, Wei; Westerhaus, Hannes; Herges, Rainer; Selhuber-Unkel, Christine

    2017-01-02

    Cell adhesion is regulated by molecularly defined protein interactions and by mechanical forces, which can activate a dynamic restructuring of adhesion sites. Previous attempts to explore the response of cell adhesion to forces have been limited to applying mechanical stimuli that involve the cytoskeleton. In contrast, we here apply a new, oscillatory type of stimulus through push-pull azobenzenes. Push-pull azobenzenes perform a high-frequency, molecular oscillation upon irradiation with visible light that has frequently been applied in polymer surface relief grating. We here use these oscillations to address single adhesion receptors. The effect of molecular oscillatory forces on cell adhesion has been analyzed using single-cell force spectroscopy and gene expression studies. Our experiments demonstrate a reinforcement of cell adhesion as well as upregulated expression levels of adhesion-associated genes as a result of the nanoscale "tickling" of integrins. This novel type of mechanical stimulus provides a previously unprecedented molecular control of cellular mechanosensing.

  3. Negative regulators of cell proliferation

    NASA Technical Reports Server (NTRS)

    Johnson, T. C.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Cell proliferation is governed by the influence of both mitogens and inhibitors. Although cell contact has long been thought to play a fundamental role in cell cycling regulation, and negative regulators have long been suspected to exist, their isolation and purification has been complicated by a variety of technical difficulties. Nevertheless, over recent years an ever-expanding list of putative negative regulators have emerged. In many cases, their biological inhibitory activities are consistent with density-dependent growth inhibition. Most likely their interactions with mitogenic agents, at an intracellular level, are responsible for either mitotic arrest or continued cell cycling. A review of naturally occurring cell growth inhibitors is presented with an emphasis on those factors shown to be residents of the cell surface membrane. Particular attention is focused on a cell surface sialoglycopeptide, isolated from intact bovine cerebral cortex cells, which has been shown to inhibit the proliferation of an unusually wide range of target cells. The glycopeptide arrest cells obtained from diverse species, both fibroblasts and epithelial cells, and a broad variety of transformed cells. Signal transduction events and a limited spectrum of cells that are refractory to the sialoglycopeptide have provided insight into the molecular events mediated by this cell surface inhibitor.

  4. Plasma polymerization for cell adhesive/anti-adhesive implant coating

    NASA Astrophysics Data System (ADS)

    Meichsner, Juergen; Testrich, Holger; Rebl, Henrike; Nebe, Barbara

    2015-09-01

    Plasma polymerization of ethylenediamine (C2H8N2, EDA) and perfluoropropane (C3F8, PFP) with admixture of argon and hydrogen, respectively, was studied using an asymmetric 13.56 MHz CCP. The analysis of the plasma chemical gas phase processes for stable molecules revealed consecutive reactions: C2H8N2 consumption, intermediate product NH3, and main final product HCN. In C3F8- H2 plasma the precursor molecule C3F8 and molecular hydrogen are consumed and HF as well as CF4 and C2F6 are found as main gaseous reaction products. The deposited plasma polymer films on the powered electrode are strongly cross-linked due to ion bombardment. The stable plasma polymerized films from EDA are characterized by high content of nitrogen with N/C ratio of about 0.35. The plasma polymerized fluorocarbon film exhibit a reduced F/C ratio of about 1.2. Adhesion tests with human osteoblast cell line MG-63 on coated Ti6Al4V samples (polished) compared with uncoated reference sample yielded both, the enhanced cell adhesion for plasma polymerized EDA and significantly reduced cell adhesion for fluorocarbon coating, respectively. Aging of the plasma polymerized EDA film, in particular due to the reactions with oxygen from air, showed no significant change in the cell adhesion. The fluorocarbon coating with low cell adhesion is of interest for temporary implants. Funded by the Campus PlasmaMed.

  5. VUV modification promotes endothelial cell proliferation on PTFE vascular grafts

    NASA Astrophysics Data System (ADS)

    Cezeaux, J. L.; Romoser, C. E.; Benson, R. S.; Buck, C. K.; Sackman, J. E.

    1998-05-01

    Small diameter (⩽6 mm ID ) synthetic vascular grafts, used as lower-limb vessel replacements in patients without suitable autologous saphenous veins, have a failure rate of 53% after 4 yr. Graft failure is due to thrombosis and intimal hyperplasia, an increase in smooth muscle cells in the lumen of the vessel which leads to progressive closing and ultimate occlusion of the vessel. In an effort to increase patency rates of synthetic grafts, investigators have seeded vascular grafts with endothelial cells prior to implantation in an attempt to control both thrombosis and smooth muscle proliferation. This technique has been successful for the development of an endothelial monolayer in animal trials, but has met with limited success in humans. The hydrophobicity, low surface energy, and weak electrical charge of expanded polytetrafluoroethylene (ePTFE) provides conditions which are not optimal for endothelial cell attachment. The purpose of this study is to evaluate the effect of vacuum ultraviolet (VUV) modification of ePTFE on endothelial cell adhesion and proliferation. Pieces of ePTFE graft material were exposed to 10, 20 or 40 W VUV radiation for 10, 20 or 40 min using a UV excimer lamp. Prior to cell adhesion and proliferation experiments, the grafts pieces were autoclaved and cut into pledgets. Half of the pledgets were precoated with fibronectin ( 20 μg/ml). Cell adhesion was measured by seeding 3H-thymidine labeled human umbilical vein endothelial cells (HUVEC) onto the pledgets for 60 min. The pledgets were then washed and the remaining radioactivity assayed using scintillation counting. For the cell proliferation experiments, pledgets were seeded with unlabeled HUVEC which were allowed to adhere to the graft material for 18 h. The cells were then exposed to 3H-thymidine ( 1 μCi/ml) for approximately 48 h and then washed to remove any unincorporated 3H-thymidine. Incorporation of 3H-thymidine was measured using scintillation counting. Four replicate

  6. Synaptic Cell Adhesion Molecules in Alzheimer's Disease

    PubMed Central

    Leshchyns'ka, Iryna

    2016-01-01

    Alzheimer's disease (AD) is a neurodegenerative brain disorder associated with the loss of synapses between neurons in the brain. Synaptic cell adhesion molecules are cell surface glycoproteins which are expressed at the synaptic plasma membranes of neurons. These proteins play key roles in formation and maintenance of synapses and regulation of synaptic plasticity. Genetic studies and biochemical analysis of the human brain tissue, cerebrospinal fluid, and sera from AD patients indicate that levels and function of synaptic cell adhesion molecules are affected in AD. Synaptic cell adhesion molecules interact with Aβ, a peptide accumulating in AD brains, which affects their expression and synaptic localization. Synaptic cell adhesion molecules also regulate the production of Aβ via interaction with the key enzymes involved in Aβ formation. Aβ-dependent changes in synaptic adhesion affect the function and integrity of synapses suggesting that alterations in synaptic adhesion play key roles in the disruption of neuronal networks in AD. PMID:27242933

  7. Cleavage and Cell Adhesion Properties of Human Epithelial Cell Adhesion Molecule (HEPCAM)*

    PubMed Central

    Tsaktanis, Thanos; Kremling, Heidi; Pavšič, Miha; von Stackelberg, Ricarda; Mack, Brigitte; Fukumori, Akio; Steiner, Harald; Vielmuth, Franziska; Spindler, Volker; Huang, Zhe; Jakubowski, Jasmine; Stoecklein, Nikolas H.; Luxenburger, Elke; Lauber, Kirsten; Lenarčič, Brigita; Gires, Olivier

    2015-01-01

    Human epithelial cell adhesion molecule (HEPCAM) is a tumor-associated antigen frequently expressed in carcinomas, which promotes proliferation after regulated intramembrane proteolysis. Here, we describe extracellular shedding of HEPCAM at two α-sites through a disintegrin and metalloprotease (ADAM) and at one β-site through BACE1. Transmembrane cleavage by γ-secretase occurs at three γ-sites to generate extracellular Aβ-like fragments and at two ϵ-sites to release human EPCAM intracellular domain HEPICD, which is efficiently degraded by the proteasome. Mapping of cleavage sites onto three-dimensional structures of HEPEX cis-dimer predicted conditional availability of α- and β-sites. Endocytosis of HEPCAM warrants acidification in cytoplasmic vesicles to dissociate protein cis-dimers required for cleavage by BACE1 at low pH values. Intramembrane cleavage sites are accessible and not part of the structurally important transmembrane helix dimer crossing region. Surprisingly, neither chemical inhibition of cleavage nor cellular knock-out of HEPCAM using CRISPR-Cas9 technology impacted the adhesion of carcinoma cell lines. Hence, a direct function of HEPCAM as an adhesion molecule in carcinoma cells is not supported and appears to be questionable. PMID:26292218

  8. Focal Adhesion-Independent Cell Migration.

    PubMed

    Paluch, Ewa K; Aspalter, Irene M; Sixt, Michael

    2016-10-06

    Cell migration is central to a multitude of physiological processes, including embryonic development, immune surveillance, and wound healing, and deregulated migration is key to cancer dissemination. Decades of investigations have uncovered many of the molecular and physical mechanisms underlying cell migration. Together with protrusion extension and cell body retraction, adhesion to the substrate via specific focal adhesion points has long been considered an essential step in cell migration. Although this is true for cells moving on two-dimensional substrates, recent studies have demonstrated that focal adhesions are not required for cells moving in three dimensions, in which confinement is sufficient to maintain a cell in contact with its substrate. Here, we review the investigations that have led to challenging the requirement of specific adhesions for migration, discuss the physical mechanisms proposed for cell body translocation during focal adhesion-independent migration, and highlight the remaining open questions for the future.

  9. Platelets: cell proliferation and atherosclerosis.

    PubMed

    Ross, R

    1979-04-01

    Intimal smooth muscle proliferation is the hallmark of the lesions of atherosclerosis. Endothelial injury is postulated to precede this intimal smooth muscle proliferative response, which is mediated by a potent mitogenic factor derived from adherence, aggregation, and release by platelets at sites of endothelial injury. Smooth muscle proliferation is accompanied by varying amounts of connective tissue formation and intracellular and extracellular lipid deposition, dependent upon the risk factors encountered in each patient. The platelet-derived mitogen (PF) is a stable, cationic, relatively low molecular weight (10,000-30,000) protein that has been partially purified by ion exchange chromotography and gel filtration. Less than 100 ng of PF/ml culture medium can stimulate sparse 3T3 cells or smooth muscle cells, but not endothelial cells, to undergo multiple cell divisions in the presence of 5% cell-free, plasma-derived serum. The latter contains no mitogenic activity. The interaction of the platelet mitogen and plasma-derived components, including lipoproteins, plays a critical role in smooth muscle proliferation in vitro and in vivo in the induction of the lesions of atherosclerosis.

  10. The right motifs for plant cell adhesion: what makes an adhesive site?

    PubMed

    Langhans, Markus; Weber, Wadim; Babel, Laura; Grunewald, Miriam; Meckel, Tobias

    2017-01-01

    Cells of multicellular organisms are surrounded by and attached to a matrix of fibrous polysaccharides and proteins known as the extracellular matrix. This fibrous network not only serves as a structural support to cells and tissues but also plays an integral part in the process as important as proliferation, differentiation, or defense. While at first sight, the extracellular matrices of plant and animals do not have much in common, a closer look reveals remarkable similarities. In particular, the proteins involved in the adhesion of the cell to the extracellular matrix share many functional properties. At the sequence level, however, a surprising lack of homology is found between adhesion-related proteins of plants and animals. Both protein machineries only reveal similarities between small subdomains and motifs, which further underlines their functional relationship. In this review, we provide an overview on the similarities between motifs in proteins known to be located at the plant cell wall-plasma membrane-cytoskeleton interface to proteins of the animal adhesome. We also show that by comparing the proteome of both adhesion machineries at the level of motifs, we are also able to identify potentially new candidate proteins that functionally contribute to the adhesion of the plant plasma membrane to the cell wall.

  11. Polyelectrolytes Multilayers to Modulate Cell Adhesion: A Study of the Influence of Film Composition and Polyelectrolyte Interdigitation on the Adhesion of the A549 Cell Line.

    PubMed

    Muzzio, Nicolás E; Pasquale, Miguel A; Gregurec, Danijela; Diamanti, Eleftheria; Kosutic, Marija; Azzaroni, Omar; Moya, Sergio E

    2016-04-01

    Polyelectrolyte multilayers (PEMs) with different polycation/polyanion pairs are fabricated by the layer-by-layer technique employing synthetic, natural, and both types of polyelectrolytes. The impact of the chemical composition of PEMs on cell adhesion is assessed by studying cell shape, spreading area, focal contacts, and cell proliferation for the A549 cell line. Cells exhibit good adhesion on PEMs containing natural polycations and poly(sodium 4-styrenesulfonate) (PSS) as polyanion, but limited adhesion is observed on PEMs fabricated from both natural polyelectrolytes. PEMs are then assembled, depositing a block of natural polyelectrolytes on top of a stiffer block with PSS as polyanion. Cell adhesion is enhanced on top of the diblock PEMs compared to purely natural PEMs. This fact could be explained by the interdigitation between polyelectrolytes from the two blocks. Diblock PEM assembly provides a simple means to tune cell adhesion on biocompatible PEMs.

  12. Cell adhesion to plasma-coated PVC.

    PubMed

    Rangel, Elidiane C; de Souza, Eduardo S; de Moraes, Francine S; Duek, Eliana A R; Lucchesi, Carolina; Schreiner, Wido H; Durrant, Steven F; Cruz, Nilson C

    2014-01-01

    To produce environments suitable for cell culture, thin polymer films were deposited onto commercial PVC plates from radiofrequency acetylene-argon plasmas. The proportion of argon in the plasmas, P(Ar), was varied from 5.3 to 65.8%. The adhesion and growth of Vero cells on the coated surfaces were examined for different incubation times. Cytotoxicity tests were performed using spectroscopic methods. Carbon, O, and N were detected in all the samples using XPS. Roughness remained almost unchanged in the samples prepared with 5.3 and 28.9% but tended to increase for the films deposited with P(Ar) between 28.9 and 55.3%. Surface free energy increased with increasing P(Ar), except for the sample prepared at 28.9% of Ar, which presented the least reactive surface. Cells proliferated on all the samples, including the bare PVC. Independently of the deposition condition there was no evidence of cytotoxicity, indicating the viability of such coatings for designing biocompatible devices.

  13. Cell Adhesion to Plasma-Coated PVC

    PubMed Central

    Rangel, Elidiane C.; de Souza, Eduardo S.; de Moraes, Francine S.; Duek, Eliana A. R.; Lucchesi, Carolina; Schreiner, Wido H.; Durrant, Steven F.; Cruz, Nilson C.

    2014-01-01

    To produce environments suitable for cell culture, thin polymer films were deposited onto commercial PVC plates from radiofrequency acetylene-argon plasmas. The proportion of argon in the plasmas, PAr, was varied from 5.3 to 65.8%. The adhesion and growth of Vero cells on the coated surfaces were examined for different incubation times. Cytotoxicity tests were performed using spectroscopic methods. Carbon, O, and N were detected in all the samples using XPS. Roughness remained almost unchanged in the samples prepared with 5.3 and 28.9% but tended to increase for the films deposited with PAr between 28.9 and 55.3%. Surface free energy increased with increasing PAr, except for the sample prepared at 28.9% of Ar, which presented the least reactive surface. Cells proliferated on all the samples, including the bare PVC. Independently of the deposition condition there was no evidence of cytotoxicity, indicating the viability of such coatings for designing biocompatible devices. PMID:25247202

  14. Biomimetic Hybrid Nanofiber Sheets Composed of RGD Peptide-Decorated PLGA as Cell-Adhesive Substrates.

    PubMed

    Shin, Yong Cheol; Lee, Jong Ho; Kim, Min Jeong; Park, Ji Hoon; Kim, Sung Eun; Kim, Jin Su; Oh, Jin-Woo; Han, Dong-Wook

    2015-05-29

    In biomedical applications, there is a need for tissue engineering scaffolds to promote and control cellular behaviors, including adhesion, proliferation and differentiation. In particular, the initial adhesion of cells has a great influence on those cellular behaviors. In this study, we concentrate on developing cell-adhesive substrates applicable for tissue engineering scaffolds. The hybrid nanofiber sheets were prepared by electrospinning poly(lactic-co-glycolic acid) (PLGA) and M13 phage, which was genetically modified to enhance cell adhesion thru expressing RGD peptides on their surface. The RGD peptide is a specific motif of extracellular matrix (ECM) for integrin receptors of cells. RGD peptide-decorated PLGA (RGD-PLGA) nanofiber sheets were characterized by scanning electron microscopy, immunofluorescence staining, contact angle measurement and differential scanning calorimetry. In addition, the initial adhesion and proliferation of four different types of mammalian cells were determined in order to evaluate the potential of RGD-PLGA nanofiber sheets as cell-adhesive substrates. Our results showed that the hybrid nanofiber sheets have a three-dimensional porous structure comparable to the native ECM. Furthermore, the initial adhesion and proliferation of cells were significantly enhanced on RGD-PLGA sheets. These results suggest that biomimetic RGD-PLGA nanofiber sheets can be promising cell-adhesive substrates for application as tissue engineering scaffolds.

  15. Cell proliferation in normal epidermis

    SciTech Connect

    Weinstein, G.D.; McCullough, J.L.; Ross, P.

    1984-06-01

    A detailed examination of cell proliferation kinetics in normal human epidermis is presented. Using tritiated thymidine with autoradiographic techniques, proliferative and differentiated cell kinetics are defined and interrelated. The proliferative compartment of normal epidermis has a cell cycle duration (Tc) of 311 h derived from 3 components: the germinative labeling index (LI), the duration of DNA synthesis (ts), and the growth fraction (GF). The germinative LI is 2.7% +/- 1.2 and ts is 14 h, the latter obtained from a composite fraction of labeled mitoses curve obtained from 11 normal subjects. The GF obtained from the literature and from human skin xenografts to nude mice is estimated to be 60%. Normal-appearing epidermis from patients with psoriasis appears to have a higher proliferation rate. The mean LI is 4.2% +/- 0.9, approximately 50% greater than in normal epidermis. Absolute cell kinetic values for this tissue, however, cannot yet be calculated for lack of other information on ts and GF. A kinetic model for epidermal cell renewal in normal epidermis is described that interrelates the rate of birth/entry, transit, and/or loss of keratinocytes in the 3 epidermal compartments: proliferative, viable differentiated (stratum malpighii), and stratum corneum. Expected kinetic homeostasis in the epidermis is confirmed by the very similar ''turnover'' rates in each of the compartments that are, respectively, 1246, 1417, and 1490 cells/day/mm2 surface area. The mean epidermal turnover time of the entire tissue is 39 days. The Tc of 311 h in normal cells in 8-fold longer than the psoriatic Tc of 36 h and is necessary for understanding the hyperproliferative pathophysiologic process in psoriasis.

  16. Contractility Modulates Cell Adhesion Strengthening Through Focal Adhesion Kinase and Assembly of Vinculin-Containing Focal Adhesions

    PubMed Central

    Dumbauld, David W.; Shin, Heungsoo; Gallant, Nathan D.; Michael, Kristin E.; Radhakrishna, Harish; García, Andrés J.

    2010-01-01

    Actin-myosin contractility modulates focal adhesion assembly, stress fiber formation, and cell migration. We analyzed the contributions of contractility to fibroblast adhesion strengthening using a hydrodynamic adhesion assay and micropatterned substrates to control cell shape and adhesive area. Serum addition resulted in adhesion strengthening to levels 30–40% higher than serum-free cultures. Inhibition of myosin light chain kinase or Rho-kinase blocked phosphorylation of myosin light chain to similar extents and eliminated the serum-induced enhancements in strengthening. Blebbistatin-induced inhibition of myosin II reduced serum-induced adhesion strength to similar levels as those obtained by blocking myosin light chain phosphorylation. Reductions in adhesion strengthening by inhibitors of contractility correlated with loss of vinculin and talin from focal adhesions without changes in integrin binding. In vinculin-null cells, inhibition of contractility did not alter adhesive force, whereas controls displayed a 20% reduction in adhesion strength, indicating that the effects of contractility on adhesive force are vinculin-dependent. Furthermore, in cells expressing FAK, inhibitors of contractility reduced serum-induced adhesion strengthening as well as eliminated focal adhesion assembly. In contrast, in the absence of FAK, these inhibitors did not alter adhesion strength or focal adhesion assembly. These results indicate that contractility modulates adhesion strengthening via FAK-dependent, vinculin-containing focal adhesion assembly. PMID:20205236

  17. Hybrid inverse opals for regulating cell adhesion and orientation

    NASA Astrophysics Data System (ADS)

    Lu, Jie; Zheng, Fuyin; Cheng, Yao; Ding, Haibo; Zhao, Yuanjin; Gu, Zhongze

    2014-08-01

    Cell adhesion and alignment are two important considerations in tissue engineering applications as they can regulate the subsequent cell proliferation activity and differentiation program. Although many effects have been applied to regulate the adhesion or alignment of cells by using physical and chemical methods, it is still a challenge to regulate these cell behaviors simultaneously. Here, we present novel substrates with tunable nanoscale patterned structures for regulating the adhesion and alignment of cells. The substrates with different degrees of pattern orientation were achieved by customizing the amount of stretching applied to polymer inverse opal films. Cells cultured on these substrates showed an adjustable morphology and alignment. Moreover, soft hydrogels, which have poor plasticity and are difficult to cast into patterned structures, were applied to infiltrate the inverse opal structure. We demonstrated that the adhesion ratio of cells could be regulated by these hybrid substrates, as well as adjusting the cell morphology and alignment. These features of functional inverse opal substrates make them suitable for important applications in tissue engineering.

  18. Hybrid inverse opals for regulating cell adhesion and orientation.

    PubMed

    Lu, Jie; Zheng, Fuyin; Cheng, Yao; Ding, Haibo; Zhao, Yuanjin; Gu, Zhongze

    2014-09-21

    Cell adhesion and alignment are two important considerations in tissue engineering applications as they can regulate the subsequent cell proliferation activity and differentiation program. Although many effects have been applied to regulate the adhesion or alignment of cells by using physical and chemical methods, it is still a challenge to regulate these cell behaviors simultaneously. Here, we present novel substrates with tunable nanoscale patterned structures for regulating the adhesion and alignment of cells. The substrates with different degrees of pattern orientation were achieved by customizing the amount of stretching applied to polymer inverse opal films. Cells cultured on these substrates showed an adjustable morphology and alignment. Moreover, soft hydrogels, which have poor plasticity and are difficult to cast into patterned structures, were applied to infiltrate the inverse opal structure. We demonstrated that the adhesion ratio of cells could be regulated by these hybrid substrates, as well as adjusting the cell morphology and alignment. These features of functional inverse opal substrates make them suitable for important applications in tissue engineering.

  19. Effect of Water-Glass Coating on HA and HA-TCP Samples for MSCs Adhesion, Proliferation, and Differentiation

    PubMed Central

    Bajpai, Indu; Kim, Duk Yeon; Kyong-Jin, Jung; Song, In-Hwan

    2016-01-01

    Ca-P and silicon based materials have become very popular as bone tissue engineering materials. In this study, water-glass (also known as sodium silicate glass) was coated on sintered hydroxyapatite (HA) and HA-TCP (TCP stands for tricalcium phosphate) samples and subsequently heat-treated at 600°C for 2 hrs. X-rays diffraction showed the presence of β- and α-TCP phases along with HA in the HA-TCP samples. Samples without coating, with water-glass coating, and heat-treated after water-glass coating were used to observe the adhesion and proliferation response of bone marrow derived-mesenchymal stem cells (MSCs). Cell culture was carried out for 4 hrs, 1 day, and 7 days. Interestingly, all samples showed similar response for cell adhesion and proliferation up to 7-day culture but fibronectin, E-cadherin, and osteogenic differentiation related genes (osteocalcin and osteopontin) were significantly induced in heat-treated water-glass coated HA-TCP samples. A water-glass coating on Ca-P samples was not found to influence the cell proliferation response significantly but activated some extracellular matrix genes and induced osteogenic differentiation in the MSCs. PMID:27429988

  20. Analytical cell adhesion chromatography reveals impaired persistence of metastatic cell rolling adhesion to P-selectin

    PubMed Central

    Oh, Jaeho; Edwards, Erin E.; McClatchey, P. Mason; Thomas, Susan N.

    2015-01-01

    ABSTRACT Selectins facilitate the recruitment of circulating cells from the bloodstream by mediating rolling adhesion, which initiates the cell–cell signaling that directs extravasation into surrounding tissues. To measure the relative efficiency of cell adhesion in shear flow for in vitro drug screening, we designed and implemented a microfluidic-based analytical cell adhesion chromatography system. The juxtaposition of instantaneous rolling velocities with elution times revealed that human metastatic cancer cells, but not human leukocytes, had a reduced capacity to sustain rolling adhesion with P-selectin. We define a new parameter, termed adhesion persistence, which is conceptually similar to migration persistence in the context of chemotaxis, but instead describes the capacity of cells to resist the influence of shear flow and sustain rolling interactions with an adhesive substrate that might modulate the probability of extravasation. Among cell types assayed, adhesion persistence to P-selectin was specifically reduced in metastatic but not leukocyte-like cells in response to a low dose of heparin. In conclusion, we demonstrate this as an effective methodology to identify selectin adhesion antagonist doses that modulate homing cell adhesion and engraftment in a cell-subtype-selective manner. PMID:26349809

  1. Bistability of cell adhesion in shear flow.

    PubMed

    Efremov, Artem; Cao, Jianshu

    2011-09-07

    Cell adhesion plays a central role in multicellular organisms helping to maintain their integrity and homeostasis. This complex process involves many different types of adhesion proteins, and synergetic behavior of these proteins during cell adhesion is frequently observed in experiments. A well-known example is the cooperation of rolling and stationary adhesion proteins during the leukocytes extravasation. Despite the fact that such cooperation is vital for proper functioning of the immune system, its origin is not fully understood. In this study we constructed a simple analytic model of the interaction between a leukocyte and the blood vessel wall in shear flow. The model predicts existence of cell adhesion bistability, which results from a tug-of-war between two kinetic processes taking place in the cell-wall contact area-bond formation and rupture. Based on the model results, we suggest an interpretation of several cytoadhesion experiments and propose a simple explanation of the existing synergy between rolling and stationary adhesion proteins, which is vital for effective cell adherence to the blood vessel walls in living organisms.

  2. Effect of molecular weight and concentration of hyaluronan on cell proliferation and osteogenic differentiation in vitro.

    PubMed

    Zhao, Ningbo; Wang, Xin; Qin, Lei; Guo, Zhengze; Li, Dehua

    2015-09-25

    Hyaluronan (HA), the simplest glycosaminoglycan and a major component of the extracellular matrix, exists in various tissues. It is involved in some critical biological procedures, including cellular signaling, cell adhesion and proliferation, and cell differentiation. The effect of molecular weight (MW) and concentration of HA on cell proliferation and differentiation was controversial. In this study, we investigated the effect of MW and concentration of HA on the proliferation and osteogenic differentiation of rabbit bone marrow-derived stem cells in vitro. Results showed that high MW HA decreased the cell adhesion rate in a concentration-dependant manner. The cell adhesion rate was decreased by increasing MW of HA. Cell proliferation was significantly enhanced by low MW HA (P < 0.05). The factorial analysis indicated that MW and concentration had an interactive effect on the cell adhesion rate and cell proliferation (P < 0.05). High MW HA increased the mRNA expressions of ALP, RUNX-2 and OCN. The higher the MW was, the higher the mRNA expressions were. The factorial analysis indicated that MW and concentration had an interactive effect on ALP mRNA expression (P < 0.05). HA of higher MW and higher concentration promoted bone formation. These findings provide some useful information in understanding the mechanism underlying the effect of MW and concentration of HA on cell proliferation and differentiation.

  3. Cell Adhesion and Growth on the Anodized Aluminum Oxide Membrane.

    PubMed

    Park, Jeong Su; Moon, Dalnim; Kim, Jin-Seok; Lee, Jin Seok

    2016-03-01

    Nanotopological cues are popular tools for in vivo investigation of the extracellular matrix (ECM) and cellular microenvironments. The ECM is composed of multiple components and generates a complex microenvironment. The development of accurate in vivo methods for the investigation of ECM are important for disease diagnosis and therapy, as well as for studies on cell behavior. Here, we fabricated anodized aluminum oxide (AAO) membranes using sulfuric and oxalic acid under controlled voltage and temperature. The membranes were designed to possess three different pore and interpore sizes, AAO-1, AAO-2, and AAO-3 membranes, respectively. These membranes were used as tools to investigate nanotopology-signal induced cell behavior. Cancerous cells, specifically, the OVCAR-8 cell-line, were cultured on porous AAO membranes and the effects of these membranes on cell shape, proliferation, and viability were studied. AAO-1 membranes bearing small sized pores were found to maintain the spreading shape of the cultured cells. Cells cultured on AAO-2 and AAO-3 membranes, bearing large pore-sized AAO membranes, changed shape from spreading to rounding. Furthermore, cellular area decreased when cells were cultured on all three AAO membranes that confirmed decreased levels of focal adhesion kinase (FAK). Additionally, OVCAR-8 cells exhibited increased proliferation on AAO membranes possessing various pore sizes, indicating the importance of the nanosurface structure in regulating cell behaviors, such as cell proliferation. Our results suggest that porous-AAO membranes induced nanosurface regulated cell behavior as focal adhesion altered the intracellular organization of the cytoskeleton. Our results may find potential applications as tools in in vivo cancer research studies.

  4. Connexin 43 expressed in endothelial cells modulates monocyte‑endothelial adhesion by regulating cell adhesion proteins.

    PubMed

    Yuan, Dongdong; Sun, Guoliang; Zhang, Rui; Luo, Chenfang; Ge, Mian; Luo, Gangjian; Hei, Ziqing

    2015-11-01

    Adhesion between circulating monocytes and vascular endothelial cells is a key initiator of atherosclerosis. In our previous studies, it was demonstrated that the expression of connexin (Cx)43 in monocytes modulates cell adhesion, however, the effects of the expression of Cx43 in endothelial cells remains to be elucidated. Therefore, the present study investigated the role of the expression of Cx43 in endothelial cells in the process of cell adhesion. A total of four different methods with distinct mechanisms were used to change the function and expression of Cx43 channels in human umbilical vein endothelial cells: Cx43 channel inhibitor (oleamide), enhancer (retinoic acid), overexpression of Cx43 by transfection with pcDNA‑Cx43 and knock‑down of the expression of Cx43 by small interfering RNA against Cx43. The results indicated that the upregulation of the expression of Cx43 enhanced monocyte‑endothelial adhesion and this was markedly decreased by downregulation of Cx43. This mechanism was associated with Cx43‑induced expression of vascular cell adhesion molecule‑1 and intercellular cell adhesion molecule‑1. The effects of Cx43 in endothelial cells was independent of Cx37 or Cx40. These experiments suggested that local regulation of endothelial Cx43 expression within the vasculature regulates monocyte‑endothelial adhesion, a critical event in the development of atherosclerosis and other inflammatory pathologies, with baseline adhesion set by the expression of Cx43. This balance may be crucial in controlling leukocyte involvement in inflammatory cascades.

  5. Control of three-dimensional cell adhesion by the chirality of nanofibers in hydrogels.

    PubMed

    Liu, Guo-Feng; Zhang, Di; Feng, Chuan-Liang

    2014-07-21

    In the three-dimensional (3D) extracellular matrix (ECM), the influence of nanofiber chirality on cell behavior is very important; the helical nanofibrous structure is closely related to the relevant biological events. Herein, we describe the use of the two enantiomers of a 1,4-benzenedicarboxamide phenylalanine derivative as supramolecular gelators to investigate the influence of the chirality of nanofibers on cell adhesion and proliferation in three dimensions. It was found that left-handed helical nanofibers can increase cell adhesion and proliferation, whereas right-handed nanofibers have the opposite effect. These effects are ascribed to the mediation of the stereospecific interaction between chiral nanofibers and fibronectin. The results stress the crucial role of the chirality of nanofibers on cell-adhesion and cell-proliferation behavior in 3D environments.

  6. Peptide-decorated chitosan derivatives enhance fibroblast adhesion and proliferation in wound healing.

    PubMed

    Patrulea, V; Hirt-Burri, N; Jeannerat, A; Applegate, L A; Ostafe, V; Jordan, O; Borchard, G

    2016-05-20

    RGD peptide sequences are known to regulate cellular activities by interacting with α5β1, αvβ5 and αvβ3 integrin, which contributes to the wound healing process. In this study, RGDC peptide was immobilized onto chitosan derivative 1,6-diaminohexane-O-carboxymethyl-N,N,N-trimethyl chitosan (DAH-CMTMC) to display RGDC-promoting adhesion for enhanced wound healing. The efficiency of N-methylation, O-carboxymethylation and spacer grafting was quantitatively and qualitatively analyzed by (1)H NMR and FTIR, yielding 0.38 degree of substitution for N-methylation and >0.85 for O-carboxymethylation. The glass transition temperatures for chitosan derivatives were also studied. Peptide immobilization was achieved through sulfhydryl groups using sulfosuccinimidyl (4-iodoacetyl)amino-benzoate (sulfo-SIAB method). RGDC immobilized peptide onto DAH-CMTMC was found to be about 15.3 μg/mg of chitosan derivative by amino acid analysis (AAA). The significant increase of human dermal fibroblast (HDF) viability in vitro over 7 days suggests that RGDC-functionalized chitosan may lead to enhanced wound healing (viability >140%). Moreover, bio-adhesion and proliferation assays confirmed that coatings of RGDC-functionalized chitosan derivatives exhibit in vitro wound healing properties by enhancing fibroblast proliferation and adhesion. These results showed that RGDC peptide-functionalized chitosan provides an optimal environment for fibroblast adhesion and proliferation.

  7. Yielding Elastic Tethers Stabilize Robust Cell Adhesion

    PubMed Central

    Whitfield, Matt J.; Luo, Jonathon P.; Thomas, Wendy E.

    2014-01-01

    Many bacteria and eukaryotic cells express adhesive proteins at the end of tethers that elongate reversibly at constant or near constant force, which we refer to as yielding elasticity. Here we address the function of yielding elastic adhesive tethers with Escherichia coli bacteria as a model for cell adhesion, using a combination of experiments and simulations. The adhesive bond kinetics and tether elasticity was modeled in the simulations with realistic biophysical models that were fit to new and previously published single molecule force spectroscopy data. The simulations were validated by comparison to experiments measuring the adhesive behavior of E. coli in flowing fluid. Analysis of the simulations demonstrated that yielding elasticity is required for the bacteria to remain bound in high and variable flow conditions, because it allows the force to be distributed evenly between multiple bonds. In contrast, strain-hardening and linear elastic tethers concentrate force on the most vulnerable bonds, which leads to failure of the entire adhesive contact. Load distribution is especially important to noncovalent receptor-ligand bonds, because they become exponentially shorter lived at higher force above a critical force, even if they form catch bonds. The advantage of yielding is likely to extend to any blood cells or pathogens adhering in flow, or to any situation where bonds are stretched unequally due to surface roughness, unequal native bond lengths, or conditions that act to unzip the bonds. PMID:25473833

  8. Investigation of Cell-Substrate Adhesion Properties of Living Chondrocyte by Measuring Adhesive Shear Force and Detachment Using AFM and Inverse FEA

    PubMed Central

    Nguyen, Trung Dung; Gu, YuanTong

    2016-01-01

    It is well-known that cell adhesion is important in many biological processes such as cell migration and proliferation. A better understanding of the cell adhesion process will shed insight into these cellular biological responses as well as cell adhesion-related diseases treatment. However, there is little research which has attempted to investigate the process of cell adhesion and its mechanism. Thus, this paper aims to study the time-dependent adhesion properties of single living chondrocytes using an advanced coupled experimental-numerical approach. Atomic Force Microscopy (AFM) tips will be used to apply lateral forces to detach chondrocytes that are seeded for three different periods. An advanced Finite Element Analysis (FEA) model combining porohyperelastic (PHE) constitutive model and cohesive zone formulation is developed to explore the mechanism of adhesion. The results revealed that the cells can resist normal traction better than tangential traction in the beginning of adhesion. This is when the cell adhesion molecules establish early attachment to the substrates. After that when the cells are spreading, stress fiber bundles generate tangential traction on the substrate to form strong adhesion. Both simulation and experimental results agree well with each other, providing a powerful tool to study the cellular adhesion process. PMID:27892536

  9. Pattern formation in cell membrane adhesion

    NASA Astrophysics Data System (ADS)

    Discher, Dennis; Hategan, A.; Sengupta, K.; Sackmann, E.

    2004-03-01

    Strong adhesion of highly active cells often nucleates focal adhesions or related structures that are, over time, reinforced by cytoskeleton (actin, etc.). Red cells lack such complex adhesion systems, but they are shown here to also exhibit complex spatial patterns within an adhesive contact zone. While strong adhesion and spreading of the red cell to a dense poly-L-lysine surface appears complete in < 1 s by reflective interference microscopy, over longer times of 10-15 min or more distinct patterns in fluorescently labeled membrane components emerge. The fluorescent lipid Fl-PE (fluorescein phosphoethanolamine), in particular, is seen to diffuse and reorganize (eg. worm-like domains of <500 nm) within the contact zone, independent of whether the cell is intact or ruptured. Lipid patterns are accompanied by visible perturbations in band 3 distribution and weaker perturbations in membrane skeleton actin. Although fluorescent poly-L-lysine is shown to be uniform under cells, pressing down on the membrane quenches the lipid patterns and reveals the topographical basis for pattern formation. Regions of strong contact are thus separated by regions where the membrane is more distant from the surface.

  10. Control of mesenchymal stem cell phenotype and differentiation depending on cell adhesion mechanism.

    PubMed

    Kang, J; Park, H M; Kim, Y W; Kim, Y H; Varghese, S; Seok, H K; Kim, Y G; Kim, S H

    2014-11-25

    Control of cell-matrix adhesion has become an important issue in the regulation of stem cell function. In this study, a maltose-binding protein (MBP)-linked basic fibroblast growth factor (FGF2)-immobilised polystyrene surface (PS-MBP-FGF2) was applied as an artificial matrix to regulate integrin-mediated signalling. We sought to characterise human mesenchymal-stem cell (hMSC) behaviour in response to two different mechanisms of cell adhesion; (i) FGF2-heparan sulphate proteoglycan (HSPG)-mediated adhesion vs. (ii) fibronectin (FN)-integrin-mediated adhesion. Heparin inhibited hMSC adhesion to PS-MBP-FGF2 but not to FN-coated surface. The phosphorylation of focal adhesion kinase, cytoskeletal re-organisation, and cell proliferation were restricted in hMSCs adhering to PS-MBP-FGF2 compared to FN-coated surface. Expression of MSC markers, such as CD105, CD90 and CD166, decreased in hMSCs expanded on PS-MBP-FGF2 compared to expression in cells expanded on FN-coated surface. hMSCs that were expanded on FN-coated surface differentiated into osteogenic and adipogenic cells more readily than those that were expanded on PS-MBP-FGF2. Furthermore, we characterised the N-linked glycan structures of hMSCs depending on the cell adhesion mechanism using mass spectrometry (MS)-based quantitative techniques. MS analysis revealed that 2,3-sialylated glycans, a potential marker of stem cell function, were more abundant on hMSCs expanded on FN-coated surface than on those expanded on PS-MBP-FGF2. Thus, the differentiation potential of hMSCs is controlled by the type of adhesion substrate that might provide an idea for the design of biomaterials to control stem cell fate. Elucidation of the glycan structure on the cell membrane may help characterise hMSC function.

  11. Cell proliferation in human coronary arteries.

    PubMed Central

    Gordon, D; Reidy, M A; Benditt, E P; Schwartz, S M

    1990-01-01

    Despite the lack of direct evidence for cell multiplication, proliferation of smooth muscle cells in human atherosclerotic lesions has been assumed to play a central role in ontogeny of the plaque. We used antibodies to cell cycle-related proteins on tissue sections of human arteries and coronary atherosclerotic plaques. Specific cell types were identified by immunochemical reagents for smooth muscle, monocyte-macrophages, and other blood cells. Low rates of smooth muscle cell proliferation were observed. Macrophages were also observed with rates of proliferation comparable to that of the smooth muscle. Additional replicating cells could not be defined as belonging to specific cell types with the reagents used in this study. These findings imply that smooth muscle replication in advanced plaques is indolent and raise the possibility of a role for proliferating leukocytes. Images PMID:1972277

  12. Decreased cell adhesion promotes angiogenesis in a Pyk2-dependent manner

    SciTech Connect

    Shen, Colette J.; Raghavan, Srivatsan; Xu, Zhe; Baranski, Jan D.; Yu, Xiang; Wozniak, Michele A.; Miller, Jordan S.; Gupta, Mudit; Buckbinder, Leonard; Chen, Christopher S.

    2011-08-01

    Angiogenesis is regulated by both soluble growth factors and cellular interactions with the extracellular matrix (ECM). While cell adhesion via integrins has been shown to be required for angiogenesis, the effects of quantitative changes in cell adhesion and spreading against the ECM remain less clear. Here, we show that angiogenic sprouting in natural and engineered three-dimensional matrices exhibited a biphasic response, with peak sprouting when adhesion to the matrix was limited to intermediate levels. Examining changes in global gene expression to determine a genetic basis for this response, we demonstrate a vascular endothelial growth factor (VEGF)-induced upregulation of genes associated with vascular invasion and remodeling when cell adhesion was limited, whereas cells on highly adhesive surfaces upregulated genes associated with proliferation. To explore a mechanistic basis for this effect, we turned to focal adhesion kinase (FAK), a central player in adhesion signaling previously implicated in angiogenesis, and its homologue, proline-rich tyrosine kinase 2 (Pyk2). While FAK signaling had some impact, our results suggested that Pyk2 can regulate both gene expression and endothelial sprouting through its enhanced activation by VEGF in limited adhesion contexts. We also demonstrate decreased sprouting of tissue explants from Pyk2-null mice as compared to wild type mice as further confirmation of the role of Pyk2 in angiogenic sprouting. These results suggest a surprising finding that limited cell adhesion can enhance endothelial responsiveness to VEGF and demonstrate a novel role for Pyk2 in the adhesive regulation of angiogenesis.

  13. Keratinocytes from APP/APLP2-deficient mice are impaired in proliferation, adhesion and migration in vitro.

    PubMed

    Siemes, Christina; Quast, Thomas; Kummer, Christiane; Wehner, Sven; Kirfel, Gregor; Müller, Ulrike; Herzog, Volker

    2006-07-01

    Growing evidence shows that the soluble N-terminal form (sAPPalpha) of the amyloid precursor protein (APP) represents an epidermal growth factor fostering keratinocyte proliferation, migration and adhesion. APP is a member of a protein family including the two mammalian amyloid precursor-like proteins APLP1 and APLP2. In the mammalian epidermis, only APP and APLP2 are expressed. APP and APLP2-deficient mice die shortly after birth but do not display a specific epidermal phenotype. In this report, we investigated the epidermis of APP and/or APLP2 knockout mice. Basal keratinocytes showed reduced proliferation in vivo by about 40%. Likewise, isolated keratinocytes exhibited reduced proliferation rates in vitro, which could be completely rescued by either exogenously added recombinant sAPPalpha, or by co-culture with dermal fibroblasts derived from APP knockout mice. Moreover, APP-knockout keratinocytes revealed reduced migration velocity resulting from severely compromised cell substrate adhesion. Keratinocytes from double knockout mice died within the first week of culture, indicating essential functions of APP-family members for survival in vitro. Our data indicate that sAPPalpha has to be considered as an essential epidermal growth factor which, however, in vivo can be functionally compensated to a certain extent by other growth factors, e.g., factors released from dermal fibroblasts.

  14. Keratinocytes from APP/APLP2-deficient mice are impaired in proliferation, adhesion and migration in vitro

    SciTech Connect

    Siemes, Christina; Quast, Thomas; Kummer, Christiane; Wehner, Sven; Kirfel, Gregor; Mueller, Ulrike; Herzog, Volker . E-mail: Herzog@uni-bonn.de

    2006-07-01

    Growing evidence shows that the soluble N-terminal form (sAPP{alpha}) of the amyloid precursor protein (APP) represents an epidermal growth factor fostering keratinocyte proliferation, migration and adhesion. APP is a member of a protein family including the two mammalian amyloid precursor-like proteins APLP1 and APLP2. In the mammalian epidermis, only APP and APLP2 are expressed. APP and APLP2-deficient mice die shortly after birth but do not display a specific epidermal phenotype. In this report, we investigated the epidermis of APP and/or APLP2 knockout mice. Basal keratinocytes showed reduced proliferation in vivo by about 40%. Likewise, isolated keratinocytes exhibited reduced proliferation rates in vitro, which could be completely rescued by either exogenously added recombinant sAPP{alpha}, or by co-culture with dermal fibroblasts derived from APP knockout mice. Moreover, APP-knockout keratinocytes revealed reduced migration velocity resulting from severely compromised cell substrate adhesion. Keratinocytes from double knockout mice died within the first week of culture, indicating essential functions of APP-family members for survival in vitro. Our data indicate that sAPP{alpha} has to be considered as an essential epidermal growth factor which, however, in vivo can be functionally compensated to a certain extent by other growth factors, e.g., factors released from dermal fibroblasts.

  15. Diversity of cell-mediated adhesions in breast cancer spheroids.

    PubMed

    Ivascu, Andrea; Kubbies, Manfred

    2007-12-01

    Due to their three dimensional (3D) architecture, multicellular tumor spheroids mimic avascular tumor areas comprising the establishment of diffusion gradients, reduced proliferation rates and increased drug resistance. We have shown recently that the spontaneous formation of spheroids is restricted to a limited number of cell lines whereas the majority grow only as aggregates of cells with loose cell-cell contacts when cultured in 3D. However, by the addition of reconstituted basement membrane (rBM, Matrigel), aggregates can be transformed into spheroids with diffusion barriers and development of quiescent therapy-resistant cells. In this report, we investigated adhesion molecules responsible for rBM-driven versus spontaneous spheroid formation in a diverse population of eight breast tumor cell lines relevant for in vitro and in vivo antitumor drug testing. Inhibition of spheroid formation was monitored in the presence of adhesion molecule functional blocking antibodies and after siRNA-mediated down-regulation of E- and N-cadherin and integrin beta1 adhesion receptors. We identified that E-cadherin mediates the spontaneous formation of spheroids in MCF7, BT-474, T-47D and MDA-MB-361 cells, whereas N-cadherin is responsible for tight packing of MDA-MB-435S cells. In contrast, the matrix protein-induced transformation of 3D aggregates into spheroids in MDA-MB-231 and SK-BR-3 cells is mediated primarily by the collagen I/integrin beta1 interaction with no cadherin involvement. A combination of both, homophilic E-cadherin and integrin beta1/collagen I interaction establishes spheroids in MDA-MB-468 cells. These findings indicate that an evolutionary diverse and complex pattern of interacting cell surface proteins exists in breast cancer cells that determines the 3D growth characteristic in vitro, thereby influencing small molecule or antibody permeation in preclinical in vitro and in vivo tumor models.

  16. Slug regulates integrin expression and cell proliferation in human epidermal keratinocytes.

    PubMed

    Turner, Frances E; Broad, Simon; Khanim, Farhat L; Jeanes, Alexa; Talma, Sonia; Hughes, Sharon; Tselepis, Chris; Hotchin, Neil A

    2006-07-28

    The human epidermis is a self-renewing epithelial tissue composed of several layers of keratinocytes. Within the epidermis there exists a complex array of cell adhesion structures, and many of the cellular events within the epidermis (differentiation, proliferation, and migration) require that these adhesion structures be remodeled. The link between cell adhesion, proliferation, and differentiation within the epidermis is well established, and in particular, there is strong evidence to link the process of terminal differentiation to integrin adhesion molecule expression and function. In this paper, we have analyzed the role of a transcriptional repressor called Slug in the regulation of adhesion molecule expression and function in epidermal keratinocytes. We report that activation of Slug, which is expressed predominantly in the basal layer of the epidermis, results in down-regulation of a number of cell adhesion molecules, including E-cadherin, and several integrins, including alpha3, beta1, and beta4. We demonstrate that Slug binds to the alpha3 promoter and that repression of alpha3 transcription by Slug is dependent on an E-box sequence within the promoter. This reduction in integrin expression is reflected in decreased cell adhesion to fibronectin and laminin-5. Despite the reduction in integrin expression and function, we do not observe any increase in differentiation. We do, however, find that activation of Slug results in a significant reduction in keratinocyte proliferation.

  17. Anandamide inhibits adhesion and migration of breast cancer cells

    SciTech Connect

    Grimaldi, Claudia; Pisanti, Simona; Laezza, Chiara; Malfitano, Anna Maria; Santoro, Antonietta; Vitale, Mario; Caruso, Maria Gabriella; Notarnicola, Maria; Iacuzzo, Irma; Portella, Giuseppe; Di Marzo, Vincenzo . E-mail: vdimarzo@icmib.na.cnr.it; Bifulco, Maurizio . E-mail: maubiful@unina.it

    2006-02-15

    The endocannabinoid system regulates cell proliferation in human breast cancer cells. We reasoned that stimulation of cannabinoid CB{sub 1} receptors could induce a non-invasive phenotype in breast mtastatic cells. In a model of metastatic spreading in vivo, the metabolically stable anandamide analogue, 2-methyl-2'-F-anandamide (Met-F-AEA), significantly reduced the number and dimension of metastatic nodes, this effect being antagonized by the selective CB{sub 1} antagonist SR141716A. In MDA-MB-231 cells, a highly invasive human breast cancer cell line, and in TSA-E1 cells, a murine breast cancer cell line, Met-F-AEA inhibited adhesion and migration on type IV collagen in vitro without modifying integrin expression: both these effects were antagonized by SR141716A. In order to understand the molecular mechanism involved in these processes, we analyzed the phosphorylation of FAK and Src, two tyrosine kinases involved in migration and adhesion. In Met-F-AEA-treated cells, we observed a decreased tyrosine phosphorylation of both FAK and Src, this effect being attenuated by SR141716A. We propose that CB{sub 1} receptor agonists inhibit tumor cell invasion and metastasis by modulating FAK phosphorylation, and that CB{sub 1} receptor activation might represent a novel therapeutic strategy to slow down the growth of breast carcinoma and to inhibit its metastatic diffusion in vivo.

  18. Redundant control of migration and adhesion by ERM proteins in vascular smooth muscle cells

    SciTech Connect

    Baeyens, Nicolas; Latrache, Iman; Yerna, Xavier; Noppe, Gauthier; Horman, Sandrine; Morel, Nicole

    2013-11-22

    Highlights: •The three ERM proteins are expressed in vascular smooth muscle cell. •ERM depletion inhibited PDGF-evoked migration redundantly. •ERM depletion increased cell adhesion redundantly. •ERM depletion did not affect PDGF-evoked Ca signal, Rac1 activation, proliferation. •ERM proteins control PDGF-induced migration by regulating adhesion. -- Abstract: Ezrin, radixin, and moesin possess a very similar structure with a C-terminal actin-binding domain and a N-terminal FERM interacting domain. They are known to be involved in cytoskeleton organization in several cell types but their function in vascular smooth muscle cells (VSMC) is still unknown. The aim of this study was to investigate the role of ERM proteins in cell migration induced by PDGF, a growth factor involved in pathophysiological processes like angiogenesis or atherosclerosis. We used primary cultured VSMC obtained from rat aorta, which express the three ERM proteins. Simultaneous depletion of the three ERM proteins with specific siRNAs abolished the effects of PDGF on cell architecture and migration and markedly increased cell adhesion and focal adhesion size, while these parameters were only slightly affected by depletion of ezrin, radixin or moesin alone. Rac1 activation, cell proliferation, and Ca{sup 2+} signal in response to PDGF were unaffected by ERM depletion. These results indicate that ERM proteins exert a redundant control on PDGF-induced VSMC migration by regulating focal adhesion turn-over and cell adhesion to substrate.

  19. Silencing of VAMP3 inhibits cell migration and integrin-mediated adhesion

    SciTech Connect

    Luftman, Kevin; Hasan, Nazarul; Day, Paul; Hardee, Deborah; Hu Chuan

    2009-02-27

    Integrins are transmembrane receptors for cell adhesion to the extracellular matrix. In cell migration, integrins are endocytosed from the plasma membrane or the cell surface, transported in vesicles and exocytosed actively at the cell front. In the present study, we examined the roles of VAMP3, a SNARE protein that mediates exocytosis, in cell migration and integrin trafficking. Small interfering RNA (siRNA)-induced silencing of VAMP3 inhibited chemotactic cell migration by more than 60% without affecting cell proliferation. VAMP3 silencing reduced the levels of {beta}1 integrin at the cell surface but had no effect on total cellular {beta}1 integrin, indicating that VAMP3 is required for trafficking of {beta}1 integrin to the plasma membrane. Furthermore, VAMP3 silencing diminished cell adhesion to laminin but not to fibronectin or collagen. Taken together, these data suggest that VAMP3-dependent integrin trafficking is crucial in cell migration and cell adhesion to laminin.

  20. Self assembling bioactive materials for cell adhesion in tissue repair

    NASA Astrophysics Data System (ADS)

    Hwang, Julia J.

    This work involved the study of biodegradable and biocompatible materials that have the potential to modify tissue engineering scaffolds through self assembly, generating multiple layers that deliver bioactivity. Diblock biomaterials containing cholesteryl moieties and oligomers of lactic acid units were found to form single crystals when precipitated from hot ethanol and smectic liquid crystalline phases when cast as a film. Cell culture experiments on these films with 3T3 and 3T6 fibroblasts indicated that these ordered materials form surfaces with specific chemistries that favored cell adhesion, spreading, and proliferation suggesting the potential of mediating human tissue repair. The author believes the cholesteryl moieties found on the surface play a key role in determining cell behavior. Cholesteryl-(L-lactic acid) diblock molecules were then functionalized with moieties including vitamin Bx, cholesterol, and the anti-inflammatory drug indomethacin. An unstable activated ester between indomethacin and the diblock molecule resulted in the release of indomethacin into the culture medium which inhibited the proliferation of 3T3 fibroblasts. Finally, a series of molecules were designed to incorporate dendrons based on amino acids at the termini of the diblock structures. It was determined that lysine, a basic amino acid, covalently coupled to cholesteryl-(L-lactic acid) can promote cell adhesion and spreading while negatively charged and zwitterionic 2nd generation dendrons based on aspartic acid do not. Incorporation of the well known arginine-glycine-aspartic acid (RGD) sequence, which is found in many adhesive proteins, to the dendrons imparted integrin-mediated cell adhesion as evidenced by the formation of stress fibers. We also explored the capacity of integrin receptors to bind to ligands that are not the linear form of RGD, but have R, G, and D spatially positioned to mimic the linear RGD environments. For this purpose, the arms of the 2 nd generation

  1. Multi-scale models for cell adhesion

    NASA Astrophysics Data System (ADS)

    Wu, Yinghao; Chen, Jiawen; Xie, Zhong-Ru

    2014-03-01

    The interactions of membrane receptors during cell adhesion play pivotal roles in tissue morphogenesis during development. Our lab focuses on developing multi-scale models to decompose the mechanical and chemical complexity in cell adhesion. Recent experimental evidences show that clustering is a generic process for cell adhesive receptors. However, the physical basis of such receptor clustering is not understood. We introduced the effect of molecular flexibility to evaluate the dynamics of receptors. By delivering new theory to quantify the changes of binding free energy in different cellular environments, we revealed that restriction of molecular flexibility upon binding of membrane receptors from apposing cell surfaces (trans) causes large entropy loss, which dramatically increases their lateral interactions (cis). This provides a new molecular mechanism to initialize receptor clustering on the cell-cell interface. By using the subcellular simulations, we further found that clustering is a cooperative process requiring both trans and cis interactions. The detailed binding constants during these processes are calculated and compared with experimental data from our collaborator's lab.

  2. Simulation of proliferation and differentiation of cells in a stem-cell niche

    NASA Astrophysics Data System (ADS)

    Zhdanov, Vladimir P.

    2008-10-01

    Stem-cell niches represent microscopic compartments formed of environmental cells that nurture stem cells and enable them to maintain tissue homeostasis. The spatio-temporal kinetics of proliferation and differentiation of cells in such niches depend on the specifics of the niche structure and on adhesion and communication between cells and may also be influenced by spatial constraints on cell division. We propose a generic lattice model, taking all these factors into account, and systematically illustrate their role. The model is motivated by the experimental data available for the niches located in the subventricular zone of adult mammalian brain. The general conclusions drawn from our Monte Carlo simulations are applicable to other niches as well. One of our main findings is that the kinetics under consideration are highly stochastic due to a relatively small number of cells proliferating and differentiating in a niche and the autocatalytic character of the symmetric cell division. In particular, the kinetics exhibit huge stochastic bursts especially if the adhesion between cells is taken into account. In addition, the results obtained show that despite the small number of cells present in stem-cell niches, their arrangement can be predetermined to appreciable extent provided that the adhesion of different cells is different so that they tend to segregate.

  3. Cell proliferation and differentiation in chemical leukemogenesis

    NASA Technical Reports Server (NTRS)

    Irons, R. D.; Stillman, W. S.; Clarkson, T. W. (Principal Investigator)

    1993-01-01

    In tissues such as bone marrow with normally high rates of cell division, proliferation is tightly coordinated with cell differentiation. Survival, proliferation and differentiation of early hematopoietic progenitor cells depend on the growth factors, interleukin 3 (IL-3) and/or granulocyte-macrophage colony stimulating factor (GM-CSF) and their synergism with other cytokines. We provide evidence that a characteristic shared by a diverse group of compounds with demonstrated leukemogenic potential is the ability to act synergistically with GM-CSF. This results in an increase in recruitment of a resting population of hematopoietic progenitor cells normally unresponsive to the cytokine and a twofold increase in the size of the proliferating cell population normally regarded to be at risk of transformation in leukemogenesis. These findings support the possibility that transient alterations in hematopoietic progenitor cell differentiation may be an important factor in the early stages of development of leukemia secondary to chemical or drug exposure.

  4. Tumor exosome-mediated promotion of adhesion to mesothelial cells in gastric cancer cells

    PubMed Central

    Arita, Tomohiro; Ichikawa, Daisuke; Konishi, Hirotaka; Komatsu, Shuhei; Shiozaki, Atsushi; Ogino, Shinpei; Fujita, Yuji; Hiramoto, Hidekazu; Hamada, Junichi; Shoda, Katsutoshi; Kosuga, Toshiyuki; Fujiwara, Hitoshi; Okamoto, Kazuma; Otsuji, Eigo

    2016-01-01

    Background Peritoneal metastasis consists of a highly complex series of steps, and the details of the underlying molecular mechanism remain largely unclear. In this study, the effects of tumor-derived exosomes (TEX) on the progression of gastric cancers were investigated in peritoneal metastasis. Results TEX were internalized in both mesothelial and gastric cancer cells in a cellular origin non-specific manner. Internalization of TEX into mesothelial cells promoted significant adhesion between mesothelial and gastric cancer cells, and TEX internalization into gastric cancer cells significantly promoted migratory ability, while internalization of mesothelial cell-derived exosomes did not. Expression of adhesion-related molecules, such as fibronectin 1 (FN1) and laminin gamma 1 (LAMC1), were increased in mesothelial cells after internalization of TEX from gastric cancer cell line and malignant pleural effusion. Methods TEX were extracted from cell-conditioned medium by ultracentrifugation. The effects of TEX on the malignant potential of gastric cancer were investigated in adhesion, invasion, and proliferation assays. PCR array as well as western blotting were performed to determine the underlying molecular mechanisms. The molecular changes in mesothelial cell after internalization of TEX derived from malignant pleural effusion were also confirmed. Conclusions TEX may play a critical role in the development of peritoneal metastasis of gastric cancer, which may be partially due to inducing increased expression of adhesion molecules in mesothelial cells. PMID:27487135

  5. Blue light inhibits proliferation of melanoma cells

    NASA Astrophysics Data System (ADS)

    Becker, Anja; Distler, Elisabeth; Klapczynski, Anna; Arpino, Fabiola; Kuch, Natalia; Simon-Keller, Katja; Sticht, Carsten; van Abeelen, Frank A.; Gretz, Norbert; Oversluizen, Gerrit

    2016-03-01

    Photobiomodulation with blue light is used for several treatment paradigms such as neonatal jaundice, psoriasis and back pain. However, little is known about possible side effects concerning melanoma cells in the skin. The aim of this study was to assess the safety of blue LED irradiation with respect to proliferation of melanoma cells. For that purpose we used the human malignant melanoma cell line SK-MEL28. Cell proliferation was decreased in blue light irradiated cells where the effect size depended on light irradiation dosage. Furthermore, with a repeated irradiation of the melanoma cells on two consecutive days the effect could be intensified. Fluorescence-activated cell sorting with Annexin V and Propidium iodide labeling did not show a higher number of dead cells after blue light irradiation compared to non-irradiated cells. Gene expression analysis revealed down-regulated genes in pathways connected to anti-inflammatory response, like B cell signaling and phagosome. Most prominent pathways with up-regulation of genes were cytochrome P450, steroid hormone biosynthesis. Furthermore, even though cells showed a decrease in proliferation, genes connected to the cell cycle were up-regulated after 24h. This result is concordant with XTT test 48h after irradiation, where irradiated cells showed the same proliferation as the no light negative control. In summary, proliferation of melanoma cells can be decreased using blue light irradiation. Nevertheless, the gene expression analysis has to be further evaluated and more studies, such as in-vivo experiments, are warranted to further assess the safety of blue light treatment.

  6. Substrate rigidity regulates human T cell activation and proliferation.

    PubMed

    O'Connor, Roddy S; Hao, Xueli; Shen, Keyue; Bashour, Keenan; Akimova, Tatiana; Hancock, Wayne W; Kam, Lance C; Milone, Michael C

    2012-08-01

    Adoptive immunotherapy using cultured T cells holds promise for the treatment of cancer and infectious disease. Ligands immobilized on surfaces fabricated from hard materials such as polystyrene plastic are commonly employed for T cell culture. The mechanical properties of a culture surface can influence the adhesion, proliferation, and differentiation of stem cells and fibroblasts. We therefore explored the impact of culture substrate stiffness on the ex vivo activation and expansion of human T cells. We describe a simple system for the stimulation of the TCR/CD3 complex and the CD28 receptor using substrates with variable rigidity manufactured from poly(dimethylsiloxane), a biocompatible silicone elastomer. We show that softer (Young's Modulus [E] < 100 kPa) substrates stimulate an average 4-fold greater IL-2 production and ex vivo proliferation of human CD4(+) and CD8(+) T cells compared with stiffer substrates (E > 2 MPa). Mixed peripheral blood T cells cultured on the stiffer substrates also demonstrate a trend (nonsignificant) toward a greater proportion of CD62L(neg), effector-differentiated CD4(+) and CD8(+) T cells. Naive CD4(+) T cells expanded on softer substrates yield an average 3-fold greater proportion of IFN-γ-producing Th1-like cells. These results reveal that the rigidity of the substrate used to immobilize T cell stimulatory ligands is an important and previously unrecognized parameter influencing T cell activation, proliferation, and Th differentiation. Substrate rigidity should therefore be a consideration in the development of T cell culture systems as well as when interpreting results of T cell activation based upon solid-phase immobilization of TCR/CD3 and CD28 ligands.

  7. Substrate rigidity regulates human T cell activation and proliferation1

    PubMed Central

    O’Connor, Roddy S.; Hao, Xueli; Shen, Keyue; Bashour, Keenan; Akimova, Tatiana; Hancock, Wayne W.; Kam, Lance; Milone, Michael C.

    2012-01-01

    Adoptive immunotherapy using cultured T cells holds promise for the treatment of cancer and infectious disease. Ligands immobilized on surfaces fabricated from hard materials such as polystyrene plastic are commonly employed for T cell culture. The mechanical properties of a culture surface can influence the adhesion, proliferation, and differentiation of stem cells and fibroblasts. We therefore explored the impact of culture substrate stiffness on the ex vivo activation and expansion of human T cells. We describe a simple system for the stimulation of the TCR/CD3 complex and the CD28 receptor using substrates with variable rigidity manufactured from poly(dimethylsiloxane) (PDMS), a biocompatible silicone elastomer. We show that softer (Young’s Modulus [E] < 100 kPa) substrates stimulate an average 4-fold greater IL-2 production and ex vivo proliferation of human CD4+ and CD8+ T cells compared with stiffer substrates (E >2 MPa). Mixed peripheral blood T cells cultured on the stiffer substrates also demonstrate a trend (non-significant) towards a greater proportion of CD62Lneg, effector-differentiated CD4+ and CD8+ T cells. Naïve CD4+ T cells expanded on softer substrates yield an average 3-fold greater proportion of IFN-γ producing TH1-like cells. These results reveal that the rigidity of the substrate used to immobilize T cell stimulatory ligands is an important and previously unrecognized parameter influencing T cell activation, proliferation and TH differentiation. Substrate rigidity should therefore be a consideration in the development of T cell culture systems as well as when interpreting results of T cell activation based upon solid-phase immobilization of TCR/CD3 and CD28 ligands. PMID:22732590

  8. Engineered electrospun poly(caprolactone)/polycaprolactone-g-hydroxyapatite nano-fibrous scaffold promotes human fibroblasts adhesion and proliferation.

    PubMed

    Keivani, F; Shokrollahi, P; Zandi, M; Irani, S; F Shokrolahi; Khorasani, S C

    2016-11-01

    Polycaprolactone (PCL)/hydroxyapatite nano-composites are among the best candidates for tissue engineering. However, interactions between nHAp and PCL are difficult to control leading to inhomogeneous dispersion of the bio-ceramic particles. Grafting of polymer chains at high density/chain length while promotes the phase compatibility may result in reduced HAp exposed surface area and therefore, bioactivity is compromised. This issue is addressed here by grafting PCL chains onto HAp nano-particles through ring opening polymerization of ε-caprolactone (PCL-g-HAp). FTIR and TGA analysis showed that PCL (6.9wt%), was successfully grafted on the HAp. PCL/PCL-g-HAp nano-fibrous scaffold showed up to 10 and 33% enhancement in tensile strength and modulus, respectively, compared to those of PCL/HAp. The effects of HAp on the in vitro HAp formation were investigated for both the PCL/HAp and PCL/PCL-g-HAp scaffolds. Precipitation of HAp on the nano-composite scaffolds observed after 15days incubation in simulated body fluid (SBF), as confirmed by scanning electron microscopy (SEM), and energy dispersive X-ray spectroscopy (EDX). Human fibroblasts were seeded on PCL, PCL/HAp and PCL/PCL-g-HAp scaffolds. According to MTT assay, the highest cell proliferation was recorded for PCL/PCL-g-HAp nano-composite, at all time intervals (1-21days, P<0.001). Fluorescent microscopy (of DAPI stained samples) and electron microscopy images showed that all nano-fibrous scaffolds (PCL, PCL/HAp, and PCL/PCL-g-HAp), were non-toxic against cells, while more cell adhesion, and the most uniform cell distribution observed on the PCL/PCL-g-HAp. Overall, grafting of relatively short chains of PCL on the surface of HAp nano-particles stimulates fibroblasts adhesion and proliferation on the PCL/PCL-g-HAp nano-composite.

  9. Expression of leukocyte-endothelial cell adhesion molecules on monocyte adhesion to human endothelial cells on plasma treated PET and PTFE in vitro.

    PubMed

    Pu, F R; Williams, R L; Markkula, T K; Hunt, J A

    2002-12-01

    We used a coculture model to evaluate the inflammatory potential of ammonia gas plasma modified PET and PTFE by flow cytometry and immunohistochemistry. In these studies, human endothelial cells from umbilical cord (HUVEC) and promonocytic U937 cells were used. HUVECs grown on polystyrene tissue culture coverslips and HUVECs stimulated with tumour necrosis factor (TNF-alpha) were used as controls. U937 adhesion to endothelium on each surface was evaluated at day 1 and day 7. To further investigate the role of leukocyte-endothelial cell adhesion molecules (CAMs) in cell-to-cell interaction on material surfaces, the expression of the leukocyte-endothelial CAMs: ICAM-1, VCAM-1, PECAM-1, and E-selectin on HUVECs were evaluated after U937 cell adhesion. The results demonstrated that plasma treated PET (T-PET) and treated PTFE (T-PTFE) did not increase U937 cell adhesion compared to the negative control. Maximal adhesion of U937 cells to HUVEC was observed on TNF-alpha stimulated endothelium with significant differences between day 1 and day 7, which is consistent with our prior observation that T-PET and T-PTFE did not cause HUVECs to increase the expression of adhesion molecules. After U937 cell adhesion, the expression of ICAM-1 and VCAM-1 of HUVECs were not different on T-PET and T-PTFE compared with the negative control. However, the expression of E-selectin was reduced on day 1, but not on day 7. The effects of plasma treated PET and PTFE on HUVEC adhesion and proliferation were also studied. On day 1 there were slight increases in the growth of HUVECs on both of T-PET and T-PTFE but this was not statistically significant. On day 7, the cell number increased significantly on the surfaces compared to the negative control. The results demonstrate that the plasma treatment of PET and PTFE with ammonia improves the adhesion and growth of endothelial cells and these surfaces do not exhibit a direct inflammatory effect in terms of monocyte adhesion and expression of

  10. Bottom-up engineering of the surface roughness of nanostructured cubic zirconia to control cell adhesion.

    PubMed

    Singh, A V; Ferri, M; Tamplenizza, M; Borghi, F; Divitini, G; Ducati, C; Lenardi, C; Piazzoni, C; Merlini, M; Podestà, A; Milani, P

    2012-11-30

    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.

  11. PI3K{gamma} activation by CXCL12 regulates tumor cell adhesion and invasion

    SciTech Connect

    Monterrubio, Maria; Mellado, Mario; Carrera, Ana C.

    2009-10-16

    Tumor dissemination is a complex process, in which certain steps resemble those in leukocyte homing. Specific chemokine/chemokine receptor pairs have important roles in both processes. CXCL12/CXCR4 is the most commonly expressed chemokine/chemokine receptor pair in human cancers, in which it regulates cell adhesion, extravasation, metastatic colonization, angiogenesis, and proliferation. All of these processes require activation of signaling pathways that include G proteins, phosphatidylinositol-3 kinase (PI3K), JAK kinases, Rho GTPases, and focal adhesion-associated proteins. We analyzed these pathways in a human melanoma cell line in response to CXCL12 stimulation, and found that PI3K{gamma} regulates tumor cell adhesion through mechanisms different from those involved in cell invasion. Our data indicate that, following CXCR4 activation after CXCL12 binding, the invasion and adhesion processes are regulated differently by distinct downstream events in these signaling cascades.

  12. Force nanoscopy of cell mechanics and cell adhesion

    NASA Astrophysics Data System (ADS)

    Dufrêne, Yves F.; Pelling, Andrew E.

    2013-05-01

    Cells are constantly exposed to mechanical stimuli in their environment and have several evolved mechanisms to sense and respond to these cues. It is becoming increasingly recognized that many cell types, from bacteria to mammalian cells, possess a diverse set of proteins to translate mechanical cues into biochemical signalling and to mediate cell surface interactions such as cell adhesion. Moreover, the mechanical properties of cells are involved in regulating cell function as well as serving as indicators of disease states. Importantly, the recent development of biophysical tools and nanoscale methods has facilitated a deeper understanding of the role that physical forces play in modulating cell mechanics and cell adhesion. Here, we discuss how atomic force microscopy (AFM) has recently been used to investigate cell mechanics and cell adhesion at the single-cell and single-molecule levels. This knowledge is critical to our understanding of the molecular mechanisms that govern mechanosensing, mechanotransduction, and mechanoresponse in living cells. While pushing living cells with the AFM tip provides a means to quantify their mechanical properties and examine their response to nanoscale forces, pulling single surface proteins with a functionalized tip allows one to understand their role in sensing and adhesion. The combination of these nanoscale techniques with modern molecular biology approaches, genetic engineering and optical microscopies provides a powerful platform for understanding the sophisticated functions of the cell surface machinery, and its role in the onset and progression of complex diseases.

  13. Biotin-Avidin Based Universal Cell-Matrix Interaction for Promoting Three-Dimensional Cell Adhesion.

    PubMed

    Dou, Xiao-Qiu; Zhang, Jia; Feng, Chuanliang

    2015-09-23

    To promote cell adhesion in three-dimensional (3D) extracellular matrix (ECM) is crucial for avoiding cell anoikis, which is one of the most important issues for fundamental cell biology. Herein, a biotin-avidin based universal cell-matrix interaction for different types of cells is developed in order to achieve the promoted adhesion in 3D ECM. For the purpose, biotinylated nanofibrous hydrogels are constructed by coassembling 1,4-benzyldicarboxamide (C2) based non-biotinylated and biotinylated supramolecular gelators. The used cells are modified by avidin (AV-cells) through biotinylating cells and then interacting with avidin. After in situ encapsulating AV-cells in the hydrogels, the adhered amount can be increased by tens of percent even with adding several percentages of the biotinylated C2 gelators in the coassembly due to the specific biotin-avidin interaction. Reverse transcription polymerase chain reaction (RT-PCR) confirms that AV-cells can proliferate without varying gene expression and denaturation. Compared with the interaction between RGD and cells, this avidin-biotin interaction should be much more universal and it is feasible to be employed to promote cell adhesion for most types of cells in 3D matrix.

  14. Inhibition of Adhesion Molecule Gene Expression and Cell Adhesion by the Metabolic Regulator PGC-1α.

    PubMed

    Minsky, Neri; Roeder, Robert G

    2016-01-01

    Cell adhesion plays an important role in determining cell shape and function in a variety of physiological and pathophysiological conditions. While links between metabolism and cell adhesion were previously suggested, the exact context and molecular details of such a cross-talk remain incompletely understood. Here we show that PGC-1α, a pivotal transcriptional co-activator of metabolic gene expression, acts to inhibit expression of cell adhesion genes. Using cell lines, primary cells and mice, we show that both endogenous and exogenous PGC-1α down-regulate expression of a variety of cell adhesion molecules. Furthermore, results obtained using mRNA stability measurements as well as intronic RNA expression are consistent with a transcriptional effect of PGC-1α on cell adhesion gene expression. Interestingly, the L2/L3 motifs of PGC-1α, necessary for nuclear hormone receptor activation, are only partly required for inhibition of several cell adhesion genes by PGC-1α. Finally, PGC-1α is able to modulate adhesion of primary fibroblasts and hepatic stellate cells to extracellular matrix proteins. Our results delineate a cross talk between a central pathway controlling metabolic regulation and cell adhesion, and identify PGC-1α as a molecular link between these two major cellular networks.

  15. Focal adhesion kinase maintains, but not increases the adhesion of dental pulp cells.

    PubMed

    Qian, Yuyan; Shao, Meiying; Zou, Wenlin; Wang, Linyan; Cheng, Ran; Hu, Tao

    2017-02-25

    Focal adhesion kinase (FAK) functions as a key enzyme in the integrin-mediated adhesion-signalling pathway. Here, we aimed to investigate the effects of FAK on adhesion of human dental pulp (HDP) cells. We transfected lentiviral vectors to silence or overexpress FAK in HDP cells ex vivo. Early cell adhesion, cell survival and focal contacts (FCs)-related proteins (FAK and paxillin) were examined. By using immunofluorescence, the formation of FCs and cytoskeleton was detected, respectively. We found that both adhesion and survival of HDP cells were suppressed by FAK inhibition. However, FAK overexpression slightly inhibited cell adhesion and exhibited no change in cell survival compared with the control. A thick rim of cytoskeleton accumulated and smaller dot-shaped FCs appeared in FAK knockdown cells. Phosphorylation of paxillin (p-paxillin) was inhibited in FAK knockdown cells, verifying that the adhesion was inhibited. Less cytoskeleton and elongated FCs were observed in FAK-overexpressed cells. However, p-paxillin had no significant difference compared with the control. In conclusion, the data suggest that FAK maintains cell adhesion, survival and cytoskeleton formation, but excessive FAK has no positive effects on these aspects.

  16. Inhibition of Adhesion Molecule Gene Expression and Cell Adhesion by the Metabolic Regulator PGC-1α

    PubMed Central

    Minsky, Neri; Roeder, Robert G.

    2016-01-01

    Cell adhesion plays an important role in determining cell shape and function in a variety of physiological and pathophysiological conditions. While links between metabolism and cell adhesion were previously suggested, the exact context and molecular details of such a cross-talk remain incompletely understood. Here we show that PGC-1α, a pivotal transcriptional co-activator of metabolic gene expression, acts to inhibit expression of cell adhesion genes. Using cell lines, primary cells and mice, we show that both endogenous and exogenous PGC-1α down-regulate expression of a variety of cell adhesion molecules. Furthermore, results obtained using mRNA stability measurements as well as intronic RNA expression are consistent with a transcriptional effect of PGC-1α on cell adhesion gene expression. Interestingly, the L2/L3 motifs of PGC-1α, necessary for nuclear hormone receptor activation, are only partly required for inhibition of several cell adhesion genes by PGC-1α. Finally, PGC-1α is able to modulate adhesion of primary fibroblasts and hepatic stellate cells to extracellular matrix proteins. Our results delineate a cross talk between a central pathway controlling metabolic regulation and cell adhesion, and identify PGC-1α as a molecular link between these two major cellular networks. PMID:27984584

  17. The evaluation of p,p'-DDT exposure on cell adhesion of hepatocellular carcinoma.

    PubMed

    Jin, Xiaoting; Chen, Meilan; Song, Li; Li, Hanqing; Li, Zhuoyu

    2014-08-01

    Many studies have found a positive association between the progression of hepatocellular carcinoma and DDT exposure. These studies mainly focus on the effect of DDT exposure on cell proliferation and epithelial to mesenchymal transition (EMT) promotion. However, the influence of DDT on cell adhesion of hepatocellular carcinoma remains to be unclear. The aim of our study was to determine the effect of p,p'-DDT on cell adhesion of hepatocellular carcinoma in vitro and in vivo. The data showed that p,p'-DDT, exposing HepG2 cells for 6 days, decreased cell-cell adhesion and elevated cell-matrix adhesion. Strikingly, p,p'-DDT increased reactive oxygen species (ROS) content, and this was accompanied by the activation of JAK/STAT3 pathway. Moreover, ROS inhibitor supplement reversed these effects significantly. However, the addition of ER inhibitor, ICI, had no effect on the p,p'-DDT-induced effects. p,p'-DDT altered the mRNA levels of related adhesion molecules, including inhibition of E-cadherin and promotion of N-cadherin along with CD29. Interestingly, the p,p'-DDT-altered adhesion molecules could be reversed with JAK inhibitor or STAT3 inhibitor. Likewise, p,p'-DDT stimulated the JAK/STAT3 pathway in nude mice, as well as altered the mRNA levels of E-cadherin, N-cadherin, and CD29. Taken together, these results indicate that p,p'-DDT profoundly promotes the adhesion process by decreasing cell-cell adhesion and inducing cell-matrix adhesion via the ROS-mediated JAK/STAT3 pathway. All these events account for the carcinogenic potential of p,p'-DDT in liver.

  18. Discoidin domain receptor 2 (DDR2) regulates proliferation of endochondral cells in mice

    SciTech Connect

    Kawai, Ikuma; Hisaki, Tomoka; Sugiura, Koji; Naito, Kunihiko; Kano, Kiyoshi

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase. Black-Right-Pointing-Pointer DDR2 regulates cell proliferation, cell adhesion, migration, and extracellular matrix remodeling. Black-Right-Pointing-Pointer We produced in vitro and in vivo model to better understand the role of DDR2. Black-Right-Pointing-Pointer DDR2 might play an inhibitory role in the proliferation of chondrocyte. -- Abstract: Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase that is activated by fibrillar collagens. DDR2 regulates cell proliferation, cell adhesion, migration, and extracellular matrix remodeling. The decrement of endogenous DDR2 represses osteoblastic marker gene expression and osteogenic differentiation in murine preosteoblastic cells, but the functions of DDR2 in chondrogenic cellular proliferation remain unclear. To better understand the role of DDR2 signaling in cellular proliferation in endochondral ossification, we inhibited Ddr2 expression via the inhibitory effect of miRNA on Ddr2 mRNA (miDdr2) and analyzed the cellular proliferation and differentiation in the prechondrocyte ATDC5 cell lines. To investigate DDR2's molecular role in endochondral cellular proliferation in vivo, we also produced transgenic mice in which the expression of truncated, kinase dead (KD) DDR2 protein is induced, and evaluated the DDR2 function in cellular proliferation in chondrocytes. Although the miDdr2-transfected ATDC5 cell lines retained normal differentiation ability, DDR2 reduction finally promoted cellular proliferation in proportion to the decreasing ratio of Ddr2 expression, and it also promoted earlier differentiation to cartilage cells by insulin induction. The layer of hypertrophic chondrocytes in KD Ddr2 transgenic mice was not significantly thicker than that of normal littermates, but the layer of proliferative chondrocytes in KD-Ddr2 transgenic mice was significantly thicker than that of normal littermates

  19. Cell adhesion: integrating cytoskeletal dynamics and cellular tension

    PubMed Central

    Parsons, J. Thomas; Horwitz, Alan Rick; Schwartz, Martin A.

    2010-01-01

    Cell migration affects all morphogenetic processes and contributes to numerous diseases, including cancer and cardiovascular disease. For most cells in most environments, movement begins with protrusion of the cell membrane followed by the formation of new adhesions at the cell front that link the actin cytoskeleton to the substratum, generation of traction forces that move the cell forwards and disassembly of adhesions at the cell rear. Adhesion formation and disassembly drive the migration cycle by activating Rho GTPases, which in turn regulate actin polymerization and myosin II activity, and therefore adhesion dynamics. PMID:20729930

  20. Cationized bovine serum albumin with pendant RGD groups forms efficient biocoatings for cell adhesion.

    PubMed

    Ng, Jeck Fei; Weil, Tanja; Jaenicke, Stephan

    2011-11-01

    Cationized bovine serum albumin (cBSA-147) has been modified by attaching the cyclic pentapeptide cRGDfK to its surface through linkers of different length. Coatings of these bioconjugates on glass surfaces were studied for their ability to stimulate cell adhesion. These chemically modified albumins combine a high number of positive charges which facilitate the initial cell adhesion to the surface with multiple Arg-Gly-Asp groups which enable focal adhesion of fibroblast cells by specific interactions with cell-surface receptors. The biocoatings are easily prepared within a few minutes by simple incubation from a dilute solution of the modified albumin. This constitutes a convenient approach for preparing surfaces for cell adhesion. Excellent focal adhesion of NIH 3T3 fibroblast cells on the biocoatings was observed. About 75% of the seeded cells attached to the cRGDfK-cBSA-147 coated surfaces, and 97% of them underwent focal adhesion. Adhering cells were able to grow and proliferate on the coated surfaces, confirming the outstanding biocompatibility of these biocoatings.

  1. Tuning cell adhesive properties via layer-by-layer assembly of chitosan and alginate.

    PubMed

    Silva, Joana M; García, José R; Reis, Rui L; García, Andrés J; Mano, João F

    2017-03-15

    Understanding the mechanisms controlling cell-multilayer film interactions is crucial to the successful engineering of these coatings for biotechnological and biomedical applications. Herein, we present a strategy to tune the cell adhesive properties of multilayers based on marine polysaccharides with and without cross-linking and/or coating with extracellular matrix proteins. Chemical cross-linking of multilayers improved mechanical properties of the coatings but also elicited changes in surface chemistry that alter the adhesion of human umbilical vein endothelial cells. We evaluated a strategy to decouple the mechanical and chemical properties of these films, enabling the transition from cell-adhesive to cell-resistant multilayers. Addition of chitosan/alginate multilayers on top of cross-linked films decreased endothelial cell adhesion, spreading, and proliferation to similar levels as uncross-linked films. Our findings highlight the key role of surface chemistry in cell-multilayer film interactions, and these engineered nanocoatings represent a tunable model of cell adhesive and non-adhesive multilayered films.

  2. Angiotensin Converting Enzyme Regulates Cell Proliferation and Migration

    PubMed Central

    Carvalho, Clarissa Coelho; Florentino, Rodrigo Machado; França, Andressa; Matias, Eveline; Guimarães, Paola Bianchi; Batista, Carolina; Freire, Valder; Carmona, Adriana Karaoglanovic; Pesquero, João Bosco; de Paula, Ana Maria; Foureaux, Giselle; Leite, Maria de Fatima

    2016-01-01

    Background The angiotensin-I converting enzyme (ACE) plays a central role in the renin-angiotensin system, acting by converting the hormone angiotensin-I to the active peptide angiotensin-II (Ang-II). More recently, ACE was shown to act as a receptor for Ang-II, and its expression level was demonstrated to be higher in melanoma cells compared to their normal counterparts. However, the function that ACE plays as an Ang-II receptor in melanoma cells has not been defined yet. Aim Therefore, our aim was to examine the role of ACE in tumor cell proliferation and migration. Results We found that upon binding to ACE, Ang-II internalizes with a faster onset compared to the binding of Ang-II to its classical AT1 receptor. We also found that the complex Ang-II/ACE translocates to the nucleus, through a clathrin-mediated process, triggering a transient nuclear Ca2+ signal. In silico studies revealed a possible interaction site between ACE and phospholipase C (PLC), and experimental results in CHO cells, demonstrated that the β3 isoform of PLC is the one involved in the Ca2+ signals induced by Ang-II/ACE interaction. Further studies in melanoma cells (TM-5) showed that Ang-II induced cell proliferation through ACE activation, an event that could be inhibited either by ACE inhibitor (Lisinopril) or by the silencing of ACE. In addition, we found that stimulation of ACE by Ang-II caused the melanoma cells to migrate, at least in part due to decreased vinculin expression, a focal adhesion structural protein. Conclusion ACE activation regulates melanoma cell proliferation and migration. PMID:27992423

  3. Cell substratum adhesion during early development of Dictyostelium discoideum.

    PubMed

    Tarantola, Marco; Bae, Albert; Fuller, Danny; Bodenschatz, Eberhard; Rappel, Wouter-Jan; Loomis, William F

    2014-01-01

    Vegetative and developed amoebae of Dictyostelium discoideum gain traction and move rapidly on a wide range of substrata without forming focal adhesions. We used two independent assays to quantify cell-substrate adhesion in mutants and in wild-type cells as a function of development. Using a microfluidic device that generates a range of hydrodynamic shear stress, we found that substratum adhesion decreases at least 10 fold during the first 6 hr of development of wild type cells. This result was confirmed using a single-cell assay in which cells were attached to the cantilever of an atomic force probe and allowed to adhere to untreated glass surfaces before being retracted. Both of these assays showed that the decrease in substratum adhesion was dependent on the cAMP receptor CAR1 which triggers development. Vegetative cells missing talin as the result of a mutation in talA exhibited slightly reduced adhesive properties compared to vegetative wild-type cells. In sharp contrast to wild-type cells, however, these talA mutant cells did not show further reduction of adhesion during development such that after 5 hr of development they were significantly more adhesive than developed wild type cells. In addition, both assays showed that substrate adhesion was reduced in 0 hr cells when the actin cytoskeleton was disrupted by latrunculin. Consistent with previous observations, substrate adhesion was also reduced in 0 hr cells lacking the membrane proteins SadA or SibA as the result of mutations in sadA or sibA. However, there was no difference in the adhesion properties between wild type AX3 cells and these mutant cells after 6 hr of development, suggesting that neither SibA nor SadA play an essential role in substratum adhesion during aggregation. Our results provide a quantitative framework for further studies of cell substratum adhesion in Dictyostelium.

  4. Molecular markers of cell adhesion in ameloblastomas. An update

    PubMed Central

    González-González, Rogelio; Molina-Frechero, Nelly; Damian-Matsumura, Pablo

    2014-01-01

    Ameloblastoma is the most common odontogenic tumor of epithelial origin, and though it is of a benign nature, it frequently infiltrates the bone, has a high rate of recurrence and could potentially become malignant. Cellular adhesion potentially plays an important role in the manifestation of these characteristics and in the tumor biology of ameloblastomas. Losses of cell-cell and extracellular matrix adhesion and cohesion are among the first events that occur in the invasion and growth of tumors of epithelial origin. The present review includes a description of the molecules that are involved in cell adhesion as reported for various types of ameloblastomas and discusses the possible roles of these molecules in the biological behaviors of this odontogenic tumor. Knowledge of the complex mechanisms in which these molecules play a role is critical for the research and discovery of future therapeutic targets. Key words:Ameloblastoma, cellular adhesion, molecular markers, cell-cell adhesion, extracellular matrix-cell adhesion. PMID:23986011

  5. The effect of elastin on chondrocyte adhesion and proliferation on poly (ɛ-caprolactone)/elastin composites.

    PubMed

    Annabi, Nasim; Fathi, Ali; Mithieux, Suzanne M; Martens, Penny; Weiss, Anthony S; Dehghani, Fariba

    2011-02-01

    The aim of this study was to demonstrate the effect of elastin on chondrocyte adhesion and proliferation within the structure of poly (ɛ-caprolactone) (PCL)/elastin composites. The homogenous 3D structure composites were constructed by using high pressure CO(2) in two stages. Porous PCL structures with average pore sizes of 540 ± 21 μm and a high degree of interconnectivity were produced using gas foaming/salt leaching. The PCL scaffolds were then impregnated with elastin and cross-linked with glutaraldehyde (GA) under high pressure CO(2). The effects of elastin and cross-linker concentrations on the characteristics of composites were investigated. Increasing the elastin concentration from 25mg/ml to 100mg/ml elevated the amount of cross-linked elastin inside the macropores of PCL. Fourier transform infrared (FTIR) analysis showed that elastin was homogeneously distributed throughout the 3D structure of all composites. The weight gain of composites increased 2-fold from 15.8 ± 0.3 to 38.3 ± 0.7 (w/w) % by increasing the elastin concentration from 25mg/ml to 50mg/ml and approached a plateau above this concentration. The presence of elastin within the pores of PCL improved the water uptake properties of PCL scaffolds; the water uptake ratio of PCL was enhanced 100-fold from 0.030 ± 0.005g liquid/g polymer to 11.80 ± 0.01g liquid/g polymer, when the elastin solution concentration was 50mg/ml. These composites exhibited lower compressive modulus and energy loss compared to pure PCL scaffolds due to their higher water content and elasticity. In vitro studies show that these composites can support primary articular cartilage chondrocyte adhesion and proliferation within the 3D structures. These results demonstrate the potential of using PCL/elastin composites for cartilage repair.

  6. Cell Adhesion in Epidermal Development and Barrier Formation

    PubMed Central

    Sumigray, Kaelyn D.; Lechler, Terry

    2015-01-01

    Cell–cell adhesions are necessary for structural integrity and barrier formation of the epidermis. Here, we discuss insights from genetic and cell biological studies into the roles of individual cell–cell junctions and their composite proteins in regulating epidermal development and function. In addition to individual adhesive functions, we will discuss emerging ideas on mechanosensation/transduction of junctions in the epidermis, noncanonical roles for adhesion proteins, and crosstalk/interdependencies between the junctional systems. These studies have revealed that cell adhesion proteins are connected to many aspects of tissue physiology including growth control, differentiation, and inflammation. PMID:25733147

  7. Nanomechanical measurement of adhesion and migration of leukemia cells with phorbol 12-myristate 13-acetate treatment.

    PubMed

    Zhou, Zhuo Long; Ma, Jing; Tong, Ming-Hui; Chan, Barbara Pui; Wong, Alice Sze Tsai; Ngan, Alfonso Hing Wan

    The adhesion and traction behavior of leukemia cells in their microenvironment is directly linked to their migration, which is a prime issue affecting the release of cancer cells from the bone marrow and hence metastasis. In assessing the effectiveness of phorbol 12-myristate 13-acetate (PMA) treatment, the conventional batch-cell transwell-migration assay may not indicate the intrinsic effect of the treatment on migration, since the treatment may also affect other cellular behavior, such as proliferation or death. In this study, the pN-level adhesion and traction forces between single leukemia cells and their microenvironment were directly measured using optical tweezers and traction-force microscopy. The effects of PMA on K562 and THP1 leukemia cells were studied, and the results showed that PMA treatment significantly increased cell adhesion with extracellular matrix proteins, bone marrow stromal cells, and human fibroblasts. PMA treatment also significantly increased the traction of THP1 cells on bovine serum albumin proteins, although the effect on K562 cells was insignificant. Western blots showed an increased expression of E-cadherin and vimentin proteins after the leukemia cells were treated with PMA. The study suggests that PMA upregulates adhesion and thus suppresses the migration of both K562 and THP1 cells in their microenvironment. The ability of optical tweezers and traction-force microscopy to measure directly pN-level cell-protein or cell-cell contact was also demonstrated.

  8. Cell adhesion molecules: detection with univalent second antibody

    PubMed Central

    1980-01-01

    Identification of cell surface molecules that play a role in cell-cell adhesion (here called cell adhesion molecules) has been achieved by demonstrating the inhibitory effect of univalent antibodies that bind these molecules in an in vitro assay of cell-cell adhesion. A more convenient reagent, intact (divalent) antibody, has been avoided because it might agglutinate the cells rather than blocking cell-cell adhesion. In this report, we show that intact rabbit immunoglobulin directed against certain cell surface molecules of Dictyostelium discoideum blocks cell-cell adhesion when the in vitro assay is performed in the presence of univalent goat anti-rabbit antibody. Under appropriate experimental conditions, the univalent second antibody blocks agglutination induced by the rabbit antibody without significantly interfering with its effect on cell-cell adhesion. This method promises to be useful for screening monoclonal antibodies raised against potential cell adhesion molecules because: (a) it allows for the screening of large numbers of antibody samples without preparation of univalent fragments; and (b) it requires much less antibody because of the greater affinity of divalent antibodies for antigens. PMID:6970200

  9. Effect of molecular weight and concentration of hyaluronan on cell proliferation and osteogenic differentiation in vitro

    SciTech Connect

    Zhao, Ningbo Wang, Xin Qin, Lei Guo, Zhengze Li, Dehua

    2015-09-25

    Hyaluronan (HA), the simplest glycosaminoglycan and a major component of the extracellular matrix, exists in various tissues. It is involved in some critical biological procedures, including cellular signaling, cell adhesion and proliferation, and cell differentiation. The effect of molecular weight (MW) and concentration of HA on cell proliferation and differentiation was controversial. In this study, we investigated the effect of MW and concentration of HA on the proliferation and osteogenic differentiation of rabbit bone marrow-derived stem cells in vitro. Results showed that high MW HA decreased the cell adhesion rate in a concentration-dependant manner. The cell adhesion rate was decreased by increasing MW of HA. Cell proliferation was significantly enhanced by low MW HA (P < 0.05). The factorial analysis indicated that MW and concentration had an interactive effect on the cell adhesion rate and cell proliferation (P < 0.05). High MW HA increased the mRNA expressions of ALP, RUNX-2 and OCN. The higher the MW was, the higher the mRNA expressions were. The factorial analysis indicated that MW and concentration had an interactive effect on ALP mRNA expression (P < 0.05). HA of higher MW and higher concentration promoted bone formation. These findings provide some useful information in understanding the mechanism underlying the effect of MW and concentration of HA on cell proliferation and differentiation. - Highlights: • Effect of hyaluronan on cell proliferation and differentiation is evaluated in vitro. • Hyaluronan of low molecular weight increases cell proliferation. • Hyaluronan of high molecular weight promotes cell osteogenic differentiation. • Molecular weight and concentration of hyaluronan show interactive effect.

  10. RP1 is a phosphorylation target of CK2 and is involved in cell adhesion.

    PubMed

    Stenner, Frank; Liewen, Heike; Göttig, Stephan; Henschler, Reinhard; Markuly, Norbert; Kleber, Sascha; Faust, Michael; Mischo, Axel; Bauer, Stefan; Zweifel, Martin; Knuth, Alexander; Renner, Christoph; Wadle, Andreas

    2013-01-01

    RP1 (synonym: MAPRE2, EB2) is a member of the microtubule binding EB1 protein family, which interacts with APC, a key regulatory molecule in the Wnt signalling pathway. While the other EB1 proteins are well characterized the cellular function and regulation of RP1 remain speculative to date. However, recently RP1 has been implicated in pancreatic cancerogenesis. CK2 is a pleiotropic kinase involved in adhesion, proliferation and anti-apoptosis. Overexpression of protein kinase CK2 is a hallmark of many cancers and supports the malignant phenotype of tumor cells. In this study we investigate the interaction of protein kinase CK2 with RP1 and demonstrate that CK2 phosphorylates RP1 at Ser(236) in vitro. Stable RP1 expression in cell lines leads to a significant cleavage and down-regulation of N-cadherin and impaired adhesion. Cells expressing a Phospho-mimicking point mutant RP1-ASP(236) show a marked decrease of adhesion to endothelial cells under shear stress. Inversely, we found that the cells under shear stress downregulate endogenous RP1, most likely to improve cellular adhesion. Accordingly, when RP1 expression is suppressed by shRNA, cells lacking RP1 display significantly increased cell adherence to surfaces. In summary, RP1 phosphorylation at Ser(236) by CK2 seems to play a significant role in cell adhesion and might initiate new insights in the CK2 and EB1 family protein association.

  11. RP1 Is a Phosphorylation Target of CK2 and Is Involved in Cell Adhesion

    PubMed Central

    Göttig, Stephan; Henschler, Reinhard; Markuly, Norbert; Kleber, Sascha; Faust, Michael; Mischo, Axel; Bauer, Stefan; Zweifel, Martin; Knuth, Alexander; Renner, Christoph; Wadle, Andreas

    2013-01-01

    RP1 (synonym: MAPRE2, EB2) is a member of the microtubule binding EB1 protein family, which interacts with APC, a key regulatory molecule in the Wnt signalling pathway. While the other EB1 proteins are well characterized the cellular function and regulation of RP1 remain speculative to date. However, recently RP1 has been implicated in pancreatic cancerogenesis. CK2 is a pleiotropic kinase involved in adhesion, proliferation and anti-apoptosis. Overexpression of protein kinase CK2 is a hallmark of many cancers and supports the malignant phenotype of tumor cells. In this study we investigate the interaction of protein kinase CK2 with RP1 and demonstrate that CK2 phosphorylates RP1 at Ser236 in vitro. Stable RP1 expression in cell lines leads to a significant cleavage and down-regulation of N-cadherin and impaired adhesion. Cells expressing a Phospho-mimicking point mutant RP1-ASP236 show a marked decrease of adhesion to endothelial cells under shear stress. Inversely, we found that the cells under shear stress downregulate endogenous RP1, most likely to improve cellular adhesion. Accordingly, when RP1 expression is suppressed by shRNA, cells lacking RP1 display significantly increased cell adherence to surfaces. In summary, RP1 phosphorylation at Ser236 by CK2 seems to play a significant role in cell adhesion and might initiate new insights in the CK2 and EB1 family protein association. PMID:23844040

  12. Effect of channel geometry on cell adhesion in microfluidic devices.

    PubMed

    Green, James V; Kniazeva, Tatiana; Abedi, Mehdi; Sokhey, Darshan S; Taslim, Mohammad E; Murthy, Shashi K

    2009-03-07

    Microfluidic channels coated with ligands are a versatile platform for the separation or enrichment of cells from small sample volumes. This adhesion-based mode of separation is mediated by ligand-receptor bonds between the cells and channel surface and also by fluid shear stress. This paper demonstrates how aspects of microchannel geometry can play an additional role in controlling cell adhesion. With a combination of computational fluid dynamics modeling and cell adhesion experiments, channels with sharp turns are shown to have regions with near-zero velocity at the turn regions where large numbers of cells adhere or become collected. The lack of uniform adhesion in the turn regions compared to other regions of these channels, together with the large variability in observed cell adhesion indicates that channels with sharp turns are not optimal for cell-capture applications where predictable cell adhesion is desired. Channels with curved turns, on the other hand are shown to provide more uniform and predictable cell adhesion provided the gap between parallel arms of the channels is sufficiently wide. The magnitude of cell adhesion in these curved channels is comparable to that in straight channels with no turns.

  13. Electrical field effects on endothelial cell adhesion and growth on conducting biomaterials surfaces

    NASA Astrophysics Data System (ADS)

    Clark, Gwen Elaine

    A major problem for vascular graft implants is poor long-term patency for small-diameter (<6 mm) prostheses. Small-diameter woven Dacron RTM or expanded polytetrafluoroethylene (e-PTFE) grafts often occlude in a short time due to thrombus or polytetrafluoroethylene intimal hyperplasia. It has generally been considered that an endothelial cell lining of such grafts might reduce thrombogenicity and thereby produce a more biomimetic prosthesis. Electrical stimulation has been studied for effects on in vitro cell growth, motility, and adhesion characteristics, as well as for in vivo wound healing. A comprehensive literature review was conducted which suggested the need for further research concerning the effects of electrical fields on endothelial cell adhesion and growth properties. The focus of these studies was therefore to determine the effect of electrical fields on the proliferation and adhesion characteristics of endothelial cells cultured on various substrates using low-voltage direct current. Voltages of 0, 0.5, and 1 volt were used for in vitro endothelial cell cultures plated at 50,000 and 100,000 cells/mL. Growth experiments were performed on glass, MylarRTM, Indium Tin Oxide (ITO)-glass, on ITO-, carbon-, and gold-palladium-coated MylarRTM, and on polypyrrole-coated DacronRTM. Proliferation of endothelial cells was determined at 12, 24, and 36 hours. Adhesion characteristics were measured using a novel flow adhesion system. Characterization was via cell staining in conjunction with optical microscopy and a 6-keto-prostaglandin-F 1alpha assay to measure cell viability. Results of these cell growth studies in electric fields indicate that low voltage stimulation moderately increased endothelial cell growth on most substrates. The release of 6-keto-prostaglandin-F1alpha decreased over time at most cell concentrations and voltage levels. Cell adhesion experiments provided contrary results to the growth studies and suggested Rested that low-voltage electric

  14. The Stochastic Theory of Cell Proliferation

    PubMed Central

    Bronk, Burt V.; Dienes, G. J.; Paskin, Arthur

    1968-01-01

    A stochastic theory of cell kinetics has been developed based on a realistic model of cell proliferation. A characteristic transit time, t̄i, has been assigned to each of the four states (G1, S, G2, M) of the cell cycle. The actual transit time, ti, for any cell is represented by a distribution around t̄i with a variance σi2. Analytic and computer formulations have been used to describe the time development of such characteristics as age distribution, labeling experiments, and response to perturbations of the system by, for example, irradiation and temperature. The decay of synchrony is analyzed in detail and is shown to proceed as a damped wave. From the first few peaks of the synchrony decay one can obtain the distribution function for the cell cycle time. The later peaks decay exponentially with a characteristic decay constant, λ, which depends only on the average cell-cycle time, T̄, and the associated variance. It is shown that the system, upon any sudden disturbance, approaches new “equilibrium” proliferation characteristics via damped periodic transients, the damping being characterized by λ. Thus, the response time of the system, T̄/λ, is as basic a parameter of the system as the cell-cycle time. PMID:5696217

  15. Mitochondrial Regulation of Cell Cycle and Proliferation

    PubMed Central

    Antico Arciuch, Valeria Gabriela; Elguero, María Eugenia; Poderoso, Juan José

    2012-01-01

    Abstract Eukaryotic mitochondria resulted from symbiotic incorporation of α-proteobacteria into ancient archaea species. During evolution, mitochondria lost most of the prokaryotic bacterial genes and only conserved a small fraction including those encoding 13 proteins of the respiratory chain. In this process, many functions were transferred to the host cells, but mitochondria gained a central role in the regulation of cell proliferation and apoptosis, and in the modulation of metabolism; accordingly, defective organelles contribute to cell transformation and cancer, diabetes, and neurodegenerative diseases. Most cell and transcriptional effects of mitochondria depend on the modulation of respiratory rate and on the production of hydrogen peroxide released into the cytosol. The mitochondrial oxidative rate has to remain depressed for cell proliferation; even in the presence of O2, energy is preferentially obtained from increased glycolysis (Warburg effect). In response to stress signals, traffic of pro- and antiapoptotic mitochondrial proteins in the intermembrane space (B-cell lymphoma-extra large, Bcl-2-associated death promoter, Bcl-2 associated X-protein and cytochrome c) is modulated by the redox condition determined by mitochondrial O2 utilization and mitochondrial nitric oxide metabolism. In this article, we highlight the traffic of the different canonical signaling pathways to mitochondria and the contributions of organelles to redox regulation of kinases. Finally, we analyze the dynamics of the mitochondrial population in cell cycle and apoptosis. Antioxid. Redox Signal. 16, 1150–1180. PMID:21967640

  16. Cell proliferation inhibition in reduced gravity

    NASA Technical Reports Server (NTRS)

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

    1994-01-01

    Extended durations of spaceflight have been shown to be deleterious on an organismic level; however, mechanisms underlying cellular sensitivity to the gravitational environment remain to be elucidated. The majority of the gravitational studies to date indicates that cell regulatory pathways may be influenced by their gravitational environment. Still, few cell biology experiments have been performed in space flight and even fewer experiments have been repeated on subsequent flights. With flight opportunities on STS-50, 54, and 57, Sf9 cells were flown in the BioServe Fluids Processing Apparatus and cell proliferation was measured with and without exposure to a cell regulatory sialoglycopeptide (CeReS) inhibitor. Results from these flights indicate that the Sf9 cells grew comparable to ground controls, that the CeReS inhibitor bound to its specific receptor, and that its signal transduction cascade was not gravity sensitive.

  17. FOXL2 suppresses proliferation, invasion and promotes apoptosis of cervical cancer cells

    PubMed Central

    Liu, Xing-Long; Meng, Yu-Han; Wang, Jian-Li; Yang, Biao-Bing; Zhang, Fan; Tang, Sheng-Jian

    2014-01-01

    FOXL2 is a transcription factor that is essential for ovarian function and maintenance, the germline mutations of which give rise to the blepharophimosis ptosis epicanthus inversus syndrome (BPES), often associated with premature ovarian failure. Recently, its mutations have been found in ovarian granulosa cell tumors (OGCTs). In this study, we measured the expression of FOXL2 in cervical cancer by immunohistochemistry and its mRNA level in cervical cancer cell lines Hela and Siha by RT-PCR. Then we overexpressed FOXL2 in Hela cells and silenced it in Siha cells by plasmid transfection and verified using western blotting. When FOXL2 was overexpressed or silenced, cells proliferation and apoptosis were determined by Brdu assay and Annexin V/PI detection kit, respectively. In addition, we investigated the effects of FOXL2 on the adhesion and invasion of Hela and Siha cells. Finally, we analyzed the influences of FOXL2 on Ki67, PCNA and FasL by flow cytometry. The results showed that FOXL2 was highly expressed in cervical squamous cancer. Overexpressing FOXL2 suppressed Hela proliferation and facilitated its apoptosis. Silencing FOXL2 enhanced Siha proliferation and inhibited its apoptosis. Meanwhile, silencing FOXL2 promoted Siha invasion, but it had no effect on cells adhesion. In addition, overexpressing FOXL2 decreased the expression of Ki67 in Hela and Siha cells. Therefore, our results suggested that FOXL2 restrained cells proliferation and enhanced cells apoptosis mainly through decreasing Ki67 expression. PMID:24817949

  18. Adhesion of Actinobacillus actinomycetemcomitans to a human oral cell line.

    PubMed Central

    Mintz, K P; Fives-Taylor, P M

    1994-01-01

    Two quantitative, rapid assays were developed to study the adhesion of Actinobacillus actinomycetemcomitans, an oral bacterium associated with periodontal disease, to human epithelial cells. The human oral carcinoma cell line KB was grown in microtiter plates, and adherent bacteria were detected by an enzyme-linked immunosorbent assay with purified anti-A. actinomycetemcomitans serum and horseradish peroxidase-conjugated secondary antibody or [3H]thymidine-labeled bacteria. Adhesion was found to be time dependent and increased linearly with increasing numbers of bacteria added. Variation in the level of adhesion was noted among strains of A. actinomycetemcomitans. Adhesion was not significantly altered by changes in pH (from pH 5 to 9) but was sensitive to sodium chloride concentrations greater than 0.15 M. Pooled human saliva was inhibitory for adhesion when bacteria were pretreated with saliva before being added to the cells. Pretreatment of the KB cells with saliva did not inhibit adhesion. Protease treatment of A. actinomycetemcomitans reduced adhesion of the bacteria to KB cells. The data are consistent with the hypothesis that a protein(s) is required for bacterial adhesion and that host components may play a role in modulating adhesion to epithelial cells. Images PMID:8063383

  19. Nanomechanical measurement of adhesion and migration of leukemia cells with phorbol 12-myristate 13-acetate treatment

    PubMed Central

    Zhou, Zhuo Long; Ma, Jing; Tong, Ming-Hui; Chan, Barbara Pui; Wong, Alice Sze Tsai; Ngan, Alfonso Hing Wan

    2016-01-01

    The adhesion and traction behavior of leukemia cells in their microenvironment is directly linked to their migration, which is a prime issue affecting the release of cancer cells from the bone marrow and hence metastasis. In assessing the effectiveness of phorbol 12-myristate 13-acetate (PMA) treatment, the conventional batch-cell transwell-migration assay may not indicate the intrinsic effect of the treatment on migration, since the treatment may also affect other cellular behavior, such as proliferation or death. In this study, the pN-level adhesion and traction forces between single leukemia cells and their microenvironment were directly measured using optical tweezers and traction-force microscopy. The effects of PMA on K562 and THP1 leukemia cells were studied, and the results showed that PMA treatment significantly increased cell adhesion with extracellular matrix proteins, bone marrow stromal cells, and human fibroblasts. PMA treatment also significantly increased the traction of THP1 cells on bovine serum albumin proteins, although the effect on K562 cells was insignificant. Western blots showed an increased expression of E-cadherin and vimentin proteins after the leukemia cells were treated with PMA. The study suggests that PMA upregulates adhesion and thus suppresses the migration of both K562 and THP1 cells in their microenvironment. The ability of optical tweezers and traction-force microscopy to measure directly pN-level cell–protein or cell–cell contact was also demonstrated. PMID:27994457

  20. Cell Adhesion on Amyloid Fibrils Lacking Integrin Recognition Motif*

    PubMed Central

    Jacob, Reeba S.; George, Edna; Singh, Pradeep K.; Salot, Shimul; Anoop, Arunagiri; Jha, Narendra Nath; Sen, Shamik; Maji, Samir K.

    2016-01-01

    Amyloids are highly ordered, cross-β-sheet-rich protein/peptide aggregates associated with both human diseases and native functions. Given the well established ability of amyloids in interacting with cell membranes, we hypothesize that amyloids can serve as universal cell-adhesive substrates. Here, we show that, similar to the extracellular matrix protein collagen, amyloids of various proteins/peptides support attachment and spreading of cells via robust stimulation of integrin expression and formation of integrin-based focal adhesions. Additionally, amyloid fibrils are also capable of immobilizing non-adherent red blood cells through charge-based interactions. Together, our results indicate that both active and passive mechanisms contribute to adhesion on amyloid fibrils. The present data may delineate the functional aspect of cell adhesion on amyloids by various organisms and its involvement in human diseases. Our results also raise the exciting possibility that cell adhesivity might be a generic property of amyloids. PMID:26742841

  1. Patterned Poly(dopamine) Films for Enhanced Cell Adhesion.

    PubMed

    Chen, Xi; Cortez-Jugo, Christina; Choi, Gwan H; Björnmalm, Mattias; Dai, Yunlu; Yoo, Pil J; Caruso, Frank

    2017-01-18

    Engineered materials that promote cell adhesion and cell growth are important in tissue engineering and regenerative medicine. In this work, we produced poly(dopamine) (PDA) films with engineered patterns for improved cell adhesion. The patterned films were synthesized via the polymerization of dopamine at the air-water interface of a floating bed of spherical particles. Subsequent dissolution of the particles yielded free-standing PDA films with tunable geometrical patterns. Our results show that these patterned PDA films significantly enhance the adhesion of both cancer cells and stem cells, thus showing promise as substrates for cell attachment for various biomedical applications.

  2. TIEG1-null tenocytes display age-dependent differences in their gene expression, adhesion, spreading and proliferation properties

    SciTech Connect

    Haddad, Oualid; Gumez, Laurie; Hawse, John R.; Subramaniam, Malayannan; Spelsberg, Thomas C.; Bensamoun, Sabine F.

    2011-07-15

    The remodeling of extracellular matrix is a crucial mechanism in tendon development and the proliferation of fibroblasts is a key factor in this process. The purpose of this study was to further elucidate the role of TIEG1 in mediating important tenocyte properties throughout the aging process. Wildtype and TIEG1 knockout tenocytes adhesion, spreading and proliferation were characterized on different substrates (fibronectin, collagen type I, gelatin and laminin) and the expression levels of various genes known to be involved with tendon development were analyzed by RT-PCR. The experiments revealed age-dependent and substrate-dependent properties for both wildtype and TIEG1 knockout tenocytes. Taken together, our results indicate an important role for TIEG1 in regulating tenocytes adhesion, spreading, and proliferation throughout the aging process. Understanding the basic mechanisms of TIEG1 in tenocytes may provide valuable information for treating multiple tendon disorders.

  3. Intercellular adhesion molecule-1 expression by skeletal muscle cells augments myogenesis

    SciTech Connect

    Goh, Qingnian; Dearth, Christopher L.; Corbett, Jacob T.; Pierre, Philippe; Chadee, Deborah N.; Pizza, Francis X.

    2015-02-15

    We previously demonstrated that the expression of intercellular adhesion molecule-1 (ICAM-1) by skeletal muscle cells after muscle overload contributes to ensuing regenerative and hypertrophic processes in skeletal muscle. The objective of the present study is to reveal mechanisms through which skeletal muscle cell expression of ICAM-1 augments regenerative and hypertrophic processes of myogenesis. This was accomplished by genetically engineering C2C12 myoblasts to stably express ICAM-1, and by inhibiting the adhesive and signaling functions of ICAM-1 through the use of a neutralizing antibody or cell penetrating peptide, respectively. Expression of ICAM-1 by cultured skeletal muscle cells augmented myoblast–myoblast adhesion, myotube formation, myonuclear number, myotube alignment, myotube–myotube fusion, and myotube size without influencing the ability of myoblasts to proliferate or differentiate. ICAM-1 augmented myotube formation, myonuclear accretion, and myotube alignment through a mechanism involving adhesion-induced activation of ICAM-1 signaling, as these dependent measures were reduced via antibody and peptide inhibition of ICAM-1. The adhesive and signaling functions of ICAM-1 also facilitated myotube hypertrophy through a mechanism involving myotube–myotube fusion, protein synthesis, and Akt/p70s6k signaling. Our findings demonstrate that ICAM-1 expression by skeletal muscle cells augments myogenesis, and establish a novel mechanism through which the inflammatory response facilitates growth processes in skeletal muscle. - Highlights: • We examined mechanisms through which skeletal muscle cell expression of ICAM-1 facilitates events of in vitro myogenesis. • Expression of ICAM-1 by cultured myoblasts did not influence their ability to proliferate or differentiate. • Skeletal muscle cell expression of ICAM-1 augmented myoblast fusion, myotube alignment, myotube–myotube fusion, and myotube size. • ICAM-1 augmented myogenic processes through

  4. Short Peptides Enhance Single Cell Adhesion and Viability on Microarrays

    PubMed Central

    Veiseh, Mandana; Veiseh, Omid; Martin, Michael C.; Asphahani, Fareid; Zhang, Miqin

    2011-01-01

    Single cell patterning holds important implications for biology, biochemistry, biotechnology, medicine, and bioinformatics. The challenge for single cell patterning is to produce small islands hosting only single cells and retaining their viability for a prolonged period of time. This study demonstrated a surface engineering approach that uses a covalently-bound short peptide as a mediator to pattern cells with improved single cell adhesion and prolonged cellular viability on gold patterned SiO2 substrates. The underlying hypothesis is that cell adhesion is regulated by the type, availability and stability of effective cell adhesion peptides, and thus covalently bound short peptides would promote cell spreading and thus, single cell adhesion and viability. The effectiveness of this approach and the underlying mechanism for the increased probability of single cell adhesion and prolonged cell viability by short peptides were studied by comparing cellular behavior of human umbilical cord vein endothelial cells on three model surfaces whose gold electrodes were immobilized with fibronectin, physically adsorbed Arg-Glu-Asp-Val-Tyr, and covalently-bound Lys-Arg-Glu-Asp-Val-Tyr, respectively. The surface chemistry and binding properties were characterized by reflectance Fourier transform infrared spectroscopy. Both short peptides were superior to fibronectin in producing adhesion of only single cells, while the covalently bound peptide also reduced apoptosis and necrosis of adhered cells. Controlling cell spreading by peptide binding domains to regulate apoptosis and viability represents a fundamental mechanism in cell-materials interaction and provides an effective strategy in engineering arrays of single cells. PMID:17371055

  5. Biofilms’ Role in Planktonic Cell Proliferation

    PubMed Central

    Bester, Elanna; Wolfaardt, Gideon M.; Aznaveh, Nahid B.; Greener, Jesse

    2013-01-01

    The detachment of single cells from biofilms is an intrinsic part of this surface-associated mode of bacterial existence. Pseudomonas sp. strain CT07gfp biofilms, cultivated in microfluidic channels under continuous flow conditions, were subjected to a range of liquid shear stresses (9.42 mPa to 320 mPa). The number of detached planktonic cells was quantified from the effluent at 24-h intervals, while average biofilm thickness and biofilm surface area were determined by confocal laser scanning microscopy and image analysis. Biofilm accumulation proceeded at the highest applied shear stress, while similar rates of planktonic cell detachment was maintained for biofilms of the same age subjected to the range of average shear rates. The conventional view of liquid-mediated shear leading to the passive erosion of single cells from the biofilm surface, disregards the active contribution of attached cell metabolism and growth to the observed detachment rates. As a complement to the conventional conceptual biofilm models, the existence of a biofilm surface-associated zone of planktonic cell proliferation is proposed to highlight the need to expand the traditional perception of biofilms as promoting microbial survival, to include the potential of biofilms to contribute to microbial proliferation. PMID:24201127

  6. The effects of cell adhesion on the growth and protein productivity of animal cells.

    PubMed

    Nishijima, K; Fujiki, T; Kojima, H; Iijima, S

    2000-07-01

    We investigated the effect of cell adhesion on cellgrowth and productivity of recombinant protein inChinese hamster ovary (CHO) cells. Cells cultured innormal tissue culture dishes attached to the dishsurfaces and grew as a monolayer, while cells culturedin non-treated dishes proliferated in suspension assingle cells without adhering to the dish surfaces. On an agarose-coated dish surface, cell aggregatesformed without attaching to the dish. Growth rates inboth suspension cultures were slightly lower thanthose in monolayer culture. Cell cycle analysisindicated that the duration of the G(1) phase insuspension cultures was longer than that in monolayerculture, suggesting that attachment to the substratummainly affected the transition from the G(1) to theS phase. Consistent with this, CDK inhibitor p27,that inhibits the G(1)S transition, was induced inthe cells cultured in suspension.To assess the productivity of recombinant proteins,CHO cells were transfected with a plasmid containingmurine interferon gamma (mIFN-gamma) under thecontrol of the cytomegalovirus promoter. Insuspension culture, mIFN-gamma productivity wasslightly lower than that in the monolayer culture. When protein kinase C was activated by phorbol ester,mIFN-gamma production was enhanced in both themonolayer and suspension cultures. However, theproductivity in the suspension culture was lower thanthat in the adherent culture even in the presence ofhigh concentrations of phorbol ester. These resultssuggested that cell adhesion to the substratum affectsvarious features of CHO cells.

  7. Myoferlin depletion elevates focal adhesion kinase and paxillin phosphorylation and enhances cell-matrix adhesion in breast cancer cells.

    PubMed

    Blackstone, B N; Li, R; Ackerman, W E; Ghadiali, S N; Powell, H M; Kniss, D A

    2015-04-15

    Breast cancer is the second leading cause of malignant death among women. A crucial feature of metastatic cancers is their propensity to lose adhesion to the underlying basement membrane as they transition to a motile phenotype and invade surrounding tissue. Attachment to the extracellular matrix is mediated by a complex of adhesion proteins, including integrins, signaling molecules, actin and actin-binding proteins, and scaffolding proteins. Focal adhesion kinase (FAK) is pivotal for the organization of focal contacts and maturation into focal adhesions, and disruption of this process is a hallmark of early cancer invasive potential. Our recent work has revealed that myoferlin (MYOF) mediates breast tumor cell motility and invasive phenotype. In this study we demonstrate that noninvasive breast cancer cell lines exhibit increased cell-substrate adhesion and that silencing of MYOF using RNAi in the highly invasive human breast cancer cell line MDA-MB-231 also enhances cell-substrate adhesion. In addition, we detected elevated tyrosine phosphorylation of FAK (FAK(Y397)) and paxillin (PAX(Y118)), markers of focal adhesion protein activation. Morphometric analysis of PAX expression revealed that RNAi-mediated depletion of MYOF resulted in larger, more elongated focal adhesions, in contrast to cells transduced with a control virus (MDA-231(LVC) cells), which exhibited smaller focal contacts. Finally, MYOF silencing in MDA-MB-231 cells exhibited a more elaborate ventral cytoskeletal structure near focal adhesions, typified by pronounced actin stress fibers. These data support the hypothesis that MYOF regulates cell adhesions and cell-substrate adhesion strength and may account for the high degree of motility in invasive breast cancer cells.

  8. Adhesions

    MedlinePlus

    Adhesions are bands of scar-like tissue. Normally, internal tissues and organs have slippery surfaces so they can shift easily as the body moves. Adhesions cause tissues and organs to stick together. They ...

  9. Adhesion

    MedlinePlus

    ... the intestines, adhesions can cause partial or complete bowel obstruction . Adhesions inside the uterine cavity, called Asherman syndrome , ... 1. Read More Appendicitis Asherman syndrome Glaucoma Infertility Intestinal obstruction Review Date 4/5/2016 Updated by: Irina ...

  10. Endothelial cells regulate the proliferation of monocytes in vitro.

    PubMed

    Pakala, R; Benedict, C R

    1999-11-01

    Monocytes (MPhis) are among the first cells to accumulate in early atherosclerotic lesions and generally are believed to be incapable of proliferation. However, recent studies indicate that the number of MPhis in atherosclerotic lesion may increase due to induction of local proliferation. Since proliferation of hematopoietic lineage cells is strongly influenced by interaction with neighboring cell types, we examined the ability of vascular endothelial cells (EC), smooth muscle cells or fibroblasts to stimulate MPhi proliferation. In this study, we show that only when seeded at high densities MPhis could proliferate in culture. However, when contact co-cultured with EC, MPhis proliferated at a higher rate (260% on day 6) than those cultured alone or co-cultured with smooth muscle cells or fibroblasts. Endothelial cells could stimulate the proliferation of MPhis even at non-proliferating densities. Only EC that were growth arrested or in lag phase could induce MPhi proliferation, whereas those in the exponential proliferating phase were non-stimulatory. Conditioned medium prepared from EC in growth arrested or lag phase failed to stimulate MPhi proliferation. Similarly physical separation of MPhis from EC also resulted in no proliferation. These results suggest that EC induced MPhi proliferation is contact dependent and no soluble factors are involved in this induction. This EC induced MPhi proliferation may have a profound effect on the rate of progression of atherosclerosis.

  11. Protein conformation as a regulator of cell-matrix adhesion.

    PubMed

    Hytönen, Vesa P; Wehrle-Haller, Bernhard

    2014-04-14

    The dynamic regulation of cell-matrix adhesion is essential for tissue homeostasis and architecture, and thus numerous pathologies are linked to altered cell-extracellular matrix (ECM) interaction and ECM scaffold. The molecular machinery involved in cell-matrix adhesion is complex and involves both sensory and matrix-remodelling functions. In this review, we focus on how protein conformation controls the organization and dynamics of cell-matrix adhesion. The conformational changes in various adhesion machinery components are described, including examples from ECM as well as cytoplasmic proteins. The discussed mechanisms involved in the regulation of protein conformation include mechanical stress, post-translational modifications and allosteric ligand-binding. We emphasize the potential role of intrinsically disordered protein regions in these processes and discuss the role of protein networks and co-operative protein interactions in the formation and consolidation of cell-matrix adhesion and extracellular scaffolds.

  12. Ultraweak sugar-sugar interactions for transient cell adhesion.

    PubMed Central

    Pincet, F; Le Bouar, T; Zhang, Y; Esnault, J; Mallet, J M; Perez, E; Sinaÿ, P

    2001-01-01

    Carbohydrate-carbohydrate interactions are rarely considered in biologically relevant situations such as cell recognition and adhesion. One Ca(2+)-mediated homotypic interaction between two Lewis(x) determinants (Le(x)) has been proposed to drive cell adhesion in murine embryogenesis. Here, we confirm the existence of this specific interaction by reporting the first direct quantitative measurements in an environment akin to that provided by membranes. The adhesion between giant vesicles functionalized with Le(x) was obtained by micropipette aspiration and contact angle measurements. This interaction is below the thermal energy, and cell-cell adhesion will require a large number of molecules, as illustrated by the Le(x) concentration peak observed at the cell membranes during the morula stage of the embryo. This adhesion is ultralow and therefore difficult to measure. Such small interactions explain why the concept of specific interactions between carbohydrates is often neglected. PMID:11222296

  13. Effects of curvature and cell-cell interaction on cell adhesion in microvessels.

    PubMed

    Yan, W W; Liu, Y; Fu, B M

    2010-10-01

    It has been found that both circulating blood cells and tumor cells are more easily adherent to curved microvessels than straight ones. This motivated us to investigate numerically the effect of the curvature of the curved vessel on cell adhesion. In this study, the fluid dynamics was carried out by the lattice Boltzmann method (LBM), and the cell dynamics was governed by the Newton's law of translation and rotation. The adhesive dynamics model involved the effect of receptor-ligand bonds between circulating cells and endothelial cells (ECs). It is found that the curved vessel would increase the simultaneous bond number, and the probability of cell adhesion is increased consequently. The interaction between traveling cells would also affect the cell adhesion significantly. For two-cell case, the simultaneous bond number of the rear cell is increased significantly, and the curvature of microvessel further enhances the probability of cell adhesion.

  14. Loss of the desmosomal cadherin desmoglein-2 suppresses colon cancer cell proliferation through EGFR signaling

    PubMed Central

    Kamekura, R; Kolegraff, KN; Nava, P; Hilgarth, RS; Feng, M; Parkos, CA; Nusrat, A

    2014-01-01

    Desmosomal cadherins mediate cell–cell adhesion in epithelial tissues and have been known to be altered in cancer. We have previously shown that one of the two intestinal epithelial desmosomal cadherins, desmocollin-2 (Dsc2) loss promotes colonic epithelial carcinoma cell proliferation and tumor formation. In this study we show that loss of the other intestinal desmosomal cadherin, desmoglein-2 (Dsg2) that pairs with Dsc2, results in decreased epithelial cell proliferation and suppressed xenograft tumor growth in mice. Dsg2-deficient cells demonstrated a compensatory increase in Dsc2 expression, and small interfering RNA-mediated loss of Dsc2 restored proliferation in Dsg2-deficient cells. Dsg2 downregulation inhibited epidermal growth factor receptor (EGFR) signaling and cell proliferation through altered phosphorylation of EGFR and downstream extracellular signal-regulated kinase activation in parallel with inhibited EGFR receptor internalization. Additionally, we demonstrated a central role of Dsc2 in controlling EGFR signaling and cell proliferation in intestinal epithelial cells. Consistent with these findings, analyses of human colon cancers demonstrated increased Dsg2 protein expression. Taken together, these data demonstrate that partner desmosomal cadherins Dsg2 and Dsc2 play opposing roles in controlling colonic carcinoma cell proliferation through differential effects on EGFR signaling. PMID:24166502

  15. Promoting Cell Proliferation Using Water Dispersible Germanium Nanowires

    PubMed Central

    Bezuidenhout, Michael; Liu, Pai; Singh, Shalini; Kiely, Maeve

    2014-01-01

    Group IV Nanowires have strong potential for several biomedical applications. However, to date their use remains limited because many are synthesised using heavy metal seeds and functionalised using organic ligands to make the materials water dispersible. This can result in unpredicted toxic side effects for mammalian cells cultured on the wires. Here, we describe an approach to make seedless and ligand free Germanium nanowires water dispersible using glutamic acid, a natural occurring amino acid that alleviates the environmental and health hazards associated with traditional functionalisation materials. We analysed the treated material extensively using Transmission electron microscopy (TEM), High resolution-TEM, and scanning electron microscope (SEM). Using a series of state of the art biochemical and morphological assays, together with a series of complimentary and synergistic cellular and molecular approaches, we show that the water dispersible germanium nanowires are non-toxic and are biocompatible. We monitored the behaviour of the cells growing on the treated germanium nanowires using a real time impedance based platform (xCELLigence) which revealed that the treated germanium nanowires promote cell adhesion and cell proliferation which we believe is as a result of the presence of an etched surface giving rise to a collagen like structure and an oxide layer. Furthermore this study is the first to evaluate the associated effect of Germanium nanowires on mammalian cells. Our studies highlight the potential use of water dispersible Germanium Nanowires in biological platforms that encourage anchorage-dependent cell growth. PMID:25237816

  16. Promoting cell proliferation using water dispersible germanium nanowires.

    PubMed

    Bezuidenhout, Michael; Liu, Pai; Singh, Shalini; Kiely, Maeve; Ryan, Kevin M; Kiely, Patrick A

    2014-01-01

    Group IV Nanowires have strong potential for several biomedical applications. However, to date their use remains limited because many are synthesised using heavy metal seeds and functionalised using organic ligands to make the materials water dispersible. This can result in unpredicted toxic side effects for mammalian cells cultured on the wires. Here, we describe an approach to make seedless and ligand free Germanium nanowires water dispersible using glutamic acid, a natural occurring amino acid that alleviates the environmental and health hazards associated with traditional functionalisation materials. We analysed the treated material extensively using Transmission electron microscopy (TEM), High resolution-TEM, and scanning electron microscope (SEM). Using a series of state of the art biochemical and morphological assays, together with a series of complimentary and synergistic cellular and molecular approaches, we show that the water dispersible germanium nanowires are non-toxic and are biocompatible. We monitored the behaviour of the cells growing on the treated germanium nanowires using a real time impedance based platform (xCELLigence) which revealed that the treated germanium nanowires promote cell adhesion and cell proliferation which we believe is as a result of the presence of an etched surface giving rise to a collagen like structure and an oxide layer. Furthermore this study is the first to evaluate the associated effect of Germanium nanowires on mammalian cells. Our studies highlight the potential use of water dispersible Germanium Nanowires in biological platforms that encourage anchorage-dependent cell growth.

  17. Beyond cell proliferation in avian facial morphogenesis

    PubMed Central

    Linde-Medina, Marta; Hallgrímsson, Benedikt; Marcucio, Ralph

    2016-01-01

    The upper jaw in vertebrates forms from several prominences that arise around the stomodeum or primitive mouth. These prominences undergo coordinated growth and morphogenesis to fuse and form the face. Undirected, regionalized cell proliferation is thought to be the driving force behind the morphogenesis of the facial prominences. However, recent findings suggest that directed cell behaviors in the mesenchyme (e.g., directed cell division, directed cell movement, convergent extension) might be required for successful face formation. Here we discuss the evidence for this view and how directed behaviors may interact with the basement membrane to regulate morphogenesis of the facial region. We believe that future research in these largely unexplored areas could significantly impact our understanding of facial morphogenesis. PMID:26637960

  18. Eimeria bovis modulates adhesion molecule gene transcription in and PMN adhesion to infected bovine endothelial cells.

    PubMed

    Hermosilla, Carlos; Zahner, Horst; Taubert, Anja

    2006-04-01

    Eimeria bovis is an important coccidian parasite of cattle causing severe diarrhea in young animals. Its first schizogony takes place in endothelial cells of the ileum resulting in the formation of macroschizonts 14-18 days p.i. This longlasting development suggests a particular immune evasion strategy of the parasite. Here, we analyse early innate immune reactions to E. bovis by determining the adhesion of polymorphonuclear neutrophils (PMN) to infected endothelial cell layers under flow conditions and the transcription of adhesion molecule genes in infected host cells. Bovine umbilical vein endothelial cells (BUVEC) were infected with E. bovis sporozoites. Sporozoites invaded BUVEC within 1h and the first mature macroschizonts occurred 14 days p.i. PMN adhesion was enhanced in E. bovis-infected BUVEC layers as early as 8h p.i.; maximum adhesion occurred 48 h p.i. Increased adhesion rates persisted until the end of the observation period at 14 days p.i. PMN adhered to both infected and uninfected cells within monolayers, suggesting paracrine cell activation. E. bovis infection upregulated the transcription of genes encoding for P-selectin, E-selectin, vascular cellular adhesion molecule 1 (VCAM-1) and intercellular adhesion molecule 1 (ICAM-1). Most marked effects concerned E-selectin followed by P-selectin, VCAM-1 and ICAM-1. Increased transcript levels were found beginning 30 min p.i. and maximum values occurred 1-2h p.i. (P-selectin) and 2-4h p.i. (E-selectin, VCAM-1, ICAM-1). By 12-24h p.i. levels had decreased to those of uninfected controls. Tumor necrosis factor alpha (TNFalpha)-induced PMN adhesion was significantly reduced in infected vs. uninfected BUVEC. Eimeria bovis also had suppressive effects on TNFalpha-mediated upregulation of adhesion molecule gene transcription. The data presented here suggest that infection of BUVEC with E. bovis on one hand induces proinflammatory reactions resulting in enhanced PMN adhesion mediated by upregulated adhesion

  19. Electrospun poly(ε-caprolactone)-based skin substitutes: In vivo evaluation of wound healing and the mechanism of cell proliferation.

    PubMed

    Augustine, Robin; Dominic, Edwin Anto; Reju, Indu; Kaimal, Balarama; Kalarikkal, Nandakumar; Thomas, Sabu

    2015-10-01

    In the present study, we have fabricated electrospun poly(ε-caprolactone)-based membranes, characterized and studied the in vivo cell migration and proliferation and wound healing activity. Moreover, we did not seed any cells prior to the animal implantation and we could observe excellent fibroblast attachment and cell proliferation. Further full thickness excision wound on guinea pig completely healed within 35 days. We could reach in an assumption that the enhanced cell proliferation and wound healing might be due to the surface degradation of the polymer under physiological conditions and the formation of functional groups like hydroxyl and carboxyl groups that promoted cell proliferation in a cell adhesion protein mediated mechanism. This study is a novel tissue engineering concept for the reconstruction of a damaged tissue without the in vitro cell seeding and proliferation prior to the in vivo implantation.

  20. Plant cell proliferation inside an inorganic host.

    PubMed

    Perullini, Mercedes; Rivero, María Mercedes; Jobbágy, Matías; Mentaberry, Alejandro; Bilmes, Sara A

    2007-01-10

    In recent years, much attention has been paid to plant cell culture as a tool for the production of secondary metabolites and the expression of recombinant proteins. Plant cell immobilization offers many advantages for biotechnological processes. However, the most extended matrices employed, such as calcium-alginate, cannot fully protect entrapped cells. Sol-gel chemistry of silicates has emerged as an outstanding strategy to obtain biomaterials in which living cells are truly protected. This field of research is rapidly developing and a large number of bacteria and yeast-entrapping ceramics have already been designed for different applications. But even mild thermal and chemical conditions employed in sol-gel synthesis may result harmful to cells of higher organisms. Here we present a method for the immobilization of plant cells that allows cell growth at cavities created inside a silica matrix. Plant cell proliferation was monitored for a 6-month period, at the end of which plant calli of more than 1 mm in diameter were observed inside the inorganic host. The resulting hybrid device had good mechanical stability and proved to be an effective barrier against biological contamination, suggesting that it could be employed for long-term plant cell entrapment applications.

  1. Cell Proliferation, Cell Death, and Size Regulation

    DTIC Science & Technology

    2000-10-01

    predicted to encode a novel 582 amino acid protein, perhaps interacting with molybdopterin. It is possible that the pie gene encodes a novel enzyme protecting against cell death during growth and development.

  2. Nanostructured conducting polymers for stiffness controlled cell adhesion

    NASA Astrophysics Data System (ADS)

    Moyen, Eric; Hama, Adel; Ismailova, Esma; Assaud, Loic; Malliaras, George; Hanbücken, Margrit; Owens, Roisin M.

    2016-02-01

    We propose a facile and reproducible method, based on ultra thin porous alumina membranes, to produce cm2 ordered arrays of nano-pores and nano-pillars on any kind of substrates. In particular our method enables the fabrication of conducting polymers nano-structures, such as poly[3,4-ethylenedioxythiophene]:poly[styrene sulfonate] (PEDOT:PSS). Here, we demonstrate the potential interest of those templates with controlled cell adhesion studies. The triggering of the eventual fate of the cell (proliferation, death, differentiation or migration) is mediated through chemical cues from the adsorbed proteins and physical cues such as surface energy, stiffness and topography. Interestingly, as well as through material properties, stiffness modifications can be induced by nano-topography, the ability of nano-pillars to bend defining an effective stiffness. By controlling the diameter, length, depth and material of the nano-structures, one can possibly tune the effective stiffness of a (nano) structured substrate. First results indicate a possible change in the fate of living cells on such nano-patterned devices, whether they are made of conducting polymer (soft material) or silicon (hard material).

  3. Intercellular Adhesion Molecule-1 Expression by Skeletal Muscle Cells Augments Myogenesis

    PubMed Central

    Goh, Qingnian; Dearth, Christopher L.; Corbett, Jacob T.; Pierre, Philippe; Chadee, Deborah N.; Pizza, Francis X.

    2014-01-01

    We previously demonstrated that the expression of intercellular adhesion molecule-1 (ICAM-1) by skeletal muscle cells after muscle overload contributes to ensuing regenerative and hypertrophic processes in skeletal muscle. The objective of the present study is to reveal mechanisms through which skeletal muscle cell expression of ICAM-1 augments regenerative and hypertrophic processes of myogenesis. This was accomplished by genetically engineering C2C12 myoblasts to stably express ICAM-1, and by inhibiting the adhesive and signaling functions of ICAM-1 through the use of a neutralizing antibody or cell penetrating peptide, respectively. Expression of ICAM-1 by cultured skeletal muscle cells augmented myoblast-myoblast adhesion, myotube formation, myonuclear number, myotube alignment, myotube-myotube fusion, and myotube size without influencing the ability of myoblasts to proliferate or differentiate. ICAM-1 augmented myotube formation, myonuclear accretion, and myotube alignment through a mechanism involving adhesion-induced activation of ICAM-1 signaling, as these dependent measures were reduced via antibody and peptide inhibition of ICAM-1. The adhesive and signaling functions of ICAM-1 also facilitated myotube hypertrophy through a mechanism involving myotube-myotube fusion, protein synthesis, and Akt/p70s6k signaling. Our findings demonstrate that ICAM-1 expression by skeletal muscle cells augments myogenesis, and establish a novel mechanism through which the inflammatory response facilitates growth processes in skeletal muscle. PMID:25281303

  4. The Proliferation Study of Hips Cell-Derived Neuronal Progenitors on Poly-Caprolactone Scaffold

    PubMed Central

    Havasi, Parvaneh; Soleimani, Masoud; Morovvati, Hassan; Bakhshandeh, Behnaz; Nabiuni, Mohammad

    2014-01-01

    Introduction The native inability of nervous system to regenerate, encourage researchers to consider neural tissue engineering as a potential treatment for spinal cord injuries. Considering the suitable characteristics of induced pluripotent stem cells (iPSCs) for tissue regeneration applications, in this study we investigated the adhesion, viability and proliferation of neural progenitors (derived from human iPSCs) on aligned poly-caprolactone (PCL) nanofibers. Methods Aligned poly-caprolactone nanofibrous scaffold was fabricated by electrospinning and characterized by scanning electron microscopy (SEM). Through neural induction, neural progenitor cells were derived from induced pluripotent stem cells. After cell seeding on the scaffolds, their proliferation was investigated on different days of culture. Results According to the SEM micrographs, the electrospun PCL scaffolds were aligned along with uniformed morphology. Evaluation of adhesion and viability of neural progenitor cells on plate (control) and PCL scaffold illustrated increasing trends in proliferation but this rate was higher in scaffold group. The statistical analyses confirmed significant differences between groups on 36h and 48h. Discussion Evaluation of cell proliferation along with morphological assessments, staining and SEM finding suggested biocompatibility of the PCL scaffolds and suitability of the combination of the mentioned scaffold and human iPS cells for neural regeneration. PMID:25337369

  5. RGD modified polymers: biomaterials for stimulated cell adhesion and beyond.

    PubMed

    Hersel, Ulrich; Dahmen, Claudia; Kessler, Horst

    2003-11-01

    Since RGD peptides (R: arginine; G: glycine; D: aspartic acid) have been found to promote cell adhesion in 1984 (Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule, Nature 309 (1984) 30), numerous materials have been RGD functionalized for academic studies or medical applications. This review gives an overview of RGD modified polymers, that have been used for cell adhesion, and provides information about technical aspects of RGD immobilization on polymers. The impacts of RGD peptide surface density, spatial arrangement as well as integrin affinity and selectivity on cell responses like adhesion and migration are discussed.

  6. Single Cell Adhesion Assay Using Computer Controlled Micropipette

    PubMed Central

    Salánki, Rita; Hős, Csaba; Orgovan, Norbert; Péter, Beatrix; Sándor, Noémi; Bajtay, Zsuzsa; Erdei, Anna; Horvath, Robert; Szabó, Bálint

    2014-01-01

    Cell adhesion is a fundamental phenomenon vital for all multicellular organisms. Recognition of and adhesion to specific macromolecules is a crucial task of leukocytes to initiate the immune response. To gain statistically reliable information of cell adhesion, large numbers of cells should be measured. However, direct measurement of the adhesion force of single cells is still challenging and today’s techniques typically have an extremely low throughput (5–10 cells per day). Here, we introduce a computer controlled micropipette mounted onto a normal inverted microscope for probing single cell interactions with specific macromolecules. We calculated the estimated hydrodynamic lifting force acting on target cells by the numerical simulation of the flow at the micropipette tip. The adhesion force of surface attached cells could be accurately probed by repeating the pick-up process with increasing vacuum applied in the pipette positioned above the cell under investigation. Using the introduced methodology hundreds of cells adhered to specific macromolecules were measured one by one in a relatively short period of time (∼30 min). We blocked nonspecific cell adhesion by the protein non-adhesive PLL-g-PEG polymer. We found that human primary monocytes are less adherent to fibrinogen than their in vitro differentiated descendants: macrophages and dendritic cells, the latter producing the highest average adhesion force. Validation of the here introduced method was achieved by the hydrostatic step-pressure micropipette manipulation technique. Additionally the result was reinforced in standard microfluidic shear stress channels. Nevertheless, automated micropipette gave higher sensitivity and less side-effect than the shear stress channel. Using our technique, the probed single cells can be easily picked up and further investigated by other techniques; a definite advantage of the computer controlled micropipette. Our experiments revealed the existence of a sub

  7. Single cell adhesion assay using computer controlled micropipette.

    PubMed

    Salánki, Rita; Hős, Csaba; Orgovan, Norbert; Péter, Beatrix; Sándor, Noémi; Bajtay, Zsuzsa; Erdei, Anna; Horvath, Robert; Szabó, Bálint

    2014-01-01

    Cell adhesion is a fundamental phenomenon vital for all multicellular organisms. Recognition of and adhesion to specific macromolecules is a crucial task of leukocytes to initiate the immune response. To gain statistically reliable information of cell adhesion, large numbers of cells should be measured. However, direct measurement of the adhesion force of single cells is still challenging and today's techniques typically have an extremely low throughput (5-10 cells per day). Here, we introduce a computer controlled micropipette mounted onto a normal inverted microscope for probing single cell interactions with specific macromolecules. We calculated the estimated hydrodynamic lifting force acting on target cells by the numerical simulation of the flow at the micropipette tip. The adhesion force of surface attached cells could be accurately probed by repeating the pick-up process with increasing vacuum applied in the pipette positioned above the cell under investigation. Using the introduced methodology hundreds of cells adhered to specific macromolecules were measured one by one in a relatively short period of time (∼30 min). We blocked nonspecific cell adhesion by the protein non-adhesive PLL-g-PEG polymer. We found that human primary monocytes are less adherent to fibrinogen than their in vitro differentiated descendants: macrophages and dendritic cells, the latter producing the highest average adhesion force. Validation of the here introduced method was achieved by the hydrostatic step-pressure micropipette manipulation technique. Additionally the result was reinforced in standard microfluidic shear stress channels. Nevertheless, automated micropipette gave higher sensitivity and less side-effect than the shear stress channel. Using our technique, the probed single cells can be easily picked up and further investigated by other techniques; a definite advantage of the computer controlled micropipette. Our experiments revealed the existence of a sub-population of

  8. Simulation of Cell Adhesion using a Particle Transport Model

    NASA Astrophysics Data System (ADS)

    Chesnutt, Jennifer

    2005-11-01

    An efficient computational method for simulation of cell adhesion through protein binding forces is discussed. In this method, the cells are represented by deformable elastic particles, and the protein binding is represented by a rate equation. The method is first developed for collision and adhesion of two similar cells impacting on each other from opposite directions. The computational method is then applied in a particle-transport model for a cloud of interacting and colliding cells, each of which are represented by particles of finite size. One application might include red blood cells adhering together to form rouleaux, which are chains of red blood cells that are found in different parts of the circulatory system. Other potential applications include adhesion of platelets to a blood vessel wall or mechanical heart valve, which is a precursor of thrombosis formation, or adhesion of cancer cells to organ walls in the lymphatic, circulatory, digestive or pulmonary systems.

  9. All-trans-retinoic acid induces integrin-independent B-cell adhesion to ADAM disintegrin domains.

    PubMed

    Bridges, Lance C; Lingo, Joshuah D; Grandon, Rachel A; Kelley, Melissa D

    2008-04-15

    Cell adhesion is an integral aspect of immunity facilitating extravasation of immune cells during homing and activation. All -trans-Retinoic acid ( t-RA) regulates leukocyte differentiation, proliferation, and transmigration. However, the role of t-RA in immune cell adhesion is poorly defined. In this study, we evaluated the impact of t-RA and its metabolism on B and T cell adhesion. Specifically, we address the impact of t-RA on the adhesive properties of the human mature B and T cell lines RPMI 8866, Daudi and Jurkats. The effect of t-RA exposure on cell adhesion to vascular cell adhesion molecule-1 (VCAM-1), a well-established integrin counter receptor involved in immunity, and to nonconventional ADAM integrin ligands was assessed. We show for the first time that t-RA potently induces B cell adhesion in an integrin-independent manner to both VCAM-1 and select ADAM disintegrin domains. Using retinoid extraction and reverse-phase HPLC analysis, we identify the retinoid that is functionally responsible for this augmented adhesion. We also provide evidence that this novel t-RA adhesive response is not prototypical of lymphocytes since both Daudi and Jurkats do not alter their adhesive properties upon t-RA treatment. Further, the t-RA metabolic profiles between these lineages is distinct with 9- cis-retinoic acid being exclusively detected in Jurkat media. This study is the first to demonstrate that t-RA directly induces B cell adhesion in an integrin-independent manner and is not contingent upon t-RA metabolism.

  10. B-cell receptor-associated protein 31 regulates human embryonic stem cell adhesion, stemness, and survival via control of epithelial cell adhesion molecule.

    PubMed

    Kim, Won-Tae; Seo Choi, Hong; Min Lee, Hyun; Jang, Young-Joo; Ryu, Chun Jeih

    2014-10-01

    B-Cell receptor-associated protein 31 (BAP31) regulates the export of secreted membrane proteins from the endoplasmic reticulum (ER) to the downstream secretory pathway. Previously, we generated a monoclonal antibody 297-D4 against the surface molecule on undifferentiated human embryonic stem cells (hESCs). Here, we found that 297-D4 antigen was localized to pluripotent hESCs and downregulated during early differentiation of hESCs and identified that the antigen target of 297-D4 was BAP31 on the hESC-surface. To investigate the functional role of BAP31 in hESCs, BAP31 expression was knocked down by small interfering RNA. BAP31 depletion impaired hESC self-renewal and pluripotency and drove hESC differentiation into multicell lineages. BAP31 depletion hindered hESC proliferation by arresting cell cycle at G0/G1 phase and inducing caspase-independent cell death. Interestingly, BAP31 depletion reduced hESC adhesion to extracellular matrix (ECM). Analysis of cell surface molecules showed decreased expression of epithelial cell adhesion molecule (EpCAM) in BAP31-depleted hESCs, while ectopic expression of BAP31 elevated the expression of EpCAM. EpCAM depletion also reduced hESC adhesion to ECM, arrested cell cycle at G0/G1 phase and induced cell death, producing similar effects to those of BAP31 depletion. BAP31 and EpCAM were physically associated and colocalized at the ER and cell surface. Both BAP31 and EpCAM depletion decreased cyclin D1 and E expression and suppressed PI3K/Akt signaling, suggesting that BAP31 regulates hESC stemness and survival via control of EpCAM expression. These findings provide, for the first time, mechanistic insights into how BAP31 regulates hESC stemness and survival via control of EpCAM expression.

  11. Proliferating cell nuclear antigen: a proteomics view.

    PubMed

    Naryzhny, S N

    2008-11-01

    Proliferating cell nuclear antigen (PCNA), a cell cycle marker protein, is well known as a DNA sliding clamp for DNA polymerase delta and as an essential component for eukaryotic chromosomal DNA replication and repair. Due to its mobility inside nuclei, PCNA is dynamically presented in a soluble or chromatin-associated form. The heterogeneity and specific modifications of PCNA may reflect its multiple functions and the presence of many binding partners in the cell. The recent proteomics approaches applied to characterizing PCNA interactions revealed multiple PCNA partners with a wide spectrum of activity and unveiled the possible existence of new PCNA functions. Since more than 100 PCNA-interacting proteins and several PCNA modifications have already been reported, a proteomics point of view seems exactly suitable to better understand the role of PCNA in cellular functions.

  12. Amygdalin influences bladder cancer cell adhesion and invasion in vitro.

    PubMed

    Makarević, Jasmina; Rutz, Jochen; Juengel, Eva; Kaulfuss, Silke; Tsaur, Igor; Nelson, Karen; Pfitzenmaier, Jesco; Haferkamp, Axel; Blaheta, Roman A

    2014-01-01

    The cyanogenic diglucoside amygdalin, derived from Rosaceae kernels, is employed by many patients as an alternative anti-cancer treatment. However, whether amygdalin indeed acts as an anti-tumor agent is not clear. Metastasis blocking properties of amygdalin on bladder cancer cell lines was, therefore, investigated. Amygdalin (10 mg/ml) was applied to UMUC-3, TCCSUP or RT112 bladder cancer cells for 24 h or for 2 weeks. Tumor cell adhesion to vascular endothelium or to immobilized collagen as well as tumor cell migration was examined. Effects of drug treatment on integrin α and β subtypes, on integrin-linked kinase (ILK) and total and activated focal adhesion kinase (FAK) were also determined. Integrin knock-down was carried out to evaluate integrin influence on migration and adhesion. A 24 h or 2 week amygdalin application distinctly reduced tumor cell adhesion and migration of UMUC-3 and RT112 cells. TCCSUP adhesion was also reduced, but migration was elevated under amygdalin. Integrin subtype expression was significantly and specifically altered by amygdalin depending on the cell line. ILK was moderately, and activated FAK strongly, lost in all tumor cell lines in the presence of amygdalin. Knock down of β1 integrin caused a significant decrease in both adhesion and migration of UMUC-3 cells, but a significant increase in TCCSUP adhesion. Knock down of β4 integrin caused a significant decrease in migration of RT112 cells. Since the different actions of amygdalin on the different cell lines was mirrored by β1 or β4 knock down, it is postulated that amygdalin influences adhesion and migratory properties of bladder cancer cells by modulating β1 or β4 integrin expression. The amygdalin induced increase in TCCSUP migratory behavior indicates that any anti-tumor benefits from amygdalin (seen with the other two cell lines) may depend upon the cancer cell type.

  13. Amygdalin Influences Bladder Cancer Cell Adhesion and Invasion In Vitro

    PubMed Central

    Makarević, Jasmina; Rutz, Jochen; Juengel, Eva; Kaulfuss, Silke; Tsaur, Igor; Nelson, Karen; Pfitzenmaier, Jesco

    2014-01-01

    The cyanogenic diglucoside amygdalin, derived from Rosaceae kernels, is employed by many patients as an alternative anti-cancer treatment. However, whether amygdalin indeed acts as an anti-tumor agent is not clear. Metastasis blocking properties of amygdalin on bladder cancer cell lines was, therefore, investigated. Amygdalin (10 mg/ml) was applied to UMUC-3, TCCSUP or RT112 bladder cancer cells for 24 h or for 2 weeks. Tumor cell adhesion to vascular endothelium or to immobilized collagen as well as tumor cell migration was examined. Effects of drug treatment on integrin α and β subtypes, on integrin-linked kinase (ILK) and total and activated focal adhesion kinase (FAK) were also determined. Integrin knock-down was carried out to evaluate integrin influence on migration and adhesion. A 24 h or 2 week amygdalin application distinctly reduced tumor cell adhesion and migration of UMUC-3 and RT112 cells. TCCSUP adhesion was also reduced, but migration was elevated under amygdalin. Integrin subtype expression was significantly and specifically altered by amygdalin depending on the cell line. ILK was moderately, and activated FAK strongly, lost in all tumor cell lines in the presence of amygdalin. Knock down of β1 integrin caused a significant decrease in both adhesion and migration of UMUC-3 cells, but a significant increase in TCCSUP adhesion. Knock down of β4 integrin caused a significant decrease in migration of RT112 cells. Since the different actions of amygdalin on the different cell lines was mirrored by β1 or β4 knock down, it is postulated that amygdalin influences adhesion and migratory properties of bladder cancer cells by modulating β1 or β4 integrin expression. The amygdalin induced increase in TCCSUP migratory behavior indicates that any anti-tumor benefits from amygdalin (seen with the other two cell lines) may depend upon the cancer cell type. PMID:25333694

  14. Hydrogen peroxide regulates cell adhesion through the redox sensor RPSA.

    PubMed

    Vilas-Boas, Filipe; Bagulho, Ana; Tenente, Rita; Teixeira, Vitor H; Martins, Gabriel; da Costa, Gonçalo; Jerónimo, Ana; Cordeiro, Carlos; Machuqueiro, Miguel; Real, Carla

    2016-01-01

    To become metastatic, a tumor cell must acquire new adhesion properties that allow migration into the surrounding connective tissue, transmigration across endothelial cells to reach the blood stream and, at the site of metastasis, adhesion to endothelial cells and transmigration to colonize a new tissue. Hydrogen peroxide (H2O2) is a redox signaling molecule produced in tumor cell microenvironment with high relevance for tumor development. However, the molecular mechanisms regulated by H2O2 in tumor cells are still poorly known. The identification of H2O2-target proteins in tumor cells and the understanding of their role in tumor cell adhesion are essential for the development of novel redox-based therapies for cancer. In this paper, we identified Ribosomal Protein SA (RPSA) as a target of H2O2 and showed that RPSA in the oxidized state accumulates in clusters that contain specific adhesion molecules. Furthermore, we showed that RPSA oxidation improves cell adhesion efficiency to laminin in vitro and promotes cell extravasation in vivo. Our results unravel a new mechanism for H2O2-dependent modulation of cell adhesion properties and identify RPSA as the H2O2 sensor in this process. This work indicates that high levels of RPSA expression might confer a selective advantage to tumor cells in an oxidative environment.

  15. The structure of cell-matrix adhesions: the new frontier.

    PubMed

    Hanein, Dorit; Horwitz, Alan Rick

    2012-02-01

    Adhesions between the cell and the extracellular matrix (ECM) are mechanosensitive multi-protein assemblies that transmit force across the cell membrane and regulate biochemical signals in response to the chemical and mechanical environment. These combined functions in force transduction, signaling and mechanosensing contribute to cellular phenotypes that span development, homeostasis and disease. These adhesions form, mature and disassemble in response to actin organization and physical forces that originate from endogenous myosin activity or external forces by the extracellular matrix. Despite advances in our understanding of the protein composition, interactions and regulation, our understanding of matrix adhesion structure and organization, how forces affect this organization, and how these changes dictate specific signaling events is limited. Insights across multiple structural levels are acutely needed to elucidate adhesion structure and ultimately the molecular basis of signaling and mechanotransduction. Here we describe the challenges and recent advances and prospects for unraveling the structure of cell-matrix adhesions and their response to force.

  16. van der Waals forces influencing adhesion of cells

    PubMed Central

    Kendall, K.; Roberts, A. D.

    2015-01-01

    Adhesion molecules, often thought to be acting by a ‘lock and key’ mechanism, have been thought to control the adhesion of cells. While there is no doubt that a coating of adhesion molecules such as fibronectin on a surface affects cell adhesion, this paper aims to show that such surface contamination is only one factor in the equation. Starting from the baseline idea that van der Waals force is a ubiquitous attraction between all molecules, and thereby must contribute to cell adhesion, it is clear that effects from geometry, elasticity and surface molecules must all add on to the basic cell attractive force. These effects of geometry, elasticity and surface molecules are analysed. The adhesion force measured between macroscopic polymer spheres was found to be strongest when the surfaces were absolutely smooth and clean, with no projecting protruberances. Values of the measured surface energy were then about 35 mJ m−2, as expected for van der Waals attractions between the non-polar molecules. Surface projections such as abrasion roughness or dust reduced the molecular adhesion substantially. Water cut the measured surface energy to 3.4 mJ m−2. Surface active molecules lowered the adhesion still further to less than 0.3 mJ m−2. These observations do not support the lock and key concept. PMID:25533101

  17. Cell adhesive peptides functionalized on CoCr alloy stimulate endothelialization and prevent thrombogenesis and restenosis.

    PubMed

    Castellanos, Maria Isabel; Guillem-Marti, Jordi; Mas-Moruno, Carlos; Díaz-Ricart, Maribel; Escolar, Ginés; Ginebra, Maria Pau; Gil, Francisco Javier; Pegueroles, Marta; Manero, Jose María

    2017-04-01

    Immobilization of bioactive peptide sequences on CoCr surfaces is an effective route to improve endothelialization, which is of great interest for cardiovascular stents. In this work, we explored the effect of physical and covalent immoblization of RGDS, YIGSR and their equimolar combination peptides on endothelial cells (EC) and smooth muscle cell (SMC) adhesion and on thrombogenicity. We extensively investigated using RT-qPCR, the expression by ECs cultured on functionalised CoCr surfaces of different genes. Genes relevant for adhesion (ICAM-1 and VCAM-1), vascularization (VEGFA, VEGFR-1 and VEGFR-2) and anti-thrombogenicity (tPA and eNOS) were over-expressed in the ECs grown to covalently functionalized CoCr surfaces compared to physisorbed and control surfaces. Pro-thrombogenic genes expression (PAI-1 and vWF) decreased over time. Cell co-cultures of ECs/SMCs found that functionalization increased the amount of adhered ECs onto modified surfaces compared to plain CoCr, independently of the used peptide and the strategy of immobilization. SMCs adhered less compared to ECs in all surfaces. All studied peptides showed a lower platelet cell adhesion compared to TCPS. Covalent functionalization of CoCr surfaces with an equimolar combination of RGDS and YIGSR represented prevailing strategy to enhance the early stages of ECs adhesion and proliferation, while preventing SMCs and platelet adhesion. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 973-983, 2017.

  18. Influence of electrospun scaffolds prepared from distinct polymers on proliferation and viability of endothelial cells

    NASA Astrophysics Data System (ADS)

    Matveeva, V. G.; Antonova, L. V.; Velikanova, E. A.; Sergeeva, E. A.; Krivkina, E. O.; Glushkova, T. V.; Kudryavtseva, Yu. A.; Barbarash, O. L.; Barbarash, L. S.

    2015-10-01

    We compared electrospun nonwoven scaffolds from polylactic acid (PLA), polycaprolactone (PCL), and polyhydroxybutyrate/valerate (PHBV)/polycaprolactone (PHBV/PCL). The surface of PHBV/PCL and PCL scaffolds was highly porous and consisted of randomly distributed fibers, whilst the surface of PLA scaffolds consisted of thin straight fibers, which located more sparsely, forming large pores. Culture of EA.hy 926 endothelial cells on these scaffolds during 7 days and further fluorescent microscopy demonstrated that the surface of PHBV/PCL scaffolds was most favorable for efficient adhesion, proliferation, and viability of endothelial cells. The lowest proliferation rate and cell viability were detected on PLA scaffolds. Therefore, PHBV/PCL electrospun nonwoven scaffolds demonstrated the best results regarding endothelial cell proliferation and viability as compared to PCL and PLA scaffolds.

  19. Influence of electrospun scaffolds prepared from distinct polymers on proliferation and viability of endothelial cells

    SciTech Connect

    Matveeva, V. G. Antonova, L. V. Velikanova, E. A.; Sergeeva, E. A.; Krivkina, E. O.; Glushkova, T. V.; Kudryavtseva, Yu. A.; Barbarash, O. L.; Barbarash, L. S.

    2015-10-27

    We compared electrospun nonwoven scaffolds from polylactic acid (PLA), polycaprolactone (PCL), and polyhydroxybutyrate/valerate (PHBV)/polycaprolactone (PHBV/PCL). The surface of PHBV/PCL and PCL scaffolds was highly porous and consisted of randomly distributed fibers, whilst the surface of PLA scaffolds consisted of thin straight fibers, which located more sparsely, forming large pores. Culture of EA.hy 926 endothelial cells on these scaffolds during 7 days and further fluorescent microscopy demonstrated that the surface of PHBV/PCL scaffolds was most favorable for efficient adhesion, proliferation, and viability of endothelial cells. The lowest proliferation rate and cell viability were detected on PLA scaffolds. Therefore, PHBV/PCL electrospun nonwoven scaffolds demonstrated the best results regarding endothelial cell proliferation and viability as compared to PCL and PLA scaffolds.

  20. Shark cartilage extract interferes with cell adhesion and induces reorganization of focal adhesions in cultured endothelial cells.

    PubMed

    Chen, J S; Chang, C M; Wu, J C; Wang, S M

    2000-06-06

    In this study, we examined the effects of shark cartilage extract on the attachment and spreading properties and the focal adhesion structure of cultured bovine pulmonary artery endothelial cells. Treatment with cartilage extract resulted in cell detachment from the substratum. Immunofluorescence staining of those treated cells that remained attached showed that, instead of being present in both central and peripheral focal adhesions as in control cells, both integrin alpha(v)beta(3) and vinculin were found only in peripheral focal adhesion and thinner actin filament bundles were seen. In addition to causing cell detachment, cartilage extract partially inhibited the initial adherence of the cells to the substratum in a dose-dependent manner. Integrin alpha(v)beta(3) and vinculin staining of these cells also showed a peripheral focal adhesion distribution pattern. Vitronectin induced cell spreading in the absence of serum, but was blocked by simultaneous incubation with cartilage extract, which was shown to inhibit both integrin alpha(v)beta(3) and vinculin recruitment to focal adhesion and the formation of stress fibers. Dot binding assays showed that these inhibitory effects on cell attachment and spreading were not due to direct binding of cartilage extract components to integrin alpha(v)beta(3) or vitronectin. Shark cartilage chondroitin sulfate had no inhibitory effect on either cell attachment or spreading of endothelial cells. These results show that the inhibitory effects of cartilage extract on cell attachment and spreading are mediated by modification of the organization of focal adhesion proteins.

  1. FGFR4 Downregulation of Cell Adhesion in Prostate Cancer

    DTIC Science & Technology

    2007-03-01

    AD_________________ Award Number: W81XWH-06-1-0385 TITLE: FGFR4 Downregulation of Cell Adhesion...2007 2. REPORT TYPE Annual 3. DATES COVERED 1 Mar 2006 – 28 Feb 2007 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER FGFR4 Downregulation of Cell...our project to examine the role of FGFR4 G388R in altering cell adhesion in prostate cancer. This includes acquiring expertise in the passage and

  2. Cell adhesion to borate glasses by colloidal probe microscopy.

    PubMed

    Wiederhorn, Sheldon M; Chae, Young-Hun; Simon, Carl G; Cahn, Jackson; Deng, Yan; Day, Delbert

    2011-05-01

    The adhesion of osteoblast-like cells to silicate and borate glasses was measured in cell growth medium using colloidal probe microscopy. The probes consisted of silicate and borate glass spheres, 25-50 μm in diameter, attached to atomic force microscope cantilevers. Variables of the study included glass composition and time of contact of the cell to the glasses. Increasing the time of contact from 15 to 900 s increased the force of adhesion. The data could be plotted linearly on a log-log plot of adhesive force versus time. Of the seven glasses tested, five had slopes close to 0.5, suggesting a square root dependence of the adhesive force on the contact time. Such behavior can be interpreted as a diffusion limited process occurring during the early stages of cell attachment. We suggest that the rate limiting step in the adhesion process is the diffusion of integrins resident in the cell membrane to the area of cell attachment. Data presented in this paper support the hypothesis of Hench et al. that strong adhesion depends on the formation of a calcium phosphate reaction layer on the surfaces of the glass. Glasses that did not form a calcium phosphate layer exhibited a weaker adhesive force relative to those glasses that did form a calcium phosphate layer.

  3. Transcriptionally Regulated Cell Adhesion Network Dictates Distal Tip Cell Directionality

    PubMed Central

    Wong, Ming-Ching; Kennedy, William P.; Schwarzbauer, Jean E.

    2015-01-01

    Background The mechanisms that govern directional changes in cell migration are poorly understood. The migratory paths of two distal tip cells (DTC) determine the U-shape of the C. elegans hermaphroditic gonad. The morphogenesis of this organ provides a model system to identify genes necessary for the DTCs to execute two stereotyped turns. Results Using candidate genes for RNAi knockdown in a DTC-specific strain, we identified two transcriptional regulators required for DTC turning: cbp-1, the CBP/p300 transcriptional coactivator homologue, and let-607, a CREBH transcription factor homologue. Further screening of potential target genes uncovered a network of integrin adhesion-related genes that have roles in turning and are dependent on cbp-1 and let-607 for expression. These genes include src-1/Src kinase, tln-1/talin, pat-2/α integrin and nmy-2, a nonmuscle myosin heavy chain. Conclusions Transcriptional regulation by means of cbp-1 and let-607 is crucial for determining directional changes during DTC migration. These regulators coordinate a gene network that is necessary for integrin-mediated adhesion. Overall, these results suggest that directional changes in cell migration rely on the precise gene regulation of adhesion. PMID:24811939

  4. Enzymatically-tailored pectins differentially influence the morphology, adhesion, cell cycle progression and survival of fibroblasts.

    PubMed

    Nagel, Marie-Danielle; Verhoef, René; Schols, Henk; Morra, Marco; Knox, J Paul; Ceccone, Giacomo; Della Volpe, Claudio; Vigneron, Pascale; Bussy, Cyrill; Gallet, Marlène; Velzenberger, Elodie; Vayssade, Muriel; Cascardo, Giovanna; Cassinelli, Clara; Haeger, Ash; Gilliland, Douglas; Liakos, Ioannis; Rodriguez-Valverde, Miguel; Siboni, Stefano

    2008-01-01

    Improved biocompatibility and performance of biomedical devices can be achieved through the incorporation of bioactive molecules on device surfaces. Five structurally distinct pectic polysaccharides (modified hairy regions (MHRs)) were obtained by enzymatic liquefaction of apple (MHR-B, MHR-A and MHR-alpha), carrot (MHR-C) and potato (MHR-P) cells. Polystyrene (PS) Petri dishes, aminated by a plasma deposition process, were surface modified by the covalent linking of the MHRs. Results clearly demonstrate that MHR-B induces cell adhesion, proliferation and survival, in contrast to the other MHRs. Moreover, MHR-alpha causes cells to aggregate, decrease proliferation and enter into apoptosis. Cells cultured in standard conditions with 1% soluble MHR-B or MHR-alpha show the opposite behaviour to the one observed on MHR-B and -alpha-grafted PS. Fibronectin was similarly adsorbed onto MHR-B and tissue culture polystyrene (TCPS) control, but poorly on MHR-alpha. The Fn cell binding site (RGD sequence) was more accessible on MHR-B than on TCPS control, but poorly on MHR-alpha. The disintegrin echistatin inhibited fibroblast adhesion and spreading on MHR-B-grafted PS, which suggests that MHRs control fibroblast behaviour via serum-adhesive proteins. This study provides a basis for the design of intelligently-tailored biomaterial coatings able to induce specific cell functions.

  5. Adhesion-mediated self-renewal abilities of Ph+ blastoma cells

    SciTech Connect

    Funayama, Keiji; Saito-Kurimoto, Yumi; Ebihara, Yasuhiro; Shimane, Miyuki; Nomura, Hitoshi; Tsuji, Ko-ichiro; Asano, Shigetaka

    2010-05-28

    The Philadelphia chromosome-positive blastoma, maintained by serial subcutaneous transplantation in nude mice, is a highly proliferating biological mass consisting of homogenous CD34{sup +}CD38{sup -} myeloblastoid cells. These cells newly evolved from pluripotent leukemia stem cells of chronic myeloid leukemia in the chronic phase. Therefore, this mass may provide a unique tool for better understanding cellular and molecular mechanisms of self-renewal of leukemia stem cells. In this paper, we demonstrated that intravenously injected blastoma cells can cause Ph+ blastic leukemia with multiple invasive foci in NOD/SCID mice but not in nude mice. In addition, using an in vitro culture system, we clearly showed that blastoma cell adhesion to OP9 stromal cells accelerates blastoma cell proliferation that is associated with up-regulation of BMI1 gene expression; increased levels of {beta}-catenin and the Notch1 intra-cellular domain; and changed the expression pattern of variant CD44 forms, which are constitutively expressed in these blastoma cells. These findings strongly suggest that adhesion of leukemic stem cells to stromal cells via CD44 might be indispensable for their cellular defense against attack by immune cells and for maintenance of their self-renewal ability.

  6. Effects of enamel matrix proteins on adherence, proliferation and migration of epithelial cells: A real-time in vitro study

    PubMed Central

    Wyganowska-Swiatkowska, Marzena; Urbaniak, Paulina; Lipinski, Daniel; Szalata, Marlena; Borysiak, Karolina; Jakun, Jerzy; Kotwicka, Malgorzata

    2017-01-01

    Enamel matrix derivative (EMD) can mimic odontogenic effects by inducing the proliferation and differentiation of connective tissue progenitor cells, stimulating bone growth and arresting epithelial cells migration. To the best of our knowledge, there is no data indicating that any active component of EMD reduces epithelial cell viability. The present study examines the impact of commercial lyophilized EMD, porcine recombinant amelogenin (prAMEL; 21.3 kDa) and tyrosine-rich amelogenin peptide (TRAP) on the adherence, proliferation and migration of human epithelial cells in real-time. The tongue carcinoma cell line SCC-25 was stimulated with EMD, porcine recombinant AMEL and TRAP, at concentrations of 12.5, 25 and 50 µg/ml. Cell adherence, migration and proliferation were monitored in real-time using the xCELLigence system. No significant effects of EMD on the morphology, adhesion, proliferation and migration of SCC-25 cells were observed. However, porcine recombinant AMEL had a dose-dependent inhibitory effect on SCC-25 cell proliferation and migration. Predominantly, no notable differences were found between control and TRAP-treated cells in terms of cell adhesion and migration, a decrease in proliferation was observed, but this was not statistically significant. EMD and its active components do not increase the tongue cancer cell viability. PMID:28123485

  7. Effects of enamel matrix proteins on adherence, proliferation and migration of epithelial cells: A real-time in vitro study.

    PubMed

    Wyganowska-Swiatkowska, Marzena; Urbaniak, Paulina; Lipinski, Daniel; Szalata, Marlena; Borysiak, Karolina; Jakun, Jerzy; Kotwicka, Malgorzata

    2017-01-01

    Enamel matrix derivative (EMD) can mimic odontogenic effects by inducing the proliferation and differentiation of connective tissue progenitor cells, stimulating bone growth and arresting epithelial cells migration. To the best of our knowledge, there is no data indicating that any active component of EMD reduces epithelial cell viability. The present study examines the impact of commercial lyophilized EMD, porcine recombinant amelogenin (prAMEL; 21.3 kDa) and tyrosine-rich amelogenin peptide (TRAP) on the adherence, proliferation and migration of human epithelial cells in real-time. The tongue carcinoma cell line SCC-25 was stimulated with EMD, porcine recombinant AMEL and TRAP, at concentrations of 12.5, 25 and 50 µg/ml. Cell adherence, migration and proliferation were monitored in real-time using the xCELLigence system. No significant effects of EMD on the morphology, adhesion, proliferation and migration of SCC-25 cells were observed. However, porcine recombinant AMEL had a dose-dependent inhibitory effect on SCC-25 cell proliferation and migration. Predominantly, no notable differences were found between control and TRAP-treated cells in terms of cell adhesion and migration, a decrease in proliferation was observed, but this was not statistically significant. EMD and its active components do not increase the tongue cancer cell viability.

  8. Cell Adhesion Molecules in Chemically-Induced Renal Injury

    PubMed Central

    Prozialeck, Walter C.; Edwards, Joshua R.

    2007-01-01

    Cell adhesion molecules are integral cell-membrane proteins that maintain cell-cell and cell-substrate adhesion, and in some cases, act as regulators of intracellular signaling cascades. In the kidney, cell adhesion molecules such as the cadherins, the catenins, ZO-1, occludin and the claudins are essential for maintaining the epithelial polarity and barrier integrity that are necessary for the normal absorption/excretion of fluid and solutes. A growing volume of evidence indicates that these cell adhesion molecules are important early targets for a variety of nephrotoxic substances including metals, drugs, and venom components. In addition, it is now widely appreciated that molecules such as ICAM-1, the integrins and selectins play important roles in the recruitment of leukocytes and inflammatory responses that are associated with nephrotoxic injury. This review summarizes the results of recent in vitro and in vivo studies indicating that these cell adhesion molecules may be primary molecular targets in many types of chemically-induced renal injury. Some of the specific agents that are discussed include Cd, Hg, Bi, cisplatin, aminoglycoside antibiotics, S-(1,2-dichlorovinyl-L-cysteine) (DCVC) and various venom toxins. This review also includes a discussion of the various mechanisms by which these substances can affect cell adhesion molecules in the kidney. PMID:17316817

  9. Research Techniques Made Simple: Techniques to Assess Cell Proliferation.

    PubMed

    Romar, George A; Kupper, Thomas S; Divito, Sherrie J

    2016-01-01

    Cell proliferation is commonly assayed in the laboratory for research purposes, but is increasingly used clinically to gauge tumor aggressiveness and potentially guide care. Therefore, both researchers and clinicians should have a basic understanding of techniques used to assess cell proliferation. Multiple cell proliferation assays exist, and the choice of method depends on the laboratory resources available, the types of cells/tissues to be studied, and the specific experimental goals. In this article, we identify four overarching categories of cell proliferation assays that signify various stages of the cell cycle: nucleoside-analog incorporation, cell cycle-associated protein detection, use of cytoplasmic proliferation dyes, and indirect measures of cell proliferation. Each method has strengths and limitations that should guide the dermatology investigator's choice of assay.

  10. Numb-deficient satellite cells have regeneration and proliferation defects

    PubMed Central

    George, Rajani M.; Biressi, Stefano; Beres, Brian J.; Rogers, Erik; Mulia, Amanda K.; Allen, Ronald E.; Rawls, Alan; Rando, Thomas A.; Wilson-Rawls, Jeanne

    2013-01-01

    The adaptor protein Numb has been implicated in the switch between cell proliferation and differentiation made by satellite cells during muscle repair. Using two genetic approaches to ablate Numb, we determined that, in its absence, muscle regeneration in response to injury was impaired. Single myofiber cultures demonstrated a lack of satellite cell proliferation in the absence of Numb, and the proliferation defect was confirmed in satellite cell cultures. Quantitative RT-PCR from Numb-deficient satellite cells demonstrated highly up-regulated expression of p21 and Myostatin, both inhibitors of myoblast proliferation. Transfection with Myostatin-specific siRNA rescued the proliferation defect of Numb-deficient satellite cells. Furthermore, overexpression of Numb in satellite cells inhibited Myostatin expression. These data indicate a unique function for Numb during the initial activation and proliferation of satellite cells in response to muscle injury. PMID:24170859

  11. Calcium channels, external calcium concentration and cell proliferation.

    PubMed

    Borowiec, Anne-Sophie; Bidaux, Gabriel; Pigat, Natascha; Goffin, Vincent; Bernichtein, Sophie; Capiod, Thierry

    2014-09-15

    Evidence for a role for calcium channel proteins in cell proliferation is numerous suggesting that calcium influx is essential in this physiological process. Several studies in the past thirty years have demonstrated that calcium channel expression levels are determinant in cell proliferation. Voltage-gated, store-operated, second messengers and receptor-operated calcium channels have been associated to cell proliferation. However, the relationship between calcium influx and cell proliferation can be uncoupled in transformed and cancer cells, resulting in an external calcium-independent proliferation. Thus, protein expression could be more important than channel function to trigger cell proliferation suggesting that additional channel functions may be responsible to reconcile calcium channel expression and cell proliferation. When needed, external calcium concentration is obviously important for calcium channel function but it also regulates calcium sensing receptor (CaSR) activity. CaSR can up- or down-regulate cell proliferation depending on physiological conditions. CaSR sensitivity to external calcium is within the 0.5 to 5 mM range and therefore, the role of these receptors in cell proliferation must be taken into account. We therefore suggest here that cell proliferation rates could depend on the relative balance between calcium influx and CaSR activation.

  12. Dissecting cell adhesion architecture using advanced imaging techniques

    PubMed Central

    Morton, Penny E

    2011-01-01

    Cell adhesion to extracellular matrix proteins or to other cells is essential for the control of embryonic development, tissue integrity, immune function and wound healing. Adhesions are tightly spatially regulated structures containing over one hundred different proteins that coordinate both dynamics and signaling events at these sites. Extensive biochemical and morphological analysis of adhesion types over the past three decades has greatly improved understanding of individual protein contributions to adhesion signaling and, in some cases, dynamics. However, it is becoming increasingly clear that these diverse macromolecular complexes contain a variety of protein sub-networks, as well as distinct sub-domains that likely play important roles in regulating adhesion behavior. Until recently, resolving these structures, which are often less than a micron in size, was hampered by the limitations of conventional light microscopy. However, recent advances in optical techniques and imaging methods have revealed exciting insight into the intricate control of adhesion structure and assembly. Here we provide an overview of the recent data arising from such studies of cell:matrix and cell:cell contact and an overview of the imaging strategies that have been applied to study the intricacies and hierarchy of proteins within adhesions. PMID:21785274

  13. Amplified effect of surface charge on cell adhesion by nanostructures

    NASA Astrophysics Data System (ADS)

    Xu, Li-Ping; Meng, Jingxin; Zhang, Shuaitao; Ma, Xinlei; Wang, Shutao

    2016-06-01

    Nano-biointerfaces with varied surface charge can be readily fabricated by integrating a template-based process with maleimide-thiol coupling chemistry. Significantly, nanostructures are employed for amplifying the effect of surface charge on cell adhesion, as revealed by the cell-adhesion performance, cell morphology and corresponding cytoskeletal organization. This study may provide a promising strategy for developing new biomedical materials with tailored cell adhesion for tissue implantation and regeneration.Nano-biointerfaces with varied surface charge can be readily fabricated by integrating a template-based process with maleimide-thiol coupling chemistry. Significantly, nanostructures are employed for amplifying the effect of surface charge on cell adhesion, as revealed by the cell-adhesion performance, cell morphology and corresponding cytoskeletal organization. This study may provide a promising strategy for developing new biomedical materials with tailored cell adhesion for tissue implantation and regeneration. Electronic supplementary information (ESI) available: Experimental details, SEM, KFM AFM, chemical modification and characterization. See DOI: 10.1039/c6nr00649c

  14. Neural cell adhesion molecule (NCAM) marks adult myogenic cells committed to differentiation

    SciTech Connect

    Capkovic, Katie L.; Stevenson, Severin; Johnson, Marc C.; Thelen, Jay J.; Cornelison, D.D.W.

    2008-04-15

    Although recent advances in broad-scale gene expression analysis have dramatically increased our knowledge of the repertoire of mRNAs present in multiple cell types, it has become increasingly clear that examination of the expression, localization, and associations of the encoded proteins will be critical for determining their functional significance. In particular, many signaling receptors, transducers, and effectors have been proposed to act in higher-order complexes associated with physically distinct areas of the plasma membrane. Adult muscle stem cells (satellite cells) must, upon injury, respond appropriately to a wide range of extracellular stimuli: the role of such signaling scaffolds is therefore a potentially important area of inquiry. To address this question, we first isolated detergent-resistant membrane fractions from primary satellite cells, then analyzed their component proteins using liquid chromatography-tandem mass spectrometry. Transmembrane and juxtamembrane components of adhesion-mediated signaling pathways made up the largest group of identified proteins; in particular, neural cell adhesion molecule (NCAM), a multifunctional cell-surface protein that has previously been associated with muscle regeneration, was significant. Immunohistochemical analysis revealed that not only is NCAM localized to discrete areas of the plasma membrane, it is also a very early marker of commitment to terminal differentiation. Using flow cytometry, we have sorted physically homogeneous myogenic cultures into proliferating and differentiating fractions based solely upon NCAM expression.

  15. Minimal synthetic cells to study integrin-mediated adhesion.

    PubMed

    Frohnmayer, Johannes P; Brüggemann, Dorothea; Eberhard, Christian; Neubauer, Stefanie; Mollenhauer, Christine; Boehm, Heike; Kessler, Horst; Geiger, Benjamin; Spatz, Joachim P

    2015-10-12

    To shed light on cell-adhesion-related molecular pathways, synthetic cells offer the unique advantage of a well-controlled model system with reduced molecular complexity. Herein, we show that liposomes with the reconstituted platelet integrin αIIb β3 as the adhesion-mediating transmembrane protein are a functional minimal cell model for studying cellular adhesion mechanisms in a defined environment. The interaction of these synthetic cells with various extracellular matrix proteins was analyzed using a quartz crystal microbalance with dissipation monitoring. The data indicated that integrin was functionally incorporated into the lipid vesicles, thus enabling integrin-specific adhesion of the engineered liposomes to fibrinogen- and fibronectin-functionalized surfaces. Then, we were able to initiate the detachment of integrin liposomes from these surfaces in the presence of the peptide GRGDSP, a process that is even faster with our newly synthesized peptide mimetic SN529, which specifically inhibits the integrin αIIb β3 .

  16. In vitro adhesion of Escherichia coli to porcine small intestinal epithelial cells: pili as adhesive factors.

    PubMed Central

    Isaacson, R E; Fusco, P C; Brinton, C C; Moon, H W

    1978-01-01

    Escherichia coli strains with pili (K99 or 987P) known to facilitate intestinal colonization adhered in vitro to porcine intestinal epithelial cells. These strains adhered equally to both ileal and jejunal epithelial cells. A laboratory E. coli strain that has type 1 pili also adhered to porcine intestinal epithelial cells. When nonpiliated cells derived from 987P+, K99+, or type 1 pilus+ strains were used for in vitro adhesion assays, they failed to adhere. The attachment of piliated bacteria to epithelial cells was a saturable process that plateaued at 30 to 40 bacterial cells attached per epithelial cell. Competitive inhibition of bacterial cell attachment to epithelial cells with purified pili showed that only purified 987P competed against the 987P+ strain and only purified type 1 pili competed against the type 1 pilus+ strain. Competition between a K99+ strain and K99 was not consistently achieved. K99+, 987P+, and type 1 pilus+ bacteria could be prevented from adhering to epithelial cells by Fab fragments specific for K99, 987P, or type 1 pili, respectively. Fab fragments specific for non-K99 bacterial surface antigens did not inhibit adhesion of the K99+ strain. It is concluded that adhesion of E. coli to porcine intestinal epithelial cells in vitro is mediated by pili and that the epithelial cells used apparently had different receptors for different pili. PMID:357285

  17. In vitro adhesion of Escherichia coli to porcine small intestinal epithelial cells: pili as adhesive factors.

    PubMed

    Isaacson, R E; Fusco, P C; Brinton, C C; Moon, H W

    1978-08-01

    Escherichia coli strains with pili (K99 or 987P) known to facilitate intestinal colonization adhered in vitro to porcine intestinal epithelial cells. These strains adhered equally to both ileal and jejunal epithelial cells. A laboratory E. coli strain that has type 1 pili also adhered to porcine intestinal epithelial cells. When nonpiliated cells derived from 987P+, K99+, or type 1 pilus+ strains were used for in vitro adhesion assays, they failed to adhere. The attachment of piliated bacteria to epithelial cells was a saturable process that plateaued at 30 to 40 bacterial cells attached per epithelial cell. Competitive inhibition of bacterial cell attachment to epithelial cells with purified pili showed that only purified 987P competed against the 987P+ strain and only purified type 1 pili competed against the type 1 pilus+ strain. Competition between a K99+ strain and K99 was not consistently achieved. K99+, 987P+, and type 1 pilus+ bacteria could be prevented from adhering to epithelial cells by Fab fragments specific for K99, 987P, or type 1 pili, respectively. Fab fragments specific for non-K99 bacterial surface antigens did not inhibit adhesion of the K99+ strain. It is concluded that adhesion of E. coli to porcine intestinal epithelial cells in vitro is mediated by pili and that the epithelial cells used apparently had different receptors for different pili.

  18. Matrix protein CCN1 is critical for prostate carcinoma cell proliferation and TRAIL-induced apoptosis.

    PubMed

    Franzen, Carrie A; Chen, Chih-Chiun; Todorović, Viktor; Juric, Vladislava; Monzon, Ricardo I; Lau, Lester F

    2009-07-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) plays an important role in immune surveillance and preferentially induces apoptosis in cancer cells over normal cells, suggesting its potential in cancer therapy. However, the molecular basis for its selective killing of cancer cells is not well understood. Recent studies have identified the CCN family of integrin-binding matricellular proteins as important regulators of cell behavior, including cell adhesion, proliferation, migration, differentiation, and survival. We show here that CCN1 (CYR61) supports the adhesion of prostatic carcinoma cells as an adhesion substrate through integrins and heparan sulfate proteoglycans. Knockdown of CCN1 expression in PC-3 and DU-145 androgen-independent prostate cancer cells strongly inhibited their proliferation without causing apoptosis, indicating that CCN1 promotes their growth. However, CCN1 also significantly enhances TRAIL-induced apoptosis through interaction with integrins alphavbeta3 and alpha6beta4 and the cell-surface heparan sulfate proteoglycan syndecan-4, acting through a protein kinase Calpha-dependent mechanism without requiring de novo protein synthesis. Knockdown of CCN1 expression in PC-3, DU-145, and LNCaP cells severely blunted their sensitivity to TRAIL, an effect that was reversed by exogenously added CCN1 protein. These findings reveal a functional dichotomy for CCN1 in prostate carcinoma cells, because it contributes to both cell proliferation and TRAIL-induced cell death and suggest that CCN1 expression status may be an important parameter in assessing the efficacy of TRAIL-dependent cancer therapy.

  19. RhoA promotes epidermal stem cell proliferation via PKN1-cyclin D1 signaling

    PubMed Central

    Wang, Fan; Zhan, Rixing; Chen, Liang; Dai, Xia; Wang, Wenping; Guo, Rui; Li, Xiaoge; Li, Zhe; Wang, Liang; Huang, Shupeng; Shen, Jie

    2017-01-01

    Objective Epidermal stem cells (ESCs) play a critical role in wound healing, but the mechanism underlying ESC proliferation is not well defined. Here, we explore the effects of RhoA on ESC proliferation and the possible underlying mechanism. Methods Human ESCs were enriched by rapid adhesion to collagen IV. RhoA(+/+)(G14V), RhoA(-/-)(T19N) and pGFP control plasmids were transfected into human ESCs. The effect of RhoA on cell proliferation was detected by cell proliferation and DNA synthesis assays. Induction of PKN1 activity by RhoA was determined by immunoblot analysis, and the effects of PKN1 on RhoA in terms of inducing cell proliferation and cyclin D1 expression were detected using specific siRNA targeting PKN1. The effects of U-46619 (a RhoA agonist) and C3 transferase (a RhoA antagonist) on ESC proliferation were observed in vivo. Results RhoA had a positive effect on ESC proliferation, and PKN1 activity was up-regulated by the active RhoA mutant (G14V) and suppressed by RhoA T19N. Moreover, the ability of RhoA to promote ESC proliferation and DNA synthesis was interrupted by PKN1 siRNA. Additionally, cyclin D1 protein and mRNA expression levels were up-regulated by RhoA G14V, and these effects were inhibited by siRNA-mediated knock-down of PKN1. RhoA also promoted ESC proliferation via PKN in vivo. Conclusion This study shows that the effect of RhoA on ESC proliferation is mediated by activation of the PKN1-cyclin D1 pathway in vitro, suggesting that RhoA may serve as a new therapeutic target for wound healing. PMID:28222172

  20. Dystrophin Dp71 in PC12 cell adhesion

    PubMed Central

    Enríquez-Aragón, Jose Arturo; Cerna-Cortés, Joel; Bermúdez de León, Mario; García-Sierra, Francisco; González, Everardo; Mornet, Dominique; Cisneros, Bulmaro

    2005-01-01

    Previously, we reported that PC12 cells with decreased Dp71 expression (antisense-Dp71 cells) display deficient nerve-growth-factor-induced neurite outgrowth. In this study, we show that disturbed neurite outgrowth of antisense-Dp71 cells is accompanied by decreased adhesion activity on laminin, collagen and fibronectin. In wild-type cells, the immunostaining of Dp71 and _1-integrin overlaps in the basal area contacting the substrate, but staining of both proteins decrease in the antisense-Dp71 cells. Morphology of antisense-Dp71 cells at the electron microscopic level is characterized by the lack of filopodia, cellular projections involved in adhesion. Our findings suggest that Dp71 is required for the efficient PC12 cell attachment to b1-integrin-dependent substrata and that decreased adhesion activity of the anti-sense-Dp71 cells could determine their deficiency to extend neurites. PMID:15706226

  1. Modulation of cell adhesion complexes by surface protein patterns.

    PubMed

    Pesen, Devrim; Haviland, David B

    2009-03-01

    Cell adhesion is an important process in several biological phenomena. To investigate the formation and organization of focal adhesions, we developed a patterning approach based on electron beam lithography. Nanodots (radius <1230 nm) and nanorings (inner radius <320 nm) of fibronectin (FN) were patterned on a K-Casein background. Intracellular vinculin immunofluorescence mirrored the FN nanopatterns. Atomic force microscopy showed that FN nanodots and nanorings organize the immediate cytoskeleton into straight fibrils and diverging fibril bundles, respectively. Our results suggest that a minimum of approximately 40 FN molecules is required for a cell to form a focal adhesion.

  2. Inhibition of focal adhesion kinase induces apoptosis in human osteosarcoma SAOS-2 cells.

    PubMed

    Wang, Jialiang; Zu, Jianing; Xu, Gongping; Zhao, Wei; Jinglong, Yan

    2014-02-01

    Focal adhesion kinase (FAK), a non-receptor tyrosine kinase protein, acts as an early modulator of integrin signaling cascade, regulating basic cellular functions. In transformed cells, unopposed FAK signaling has been considered to promote tumor growth, progression, and metastasis. The aim of this study was to assess the role of focal adhesion kinase in human osteosarcoma SAOS-2 cells. SAOS-2 cells were transfected with PGPU6/GFP/shNC, and PGPU6/GFP/FAK-334 (shRNA-334), respectively. Expression of FAK was detected by real-time PCR and western blots. MTT assay was used to examine changes in cell proliferation. Cell apoptosis was analyzed by flow cytometry. The expression of caspase-3,-7,-9 was measured by Western blots. The expression of FAK in SAOS-2 cells significantly decreased in shRNA-334 group contrast to the control group (P < 0.01). Cells proliferation was inhibited by shRNA-334 and shRNA-334 + cisplatin, and the effects were clearly enhanced when cells treated with the anticancer agents. The level of cell apoptosis in shRNA-334 and shRNA-334 + cisplatin group was higher than in the control group (P < 0.01). The current data support evidence that down-regulation of FAK could induce SAOS-2 apoptosis through the caspase-dependent cell death pathway. Inhibition of the kinases may be important for therapies designed to enhance the apoptosis in osteosarcoma.

  3. A novel role for junctional adhesion molecule-A in tumor proliferation: modulation by an anti-JAM-A monoclonal antibody.

    PubMed

    Goetsch, Liliane; Haeuw, Jean-François; Beau-Larvor, Charlotte; Gonzalez, Alexandra; Zanna, Laurence; Malissard, Martine; Lepecquet, Anne-Marie; Robert, Alain; Bailly, Christian; Broussas, Matthieu; Corvaia, Nathalie

    2013-03-15

    To identify new potential targets in oncology, functional approaches were developed using tumor cells as immunogens to select monoclonal antibodies targeting membrane receptors involved in cell proliferation. For that purpose cancer cells were injected into mice and resulting hybridomas were screened for their ability to inhibit cell proliferation in vitro. Based on this functional approach coupled to proteomic analysis, a monoclonal antibody specifically recognizing the human junctional adhesion molecule-A (JAM-A) was defined. Interestingly, compared to both normal and tumor tissues, we observed that JAM-A was mainly overexpressed on breast, lung and kidney tumor tissues. In vivo experiments demonstrated that injections of anti-JAM-A antibody resulted in a significant tumor growth inhibition of xenograft human tumors. Treatment with monoclonal antibody induced a decrease of the Ki67 expression and downregulated JAM-A levels. All together, our results show for the first time that JAM-A can interfere with tumor proliferation and suggest that JAM-A is a potential novel target in oncology. The results also demonstrate that a functional approach coupled to a robust proteomic analysis can be successful to identify new antibody target molecules that lead to promising new antibody-based therapies against cancers.

  4. The molecular mechanism of mediation of adsorbed serum proteins to endothelial cells adhesion and growth on biomaterials.

    PubMed

    Yang, Dayun; Lü, Xiaoying; Hong, Ying; Xi, Tingfei; Zhang, Deyuan

    2013-07-01

    To explore molecular mechanism of mediation of adsorbed proteins to cell adhesion and growth on biomaterials, this study examined endothelial cell adhesion, morphology and viability on bare and titanium nitride (TiN) coated nickel titanium (NiTi) alloys and chitosan film firstly, and then identified the type and amount of serum proteins adsorbed on the three surfaces by proteomic technology. Subsequently, the mediation role of the identified proteins to cell adhesion and growth was investigated with bioinformatics analyses, and further confirmed by a series of cellular and molecular biological experiments. Results showed that the type and amount of adsorbed serum proteins associated with cell adhesion and growth was obviously higher on the alloys than on the chitosan film, and these proteins mediated endothelial cell adhesion and growth on the alloys via four ways. First, proteins such as adiponectin in the adsorbed protein layer bound with cell surface receptors to generate signal transduction, which activated cell surface integrins through increasing intracellular calcium level. Another way, thrombospondin 1 in the adsorbed protein layer promoted TGF-β signaling pathway activation and enhanced integrins expression. The third, RGD sequence containing proteins such as fibronectin 1, vitronectin and thrombospondin 1 in the adsorbed protein layer bound with activated integrins to activate focal adhesion pathway, increased focal adhesion formation and actin cytoskeleton organization and mediated cell adhesion and spreading. In addition, the activated focal adhesion pathway promoted the expression of cell growth related genes and resulted in cell proliferation. The fourth route, coagulation factor II (F2) and fibronectin 1 in the adsorbed protein layer bound with cell surface F2 receptor and integrin, activated regulation of actin cytoskeleton pathway and regulated actin cytoskeleton organization.

  5. Charge displacement by adhesion and spreading of a cell.

    PubMed

    Svetlicić, V; Ivosević, N; Kovac, S; Zutić, V

    2001-01-01

    The potentiostatic control of surface charge density and interfacial tension of an electrode immersed in an aqueous electrolyte solution offers a possibility for direct studies of non-specific interactions in cell adhesion. Unicellular marine alga, Dunaliella tertiolecta (Chlorophyceae) of micrometer size and flexible cell envelope was used as a model cell and 0.1 M NaCl as supporting electrolyte. The dropping mercury electrode acted as in situ adhesion sensor and the electrochemical technique of chronoamperometry allowed measurement of the spread cell-electrode interface area and the distance of the closest approach of a cell. The adhesion and spreading of a single cell at the mercury electrode causes a displacement of counter-ions from the electrical double layer over a broad range of the positive and negative surface charge densities (from +16.0 to -8.2 microC/cm2). The flow of compensating current reflects the dynamics of adhesive contact formation and subsequent spreading of a cell. The adhesion and spreading rates are enhanced by the hydrodynamic regime of electrode's growing fluid interface. The distance of the closest approach of an adherent cell is smaller or equal to the distance of the outer Helmholz plane within the electrical double layer, i.e. 0.3-0.5 nm. There is a clear evidence of cell rupture for the potentials of maximum attraction as the area of the contact interface exceeded up to 100 times the cross-section area of a free cell.

  6. Photoinitiator-Free Synthesis of Endothelial Cell Adhesive and Enzymatically Degradable Hydrogels

    PubMed Central

    Jones, Derek R.; Marchant, Roger E.; von Recum, Horst; Gupta, Anirban Sen; Kottke-Marchant, Kandice

    2015-01-01

    We report on a photoinitiator-free synthetic method of incorporating bioactivity into poly(ethylene glycol) (PEG) hydrogels in order to control physical properties, enzymatic biodegradability and cell-specific adhesiveness of the polymer network, while eliminating the need for UV-mediated photopolymerization. To accomplish this, hydrogel networks were polymerized using Michael addition with four-arm PEG acrylate (10 kDa), using a collagenase sensitive peptide (CSP) as a crosslinker, and introducing an endothelial cell adhesive peptide either terminally (RGD) or attached to the crosslinking peptide sequence (CSP-RGD). The efficiency of the Michael addition reactions were determined by NMR and Ellman’s assay. Successful decoupling of cell adhesivity and physical properties was demonstrated by quantifying and comparing the swelling ratios and Young’s Moduli of various hydrogel formulations. Degradation profiles were established by incubating functionalized hydrogels in collagenase solutions (0.0 – 1.0 µg/mL), demonstrating that functionalized hydrogels degraded at a rate dependent upon collagenase concentration. Moreover, it was shown that the degradation rate was independent of CSP-RGD concentration. Cell attachment and proliferation on functionalized hydrogels were compared for various RGD concentrations, providing evidence that cell attachment and proliferation were directly related to relative amounts of the CSP-RGD combination peptide. An increase in cell viability was achieved using Michael addition techniques when compared to UV-polymerization, and was assessed by a LIVE/DEAD fluorescence assay. This photoinitiator-free method shows promise in creating hydrogel-based tissue engineering scaffolds allow for decoupled cell adhesivity and physical properties and that render greater cell viability. PMID:25462848

  7. Cadherin-mediated cell adhesion is critical for the closing of the mouse optic fissure.

    PubMed

    Chen, Shuyi; Lewis, Brandy; Moran, Andrea; Xie, Ting

    2012-01-01

    Coloboma is a congenital disease that contributes significantly to childhood blindness. It results from the failure in closing the optic fissure, a transient opening on the ventral side of the developing eye. Although human and mouse genetic studies have identified a number of genes associated with coloboma, the detailed cellular mechanisms underlying the optic fissure closure and coloboma formation remain largely undefined. N-cadherin-mediated cell adhesion has been shown to be important for the optic fissure closure in zebrafish, but it remains to be determined experimentally how cell-cell adhesions are involved in the mammalian optic fissure closing process. α-Catenin is required for cell adhesion mediated by all of the classic cadherin molecules, including N-cadherin. In this study, we used the Cre-mediated conditional knockout technique to specifically delete α-catenin from the developing mouse eye to show that it is required for the successful closing of the optic fissure. In α-catenin conditional mutant optic cups, the major cell fates, including the optic fissure margin, neural retina and retinal pigmented epithelium, are specified normally, and the retinal progenitor cells proliferate normally. However, adherens junctions components, including N-cadherin, β-catenin and filamentous actin, fail to accumulate on the apical side of α-catenin mutant retinal progenitor cells, where adherens junctions are normally abundant, and the organization of the neural retina and the optic fissure margin is disrupted. Finally, the α-catenin mutant retina gradually degenerates in the adult mouse eye. Therefore, our results show that α-catenin-mediated cell adhesion and cell organization are important for the fissure closure in mice, and further suggest that genes that regulate cell adhesion may underlie certain coloboma cases in humans.

  8. Single-cell force spectroscopy of pili-mediated adhesion

    NASA Astrophysics Data System (ADS)

    Sullan, Ruby May A.; Beaussart, Audrey; Tripathi, Prachi; Derclaye, Sylvie; El-Kirat-Chatel, Sofiane; Li, James K.; Schneider, Yves-Jacques; Vanderleyden, Jos; Lebeer, Sarah; Dufrêne, Yves F.

    2013-12-01

    Although bacterial pili are known to mediate cell adhesion to a variety of substrates, the molecular interactions behind this process are poorly understood. We report the direct measurement of the forces guiding pili-mediated adhesion, focusing on the medically important probiotic bacterium Lactobacillus rhamnosus GG (LGG). Using non-invasive single-cell force spectroscopy (SCFS), we quantify the adhesion forces between individual bacteria and biotic (mucin, intestinal cells) or abiotic (hydrophobic monolayers) surfaces. On hydrophobic surfaces, bacterial pili strengthen adhesion through remarkable nanospring properties, which - presumably - enable the bacteria to resist high shear forces under physiological conditions. On mucin, nanosprings are more frequent and adhesion forces larger, reflecting the influence of specific pili-mucin bonds. Interestingly, these mechanical responses are no longer observed on human intestinal Caco-2 cells. Rather, force curves exhibit constant force plateaus with extended ruptures reflecting the extraction of membrane nanotethers. These single-cell analyses provide novel insights into the molecular mechanisms by which piliated bacteria colonize surfaces (nanosprings, nanotethers), and offer exciting avenues in nanomedicine for understanding and controlling the adhesion of microbial cells (probiotics, pathogens).

  9. Titanium phosphate glass microcarriers induce enhanced osteogenic cell proliferation and human mesenchymal stem cell protein expression

    PubMed Central

    Lakhkar, Nilay J; M Day, Richard; Kim, Hae-Won; Ludka, Katarzyna; Mordan, Nicola J; Salih, Vehid; Knowles, Jonathan C

    2015-01-01

    In this study, we have developed 50- to 100-µm-sized titanium phosphate glass microcarriers (denoted as Ti5) that show enhanced proliferation of human mesenchymal stem cells and MG63 osteosarcoma cells, as well as enhanced human mesenchymal stem cell expression of bone differentiation markers, in comparison with commercially available glass microspheres at all time points. We also demonstrate that these microcarriers provide superior human mesenchymal stem cell proliferation with conventional Dulbecco’s Modified Eagle medium than with a specially developed commercial stem cell medium. The microcarrier proliferative capacity is revealed by a 24-fold increase in MG63 cell numbers in spinner flask bioreactor studies performed over a 7-day period, versus only a 6-fold increase in control microspheres under the same conditions; the corresponding values of Ti5 and control microspheres under static culture are 8-fold and 7-fold, respectively. The capability of guided osteogenic differentiation is confirmed by ELISAs for bone morphogenetic protein-2 and osteopontin, which reveal significantly greater expression of these markers, especially osteopontin, by human mesenchymal stem cells on the Ti5 microspheres than on the control. Scanning electron microscopy and confocal laser scanning microscopy images reveal favorable MG63 and human mesenchymal stem cell adhesion on the Ti5 microsphere surfaces. Thus, the results demonstrate the suitability of the developed microspheres for use as microcarriers in bone tissue engineering applications. PMID:26668711

  10. Mast cell mediators and peritoneal adhesion formation in the rat.

    PubMed

    Langer, J C; Liebman, S M; Monk, P K; Pelletier, G J

    1995-09-01

    We have previously shown that mast cell stabilization attenuates peritoneal adhesion formation in the rat. The present study investigated the mechanism of this protection. Adhesions were created in weanling rats using cecal scraping and application of 95% ethanol. Rats received specific blockers for the mast cell products histamine, serotonin (5HT), leukotriene D4, and platelet activating factor intraperitoneally 30 min before laparotomy and at the time of abdominal closure. Control animals received saline. Adhesions were assessed blindly 1 week later using a standardized scale. Adhesion formation was not affected by histamine blockade using combined mepyramine and ranitidine, 5-HT1 blockade using methysergide, 5-HT3 blockade using ondansetron, leukotriene D4 blockade using MK-571, or platelet activating factor blockade using WEB-2086. However, blockade of the 5-HT2 receptor using ketanserin resulted in significant dose-dependent attenuation of adhesions compared to saline. These data suggest that mast cells mediate peritoneal adhesion formation in the rat through release of serotonin acting on 5HT2 receptors. Further understanding of this process may lead to new strategies for the prevention of postoperative adhesions.

  11. Quantifying Cell Adhesion through Impingement of a Controlled Microjet

    PubMed Central

    Visser, Claas Willem; Gielen, Marise V.; Hao, Zhenxia; Le Gac, Séverine; Lohse, Detlef; Sun, Chao

    2015-01-01

    The impingement of a submerged, liquid jet onto a cell-covered surface allows assessing cell attachment on surfaces in a straightforward and quantitative manner and in real time, yielding valuable information on cell adhesion. However, this approach is insufficiently characterized for reliable and routine use. In this work, we both model and measure the shear stress exerted by the jet on the impingement surface in the micrometer-domain, and subsequently correlate this to jet-induced cell detachment. The measured and numerically calculated shear stress data are in good agreement with each other, and with previously published values. Real-time monitoring of the cell detachment reveals the creation of a circular cell-free area upon jet impingement, with two successive detachment regimes: 1), a dynamic regime, during which the cell-free area grows as a function of both the maximum shear stress exerted by the jet and the jet diameter; followed by 2), a stationary regime, with no further evolution of the cell-free area. For the latter regime, which is relevant for cell adhesion strength assessment, a relationship between the jet Reynolds number, the cell-free area, and the cell adhesion strength is proposed. To illustrate the capability of the technique, the adhesion strength of HeLa cervical cancer cells is determined ((34 ± 14) N/m2). Real-time visualization of cell detachment in the dynamic regime shows that cells detach either cell-by-cell or by collectively (for which intact parts of the monolayer detach as cell sheets). This process is dictated by the cell monolayer density, with a typical threshold of (1.8 ± 0.2) × 109 cells/m2, above which the collective behavior is mostly observed. The jet impingement method presents great promises for the field of tissue engineering, as the influence of both the shear stress and the surface characteristics on cell adhesion can be systematically studied. PMID:25564849

  12. Proliferation status defines functional properties of endothelial cells.

    PubMed

    Lipps, Christoph; Badar, Muhammad; Butueva, Milada; Dubich, Tatyana; Singh, Vivek Vikram; Rau, Sophie; Weber, Axel; Kracht, Michael; Köster, Mario; May, Tobias; Schulz, Thomas F; Hauser, Hansjörg; Wirth, Dagmar

    2017-04-01

    Homeostasis of solid tissue is characterized by a low proliferative activity of differentiated cells while special conditions like tissue damage induce regeneration and proliferation. For some cell types it has been shown that various tissue-specific functions are missing in the proliferating state, raising the possibility that their proliferation is not compatible with a fully differentiated state. While endothelial cells are important players in regenerating tissue as well as in the vascularization of tumors, the impact of proliferation on their features remains elusive. To examine cell features in dependence of proliferation, we established human endothelial cell lines in which proliferation is tightly controlled by a doxycycline-dependent, synthetic regulatory unit. We observed that uptake of macromolecules and establishment of cell-cell contacts was more pronounced in the growth-arrested state. Tube-like structures were formed in vitro in both proliferating and non-proliferating conditions. However, functional vessel formation upon transplantation into immune-compromised mice was restricted to the proliferative state. Kaposi's sarcoma-associated herpes virus (KSHV) infection resulted in reduced expression of endothelial markers. Upon transplantation of infected cells, drastic differences were observed: proliferation arrested cells acquired a high migratory activity while the proliferating counterparts established a tumor-like phenotype, similar to Kaposi Sarcoma lesions. The study gives evidence that proliferation governs endothelial functions. This suggests that several endothelial functions are differentially expressed during angiogenesis. Moreover, since proliferation defines the functional properties of cells upon infection with KSHV, this process crucially affects the fate of virus-infected cells.

  13. A novel culture platform for fast proliferation of human annulus fibrosus cells.

    PubMed

    Xiao, Li; Ding, Mengmeng; Saadoon, Osama; Vess, Eric; Fernandez, Andrew; Zhao, Ping; Jin, Li; Li, Xudong

    2017-02-01

    Tissue engineering provides a promising approach to treat degenerative disc disease, which usually requires a large quantity of seed cells. A simple and reliable in vitro culture system to expand seed cells in a timely fashion is necessary to implement the application clinically. Here, we sought to establish a cost-effective culture system for expanding human annulus fibrosus cells using extracellular matrix (ECM) proteins as culture substrates. Cells were cultured onto a plastic surface coated with various types of ECMs, including fibronectin, vitronectin, collagen type I, gelatin and cell-free matrix deposited by human nucleus pulposus cells. AF cell morphology, growth, adhesion and phenotype (anabolic and catabolic markers) were assessed by microscopy, real-time RT-PCR, western blotting, zymography, immunofluorescence staining and biochemical assays. Fibronectin, collagen and gelatin promoted cell proliferation and adhesion in a dose-dependent manner. Fibronectin elevated mRNA expression of proteoglycan and enhanced glycosaminoglycan production. Both collagen and gelatin increased protein expression of type II collagen. Consistent with increased cell adhesion, collagen and fibronectin promoted formation of focal adhesion complexes in the cell-matrix junction, suggesting enhanced binding of the actin network with both ECM substrates. On the other hand, fibronectin, collagen and gelatin decreased expression of matrix metalloproteinase-2 and matrix metalloproteinase-9 in media. Finally, a mixture of fibronectin (1.7 μg/mL) and collagen (1.3 μg/mL) was identified as the most promising in vitro culture substrate system in promoting proliferation and maintaining anabolic-catabolic balance. Our method provides a simple and cost-effective platform for tissue engineering applications in intervertebral disc research.

  14. Inhibition by Tyroserleutide (YSL) on the Invasion and Adhesion of the Mouse Melanoma Cell

    PubMed Central

    Yao, Zhi; Che, Xu-chun; Lu, Rong; Zheng, Min-na; Zhu, Zhi-feng; Li, Jin-ping; Jian, Xu; Shi, Lin-xi; Liu, Jun-yan; Gao, Wen-yuan

    2007-01-01

    Tyroserleutide (YSL) is an active, low-molecular-weight polypeptide, comprised of three amino acids, that has shown antitumor effects on human hepatocarcinoma BEL-7402 in vitro and in vivo. In this study, we evaluated the inhibition of YSL on invasion and adhesion of the mouse B16-F10 melanoma cell line by injecting B16-F10 cells into the tail veins of C57BL/6 mice to establish an experimental lung metastasis model. YSL inhibited B16-F10 cell metastasis to lung, reducing the number and area of metastasis lesions. When we treated B16-F10 cells with YSL (0.01, 0.1, 1, 10, or 100 μg/mL) in vitro, we found that YSL inhibited the proliferation of B16-F10 cells with a 28.11% rate of inhibition. YSL significantly decreased the adhesiveness of B16-F10 cells to Matrigel with a 29.15% inhibition rate; YSL also significantly inhibited the invasion of B16-F10 cells, producing an inhibition of 35.31%. By analyses with Western blot and real-time RT-PCR, we found that YSL markedly inhibited the expression of ICAM-1 in B16-F10 cells. These data suggest that YSL inhibits the growth, invasion, and adhesion of B16-F10 cells. PMID:17515953

  15. Inhibition of cell adhesion by phosphorylated Ezrin/Radixin/Moesin.

    PubMed

    Tachibana, Kouichi; Haghparast, Seyed Mohammad Ali; Miyake, Jun

    2015-01-01

    Altered phosphorylation status of the C-terminal Thr residues of Ezrin/Radixin/Moesin (ERM) is often linked to cell shape change. To determine the role of phophorylated ERM, we modified phosphorylation status of ERM and investigated changes in cell adhesion and morphology. Treatment with Calyculin-A (Cal-A), a protein phosphatase inhibitor, dramatically augmented phosphorylated ERM (phospho-ERM). Cal-A-treatment or expression of phospho-mimetic Moesin mutant (Moesin-TD) induced cell rounding in adherent cells. Moreover, reattachment of detached cells to substrate was inhibited by either treatment. Phospho-ERM, Moesin-TD and actin cytoskeleton were observed at the plasma membrane of such round cells. Augmented cell surface rigidity was also observed in both cases. Meanwhile, non-adherent KG-1 cells were rather rich in phospho-ERM. Treatment with Staurosporine, a protein kinase inhibitor that dephosphorylates phospho-ERM, up-regulated the integrin-dependent adhesion of KG-1 cells to substrate. These findings strongly suggest the followings: (1) Phospho-ERM inhibit cell adhesion, and therefore, dephosphorylation of ERM proteins is essential for cell adhesion. (2) Phospho-ERM induce formation and/or maintenance of spherical cell shape. (3) ERM are constitutively both phosphorylated and dephosphorylated in cultured adherent and non-adherent cells.

  16. Aging affects initiation and continuation of T cell proliferation.

    PubMed

    Jiang, Jiu; Gross, Diara; Elbaum, Philip; Murasko, Donna M

    2007-04-01

    Aging is associated with a decline in immune responses, particularly within the T cell compartment. While the expansion of specific T cells in response to virus infections is consistently decreased in aged mice, the differences in T cell activation between young and aged mice as demonstrated in each round of proliferation remain poorly defined. In the present study, we utilized the T cell mitogen, ConA, to explore if fewer T cells of aged mice initiate proliferation upon mitogen stimulation or if similar numbers of T cells of aged mice begin proliferation but undergo fewer rounds of division. We also examined whether these age-associated changes in proliferation are reflected by differences in T cell activation by comparing activation markers (CD25, CD69, CD44, and CD62L) on T cells of young and aged mice at each round of proliferation. Not only was the kinetics of the expression of these markers greatly different between young and aged mice on the entire CD8 T cell population, but also at each round of proliferation. Our results demonstrate that a larger percentage of CD8 T cells of aged mice do not proliferate at all upon stimulation. Of the CD8 T cells of aged mice that do proliferate, a larger percentage start later and stop sooner. These results suggest that multiple levels of alteration may need to be considered when trying to maximize the immune response of aged individuals.

  17. Adhesion and migration of cells responding to microtopography.

    PubMed

    Estévez, Maruxa; Martínez, Elena; Yarwood, Stephen J; Dalby, Matthew J; Samitier, Josep

    2015-05-01

    It is known that cells respond strongly to microtopography. However, cellular mechanisms of response are unclear. Here, we study wild-type fibroblasts responding to 25 µm(2) posts and compare their response to that of FAK(-/-) fibroblasts and fibroblasts with PMA treatment to stimulate protein kinase C (PKC) and the small g-protein Rac. FAK knockout cells modulated adhesion number and size in a similar way to cells on topography; that is, they used more, smaller adhesions, but migration was almost completely stalled demonstrating the importance of FAK signaling in contact guidance and adhesion turnover. Little similarity, however, was observed to PKC stimulated cells and cells on the topography. Interestingly, with PKC stimulation the cell nuclei became highly deformable bringing focus on these surfaces to the study of metastasis. Surfaces that aid the study of cellular migration are important in developing understanding of mechanisms of wound healing and repair in aligned tissues such as ligament and tendon.

  18. Cell adhesion in zebrafish embryos is modulated by March 8.

    PubMed

    Kim, Mi Ha; Rebbert, Martha L; Ro, Hyunju; Won, Minho; Dawid, Igor B

    2014-01-01

    March 8 is a member of a family of transmembrane E3 ubiquitin ligases that have been studied mostly for their role in the immune system. We find that March 8 is expressed in the zebrafish egg and early embryo, suggesting a role in development. Both knock-down and overexpression of March 8 leads to abnormal development. The phenotype of zebrafish embryos and Xenopus animal explants overexpressing March 8 implicates impairment of cell adhesion as a cause of the effect. In zebrafish embryos and in cultured cells, overexpression of March 8 leads to a reduction in the surface levels of E-cadherin, a major cell-cell adhesion molecule. Experiments in cell culture further show that E-cadherin can be ubiquitinated by March 8. On the basis of these observations we suggest that March 8 functions in the embryo to modulate the strength of cell adhesion by regulating the localization of E-cadherin.

  19. Adhesion and Invasion of Gastric Mucosa Epithelial Cells by Helicobacter pylori

    PubMed Central

    Huang, Ying; Wang, Qi-long; Cheng, Dan-dan; Xu, Wen-ting; Lu, Nong-hua

    2016-01-01

    Helicobacter pylori is the main pathogenic bacterium involved in chronic gastritis and peptic ulcer and a class 1 carcinogen in gastric cancer. Current research focuses on the pathogenicity of H. pylori and the mechanism by which it colonizes the gastric mucosa. An increasing number of in vivo and in vitro studies demonstrate that H. pylori can invade and proliferate in epithelial cells, suggesting that this process might play an important role in disease induction, immune escape and chronic infection. Therefore, to explore the process and mechanism of adhesion and invasion of gastric mucosa epithelial cells by H. pylori is particularly important. This review examines the relevant studies and describes evidence regarding the adhesion to and invasion of gastric mucosa epithelial cells by H. pylori. PMID:27921009

  20. Twist1-positive epithelial cells retain adhesive and proliferative capacity throughout dissemination

    PubMed Central

    Shamir, Eliah R.; Coutinho, Kester; Georgess, Dan; Auer, Manfred

    2016-01-01

    ABSTRACT Dissemination is the process by which cells detach and migrate away from a multicellular tissue. The epithelial-to-mesenchymal transition (EMT) conceptualizes dissemination in a stepwise fashion, with downregulation of E-cadherin leading to loss of intercellular junctions, induction of motility, and then escape from the epithelium. This gain of migratory activity is proposed to be mutually exclusive with proliferation. We previously developed a dissemination assay based on inducible expression of the transcription factor Twist1 and here utilize it to characterize the timing and dynamics of intercellular adhesion, proliferation and migration during dissemination. Surprisingly, Twist1+ epithelium displayed extensive intercellular junctions, and Twist1– luminal epithelial cells could still adhere to disseminating Twist1+ cells. Although proteolysis and proliferation were both observed throughout dissemination, neither was absolutely required. Finally, Twist1+ cells exhibited a hybrid migration mode; their morphology and nuclear deformation were characteristic of amoeboid cells, whereas their dynamic protrusive activity, pericellular proteolysis and migration speeds were more typical of mesenchymal cells. Our data reveal that epithelial cells can disseminate while retaining competence to adhere and proliferate. PMID:27402962

  1. Possible involvement of queuine in regulation of cell proliferation.

    PubMed

    Pathak, Chandramani; Jaiswal, Yogesh K; Vinayak, Manjula

    2007-01-01

    An increase in cell number is one of the most prominent characteristics of cancer cells. This may be caused by an increase in cell proliferation or decrease in cell death. Queuine is one of the modified base which is found at first anticodon position of specific tRNAs. It is ubiquitously present throughout the living system except mycoplasma and yeast. The tRNAs of Q-family are completely modified to Q-tRNAs in terminally differentiated somatic cells, however hypomodification of Q-tRNA is closely associated with cell proliferation and malignancy. Queuine participates at various cellular functions such as regulation of cell proliferation, cell signaling and alteration in the expression of growth associated proto-oncogenes. Like other proto-oncogenes bcl2 is known to involve in cell survival by inhibiting apoptosis. Queuine or Q-tRNA is suggested to inhibit cell proliferation but the mechanism of regulation of cell proliferation by queuine or Q-tRNA is not well understood. Therefore, in the present study regulation in cell proliferation by queuine in vivo and in vitro as well as the expression of cell death regulatory protein Bcl2 are investigated. For this DLAT cancerous mouse, U87 cell line and HepG2 cell line are treated with different concentrations of queuine and the effect of queuine on cell proliferation and apoptosis are studied. The results indicate that queuine down regulates cell proliferation and expression of Bcl2 protein, suggesting that queuine promotes cell death and participates in the regulation of cell proliferation.

  2. Proteomic Profiling of Neuroblastoma Cells Adhesion on Hyaluronic Acid-Based Surface for Neural Tissue Engineering

    PubMed Central

    Chen, Ko-Chin; Chiang, Pei-Wen; Chu, Pei-Yu; Lu, Yi-Shan; Yuan, Cheng-Hui; Wang, Ming-Chen; Lin, Chia-Yang; Huang, Ying-Fong; Jong, Shiang-Bin; Lin, Po-Chiao

    2016-01-01

    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. PMID:28053978

  3. Proteomic Profiling of Neuroblastoma Cells Adhesion on Hyaluronic Acid-Based Surface for Neural Tissue Engineering.

    PubMed

    Yang, Ming-Hui; Chen, Ko-Chin; Chiang, Pei-Wen; Chung, Tze-Wen; Chen, Wan-Jou; Chu, Pei-Yu; Chen, Sharon Chia-Ju; Lu, Yi-Shan; Yuan, Cheng-Hui; Wang, Ming-Chen; Lin, Chia-Yang; Huang, Ying-Fong; Jong, Shiang-Bin; Lin, Po-Chiao; Tyan, Yu-Chang

    2016-01-01

    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.

  4. Skin cell proliferation stimulated by microneedles.

    PubMed

    Liebl, Horst; Kloth, Luther C

    2012-03-01

    A classical wound may be defined as a disruption of tissue integrity. Wounds, caused by trauma from accidents or surgery, that close via secondary intention rely on the biological phases of healing, i.e., hemostasis, inflammation, proliferation, and remodeling (HIPR). Depending on the wound type and severity, the inflammation phase begins immediately after injury and may last for an average of 7-14 days. Concurrent with the inflammation phase or slightly delayed, cell proliferation is stimulated followed by the activation of the remodeling (maturation) phase. The latter phase can last as long as 1 year or more, and the final healed state is represented by a scar tissue, a cross-linked collagen formation that usually aligns collagen fibers in a single direction. One may assume that skin microneedling that involves the use of dozens or as many as 200 needles that limit penetration to 1.5 mm over 1 cm(2) of skin would cause trauma and bleeding followed by the classical HIPR. However, this is not the case or at least the HIPR phases are significantly curtailed and healing never ends in a scar formation. Conversely dermabrasion used in aesthetic medicine for improving skin quality is based on "ablation" (destruction or wounding of superficial skin layers), which requires several weeks for healing that involves formation of new skin layers. Such procedures provoke an acute inflammatory response. We believe that a less intense inflammatory response occurs following microneedle perforation of the skin. However, the mechanism of action of microneedling appears to be different. Here we review the potential mechanisms by which microneedling of the skin facilitates skin repair without scarring after the treatment of superficial burns, acne, hyperpigmentation, and the non-advancing periwound skin surrounding the chronic ulcerations of the integument.

  5. Skin Cell Proliferation Stimulated by Microneedles

    PubMed Central

    Liebl, Horst; Kloth, Luther C.

    2012-01-01

    A classical wound may be defined as a disruption of tissue integrity. Wounds, caused by trauma from accidents or surgery, that close via secondary intention rely on the biological phases of healing, i.e., hemostasis, inflammation, proliferation, and remodeling (HIPR). Depending on the wound type and severity, the inflammation phase begins immediately after injury and may last for an average of 7–14 days. Concurrent with the inflammation phase or slightly delayed, cell proliferation is stimulated followed by the activation of the remodeling (maturation) phase. The latter phase can last as long as 1 year or more, and the final healed state is represented by a scar tissue, a cross-linked collagen formation that usually aligns collagen fibers in a single direction. One may assume that skin microneedling that involves the use of dozens or as many as 200 needles that limit penetration to 1.5 mm over 1 cm2 of skin would cause trauma and bleeding followed by the classical HIPR. However, this is not the case or at least the HIPR phases are significantly curtailed and healing never ends in a scar formation. Conversely dermabrasion used in aesthetic medicine for improving skin quality is based on “ablation” (destruction or wounding of superficial skin layers), which requires several weeks for healing that involves formation of new skin layers. Such procedures provoke an acute inflammatory response. We believe that a less intense inflammatory response occurs following microneedle perforation of the skin. However, the mechanism of action of microneedling appears to be different. Here we review the potential mechanisms by which microneedling of the skin facilitates skin repair without scarring after the treatment of superficial burns, acne, hyperpigmentation, and the non-advancing periwound skin surrounding the chronic ulcerations of the integument. PMID:24527373

  6. Peptide array-based screening of human mesenchymal stem cell-adhesive peptides derived from fibronectin type III domain

    SciTech Connect

    Okochi, Mina; Nomura, Shigeyuki; Kaga, Chiaki; Honda, Hiroyuki

    2008-06-20

    Human mesenchymal stem cell-adhesive peptides were screened based on the amino acid sequence of fibronectin type III domain 8-11 (FN-III{sub 8-11}) using a peptide array synthesized by the Fmoc-chemistry. Using hexameric peptide library of FN-III{sub 8-11} scan, we identified the ALNGR (Ala-Leu-Asn-Gly-Arg) peptide that induced cell adhesion as well as RGDS (Arg-Gly-Asp-Ser) peptide. After incubation for 2 h, approximately 68% of inoculated cells adhere to the ALNGR peptide disk. Adhesion inhibition assay with integrin antibodies showed that the ALNGR peptide interacts with integrin {beta}1 but not with {alpha}v{beta}3, indicating that the receptors for ALNGR are different from RGDS. Additionally, the ALNGR peptide expressed cell specificities for adhesion: cell adhesion was promoted for fibroblasts but not for keratinocytes or endotherial cells. The ALNGR peptide induced cell adhesion and promoted cell proliferation without changing its property. It is therefore useful for the construction of functional biomaterials.

  7. Discoidin domain receptor 2 (DDR2) regulates proliferation of endochondral cells in mice.

    PubMed

    Kawai, Ikuma; Hisaki, Tomoka; Sugiura, Koji; Naito, Kunihiko; Kano, Kiyoshi

    2012-10-26

    Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase that is activated by fibrillar collagens. DDR2 regulates cell proliferation, cell adhesion, migration, and extracellular matrix remodeling. The decrement of endogenous DDR2 represses osteoblastic marker gene expression and osteogenic differentiation in murine preosteoblastic cells, but the functions of DDR2 in chondrogenic cellular proliferation remain unclear. To better understand the role of DDR2 signaling in cellular proliferation in endochondral ossification, we inhibited Ddr2 expression via the inhibitory effect of miRNA on Ddr2 mRNA (miDdr2) and analyzed the cellular proliferation and differentiation in the prechondrocyte ATDC5 cell lines. To investigate DDR2's molecular role in endochondral cellular proliferation in vivo, we also produced transgenic mice in which the expression of truncated, kinase dead (KD) DDR2 protein is induced, and evaluated the DDR2 function in cellular proliferation in chondrocytes. Although the miDdr2-transfected ATDC5 cell lines retained normal differentiation ability, DDR2 reduction finally promoted cellular proliferation in proportion to the decreasing ratio of Ddr2 expression, and it also promoted earlier differentiation to cartilage cells by insulin induction. The layer of hypertrophic chondrocytes in KD Ddr2 transgenic mice was not significantly thicker than that of normal littermates, but the layer of proliferative chondrocytes in KD-Ddr2 transgenic mice was significantly thicker than that of normal littermates. Taken together, our data demonstrated that DDR2 might play a local and essential role in the proliferation of chondrocytes.

  8. Combinatorial plasma polymerization approach to produce thin films for testing cell proliferation.

    PubMed

    Antonini, V; Torrengo, S; Marocchi, L; Minati, L; Dalla Serra, M; Bao, G; Speranza, G

    2014-01-01

    Plasma enhanced physical vapor depositions are extensively used to fabricate substrates for cell culture applications. One peculiarity of the plasma processes is the possibility to deposit thin films with reproducible chemical and physical properties. In the present work, a combinatorial plasma polymerization process was used to deposit thin carbon based films to promote cell adhesion, in the interest of testing cell proliferation as a function of the substrate chemical properties. Peculiarity of the combinatorial approach is the possibility to produce in just one deposition experiment, a set of surfaces of varying chemical moieties by changing the precursor composition. A full characterization of the chemical, physical and thermodynamic properties was performed for each set of the synthesized surfaces. X-ray photoelectron spectroscopy was used to measure the concentration of carboxyl, hydroxyl and amine functional groups on the substrate surfaces. A perfect linear trend between polar groups' density and precursors' concentration was found. Further analyses reveled that also contact angles and the correspondent surface energies of all deposited thin films are linearly dependent on the precursor concentration. To test the influence of the surface composition on the cell adhesion and proliferation, two cancer cell lines were utilized. The cell viability was assessed after 24 h and 48 h of cell culture. Experiments show that we are able to control the cell adhesion and proliferation by properly changing the thin film deposition conditions i.e. the concentration and the kind of chemical moiety on the substrate surface. The results also highlight that physical and chemical factors of biomaterial surface, including surface hydrophobicity and free energy, chemical composition, and topography, can altered cell attachment.

  9. Controlling the cell adhesion property of silk films by graft polymerization.

    PubMed

    Dhyani, Vartika; Singh, Neetu

    2014-04-09

    We report here a graft polymerization method to improve the cell adhesion property of Bombyx mori silk fibroin films. B. mori silk has evolved as a promising material for tissue engineering because of its biocompatibility and biodegradability. However, silk's hydrophobic character makes cell adhesion and proliferation difficult. Also, the lack of sufficient reactive amino acid residues makes biofunctionalization via chemical modification challenging. Our study describes a simple method that provides increased chemical handles for tuning of the surface chemistry of regenerated silk films (SFs), thus allowing manipulation of their bioactivity. By grafting pAAc and pHEMA via plasma etching, we have increased carboxylic acid and hydroxyl groups on silk, respectively. These modifications allowed us to tune the hydrophilicity of SFs and provide functional groups for bioconjugation. Our strategy also allowed us to develop silk-based surface coatings, where spatial control over cell adhesion can be achieved. This control over cell adhesion in a particular region of the SFs is difficult to obtain via existing methods of modifying the silk fibroin instead of the SF surface. Thus, our strategy will be a valuable addition to the toolkit of biofunctionalization for enhancing SFs' tissue engineering applications.

  10. β-Lactoglobulin Influences Human Immunity and Promotes Cell Proliferation

    PubMed Central

    Tai, Chun San; Chen, Yi Yun

    2016-01-01

    β-Lactoglobulin (LG) is suspected to enhance or modulate human immune responses. Moreover, LG is also hypothesized to increase human cell proliferation. However, these potential functions of LG have not been directly or thoroughly addressed. In this study, we demonstrated that LG is a potent stimulator of cell proliferation using a hybridoma cell (a splenocyte fused with a myeloma cell) model. LG's ability to promote cell proliferation was lost when the protein is denatured. To further investigate the influence of LG's conformation on cell proliferation, we chemically modified LG by either carboxymethylation (CM) or acetylation and observed significantly reduced cell proliferation when the protein structure was altered. Furthermore, we proved that LG enhances cell proliferation via receptor-mediated membrane IgM receptor. These data indicated that nondenatured LG is the major component in milk that modulates cell proliferation. Collectively, our study showed that LG plays a key role in enhancing immune responses by promoting cell proliferation through IgM receptor. PMID:27957499

  11. Satellite cell proliferation in adult skeletal muscle

    NASA Technical Reports Server (NTRS)

    Booth, Frank W. (Inventor); Thomason, Donald B. (Inventor); Morrison, Paul R. (Inventor); Stancel, George M. (Inventor)

    1995-01-01

    Novel methods of retroviral-mediated gene transfer for the in vivo corporation and stable expression of eukaryotic or prokaryotic foreign genes in tissues of living animals is described. More specifically, methods of incorporating foreign genes into mitotically active cells are disclosed. The constitutive and stable expression of E. coli .beta.-galactosidase gene under the promoter control of the Moloney murine leukemia virus long terminal repeat is employed as a particularly preferred embodiment, by way of example, establishes the model upon which the incorporation of a foreign gene into a mitotically-active living eukaryotic tissue is based. Use of the described methods in therapeutic treatments for genetic diseases, such as those muscular degenerative diseases, is also presented. In muscle tissue, the described processes result in genetically-altered satellite cells which proliferate daughter myoblasts which preferentially fuse to form a single undamaged muscle fiber replacing damaged muscle tissue in a treated animal. The retroviral vector, by way of example, includes a dystrophin gene construct for use in treating muscular dystrophy. The present invention also comprises an experimental model utilizable in the study of the physiological regulation of skeletal muscle gene expression in intact animals.

  12. Hyperbranched Polyester Hydrogels with Controlled Drug Release and Cell Adhesion Properties

    PubMed Central

    Zhang, Hongbin; Patel, Alpesh; Gaharwar, Akhilesh K.; Mihaila, Silvia M.; Iviglia, Giorgio; Mukundan, Shilpaa; Bae, Hojae; Yang, Huai; Khademhosseini, Ali

    2013-01-01

    Hyperbranched polyesters (HPE) have a high efficiency to encapsulate bioactive agents, including drugs, genes and proteins, due to their globe-like nanostructure. However, the use of these highly branched polymeric systems for tissue engineering applications has not been broadly investigated. Here, we report synthesis and characterization of photocrosslinkable HPE hydrogels with sustained drug release characteristics for cellular therapies. These HPE can encapsulate hydrophobic drug molecules within the HPE cavities, due to the presence of hydrophobic inner structure that is otherwise difficult to achieve in conventional hydrogels. The functionalization of HPE with photocrosslinkable acrylate moieties renders the formation of hydrogels with highly porous interconnected structure, and mechanically tough network. The compressive modulus of HPE hydrogels was tunable by changing the crosslinking density. The feasibility of using these HPE networks for cellular therapies was investigated by evaluating cell adhesion, spreading and proliferation on hydrogel surface. Highly crosslinked and mechanically stiff HPE hydrogels have higher cell adhesion, spreading, proliferation compared to soft and complaint HPE hydrogels. Overall, we showed that hydrogels made from HPE could be used for biomedical applications that require control cell adhesion and control release of hydrophobic clues. PMID:23394067

  13. Biofunctionalized 3-D Carbon Nano-Network Platform for Enhanced Fibroblast Cell Adhesion

    PubMed Central

    Chowdhury, A. K. M. Rezaul Haque; Tavangar, Amirhossein; Tan, Bo; Venkatakrishnan, Krishnan

    2017-01-01

    Carbon nanomaterials have been investigated for various biomedical applications. In most cases, however, these nanomaterials must be functionalized biologically or chemically due to their biological inertness or possible cytotoxicity. Here, we report the development of a new carbon nanomaterial with a bioactive phase that significantly promotes cell adhesion. We synthesize the bioactive phase by introducing self-assembled nanotopography and altered nano-chemistry to graphite substrates using ultrafast laser. To the best of our knowledge, this is the first time that such a cytophilic bio-carbon is developed in a single step without requiring subsequent biological/chemical treatments. By controlling the nano-network concentration and chemistry, we develop platforms with different degrees of cell cytophilicity. We study quantitatively and qualitatively the cell response to nano-network platforms with NIH-3T3 fibroblasts. The findings from the in vitro study indicate that the platforms possess excellent biocompatibility and promote cell adhesion considerably. The study of the cell morphology shows a healthy attachment of cells with a well-spread shape, overextended actin filaments, and morphological symmetry, which is indicative of a high cellular interaction with the nano-network. The developed nanomaterial possesses great biocompatibility and considerably stimulates cell adhesion and subsequent cell proliferation, thus offering a promising path toward engineering various biomedical devices. PMID:28287138

  14. Biofunctionalized 3-D Carbon Nano-Network Platform for Enhanced Fibroblast Cell Adhesion

    NASA Astrophysics Data System (ADS)

    Chowdhury, A. K. M. Rezaul Haque; Tavangar, Amirhossein; Tan, Bo; Venkatakrishnan, Krishnan

    2017-03-01

    Carbon nanomaterials have been investigated for various biomedical applications. In most cases, however, these nanomaterials must be functionalized biologically or chemically due to their biological inertness or possible cytotoxicity. Here, we report the development of a new carbon nanomaterial with a bioactive phase that significantly promotes cell adhesion. We synthesize the bioactive phase by introducing self-assembled nanotopography and altered nano-chemistry to graphite substrates using ultrafast laser. To the best of our knowledge, this is the first time that such a cytophilic bio-carbon is developed in a single step without requiring subsequent biological/chemical treatments. By controlling the nano-network concentration and chemistry, we develop platforms with different degrees of cell cytophilicity. We study quantitatively and qualitatively the cell response to nano-network platforms with NIH-3T3 fibroblasts. The findings from the in vitro study indicate that the platforms possess excellent biocompatibility and promote cell adhesion considerably. The study of the cell morphology shows a healthy attachment of cells with a well-spread shape, overextended actin filaments, and morphological symmetry, which is indicative of a high cellular interaction with the nano-network. The developed nanomaterial possesses great biocompatibility and considerably stimulates cell adhesion and subsequent cell proliferation, thus offering a promising path toward engineering various biomedical devices.

  15. Osteopontin improves adhesion and migration of human primary renal cortical epithelial cells during wound healing

    PubMed Central

    Wu, Jinfeng; Wang, Zuolin

    2016-01-01

    The aim of the present study was to investigate the effect of osteopontin (OPN) on adhesion and migration in human primary renal cortical epithelial cells during wound healing and Transwell assays. MTT assay was used to examine the cell viability and western blot analysis was used to examine the expression of cytoskeletal proteins and cell adhesion molecules. The results showed that overexpression of OPN had positive effects on the viability, proliferation, adhesion and migration of the human primary renal cortical epithelial cells. In addition, the integrity of the cell membrane and cytoskeleton of the epithelial cells was negatively affected by knockdown of OPN expression. The Transwell migration and a wound healing assays performed using OPN-knockdown cells suggested that OPN had a significant impact on cell migration (P=0.0421) and wound healing (P=0.0333). Therefore, OPN may be a potential target for the therapeutic modulation of skin repair to improve the healing rate and quality of wound healing. PMID:28101213

  16. Cell-cell and cell-ECM adhesions cooperate to organize actomyosin networks and maintain force transmission during Dorsal Closure.

    PubMed

    Goodwin, Katharine; Lostchuck, Emily E; Cramb, Kaitlyn M L; Zulueta-Coarasa, Teresa; Fernandez-Gonzalez, Rodrigo; Tanentzapf, Guy

    2017-03-22

    Tissue morphogenesis relies on the coordinated action of actin networks, cell-cell adhesions, and cell-ECM adhesions. Such coordination can be achieved through crosstalk between cell-cell and cell-ECM adhesions. Drosophila Dorsal Closure (DC), a morphogenetic process wherein an extra-embryonic tissue called the amnioserosa contracts and ingresses to close a discontinuity in the dorsal epidermis of the embryo, requires both cell-cell and cell-ECM adhesions. However, whether the function of these two types of adhesion is coordinated during DC is not known. Here, we analyzed possible interdependence between cell-cell and cell-ECM adhesions during DC, and its effect on the actomyosin network. We find that loss of cell-ECM adhesion results in aberrant distributions of cadherin-mediated adhesions and actin networks in the amnioserosa; and subsequent disruption of myosin recruitment and dynamics. Moreover, loss of cell-cell adhesion caused an upregulation of cell-ECM adhesion, leading to reduced cell deformation and force transmission across amnioserosa cells. Our results show how interdependence between cell-cell and cell-ECM adhesions is important in regulating cell behaviours, force generation and force transmission critical for tissue morphogenesis.

  17. Quantification of Depletion-Induced Adhesion of Red Blood Cells

    NASA Astrophysics Data System (ADS)

    Steffen, P.; Verdier, C.; Wagner, C.

    2013-01-01

    Red blood cells (RBCs) are known to form aggregates in the form of rouleaux due to the presence of plasma proteins under physiological conditions. The formation of rouleaux can also be induced in vitro by the addition of macromolecules to the RBC suspension. Current data on the adhesion strength between red blood cells in their natural discocyte shapes mostly originate from indirect measurements such as flow chamber experiments, but data is lacking at the single cell level. Here, we present measurements on the dextran-induced aggregation of red blood cells using atomic force microscopy-based single cell force spectroscopy. The effects of dextran concentration and molecular weight on the interaction energy of adhering RBCs were determined. The results on adhesion energy are in excellent agreement with a model based on the depletion effect and previous experimental studies. Furthermore, our method allowed to determine the adhesion force, a quantity that is needed in theoretical investigations on blood flow.

  18. Thinking outside the cell: how cadherins drive adhesion

    PubMed Central

    Brasch, Julia; Harrison, Oliver J.; Honig, Barry; Shapiro, Lawrence

    2012-01-01

    Cadherins embody a superfamily of cell-surface glycoproteins whose ectodomains contain multiple repeats of β-sandwich EC (extracellular cadherin) domains that adopt a similar fold to immunoglobulin domains. The best characterized cadherins are the vertebrate “classical” cadherins, which mediate adhesion via trans homodimerization between their membrane-distal EC1 domains that extend from apposed cells, and assemble intercellular adherens junctions through cis clustering. To form mature trans adhesive dimers, cadherin domains from apposed cells dimerize in a “strand-swapped” conformation. This occurs in a two-step binding process involving a fast-binding intermediate called the “X-dimer”. Trans dimers are less flexible than cadherin monomers, a factor which drives junction assembly following cell-cell contact by reducing the entropic cost associated with the formation of lateral cis oligomers. Cadherins outside of the classical subfamily appear to have evolved distinct adhesive mechanisms which are just now beginning to be understood. PMID:22555008

  19. Thinking outside the cell: how cadherins drive adhesion.

    PubMed

    Brasch, Julia; Harrison, Oliver J; Honig, Barry; Shapiro, Lawrence

    2012-06-01

    Cadherins are a superfamily of cell surface glycoproteins whose ectodomains contain multiple repeats of β-sandwich extracellular cadherin (EC) domains that adopt a similar fold to immunoglobulin domains. The best characterized cadherins are the vertebrate 'classical' cadherins, which mediate adhesion via trans homodimerization between their membrane-distal EC1 domains that extend from apposed cells, and assemble intercellular adherens junctions through cis clustering. To form mature trans adhesive dimers, cadherin domains from apposed cells dimerize in a 'strand-swapped' conformation. This occurs in a two-step binding process involving a fast-binding intermediate called the 'X-dimer'. Trans dimers are less flexible than cadherin monomers, a factor that drives junction assembly following cell-cell contact by reducing the entropic cost associated with the formation of lateral cis oligomers. Cadherins outside the classical subfamily appear to have evolved distinct adhesive mechanisms that are only now beginning to be understood.

  20. PBRM1 Regulates the Expression of Genes Involved in Metabolism and Cell Adhesion in Renal Clear Cell Carcinoma

    PubMed Central

    Chowdhury, Basudev; Porter, Elizabeth G.; Stewart, Jane C.; Ferreira, Christina R.; Schipma, Matthew J.; Dykhuizen, Emily C.

    2016-01-01

    Polybromo-1 (PBRM1) is a component of the PBAF (Polybromo-associated-BRG1- or BRM-associated factors) chromatin remodeling complex and is the second most frequently mutated gene in clear-cell renal cell Carcinoma (ccRCC). Mutation of PBRM1 is believed to be an early event in carcinogenesis, however its function as a tumor suppressor is not understood. In this study, we have employed Next Generation Sequencing to profile the differentially expressed genes upon PBRM1 re-expression in a cellular model of ccRCC. PBRM1 re-expression led to upregulation of genes involved in cellular adhesion, carbohydrate metabolism, apoptotic process and response to hypoxia, and a downregulation of genes involved in different stages of cell division. The decrease in cellular proliferation upon PBRM1 re-expression was confirmed, validating the functional role of PBRM1 as a tumor suppressor in a cell-based model. In addition, we identified a role for PBRM1 in regulating metabolic pathways known to be important for driving ccRCC, including the regulation of hypoxia response genes, PI3K signaling, glucose uptake, and cholesterol homeostasis. Of particular novelty is the identification of cell adhesion as a major downstream process uniquely regulated by PBRM1 expression. Cytoskeletal reorganization was induced upon PBRM1 reexpression as evidenced from the increase in the number of cells displaying cortical actin, a hallmark of epithelial cells. Genes involved in cell adhesion featured prominently in our transcriptional dataset and overlapped with genes uniquely regulated by PBRM1 in clinical specimens of ccRCC. Genes involved in cell adhesion serve as tumor suppressor and maybe involved in inhibiting cell migration. Here we report for the first time genes linked to cell adhesion serve as downstream targets of PBRM1, and hope to lay the foundation of future studies focusing on the role of chromatin remodelers in bringing about these alterations during malignancies. PMID:27100670

  1. Rocking adhesion assay system to study adhesion and transendothelial migration of cancer cells.

    PubMed

    Bapu, Deepashree; Khadim, Munira; Brooks, Susan A

    2014-01-01

    Adhesion of metastatic cancer cells to the vascular endothelium of the target organs and their subsequent transendothelial migration is one of the critical, yet poorly understood, steps of the metastatic cascade. Conventionally, the mechanisms of this complex process have been studied using static adhesion systems or flow assay systems. Static assay systems are easy to set up and perform but do not mimic the physiological conditions of blood flow. Flow assays closely mimic physiological conditions of flow but are time consuming and require specialist equipment. In this chapter we describe the rocking adhesion system which incorporates the key advantages of both the static and flow assay systems and not only is easy to set up and perform but also mimics conditions of blood flow.

  2. Splicing factors PTBP1 and PTBP2 promote proliferation and migration of glioma cell lines

    PubMed Central

    Cheung, Hannah C.; Hai, Tao; Zhu, Wen; Baggerly, Keith A.; Tsavachidis, Spiridon; Krahe, Ralf

    2009-01-01

    Polypyrimidine tract-binding protein 1 (PTBP1) is a multi-functional RNA-binding protein that is aberrantly overexpressed in glioma. PTBP1 and its brain-specific homologue polypyrimidine tract-binding protein 2 (PTBP2) regulate neural precursor cell differentiation. However, the overlapping and non-overlapping target transcripts involved in this process are still unclear. To determine why PTBP1 and not PTBP2 would promote glial cell-derived tumours, both PTBP1 and PTBP2 were knocked down in the human glioma cell lines U251 and LN229 to determine the role of these proteins in cell proliferation, migration, and adhesion. Surprisingly, removal of both PTBP1 and PTBP2 slowed cell proliferation, with the double knockdown having no additive effects. Decreased expression of both proteins individually and in combination inhibited cell migration and increased adhesion of cells to fibronectin and vitronectin. A global survey of differential exon expression was performed following PTBP1 knockdown in U251 cells using the Affymetrix Exon Array to identify PTBP1-specific splicing targets that enhance gliomagenesis. In the PTBP1 knockdown, previously determined targets were unaltered in their splicing patterns. A single gene, RTN4 (Nogo) had significantly enhanced inclusion of exon 3 when PTBP1 was removed. Overexpression of the splice isoform containing exon 3 decreased cell proliferation to a similar degree as the removal of PTBP1. These results provide the first evidence that RNA-binding proteins affect the invasive and rapid growth characteristics of glioma cell lines. Its actions on proliferation appear to be mediated, in part, through alternative splicing of RTN4. PMID:19506066

  3. Non-Cell-Adhesive Substrates for Printing of Arrayed Biomaterials

    PubMed Central

    Appel, Eric A.; Larson, Benjamin L.; Luly, Kathryn M.; Kim, Jinseong D.

    2015-01-01

    Cellular microarrays have become extremely useful in expediting the investigation of large libraries of (bio)materials for both in vitro and in vivo biomedical applications. We have developed an exceedingly simple strategy for the fabrication of non-cell-adhesive substrates supporting the immobilization of diverse (bio)material features, including both monomeric and polymeric adhesion molecules (e.g. RGD and polylysine), hydrogels, and polymers. PMID:25430948

  4. Nukbone® promotes proliferation and osteoblastic differentiation of mesenchymal stem cells from human amniotic membrane

    SciTech Connect

    Rodríguez-Fuentes, Nayeli; Rodríguez-Hernández, Ana G.; Enríquez-Jiménez, Juana; Alcántara-Quintana, Luz E.; Fuentes-Mera, Lizeth; Piña-Barba, María C.; Zepeda-Rodríguez, Armando; and others

    2013-05-10

    Highlights: •Nukbone showed to be a good scaffold for adhesion, proliferation and differentiation of stem cells. •Nukbone induced osteoblastic differentiation of human mesenchymal stem cells. •Results showed that Nukbone offer an excellent option for bone tissue regeneration due to properties. -- Abstract: Bovine bone matrix Nukbone® (NKB) is an osseous tissue-engineering biomaterial that retains its mineral and organic phases and its natural bone topography and has been used as a xenoimplant for bone regeneration in clinics. There are not studies regarding its influence of the NKB in the behavior of cells during the repairing processes. The aim of this research is to demonstrate that NKB has an osteoinductive effect in human mesenchymal stem cells from amniotic membrane (AM-hMSCs). Results indicated that NKB favors the AM-hMSCs adhesion and proliferation up to 7 days in culture as shown by the scanning electron microscopy and proliferation measures using an alamarBlue assay. Furthermore, as demonstrated by reverse transcriptase polymerase chain reaction, it was detected that two gene expression markers of osteoblastic differentiation: the core binding factor and osteocalcin were higher for AM-hMSCs co-cultured with NKB in comparison with cultivated cells in absence of the biomaterial. As the results indicate, NKB possess the capability for inducing successfully the osteoblastic differentiation of AM-hMSC, so that, NKB is an excellent xenoimplant option for repairing bone tissue defects.

  5. Knockdown of Golgi phosphoprotein 2 inhibits hepatocellular carcinoma cell proliferation and motility

    PubMed Central

    Liu, Yiming; Zhang, Xiaodi; Sun, Ting; Jiang, Junchang; Li, Ying; Chen, Mingliang; Wei, Zhen; Jiang, Weiqin; Zhou, Linfu

    2016-01-01

    Golgi phosphoprotein 2 (GP73) is highly expressed in hepatocellular carcinoma (HCC) cells, where it serves as a biomarker and indicator of disease progression. We used MTS assays, anchorage-independent cell colony formation assays and a xenograft tumor model to show that GP73-specific siRNAs inhibit HCC proliferation in HepG2, SMMC-7721, and Huh7 cell lines and in vivo. Following GP73 silencing, levels of p-Rb, a factor related to metastasis, were reduced, but cell cycle progression was unaffected. Our results suggest that GP73 silencing may not directly suppress proliferation, but may instead inhibit cell motility. Results from proliferation assays suggest GP73 reduces expression of epithelial mesenchymal transition (EMT)-related factors and promotes cell motility, while transwell migration and invasion assays indicated a possible role in metastasis. Immunofluorescence co-localization microscopy and immunoblotting showed that GP73 decreases expression of N-cadherin and E-cadherin, two key factors in EMT, which may in turn decrease intracellular adhesive forces and promote cell motility. This study confirmed that GP73 expression leads to increased expression of EMT-related proteins and that GP73 silencing reduces HCC cell migration in vitro. These findings suggest that GP73 silencing through siRNA delivery may provide a novel low-toxicity therapy for the inhibition of tumor proliferation and metastasis. PMID:26870893

  6. Silencing of WWP2 inhibits adhesion, invasion, and migration in liver cancer cells.

    PubMed

    Qin, Yong; Xu, Sheng-Qian; Pan, De-Biao; Ye, Guan-Xiong; Wu, Cheng-Jun; Wang, Shi; Wang, Chao-Jun; Jiang, Jin-Yan; Fu, Jing

    2016-05-01

    The role and clinical implication of the WWP2 E3 ubiquitin ligase in liver cancer are poorly understood. In the current study, we investigated the expression level of WWP2 and its functions in cell adhesion, invasion, and migration in liver cancer. We used real-time PCR to detect the expression of WWP2 in liver cancer and adjacent samples from the People's Hospital of Lishui and also analyzed The Cancer Genome Atlas (TCGA) RNA-seq data by bioinformatics. Migration and invasion were detected by transwell analysis. We detected a strong WWP2 expression in tumor tissues of the People's Hospital of Lishui, and the survival rate was significantly higher in patients with lower WWP2-expressing tumors. WWP2 small hairpin RNA (shRNA) lentivirus stably infected cells (shWWP2), Huh7, showed slower growth speed compared with scramble control-infected cells in a xenograft mouse model. Knockdown of WWP2 Huh7 and BEL-7404 cells demonstrated a reduction in adhesion, invasion, and migration. Gene set enrichment analysis (GSEA) showed that WWP2 is positively correlated to cancer-related pathways including the chemokine signaling pathway. WWP2 also regulated MMP-9, caspase-9, CXCR3, and CCR5 expression in liver cancer cells. In addition, knockdown of CXCR3 and CCR5 significantly inhibited cell proliferation, adhesion, invasion, and migration in Huh7 and BEL-7404 cells. Our data suggest that targeting of WWP2 may be a therapeutic strategy for liver cancer treatment.

  7. Role of Periostin in Adhesion and Migration of Bone Remodeling Cells.

    PubMed

    Cobo, Teresa; Viloria, Cristina G; Solares, Laura; Fontanil, Tania; González-Chamorro, Elena; De Carlos, Félix; Cobo, Juan; Cal, Santiago; Obaya, Alvaro J

    2016-01-01

    Periostin is an extracellular matrix protein highly expressed in collagen-rich tissues subjected to continuous mechanical stress. Functionally, periostin is involved in tissue remodeling and its altered function is associated to numerous pathological processes. In orthodontics, periostin plays key roles in the maintenance of dental tissues and it is mainly expressed in those areas where tension or pressing forces are taking place. In this regard, high expression of periostin is essential to promote migration and proliferation of periodontal ligament fibroblasts. However little is known about the participation of periostin in migration and adhesion processes of bone remodeling cells. In this work we employ the mouse pre-osteoblastic MC3T3-E1 and the macrophage-like RAW 264.7 cell lines to overexpress periostin and perform different cell-based assays to study changes in cell behavior. Our data indicate that periostin overexpression not only increases adhesion capacity of MC3T3-E1 cells to different matrix proteins but also hampers their migratory capacity. Changes on RNA expression profile of MC3T3-E1 cells upon periostin overexpression have been also analyzed, highlighting the alteration of genes implicated in processes such as cell migration, adhesion or bone metabolism but not in bone differentiation. Overall, our work provides new evidence on the impact of periostin in osteoblasts physiology.

  8. Role of Periostin in Adhesion and Migration of Bone Remodeling Cells

    PubMed Central

    Cobo, Teresa; Viloria, Cristina G.; Solares, Laura; Fontanil, Tania; González-Chamorro, Elena; De Carlos, Félix; Cobo, Juan; Cal, Santiago; Obaya, Alvaro J.

    2016-01-01

    Periostin is an extracellular matrix protein highly expressed in collagen-rich tissues subjected to continuous mechanical stress. Functionally, periostin is involved in tissue remodeling and its altered function is associated to numerous pathological processes. In orthodontics, periostin plays key roles in the maintenance of dental tissues and it is mainly expressed in those areas where tension or pressing forces are taking place. In this regard, high expression of periostin is essential to promote migration and proliferation of periodontal ligament fibroblasts. However little is known about the participation of periostin in migration and adhesion processes of bone remodeling cells. In this work we employ the mouse pre-osteoblastic MC3T3-E1 and the macrophage-like RAW 264.7 cell lines to overexpress periostin and perform different cell-based assays to study changes in cell behavior. Our data indicate that periostin overexpression not only increases adhesion capacity of MC3T3-E1 cells to different matrix proteins but also hampers their migratory capacity. Changes on RNA expression profile of MC3T3-E1 cells upon periostin overexpression have been also analyzed, highlighting the alteration of genes implicated in processes such as cell migration, adhesion or bone metabolism but not in bone differentiation. Overall, our work provides new evidence on the impact of periostin in osteoblasts physiology. PMID:26809067

  9. MAPs/bFGF-PLGA microsphere composite-coated titanium surfaces promote increased adhesion and proliferation of fibroblasts.

    PubMed

    Wang, Zhongshan; Wu, Guofeng; Bai, Shizhu; Feng, Zhihong; Dong, Yan; Zhou, Jian; Qin, Haiyan; Zhao, Yimin

    2014-06-01

    Infection and epithelial downgrowth are two major problems with maxillofacial transcutaneous implants, and both are mainly due to lack of stable closure of soft tissues at transcutaneous sites. Fibroblasts have been shown to play a key role in the formation of biological seals. In this work, titanium (Ti) model surfaces were coated with mussel adhesive proteins (MAPs) utilizing its unique adhesion ability on diverse inorganic and organic surfaces in wet environments. Prepared basic fibroblast growth factor (bFGF)-poly(lactic-co-glycolic acid) (PLGA) microspheres can be easily synthesized and combined onto MAPs-coated Ti surfaces, due to the negative surface charges of microspheres in aqueous solution, which is in contrast to the positive charges of MAPs. Titanium model surfaces were divided into three groups. Group A: MAPs/bFGF-PLGA microspheres composite-coated Ti surfaces. Group B: MAPs-coated Ti surfaces. Group C: uncoated Ti surfaces. The effects of coated Ti surfaces on adhesion of fibroblasts, cytoskeletal organization, proliferation, and extracellular matrix (ECM)-related gene expressions were examined. The results revealed increased adhesion (P < 0.05), enhanced actin cytoskeletal organization, and up-regulated ECM-related gene expressions in groups A and B compared with group C. Increased proliferation of fibroblasts during five days of incubation was observed in group A compared with groups B and C (P < 0.05). Collectively, the results from this in vitro study demonstrated that MAPs/bFGF-PLGA microspheres composite-coated Ti surfaces had the ability to increase fibroblast functionality. In addition, MAPs/bFGF-PLGA microsphere composite-coated Ti surfaces should be studied further as a method of promoting formation of stable biological seals around transcutaneous sites.

  10. Platelet adhesion to human umbilical vein endothelial cells cultured on anionic hydrogel scaffolds.

    PubMed

    Chen, Yong Mei; Tanaka, Masaru; Gong, Jian Ping; Yasuda, Kazunori; Yamamoto, Sadaaki; Shimomura, Masatsugu; Osada, Yoshihito

    2007-04-01

    In this work we describe experiments designed to understand the human platelet adhesion to human umbilical vein endothelial cells (HUVECs) cultured on various kinds of chemically cross-linked anionic hydrogels, which were synthesized by radical polymerization. HUVECs could proliferate to sub-confluent or confluent on poly(acrylic acid) (PAA), poly(2-acrylamido-2-methyl-propane sulfonic acid sodium salt) (PNaAMPS), and poly(sodium p-styrene sulfonate) (PNaSS) gels. The proliferation behavior was not sensitive to the cross-linker concentration of the gels. However, the platelet adhesion on the HUVECs cultured on these gels showed different behavior, as revealed by human platelet adhesion test in static conditions. Only a few platelets adhered on the HUVEC sheets cultured on PNaAMPS gels with 4 and 10mol% cross-linker concentrations, and completely no platelet adhered on the HUVEC sheets cultured on PNaSS gels with 4 and 10mol% cross-linker concentrations. On the other hand, a large number of platelets adhered on the HUVECs cultured on PAA gels with 1, 2mol% cross-linker concentrations and PNaAMPS gel with 2mol% cross-linker concentration. Furthermore, the study showed that promote of the glycocalyx of HUVECs with transforming growth factor-beta(1) (TGF-beta(1)) decreased platelet adhesion, and degrade the glycocalyx with heparinase I increased platelet adhesion. The results suggested that the glycocalyx of cultured HUVECs modulates platelet compatibility, and the amount of glycocalyx secreted by HUVECs dependents on the chemical structure and cross-linker concentration of gel scaffolds. This result should be applied to make the hybrid artificial blood vessel composes of gels and endothelial cells with high platelet compatibility.

  11. Combinatorial growth of oxide nanoscaffolds and its influence in osteoblast cell adhesion

    PubMed Central

    Acevedo-Morantes, Claudia Y.; Irizarry-Ortiz, Roberto A.; Caceres-Valencia, Pablo G.; Singh, Surinder P.; Ramirez-Vick, Jaime E.

    2012-01-01

    We report a novel method for high-throughput investigations on cell-material interactions based on metal oxide nanoscaffolds. These scaffolds possess a continuous gradient of various titanium alloys allowing the compositional and morphological variation that could substantially improve the formation of an osseointegrative interface with bone. The model nanoscaffold has been fabricated on commercially pure titanium (cp-Ti) substrate with a compositional gradients of tin (Sn), chromium (Cr), and niobium (Nb) deposited using a combinatorial approach followed by annealing to create native oxide surface. As an invitro test system, the human fetal osteoblastic cell line (hFOB 1.19) has been used. Cell-adhesion of hFOB 1.19 cells and the suitability of these alloys have been evaluated for cell-morphology, cell-number, and protein adsorption. Although, cell-morphology was not affected by surface composition, cell-proliferation rates varied significantly with surface metal oxide composition; with the Sn- and Nb-rich regions showing the highest proliferation rate and the Cr-rich regions presenting the lowest. The results suggest that Sn and Nb rich regions on surface seems to promote hFOB 1.19 cell proliferation and may therefore be considered as implant material candidates that deserve further analysis. PMID:22670064

  12. Combinatorial growth of oxide nanoscaffolds and its influence in osteoblast cell adhesion

    NASA Astrophysics Data System (ADS)

    Acevedo-Morantes, Claudia Y.; Irizarry-Ortiz, Roberto A.; Caceres-Valencia, Pablo G.; Singh, Surinder P.; Ramirez-Vick, Jaime E.

    2012-05-01

    We report a novel method for high-throughput investigations on cell-material interactions based on metal oxide nanoscaffolds. These scaffolds possess a continuous gradient of various titanium alloys allowing the compositional and morphological variation that could substantially improve the formation of an osseointegrative interface with bone. The model nanoscaffold has been fabricated on commercially pure titanium (cp-Ti) substrate with a compositional gradients of tin (Sn), chromium (Cr), and niobium (Nb) deposited using a combinatorial approach followed by annealing to create native oxide surface. As an invitro test system, the human fetal osteoblastic cell line (hFOB 1.19) has been used. Cell-adhesion of hFOB 1.19 cells and the suitability of these alloys have been evaluated for cell-morphology, cell-number, and protein adsorption. Although, cell-morphology was not affected by surface composition, cell-proliferation rates varied significantly with surface metal oxide composition; with the Sn- and Nb-rich regions showing the highest proliferation rate and the Cr-rich regions presenting the lowest. The results suggest that Sn and Nb rich regions on surface seems to promote hFOB 1.19 cell proliferation and may therefore be considered as implant material candidates that deserve further analysis.

  13. Biomechanics of cell rolling: shear flow, cell-surface adhesion, and cell deformability.

    PubMed

    Dong, C; Lei, X X

    2000-01-01

    The mechanics of leukocyte (white blood cell; WBC) deformation and adhesion to endothelial cells (EC) has been investigated using a novel in vitro side-view flow assay. HL-60 cell rolling adhesion to surface-immobilized P-selectin was used to model the WBC-EC adhesion process. Changes in flow shear stress, cell deformability, or substrate ligand strength resulted in significant changes in the characteristic adhesion binding time, cell-surface contact and cell rolling velocity. A 2-D model indicated that cell-substrate contact area under a high wall shear stress (20 dyn/cm2) could be nearly twice of that under a low stress (0.5 dyn/cm2) due to shear flow-induced cell deformation. An increase in contact area resulted in more energy dissipation to both adhesion bonds and viscous cytoplasm, whereas the fluid energy that inputs to a cell decreased due to a flattened cell shape. The model also predicted a plateau of WBC rolling velocity as flow shear stresses further increased. Both experimental and computational studies have described how WBC deformation influences the WBC-EC adhesion process in shear flow.

  14. Cell Adhesion Molecules and Ubiquitination—Functions and Significance

    PubMed Central

    Homrich, Mirka; Gotthard, Ingo; Wobst, Hilke; Diestel, Simone

    2015-01-01

    Cell adhesion molecules of the immunoglobulin (Ig) superfamily represent the biggest group of cell adhesion molecules. They have been analyzed since approximately 40 years ago and most of them have been shown to play a role in tumor progression and in the nervous system. All members of the Ig superfamily are intensively posttranslationally modified. However, many aspects of their cellular functions are not yet known. Since a few years ago it is known that some of the Ig superfamily members are modified by ubiquitin. Ubiquitination has classically been described as a proteasomal degradation signal but during the last years it became obvious that it can regulate many other processes including internalization of cell surface molecules and lysosomal sorting. The purpose of this review is to summarize the current knowledge about the ubiquitination of cell adhesion molecules of the Ig superfamily and to discuss its potential physiological roles in tumorigenesis and in the nervous system. PMID:26703751

  15. Endoplasmic Reticulum Calcium, Stress and Cell-to-Cell Adhesion

    PubMed Central

    Mauro, Theodora

    2014-01-01

    Darier's Disease (DD) is caused by mutations in the endoplasmic reticulum (ER) Ca2+ ATPase ATP2A2 (protein SERCA2). Current treatment modalities are ineffective for many patients. This report shows that impaired SERCA2 function, both in DD keratinocytes and in normal keratinocytes treated with the SERCA2-inhibitor thapsigargin, depletes ER Ca2+ stores, leading to constitutive ER stress and increased sensitivity to ER stressors. ER stress, in turn, leads to abnormal cell-to-cell adhesion via impaired redistribution of desmoplakin, desmoglein 3, desmocollin 3 and E-cadherin to the plasma membrane. This report illustrates how ER Ca2+ depletion and the resulting ER stress are central to the pathogenesis of the disease. Additionally, the authors introduce a possible new therapeutic agent, Miglustat. PMID:24924761

  16. Use of nano-sized clay crystallites to restore adhesion among tumor and aging stem cells - a molecular simulations approach

    PubMed Central

    Ahmed, Habib-ur-Rehman; Abduljauwad, Sahel N

    2016-01-01

    Adhesion of cells to the ECM is key to the regulation of cellular morphology, migration, proliferation, survival, and differentiation. The decrease in or loss of the cell’s ability of mutual adhesiveness has been considered as one of the specific abnormalities in the surface properties of malignant cells. A change in the association of plasma membrane with cytoskeletal structures also seems to have a close relation with these abnormalities. Similar to the role of adhesions in tumor cells, stem cells’ self-renewal is also tightly controlled by the concerted action of stem cell-intrinsic factors and signals within the niche. This study has demonstrated through molecular simulations the potential use of smectite (Na-montmorillonite) clay crystallites to create adhesions among tumor and stem cells. High electrostatic energies and cohesive energy densities measured in the simulations after the sorption of clay crystallites on cell-cell and cell-ECM complexes validate the concept of using these crystallites for the purposes. As results of this study are quite promising and clay crystallites could be considered as an option to restore adhesions in tumor and stem cells, other confirmatory tests and live cell culture studies are in process for the final validation. PMID:28078181

  17. Simple surface engineering of polydimethylsiloxane with polydopamine for stabilized mesenchymal stem cell adhesion and multipotency

    PubMed Central

    Chuah, Yon Jin; Koh, Yi Ting; Lim, Kaiyang; Menon, Nishanth V.; Wu, Yingnan; Kang, Yuejun

    2015-01-01

    Polydimethylsiloxane (PDMS) has been extensively exploited to study stem cell physiology in the field of mechanobiology and microfluidic chips due to their transparency, low cost and ease of fabrication. However, its intrinsic high hydrophobicity renders a surface incompatible for prolonged cell adhesion and proliferation. Plasma-treated or protein-coated PDMS shows some improvement but these strategies are often short-lived with either cell aggregates formation or cell sheet dissociation. Recently, chemical functionalization of PDMS surfaces has proved to be able to stabilize long-term culture but the chemicals and procedures involved are not user- and eco-friendly. Herein, we aim to tailor greener and biocompatible PDMS surfaces by developing a one-step bio-inspired polydopamine coating strategy to stabilize long-term bone marrow stromal cell culture on PDMS substrates. Characterization of the polydopamine-coated PDMS surfaces has revealed changes in surface wettability and presence of hydroxyl and secondary amines as compared to uncoated surfaces. These changes in PDMS surface profile contribute to the stability in BMSCs adhesion, proliferation and multipotency. This simple methodology can significantly enhance the biocompatibility of PDMS-based microfluidic devices for long-term cell analysis or mechanobiological studies. PMID:26647719

  18. The Src homology 2 protein Shb promotes cell cycle progression in murine hematopoietic stem cells by regulation of focal adhesion kinase activity

    SciTech Connect

    Gustafsson, Karin; Heffner, Garrett; Wenzel, Pamela L.; Curran, Matthew; Grawé, Jan; McKinney-Freeman, Shannon L.; Daley, George Q.; Welsh, Michael

    2013-07-15

    The widely expressed adaptor protein Shb has previously been reported to contribute to T cell function due to its association with the T cell receptor and furthermore, several of Shb's known interaction partners are established regulators of blood cell development and function. In addition, Shb deficient embryonic stem cells displayed reduced blood cell colony formation upon differentiation in vitro. The aim of the current study was therefore to explore hematopoietic stem and progenitor cell function in the Shb knockout mouse. Shb deficient bone marrow contained reduced relative numbers of long-term hematopoietic stem cells (LT-HSCs) that exhibited lower proliferation rates. Despite this, Shb knockout LT-HSCs responded promptly by entering the cell cycle in response to genotoxic stress by 5-fluorouracil treatment. In competitive LT-HSC transplantations, Shb null cells initially engrafted as well as the wild-type cells but provided less myeloid expansion over time. Moreover, Shb knockout bone marrow cells exhibited elevated basal activities of focal adhesion kinase/Rac1/p21-activated kinase signaling and reduced responsiveness to Stem Cell Factor stimulation. Consequently, treatment with a focal adhesion kinase inhibitor increased Shb knockout LT-HSC proliferation. The altered signaling characteristics thus provide a plausible mechanistic explanation for the changes in LT-HSC proliferation since these signaling intermediates have all been shown to participate in LT-HSC cell cycle control. In summary, the loss of Shb dependent signaling in bone marrow cells, resulting in elevated focal adhesion kinase activity and reduced proliferative responses in LT-HSCs under steady state hematopoiesis, confers a disadvantage to the maintenance of LT-HSCs over time. -- Highlights: • Shb is an adaptor protein operating downstream of tyrosine kinase receptors. • Shb deficiency reduces hematopoietic stem cell proliferation. • The proliferative effect of Shb occurs via increased

  19. Cell-cell adhesion in the cnidaria: insights into the evolution of tissue morphogenesis.

    PubMed

    Magie, Craig R; Martindale, Mark Q

    2008-06-01

    Cell adhesion is a major aspect of cell biology and one of the fundamental processes involved in the development of a multicellular animal. Adhesive mechanisms, both cell-cell and between cell and extracellular matrix, are intimately involved in assembling cells into the three-dimensional structures of tissues and organs. The modulation of adhesive complexes could therefore be seen as a central component in the molecular control of morphogenesis, translating information encoded within the genome into organismal form. The availability of whole genomes from early-branching metazoa such as cnidarians is providing important insights into the evolution of adhesive processes by allowing for the easy identification of the genes involved in adhesion in these organisms. Discovery of the molecular nature of cell adhesion in the early-branching groups, coupled with comparisons across the metazoa, is revealing the ways evolution has tinkered with this vital cellular process in the generation of the myriad forms seen across the animal kingdom.

  20. Chemical and physical modifications to poly(dimethylsiloxane) surfaces affect adhesion of Caco-2 cells.

    PubMed

    Wang, Lin; Sun, Bing; Ziemer, Katherine S; Barabino, Gilda A; Carrier, Rebecca L

    2010-06-15

    Polydimethylsiloxane (PDMS) silicone elastomer is extensively used in soft lithography processes to fabricate microscale or nano scale systems for microfluidic or cell culture applications. Though PDMS is biocompatible, it is not an ideal material for cell culture due to its poor cell adhesion properties. In this study, PDMS surfaces were modified to promote intestinal cell adhesion, in the interest of testing feasibility of using microfabricated PDMS systems for high throughput drug screening. Modification techniques included changing chemical composition of PDMS (i.e., varying curing to mixing agent ratio, and oxidization of PDMS surface by oxygen plasma), surface treatment of PDMS by coating with charged molecules (i.e., poly-D-lysine, L-alpha-phosphatidylcholine, and a layer bylayer coating), and deposition of extracellular matrix (ECM) proteins (i.e., laminin, fibronectin, and collagen). The influence of these modifications on PDMS properties, including elastic modulus and surface properties (wettability, chemical composition, topography, and protein adsorption) were characterized. Modification techniques were all found to change PDMS properties and influence the attachment and proliferation of Caco-2 cells over three days of culture to varying degrees. Generally, Caco-2 cells preferred to attach on collagen-coated, fibronectin-coated, and fibronectin-coated oxygen-plasma treated PDMS. The results highlight the importance of considering multiple physical and chemical factors that may be influenced by biomaterial modification and result in altered cell attachment to microfabricated systems, including surface hydrophobicity, chemical composition, stiffness, and topography. This study provides a foundation for further miniaturization, utilizing soft lithography techniques, of Caco-2 cell-based system for high-throughput screening of drug intestinal absorption during lead optimization in drug discovery. The understanding of different surface modifications on

  1. HOXA10 controls proliferation, migration and invasion in oral squamous cell carcinoma

    PubMed Central

    Carrera, Manoela; Bitu, Carolina C; de Oliveira, Carine Ervolino; Cervigne, Nilva K; Graner, Edgard; Manninen, Aki; Salo, Tuula; Coletta, Ricardo D

    2015-01-01

    Although HOX genes are best known for acting in the regulation of important events during embryogenesis, including proliferation, differentiation and migration, alterations in their expression patterns have been frequently described in cancers. In previous studies we analyzed the expression profile of the members of the HOX family of homeobox genes in oral samples of normal mucosa and squamous cell carcinoma (OSCC) and identified differently expressed genes such as HOXA10. The present study aimed to validate the increased expression of HOXA10 in OSCCs, and to investigate the effects arising from its knockdown in OSCC cells. The levels of HOXA10 mRNA were determined in human OSCC samples and cell lines by quantitative PCR, and HOXA10-mediated effects on proliferation, apoptosis, adhesion, epithelial-mesenchymal transition (EMT), migration and invasion were studied in HSC-3 tongue carcinoma cells by using retrovirus-mediated RNA interference. Higher expression of HOXA10 mRNA was observed in OSCC cell lines and in tumor tissues compared to normal controls. HOXA10 knockdown significantly reduced the proliferation of the tumor cells which was accompanied by increased levels of p21. HOXA10 silencing also significantly induced the expression of EMT markers and enhanced the adhesion, migration and invasion of HSC-3 cells. No effects on cell death were observed after HOXA10 knockdown. The results of the current study confirm the overexpression of HOXA10 in OSCCs, and further demonstrate that its expression is functionally associated with several important biological processes related to oral tumorigenesis, such as proliferation, migration and invasion. PMID:26097543

  2. Regulation of global gene expression and cell proliferation by APP

    PubMed Central

    Wu, Yili; Zhang, Si; Xu, Qin; Zou, Haiyan; Zhou, Weihui; Cai, Fang; Li, Tingyu; Song, Weihong

    2016-01-01

    Down syndrome (DS), caused by trisomy of chromosome 21, is one of the most common genetic disorders. Patients with DS display growth retardation and inevitably develop characteristic Alzheimer’s disease (AD) neuropathology, including neurofibrillary tangles and neuritic plaques. The expression of amyloid precursor protein (APP) is increased in both DS and AD patients. To reveal the function of APP and elucidate the pathogenic role of increased APP expression in DS and AD, we performed gene expression profiling using microarray method in human cells overexpressing APP. A set of genes are significantly altered, which are involved in cell cycle, cell proliferation and p53 signaling. We found that overexpression of APP inhibits cell proliferation. Furthermore, we confirmed that the downregulation of two validated genes, PSMA5 and PSMB7, inhibits cell proliferation, suggesting that the downregulation of PSMA5 and PSMB7 is involved in APP-induced cell proliferation impairment. Taken together, this study suggests that APP regulates global gene expression and increased APP expression inhibits cell proliferation. Our study provides a novel insight that APP overexpression may contribute to the growth impairment in DS patients and promote AD pathogenesis by inhibiting cell proliferation including neural stem cell proliferation and neurogenesis. PMID:26936520

  3. Regulation of global gene expression and cell proliferation by APP.

    PubMed

    Wu, Yili; Zhang, Si; Xu, Qin; Zou, Haiyan; Zhou, Weihui; Cai, Fang; Li, Tingyu; Song, Weihong

    2016-03-03

    Down syndrome (DS), caused by trisomy of chromosome 21, is one of the most common genetic disorders. Patients with DS display growth retardation and inevitably develop characteristic Alzheimer's disease (AD) neuropathology, including neurofibrillary tangles and neuritic plaques. The expression of amyloid precursor protein (APP) is increased in both DS and AD patients. To reveal the function of APP and elucidate the pathogenic role of increased APP expression in DS and AD, we performed gene expression profiling using microarray method in human cells overexpressing APP. A set of genes are significantly altered, which are involved in cell cycle, cell proliferation and p53 signaling. We found that overexpression of APP inhibits cell proliferation. Furthermore, we confirmed that the downregulation of two validated genes, PSMA5 and PSMB7, inhibits cell proliferation, suggesting that the downregulation of PSMA5 and PSMB7 is involved in APP-induced cell proliferation impairment. Taken together, this study suggests that APP regulates global gene expression and increased APP expression inhibits cell proliferation. Our study provides a novel insight that APP overexpression may contribute to the growth impairment in DS patients and promote AD pathogenesis by inhibiting cell proliferation including neural stem cell proliferation and neurogenesis.

  4. Cell-substrate impedance fluctuations of single amoeboid cells encode cell-shape and adhesion dynamics

    NASA Astrophysics Data System (ADS)

    Leonhardt, Helmar; Gerhardt, Matthias; Höppner, Nadine; Krüger, Kirsten; Tarantola, Marco; Beta, Carsten

    2016-01-01

    We show systematic electrical impedance measurements of single motile cells on microelectrodes. Wild-type cells and mutant strains were studied that differ in their cell-substrate adhesion strength. We recorded the projected cell area by time-lapse microscopy and observed irregular oscillations of the cell shape. These oscillations were correlated with long-term variations in the impedance signal. Superposed to these long-term trends, we observed fluctuations in the impedance signal. Their magnitude clearly correlated with the adhesion strength, suggesting that strongly adherent cells display more dynamic cell-substrate interactions.

  5. Graphical analysis of mammalian cell adhesion in vitro.

    PubMed

    Huang, Qiaoling; Antensteiner, Martin; Liu, Xiang Yang; Lin, Changjian; Vogler, Erwin A

    2016-12-01

    Short-term (<2h) cell adhesion kinetics of 3 different mammalian cell types: MDCK (epithelioid), MC3T3-E1 (osteoblastic), and MDA-MB-231 (cancerous) on 7 different substratum surface chemistries spanning the experimentally-observable range of water wettability (surface energy) are graphically analyzed to qualitatively elucidate commonalities and differences among cell/surface/suspending media combinations. We find that short-term mammalian cell attachment/adhesion in vitro correlates with substratum surface energy as measured by water adhesion tension, τ≡γlvcosθ, where γlv is water liquid-vapor interfacial energy (72.8   mJ/m(2)) and cosθ is the cosine of the advancing contact angle subtended by a water droplet on the substratum surface. No definitive functional relationships among cell-adhesion kinetic parameters and τ were observed as in previous work, but previously-observed general trends were reproduced, especially including a sharp transition in the magnitude of kinetic parameters from relatively low-to-high near τ=0mJ/m(2), although the exact adhesion tension at which this transition occurs is difficult to accurately estimate from the current data set. We note, however, that the transition is within the hydrophobic range based on the τ=30mJ/m(2) surface-energetic dividing line that has been proposed to differentiate "hydrophobic" surfaces from "hydrophilic". Thus, a rather sharp hydrophobic/hydrophilic contrast is observed for cell adhesion for disparate cell/surface combinations.

  6. Epithelial cell adhesion and gastrointestinal colonization of Lactobacillus in poultry.

    PubMed

    Spivey, Megan A; Dunn-Horrocks, Sadie L; Duong, Tri

    2014-11-01

    Administration of probiotic Lactobacillus cultures is an important alternative to the use of antibiotic growth promoters and has been demonstrated to improve animal health, growth performance, and preharvest food safety in poultry production. Whereas gastrointestinal colonization is thought to be critical to their probiotic functionality, factors important to Lactobacillus colonization in chickens are not well understood. In this study we investigate epithelial cell adhesion in vitro and colonization of Lactobacillusin vivo in broiler chickens. Adhesion of Lactobacillus cultures to epithelial cells was evaluated using the chicken LMH cell line. Lactobacillus cultures were able adhere effectively to LMH cells relative to Bacillus subtilis and Salmonella Typhimurium. Epithelial cell adhesion was similar for Lactobacillus crispatus TDCC 75, L. cristpatus TDCC 76, and Lactobacillus gallinarum TDCC 77, and all 3 were more adherent than L. gallinarum TDCC 78. However, when colonization was evaluated in the ileum and cecum of broiler chicks, L. crispatus TDCC 75 and L. gallinarum TDCC 77 were more persistent than L. crispatus TDCC 76 and L. gallinarum TDCC 78. The reduction of growth in medium supplemented with oxgal was greater for L. gallinarum TDCC 78 than L. gallinarum TDCC 77, suggesting that whereas adhesion was similar for the 2 strains, the difference in colonization between L. gallinarum strains may be due in part to their bile sensitivity. This study demonstrates that whereas adhesion to epithelial cells may be important in predicting gastrointestinal colonization, other factors including bile tolerance may also contribute to the colonization of Lactobacillus in poultry. Additionally, the chicken LMH cell line is expected to provide a platform for investigating mechanisms of Lactobacillus adhesion to epithelial tissue and evaluating the probiotic potential Lactobacillus in poultry.

  7. Lymphocyte adhesion-dependent calcium signaling in human endothelial cells

    PubMed Central

    1995-01-01

    Vascular endothelial cells (ECs) can undergo dramatic phenotypic and functional alterations in response to humoral and cellular stimuli. These changes promote endothelial participation in the inflammatory response through active recruitment of immune effector cells, increased vascular permeability, and alteration in vascular tone. In an attempt to define early events in lymphocyte-mediated EC signaling, we investigated cytosolic-free calcium (Ca2+) changes in single, Fluo-3- labeled human umbilical vein ECs (HUVECs), using an ACAS interactive laser cytometer. Of all lymphocyte subsets tested, allogeneic CD3-, CD56+ natural killer (NK) cells uniquely elicited oscillatory EC Ca2+ signals in cytokine (interleukin [IL]-1- or tumor necrosis factor [TNF])-treated ECs. The induction of these signals required avid intercellular adhesion, consisted of both Ca2+ mobilization and extracellular influx, and was associated with EC inositol phosphate (IP) generation. Simultaneous recording of NK and EC Ca2+ signals using two-color fluorescence detection revealed that, upon adhesion, NK cells flux prior to EC. Lymphocyte Ca2+ buffering with 1,2-bis-5-methyl-amino- phenoxylethane-N,N,N'-tetra-acetoxymethyl acetate (MAPTAM) demonstrated that lymphocyte fluxes are, in fact, prerequisites for the adhesion- dependent EC signals. mAb studies indicate that the beta 2 integrin- intercellular adhesion molecule (ICAM)-1 adhesion pathway is critically involved. However, ICAM-1 antisense oligonucleotide inhibition of IL-1- mediated ICAM-1 hyperinduction had no effect on EC Ca2+ signaling in lymphocyte-EC conjugates, indicating that additional cytokine-induced EC alteration is required. These experiments combine features of lymphocyte-endothelial interactions, intercellular adhesion, EC cytokine activation and transmembrane signaling. The results implicate the IP/Ca2+ second messenger pathway in EC outside-in signaling induced by cytotoxic lymphocytes, and suggest that these signals may play a

  8. Adhesion of platelets to artificial surfaces: effect of red cells.

    PubMed

    Brash, J L; Brophy, J M; Feuerstein, I A

    1976-05-01

    Adhesion of platelets to several polymer- and protein-coated glass surfaces has been studied in vitro. The apparatus consists of a cylindrical probe rotating in a test tube containing the platelet medium and allows close control of fluid shear and mass transport. Suspensions of washed pig platelets constitute the basic platelet medium, and can be modified by adding back red cells and plasma proteins. Adhesion is measured via 51Cr-labeling of platelets. In the absence of red cells, identical low levels of adhesion were seen on all surfaces and saturation was reached within 2 min. In the presence of red cells, adhesion was greater. Saturation on all surfaces except fibrinogen and collagen again occurred within 2 min. The adhesion levels on polymer surfaces and glass were indistinguishable, while those on albumin were lower and those on fibrinogen were higher. Collagen was the most reactive surface. It did not equilibrate within 15 min., and kinetic data indicated a platelet diffusivity strongly dependent on hematocrit. These effects were attributed to rotational and translational motion of the red cells causing increased diffusion and surface-platelet collision energy.

  9. Flexible nanopillars to regulate cell adhesion and movement

    NASA Astrophysics Data System (ADS)

    Chien, Fan-Ching; Dai, Yang-Hong; Kuo, Chiung Wen; Chen, Peilin

    2016-11-01

    Flexible polymer nanopillar substrates were used to systematically demonstrate cell alignment and migration guided by the directional formation of focal adhesions. The polymer nanopillar substrates were constructed to various height specifications to provide an extensive variation of flexibility; a rectangular arrangement created spatial confinement between adjacent nanopillars, providing less spacing in the horizontal and vertical directions. Three polymer nanopillar substrates with the diameter of 400 nm and the heights of 400, 800, and 1200 nm were fabricated. Super-resolution localization imaging and protein pair-distance analysis of vinculin proteins revealed that Chinese hamster ovary (CHO) cells formed mature focal adhesions on 1200 nm high nanopillar substrates by bending adjacent nanopillars to link dot-like adhesions. The spacing confinement of the adjacent nanopillars enhanced the orthogonal directionality of the formation tendency of the mature focal adhesions. The directional formation of the mature focal adhesions also facilitated the organization of actin filaments in the horizontal and vertical directions. Moreover, 78% of the CHO cells were aligned in these two directions, in conformity with the flexibility and nanotopographical cues of the nanopillars. Biased cell migration was observed on the 1200 nm high nanopillar substrates.

  10. Spatially controlled cell adhesion on three-dimensional substrates.

    PubMed

    Richter, Christine; Reinhardt, Martina; Giselbrecht, Stefan; Leisen, Daniel; Trouillet, Vanessa; Truckenmüller, Roman; Blau, Axel; Ziegler, Christiane; Welle, Alexander

    2010-10-01

    The microenvironment of cells in vivo is defined by spatiotemporal patterns of chemical and biophysical cues. Therefore, one important goal of tissue engineering is the generation of scaffolds with defined biofunctionalization in order to control processes like cell adhesion and differentiation. Mimicking extrinsic factors like integrin ligands presented by the extracellular matrix is one of the key elements to study cellular adhesion on biocompatible scaffolds. By using special thermoformable polymer films with anchored biomolecules micro structured scaffolds, e.g. curved and micro-patterned substrates, can be fabricated. Here, we present a novel strategy for the fabrication of micro-patterned scaffolds based on the "Substrate Modification and Replication by Thermoforming" (SMART) technology: The surface of a poly lactic acid membrane, having a low forming temperature of 60 degrees C and being initially very cell attractive, was coated with a photopatterned layer of poly(L-lysine) (PLL) and hyaluronic acid (VAHyal) to gain spatial control over cell adhesion. Subsequently, this modified polymer membrane was thermoformed to create an array of spherical microcavities with diameters of 300 microm for 3D cell culture. Human hepatoma cells (HepG2) and mouse fibroblasts (L929) were used to demonstrate guided cell adhesion. HepG2 cells adhered and aggregated exclusively within these cavities without attaching to the passivated surfaces between the cavities. Also L929 cells adhering very strongly on the pristine substrate polymer were effectively patterned by the cell repellent properties of the hyaluronic acid based hydrogel. This is the first time cell adhesion was controlled by patterned functionalization of a polymeric substrate with UV curable PLL-VAHyal in thermoformed 3D microstructures.

  11. Spatially controlled cell adhesion on three-dimensional substrates

    PubMed Central

    Richter, Christine; Reinhardt, Martina; Giselbrecht, Stefan; Leisen, Daniel; Trouillet, Vanessa; Truckenmüller, Roman; Blau, Axel; Ziegler, Christiane

    2010-01-01

    The microenvironment of cells in vivo is defined by spatiotemporal patterns of chemical and biophysical cues. Therefore, one important goal of tissue engineering is the generation of scaffolds with defined biofunctionalization in order to control processes like cell adhesion and differentiation. Mimicking extrinsic factors like integrin ligands presented by the extracellular matrix is one of the key elements to study cellular adhesion on biocompatible scaffolds. By using special thermoformable polymer films with anchored biomolecules micro structured scaffolds, e.g. curved and µ-patterned substrates, can be fabricated. Here, we present a novel strategy for the fabrication of µ-patterned scaffolds based on the “Substrate Modification and Replication by Thermoforming” (SMART) technology: The surface of a poly lactic acid membrane, having a low forming temperature of 60°C and being initially very cell attractive, was coated with a photopatterned layer of poly(L-lysine) (PLL) and hyaluronic acid (VAHyal) to gain spatial control over cell adhesion. Subsequently, this modified polymer membrane was thermoformed to create an array of spherical microcavities with diameters of 300 µm for 3D cell culture. Human hepatoma cells (HepG2) and mouse fibroblasts (L929) were used to demonstrate guided cell adhesion. HepG2 cells adhered and aggregated exclusively within these cavities without attaching to the passivated surfaces between the cavities. Also L929 cells adhering very strongly on the pristine substrate polymer were effectively patterned by the cell repellent properties of the hyaluronic acid based hydrogel. This is the first time cell adhesion was controlled by patterned functionalization of a polymeric substrate with UV curable PLL-VAHyal in thermoformed 3D microstructures. PMID:20480241

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

    SciTech Connect

    Wu, C.-C.; Su, H.-W.; Lee, C.-C.; Tang, M.-J.; Su, F.-C. . E-mail: fcsu@mail.ncku.edu.tw

    2005-04-01

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

  13. Molecular markers of cell adhesion in ameloblastomas. An update.

    PubMed

    González-González, Rogelio; Molina-Frechero, Nelly; Damian-Matsumura, Pablo; Bologna-Molina, Ronell

    2014-01-01

    Ameloblastoma is the most common odontogenic tumor of epithelial origin, and though it is of a benign nature, it frequently infiltrates the bone, has a high rate of recurrence and could potentially become malignant. Cellular adhesion potentially plays an important role in the manifestation of these characteristics and in the tumor biology of ameloblastomas. Losses of cell-cell and extracellular matrix adhesion and cohesion are among the first events that occur in the invasion and growth of tumors of epithelial origin. The present review includes a description of the molecules that are involved in cell adhesion as reported for various types of ameloblastomas and discusses the possible roles of these molecules in the biological behaviors of this odontogenic tumor. Knowledge of the complex mechanisms in which these molecules play a role is critical for the research and discovery of future therapeutic targets.

  14. Relationship between neuronal migration and cell-substratum adhesion: laminin and merosin promote olfactory neuronal migration but are anti- adhesive

    PubMed Central

    1991-01-01

    Regulation by the extracellular matrix (ECM) of migration, motility, and adhesion of olfactory neurons and their precursors was studied in vitro. Neuronal cells of the embryonic olfactory epithelium (OE), which undergo extensive migration in the central nervous system during normal development, were shown to be highly migratory in culture as well. Migration of OE neuronal cells was strongly dependent on substratum- bound ECM molecules, being specifically stimulated and guided by laminin (or the laminin-related molecule merosin) in preference to fibronectin, type I collagen, or type IV collagen. Motility of OE neuronal cells, examined by time-lapse video microscopy, was high on laminin-containing substrata, but negligible on fibronectin substrata. Quantitative assays of adhesion of OE neuronal cells to substrata treated with different ECM molecules demonstrated no correlation, either positive or negative, between the migratory preferences of cells and the strength of cell-substratum adhesion. Moreover, measurements of cell adhesion to substrata containing combinations of ECM proteins revealed that laminin and merosin are anti-adhesive for OE neuronal cells, i.e., cause these cells to adhere poorly to substrata that would otherwise be strongly adhesive. The evidence suggests that the anti- adhesive effect of laminin is not the result of interactions between laminin and other ECM molecules, but rather an effect of laminin on cells, which alters the way in which cells adhere. Consistent with this view, laminin was found to interfere strongly with the formation of focal contacts by OE neuronal cells. PMID:1918163

  15. Non-viral gene delivery regulated by stiffness of cell adhesion substrates

    NASA Astrophysics Data System (ADS)

    Kong, Hyun Joon; Liu, Jodi; Riddle, Kathryn; Matsumoto, Takuya; Leach, Kent; Mooney, David J.

    2005-06-01

    Non-viral gene vectors are commonly used for gene therapy owing to safety concerns with viral vectors. However, non-viral vectors are plagued by low levels of gene transfection and cellular expression. Current efforts to improve the efficiency of non-viral gene delivery are focused on manipulations of the delivery vector, whereas the influence of the cellular environment in DNA uptake is often ignored. The mechanical properties (for example, rigidity) of the substrate to which a cell adheres have been found to mediate many aspects of cell function including proliferation, migration and differentiation, and this suggests that the mechanics of the adhesion substrate may regulate a cell's ability to uptake exogeneous signalling molecules. In this report, we present a critical role for the rigidity of the cell adhesion substrate on the level of gene transfer and expression. The mechanism relates to material control over cell proliferation, and was investigated using a fluorescent resonance energy transfer (FRET) technique. This study provides a new material-based control point for non-viral gene therapy.

  16. Simvastatin suppresses breast cancer cell proliferation induced by senescent cells

    PubMed Central

    Liu, Su; Uppal, Harpreet; Demaria, Marco; Desprez, Pierre-Yves; Campisi, Judith; Kapahi, Pankaj

    2015-01-01

    Cellular senescence suppresses cancer by preventing the proliferation of damaged cells, but senescent cells can also promote cancer though the pro-inflammatory senescence-associated secretory phenotype (SASP). Simvastatin, an HMG-coA reductase inhibitor, is known to attenuate inflammation and prevent certain cancers. Here, we show that simvastatin decreases the SASP of senescent human fibroblasts by inhibiting protein prenylation, without affecting the senescent growth arrest. The Rho family GTPases Rac1 and Cdc42 were activated in senescent cells, and simvastatin reduced both activities. Further, geranylgeranyl transferase, Rac1 or Cdc42 depletion reduced IL-6 secretion by senescent cells. We also show that simvastatin mitigates the effects of senescent conditioned media on breast cancer cell proliferation and endocrine resistance. Our findings identify a novel activity of simvastatin and mechanism of SASP regulation. They also suggest that senescent cells, which accumulate after radio/chemo therapy, promote endocrine resistance in breast cancer and that simvastatin might suppress this resistance. PMID:26658759

  17. Adhesion between peptides/antibodies and breast cancer cells

    NASA Astrophysics Data System (ADS)

    Meng, J.; Paetzell, E.; Bogorad, A.; Soboyejo, W. O.

    2010-06-01

    Atomic force microscopy (AFM) techniques were used to measure the adhesion forces between the receptors on breast cancer cells specific to human luteinizing hormone-releasing hormone (LHRH) peptides and antibodies specific to the EphA2 receptor. The adhesion forces between LHRH-coated AFM tips and human MDA-MB-231 cells (breast cancer cells) were shown to be about five times greater than those between LHRH-coated AFM tips and normal Hs578Bst breast cells. Similarly, those between EphA2 antibody-coated AFM tips and breast cancer cells were over five times greater than those between EphA2 antibody-coated AFM tips and normal breast cells. The results suggest that AFM can be used for the detection of breast cancer cells in biopsies. The implications of the results are also discussed for the early detection and localized treatment of cancer.

  18. miR-24 triggers epidermal differentiation by controlling actin adhesion and cell migration

    PubMed Central

    Amelio, Ivano; Lena, Anna Maria; Viticchiè, Giuditta; Shalom-Feuerstein, Ruby; Terrinoni, Alessandro; Dinsdale, David; Russo, Giandomenico; Fortunato, Claudia; Bonanno, Elena; Spagnoli, Luigi Giusto; Aberdam, Daniel; Knight, Richard Austen

    2012-01-01

    During keratinocyte differentiation and stratification, cells undergo extensive remodeling of their actin cytoskeleton, which is important to control cell mobility and to coordinate and stabilize adhesive structures necessary for functional epithelia. Limited knowledge exists on how the actin cytoskeleton is remodeled in epithelial stratification and whether cell shape is a key determinant to trigger terminal differentiation. In this paper, using human keratinocytes and mouse epidermis as models, we implicate miR-24 in actin adhesion dynamics and demonstrate that miR-24 directly controls actin cable formation and cell mobility. miR-24 overexpression in proliferating cells was sufficient to trigger keratinocyte differentiation both in vitro and in vivo and directly repressed cytoskeletal modulators (PAK4, Tks5, and ArhGAP19). Silencing of these targets recapitulated the effects of miR-24 overexpression. Our results uncover a new regulatory pathway involving a differentiation-promoting microribonucleic acid that regulates actin adhesion dynamics in human and mouse epidermis. PMID:23071155

  19. Hyaluronan Stabilizes Focal Adhesions, Filopodia, and the Proliferative Phenotype in Esophageal Squamous Carcinoma Cells*

    PubMed Central

    Twarock, Sören; Tammi, Markku I.; Savani, Rashmin C.; Fischer, Jens W.

    2010-01-01

    Hyaluronan (HA) is a polysaccharide component in the parenchyma and stroma of human esophageal squamous cell carcinoma (ESCC). Clinically, esophageal cancer represents a highly aggressive tumor type with poor prognosis resulting in a 5-year survival rate of 5%. The aim of the present study was the detailed analysis of the role of HA synthesis for ESCC phenotype in vitro using the ESCC cell line OSC1. In OSC1 cells, pericellular HA-matrix surrounding extended actin-dependent filopodia was detected. The small molecule inhibitor of HA synthesis, 4-methylumbelliferone (4-MU, 0.3 mm) caused loss of these filopodia and focal adhesions and inhibited proliferation and migration. In search of the underlying mechanism cleavage of focal adhesion kinase (FAK) was detected by immunoblotting. In addition, displacing HA by an HA-binding peptide (Pep-1, 500 μg/ml) and digestion of pericellular HA by hyaluronidase resulted in cleavage of focal adhesions. Furthermore, real-time reverse transcription PCR revealed that HA synthase 3 (HAS3) > HAS2 are the predominant HA-synthases in OSC1. Lentiviral transduction with shHAS3, and to a lesser extent with shHAS2, reduced intact FAK protein and filopodia as well as proliferation and migration. Furthermore, down-regulation by lentiviral shRNA of RHAMM (receptor of HA-mediated motility) but not CD44 induced loss of filopodia and caused FAK cleavage. In contrast, knockdown of both HA receptors inhibited proliferation and migration of OSC1. In conclusion, HA synthesis and, in turn, RHAMM and CD44 signaling promoted an activated phenotype of OSC1. Because RHAMM appears to support both filopodia, FAK, and the proliferative and migratory phenotype, it may be promising to explore RHAMM as a potential therapeutic target in esophageal cancer. PMID:20463012

  20. Thermodynamics of short-term cell adhesion in vitro.

    PubMed Central

    Vogler, E A

    1988-01-01

    A thermodynamic theory of short-term (less than 2 hr) in vitro cell adhesion has been developed which allows calculation of reversible work of adhesion and estimation of a term proportional to cell-substrate contact area. The theory provides a means of determining a parameter related to membrane wetting tension for microscopic cells that does not require special manipulations which might desiccate or denature delicate cell membranes. Semiquantitative agreement between predicted and experimentally-measured cell adhesion obtained for three different cell types (MDCK, RBL-1, and HCT-15) in two different liquid phase compositions of surfactants (Tween-80 and fetal bovine serum) supports concepts and approximations utilized in development of theory. Cell-substrate contact areas were largest for wettable surfaces treated with ionizing corona or plasma discharges and smallest for hydrophobic materials for each cell type studied. Contact area for the continuous dog-kidney cell line MDCK was larger than that of either the leukemic blood cell RBL-1 or the anaplastic human colon cell HCT-15. PMID:3390519

  1. Androgens inhibit tumor necrosis factor-α-induced cell adhesion and promote tube formation of human coronary artery endothelial cells.

    PubMed

    Liao, Chun-Hou; Lin, Feng-Yen; Wu, Yi-No; Chiang, Han-Sun

    2012-06-01

    Endothelial cells contribute to the function and integrity of the vascular wall, and a functional aberration may lead to atherogenesis. There is increasing evidence on the atheroprotective role of androgens. Therefore, we studied the effect of the androgens-testosterone and dihydrotestosterone-and estradiol on human coronary artery endothelial cell (HCAEC) function. We found by MTT assay that testosterone is not cytotoxic and enhances HCAEC proliferation. The effect of testosterone (10-50 nM), dihydrotestosterone (5-50 nM), and estradiol (0.1-0.4 nM) on the adhesion of tumor necrosis factor-α (TNF-α)-stimulated HCAECs was determined at different time points (12-96 h) by assessing their binding with human monocytic THP-1 cells. In addition, the expression of adhesion molecules, vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1), was determined by ELISA and Western blot analysis. Both testosterone and dihydrotestosterone attenuated cell adhesion and the expression of VCAM-1 and ICAM-1 in a dose- and time-dependent manner. Furthermore, androgen treatment for a longer duration inhibited cell migration, as demonstrated by wound-healing assay, and promoted tube formation on a Matrigel. Western blot analysis demonstrated that the expression of phosphorylated endothelial nitric oxide synthase (eNOS) increased, whereas that of inducible nitric oxide synthase (iNOS) decreased following the 96-h steroid treatment of TNF-α-stimulated HCAECs. Our findings suggest that androgens modulate endothelial cell functions by suppressing the inflammatory process and enhancing wound-healing and regenerative angiogenesis, possibly through an androgen receptor (AR)-dependent mechanism.

  2. DLC-1, a GTPase-activating protein for Rho, is associated with cell proliferation, morphology, and migration in human hepatocellular carcinoma

    SciTech Connect

    Kim, Tai Young; Lee, Jung Weon; Kim, Hwang-Phill; Jong, Hyun-Soon; Kim, Tae-You; Jung, Mira; Bang, Yung-Jue; E-mail: bangyj@plaza.snu.ac.kr

    2007-03-30

    DLC-1 (deleted in liver cancer-1) is a tumor suppressor gene for hepatocellular carcinoma and other cancers. To characterize its functions, we constructed recombinant adenovirus encoding the wild-type DLC-1 and examined its effects on behaviors of a hepatocellular carcinoma cell line (SNU-368), which does not express DLC-1. Here, we found that restoration of DLC-1 expression in the SNU-368 cells caused an inhibition of cell proliferation with an increase of a subG1 population. Furthermore, DLC-1 overexpression induced disassembly of stress fibers and extensive membrane protrusions around cells on laminin-1. DLC-1 overexpression also inhibited cell migration and dephosphorylated focal adhesion proteins such as focal adhesion kinase (FAK), Cas (p130Cas; Crk-associated substrate), and paxillin. These observations suggest that DLC-1 plays important roles in signal transduction pathway regulating cell proliferation, cell morphology, and cell migration by affecting Rho family GTPases and focal adhesion proteins.

  3. Improvement in cell proliferation on silicone rubber by carbon nanotube coating.

    PubMed

    Matsuoka, Makoto; Akasaka, Tsukasa; Hashimoto, Takeshi; Totsuka, Yasunori; Watari, Fumio

    2009-01-01

    Silicone rubbers are widely used as tissue implants because of their flexibility and chemical stability. However, they have limited cellular adhesiveness and may cause problems in the long term. In this study, a coating of carbon nanotubes (CNTs) was applied to silicone rubber to improve its cellular adhesiveness. Scanning electron micrograph of this coating revealed that CNTs had formed a densely packed meshwork; the Ra values and protein adsorption capacity were enhanced. Although the contact angle did not change after coating, it decreased after immersion into a culture medium. After cultivation for 6 d, while Saos-2 cells were hardly observed on untreated silicone, the cells proliferated on CNT-coated silicone. Thus, CNT coating might be a simple and effective solution to problems associated with silicone implants.

  4. Metformin inhibits the proliferation of benign prostatic epithelial cells

    PubMed Central

    Ge, Rongbin; Li, Jijun; Johnson, Cameron W.; Rassoulian, Cyrus; Olumi, Aria F.

    2017-01-01

    Objective Benign prostatic hyperplasia (BPH) is the most common proliferative abnormality of the prostate affecting elderly men throughout the world. Epidemiologic studies have shown that diabetes significantly increases the risk of developing BPH, although whether anti-diabetic medications preventing the development of BPH remains to be defined. We have previously found that stromally expressed insulin-like growth factor 1 (IGF-1) promotes benign prostatic epithelial cell proliferation through paracrine mechanisms. Here, we seek to understand if metformin, a first line medication for the treatment of type 2 diabetes, inhibits the proliferation of benign prostatic epithelial cells through reducing the expression of IGF-1 receptor (IGF-1R) and regulating cell cycle. Methods BPE cell lines BPH-1 and P69, murine fibroblasts3T3 and primary human prostatic fibroblasts were cultured and tested in this study. Cell proliferation and the cell cycle were analyzed by MTS assay and flow cytometry, respectively. The expression of IGF-1R was determined by western-blot and immunocytochemistry. The level of IGF-1 secretion in culture medium was measured by ELISA. Results Metformin (0.5-10mM, 6-48h) significantly inhibited the proliferation of BPH-1 and P69 cells in a dose-dependent and time-dependent manner. Treatment with metformin for 24 hours lowered the G2/M cell population by 43.24% in P69 cells and 24.22% in BPH-1 cells. On the other hand, IGF-1 (100ng/mL, 24h) stimulated the cell proliferation (increased by 28.81% in P69 cells and 20.95% in BPH-1 cells) and significantly enhanced the expression of IGF-1R in benign prostatic epithelial cells. Metformin (5mM) abrogated the proliferation of benign prostatic epithelial cells induced by IGF-1. In 3T3 cells, the secretion of IGF-1 was significantly inhibited by metformin from 574.31pg/ml to 197.61pg/ml. The conditioned media of 3T3 cells and human prostatic fibroblasts promoted the proliferation of epithelial cells and the

  5. Collective Cell Behaviour with Neighbour-Dependent Proliferation, Death and Directional Bias.

    PubMed

    Binny, Rachelle N; James, Alex; Plank, Michael J

    2016-11-01

    Collective cell migration and proliferation are integral to tissue repair, embryonic development, the immune response and cancer. Central to collective cell migration and proliferation are interactions among neighbouring cells, such as volume exclusion, contact inhibition and adhesion. These individual-level processes can have important effects on population-level outcomes, such as growth rate and equilibrium density. We develop an individual-based model of cell migration and proliferation that includes these interactions. This is an extension of a previous model with neighbour-dependent directional bias to incorporate neighbour-dependent proliferation and death. A deterministic approximation to this individual-based model is derived using a spatial moment dynamics approach, which retains information about the spatial structure of the cell population. We show that the individual-based model and spatial moment model match well across a range of parameter values. The spatial moment model allows insight into the two-way interaction between spatial structure and population dynamics that cannot be captured by traditional mean-field models.

  6. Ultrasound stimulation increases proliferation of MC3T3-E1 preosteoblast-like cells

    PubMed Central

    2014-01-01

    Background Mechanical stimulation of bone increases bone mass and fracture healing, at least in part, through increases in proliferation of osteoblasts and osteoprogenitor cells. Researchers have previously performed in vitro studies of ultrasound-induced osteoblast proliferation but mostly used fixed ultrasound settings and have reported widely varying and inconclusive results. Here we critically investigated the effects of the excitation parameters of low-intensity pulsed ultrasound (LIPUS) stimulation on proliferation of MC3T3-E1 preosteoblastic cells in monolayer cultures. Methods We used a custom-designed ultrasound exposure system to vary the key ultrasound parameters—intensity, frequency and excitation duration. MC3T3-E1 cells were seeded in 12-well cell culture plates. Unless otherwise specified, treated cells, in groups of three, were excited twice for 10 min with an interval of 24 h in between after cell seeding. Proliferation rates of these cells were determined using BrdU and MTS assays 24 h after the last LIPUS excitation. All data are presented as the mean ± standard error. The statistical significance was determined using Student's two-sample two-tailed t tests. Results Using discrete LIPUS intensities ranging from 1 to 500 mW/cm2 (SATA, spatial average-temporal average), we found that approximately 75 mW/cm2 produced the greatest increase in osteoblast proliferation. Ultrasound exposures at higher intensity (approximately 465 mW/cm2) significantly reduced proliferation in MC3T3-E1 cells, suggesting that high-intensity pulsed ultrasound may increase apoptosis or loss of adhesion in these cells. Variation in LIPUS frequency from 0.5 MHz to 5 MHz indicated that osteoblast proliferation rate was not frequency dependent. We found no difference in the increase in proliferation rate if LIPUS was applied for 30 min/day or 10 min/day, indicating a habituation response. Conclusion This study concludes that a short-term stimulation with optimum intensity

  7. Viability and proliferation of endothelial cells upon exposure to GaN nanoparticles

    PubMed Central

    Braniste, Tudor; Tiginyanu, Ion; Horvath, Tibor; Raevschi, Simion; Cebotari, Serghei; Lux, Marco; Haverich, Axel

    2016-01-01

    Summary Nanotechnology is a rapidly growing and promising field of interest in medicine; however, nanoparticle–cell interactions are not yet fully understood. The goal of this work was to examine the interaction between endothelial cells and gallium nitride (GaN) semiconductor nanoparticles. Cellular viability, adhesion, proliferation, and uptake of nanoparticles by endothelial cells were investigated. The effect of free GaN nanoparticles versus the effect of growing endothelial cells on GaN functionalized surfaces was examined. To functionalize surfaces with GaN, GaN nanoparticles were synthesized on a sacrificial layer of zinc oxide (ZnO) nanoparticles using hydride vapor phase epitaxy. The uptake of GaN nanoparticles by porcine endothelial cells was strongly dependent upon whether they were fixed to the substrate surface or free floating in the medium. The endothelial cells grown on surfaces functionalized with GaN nanoparticles demonstrated excellent adhesion and proliferation, suggesting good biocompatibility of the nanostructured GaN. PMID:27826507

  8. ZF21 protein regulates cell adhesion and motility.

    PubMed

    Nagano, Makoto; Hoshino, Daisuke; Sakamoto, Takeharu; Kawasaki, Noritaka; Koshikawa, Naohiko; Seiki, Motoharu

    2010-07-02

    Cell migration on an extracellular matrix (ECM) requires continuous formation and turnover of focal adhesions (FAs) along the direction of cell movement. However, our knowledge of the components of FAs and the mechanism of their regulation remains limited. Here, we identify ZF21, a member of a protein family characterized by the presence of a phosphatidylinositol 3-phosphate-binding FYVE domain, to be a new regulator of FAs and cell movement. Knockdown of ZF21 expression in cells increased the number of FAs and suppressed cell migration. Knockdown of ZF21 expression also led to a significant delay in FA disassembly following induction of synchronous disassembly of FAs by nocodazole treatment. ZF21 bound to focal adhesion kinase, localized to FAs, and was necessary for dephosphorylation of FAK at Tyr(397), which is important for disassembly of FAs. Thus, ZF21 represents a new component of FAs, mediates disassembly of FAs, and thereby regulates cell motility.

  9. How to let go: pectin and plant cell adhesion.

    PubMed

    Daher, Firas Bou; Braybrook, Siobhan A

    2015-01-01

    Plant cells do not, in general, migrate. They maintain a fixed position relative to their neighbors, intimately linked through growth and differentiation. The mediator of this connection, the pectin-rich middle lamella, is deposited during cell division and maintained throughout the cell's life to protect tissue integrity. The maintenance of adhesion requires cell wall modification and is dependent on the actin cytoskeleton. There are developmental processes that require cell separation, such as organ abscission, dehiscence, and ripening. In these instances, the pectin-rich middle lamella must be actively altered to allow cell separation, a process which also requires cell wall modification. In this review, we will focus on the role of pectin and its modification in cell adhesion and separation. Recent insights gained in pectin gel mechanics will be discussed in relation to existing knowledge of pectin chemistry as it relates to cell adhesion. As a whole, we hope to begin defining the physical mechanisms behind a cells' ability to hang on, and how it lets go.

  10. Numerical analysis of cell adhesion in capillary flow

    NASA Astrophysics Data System (ADS)

    Takeishi, Naoki; Imai, Yohsuke; Ishida, Shunichi; Omori, Toshihiro; Kamm, Roger; Ishikawa, Takuji

    2016-11-01

    Numerical simulation of cell adhesion was performed for capillaries whose diameter is comparable to or smaller than that of the cell. Despite a lot of works about leukocyte and tumor cell rolling, cell motion in capillaries has remained unclear. The solid and fluid mechanics of a cell in flow was coupled with a slip bond model of ligand-receptor interactions. When the size of a capillary was reduced, the cell always transitioned to "bullet-like" motion, with a consequent decrease in the velocity of the cell. A state diagram is obtained for various values of capillary diameter and receptor density. According to our numerical results, bullet motion enables firm adhesion of a cell to the capillary wall even for a weak ligand-receptor binding. We also quantified effects of various parameters, including the dissociation rate constant, the spring constant, and the reactive compliance on the characteristics of cell motion. Our results suggest that even under the interaction between PSGL-1 and P-selectin, which is mainly responsible for leukocyte rolling, a cell is able to show firm adhesion in a small capillary. These findings may help in understanding such phenomena as leukocyte plugging and cancer metastasis. This research was supported by JSPS KAKENHI Grant Numbers 25000008, 26107703, 14J03967. We also acknowledge support from the Tohoku University Division for International Advanced Research and Education Organization.

  11. A Neural Network Based Workstation for Automated Cell Proliferation Analysis

    DTIC Science & Technology

    2001-10-25

    proliferation analysis, of cytological microscope images. The software of the system assists the expert biotechnologist during cell proliferation and...work was supported by the Programa de Apoyo a Proyectos de Desarrollo e Investigacíon en Informática REDII 2000. We thank Blanca Itzel Taboada for

  12. Interferon-alpha and dexamethasone inhibit adhesion of T cells to endothelial cells and synovial cells

    PubMed Central

    Eguchi, K.; Kawakami, A.; Nakashima, M.; Ida, H.; Sakito, S.; Matsuoka, N.; Terada, K.; Sakai, M.; Kawabe, Y.; Fukuda, T.; Ishimaru, T.; Kurouji, K.; Fujita, N.; Aoyagi, T.; Maeda, K.; Nagataki, S.

    1992-01-01

    We investigated whether interferon-gamma (IFN-γ), interferon-alpha (IFN-α) and glucocorticoids affected the adhesion of T cells to human umbilical endothelial cells or human synovial cells. About 30% of peripheral blood T cells could bind to unstimulated endothelial cells, but only a few T cells could bind to unstimulated synovial cells. When both endothelial cells and synovial cells were cultured with recombinant IFN-γ (rIFN-γ), the percentage of T cell binding to both types of cells increased in a dose-dependent manner. rIFN-α and dexamethasone blocked the T cell binding to unstimulated endothelial cells. Furthermore, rIFN-α and dexamethasone suppressed T cell binding to both endothelial cells and synovial cells stimulated by IFN-γ, and also inhibited intercellular adhesion molecule-1 (ICAM-1) expression on both endothelial cells and synovial cells stimulated by IFN-γ. These results suggest that IFN-α and glucocorticoids may inhibit T cell binding to endothelial cells or synovial cells by modulating adhesion molecule expression on these cells. PMID:1606729

  13. Multiparticle adhesive dynamics. Interactions between stably rolling cells.

    PubMed Central

    King, M R; Hammer, D A

    2001-01-01

    A novel numerical simulation of adhesive particles (cells) reversibly interacting with an adhesive surface under flow is presented. Particle--particle and particle--wall hydrodynamic interactions in low Reynolds number Couette flow are calculated using a boundary element method that solves an integral representation of the Stokes equation. Molecular bonds between surfaces are modeled as linear springs and stochastically formed and broken according to postulated descriptions of force-dependent kinetics. The resulting simulation, Multiparticle Adhesive Dynamics, is applied to the problem of selectin-mediated rolling of hard spheres coated with leukocyte adhesion molecules (cell-free system). Simulation results are compared to flow chamber experiments performed with carbohydrate-coated spherical beads rolling on P-selectin. Good agreement is found between theory and experiment, with the main observation being a decrease in rolling velocity with increasing concentration of rolling cells or increasing proximity between rolling cells. Pause times are found to increase and deviation motion is found to decrease as pairs of rolling cells become closer together or align with the flow. PMID:11463626

  14. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons.

    PubMed

    Park, Kyoung Ho; Yeo, Sang Won; Troy, Frederic A

    2014-10-17

    During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia-NCAMs) modulate cell-cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia-NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb's to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell-cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.

  15. Nanovesicles engineered from ES cells for enhanced cell proliferation.

    PubMed

    Jeong, Dayeong; Jo, Wonju; Yoon, Jaewoong; Kim, Junho; Gianchandani, Sachi; Gho, Yong Song; Park, Jaesung

    2014-11-01

    Extracellular vesicles (Exosomes and microvesicles) have drawn wide attentions in both diagnostic and therapeutic applications, since they are considered to shuttle biological signals intercellularly. However, further research on exosomes is limited by their rarity and heterogeneity even after lengthy isolation processes. In particular, these limitations are challenging in therapeutic applications. To meet these demands, cell-derived nanovesicles that mimic exosomes were generated by extruding living embryonic stem cells through micro-filters. These nanovesicles have an enclosed lipid bilayer and contain cellular contents. The present study investigated the ability of these nanovesicles to improve proliferation by treating primary murine skin fibroblasts with the nanovesicles. The treated skin fibroblasts showed higher expression levels of mRNA, VEGF-α, protein levels of TGF-β collagen I, PCNA, and Ki-67, as well as enhanced cell proliferation rate and number, compared to non-treated cells. The results indicate that treatment with the nanovesicles could potentially contribute to recovery or wound healing process of tissues.

  16. Scalable topographies to support proliferation and Oct4 expression by human induced pluripotent stem cells

    PubMed Central

    Reimer, Andreas; Vasilevich, Aliaksei; Hulshof, Frits; Viswanathan, Priyalakshmi; van Blitterswijk, Clemens A.; de Boer, Jan; Watt, Fiona M.

    2016-01-01

    It is well established that topographical features modulate cell behaviour, including cell morphology, proliferation and differentiation. To define the effects of topography on human induced pluripotent stem cells (iPSC), we plated cells on a topographical library containing over 1000 different features in medium lacking animal products (xeno-free). Using high content imaging, we determined the effect of each topography on cell proliferation and expression of the pluripotency marker Oct4 24 h after seeding. Features that maintained Oct4 expression also supported proliferation and cell-cell adhesion at 24 h, and by 4 days colonies of Oct4-positive, Sox2-positive cells had formed. Computational analysis revealed that small feature size was the most important determinant of pluripotency, followed by high wave number and high feature density. Using this information we correctly predicted whether any given topography within our library would support the pluripotent state at 24 h. This approach not only facilitates the design of substrates for optimal human iPSC expansion, but also, potentially, identification of topographies with other desirable characteristics, such as promoting differentiation. PMID:26757610

  17. Cytotoxicity of denture adhesives.

    PubMed

    de Gomes, Pedro Sousa; Figueiral, Maria Helena; Fernandes, Maria Helena R; Scully, Crispian

    2011-12-01

    Ten commercially available denture adhesives, nine soluble formulations (six creams, three powders) and one insoluble product (pad), were analyzed regarding the cytotoxicity profile in direct and indirect assays using L929 fibroblast cells. In the direct assay, fibroblasts were seeded over the surface of a thick adhesive gel (5%, creams; 2.5%, powders and pad). In the indirect assay, cells were cultured in the presence of adhesive extracts prepared in static and dynamic conditions (0.5-2%, creams; 0.25-1%, powders and pad). Cell toxicity was assessed for cell viability/proliferation (MTT assay) and cell morphology (observation of the F-actin cytoskeleton organization by confocal laser scanning microscopy). Direct contact of the L929 fibroblasts with the thick adhesive gels caused no, or only a slight, decrease in cell viability/proliferation. The adhesive extracts (especially those prepared in dynamic conditions) caused significantly higher growth inhibition of fibroblasts and, in addition, caused dose- and time-dependent effects, throughout the 6-72 h exposure time. Also, dose-dependent effects on cell morphology, with evident disruption of the F-actin cytoskeleton organization, were seen in the presence of most adhesives. In conclusion, the adhesives possessed different degrees of cytotoxicity, but similar dose- and time-dependent biological profiles.

  18. Hyaluronic acid influence on platelet-induced airway smooth muscle cell proliferation

    SciTech Connect

    Svensson Holm, Ann-Charlotte B.; Bengtsson, Torbjoern; Grenegard, Magnus; Lindstroem, Eva G.

    2012-03-10

    Hyaluronic acid (HA) is one of the main components of the extracellular matrix (ECM) and is expressed throughout the body including the lung and mostly in areas surrounding proliferating and migrating cells. Furthermore, platelets have been implicated as important players in the airway remodelling process, e.g. due to their ability to induce airway smooth muscle cell (ASMC) proliferation. The aim of the present study was to investigate the role of HA, the HA-binding surface receptor CD44 and focal adhesion kinase (FAK) in platelet-induced ASMC proliferation. Proliferation of ASMC was measured using the MTS-assay, and we found that the CD44 blocking antibody and the HA synthase inhibitor 4-Methylumbelliferone (4-MU) significantly inhibited platelet-induced ASMC proliferation. The interaction between ASMC and platelets was studied by fluorescent staining of F-actin. In addition, the ability of ASMC to synthesise HA was investigated by fluorescent staining using biotinylated HA-binding protein and a streptavidin conjugate. We observed that ASMC produced HA and that a CD44 blocking antibody and 4-MU significantly inhibited platelet binding to the area surrounding the ASMC. Furthermore, the FAK-inhibitor PF 573228 inhibited platelet-induced ASMC proliferation. Co-culture of ASMC and platelets also resulted in increased phosphorylation of FAK as detected by Western blot analysis. In addition, 4-MU significantly inhibited the increased FAK-phosphorylation. In conclusion, our findings demonstrate that ECM has the ability to influence platelet-induced ASMC proliferation. Specifically, we propose that HA produced by ASMC is recognised by platelet CD44. The platelet/HA interaction is followed by FAK activation and increased proliferation of co-cultured ASMC. We also suggest that the mitogenic effect of platelets represents a potential important and novel mechanism that may contribute to airway remodelling.

  19. Computer simulations of cell sorting due to differential adhesion.

    PubMed

    Zhang, Ying; Thomas, Gilberto L; Swat, Maciej; Shirinifard, Abbas; Glazier, James A

    2011-01-01

    The actions of cell adhesion molecules, in particular, cadherins during embryonic development and morphogenesis more generally, regulate many aspects of cellular interactions, regulation and signaling. Often, a gradient of cadherin expression levels drives collective and relative cell motions generating macroscopic cell sorting. Computer simulations of cell sorting have focused on the interactions of cells with only a few discrete adhesion levels between cells, ignoring biologically observed continuous variations in expression levels and possible nonlinearities in molecular binding. In this paper, we present three models relating the surface density of cadherins to the net intercellular adhesion and interfacial tension for both discrete and continuous levels of cadherin expression. We then use then the Glazier-Graner-Hogeweg (GGH) model to investigate how variations in the distribution of the number of cadherins per cell and in the choice of binding model affect cell sorting. We find that an aggregate with a continuous variation in the level of a single type of cadherin molecule sorts more slowly than one with two levels. The rate of sorting increases strongly with the interfacial tension, which depends both on the maximum difference in number of cadherins per cell and on the binding model. Our approach helps connect signaling at the molecular level to tissue-level morphogenesis.

  20. Endothelial cell migration on surfaces modified with immobilized adhesive peptides.

    PubMed

    Kouvroukoglou, S; Dee, K C; Bizios, R; McIntire, L V; Zygourakis, K

    2000-09-01

    Endothelial cell (EC) migration has been studied on aminophase surfaces with covalently bound RGDS and YIGSRG cell adhesion peptides. The fluorescent marker dansyl chloride was used to quantify the spatial distribution of the peptides on the modified surfaces. Peptides appeared to be distributed in uniformly dispersed large clusters separated by areas of lower peptide concentrations. We employed digital time-lapse video microscopy and image analysis to monitor EC migration on the modified surfaces and to reconstruct the cell trajectories. The persistent random walk model was then applied to analyze the cell displacement data and compute the mean root square speed, the persistence time, and the random motility coefficient of EC. We also calculated the time-averaged speed of cell locomotion. No differences in the speed of cell locomotion on the various substrates were noted. Immobilization of the cell adhesion peptides (RGDS and YIGSRG), however, significantly increased the persistence of cell movement and, thus, the random motility coefficient. These results suggest that immobilization of cell adhesion peptides on the surface of implantable biomaterials may lead to enhanced endothelization rates.

  1. Amygdalin Regulates Apoptosis and Adhesion in Hs578T Triple-Negative Breast Cancer Cells.

    PubMed

    Lee, Hye Min; Moon, Aree

    2016-01-01

    Amygdalin, D-mandelonitrile-β-D-glucoside-6-β-glucoside, belongs to aromatic cyanogenic glycoside group derived from rosaceous plant seed. Mounting evidence has supported the anti-cancer effects of amygdalin. However, whether amygdalin indeed acts as an anti-tumor agent against breast cancer cells is not clear. The present study aimed to investigate the effect of amygdalin on the proliferation of human breast cancer cells. Here, we show that amygdalin exerted cytotoxic activities on estrogen receptors (ER)-positive MCF7 cells, and MDA-MB-231 and Hs578T triple-negative breast cancer (TNBC) cells. Amygdalin induced apoptosis of Hs578T TNBC cells. Amygdalin downregulated B-cell lymphoma 2 (Bcl-2), upregulated Bcl-2-associated X protein (Bax), activated of caspase-3 and cleaved poly ADP-ribose polymerase (PARP). Amygdalin activated a pro-apoptotic signaling molecule p38 mitogen-activated protein kinases (p38 MAPK) in Hs578T cells. Treatment of amygdalin significantly inhibited the adhesion of Hs578T cells, in which integrin α5 may be involved. Taken together, this study demonstrates that amygdalin induces apoptosis and inhibits adhesion of breast cancer cells. The results suggest a potential application of amygdalin as a chemopreventive agent to prevent or alleviate progression of breast cancer, especially TNBC.

  2. Amygdalin Regulates Apoptosis and Adhesion in Hs578T Triple-Negative Breast Cancer Cells

    PubMed Central

    Lee, Hye Min; Moon, Aree

    2016-01-01

    Amygdalin, D-mandelonitrile-β-D-glucoside-6-β-glucoside, belongs to aromatic cyanogenic glycoside group derived from rosaceous plant seed. Mounting evidence has supported the anti-cancer effects of amygdalin. However, whether amygdalin indeed acts as an anti-tumor agent against breast cancer cells is not clear. The present study aimed to investigate the effect of amygdalin on the proliferation of human breast cancer cells. Here, we show that amygdalin exerted cytotoxic activities on estrogen receptors (ER)-positive MCF7 cells, and MDA-MB-231 and Hs578T triple-negative breast cancer (TNBC) cells. Amygdalin induced apoptosis of Hs578T TNBC cells. Amygdalin downregulated B-cell lymphoma 2 (Bcl-2), upregulated Bcl-2-associated X protein (Bax), activated of caspase-3 and cleaved poly ADP-ribose polymerase (PARP). Amygdalin activated a pro-apoptotic signaling molecule p38 mitogen-activated protein kinases (p38 MAPK) in Hs578T cells. Treatment of amygdalin significantly inhibited the adhesion of Hs578T cells, in which integrin α5 may be involved. Taken together, this study demonstrates that amygdalin induces apoptosis and inhibits adhesion of breast cancer cells. The results suggest a potential application of amygdalin as a chemopreventive agent to prevent or alleviate progression of breast cancer, especially TNBC. PMID:26759703

  3. Antibacterial and cell-adhesive polypeptide and poly(ethylene glycol) hydrogel as a potential scaffold for wound healing.

    PubMed

    Song, Airong; Rane, Aboli A; Christman, Karen L

    2012-01-01

    The ideal wound-healing scaffold should provide the appropriate physical and mechanical properties to prevent secondary infection, as well as an excellent physiological environment to facilitate cell adhesion, proliferation and/or differentiation. Therefore, we developed a synthetic cell-adhesive polypeptide hydrogel with inherent antibacterial activity. A series of polypeptides, poly(Lys)(x)(Ala)(y) (x+y=100), with varied hydrophobicity via metal-free ring-opening polymerization of NCA-Lys(Boc) and NCA-Ala monomers (NCA=N-carboxylic anhydride) mediated by hexamethyldisilazane (HMDS) were synthesized. These polypeptides were cross-linked with 6-arm polyethylene glycol (PEG)-amide succinimidyl glutarate (ASG) (M(w)=10K) to form hydrogels with a gelation time of five minutes and a storage modulus (G') of 1400-3000 Pa as characterized by rheometry. The hydrogel formed by cross-linking of poly(Lys)(60)(Ala)(40) (5 wt.%) and 6-arm PEG-ASG (16 wt.%) (Gel-III) exhibited cell adhesion and cell proliferation activities superior to other polypeptide hydrogels. In addition, Gel-III displays significant antibacterial activity against Escherichia coli JM109 and Staphylococcus aureus ATCC25923. Thus, we have developed a novel, cell-adhesive hydrogel with inherent antibacterial activity as a potential scaffold for cutaneous wound healing.

  4. Antibacterial and Cell-adhesive Polypeptide and Poly(ethylene glycol) Hydrogel as a Potential Scaffold for Wound Healing

    PubMed Central

    Song, Airong; Rane, Aboli A.; Christman, Karen L.

    2011-01-01

    The ideal wound healing scaffold should provide the appropriate physical and mechanical properties to prevent secondary infection, as well as an excellent physiological environment to facilitate cell adhesion, proliferation and/or differentiation. Therefore, we developed a synthetic cell-adhesive polypeptide hydrogel with inherent antibacterial activity. A series of polypeptides, poly(Lys)x(Ala)y (x+y=100) with varied hydrophobicity via metal-free ring-opening polymerization of NCA-Lys(Boc) and NCA-Ala monomers (NCA = N-carboxylic anhydride) mediated by hexamethyldisilazane (HMDS) were synthesized. These polypeptides were cross-linked with 6-arm PEG-amide succinimidyl glutarate (ASG) (Mw = 10K) to form hydrogels with a gelation time of five minutes and a storage modulus (G') of 1400–3000 Pa as characterized by rheometry. The hydrogel formed by cross-linking of poly(Lys)60(Ala)40 (5 wt%) and 6-arm PEG-ASG (16 wt%) (Gel-III) exhibited cell adhesion and cell proliferation activities superior to other polypeptide hydrogels. In addition, Gel-III displays significant antibacterial activity against E. coli JM109 and S. aureus ATCC25923. Thus, we have developed a novel, cell-adhesive hydrogel with inherent antibacterial activity as a potential scaffold for cutaneous wound healing. PMID:22023748

  5. Quantitative studies of endothelial cell adhesion. Directional remodeling of focal adhesion sites in response to flow forces.

    PubMed Central

    Davies, P F; Robotewskyj, A; Griem, M L

    1994-01-01

    Focal adhesion sites were observed in cultured endothelial cells by tandem scanning confocal microscopy and digitized image analysis, techniques that provide real-time images of adhesion site area and topography in living cells. Image subtraction demonstrated that in the presence of unidirectional steady laminar flow (shear stress [tau] = 10 dyn/cm2) a substantial fraction of focal adhesion sites remodeled in the direction of flow. In contrast, focal adhesions of control (no flow) cells remodeled without preferred direction. In confluent monolayers subjected to shear stresses of 10 dyn/cm2, cells began to realign in the direction of flow after 7-9 h. This was accompanied by redistribution of intracellular stress fibers, alignment of individual focal adhesion sites, and the coalescence of smaller sites resulting in fewer, but larger, focal adhesions per cell. Cell adhesion, repeatedly calculated in the same cells as a function of the areas of focal contact and the separation distances between membrane and substratum, varied by < 10% during both short (30 min), or prolonged (< or = 24 h), periods of exposure to flow. Consistent with these measurements, the gains and losses of focal adhesion area as each site remodeled were approximately equivalent. When the glass substratum was coated with gelatin, rates of remodeling were inhibited by 47% during flow (tau = 10 dyn/cm2). These studies: (a) reveal the dynamic nature of focal adhesion; (b) demonstrate that these sites at the ablumenal endothelial membrane are both acutely and chronically responsive to frictional shear stress forces applied to the opposite (lumenal) cell surface; and (c) suggest that components of the focal adhesion complex may be mechanically responsive elements coupled to the cytoskeleton. Images PMID:8182135

  6. The MRL proteins: adapting cell adhesion, migration and growth.

    PubMed

    Coló, Georgina P; Lafuente, Esther M; Teixidó, Joaquin

    2012-01-01

    MIG-10, RIAM and Lamellipodin (Lpd) are the founding members of the MRL family of multi-adaptor molecules. These proteins have common domain structures but display distinct functions in cell migration and adhesion, signaling, and in cell growth. The binding of RIAM with active Rap1 and with talin provides these MRL molecules with important regulatory roles on integrin-mediated cell adhesion and migration. Furthermore, RIAM and Lpd can regulate actin dynamics through their binding to actin regulatory Ena/VASP proteins. Recent data generated with the Drosophila MRL ortholog called Pico and with RIAM in melanoma cells indicate that these proteins can also regulate cell growth. As MRL proteins represent a relatively new family, many questions on their structure-function relationships remain unanswered, including regulation of their expression, post-translational modifications, new interactions, involvement in signaling and their knockout mice phenotype.

  7. Overexpression of flotillin-1 is involved in proliferation and recurrence of bladder transitional cell carcinoma.

    PubMed

    Guan, Yawei; Song, Haiyan; Zhang, Guohui; Ai, Xing

    2014-08-01

    Flotillin-1 (FLOT1) is known to have a role in tumorigenesis; however, the effect of FLOT1 on proliferation and recurrence of human transitional cell carcinoma (TCC) is unclear. Samples from 156 TCC patients and 142 patients undergoing open bladder surgery for indications other than TCC were used in the present study. FLOT1 protein expression was determined by immunohistochemistry and western blot analysis, and mRNA expression was detected by RT-PCR and real-time PCR. A FLOT1-expressing pcDEF3 vector was stably transfected into 4 TCC cell lines and FLOT1 expression was decreased by RNAi. Proliferative analysis of TCC cells was detected by the WST-1 assay and a xenograft model using BALB/C nude mice. The association between FLOT1 expression and TCC recurrence was also analyzed by adhesion, migration and invasion assays. FLOT1 expression in TCC was significantly overexpressed compared to normal urothelial tissue, and the level of FLOT1 expression was significantly correlated with tumor size, pathologic grade, clinical stage and recurrence. In addition, FLOT1 significantly increased the proliferative ability of TCC cells in vitro and in vivo. TCC cells with a high level of FLOT1 expression exhibited a higher level of adhesion, migration and invasion. FLOT1 expression was shown to be upregulated in human TCC. These findings suggest that FLOT1 plays an important role in the proliferation and recurrence of TCC and that silencing FLOT1 expression might be a novel therapeutic strategy.

  8. Epac Activation Regulates Human Mesenchymal Stem Cells Migration and Adhesion.

    PubMed

    Yu, Jiao-Le; Deng, Ruixia; Chung, Sookja K; Chan, Godfrey Chi-Fung

    2016-04-01

    How to enhance the homing of human mesenchymal stem cells (hMSCs) to the target tissues remains a clinical challenge nowadays. To overcome this barrier, the mechanism responsible for the hMSCs migration and engraftment has to be defined. Currently, the exact mechanism involved in migration and adhesion of hMSCs remains unknown. Exchange protein directly activated by cAMP (Epac), a novel protein discovered in cAMP signaling pathway, may have a potential role in regulating cells adhesion and migration by triggering the downstream Rap family signaling cascades. However, the exact role of Epac in cells homing is elusive. Our study evaluated the role of Epac in the homing of hMSCs. We confirmed that hMSCs expressed functional Epac and its activation enhanced the migration and adhesion of hMSCs significantly. The Epac activation was further found to be contributed directly to the chemotactic responses induced by stromal cell derived factor-1 (SDF-1) which is a known chemokine in regulating hMSCs homing. These findings suggested Epac is connected to the SDF-1 signaling cascades. In conclusion, our study revealed that Epac plays a role in hMSCs homing by promoting adhesion and migration. Appropriate manipulation of Epac may enhance the homing of hMSCs and facilitate their future clinical applications.

  9. Potentialities of ultrasounds for the nondestructive evaluation of cell adhesion.

    PubMed

    Myrdycz, A; Lefebvre, F; Ouaftouh, M; Monchau, F; Callens, D; Hildebrand, H F

    1999-08-01

    The aim of this paper is to present the potentialities of ultrasounds to investigate the mechanical properties of a cell/substrate interface. The adhesion process plays a major role in the development of osteoblastic cells on various substrates used in orthopedic applications such as metals, bioceramics, etc. Particularly, cell adherence appears to be a critical factor in the colonization process. High-frequency and low-power ultrasounds seem to be an appropriate tool for a nondestructive evaluation of interface properties. First, we present the results obtained with bulk longitudinal and shear waves under an arbitrary incidence over an aluminum-adhesive interface. This study was performed for an industrial application of bonding. The results clearly show the sensitivity of shear waves for the evaluation of the adhesion quality owing to the shear solicitations at the interface they induce. A model of ultrasound interactions with a boundary subject to varying degrees of adhesion has been developed and compared to the experiments. Second, we investigated osteoblastic cell cultures with a high-frequency acoustic microscope working at 50 MHz. The images obtained in the shear mode reveal a better contrast than those obtained in the longitudinal mode. For the time being, these results are qualitative, and theoretical models have to be developed according to the point of view of biologists.

  10. How to let go: pectin and plant cell adhesion

    PubMed Central

    Daher, Firas Bou; Braybrook, Siobhan A.

    2015-01-01

    Plant cells do not, in general, migrate. They maintain a fixed position relative to their neighbors, intimately linked through growth and differentiation. The mediator of this connection, the pectin-rich middle lamella, is deposited during cell division and maintained throughout the cell’s life to protect tissue integrity. The maintenance of adhesion requires cell wall modification and is dependent on the actin cytoskeleton. There are developmental processes that require cell separation, such as organ abscission, dehiscence, and ripening. In these instances, the pectin-rich middle lamella must be actively altered to allow cell separation, a process which also requires cell wall modification. In this review, we will focus on the role of pectin and its modification in cell adhesion and separation. Recent insights gained in pectin gel mechanics will be discussed in relation to existing knowledge of pectin chemistry as it relates to cell adhesion. As a whole, we hope to begin defining the physical mechanisms behind a cells’ ability to hang on, and how it lets go. PMID:26236321

  11. Understanding dynamic changes in live cell adhesion with neutron reflectometry

    SciTech Connect

    Junghans, Ann; Waltman, Mary Jo; Smith, Hillary L.; Pocivavsek, Luka; Zebda, Noureddine; Birukov, Konstantin; Viapiano, Mariano; Majewski, Jaroslaw

    2014-12-10

    In this study, neutron reflectometry (NR) was used to examine various live cells' adhesion to quartz substrates under different environmental conditions, including flow stress. To the best of our knowledge, these measurements represent the first successful visualization and quantization of the interface between live cells and a substrate with sub-nanometer resolution. In our first experiments, we examined live mouse fibroblast cells as opposed to past experiments using supported lipids, proteins, or peptide layers with no associated cells. We continued the NR studies of cell adhesion by investigating endothelial monolayers and glioblastoma cells under dynamic flow conditions. We demonstrated that neutron reflectometry is a powerful tool to study the strength of cellular layer adhesion in living tissues, which is a key factor in understanding the physiology of cell interactions and conditions leading to abnormal or disease circumstances. Continuative measurements, such as investigating changes in tumor cell — surface contact of various glioblastomas, could impact advancements in tumor treatments. In principle, this can help us to identify changes that correlate with tumor invasiveness. Pursuit of these studies can have significant medical impact on the understanding of complex biological problems and their effective treatment, e.g. for the development of targeted anti-invasive therapies.

  12. Understanding dynamic changes in live cell adhesion with neutron reflectometry

    DOE PAGES

    Junghans, Ann; Waltman, Mary Jo; Smith, Hillary L.; ...

    2014-12-10

    In this study, neutron reflectometry (NR) was used to examine various live cells' adhesion to quartz substrates under different environmental conditions, including flow stress. To the best of our knowledge, these measurements represent the first successful visualization and quantization of the interface between live cells and a substrate with sub-nanometer resolution. In our first experiments, we examined live mouse fibroblast cells as opposed to past experiments using supported lipids, proteins, or peptide layers with no associated cells. We continued the NR studies of cell adhesion by investigating endothelial monolayers and glioblastoma cells under dynamic flow conditions. We demonstrated that neutronmore » reflectometry is a powerful tool to study the strength of cellular layer adhesion in living tissues, which is a key factor in understanding the physiology of cell interactions and conditions leading to abnormal or disease circumstances. Continuative measurements, such as investigating changes in tumor cell — surface contact of various glioblastomas, could impact advancements in tumor treatments. In principle, this can help us to identify changes that correlate with tumor invasiveness. Pursuit of these studies can have significant medical impact on the understanding of complex biological problems and their effective treatment, e.g. for the development of targeted anti-invasive therapies.« less

  13. Understanding dynamic changes in live cell adhesion with neutron reflectometry

    PubMed Central

    JUNGHANS, ANN; WALTMAN, MARY JO; SMITH, HILLARY L.; POCIVAVSEK, LUKA; ZEBDA, NOUREDDINE; BIRUKOV, KONSTANTIN; VIAPIANO, MARIANO; MAJEWSKI, JAROSLAW

    2015-01-01

    Neutron reflectometry (NR) was used to examine various live cells adhesion to quartz substrates under different environmental conditions, including flow stress. To the best of our knowledge, these measurements represent the first successful visualization and quantization of the interface between live cells and a substrate with sub-nanometer resolution. In our first experiments, we examined live mouse fibroblast cells as opposed to past experiments using supported lipids, proteins, or peptide layers with no associated cells. We continued the NR studies of cell adhesion by investigating endothelial monolayers and glioblastoma cells under dynamic flow conditions. We demonstrated that neutron reflectometry is a powerful tool to study the strength of cellular layer adhesion in living tissues, which is a key factor in understanding the physiology of cell interactions and conditions leading to abnormal or disease circumstances. Continuative measurements, such as investigating changes in tumor cell – surface contact of various glioblastomas, could impact advancements in tumor treatments. In principle, this can help us to identify changes that correlate with tumor invasiveness. Pursuit of these studies can have significant medical impact on the understanding of complex biological problems and their effective treatment, e.g. for the development of targeted anti-invasive therapies. PMID:25705067

  14. Understanding dynamic changes in live cell adhesion with neutron reflectometry

    NASA Astrophysics Data System (ADS)

    Junghans, Ann; Waltman, Mary Jo; Smith, Hillary L.; Pocivavsek, Luka; Zebda, Noureddine; Birukov, Konstantin; Viapiano, Mariano; Majewski, Jaroslaw

    2014-12-01

    Neutron reflectometry (NR) was used to examine various live cells' adhesion to quartz substrates under different environmental conditions, including flow stress. To the best of our knowledge, these measurements represent the first successful visualization and quantization of the interface between live cells and a substrate with sub-nanometer resolution. In our first experiments, we examined live mouse fibroblast cells as opposed to past experiments using supported lipids, proteins, or peptide layers with no associated cells. We continued the NR studies of cell adhesion by investigating endothelial monolayers and glioblastoma cells under dynamic flow conditions. We demonstrated that neutron reflectometry is a powerful tool to study the strength of cellular layer adhesion in living tissues, which is a key factor in understanding the physiology of cell interactions and conditions leading to abnormal or disease circumstances. Continuative measurements, such as investigating changes in tumor cell — surface contact of various glioblastomas, could impact advancements in tumor treatments. In principle, this can help us to identify changes that correlate with tumor invasiveness. Pursuit of these studies can have significant medical impact on the understanding of complex biological problems and their effective treatment, e.g. for the development of targeted anti-invasive therapies.

  15. The endogenous cannabinoid anandamide inhibits human breast cancer cell proliferation

    PubMed Central

    De Petrocellis, Luciano; Melck, Dominique; Palmisano, Antonella; Bisogno, Tiziana; Laezza, Chiara; Bifulco, Maurizio; Di Marzo, Vincenzo

    1998-01-01

    Anandamide was the first brain metabolite shown to act as a ligand of “central” CB1 cannabinoid receptors. Here we report that the endogenous cannabinoid potently and selectively inhibits the proliferation of human breast cancer cells in vitro. Anandamide dose-dependently inhibited the proliferation of MCF-7 and EFM-19 cells with IC50 values between 0.5 and 1.5 μM and 83–92% maximal inhibition at 5–10 μM. The proliferation of several other nonmammary tumoral cell lines was not affected by 10 μM anandamide. The anti-proliferative effect of anandamide was not due to toxicity or to apoptosis of cells but was accompanied by a reduction of cells in the S phase of the cell cycle. A stable analogue of anandamide (R)-methanandamide, another endogenous cannabinoid, 2-arachidonoylglycerol, and the synthetic cannabinoid HU-210 also inhibited EFM-19 cell proliferation, whereas arachidonic acid was much less effective. These cannabimimetic substances displaced the binding of the selective cannabinoid agonist [3H]CP 55,940 to EFM-19 membranes with an order of potency identical to that observed for the inhibition of EFM-19 cell proliferation. Moreover, anandamide cytostatic effect was inhibited by the selective CB1 receptor antagonist SR 141716A. Cell proliferation was arrested by a prolactin mAb and enhanced by exogenous human prolactin, whose mitogenic action was reverted by very low (0.1–0.5 μM) doses of anandamide. Anandamide suppressed the levels of the long form of the prolactin receptor in both EFM-19 and MCF-7 cells, as well as a typical prolactin-induced response, i.e., the expression of the breast cancer cell susceptibility gene brca1. These data suggest that anandamide blocks human breast cancer cell proliferation through CB1-like receptor-mediated inhibition of endogenous prolactin action at the level of prolactin receptor. PMID:9653194

  16. Effects of plasma treated PET and PTFE on expression of adhesion molecules by human endothelial cells in vitro.

    PubMed

    Pu, F R; Williams, R L; Markkula, T K; Hunt, J A

    2002-06-01

    The aim of this study was to evaluate the expression of adhesion molecules on the surface of human endothelial cells in response to the systematic variation in materials properties by the ammonia plasma modification of polyethylene terephthalate (PET) and polytetrafluorethylene (PTFE). These adhesion molecules act as mediators of cell adhesion, play a role in the modulation of cell adhesion on biomaterials and therefore condition the response of tissues to implants. First and second passage human umbilical vein endothelial cells (HUVECs) were cultured on plasma treated and untreated PET and PTFE. HUVECs grown on polystyrene tissue culture coverslips and HUVECs stimulated with tumour necrosis factor (TNF-alpha) were used as controls. After 1 day and 7 days, the expression of adhesion molecules platelet endothelial cell adhesion molecule-1 (PECAM-1), intercellular adhesion molecule-1 (ICAM-1), Integrin alphavbeta3, vascular cell adhesion molecule-1 (VCAM-1), E-selectin, P-selectin and L-selectin were evaluated using flow cytometry and immunohistochemistry. There was a slight increase in positive cell numbers expressing the adhesion molecules ICAM-1 and VCAM-1 on plasma treated PET and PTFE. A significant increase in E-selectin positive cells on untreated PTFE was demonstrated after 7 days. Stimulation with TNF-alpha demonstrated a significant increase in the proportion of ICAM-1. VCAM-1 and E-selectin positive cells. Almost all cells expressed PECAM-1 and integrin alphavbeta3, on both materials and controls but did not express P- and L-selectin on any surface. When second passage cells were used, the expression of the adhesion molecules ICAM-1 and VCAM-1 was markedly increased on all surfaces but not with TNF-alpha. These significant differences were not observed in other adhesion molecules. These results were supported by immunohistochemical studies. The effects of plasma treated PET and PTFE on cell adhesion and proliferation was also studied. There was a 1.3-fold

  17. Chitosan Feasibility to Retain Retinal Stem Cell Phenotype and Slow Proliferation for Retinal Transplantation

    PubMed Central

    Srivastava, Girish K.; Rodriguez-Crespo, David; Singh, Amar K.; Casado-Coterillo, Clara; Garcia-Gutierrez, Maria T.; Coronas, Joaquin; Pastor, J. Carlos

    2014-01-01

    Retinal stem cells (RSCs) are promising in cell replacement strategies for retinal diseases. RSCs can migrate, differentiate, and integrate into retina. However, RSCs transplantation needs an adequate support; chitosan membrane (ChM) could be one, which can carry RSCs with high feasibility to support their integration into retina. RSCs were isolated, evaluated for phenotype, and subsequently grown on sterilized ChM and polystyrene surface for 8 hours, 1, 4, and 11 days for analysing cell adhesion, proliferation, viability, and phenotype. Isolated RSCs expressed GFAP, PKC, isolectin, recoverin, RPE65, PAX-6, cytokeratin 8/18, and nestin proteins. They adhered (28 ± 16%, 8 hours) and proliferated (40 ± 20 cells/field, day 1 and 244 ± 100 cells/field, day 4) significantly low (P < 0.05) on ChM. However, they maintained similar viability (>95%) and phenotype (cytokeratin 8/18, PAX6, and nestin proteins expression, day 11) on both surfaces (ChM and polystyrene). RSCs did not express alpha-SMA protein on both surfaces. RSCs express proteins belonging to epithelial, glial, and neural cells, confirming that they need further stimulus to reach a final destination of differentiation that could be provided in in vivo condition. ChM does not alternate RSCs behaviour and therefore can be used as a cell carrier so that slow proliferating RSCs can migrate and integrate into retina. PMID:24719852

  18. The Evolutionary Origin of Epithelial Cell-Cell Adhesion Mechanisms

    PubMed Central

    Miller, Phillip W.; Clarke, Donald N.; Weis, William I.; Lowe, Christopher J.; Nelson, W. James

    2014-01-01

    SUMMARY A simple epithelium forms a barrier between the outside and the inside of an organism, and is the first organized multicellular tissue found in evolution. We examine the relationship between the evolution of epithelia and specialized cell-cell adhesion proteins comprising the classical cadherin/β-catenin/α-catenin complex (CCC). A review of the divergent functional properties of the CCC in metazoans and non-metazoans, and an updated phylogenetic coverage of the CCC using recent genomic data reveal: 1) The core CCC likely originated before the last common ancestor of unikonts and their closest bikont sister taxa. 2) Formation of the CCC may have constrained sequence evolution of the classical cadherin cytoplasmic domain and β-catenin in metazoa. 3) The α-catenin binding domain in β-catenin appears to be the favored mutation site for disrupting β-catenin function in the CCC. 4) The ancestral function of the α/β-catenin heterodimer appears to be an actin-binding module. In some metazoan groups, more complex functions of α-catenin were gained by sequence divergence in the non-actin binding (N-, M-) domains. 5) Allosteric regulation of α-catenin, rather than loss of function mutations, may have evolved for more complex regulation of the actin cytoskeleton. PMID:24210433

  19. Different effects of 25-kDa amelogenin on the proliferation, attachment and migration of various periodontal cells

    SciTech Connect

    Li, Xiting; Shu, Rong; Liu, Dali; Jiang, Shaoyun

    2010-04-09

    Previous studies have assumed that amelogenin is responsible for the therapeutic effect of the enamel matrix derivative (EMD) in periodontal tissue healing and regeneration. However, it is difficult to confirm this hypothesis because both the EMD and the amelogenins are complex mixtures of multiple proteins. Further adding to the difficulties is the fact that periodontal tissue regeneration involves various types of cells and a sequence of associated cellular events including the attachment, migration and proliferation of various cells. In this study, we investigated the potential effect of a 25-kDa recombinant porcine amelogenin (rPAm) on primarily cultured periodontal ligament fibroblasts (PDLF), gingival fibroblasts (GF) and gingival epithelial cells (GEC). The cells were treated with 25-kDa recombinant porcine amelogenin at a concentration of 10 {mu}g/mL. We found that rPAm significantly promoted the proliferation and migration of PDLF, but not their adhesion. Similarly, the proliferation and adhesion of GF were significantly enhanced by treatment with rPAm, while migration was greatly inhibited. Interestingly, this recombinant protein inhibited the growth rate, cell adhesion and migration of GEC. These data suggest that rPAm may play an essential role in periodontal regeneration through the activation of periodontal fibroblasts and inhibition of the cellular behaviors of gingival epithelial cells.

  20. Differential migration and proliferation of geometrical ensembles of cell clusters

    SciTech Connect

    Kumar, Girish; Chen, Bo; Co, Carlos C.; Ho, Chia-Chi

    2011-06-10

    Differential cell migration and growth drives the organization of specific tissue forms and plays a critical role in embryonic development, tissue morphogenesis, and tumor invasion. Localized gradients of soluble factors and extracellular matrix have been shown to modulate cell migration and proliferation. Here we show that in addition to these factors, initial tissue geometry can feedback to generate differential proliferation, cell polarity, and migration patterns. We apply layer by layer polyelectrolyte assembly to confine multicellular organization and subsequently release cells to demonstrate the spatial patterns of cell migration and growth. The cell shapes, spreading areas, and cell-cell contacts are influenced strongly by the confining geometry. Cells within geometric ensembles are morphologically polarized. Symmetry breaking was observed for cells on the circular pattern and cells migrate toward the corners and in the direction parallel to the longest dimension of the geometric shapes. This migration pattern is disrupted when actomyosin based tension was inhibited. Cells near the edge or corner of geometric shapes proliferate while cells within do not. Regions of higher rate of cell migration corresponded to regions of concentrated growth. These findings demonstrate that multicellular organization can result in spatial patterns of migration and proliferation.

  1. Scaffold architecture and fibrin gels promote meniscal cell proliferation

    SciTech Connect

    Pawelec, K. M. E-mail: jw626@cam.ac.uk; Best, S. M.; Cameron, R. E.; Wardale, R. J. E-mail: jw626@cam.ac.uk

    2015-01-01

    Stability of the knee relies on the meniscus, a complex connective tissue with poor healing ability. Current meniscal tissue engineering is inadequate, as the signals for increasing meniscal cell proliferation have not been established. In this study, collagen scaffold structure, isotropic or aligned, and fibrin gel addition were tested. Metabolic activity was promoted by fibrin addition. Cellular proliferation, however, was significantly increased by both aligned architectures and fibrin addition. None of the constructs impaired collagen type I production or triggered adverse inflammatory responses. It was demonstrated that both fibrin gel addition and optimized scaffold architecture effectively promote meniscal cell proliferation.

  2. Intrinsic effects of gold nanoparticles on proliferation and invasion activity in SGC-7901 cells.

    PubMed

    Wu, Yucheng; Zhang, Qingqing; Ruan, Zhongbao; Yin, Yigang

    2016-03-01

    Although biomedical applications of functionalized nanoparticles have taken significant strides, biological characterization of unmodified nanoparticles remains unclear. In the present study, we investigated the cell viability and invasion activity of gastric cancer cells after treatment with gold nanoparticles. The growth of SGC-7901 cells was inhibited significantly after treatment with 5-nm gold nanoparticles, and the cell invasion decreased markedly. These effects were not seen by different size gold nanoparticles (10, 20 and 40 nm). The attenuated invasion activity may be associated with the decreased expression of matrix metalloproteinase 9 and intercellular adhesion molecule-1. These data indicated that the response of SGC-7901 cells to gold nanoparticles was strongly associated with their unique size-dependent physiochemical properties. Therefore, we provided new evidence for the effect of gold nanoparticles on gastric cancer cell proliferation and invasion in vitro, making a contribution to the application of gold nanoparticles to novel therapies in gastric cancer.

  3. Endoglin regulates mural cell adhesion in the circulatory system.

    PubMed

    Rossi, Elisa; Smadja, David M; Boscolo, Elisa; Langa, Carmen; Arevalo, Miguel A; Pericacho, Miguel; Gamella-Pozuelo, Luis; Kauskot, Alexandre; Botella, Luisa M; Gaussem, Pascale; Bischoff, Joyce; Lopez-Novoa, José M; Bernabeu, Carmelo

    2016-04-01

    The circulatory system is walled off by different cell types, including vascular mural cells and podocytes. The interaction and interplay between endothelial cells (ECs) and mural cells, such as vascular smooth muscle cells or pericytes, play a pivotal role in vascular biology. Endoglin is an RGD-containing counter-receptor for β1 integrins and is highly expressed by ECs during angiogenesis. We find that the adhesion between vascular ECs and mural cells is enhanced by integrin activators and inhibited upon suppression of membrane endoglin or β1-integrin, as well as by addition of soluble endoglin (SolEng), anti-integrin α5β1 antibody or an RGD peptide. Analysis of different endoglin mutants, allowed the mapping of the endoglin RGD motif as involved in the adhesion process. In Eng (+/-) mice, a model for hereditary hemorrhagic telangectasia type 1, endoglin haploinsufficiency induces a pericyte-dependent increase in vascular permeability. Also, transgenic mice overexpressing SolEng, an animal model for preeclampsia, show podocyturia, suggesting that SolEng is responsible for podocytes detachment from glomerular capillaries. These results suggest a critical role for endoglin in integrin-mediated adhesion of mural cells and provide a better understanding on the mechanisms of vessel maturation in normal physiology as well as in pathologies such as preeclampsia or hereditary hemorrhagic telangiectasia.

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

    NASA Astrophysics Data System (ADS)

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

    2008-12-01

    Current small diameter (<5 mm) synthetic vascular graft materials exhibit poor long-term patency due to thrombosis and intimal hyperplasia. Tissue engineered solutions have yielded functional vascular tissue, but some require an eight-week in vitro culture period prior to implantation—too long for immediate clinical bedside applications. Previous in vitro studies have shown that nanostructured poly(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.

  5. Inhibition of brain tumor cell proliferation by alternating electric fields

    SciTech Connect

    Jeong, Hyesun; Oh, Seung-ick; Hong, Sunghoi E-mail: radioyoon@korea.ac.kr; Sung, Jiwon; Jeong, Seonghoon; Yoon, Myonggeun E-mail: radioyoon@korea.ac.kr; Koh, Eui Kwan

    2014-11-17

    This study was designed to investigate the mechanism by which electric fields affect cell function, and to determine the optimal conditions for electric field inhibition of cancer cell proliferation. Low-intensity (<2 V/cm) and intermediate-frequency (100–300 kHz) alternating electric fields were applied to glioblastoma cell lines. These electric fields inhibited cell proliferation by inducing cell cycle arrest and abnormal mitosis due to the malformation of microtubules. These effects were significantly dependent on the intensity and frequency of applied electric fields.

  6. Cell adhesion defines the topology of endocytosis and signaling

    PubMed Central

    Grossier, Jean-Philippe; Xouri, Georgia; Goud, Bruno; Schauer, Kristine

    2014-01-01

    Preferred sites of endocytosis have been observed in various cell types, but whether they occur randomly or are linked to cellular cues is debated. Here, we quantified the sites of endocytosis of transferrin (Tfn) and epidermal growth factor (EGF) in cells whose adhesion geometry was defined by micropatterns. 3D probabilistic density maps revealed that Tfn was enriched in adhesive sites during uptake, whereas EGF endocytosis was restricted to the dorsal cellular surface. This spatial separation was not due to distributions of corresponding receptors but was regulated by uptake mechanisms. Asymmetric uptake of Tfn resulted from the enrichment of clathrin and adaptor protein 2 at adhesive areas. Asymmetry in EGF uptake was strongly dependent on the actin cytoskeleton and led to asymmetry in EGF receptor activation. Mild alteration of actin dynamics abolished asymmetry in EGF uptake and decreased EGF-induced downstream signaling, suggesting that cellular adhesion cues influence signal propagation. We propose that restriction of endocytosis at distinct sites allows cells to sense their environment in an “outside-in” mechanism. PMID:24366944

  7. EDA-Containing Fibronectin Increases Proliferation of Embryonic Stem Cells

    PubMed Central

    Losino, Noelia; Waisman, Ariel; Solari, Claudia; Luzzani, Carlos; Espinosa, Darío Fernández; Sassone, Alina; Muro, Andrés F.; Miriuka, Santiago; Sevlever, Gustavo; Barañao, Lino; Guberman, Alejandra

    2013-01-01

    Embryonic stem cells (ESC) need a set of specific factors to be propagated. They can also grow in conditioned medium (CM) derived from a bovine granulosa cell line BGC (BGC-CM), a medium that not only preserves their main features but also increases ESC´s proliferation rate. The mitogenic properties of this medium were previously reported, ascribing this effect to an alternative spliced generated fibronectin isoform that contains the extra domain A (FN EDA+). Here, we investigated if the FN EDA+ isoform increased proliferation of mouse and human ES cells. We analyzed cell proliferation using conditioned media produced by different mouse embryonic fibroblast (MEF) lines genetically engineered to express FN constitutively including or excluding the EDA domain (FN EDA-), and in media supplemented with recombinant peptides containing or not the EDA. We found that the presence of EDA in the medium increased mouse and human ESC’s proliferation rate. Here we showed for the first time that this FN isoform enhances ESC’s proliferation. These findings suggest a possible conserved behavior for regulation of ES cells proliferation by this FN isoform and could contribute to improve their culturing conditions both for research and cell therapy. PMID:24244705

  8. EDA-containing fibronectin increases proliferation of embryonic stem cells.

    PubMed

    Losino, Noelia; Waisman, Ariel; Solari, Claudia; Luzzani, Carlos; Espinosa, Darío Fernández; Sassone, Alina; Muro, Andrés F; Miriuka, Santiago; Sevlever, Gustavo; Barañao, Lino; Guberman, Alejandra

    2013-01-01

    Embryonic stem cells (ESC) need a set of specific factors to be propagated. They can also grow in conditioned medium (CM) derived from a bovine granulosa cell line BGC (BGC-CM), a medium that not only preserves their main features but also increases ESC´s proliferation rate. The mitogenic properties of this medium were previously reported, ascribing this effect to an alternative spliced generated fibronectin isoform that contains the extra domain A (FN EDA(+)). Here, we investigated if the FN EDA(+) isoform increased proliferation of mouse and human ES cells. We analyzed cell proliferation using conditioned media produced by different mouse embryonic fibroblast (MEF) lines genetically engineered to express FN constitutively including or excluding the EDA domain (FN EDA(-)), and in media supplemented with recombinant peptides containing or not the EDA. We found that the presence of EDA in the medium increased mouse and human ESC's proliferation rate. Here we showed for the first time that this FN isoform enhances ESC's proliferation. These findings suggest a possible conserved behavior for regulation of ES cells proliferation by this FN isoform and could contribute to improve their culturing conditions both for research and cell therapy.

  9. Ameloblastin regulates cell attachment and proliferation through RhoA and p27.

    PubMed

    Zhang, Youbin; Zhang, Xu; Lu, Xuanyu; Atsawasuwan, Phimon; Luan, Xianghong

    2011-12-01

    The matrix adhesion protein ameloblastin (AMBN) is one of the unique components of the mineralizing matrix of bones and teeth. Here we focused on two types of cells expressing AMBN - mouse dental follicle cells (mDF) and mouse periodontal ligament cells (mPDL) - to decipher AMBN function in developing dental, periodontal, and bone tissues. To test AMBN function, cell culture dishes of mDF and mPDL were exposed to either full-length or C-terminal (amino acids 137-407) recombinant Ambn protein. Alternatively, cells were subjected to transient transfection using an Ambn-small hairpin (sh) RNA vector. Our cell culture studies documented that dishes coated with full-length AMBN promoted the attachment of mPDL and mDF cells as early as 1 h after seeding. In order to identify potential intermediaries that might aid the effect of AMBN on adhesion, RhoA expression levels in AMBN-coated and uncoated control dishes were assessed. These studies indicated that AMBN induced RhoA expression 4 h after seeding, especially in mPDL cells. After 4 h of culture, the cell cycle inhibitor p27 was also up-regulated. In addition, exogenous AMBN and its C-terminal fragment reduced the proliferation of mDF and mPDL. Finally, transient transfection of mDF and mPDL cells with the Ambn-shRNA vector resulted in the down-regulation of p27 in mPDL cells. Together, these data indicate that AMBN affects cell adhesion via RhoA and cell cycle progression through p27.

  10. Proliferation, behavior, and cytokine gene expression of human umbilical vascular endothelial cells in response to different titanium surfaces.

    PubMed

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

    2010-04-01

    Success of dental implantation is initially affected by wound healing of both, hard and soft tissues. Endothelial cells (ECs) are involved as crucial cells in the angiogenesis and inflammation process of wound healing. In the present study, proliferation, mobility, cluster formation, and gene expression of angiogenesis-related molecules of human umbilical vascular endothelial cells (HUVECs) were investigated on titanium surfaces with different roughnesses: acid-etched (A), coarse-grit-blasted and acid-etched (SLA) surfaces, as well as on hydrophilic modified modA and modSLA surfaces. Cell behaviors were analyzed by proliferation assay and time-lapse microscopy, gene expression was analyzed by real time PCR. Results showed that cell proliferation, mobility, and cluster formation were highest on modA surfaces compared with all other surfaces. HUVECs moved slowly and exhibited seldom cell aggregation on SLA and modSLA surfaces during the whole observing period of 120 h. The gene expressions of the angiogenesis-related factors von Willebrand factor, thrombomodulin, endothelial cell protein C receptor, and adhesion molecules intercellular adhesion molecule-1 and E-selectin were most enhanced on modSLA surfaces. These results suggest that modA surface is optimal for proliferation and angiogenic behavior of ECs. However, modSLA surface seems to promote ECs to express angiogenesis-related factor genes, which play essential roles in controlling inflammation and revascularization of wound healing.

  11. Topographic cell instructive patterns to control cell adhesion, polarization and migration

    PubMed Central

    Ventre, Maurizio; Natale, Carlo Fortunato; Rianna, Carmela; Netti, Paolo Antonio

    2014-01-01

    Topographic patterns are known to affect cellular processes such as adhesion, migration and differentiation. However, the optimal way to deliver topographic signals to provide cells with precise instructions has not been defined yet. In this work, we hypothesize that topographic patterns may be able to control the sensing and adhesion machinery of cells when their interval features are tuned on the characteristic lengths of filopodial probing and focal adhesions (FAs). Features separated by distance beyond the length of filopodia cannot be readily perceived; therefore, the formation of new adhesions is discouraged. If, however, topographic features are separated by a distance within the reach of filopodia extension, cells can establish contact between adjacent topographic islands. In the latter case, cell adhesion and polarization rely upon the growth of FAs occurring on a specific length scale that depends on the chemical properties of the surface. Topographic patterns and chemical properties may interfere with the growth of FAs, thus making adhesions unstable. To test this hypothesis, we fabricated different micropatterned surfaces displaying feature dimensions and adhesive properties able to interfere with the filopodial sensing and the adhesion maturation, selectively. Our data demonstrate that it is possible to exert a potent control on cell adhesion, elongation and migration by tuning topographic features’ dimensions and surface chemistry. PMID:25253035

  12. Enzymatically crosslinked gelatin hydrogel promotes the proliferation of adipose tissue-derived stromal cells

    PubMed Central

    Ren, Xiaomei; Long, Haiyan; Qian, Hong; Ma, Kunlong

    2016-01-01

    Gelatin hydrogel crosslinked by microbial transglutaminase (mTG) exhibits excellent performance in cell adhesion, proliferation, and differentiation. We examined the gelation time and gel strength of gelatin/mTG hydrogels in various proportions to investigate their physical properties and tested their degradation performances in vitro. Cell morphology and viability of adipose tissue-derived stromal cells (ADSCs) cultured on the 2D gel surface or in 3D hydrogel encapsulation were evaluated by immunofluorescence staining. Cell proliferation was tested via Alamar Blue assay. To investigate the hydrogel effect on cell differentiation, the cardiac-specific gene expression levelsof Nkx2.5, Myh6, Gja1, and Mef2c in encapsulated ADSCs with or without cardiac induction medium were detected by real-time RT-PCR. Cell release from the encapsulated status and cell migration in a 3D hydrogel model were assessed in vitro. Results show that the gelatin/mTG hydrogels are not cytotoxic and that their mechanical properties are adjustable. Hydrogel degradation is related to gel concentration and the resident cells. Cell growth morphology and proliferative capability in both 2D and 3D cultures were mainly affected by gel concentration. PCR result shows that hydrogel modulus together with induction medium affects the cardiac differentiation of ADSCs. The cell migration experiment and subcutaneous implantation show that the hydrogels are suitable for cell delivery. PMID:27703850

  13. Enzymatically crosslinked gelatin hydrogel promotes the proliferation of adipose tissue-derived stromal cells.

    PubMed

    Yang, Gang; Xiao, Zhenghua; Ren, Xiaomei; Long, Haiyan; Qian, Hong; Ma, Kunlong; Guo, Yingqiang

    2016-01-01

    Gelatin hydrogel crosslinked by microbial transglutaminase (mTG) exhibits excellent performance in cell adhesion, proliferation, and differentiation. We examined the gelation time and gel strength of gelatin/mTG hydrogels in various proportions to investigate their physical properties and tested their degradation performances in vitro. Cell morphology and viability of adipose tissue-derived stromal cells (ADSCs) cultured on the 2D gel surface or in 3D hydrogel encapsulation were evaluated by immunofluorescence staining. Cell proliferation was tested via Alamar Blue assay. To investigate the hydrogel effect on cell differentiation, the cardiac-specific gene expression levelsof Nkx2.5, Myh6, Gja1, and Mef2c in encapsulated ADSCs with or without cardiac induction medium were detected by real-time RT-PCR. Cell release from the encapsulated status and cell migration in a 3D hydrogel model were assessed in vitro. Results show that the gelatin/mTG hydrogels are not cytotoxic and that their mechanical properties are adjustable. Hydrogel degradation is related to gel concentration and the resident cells. Cell growth morphology and proliferative capability in both 2D and 3D cultures were mainly affected by gel concentration. PCR result shows that hydrogel modulus together with induction medium affects the cardiac differentiation of ADSCs. The cell migration experiment and subcutaneous implantation show that the hydrogels are suitable for cell delivery.

  14. A Review of Cell Adhesion Studies for Biomedical and Biological Applications

    PubMed Central

    Ahmad Khalili, Amelia; Ahmad, Mohd Ridzuan

    2015-01-01

    Cell adhesion is essential in cell communication and regulation, and is of fundamental importance in the development and maintenance of tissues. The mechanical interactions between a cell and its extracellular matrix (ECM) can influence and control cell behavior and function. The essential function of cell adhesion has created tremendous interests in developing methods for measuring and studying cell adhesion properties. The study of cell adhesion could be categorized into cell adhesion attachment and detachment events. The study of cell adhesion has been widely explored via both events for many important purposes in cellular biology, biomedical, and engineering fields. Cell adhesion attachment and detachment events could be further grouped into the cell population and single cell approach. Various techniques to measure cell adhesion have been applied to many fields of study in order to gain understanding of cell signaling pathways, biomaterial studies for implantable sensors, artificial bone and tooth replacement, the development of tissue-on-a-chip and organ-on-a-chip in tissue engineering, the effects of biochemical treatments and environmental stimuli to the cell adhesion, the potential of drug treatments, cancer metastasis study, and the determination of the adhesion properties of normal and cancerous cells. This review discussed the overview of the available methods to study cell adhesion through attachment and detachment events. PMID:26251901

  15. Role of Calmodulin in Cell Proliferation

    NASA Technical Reports Server (NTRS)

    Chafouleas, J.

    1983-01-01

    Calmodulin levels were found to increase as cells enter plateau. The data suggest that the cells are exiting the cell cycle late in the G sub 1 phase, or that the calmodulin levels in plateau cells are uncoupled to progression into S phase in plateau cells. Upon release, calmodulin levels rapidly decrease. Following this decrease, there is a increase prior to S phase.

  16. The role of adhesion energy in controlling cell–cell contacts

    PubMed Central

    Maître, Jean-Léon; Heisenberg, Carl-Philipp

    2011-01-01

    Recent advances in microscopy techniques and biophysical measurements have provided novel insight into the molecular, cellular and biophysical basis of cell adhesion. However, comparably little is known about a core element of cell–cell adhesion—the energy of adhesion at the cell–cell contact. In this review, we discuss approaches to understand the nature and regulation of adhesion energy, and propose strategies to determine adhesion energy between cells in vitro and in vivo. PMID:21807491

  17. Nanocarbon Allotropes-Graphene and Nanocrystalline Diamond-Promote Cell Proliferation.

    PubMed

    Verdanova, Martina; Rezek, Bohuslav; Broz, Antonin; Ukraintsev, Egor; Babchenko, Oleg; Artemenko, Anna; Izak, Tibor; Kromka, Alexander; Kalbac, Martin; Hubalek Kalbacova, Marie

    2016-05-01

    Two profoundly different carbon allotropes - nanocrystalline diamond and graphene - are of considerable interest from the viewpoint of a wide range of biomedical applications including implant coating, drug and gene delivery, cancer therapy, and biosensing. Osteoblast adhesion and proliferation on nanocrystalline diamond and graphene are compared under various conditions such as differences in wettability, topography, and the presence or absence of protein interlayers between cells and the substrate. The materials are characterized in detail by means of scanning electron microscopy, atomic force microscopy, photoelectron spectroscopy, Raman spectroscopy, and contact angle measurements. In vitro experiments have revealed a significantly higher degree of cell proliferation on graphene than on nanocrystalline diamond and a tissue culture polystyrene control material. Proliferation is promoted, in particular, by hydrophobic graphene with a large number of nanoscale wrinkles independent of the presence of a protein interlayer, i.e., substrate fouling is not a problematic issue in this respect. Nanowrinkled hydrophobic graphene, thus, exhibits superior characteristics for those biomedical applications where high cell proliferation is required under differing conditions.

  18. Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro

    SciTech Connect

    Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang Zhang, Yi

    2013-11-01

    Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients.

  19. Effects of ciglitazone and troglitazone on the proliferation of human stomach cancer cells

    PubMed Central

    Cheon, Chan Woo; Kim, Dae Hwan; Kim, Dong Heon; Cho, Yong Hoon; Kim, Jae Hun

    2009-01-01

    , and cell adhesion, and were also associated with reductions in cell proliferation, the cell cycle, nuclear metabolism, and phosphorylation. CONCLUSION: Troglitazone and ciglitazone suppress the proliferation of stomach cancer cells via a PPAR-γ-independent pathway. PMID:19140230

  20. Understanding dynamic changes in live cell adhesion with neutron reflectometry

    NASA Astrophysics Data System (ADS)

    Junghans, Ann

    Understanding the structure and functionality of biological systems on a nanometer-resolution and short temporal scales is important for solving complex biological problems, developing innovative treatment, and advancing the design of highly functionalized biomimetic materials. For example, adhesion of cells to an underlying substrate plays a crucial role in physiology and disease development, and has been investigated with great interest for several decades. In the talk, we would like to highlight recent advances in utilizing neutron scattering to study bio-related structures in dynamic conditions (e . g . under the shear flow) including in-situ investigations of the interfacial properties of living cells. The strength of neutron reflectometry is its non-pertubative nature, the ability to probe buried interfaces with nanometer resolution and its sensitivity to light elements like hydrogen and carbon. That allows us to study details of cell - substrate interfaces that are not accessible with any other standard techniques. We studied the adhesion of human brain tumor cells (U251) to quartz substrates and their responses to the external mechanical forces. Such cells are isolated within the central nervous system which makes them difficult to reach with conventional therapies and therefore making them highly invasive. Our results reveal changes in the thickness and composition of the adhesion layer (a layer between the cell lipid membrane and the quartz substrate), largely composed of hyaluronic acid and associated proteoglycans, when the cells were subjected to shear stress. Further studies will allow us to determine more conditions triggering changes in the composition of the bio-material in the adhesion layer. This, in turn, can help to identify changes that correlate with tumor invasiveness, which can have significant medical impact for the development of targeted anti-invasive therapies.

  1. ADAMTS-10 and -6 differentially regulate cell-cell junctions and focal adhesions

    PubMed Central

    Cain, Stuart A.; Mularczyk, Ewa J.; Singh, Mukti; Massam-Wu, Teresa; Kielty, Cay M.

    2016-01-01

    ADAMTS10 and ADAMTS6 are homologous metalloproteinases with ill-defined roles. ADAMTS10 mutations cause Weill-Marchesani syndrome (WMS), implicating it in fibrillin microfibril biology since some fibrillin-1 mutations also cause WMS. However little is known about ADAMTS6 function. ADAMTS10 is resistant to furin cleavage, however we show that ADAMTS6 is effectively processed and active. Using siRNA, over-expression and mutagenesis, it was found ADAMTS6 inhibits and ADAMTS10 is required for focal adhesions, epithelial cell-cell junction formation, and microfibril deposition. Either knockdown of ADAMTS6, or disruption of its furin processing or catalytic sites restores focal adhesions, implicating its enzyme activity acts on targets in the focal adhesion complex. In ADAMTS10-depleted cultures, expression of syndecan-4 rescues focal adhesions and cell-cell junctions. Recombinant C-termini of ADAMTS10 and ADAMTS6, both of which induce focal adhesions, bind heparin and syndecan-4. However, cells overexpressing full-length ADAMTS6 lack heparan sulphate and focal adhesions, whilst depletion of ADAMTS6 induces a prominent glycocalyx. Thus ADAMTS10 and ADAMTS6 oppositely affect heparan sulphate-rich interfaces including focal adhesions. We previously showed that microfibril deposition requires fibronectin-induced focal adhesions, and cell-cell junctions in epithelial cultures. Here we reveal that ADAMTS6 causes a reduction in heparan sulphate-rich interfaces, and its expression is regulated by ADAMTS10. PMID:27779234

  2. Laser-based microfabrication for cell adhesion and migration

    NASA Astrophysics Data System (ADS)

    Miller, Jordan S.

    Mammalian cell adhesion and migration impact a multitude of cellular behaviors and tissue remodeling processes. Over the past several decades, investigators have methodically improved in vitro systems as mimics of the extracellular microenvironment to study these biologic phenomena. Experiments have progressed from early studies on bifunctional inorganic surfaces to those with purified adhesive proteins against an organic, non-adhesive background. Recently, subcellular geometric patterns of adhesive proteins have proven useful to restrict and direct focal contact formation, cell survival, lamellopodia extension, and the maturation of "supermature" focal contacts. The vast majority of recent studies have involved the construction of hydrophobic patches with adsorbed fibronectin as the adhesive constraint of choice. However, the extracellular matrix (ECM) in which cells operate is a complex and diverse environment where numerous signals interact with a cell simultaneously; signals that the cell must integrate and that directly impact these processes. Microfabrication methods to approximate the extracellular milieu have significant limitations in their potential to be extended to pattern multiple bioactive ligands with high precision. Current techniques require multi-step processes which lose feature fidelity at every pattern transfer step, while simultaneously increasing logistical complexity and the chance of technical missteps. We have developed a family of complementary techniques using the raster-scanning laser of a confocal microscope to address a number of current challenges in improving microfabrication. For our work with thin films of self-assembled organic monolayers, we systematically removed the multi-step processing requirements of conventional photolithographic microfabrication and characterized and verified the technical advantages of our new patterning techniques. For 3D work, we developed and demonstrated micron-scale biochemical and mechanical

  3. CRISPR-Cas9 Mediated NOX4 Knockout Inhibits Cell Proliferation and Invasion in HeLa Cells

    PubMed Central

    Park, Rackhyun; Li, Liqing; Jang, Minsu; Morris, Andrew J.; Huang, Cai

    2017-01-01

    Increased expression of NOX4 protein is associated with cancer progression and metastasis but the role of NOX4 in cell proliferation and invasion is not fully understood. We generated NOX4 knockout HeLa cell lines using the CRISPR-Cas9 gene editing system to explore the cellular functions of NOX4. After transfection of CRISPR-Cas9 construct, we performed T7 endonuclease 1 assays and DNA sequencing to generate and identify insertion and deletion of the NOX4 locus. We confirmed the knockout of NOX4 by Western blotting. NOX4 knockout cell lines showed reduced cell proliferation with an increase of sub-G1 cell population and the decrease of S/G2/M population. Moreover, NOX4 deficiency resulted in a dramatic decrease in invadopodium formation and the invasive activity. In addition, NOX4 deficiency also caused a decrease in focal adhesions and cell migration in HeLa cells. These results suggest that NOX4 is required for both efficient proliferation and invasion of HeLa cells. PMID:28099519

  4. Adhesion receptors as therapeutic targets for circulating tumor cells

    PubMed Central

    Li, Jiahe; King, Michael R.

    2012-01-01

    Metastasis contributes to >90% of cancer-associated mortality. Though primary tumors can be removed by surgical resection or chemo/radiotherapy, metastatic disease is a great challenge to treatment due to its systemic nature. As metastatic “seeds,” circulating tumor cells (CTCs) are believed to be responsible for dissemination from a primary tumor to anatomically distant organs. Despite the possibility of physical trapping of CTCs in microvessels, recent advances have provided insights into the involvement of a variety of adhesion molecules on CTCs. Such adhesion molecules facilitate direct interaction with the endothelium in specific tissues or indirectly through leukocytes. Importantly, significant progress has been made in understanding how these receptors confer enhanced invasion and survival advantage during hematogenous circulation of CTCs through recruitment of macrophages, neutrophils, platelets, and other cells. This review highlights the identification of novel adhesion molecules and how blocking their function can compromise successful seeding and colonization of CTCs in new microenvironment. Encouraged by existing diagnostic tools to identify and isolate CTCs, strategic targeting of these adhesion molecules to deliver conventional chemotherapeutics or novel apoptotic signals is discussed for the neutralization of CTCs in the circulation. PMID:22837985

  5. Modeling keratinocyte wound healing dynamics: Cell-cell adhesion promotes sustained collective migration.

    PubMed

    Nardini, John T; Chapnick, Douglas A; Liu, Xuedong; Bortz, David M

    2016-07-07

    The in vitro migration of keratinocyte cell sheets displays behavioral and biochemical similarities to the in vivo wound healing response of keratinocytes in animal model systems. In both cases, ligand-dependent Epidermal Growth Factor Receptor (EGFR) activation is sufficient to elicit collective cell migration into the wound. Previous mathematical modeling studies of in vitro wound healing assays assume that physical connections between cells have a hindering effect on cell migration, but biological literature suggests a more complicated story. By combining mathematical modeling and experimental observations of collectively migrating sheets of keratinocytes, we investigate the role of cell-cell adhesion during in vitro keratinocyte wound healing assays. We develop and compare two nonlinear diffusion models of the wound healing process in which cell-cell adhesion either hinders or promotes migration. Both models can accurately fit the leading edge propagation of cell sheets during wound healing when using a time-dependent rate of cell-cell adhesion strength. The model that assumes a positive role of cell-cell adhesion on migration, however, is robust to changes in the leading edge definition and yields a qualitatively accurate density profile. Using RNAi for the critical adherens junction protein, α-catenin, we demonstrate that cell sheets with wild type cell-cell adhesion expression maintain migration into the wound longer than cell sheets with decreased cell-cell adhesion expression, which fails to exhibit collective migration. Our modeling and experimental data thus suggest that cell-cell adhesion promotes sustained migration as cells pull neighboring cells into the wound during wound healing.

  6. Cell proliferation within small intestinal crypts is the principal driving force for cell migration on villi.

    PubMed

    Parker, Aimee; Maclaren, Oliver J; Fletcher, Alexander G; Muraro, Daniele; Kreuzaler, Peter A; Byrne, Helen M; Maini, Philip K; Watson, Alastair J M; Pin, Carmen

    2017-02-01

    The functional integrity of the intestinal epithelial barrier relies on tight coordination of cell proliferation and migration, with failure to regulate these processes resulting in disease. It is not known whether cell proliferation is sufficient to drive epithelial cell migration during homoeostatic turnover of the epithelium. Nor is it known precisely how villus cell migration is affected when proliferation is perturbed. Some reports suggest that proliferation and migration may not be related while other studies support a direct relationship. We used established cell-tracking methods based on thymine analog cell labeling and developed tailored mathematical models to quantify cell proliferation and migration under normal conditions and when proliferation is reduced and when it is temporarily halted. We found that epithelial cell migration velocities along the villi are coupled to cell proliferation rates within the crypts in all conditions. Furthermore, halting and resuming proliferation results in the synchronized response of cell migration on the villi. We conclude that cell proliferation within the crypt is the primary force that drives cell migration along the villus. This methodology can be applied to interrogate intestinal epithelial dynamics and characterize situations in which processes involved in cell turnover become uncoupled, including pharmacological treatments and disease models.-Parker, A., Maclaren, O. J., Fletcher, A. G., Muraro, D., Kreuzaler, P. A., Byrne, H. M., Maini, P. K., Watson, A. J. M., Pin, C. Cell proliferation within small intestinal crypts is the principal driving force for cell migration on villi.

  7. Cell proliferation within small intestinal crypts is the principal driving force for cell migration on villi

    PubMed Central

    Parker, Aimee; Maclaren, Oliver J.; Fletcher, Alexander G.; Muraro, Daniele; Kreuzaler, Peter A.; Byrne, Helen M.; Maini, Philip K.; Watson, Alastair J. M.; Pin, Carmen

    2017-01-01

    The functional integrity of the intestinal epithelial barrier relies on tight coordination of cell proliferation and migration, with failure to regulate these processes resulting in disease. It is not known whether cell proliferation is sufficient to drive epithelial cell migration during homoeostatic turnover of the epithelium. Nor is it known precisely how villus cell migration is affected when proliferation is perturbed. Some reports suggest that proliferation and migration may not be related while other studies support a direct relationship. We used established cell-tracking methods based on thymine analog cell labeling and developed tailored mathematical models to quantify cell proliferation and migration under normal conditions and when proliferation is reduced and when it is temporarily halted. We found that epithelial cell migration velocities along the villi are coupled to cell proliferation rates within the crypts in all conditions. Furthermore, halting and resuming proliferation results in the synchronized response of cell migration on the villi. We conclude that cell proliferation within the crypt is the primary force that drives cell migration along the villus. This methodology can be applied to interrogate intestinal epithelial dynamics and characterize situations in which processes involved in cell turnover become uncoupled, including pharmacological treatments and disease models.—Parker, A., Maclaren, O. J., Fletcher, A. G., Muraro, D., Kreuzaler, P. A., Byrne, H. M., Maini, P. K., Watson, A. J. M., Pin, C. Cell proliferation within small intestinal crypts is the principal driving force for cell migration on villi. PMID:27811059

  8. Cell cycles and proliferation patterns in Haematococcus pluvialis

    NASA Astrophysics Data System (ADS)

    Zhang, Chunhui; Liu, Jianguo; Zhang, Litao

    2016-09-01

    Most studies on Haematococcus pluvialis have been focused on cell growth and astaxanthin accumulation; far less attention has been paid to cell cycles and proliferation patterns. The purpose of this study was to clarify cell cycles and proliferation patterns in H. pluvialis microscopically using a camera and video recorder system. The complicated life history of H. pluvialis can be divided into two stages: the motile stage and the non-motile stage. All the cells can be classified into forms as follows: motile cell, non-motile cell, zoospore and aplanospore. The main cell proliferation, both in the motile phase and non-motile phase in H. pluvialis, is by asexual reproduction. Under normal growth conditions, a motile cell usually produces two, sometimes four, and exceptionally eight zoospores. Under unfavorable conditions, the motile cell loses its flagella and transforms into a non-motile cell, and the non-motile cell usually produces 2, 4 or 8 aplanospores, and occasionally 20-32 aplanospores, which further develop into non-motile cells. Under suitable conditions, the non-motile cell is also able to release zoospores. The larger non-motile cells produce more than 16 zoospores, and the smaller ones produce 4 or 8 zoospores. Vegetative reproduction is by direct cell division in the motile phase and by occasional cell budding in the non-motile phase. There is, as yet, no convincing direct evidence for sexual reproduction.

  9. The influence of nanostructured features on bacterial adhesion and bone cell functions on severely shot peened 316L stainless steel.

    PubMed

    Bagherifard, Sara; Hickey, Daniel J; de Luca, Alba C; Malheiro, Vera N; Markaki, Athina E; Guagliano, Mario; Webster, Thomas J

    2015-12-01

    Substrate grain structure and topography play major roles in mediating cell and bacteria activities. Severe plastic deformation techniques, known as efficient metal-forming and grain refining processes, provide the treated material with novel mechanical properties and can be adopted to modify nanoscale surface characteristics, possibly affecting interactions with the biological environment. This in vitro study evaluates the capability of severe shot peening, based on severe plastic deformation, to modulate the interactions of nanocrystallized metallic biomaterials with cells and bacteria. The treated 316L stainless steel surfaces were first investigated in terms of surface topography, grain size, hardness, wettability and residual stresses. The effects of the induced surface modifications were then separately studied in terms of cell morphology, adhesion and proliferation of primary human osteoblasts (bone forming cells) as well as the adhesion of multiple bacteria strains, specifically Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, and ampicillin-resistant Escherichia coli. The results indicated a significant enhancement in surface work hardening and compressive residual stresses, maintenance of osteoblast adhesion and proliferation as well as a remarkable decrease in the adhesion and growth of gram-positive bacteria (S. aureus and S. epidermidis) compared to non-treated and conventionally shot peened samples. Impressively, the decrease in bacteria adhesion and growth was achieved without the use of antibiotics, for which bacteria can develop a resistance towards anyway. By slightly grinding the surface of severe shot peened samples to remove differences in nanoscale surface roughness, the effects of varying substrate grain size were separated from those of varying surface roughness. The expression of vinculin focal adhesions from osteoblasts was found to be singularly and inversely related to grain size, whereas the attachment of gram

  10. Ethylene Inhibits Cell Proliferation of the Arabidopsis Root Meristem.

    PubMed

    Street, Ian H; Aman, Sitwat; Zubo, Yan; Ramzan, Aleena; Wang, Xiaomin; Shakeel, Samina N; Kieber, Joseph J; Schaller, G Eric

    2015-09-01

    The root system of plants plays a critical role in plant growth and survival, with root growth being dependent on both cell proliferation and cell elongation. Multiple phytohormones interact to control root growth, including ethylene, which is primarily known for its role in controlling root cell elongation. We find that ethylene also negatively regulates cell proliferation at the root meristem of Arabidopsis (Arabidopsis thaliana). Genetic analysis indicates that the inhibition of cell proliferation involves two pathways operating downstream of the ethylene receptors. The major pathway is the canonical ethylene signal transduction pathway that incorporates CONSTITUTIVE TRIPLE RESPONSE1, ETHYLENE INSENSITIVE2, and the ETHYLENE INSENSITIVE3 family of transcription factors. The secondary pathway is a phosphorelay based on genetic analysis of receptor histidine kinase activity and mutants involving the type B response regulators. Analysis of ethylene-dependent gene expression and genetic analysis supports SHORT HYPOCOTYL2, a repressor of auxin signaling, as one mediator of the ethylene response and furthermore, indicates that SHORT HYPOCOTYL2 is a point of convergence for both ethylene and cytokinin in negatively regulating cell proliferation. Additional analysis indicates that ethylene signaling contributes but is not required for cytokinin to inhibit activity of the root meristem. These results identify key elements, along with points of cross talk with cytokinin and auxin, by which ethylene negatively regulates cell proliferation at the root apical meristem.

  11. Promotion of Cell Growth and Adhesion of a Peptide Hydrogel Scaffold via mTOR/Cadherin Signaling.

    PubMed

    Wei, Guojun; Wang, Liping; Dong, Daming; Teng, Zhaowei; Shi, Zuowei; Wang, Kaifu; An, Gang; Guan, Ying; Han, Bo; Yao, Meng; Xian, Cory J

    2017-02-18

    Understanding neurite outgrowth, orientation, and migration is important for the design of biomaterials that interface with the neural tissue. However, the molecular signaling alternations have not been well elucidated to explain the impact of hydrogels on cell morphology. In our previous studies, a silk fibroin peptide (SF16) hydrogel was found to be an effective matrix for the viability, morphology and proliferation of PC12 rat pheocrhomocytoma cells. We found that PC12 cells in the peptide hydrogel exhibited adhesive morphology compared to those cultured in agarose or collagen. Moreover, we identified that cell adhesion molecules (E- and N-cadherin) controlled by mTOR signaling were highly induced in PC12 cells cultured in the SF16 peptide hydrogel. Our findings suggest that the SF16 peptide might be suitable to be a cell-adhesion material in cell culture or tissue engineering, and mTOR/cadherin signaling is required for the cell adhesion in the SF16-peptide hydrogel. This article is protected by copyright. All rights reserved.

  12. Physiology and pathophysiology of selectins, integrins, and IgSF cell adhesion molecules focusing on inflammation. A paradigm model on infectious endocarditis.

    PubMed

    Golias, Christos; Batistatou, Anna; Bablekos, Georgios; Charalabopoulos, Alexandros; Peschos, Dimitrios; Mitsopoulos, Panagiotis; Charalabopoulos, Konstantinos

    2011-06-01

    The development of adhesion bonds, either among cells or among cells and components of the extracellular matrix, is a crucial process. These interactions are mediated by some molecules collectively known as adhesion molecules (CAMs). CAMs are ubiquitously expressed proteins playing a central role in controlling cell migration, proliferation, survival, and apoptosis. Besides their key function in physiological maintenance of tissue integrity, CAMs play an eminent role in various pathological processes such as cardiovascular disorders, atherogenesis, atherosclerotic plaque progression and regulation of the inflammatory response. CAMs such as selectins, integrins, and immunoglobulin superfamily take part in interactions between leukocyte and vascular endothelium (leukocyte rolling, arrest, firm adhesion, migration). Experimental data and pathologic observations support the assumption that pathogenic microorganisms attach to vascular endothelial cells or sites of vascular injury initiating intravascular infections. In this review a paradigm focusing on cell adhesion molecules pathophysiology and infective endocarditis development is given.

  13. Polymer films with surfaces unmodified and modified by non-thermal plasma as new substrates for cell adhesion.

    PubMed

    Borges, A M G; Benetoli, L O; Licínio, M A; Zoldan, V C; Santos-Silva, M C; Assreuy, J; Pasa, A A; Debacher, N A; Soldi, V

    2013-04-01

    The surface properties of biomaterials, such as wettability, polar group distribution, and topography, play important roles in the behavior of cell adhesion and proliferation. Gaseous plasma discharges are among the most common means to modify the surface of a polymer without affecting its properties. Herein, we describe the surface modification of poly(styrene) (PS) and poly(methyl methacrylate) (PMMA) films using atmospheric pressure plasma processing through exposure to a dielectric barrier discharge (DBD). After treatment the film surface showed significant changes from hydrophobic to hydrophilic as the water contact angle decreasing from 95° to 37°. All plasma-treated films developed more hydrophilic surfaces compared to untreated films, although the reasons for the change in the surface properties of PS and PMMA differed, that is, the PS showed chemical changes and in the case of PMMA they were topographical. Excellent adhesion and cell proliferation were observed in all films. In vitro studies employing flow cytometry showed that the proliferation of L929 cells was higher in the film formed by a 1:1 mixture of PS/PMMA, which is consistent with the results of a previous study. These findings suggest better adhesion of L929 onto the 1:1 PS/PMMA modified film, indicating that this system is a new candidate biomaterial for tissue engineering.

  14. N-Cadherin adhesive interactions modulate matrix mechanosensing and fate commitment of mesenchymal stem cells

    PubMed Central

    Cosgrove, Brian D.; Mui, Keeley L.; Driscoll, Tristan P.; Caliari, Steven R.; Mehta, Kush D.; Assoian, Richard K.; Burdick, Jason A.; Mauck, Robert L.

    2016-01-01

    During mesenchymal development, the microenvironment gradually transitions from one that is rich in cell-cell interactions to one that is dominated by cell-extracellular-matrix (ECM) interactions. Because these cues cannot readily be decoupled in vitro or in vivo, how they converge to regulate mesenchymal stem cell (MSC) mechanosensing is not fully understood. Here, we show that a hyaluronic acid hydrogel system enables, across a physiological range of ECM stiffness, the independent co-presentation of the HAVDI adhesive motif from the EC1 domain of N-Cadherin and the RGD adhesive motif from fibronectin. Decoupled presentation of these cues revealed that HAVDI ligation (at constant RGD ligation) reduced the contractile state and thereby nuclear YAP/TAZ localization in MSCs, resulting in altered interpretation of ECM stiffness and subsequent changes in downstream cell proliferation and differentiation. Our findings reveal that, in an evolving developmental context, HAVDI/N-Cadherin interactions can alter stem cell perception of the stiffening extracellular microenvironment. PMID:27525568

  15. N-cadherin adhesive interactions modulate matrix mechanosensing and fate commitment of mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    Cosgrove, Brian D.; Mui, Keeley L.; Driscoll, Tristan P.; Caliari, Steven R.; Mehta, Kush D.; Assoian, Richard K.; Burdick, Jason A.; Mauck, Robert L.

    2016-12-01

    During mesenchymal development, the microenvironment gradually transitions from one that is rich in cell-cell interactions to one that is dominated by cell-ECM (extracellular matrix) interactions. Because these cues cannot readily be decoupled in vitro or in vivo, how they converge to regulate mesenchymal stem cell (MSC) mechanosensing is not fully understood. Here, we show that a hyaluronic acid hydrogel system enables, across a physiological range of ECM stiffness, the independent co-presentation of the HAVDI adhesive motif from the EC1 domain of N-cadherin and the RGD adhesive motif from fibronectin. Decoupled presentation of these cues revealed that HAVDI ligation (at constant RGD ligation) reduced the contractile state and thereby nuclear YAP/TAZ localization in MSCs, resulting in altered interpretation of ECM stiffness and subsequent changes in downstream cell proliferation and differentiation. Our findings reveal that, in an evolving developmental context, HAVDI/N-cadherin interactions can alter stem cell perception of the stiffening extracellular microenvironment.

  16. Cell adhesive and antifouling polyvinyl chloride surfaces via wet chemical modification.

    PubMed

    Gabriel, Matthias; Strand, Dennis; Vahl, Christian-Friedrich

    2012-09-01

    Polyvinyl chloride (PVC) is one of the most frequently used polymers for the manufacturing of medical devices. Limitations for its usage are based upon unfavorable surface properties of the polymer including its hydrophobicity and lack of functionalities in order to increase its versatility. To address this issue, wet chemical modification of PVC was performed through surface amination using the bifunctional compound ethylene diamine. The reaction was conducted in order to achieve maximum surface amination while leaving the bulk material unaffected. The initial activation step was characterized by means of various methods including contact angle measurements, colorimetric amine quantification, infrared spectroscopy, and gel permeation chromatography. Depth profiles were obtained by a confocal microscopic method using fluorescence labeling. Exclusive surface modification was thus confirmed. To demonstrate biological applications of the presented technique, two examples were chosen: The covalent immobilization of the cell adhesive Asp-Gly-Asp-Ser-peptide (RGD) onto PVC samples yielded a surface that strongly supported cellular adhesion and proliferation of fibroblasts. In contrast, the decoration of PVC with the hydrophilic polymer polyethylene glycol prevented cellular adhesion to a large extent. The impact of these modifications was demonstrated by cell culture experiments.

  17. Sickle cell disease biochip: a functional red blood cell adhesion assay for monitoring sickle cell disease

    PubMed Central

    ALAPAN, YUNUS; KIM, CEONNE; ADHIKARI, ANIMA; GRAY, KAYLA E.; GURKAN-CAVUSOGLU, EVREN; LITTLE, JANE A.; GURKAN, UMUT A.

    2016-01-01

    Sickle cell disease (SCD) afflicts millions of people worldwide and is associated with considerable morbidity and mortality. Chronic and acute vaso-occlusion are the clinical hallmarks of SCD and can result in pain crisis, widespread organ damage, and early movtality. Even though the molecular underpinnings of SCD were identified more than 60 years ago, there are no molecular or biophysical markers of disease severity that are feasibly measured in the clinic. Abnormal cellular adhesion to vascular endothelium is at the root of vaso-occlusion. However, cellular adhesion is not currently evaluated clinically. Here, we present a clinically applicable microfluidic device (SCD biochip) that allows serial quantitative evaluation of red blood cell (RBC) adhesion to endothelium-associated protein-immobilized microchannels, in a closed and preprocessing-free system. With the SCD biochip, we have analyzed blood samples from more than 100 subjects and have shown associations between the measured RBC adhesion to endothelium-associated proteins (fibronectin and laminin) and individual RBC characteristics, including hemoglobin content, fetal hemoglobin concentration, plasma lactate dehydrogenase level, and reticulocyte count. The SCD biochip is a functional adhesion assay, reflecting quantitative evaluation of RBC adhesion, which could be used at baseline, during crises, relative to various long-term complications, and before and after therapeutic interventions. PMID:27063958

  18. OSTEOBLAST ADHESION OF BREAST CANCER CELLS WITH SCANNING ACOUSTIC MICROSCOPY

    SciTech Connect

    Chiaki Miyasaka; Robyn R. Mercer; Andrea M. Mastro; Ken L. Telschow

    2005-03-01

    Breast cancer frequently metastasizes to the bone. Upon colonizing bone tissue, the cancer cells stimulate osteoclasts (cells that break bone down), resulting in large lesions in the bone. The breast cancer cells also affect osteoblasts (cells that build new bone). Conditioned medium was collected from a bone-metastatic breast cancer cell line, MDA-MB-231, and cultured with an immature osteoblast cell line, MC3T3-E1. Under these conditions the osteoblasts acquired a changed morphology and appeared to adherer in a different way to the substrate and to each other. To characterize cell adhesion, MC3T3-E1 osteoblasts were cultured with or without MDA-MB-231 conditioned medium for two days, and then assayed with a mechanical scanning acoustic reflection microscope (SAM). The SAM indicated that in normal medium the MC3T3-E1 osteoblasts were firmly attached to their plastic substrate. However, MC3T3-E1 cells cultured with MDA-MB-231 conditioned medium displayed both an abnormal shape and poor adhesion at the substrate interface. The cells were fixed and stained to visualize cytoskeletal components using optical microscopic techniques. We were not able to observe these differences until the cells were quite confluent after 7 days of culture. However, using the SAM, we were able to detect these changes within 2 days of culture with MDA-MB-231 conditioned medium

  19. Inflammation and Proliferation Act Together to Mediate Intestinal Cell Fusion

    PubMed Central

    Swain, John R.; Wong, Melissa H.

    2009-01-01

    Cell fusion between circulating bone marrow-derived cells (BMDCs) and non-hematopoietic cells is well documented in various tissues and has recently been suggested to occur in response to injury. Here we illustrate that inflammation within the intestine enhanced the level of BMDC fusion with intestinal progenitors. To identify important microenvironmental factors mediating intestinal epithelial cell fusion, we performed bone marrow transplantation into mouse models of inflammation and stimulated epithelial proliferation. Interestingly, in a non-injury model or in instances where inflammation was suppressed, an appreciable baseline level of fusion persisted. This suggests that additional mediators of cell fusion exist. A rigorous temporal analysis of early post-transplantation cellular dynamics revealed that GFP-expressing donor cells first trafficked to the intestine coincident with a striking increase in epithelial proliferation, advocating for a required fusogenic state of the host partner. Directly supporting this hypothesis, induction of augmented epithelial proliferation resulted in a significant increase in intestinal cell fusion. Here we report that intestinal inflammation and epithelial proliferation act together to promote cell fusion. While the physiologic impact of cell fusion is not yet known, the increased incidence in an inflammatory and proliferative microenvironment suggests a potential role for cell fusion in mediating the progression of intestinal inflammatory diseases and cancer. PMID:19657387

  20. Cholesterol induces proliferation of chicken primordial germ cells.

    PubMed

    Chen, Dongyang; Chen, Meijuan; Lu, Zhenping; Yang, Mengmeng; Xie, Long; Zhang, Wenxin; Xu, Huiyan; Lu, Kehuan; Lu, Yangqing

    2016-08-01

    Primordial germ cells (PGCs) are the precursors of sperm and eggs and may serve as suitable cells for use in research in developmental biology and transgenic animals. However, the long-term propagation of PGCs in vitro has so far been plagued by the loss of their germ cell characteristics. This is largely because of the scarcity of knowledge concerning cell division and proliferation in these cells and the poor optimization of the culture medium. The sonic hedgehog (SHH) signaling pathway is involved in proliferation of many types of cells, but little is known about its role in chicken PGCs. The results of the current study indicate that the proliferation of chicken PGCs increases significantly when cholesterol, a molecule that facilitates the trafficking of HH ligands, is supplemented in the culture medium. This effect was attenuated when an SHH antagonist, cyclopamine was added, suggesting the involvement of SHH signaling in this process. The characterization of PGCs treated with cholesterol has shown that these cells express germ-cell-related markers and retain their capability to colonize the embryonic gonad after re-introduction to vasculature of stage-15 HH embryos, indicating that proliferation of PGCs induced by cholesterol does not alter the germ cell characteristics of these cells.

  1. Lysophosphatidic acid possesses dual action in cell proliferation.

    PubMed Central

    Tigyi, G; Dyer, D L; Miledi, R

    1994-01-01

    Lysophosphatidic acid (LPA) induces mitogenic responses in cultured fibroblasts through a pertussis toxin-sensitive signaling pathway. In contrast, we have shown that LPA inhibits the proliferation of Sp2/0-Ag14 myeloma cells. To resolve this apparent controversy, LPA-elicited responses in cell proliferation and the underlying second messenger mechanisms were compared in Sp2/0-Ag14 myeloma and NIH 3T3 fibroblast cells. The antimitogenic response was not elicited by micromolar concentrations of phosphatidic acid, phosphatidylglycerol, or diacylglycerol. In NIH 3T3 and Sp2 cells, LPA elicited an increase in inositol trisphosphate and a subsequent transient increase in free cytoplasmic Ca2+. Unlike the mitogenic response in NIH 3T3 cells, the antimitogenic effect was not affected by pertussis toxin; on the contrary, it was accompanied by an increase in cAMP. In Sp2 cells, cAMP analogs, forskolin, and isobutylmethylxanthine inhibited cell proliferation and enhanced LPA action in an additive manner, suggesting that an LPA-elicited increase in cAMP-mediated signaling was responsible for the antimitogenic response. In addition to the mitogenic response in fibroblasts and the antimitogenic response in tumor cell lines, there are some cell types (Jurkat T-cell lymphoma and primary astrocytes) in which LPA is ineffective in altering cell proliferation. The cell-type-specific dual action of LPA suggests that this endogenous lipid mediator when released from activated cells might play an important role as a regulator, rather than a ubiquitous inducer, of cell proliferation. Images PMID:8127904

  2. Temporal gene expression profile of human precursor B leukemia cells induced by adhesion receptor: identification of pathways regulating B-cell survival.

    PubMed

    Astier, Anne Laurence; Xu, Ronghui; Svoboda, Marek; Hinds, Esther; Munoz, Olivier; de Beaumont, Rosalie; Crean, Colin Daniel; Gabig, Theodore; Freedman, Arnold Stephen

    2003-02-01

    The physical interactions between B cells and stromal cells from the lymphoid tissue microenvironment are critical to the survival of normal and malignant B cells. They are principally mediated by integrins expressed on B cells and counterreceptors on stromal cells. Specifically, alpha4beta1 integrin engagement rescues B cells from physiological or drug-induced apoptosis. Therefore, in order to understand the mechanisms by which integrins prevent apoptosis in leukemia B cells, we compared the temporal gene expression profiles induced by beta1-integrin ligation with fibronectin (Fn) or adhesion by poly-L-Lysine in serum-starved precursor B leukemia cells. Among the 38 selected differentially expressed genes, 6 genes involved in adhesion (VAV2, EPB41L1, CORO1A), proliferation (FRAP1, CCT4), and intercellular communication (GJB3) were validated by real-time quantitative polymerase chain reaction (RT-Q-PCR). Gene expression modulation could also be validated at the protein level for 5 other genes. We show that integrin stimulation up-regulated FBI-1 expression but inhibited CD79a, Requiem, c-Fos, and caspase 7 induction when the cells underwent apoptosis. We further demonstrate that Fn stimulation also inhibits caspase 3 activation but increases XIAP and survivin expression. Moreover, integrin stimulation also prevents caspase activation induced by doxorubicin. Therefore, we identified genes modulated by adhesion of human precursor B leukemia cells that regulate proliferation and apoptosis, highlighting new pathways that might provide insights into future therapy aiming at targeting apoptosis of leukemia cells.

  3. Specific β-containing Integrins Exert Differential Control on Proliferation and Two-dimensional Collective Cell Migration in Mammary Epithelial Cells*

    PubMed Central

    Jeanes, Alexa I.; Wang, Pengbo; Moreno-Layseca, Paulina; Paul, Nikki; Cheung, Julia; Tsang, Ricky; Akhtar, Nasreen; Foster, Fiona M.; Brennan, Keith; Streuli, Charles H.

    2012-01-01

    Understanding how cell cycle is regulated in normal mammary epithelia is essential for deciphering defects of breast cancer and therefore for developing new therapies. Signals provided by both the extracellular matrix and growth factors are essential for epithelial cell proliferation. However, the mechanisms by which adhesion controls cell cycle in normal epithelia are poorly established. In this study, we describe the consequences of removing the β1-integrin gene from primary cultures of mammary epithelial cells in situ, using CreER. Upon β1-integrin gene deletion, the cells were unable to progress efficiently through S-phase, but were still able to undergo collective two-dimensional migration. These responses are explained by the presence of β3-integrin in β1-integrin-null cells, indicating that integrins containing different β-subunits exert differential control on mammary epithelial proliferation and migration. β1-Integrin deletion did not inhibit growth factor signaling to Erk or prevent the recruitment of core adhesome components to focal adhesions. Instead the S-phase arrest resulted from defective Rac activation and Erk translocation to the nucleus. Rac inhibition prevented Erk translocation and blocked proliferation. Activated Rac1 rescued the proliferation defect in β1-integrin-depleted cells, indicating that this GTPase is essential in propagating proliferative β1-integrin signals. These results show that β1-integrins promote cell cycle in mammary epithelial cells, whereas β3-integrins are involved in migration. PMID:22511753

  4. Knockdown of fucosyltransferase III disrupts the adhesion of circulating cancer cells to E-selectin without affecting hematopoietic cell adhesion.

    PubMed

    Yin, Xiaoyan; Rana, Kuldeepsinh; Ponmudi, Varun; King, Michael R

    2010-11-02

    Adhesive interactions between selectins and their ligands play an essential role during cancer extravasation. Fucosylation of these proteins by fucosyltransferases, or FUTs, is critical for their functions. Using quantitative RT-PCR, we demonstrated that FUT4 and FUT7 are the predominant FUTs expressed in hematopoietic cell line, while FUT3 is heavily expressed by multiple cancer cell lines including the prostate cancer cell line MDA PCa2b. Knockdown of FUT3 expression in MDA PCa2b cells by small interference RNA (siRNA) significantly reduced FUT3 expression. Cell-surface sialyl Lewis antigens were largely abolished. Cell adhesion and cell rolling on the blood vessel wall were simulated by perfusing cancer cells through microtubes coated with recombinant human E-selectin. At physiological levels of wall shear stress, the number of flowing cancer cells recruited to the microtube surface was dramatically reduced by FUT3 knockdown. Higher rolling velocity was also observed, which is consistent with reduced E-selectin binding activity. Interestingly, FUT3 siRNA treatment also significantly reduced the cell growth rate. Combined with the novel siRNA delivery platform recently developed in our laboratory, FUT3 siRNA could be a promising conjunctive therapy aiming at reducing the metastatic virulence of circulating epithelial cancer cells.

  5. Analysis of Proliferation of Melanoma Cells and Mesenchymal Stem Cells in Co-Culture and Contribution of Experimental Conditions into Interpretation of the Results.

    PubMed

    Kandarakov, O F; Kopantseva, E E; Belyavsky, A V

    2016-11-01

    A series of experiments on co-culturing of Mel IL melanoma cells and mesenhymal stem cells showed that these cells do not influence proliferation of each other, but we observed weaker adhesion of stromal stem cells to plastic in cocultures where with melanoma cells were grown on mesenhymal stem cells feeder. Cell proliferation was also considerably influenced by experimental conditions, which should be taken into account for correct interpretation of obtained results. The principles of experiments on co-culturing of cancer and stromal cells are formulated that take into account the most important factors influencing cell behavior and minimize the probability of artifact results. It was concluded that co-culturing conditions cells significantly affect the experimental results and can be the source of conflicting conclusions on mutual influence of stromal and cancer cells in vitro.

  6. Stretched cell cycle model for proliferating lymphocytes

    PubMed Central

    Dowling, Mark R.; Kan, Andrey; Heinzel, Susanne; Zhou, Jie H. S.; Marchingo, Julia M.; Wellard, Cameron J.; Markham, John F.; Hodgkin, Philip D.

    2014-01-01

    Stochastic variation in cell cycle time is a consistent feature of otherwise similar cells within a growing population. Classic studies concluded that the bulk of the variation occurs in the G1 phase, and many mathematical models assume a constant time for traversing the S/G2/M phases. By direct observation of transgenic fluorescent fusion proteins that report the onset of S phase, we establish that dividing B and T lymphocytes spend a near-fixed proportion of total division time in S/G2/M phases, and this proportion is correlated between sibling cells. This result is inconsistent with models that assume independent times for consecutive phases. Instead, we propose a stretching model for dividing lymphocytes where all parts of the cell cycle are proportional to total division time. Data fitting based on a stretched cell cycle model can significantly improve estimates of cell cycle parameters drawn from DNA labeling data used to monitor immune cell dynamics. PMID:24733943

  7. Activation of the canonical Wnt/{beta}-catenin pathway enhances monocyte adhesion to endothelial cells

    SciTech Connect

    Lee, Dong Kun . E-mail: leedk@memorialhealthsource.com; Nathan Grantham, R.; Trachte, Aaron L.; Mannion, John D.; Wilson, Colleen L.

    2006-08-18

    Monocyte adhesion to vascular endothelium has been reported to be one of the early processes in the development of atherosclerosis. In an attempt to develop strategies to prevent or delay atherosclerosis progression, we analyzed effects of the Wnt/{beta}-catenin signaling pathway on monocyte adhesion to various human endothelial cells. Adhesion of fluorescein-labeled monocytes to various human endothelial cells was analyzed under a fluorescent microscope. Unlike sodium chloride, lithium chloride enhanced monocyte adhesion to endothelial cells in a dose-dependent manner. We further demonstrated that inhibitors for glycogen synthase kinase (GSK)-3{beta} or proteosome enhanced monocyte-endothelial cell adhesion. Results of semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) indicated that activation of Wnt/{beta}-catenin pathway did not change expression levels of mRNA for adhesion molecules. In conclusion, the canonical Wnt/{beta}-catenin pathway enhanced monocyte-endothelial cell adhesion without changing expression levels of adhesion molecules.

  8. Mechanism of inhibition of cell proliferation by Vinca alkaloids.

    PubMed

    Jordan, M A; Thrower, D; Wilson, L

    1991-04-15

    We have used a structure-activity approach to investigate whether the Vinca alkaloids inhibit cell proliferation primarily by means of their effects on mitotic spindle microtubules or by another mechanism or by a combination of mechanisms. Five Vinca alkaloids were used to investigate the relationship in HeLa cells between inhibition of cell proliferation and blockage of mitosis, alteration of spindle organization, and depolymerization of microtubules. Indirect immunofluorescence staining of microtubules and 4,6-diamidino-2-phenylindole staining of chromatin were used to characterize the effects of the drugs on the distributions of cells in stages of the cell cycle and on the organization of microtubules and chromosomes in metaphase spindles. The microtubule polymer was isolated from cells and quantified using a competitive enzyme-linked immunoadsorbent assay for tubulin. We observed a nearly perfect coincidence between the concentration of each Vinca derivative that inhibited cell proliferation and the concentration that caused 50% accumulation of cells at metaphase, despite the fact that the antiproliferative potencies of the drugs varied over a broad concentration range. Inhibition of cell proliferation and blockage of cells at metaphase at the lowest effective concentrations of all Vinca derivatives occurred with little or no microtubule depolymerization or spindle disorganization. With increasing drug concentrations, the organization of microtubules and chromosomes in arrested mitotic spindles deteriorated in a manner that was common to all five congeners. These results indicate that the antiproliferative activity of the Vinca alkaloids at their lowest effective concentrations in HeLa cells is due to inhibition of mitotic spindle function. The results suggest further that the Vinca alkaloids inhibit cell proliferation by altering the dynamics of tubulin addition and loss at the ends of mitotic spindle microtubules rather than by depolymerizing the microtubules

  9. Hepatic stellate cells promote upregulation of epithelial cell adhesion molecule and epithelial-mesenchymal transition in hepatic cancer cells.

    PubMed

    Nagahara, Teruya; Shiraha, Hidenori; Sawahara, Hiroaki; Uchida, Daisuke; Takeuchi, Yasuto; Iwamuro, Masaya; Kataoka, Junro; Horiguchi, Shigeru; Kuwaki, Takeshi; Onishi, Hideki; Nakamura, Shinichiro; Takaki, Akinobu; Nouso, Kazuhiro; Yamamoto, Kazuhide

    2015-09-01

    Microenvironment plays an important role in epithelial-mesenchymal transition (EMT) and stemness of cells in hepatocellular carcinoma (HCC). Epithelial cell adhesion molecule (EpCAM) is known as a tumor stemness marker of HCC. To investigate the relationship between microenvironment and stemness, we performed an in vitro co-culture assay. Four HCC cell lines (HepG2, Hep3B, HuH-7 and PLC/PRF/5) were co-cultured with the TWNT-1 immortalized hepatic stellate cells (HSCs), which create a microenvironment with HCC. Cell proliferation ability was analyzed by flow cytometry (FCM) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, while migration ability was assessed by a wound healing assay. Expression of EpCAM was analyzed by immunoblotting and FCM. HCC cell lines were co-cultured with TWNT-1 treated with small interfering RNA (siRNA) for TGF-β and HB-EGF; we then analyzed proliferation, migration ability and protein expression using the methods described above. Proliferation ability was unchanged in HCC cell lines co-cultured with TWNT-1. Migration ability was increased in HCC cell lines (HepG2, Hep3B, HuH-7 and PLC/PRF/5) directly (216.2±67.0, 61.0±22.0, 124.0±66.2 and 51.5±40.3%) and indirectly (102.5±22.0, 84.6±30.9, 86.1±25.7 and 73.9±29.7%) co-cultured with TWNT-1 compared with the HCC uni-culture. Immunoblot analysis revealed increased EpCAM expression in the HCC cell lines co-cultured with TWNT-1. Flow cytometry revealed that the population of E-cadherin-/N-cadherin+ and EpCAM-positive cells increased and accordingly, EMT and stemness in the HCC cell line were activated. These results were similar in the directly and indirectly co-cultured samples, indicating that humoral factors were at play. Conversely, HCC cell lines co-cultured with siRNA‑treated TWNT-1 showed decreased migration ability, a decreased population of EpCAM-positive and E-cadherin-/N-cadherin+ cells. Taken together, humoral factors secreted from TWNT-1

  10. Evaluating fundamental position-dependent differences in wood cell wall adhesion using nanoindentation.

    PubMed

    Obersriebnig, Michael; Konnerth, Johannes; Gindl-Altmutter, Wolfgang

    2013-01-01

    Spruce wood specimens were bonded with one-component polyurethane (PUR) and urea-formaldehyde (UF) adhesive, respectively. The adhesion of the adhesives to the wood cell wall was evaluated at two different locations by means of a new micromechanical assay based on nanoindentation. One location tested corresponded to the interface between the adhesive and the natural inner cell wall surface of the secondary cell wall layer 3 (S3), whereas the second location corresponded to the interface between the adhesive and the freshly cut secondary cell wall layer 2 (S2). Overall, a trend towards reduced cell wall adhesion was found for PUR compared to UF. Position-resolved examination revealed excellent adhesion of UF to freshly cut cell walls (S2) but significantly diminished adhesion to the inner cell wall surface (S3). In contrast, PUR showed better adhesion to the inner cell wall surface and less adhesion to freshly cut cell walls. Atomic force microscopy revealed a less polar character for the inner cell wall surface (S3) compared to freshly cut cell walls (S2). It is proposed that differences in the polarity of the used adhesives and the surface chemistry of the two cell wall surfaces examined account for the observed trends.

  11. Lateral adhesion drives reintegration of misplaced cells into epithelial monolayers

    PubMed Central

    St Johnston, Daniel

    2016-01-01

    Cells in simple epithelia orient their mitotic spindles in the plane of the epithelium so that both daughter cells are born within the epithelial sheet. This is assumed to be important to maintain epithelial integrity and prevent hyperplasia, because misaligned divisions give rise to cells outside the epithelium1,2. Here we test this assumption in three types of Drosophila epithelia; the cuboidal follicle epithelium, the columnar early embryonic ectoderm, and the pseudostratified neuroepithelium. Ectopic expression of Inscuteable in these tissues reorients mitotic spindles, resulting in one daughter cell being born outside of the epithelial layer. Live imaging reveals that these misplaced cells reintegrate into the tissue. Reducing the levels of the lateral homophilic adhesion molecules Neuroglian or Fasciclin 2 disrupts reintegration, giving rise to extra-epithelial cells, whereas disruption of adherens junctions has no effect. Thus, the reinsertion of misplaced cells appears to be driven by lateral adhesion, which pulls cells born outside the epithelia layer back into it. Our findings reveal a robust mechanism that protects epithelia against the consequences of misoriented divisions. PMID:26414404

  12. Effect of testosterone incorporation on cell proliferation and differentiation for polymer-bioceramic composites.

    PubMed

    da Costa, Kelen Jorge Rodrigues; Passos, Joel J; Gomes, Alinne D M; Sinisterra, Rubén D; Lanza, Célia R M; Cortés, Maria Esperanza

    2012-11-01

    In the current study, we characterized the polycaprolactone (PCL), poly(lactic acid-co-glycolic acid) (PLGA), and biphasic calcium phosphate (BCP) composites coated with testosterone propionate (T) using Fourier transform infrared spectroscopy (FTIR) and powder X-ray diffraction (XRD). Osteoblastic cells were seeded with PCL/BCP, PCL/BCP/T, PLGA/PCL/BCP and PLGA/PCL/BCP/T scaffolds, and cell viability, proliferation, differentiation and adhesion were analyzed. The results of physic-chemical experiments showed no displacements or suppression of bands in the FTIR spectra of scaffolds. The XRD patterns of the scaffolds showed an amorphous profile. The osteoblastic cells viability and proliferation increased in the presence of composites with testosterone over 72 h, and were significantly greater when PLGA/PCL/BCP/T scaffold was tested against PCL/BCP/T. Furthermore alkaline phosphatase production was significantly greater in the same group. In conclusion, the PLGA/PCL/BCP scaffold with testosterone could be a promising option for bone tissue applications due to its biocompatibility and its stimulatory effect on cell proliferation.

  13. Effect of a hot water extract of Chlorella vulgaris on proliferation of IEC-6 cells.

    PubMed

    Song, Seo-Hyeon; Kim, In-Hye; Nam, Taek-Jeong

    2012-05-01

    Chlorella vulgaris, a unicellular microalgae, exerts various biological effects; however their effect on proliferation signaling pathways in normal cells has not been studied. We investigated the effect of hot water extracts of Chlorella vulgaris (CVE) on cell proliferation