Key Specifications Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, M, Po, R||IP, WB||Rb||Serum||Polyclonal Antibody|
|Description||Anti-Integrin α5 Antibody|
|Presentation||Serum. Liquid containing no preservatives.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||Maintain frozen at -20°C in undiluted aliquots for up to 12 months from date of receipt.|
|Material Size||100 µL|
|RABBIT ANTI-INTEGRIN ALPHA 5 -2689062||2689062|
|RABBIT ANTI-INTEGRIN ALPHA 5 POLYCLONAL ANTIBODY - 2041545||2041545|
|RABBIT ANTI-INTEGRIN ALPHA 5 POLYCLONAL ANTIBODY - 2191929||2191929|
References | 20 Available | See All References
|Reference overview||Application||Species||Pub Med ID|
|Mesoangioblast delivery of miniagrin ameliorates murine model of merosin-deficient congenital muscular dystrophy type 1A. |
Domi, T; Porrello, E; Velardo, D; Capotondo, A; Biffi, A; Tonlorenzi, R; Amadio, S; Ambrosi, A; Miyagoe-Suzuki, Y; Takeda, S; Ruegg, MA; Previtali, SC
Skeletal muscle 5 30 2015
Merosin-deficient congenital muscular dystrophy type-1A (MDC1A) is characterized by progressive muscular dystrophy and dysmyelinating neuropathy caused by mutations of the α2 chain of laminin-211, the predominant laminin isoform of muscles and nerves. MDC1A has no available treatment so far, although preclinical studies showed amelioration of the disease by the overexpression of miniagrin (MAG). MAG reconnects orphan laminin-211 receptors to other laminin isoforms available in the extracellular matrix of MDC1A mice.Mesoangioblasts (MABs) are vessel-associated progenitors that can form the skeletal muscle and have been shown to restore defective protein levels and motor skills in animal models of muscular dystrophies. As gene therapy in humans still presents challenging technical issues and limitations, we engineered MABs to overexpress MAG to treat MDC1A mouse models, thus combining cell to gene therapy.MABs synthesize and secrete only negligible amount of laminin-211 either in vitro or in vivo. MABs engineered to deliver MAG and injected in muscles of MDC1A mice showed amelioration of muscle histology, increased expression of laminin receptors in muscle, and attenuated deterioration of motor performances. MABs did not enter the peripheral nerves, thus did not affect the associated peripheral neuropathy.Our study demonstrates the potential efficacy of combining cell with gene therapy to treat MDC1A.
|SEL1L regulates adhesion, proliferation and secretion of insulin by affecting integrin signaling. |
Diaferia, GR; Cirulli, V; Biunno, I
PloS one 8 e79458 2013
SEL1L, a component of the endoplasmic reticulum associated degradation (ERAD) pathway, has been reported to regulate the (i) differentiation of the pancreatic endocrine and exocrine tissue during the second transition of mouse embryonic development, (ii) neural stem cell self-renewal and lineage commitment and (iii) cell cycle progression through regulation of genes related to cell-matrix interaction. Here we show that in the pancreas the expression of SEL1L is developmentally regulated, such that it is readily detected in developing islet cells and in nascent acinar clusters adjacent to basement membranes, and becomes progressively restricted to the islets of Langherans in post-natal life. This peculiar expression pattern and the presence of two inverse RGD motifs in the fibronectin type II domain of SEL1L protein indicate a possible interaction with cell adhesion molecules to regulate islets architecture. Co-immunoprecipitation studies revealed SEL1L and ß1-integrin interaction and, down-modulation of SEL1L in pancreatic ß-cells, negatively influences both cell adhesion on selected matrix components and cell proliferation likely due to altered ERK signaling. Furthermore, the absence of SEL1L protein strongly inhibits glucose-stimulated insulin secretion in isolated mouse pancreatic islets unveiling an important role of SEL1L in insulin trafficking. This phenotype can be rescued by the ectopic expression of the ß1-integrin subunit confirming the close interaction of these two proteins in regulating the cross-talk between extracellular matrix and insulin signalling to create a favourable micro-environment for ß-cell development and function.
|ETV5 transcription factor is overexpressed in ovarian cancer and regulates cell adhesion in ovarian cancer cells. |
Llauradó M, Abal M, Castellví J, Cabrera S, Gil-Moreno A, Pérez-Benavente A, Colás E, Doll A, Dolcet X, Matias-Guiu X, Vazquez-Levin M, Reventós J, Ruiz A
International journal of cancer Journal international du cancer 2011
Epithelial ovarian cancer is the most lethal gynecological malignancy and the fifth leading cause of cancer deaths in women in the Western world. ETS transcription factors are known to act as positive or negative regulators of the expression of genes that are involved in various biological processes, including those that control cellular proliferation, differentiation, apoptosis, tissue remodelling, angiogenesis and transformation. ETV5 belongs to the PEA3 subfamily. PEA3 subfamily members are able to activate the transcription of proteases, MMPs and TIMPs, which is central to both tumor invasion and angiogenesis. Here, we examined the role of the ETV5 transcription factor in epithelial ovarian cancer and we found ETV5 upregulated in ovarian tumor samples compared to ovarian tissue controls. The in vitro inhibition of ETV5 decreased cell proliferation in serum-deprived conditions, induced EMT and cell migration and decreased cell adhesion to extracellular matrix components. ETV5 inhibition also decreased cell-cell adhesion and induced apoptosis in anchorage independent conditions. Accordingly, ETV5 upregulation induced the expression of cell adhesion molecules and enhanced cell survival in a spheroid model. Our findings suggest that the overexpression of ETV5 detected in ovarian cancer cells may contribute to ovarian tumor progression through the ability of ETV5 to enhance ovarian cancer cell proliferation. In addition, ETV5 upregulation would play a role in ovarian cancer cell dissemination and metastasis into the peritoneal cavity by protecting ovarian cancer cells from apoptosis and by increasing the adhesion of ovarian cancer cells to the peritoneal wall through the regulation of cell adhesion molecules.Copyright © 2011 UICC.
|Alteration of pulmonary artery integrin levels in chronic hypoxia and monocrotaline-induced pulmonary hypertension. |
Umesh, A; Paudel, O; Cao, YN; Myers, AC; Sham, JS
Journal of vascular research 48 525-37 2011
Pulmonary hypertension is associated with vascular remodeling and increased extracellular matrix (ECM) deposition. While the contribution of ECM in vascular remodeling is well documented, the roles played by their receptors, integrins, in pulmonary hypertension have received little attention. Here we characterized the changes of integrin expression in endothelium-denuded pulmonary arteries (PAs) and aorta of chronic hypoxia as well as monocrotaline-treated rats.Immunoblot showed increased α(1)-, α(8)- and α(v)-integrins, and decreased α(5)-integrin levels in PAs of both models. β(1)- and β(3)-integrins were reduced in PAs of chronic hypoxia and monocrotaline-treated rats, respectively. Integrin expression in aorta was minimally affected. Differential expression of α(1)- and α(5)-integrins induced by chronic hypoxia was further examined. Immunostaining showed that they were expressed on the surface of PA smooth muscle cells (PASMCs), and their distribution was unaltered by chronic hypoxia. Phosphorylation of focal adhesion kinase was augmented in PAs of chronic hypoxia rats, and in chronic hypoxia PASMCs cultured on the α(1)-ligand collagen IV. Moreover, α(1)-integrin binding hexapeptide GRGDTP elicited an enhanced Ca(2+) response, whereas the response to α(5)-integrin binding peptide GRGDNP was reduced in CH-PASMCs.Integrins in PASMCs are differentially regulated in pulmonary hypertension, and the dynamic integrin-ECM interactions may contribute to the vascular remodeling accompanying disease progression.
|Coordinate regulation of estrogen-mediated fibronectin matrix assembly and epidermal growth factor receptor transactivation by the G protein-coupled receptor, GPR30. |
Quinn, JA; Graeber, CT; Frackelton, AR; Kim, M; Schwarzbauer, JE; Filardo, EJ
Molecular endocrinology (Baltimore, Md.) 23 1052-64 2009
Estrogen promotes changes in cytoskeletal architecture not easily attributed to the biological action of estrogen receptors, ERalpha and ERbeta. The Gs protein-coupled transmembrane receptor, GPR30, is linked to specific estrogen binding and rapid estrogen-mediated release of heparin-bound epidermal growth factor. Using marker rescue and dominant interfering mutant strategies, we show that estrogen action via GPR30 promotes fibronectin (FN) matrix assembly by human breast cancer cells. Stimulation with 17beta-estradiol or the ER antagonist, ICI 182, 780, results in the recruitment of FN-engaged integrin alpha5beta1 conformers to fibrillar adhesions and the synthesis of FN fibrils. Concurrent with this cellular response, GPR30 promotes the formation of Src-dependent, Shc-integrin alpha5beta1 complexes. Function-blocking antibodies directed against integrin alpha5beta1 or soluble Arg-Gly-Asp peptide fragments derived from FN specifically inhibited GPR30-mediated epidermal growth factor receptor transactivation. Estrogen-mediated FN matrix assembly and epidermal growth factor receptor transactivation were similarly disrupted in integrin beta1-deficient GE11 cells, whereas reintroduction of integrin beta1 into GE11 cells restored these responses. Mutant Shc (317Y/F) blocked GPR30-induced FN matrix assembly and tyrosyl phosphorylation of erbB1. Interestingly, relative to recombinant wild-type Shc, 317Y/F Shc was more readily retained in GPR30-induced integrin alpha5beta1 complexes, yet this mutant did not prevent endogenous Shc-integrin alpha5beta1 complex formation. Our results suggest that GPR30 coordinates estrogen-mediated FN matrix assembly and growth factor release in human breast cancer cells via a Shc-dependent signaling mechanism that activates integrin alpha5beta1.Full Text Article
|Alterations in integrin expression modulates invasion of pancreatic cancer cells. |
Walsh, N; Clynes, M; Crown, J; O'Donovan, N
Journal of experimental & clinical cancer research : CR 28 140 2009
Factors mediating the invasion of pancreatic cancer cells through the extracellular matrix (ECM) are not fully understood.In this study, sub-populations of the human pancreatic cancer cell line, MiaPaCa-2 were established which displayed differences in invasion, adhesion, anoikis, anchorage-independent growth and integrin expression.Clone #3 displayed higher invasion with less adhesion, while Clone #8 was less invasive with increased adhesion to ECM proteins compared to MiaPaCa-2. Clone #8 was more sensitive to anoikis than Clone #3 and MiaPaCa-2, and displayed low colony-forming efficiency in an anchorage-independent growth assay. Integrins beta 1, alpha 5 and alpha 6 were over-expressed in Clone #8. Using small interfering RNA (siRNA), integrin beta1 knockdown in Clone #8 cells increased invasion through matrigel and fibronectin, increased motility, decreased adhesion and anoikis. Integrin alpha 5 and alpha 6 knockdown also resulted in increased motility, invasion through matrigel and decreased adhesion.Our results suggest that altered expression of integrins interacting with different extracellular matrixes may play a significant role in suppressing the aggressive invasive phenotype. Analysis of these clonal populations of MiaPaCa-2 provides a model for investigations into the invasive properties of pancreatic carcinoma.Full Text Article
|The regulation of integrin-mediated osteoblast focal adhesion and focal adhesion kinase expression by nanoscale topography. |
Jung Yul Lim,Andrea D Dreiss,Zhiyi Zhou,Joshua C Hansen,Christopher A Siedlecki,Robert W Hengstebeck,Juan Cheng,Nicholas Winograd,Henry J Donahue
Biomaterials 28 2007
An important consideration in developing physical biomimetic cell-stimulating cues is that the in vivo extracellular milieu includes nanoscale topographic interfaces. We investigated nanoscale topography regulation of cell functions using human fetal osteoblastic (hFOB) cell culture on poly(l-lactic acid) and polystyrene (50/50 w/w) demixed nanoscale pit textures (14, 29, and 45nm deep pits). Secondary ion mass spectroscopy revealed that these nanotopographic surfaces had similar surface chemistries to that of pure PLLA because of PLLA component surface segregation during spin casting. We observed that 14 and 29nm deep pit surfaces increased hFOB cell attachment, spreading, selective integrin subunit expression (e.g., alphav relative to alpha5, beta1, or beta3), focal adhesive paxillin protein synthesis and paxillin colocalization with cytoskeletal actin stress fibers, and focal adhesion kinase (FAK) and phosphorylated FAK (pY397) expression to a greater degree than did 45nm deep pits or flat PLLA surfaces. Considering the important role of integrin-mediated focal adhesion and intracellular signaling in anchorage-dependent cell function, our results suggest a mechanism by which nanostructured physical signals regulate cell function. Modulation of integrin-mediated focal adhesion and related cell signaling by altering nanoscale substrate topography will have powerful applications in biomaterials science and tissue engineering.
|N-glycosylation of the beta-propeller domain of the integrin alpha5 subunit is essential for alpha5beta1 heterodimerization, expression on the cell surface, and its biological function. |
Isaji, T; Sato, Y; Zhao, Y; Miyoshi, E; Wada, Y; Taniguchi, N; Gu, J
The Journal of biological chemistry 281 33258-67 2006
The N-glycosylation of integrin alpha5beta1 is thought to play crucial roles in cell spreading, cell migration, ligand binding, and dimer formation, but the underlying mechanism remains unclear. To investigate the importance of the N-glycans of this integrin in detail, sequential site-directed mutagenesis was carried out to remove single or combined putative N-glycosylation sites on the alpha5 integrin. Removal of the putative N-glycosylation sites on the beta-propeller, Thigh, Calf-1, or Calf-2 domains of the alpha5 subunit resulted in a decrease in molecular weight compared with the wild type, suggesting that all of these domains contain attached N-glycans. Importantly, the absence of N-glycosylation sites (sites 1-5) on the beta-propeller resulted in the persistent association of integrin subunit with calnexin in the endoplasmic reticulum, which subsequently blocked heterodimerization and its expression on the cell surface. Interestingly, the activities for cell spreading and migration for the alpha5 subunit carrying only three potential N-glycosylation sites (3-5 sites) on the beta-propeller were comparable with those of the wild type. In contrast, mutation of these three sites resulted in a significant decrease in cell spreading as well as functional expression, although the total expression level of the Delta3-5 mutant on the cell surface was comparable with that of wild type. Furthermore, we found that site 5 is a most important site for its expression on the cell surface, whereas the S5 mutant did not show any biological functions. Taken together, this study reveals for the first time that the N-glycosylation on the beta-propeller domain of the alpha5 subunit is essential for heterodimerization and biological functions of alpha5beta1 integrin and might also be useful for studies of the molecular structure.
|Integrin expression and osteopontin regulation in human fetal osteoblastic cells mediated by substratum surface characteristics. |
Jung Yul Lim, Amanda F Taylor, Zhongyong Li, Erwin A Vogler, Henry J Donahue
Tissue engineering 11 19-29 2005
Integrin-mediated adhesion of anchorage-dependent cells to scaffolds is a critical component of tissue engineering. We investigated integrin expression by the human fetal osteoblastic cell line, hFOB 1.19 (hFOB), as a function of substratum surface wettability. The influence of surface wettability on bone cell phenotype was also examined. Plasma-treated quartz (PTQ) and glass (PTG) (hydrophilic, contact angles of 0 degrees), octadecyltrichlorosilane-treated quartz (STQ) and glass (STG) (hydrophobic, contact angles above about 100 degrees), and tissue culture polystyrene were used for cell culture. hFOB cells cultured on hydrophilic substrata displayed well-developed actin stress fibers relative to cells on hydrophobic substrata. Western blot analysis revealed that hFOB cells cultured on hydrophobic substrata (STQ or STG) express lower levels of alphav and beta3 integrin subunits than do cells on hydrophilic substrata (PTQ or PTG). This effect was more pronounced in cells on STQ than on STG. These variations in integrin expression were lessened by extended culture time. Double- labeled integrin/actin immunofluorescence confirmed Western blot results, that is, cells cultured on PTQ displayed distinct, large plaques of alphav and beta3 subunits and integrin alphavbeta3, as well as their colocalization with actin stress fiber ends, whereas cells on STQ did not display integrin plaques after 24 h and displayed only minimal plaque formation after 3 days. Vinculin, a focal adhesion protein that mediates binding between the integrin and actin cytoskeleton, appeared in Western blots to mimic the variations of alphav and beta3 expression with respect to surface wettability. Interestingly, real-time RT-PCR analysis showed that hFOB cultured on hydrophobic substrata, which have downregulated alphav and beta3 integrin subunits, displayed greater steady state mRNA levels of osteopontin, an extracellular matrix (ECM) protein containing the Arg-Gly-Asp (RGD) integrin recognition sequence, than did cells cultured on hydrophilic substrata. Our results imply that substratum surface wettability regulates integrin-mediated bone cell adhesion and further influences the expression of bone cell-ECM complexes.
|Visinin-like protein-1 is a potent inhibitor of cell adhesion and migration in squamous carcinoma cells. |
Gonzalez Guerrico, AM; Jaffer, ZM; Page, RE; Braunewell, KH; Chernoff, J; Klein-Szanto, AJ
Oncogene 24 2307-16 2005
Tumor cell invasion is a highly integrated and complex process comprising several biologically distinct functions such as cell adhesion, motility, proteolysis, etc. Visinin-like protein-1 (VILIP-1), a member of the neuronal EF-hand calcium-sensor protein family, plays a role in regulating tumor cell invasiveness of mouse squamous cell carcinoma (SCC). VILIP-1 enhances cyclic adenosine monophosphate levels through PKA induction. However, the mechanism by which VILIP-1 reduces cell invasiveness is not well understood. In this study, we show that VILIP-1 decreased cell adhesion and migration/invasiveness of highly invasive mouse SCC cells. Forced expression of VILIP-1 reduced cell adhesion to fibronectin in parallel to downregulating alphav and alpha5 integrin subunit levels. VILIP-1 overexpression also led to decreased migration ability. Conversely, short hairpin RNA-mediated VILIP-1 knock-down of SCC cells' characterized by little or no invasiveness, correlated with increased adhesion to fibronectin and enhanced expression of alphav and alpha5 integrin subunits together with increased cell migration. Function-blocking assays with inhibitory anti-alpha5 and anti-alphav integrin antibodies showed that both subunits contributed to cell adhesion, migration, and invasiveness of highly invasive SCC cell lines. These results point to a critical role of VILIP-1 in regulating cell adhesion and migration by downregulation of fibronectin receptor expression. Decreased or absent VILIP-1 expression in SCC cell subpopulations may lead to a more advanced malignant phenotype characterized by changes in adhesive ability and increased cell motility, suggestive of a tumor suppressor function.
|Squamous cell carcinoma cell aggregates escape suspension-induced, p53-mediated anoikis: fibronectin and integrin alphav mediate survival signals through focal adhesion kinase. |
Zhang, Y; Lu, H; Dazin, P; Kapila, Y
The Journal of biological chemistry 279 48342-9 2004
Resistance to anoikis, or apoptosis triggered by detachment from the extracellular matrix (ECM), lengthens the survival of malignant cells, facilitating reattachment and colonization of secondary sites. To examine the molecular mechanisms underlying resistance to anoikis in human oral squamous cell carcinoma (SCC) cells, we cultured human squamous carcinoma (HSC-3) cells in suspension on plates coated with poly-2-hydroxyethyl methacrylate, which blocks access to the ECM. Cells in suspension that formed multicellular aggregates had significantly lower levels of apoptosis than single cells. Aggregates, but not single cells, had high levels of fibronectin. Preincubation with a cyclic arginine-glycine-aspartic acid peptide or fibronectin-blocking antibody significantly increased anoikis. Single cells had markedly lower expression of the integrin alpha(v) receptor than aggregates. Blocking alpha(v) function with a blocking antibody or by transfection with an antisense oligonucleotide increased apoptosis and inhibited aggregation. In single cells but not aggregates, phosphorylation of the integrin-associated focal adhesion kinase (FAK) at tyrosine 397 was reduced, and p53 levels were increased. Apoptosis was increased by blocking FAK with an antisense oligonucleotide and reduced by blocking p53. These findings show that SCC cells escape suspension-induced anoikis by forming multicellular aggregates that avail themselves of fibronectin survival signals mediated by integrin alpha(v). Single cells in suspension that do not form aggregates undergo anoikis because of decreased FAK phosphorylation and increased p53 levels. Thus, SCC cells appear to use neighboring cells and the ECM molecule FN to promote the metastatic phenotype.
|Reversion of the Jun-induced oncogenic phenotype by enhanced synthesis of sialosyllactosylceramide (GM3 ganglioside). |
Miura, Y; Kainuma, M; Jiang, H; Velasco, H; Vogt, PK; Hakomori, S
Proceedings of the National Academy of Sciences of the United States of America 101 16204-9 2004
In the mouse fibroblast cell line C3H 10T1/2 and the chicken fibroblast cell line DF1, the ganglioside GM3 is the major glycosphingolipid component of the plasma membrane. Expression of the viral oncoprotein Jun (v-Jun) induces transformed cell clones with greatly reduced levels of GM3 and GM3 synthase (lactosylceramide alpha2,3-sialyltransferase) mRNA in both 10T1/2 and DF1 cell cultures. Compared with nontransformed controls, v-Jun transfectants show enhanced ability of anchorage-independent growth, and their growth rates as adherent cells are increased. When the mouse GM3 synthase gene is transfected with the pcDNA vector into v-Jun-transformed 10T1/2 cells, the levels of GM3 synthase and corresponding mRNA are restored to those of control cells. Reexpression of GM3 correlates with a reduced ability of the cells to form colonies in nutrient agar. Similarly, when the newly cloned chicken GM3 synthase gene is transfected into v-Jun-transformed DF1 with the pcDNA vector, the GM3 synthase level is restored to that of control cells, and the ability of the cells to form agar colonies is reduced. The levels of GM3 in the cell also affect membrane microdomains. The complex of GM3 with tetraspanin CD9 and integrin alpha5beta1 inhibits motility and invasiveness. The amounts of this complex are greatly reduced in transformed cells. Expression of GM3 and consequent reversion of the transformed phenotype results in increased levels of that microdomain complex.Full Text Article
|Alterations in cell-adhesive and migratory properties of proximal tubule and collecting duct cells from bcl-2 -/- mice. |
Ziehr, J; Sheibani, N; Sorenson, CM
American journal of physiology. Renal physiology 287 F1154-63 2004
Bcl-2 protects cells from apoptosis initiated by a variety of stimuli including loss of cell adhesion. Bcl-2 -/- mice develop renal hypoplastic/cystic dysplasia with renal cyst formation coinciding with renal maturation in normal mice. To gain a better understanding of the role cell-adhesive mechanisms play during renal maturation, we generated proximal tubule and collecting duct cell lines from postnatal day 10 (P10) and P20 bcl-2 +/+ and bcl-2 -/- mice. Very little is known about the role cell-adhesive and migratory mechanisms play during renal maturation. We observed that modulation of cell-adhesive properties, which normally occur in a nephron segment-specific manner during renal maturation, and cell migration were altered in cells from bcl-2 -/- mice. Enhanced migration of bcl-2 -/- proximal tubule cells in a scratch wound assay was completely inhibited by incubation with PP1 (Src inhibitor) and moderately affected by incubation with SB-203580 (p38 inhibitor). These cells expressed increased levels of fibronectin and had numerous central focal adhesions. P20 bcl-2 -/- proximal tubule cells adhered to fibronectin but adhered poorly to collagen, vitronectin, or laminin. Collecting duct cells, similar to proximal tubule cells from bcl-2 -/- mice, demonstrated enhanced migration in a scratch wound assay that was inhibited by incubation with PP1. Migration of these cells was moderately affected by incubation with PD-98059 (MEK inhibitor) or LY-294002 (PI3 kinase inhibitor), whereas incubation with SB-203580 had no effect. P10 bcl-2 -/- collecting duct cells also expressed increased levels of fibronectin but decreased levels of thrombospondin-1 and demonstrated precocious binding to fibronectin and vitronectin compared with bcl-2 +/+ cells. The ability of P20 bcl-2 +/+ collecting duct cells to adhere to fibronectin and vitronectin corresponded with a decline in thrombospondin-1 expression. Therefore, alterations in cell-adhesive and migratory characteristics may be an early indicator of aberrant renal epithelial cell differentiation.
|Fluorescence Activated Cell Sorting (FACS)||15292044|
|Effect of quiescence on integrin alpha5beta1 expression in human retinal pigment epithelium. |
Proulx, Stephanie, et al.
Mol. Vis., 9: 473-81 (2003) 2003
PURPOSE: The retinal pigment epithelium (RPE) is differentiated and mitotically inactive in the normal eye, but several pathologies such as proliferative vitreoretinopathy (PVR) cause RPE cells to dedifferentiate and resume proliferation. Integrins, a family of cell surface glycoproteins that mediate cell proliferation and differentiation, are thought to play fundamental roles in PVR. The aim of this study was to evaluate protein expression and gene regulation of the integrin alpha5 subunit in proliferating and quiescent RPE cells. METHODS: Protein expression was studied in situ by immunohistochemistry and in vitro at different cell confluences by immunoprecipitation. Semi-quantitative RT-PCR and transient transfections were used to determine whether increasing cell confluence also affected alpha5 subunit mRNA levels and promoter activity, respectively. RESULTS: We demonstrated that the integrin alpha5 subunit is present at the RPE cell surface both in situ and in vitro, and that alpha5 protein level is influenced by confluence. Levels of integrin alpha5 transcripts are similar for sub-confluent and confluent cells, and a small increase in the promoter activity was observed between sub-confluent and confluent cells. However, both the integrin alpha5 subunit transcript and the alpha5 promoter activity decreased when cells reached post-confluence. CONCLUSIONS: We demonstrated that cell confluence affected protein and gene expression of the integrin alpha5 subunit. Proliferating RPE cells expressed high levels of both the alpha5 protein and mRNA transcripts and showed a high promoter activity. However, when cells reached quiescence, alpha5 gene expression was substantially reduced and RPE cells expressed little alpha5 protein at their cell surface.
|Active detachment involves inhibition of cell-matrix contacts of malignant melanoma cells by secretion of melanoma inhibitory activity. |
Anja-Katrin Bosserhoff, Raphael Stoll, Jonathan P Sleeman, Frauke Bataille, Reinhard Buettner, Tad A Holak
Laboratory investigation; a journal of technical methods and pathology 83 1583-94 2003
Melanoma inhibitory activity (MIA) has been identified as a small protein secreted from malignant melanoma cells. Recent results revealed a direct interaction of MIA and epitopes within extracellular matrix proteins including fibronectin. The aim of this study was to analyze functional consequences mediated by this interaction. Here we show that MIA interferes specifically with attachment of melanoma cells to fibronectin, a phenomenon we refer to as active detachment. Antibodies inhibiting binding of alpha4beta1 and alpha5beta1 integrins to fibronectin cross-react specifically with MIA, suggesting that MIA shares significant structural homology with the binding pockets of these integrins and thereby masks the respective epitopes on extracellular matrix molecules. Several peptides derived from fibronectin and from a phage display screening were tested with respect to a potential MIA-inhibitory effect. In vitro tests identified two peptides affecting MIA function; both inhibited growth of melanoma metastases in vivo. In summary, we conclude that MIA may play a role in tumor progression and spread of malignant melanomas via mediating active detachment of cells from extracellular matrix molecules within their local milieu. Further, our results suggest that inhibiting MIA functions in vivo may provide a novel therapeutic strategy for metastatic melanoma disease.
|Polarized distribution of alpha5 integrin in dendrites of hippocampal and cortical neurons. |
X Bi, G Lynch, J Zhou, C M Gall, X Bi, G Lynch, J Zhou, C M Gall
The Journal of comparative neurology 435 184-93 2001
The distribution of immunoreactivity for the alpha5 subunit of the fibronectin receptor was evaluated in adult rat brain with particular interest in the cellular localization of immunostaining in the hippocampal formation and neocortex. Beyond localization to neuronal perikarya and short dendritic fragments within most brain areas, alpha5 immunoreactivity (-ir) was particularly dense within primary apical dendrites of pyramidal cells in both hippocampus and neocortex and within the dendritic arbors of cerebellar Purkinje cells. In hippocampal and cortical pyramidal cells, immunostaining was clearly polarized: alpha5-ir was not detectable in basal dendrites in hippocampal neurons and was limited to proximal arbors or absent from basal dendrites in pyramidal cells in superficial and deep layers of neocortex. Beyond this, alpha5-ir was distributed within the dendritic ramifications of the dentate gyrus granule cells and within perikarya and dendrites of occasional nonpyramidal neurons. Developmental studies demonstrated that, in both hippocampus and neocortex, alpha5-ir appears first within perikarya and is distributed to dendrites during the second postnatal week. These results are in accord with the broad hypothesis that integrins contribute to apical-basal differences in dendrites and that the integrin fibronectin (alpha5beta1) receptor, in particular, contributes to some late developing features of dendritic structure or function.
|Evidence that integrins contribute to multiple stages in the consolidation of long term potentiation in rat hippocampus. |
D Chun, C M Gall, X Bi, G Lynch
Neuroscience 105 815-29 2001
Three structurally distinct groups of antagonists were used to test the hypothesis that integrin adhesion receptors play an essential role in consolidating (stabilizing) long term potentiation of the Schaffer collaterals in rat hippocampus. Comparisons were made of percent potentiation at antagonist-treated versus control sites within CA1 stratum radiatum of the same hippocampal slice. Function blocking antibodies against the alpha5 subunit of the fibronectin receptor had no effect on baseline responses or initial potentiation but resulted in a >30% reduction, relative to within-slice control long term potentiation, 45 min later. Larger reductions were recorded in separate experiments continued for 4 h after the induction of potentiation. Alpha(v) and alpha2 subunit antibodies did not reliably affect the stabilization of potentiation. An antagonist peptide with preference for beta1 integrins produced a slowly developing decline of the type seen with alpha5 antibodies. A cyclic peptide antagonist reduced potentiation within 10 min of induction and caused an almost 40% decrease over 45 min. Two disintegrins (snake toxins that potently block integrins) were very effective in preventing the consolidation of long term potentiation: echistatin reduced potentiation by >70%, while triflavin caused approximately 50% decrease. The suppressing effects of echistatin were concentration-dependent, obtained with treatment after induction, and much more rapid than the effects of antibodies. Rapid declines in potentiation were particularly evident when the two disintegrins were applied together.These results indicate that hippocampal fibronectin receptors (alpha5/beta1 integrin) contribute importantly to a slowly developing phase of long term potentiation consolidation. They also suggest that other integrins are critical to aspects of consolidation occurring in the first few minutes after induction.
|Short-term binding of fibroblasts to fibronectin: optical tweezers experiments and probabilistic analysis |
Thoumine, O, et al
Eur Biophys J, 29:398-408 (2000) 2000
|Dynamics of adhesive rupture between fibroblasts and fibronectin: microplate manipulations and deterministic model |
Thoumine, O and Meister, J J
Eur Biophys J, 29:409-19 (2000) 2000
|Glycosylation affects translocation of integrin, Src, and caveolin into or out of GEM. |
A Kazui, M Ono, K Handa, S Hakomori, A Kazui, M Ono, K Handa, S Hakomori
Biochemical and biophysical research communications 273 159-63 2000
Endogenous GM3 synthesis and full N-glycosylation in membrane receptors occurred in 4-epimerase-less ldlD (Krieger's CHO mutant) cells cultured in Gal-containing medium, whereby components of detergent-insoluble, low-density, buoyant membrane fraction, termed glycolipid-enriched microdomain (GEM), varied significantly by translocation into or out of GEM. Integrins alpha3 and alpha5 were translocated into GEM in the presence of 0.5 or 0.25% Triton X-100, particularly in the absence of Gal, whereby integrins are underglycosylated and GlcCer is the major glycolipid component in GEM. Src family kinase was translocated into and enriched in GEM fractions when prepared in 0.5 or 0.25% Triton X-100 from cells grown in Gal-containing medium, whereby GM3 synthesis is induced. In contrast, caveolin is highly enriched in GEM when GM3 synthesis does not occur, and is translocated into high-density membrane fraction when GM3 synthesis occurs. The results suggest that levels of key molecules controlling cell adhesion and signaling are defined by translocation into or out of GEM, which depends on glycosylation state.
|Anti-Integrin alpha5 - Data Sheet|