SEARCH · Search PubMed

Results for “Vitronectin”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 793 records · Page 44Linked to original sources

PubMed1997

Mechanism of initial attachment of corneal epithelial cells to polymeric surfaces.

The initial attachment of cultured bovine corneal epithelial cells and stromal fibroblasts to two oxygen-containing synthetic polymers was studied. Cultured epithelial cells and stromal fibroblasts were seeded onto two oxygen-containing surfaces: 'tissue culture' polystyrene (TCPS) and a polymer film deposited by RF plasma deposition using a methylmethacrylate monomer (MMA/FEP). To establish the mechanism of cell attachment, the effect of the selective removal of the vitronectin and fibronectin from the serum used in the culture medium was tested. The attachment of cultured epithelial cells during the first 90 min of culture was reduced by 40% (TCPS)-80% (MMA/FEP) as a result of removing vitronectin from the medium. Attachment of these cells to TCPS was reduced by 85-95% when the serum was depleted of both fibronectin and vitronectin. However, depletion of fibronectin reduced cell attachment to TCPS by 20%, whilst on MMA/FEP cell attachment was equivalent, or higher, than that for intact serum. The attachment of cultured corneal stromal fibroblasts was similarly dependent on vitronectin but less dependent on fibronectin. Therefore, for the attachment of both cultured epithelial cells and fibroblasts to oxygen-containing surfaces in the presence of serum, there is a high requirement for serum vitronectin but a lesser requirement for fibronectin. The effects of the establishment of corneal epithelial cells in culture and the site of origin of the cells, were determined. Primary isolates of epithelial cells isolated from the limbal, central or peripheral regions of the cornea were less dependent on vitronectin for initial attachment to TCPS than were these cells after several passages in culture. Furthermore, the primary isolates were dramatically less responsive to vitronectin than the cultured cells. These results indicate that the mechanism of attachment of corneal epithelial cells to TCPS varies with the culture experience of the cells. Cells that are culture neophytes can employe endogenous mechanisms for the initial attachment to TCPS, whereas cells established in culture are dependent on exogenous vitronectin in order to attach.

Animals↗

PubMed2002

Expression, regulation, and function of alpha V integrins in hepatocellular carcinoma: an in vivo and in vitro study.

The expression of alpha V integrins by neoplastic cells contributes to the promotion of local invasion and metastasis. The most characteristic extracellular ligands of alpha V integrins are vitronectin and fibronectin. Hepatocytes are the main source of vitronectin, and the capacity to synthesize and secrete vitronectin is usually retained in hepatocellular carcinoma. The aim of this study was to explore the expression, regulation, and functional role of alpha V integrins in hepatocellular carcinoma. We first analyzed the expression of alpha V integrins and their ligands fibronectin and vitronectin in 80 cases of hepatocellular carcinoma. alpha V integrin chain was detected in 44 cases and vitronectin in 50. Twenty-four of the 44 alpha V-positive tumors contained large amounts of vitronectin. These cases presented more frequently with adverse histoprognostic factors, including infiltrative growth pattern (62.5%), lack of capsule (71%), presence of capsular invasion (57%), and satellite nodules (50%). We then used HepG2 and Hep3B cell lines as in vitro models to study alpha V integrin regulation and function. HepG2 and Hep3B cells expressed alpha V integrin chain and used alpha V beta 1 and alpha V beta 5 for adhesion and migration on vitronectin. Tumor necrosis factor (TNF) alpha and transforming growth factor (TGF) beta significantly increased the expression levels of alpha V integrins and stimulated the adhesion and migration of both HepG2 and Hep3B cell lines on vitronectin. The effects of growth factors on cell adhesion and migration were reproduced by incubation with conditioned medium from rat liver myofibroblasts. In conclusion, our results support the existence of an alpha V integrin/vitronectin connection in hepatocellular carcinoma and suggest that this connection may be an adverse prognostic factor.

Adult↗

PubMed1995

Soluble ligands of the alpha v beta 3 integrin mediate enhanced tyrosine phosphorylation of multiple proteins in adherent bovine pulmonary artery endothelial cells.

Binding of substrate-bound extracellular matrix proteins to cell surface integrins results in a variety of cellular responses including adhesion, cytoskeletal reorganization, and gene expression. We have previously shown that addition of soluble SC5b-9, the complement-vitronectin complex, resulted in an RGD-dependent increase in lung venular hydraulic conductivity (Ishikawa, S., Tsukada, H., and Bhattacharya, J. (1993) J. Clin. Invest. 91, 103-109). To identify specific integrin(s) and signal transduction pathways that are responsive to soluble vitronectin-containing ligands, we exposed confluent bovine pulmonary artery cells to purified soluble human mono- or multimeric vitronectin, or SC5b-9, and determined the extent of endothelial cell protein tyrosine phosphorylation. Monomeric vitronectin (Vn) did not induce enhanced protein tyrosine phosphorylation. However, multimeric Vn and SC5b-9 elicited time- and concentration-dependent increases in tyrosine phosphorylation of numerous proteins. Antiserum against vitronectin, RGD peptides, and monoclonal and polyclonal antibodies against the alpha v beta 3 integrin blocked the vitronectin- or SC5b-9-induced enhanced accumulation of tyrosine phosphoproteins, while antibodies against beta 1 integrins and the alpha v beta 5 integrin did not. Clustering of the alpha v beta 3 integrin using monoclonal antibody LM609 caused a pattern of enhanced tyrosine phosphorylation similar to that caused by multimeric Vn and SC5b-9, suggesting that aggregation of alpha v beta 3 was critical for signaling. Among the proteins that underwent enhanced tyrosine phosphorylation in response to vitronectin were the cytoskeletal proteins paxillin, cortactin, and ezrin, as well as the SH2 domain-containing protein Shc, and p125FAK. We conclude that ligation of the alpha v beta 3 integrin by soluble ligands promotes enhanced phosphorylation of several proteins implicated in tyrosine kinase signaling and suggest that this pathway may be important in inflammatory states which are accompanied by accumulation of SC5b-9.

Animals↗

PubMed1999

Distinct roles for astrocyte alphavbeta5 and alphavbeta8 integrins in adhesion and migration.

The alphav integrins are likely to be an important group of molecules for regulating astrocyte behaviour within the central nervous system. Together with their ligand vitronectin, they are expressed by astrocytes in vivo and are further upregulated during neurological disease. Here we have characterised the expression of alphav integrins on primary astrocytes from both rat and mouse, and shown that they express just two members, alphavbeta5 and alphavbeta8. By using RGD peptides and function-blocking antibodies against the beta1 integrins and alphavbeta5, we find that both alphavbeta5 and alphavbeta8 can act as functional receptors for vitronectin. However, while alphavbeta5 is largely responsible for astrocyte adhesion to vitronectin this integrin appears to play no role in migration on vitronectin, with alphavbeta8 playing the dominant role in promoting migration on this substrate. beta1 integrins are not involved in mediating interactions between astrocytes and vitronectin. These results were confirmed in experiments with astrocytes derived from mice in which the beta5 gene had been deleted by homologous recombination. beta5 null astrocytes attached to vitronectin at a reduced rate, but showed no defect in migration on vitronectin relative to wild-type astrocytes. These data provide the first evidence that alphavbeta8 regulates migration and show that astrocyte alphavbeta5 and alphavbeta8 integrins have distinct functions.

Animals↗

PubMed2001

Identification of a target site in plasminogen activator inhibitor-1 that allows neutralization of its inhibitor properties concomitant with an allosteric up-regulation of its antiadhesive properties.

The serpin plasminogen activator inhibitor-1 (PAI-1) has a dual function: 1) it plays an important role as a direct inhibitor of the plasminogen activation system, and 2) its interaction with the adhesive glycoprotein vitronectin suggests a role in tissue remodeling and metastasis, independent from its proteinase inhibitory properties. Unique to this serpin is the close association between its conformational and functional properties. Indeed, PAI-1 can occur in an active and a latent conformation, but both functions are exclusively present in the active conformation. We report here the epitope localization and functional effects of a monoclonal antibody (MA-124K1) that inhibits rat PAI-1 activity and simultaneously increases the binding of inactive PAI-1 to vitronectin (the affinity constant of PAI-1 for vitronectin is 2 x 10(7) m(-1) in the absence of MA-124K1 and 160 x 10(7) m(-1) in the presence of MA-124K1). To the best of our knowledge, this is the first monoclonal antibody dissociating the proteinase inhibitory properties from the vitronectin binding properties in PAI-1. Mutation of Glu(212) and/or Glu(220) in rat PAI-1 to Ala results in a strongly reduced affinity or absence of binding to MA-124K1. The three-dimensional structure of PAI-1 reveals that these residues constitute a conformational epitope close to the reactive-site loop and compatible with the effect of MA-124K1 on the inhibitory properties of PAI-1. However, the vitronectin binding site is localized at the opposite site of the molecule, indicating that the effect of MA-124K1 involves an allosteric modulation of the vitronectin binding site. Cell culture experiments revealed a significant reduction of cell attachment and migration in the presence of MA-124K1, providing evidence for the functional relevance of this antibody-mediated up-regulation of the vitronectin binding properties of PAI-1. In conclusion, a novel mechanism for interference with PAI-1 functions has been identified and is of importance in the modulation of cell migration and related events (e.g. tumor metastasis).

Allosteric Regulation↗

PubMed2004

Regulation of Ca2+-dependent K+ current by alphavbeta3 integrin engagement in vascular endothelium.

Interactions between endothelial cells and extracellular matrix proteins are important determinants of endothelial cell signaling. Endothelial adhesion to fibronectin through alpha(v)beta(3) integrins or the engagement and aggregation of luminal alpha(v)beta(3) receptors by vitronectin triggers Ca2+ influx. However, the underlying signaling mechanisms are unknown. The electrophysiological basis of alpha(v)beta(3) integrin-mediated changes in endothelial cell Ca2+ signaling was studied using whole cell patch clamp and microfluorimetry. The resting membrane potential of bovine pulmonary artery endothelial cells averaged -60 +/- 3 mV. In the absence of intracellular Ca2+ buffering, the application of soluble vitronectin (200 microg/ml) resulted in activation of an outwardly rectifying K+ current at holding potentials from -50 to +50 mV. Neither a significant shift in reversal potential (in voltage clamp mode) nor a change in membrane potential (in current clamp mode) occurred in response to vitronectin. Vitronectin-activated current was significantly inhibited by pretreatment with the alpha(v)beta(3) integrin antibody LM609 by exchanging extracellular K+ with Cs+ or by the application of iberiotoxin, a selective inhibitor of large-conductance, Ca2+-activated K+ channels. With intracellular Ca2+ buffered by EGTA in the recording pipette, vitronectin-activated K+ current was abolished. Fura-2 microfluorimetry revealed that vitronectin induced a significant and sustained increase in intracellular Ca2+ concentration, although vitronectin-induced Ca2+ current could not be detected. This is the first report to show that an endothelial cell ion channel is regulated by integrin activation, and this K+ current likely plays a crucial role in maintaining membrane potential and a Ca2+ driving force during engagement and activation of endothelial cell alpha(v)beta(3) integrin.

Animals↗

PubMed1988

Inhibition of in vitro tumor cell invasion by Arg-Gly-Asp-containing synthetic peptides.

The interaction of cells with extracellular matrix components such as fibronectin, vitronectin, and type I collagen has been shown to be mediated through a family of cell-surface receptors that specifically recognize an arginine-glycine-aspartic acid (RGD) amino acid sequence within each protein. Synthetic peptides containing the RGD sequence can inhibit these receptor-ligand interactions. Here, we use novel RGD-containing synthetic peptides with different inhibition properties to investigate the role of the various RGD receptors in tumor cell invasion. The RGD-containing peptides used include peptides that inhibit the attachment of cells to fibronectin and vitronectin, a peptide that inhibits attachment to fibronectin but not to vitronectin, a cyclic peptide with the opposite specificity, and a peptide, GRGDTP, that inhibits attachment to type I collagen in addition to inhibiting attachment to fibronectin and vitronectin. The penetration of two human melanoma cell lines and a glioblastoma cell line through the human amniotic basement membrane and its underlying stroma was inhibited by all of the RGD-containing peptides except for the one that inhibits only the vitronectin attachment. Various control peptides lacking RGD showed essentially no inhibition. This inhibitory effect on cell invasion was dose-dependent and nontoxic. A hexapeptide, GRGDTP, that inhibits the attachment of cells to type I collagen in addition to inhibiting fibronectin- and vitronectin-mediated attachment was more inhibitory than those RGD peptides that inhibit only fibronectin and vitronectin attachment. Analysis of the location of these cells that were prevented from invading indicated that they attached to the amniotic basement membrane but did not proceed further into the tissue. These results suggest that interactions between RGD-containing extracellular matrix adhesion proteins and cells are necessary for cell invasion through tissues and that fibronectin and type I collagen are important for this process.

Amino Acid Sequence↗

PubMed1993

Integrin beta 1- and beta 3-mediated endothelial cell migration is triggered through distinct signaling mechanisms.

Human umbilical vein endothelial cell attachment, spreading and migration on collagen and vitronectin are mediated by integrins alpha 2 beta 1 and alpha v beta 3, respectively, and these events take place in the absence of cytokines, growth factors, or chemoattractants. Cell attachment and spreading on these ligands occur in the absence of extracellular calcium, as does migration on collagen. In contrast, vitronectin-mediated migration is absolutely dependent on the presence of extracellular calcium. Cell contact with immobilized vitronectin or anti-alpha v beta 3 mAbs promotes a measurable rise in [Ca2+]i which requires an extracellular calcium source, whereas collagen, or anti-alpha 2 beta 1 mAbs fail to promote this signaling event. In fact, vitronectin-mediated migration and the rise in intracellular calcium showed the same dose dependence on extracellular calcium. While vitronectin and collagen differ in their ability to induce a calcium influx both ligands or antibodies to their respective integrins promote an equivalent increase in intracellular pH consistent with activation of the Na/H antiporter an event independent of extracellular calcium. These results support two salient conclusions. Firstly, collagen and vitronectin, through their respective integrins, promote distinct intracellular signaling events. Secondly, the alpha v beta 3 specific influx of calcium is not required for cell spreading yet appears to facilitate cellular migration on vitronectin.

Calcium↗

PubMed2006

Effect of pharmacologic plasminogen activator inhibitor-1 inhibition on cell motility and tumor angiogenesis.

BACKGROUND: Plasminogen activator inhibitor-1 (PAI-1) is integrally involved in tumorigenesis by impacting on both proteolytic activity and cell migration during angiogenesis. OBJECTIVES: We hypothesized that an orally active small molecule inhibitor of PAI-1 (PAI-039; tiplaxtinin) could affect smooth muscle cell (SMC) attachment and migration in vitro on a vitronectin matrix, and exhibit antiangiogenic activity in vivo. METHODS: In vitro assays were used to assess the mechanism of inhibition of PAI-1 by PAI-039 using wild-type PAI-1 in the presence or absence of vitronectin and wild-type PAI-1 and specific PAI-1 mutants in SMC adhesion and migration assays. An in vivo tumor angiogenesis model was used to assess the effect of PAI-039 administration on neovascularization in a Matrigel implant. RESULTS: PAI-039 dose-dependently inhibited soluble, but not vitronectin-bound, PAI-1. Cell adhesion assays using PAI-1 mutants unable to bind vitronectin (PAI-1K) or inactivate proteases (PAI-1R) further suggested that PAI-039 inactivated PAI-1 by binding near its vitronectin domain. In a tumor angiogenesis model, PAI-039 treatment of wild-type mice dose-dependently decreased hemoglobin concentration and endothelial cell staining within the Matrigel implant, indicating reduced angiogenesis, but exhibited no in vivo efficacy in PAI-1 null mice. CONCLUSIONS: Administration of an orally active PAI-1 inhibitor prevented angiogenesis in a Matrigel implant. The lack of activity of PAI-039 against wild-type PAI-1 bound to vitronectin and PAI-1K suggests PAI-039 binding near the vitronectin-binding site. Our studies further substantiate a role for PAI-1 in cellular motility and tumor angiogenesis, and suggest for the first time that these effects can be modulated pharmacologically.

Angiogenesis Inhibitors↗

PubMed2001

Plasminogen activator inhibitor-1 promotes angiogenesis by stimulating endothelial cell migration toward fibronectin.

Increased expression of plasminogen activator inhibitor-1 (PAI-1) in cancer patients is associated with unfavorable outcome, and the reason for this paradox has been poorly understood. We have previously reported elevated levels of PAI-1 in primary tumors of advanced neuroblastomas (Y. Sugiura et al., Cancer Res., 59: 1327-1336, 1999). Here we demonstrate that PAI-1 is coexpressed with the angiogenesis marker alpha(v)beta3 integrin in blood vessels of primary neuroblastoma tumors, suggesting that PAI-1 plays a role in angiogenesis. Using human brain microvascular endothelial cells (HBMECs), we found that PAI-1 inhibits alpha(v)beta3 integrin-mediated cell adhesion to vitronectin but promotes alpha5beta1-mediated migration from vitronectin toward fibronectin. Inhibition of vitronectin adhesion by PAI-1 did not induce HBMEC apoptosis. PAI-1 also inhibited endothelial tube formation on Matrigel in the presence of vitronectin but had a stimulatory effect in the presence of fibronectin. This effect of PAI-1 on microvascular endothelial cells is primarily related to the ability of PAI-1 to bind to vitronectin via its NH2-terminal domain and to interfere with cell adhesion to vitronectin. We propose that PAI-1 acts as a positive switch for angiogenesis by promoting endothelial cell migration away from their vitronectin-containing perivascular space toward fibronectin-rich tumor tissue. These observations provide a novel explanation for the enhancing effect of PAI-1 in cancer progression.

Apoptosis↗

PubMed1990

Localization of beta 1, beta 3, alpha 5, alpha v, and alpha IIb subunits of the integrin family in spreading human erythroleukemia cells.

The localization of five integrin subunit proteins was studied in human erythroleukemia (HEL) cells spreading on various culture substrata in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) and the absence of serum. The cells readily adhered on fibronectin, but TPA was needed for adherence on vitronectin and for the spreading of the cells on both substrata. Indirect immunofluorescence microscopy showed that in the spread cells cultured on vitronectin or fibronectin for 2 hours, beta 1, beta 3, alpha 5, and alpha IIb integrin subunits were localized at focal adhesions as identified by talin-immunoreactivity. The alpha v integrin immunoreactivity was initially found at the focal adhesions when the cells were cultured on vitronectin, but was also found later in cells cultured on fibronectin. The alpha IIb integrin immunoreactivity disappeared from focal adhesions within 24 hours. The alpha 5 and beta 1 integrin immunoreactivities disappeared from the focal adhesions in cells cultured on vitronectin, but not in cells cultured on fibronectin. When the cells were plated on glass substratum in the presence of TPA, they spread much slower than on vitronectin or fibronectin, but some cells showed focal adhesions after only 8 hours in culture. In this case, the alpha v and beta 3 integrin subunits were found at focal adhesions. After TPA treatment, HEL cells deposited thrombospondin-immunoreactive material onto their culture substratum, but synthesis of fibronectin, vitronectin, fibrinogen, or von Willebrand factor was not detected. Thus, the results suggest that TPA would activate several integrin receptors in HEL cells and also stimulate the secretion of thrombospondin, which might be used as an adhesion ligand for the integrin vitronectin receptor alpha v/beta 3 complex.

Cell Adhesion↗

PubMed2003

The pattern of enhancement of Src kinase activity on platelet-derived growth factor stimulation of glioblastoma cells is affected by the integrin engaged.

Enhanced expression of both integrin alpha v beta 3 and platelet-derived growth factor receptor (PDGFr) has been described in glioblastoma tumors. We therefore explored the possibility that integrin alpha v beta 3 cooperates with PDGFr to promote cell migration in glioblastoma cells, and extended the study to identify the Src family members that are activated on PDGF stimulation. Glioblastoma cells utilize integrins alpha v beta 3 and alpha v beta 5 to mediate vitronectin attachment. We found that physiologic PDGF stimulation (83 pm, 10 min) of vitronectin-adherent cells promoted the specific recruitment of integrin alpha v beta 3-containing focal adhesions to the cell cortex and alpha v beta 3-mediated cell motility. Analysis of PDGFr immunoprecipitates indicated an association of the PDGFr beta with integrin alpha v beta 3, but not integrin alpha v beta 5. Cells plated onto collagen or laminin, which engage different integrins, exhibited significantly less migration on PDGF stimulation, indicating a cooperation of alpha v beta 3 and the PDGFr beta in glioblastoma cells that promotes migration. Further analysis of the cells plated onto vitronectin indicated that PDGF stimulation caused an increase in Src kinase activity, which was associated with integrin alpha v beta 3. In the vitronectin-adherent cells, Lyn was associated preferentially with alpha v beta 3 both in the presence and absence of PDGF stimulation. In contrast, Fyn was associated with both alpha v beta 3 and alpha v beta 5. Moreover, PDGF stimulation increased the activity of Lyn, but not Fyn, in vitronectin-adherent cells, and the activity of Fyn, but not Lyn, in laminin-adherent cells. Using cells attached to mAb anti-alpha v beta 3 or mAb anti-integrin alpha 6, we confirmed the activation of specific members of the Src kinase family with PDGF stimulation. Down-regulation of Lyn expression by siRNA significantly inhibited the cell migration mediated by integrin alpha v beta 3 in PDGF-stimulated cells, demonstrating the PDGFr beta cooperates with integrin alpha v beta 3 in promoting the motility of vitronectin-adherent glioblastoma cells through a Lyn kinase-mediated pathway. Notably, the data indicate that engagement of different integrins alters the identity of the Src family members that are activated on stimulation with PDGF.

Base Sequence↗

PubMed1988

Identification of a bone sialoprotein receptor in osteosarcoma cells.

Bone sialoprotein (BSP) is an extracellular matrix glycoprotein associated with the mineral bone matrix. The amino acid sequence of BSP contains an Arg-Gly-Asp (RGD) sequence which confers to the protein cell binding properties (Oldberg, A., Franzén, A., and Heinegård, D. (1988) J. Biol. Chem. 263, 19430-19432). When BSP was used as an affinity matrix to isolate a cell surface receptor from rat osteosarcoma cells, a protein composed of polypeptides similar in size to those of a previously characterized vitronectin receptor was obtained. This putative BSP receptor, like the vitronectin receptor, bound also to an affinity matrix made of an RGD-containing heptapeptide. Moreover, similar patterns of inhibition of cell attachment to BSP and vitronectin was obtained with variant RGD-containing peptides, with BSP and with vitronectin. Finally, an anti-vitronectin receptor antiserum immunoprecipitated a receptor identical in size to the receptor bound to a BSP affinity matrix. These results show that BSP is recognized by an RGD-directed receptor and that both vitronectin and BSP can bind to this receptor.

Animals↗

PubMed1995

Human mast cells derived from fetal liver cells cultured with stem cell factor express a functional CD51/CD61 (alpha v beta 3) integrin.

Human fetal livers contain progenitor cells that become mast cells after 4 weeks of culture with recombinant human stem cell factor. Expression of cell surface CD29 (beta 1), CD18 (beta 2), CD61 (beta 3), and beta 5 integrins was investigated on such cells by flow cytometry and adhesion measurements. High surface expression of CD49e, CD51, and CD61 along with kit was apparent by 4 weeks of culture, whereas expression of each at day 0 was low to undetectable. CD29 and CD49d were detected on cells from day 0 to 4 weeks of culture; CD49b, CD49c, CD49f, CD18, and CD54 expression was negligible. The fetal liver-derived mast cells spontaneously adhered to vitronectin. No evidence for degranulation was found during vitronectin-dependent adhesion. Adhesion occurred in part through the CD61/CD51 receptor. No evidence for adhesion to vitronectin through CD29 and beta 5 integrins was obtained. Almost all of the vitronectin-adherent cells expressed CD51, CD61, kit, and tryptase, and exhibited metachromasia with toluidine blue. Thus, among the fetal liver-derived cells, developing mast cells were selectively adherent to vitronectin. These mast cells and the other cell types present also adhere spontaneously to fibronectin and to laminin, this adhesion being partially inhibited by antibodies against CD61 and CD29 integrins. In conclusion, human mast cells acquire functional vitronectin receptors as they develop from fetal liver progenitors under the influence of rhSCF. This may be important for the recruitment, localization, and retention of developing mast cells.

Amino Acid Sequence↗

PubMed1996

Involvement of integrin alphavbeta3 in cell adhesion, motility, and liver metastasis of murine RAW117 large cell lymphoma.

The molecules that mediate metastatic cell homing to specific organ sites remain largely unidentified. As a target organ for metastasis, the liver is a unique environment characterized by fenestrated sinusoidal endothelium, lack of a complete basement membrane, and production of serum components, including fibronectin and vitronectin. We examined a series of marine RAW117 large cell lymphoma variants selected in vivo for liver-colonizing properties (H10 >> L17 > P). Compared with L17 or P cells, the highly liver-colonizing H1O cells expressed much higher levels of surface integrin alphavbeta3, as shown by affinity chromatography, immunoprecipitation, and flow cytometry. H10 cells adhered at higher rates to vitronectin and fibronectin than to fibrinogen, fibrin, laminin, and type I collagen. Among the RGD peptides, H10 cells adhered at significantly higher rates to the polymeric RGD peptide (glycyl-arginyl-glycyl-aspartyl-serine)4 than to monomeric RGD peptides. H10 cells were able to spread on immobilized vitronectin with highly polarized morphology but not on fibronectin. In contrast, the poorly liver-metastatic P and L17 cells did not adhere or spread well on vitronectin or fibronectin. H10 cells also migrated toward vitronectin concentration gradients. Blocking cell surface alphavbeta3 molecules with specific anti-beta3 monoclonal antibodies resulted in significant decreases in the adhesion of H10 cells to vitronectin and (glycyl-arginyl-glycyl-aspartyl-serine)4 and significant inhibition of the formation of experimental liver metastases. These data suggest an important role of integrin alphavbeta3 in the metastasis of RAW117 cells to the liver.

Amino Acid Sequence↗

PubMed1996

Mouse polymorphonuclear granulocyte binding to extracellular matrix molecules involves beta 1 integrins.

The mechanism of adhesion of purified mouse polymorphonuclear granulocytes (PMN) to extracellular matrix proteins characteristic of basement membranes and the interstitium has been investigated and compared with the adhesion of a mouse progranulocytic cell line, 32DC13, and a mouse monocytic cell line, WEHI 78/24. All three cell types bound specifically to fibronectin and vitronectin to different degrees under different cellular activation states. 32DC13 bound to fibronectin and vitronectin strongly, and this binding increased upon cellular activation with phorbol 12-myristate-13-acetate (PMA) but not with formyl-Met-Leu-Phe. Only 32DC13 showed significant binding to laminin-1. By contrast, WEHI 78/24 and PMN bound only fibronectin and vitronectin; this binding was weak and was altered only marginally upon activation with PMA. In the case of WEHI 78/24, a slight increase in adhesion both to fibronectin and to vitronectin was observed after cellular activation with PMA, while PMN adhesion to both substrates was slightly reduced. The mechanism of binding to fibronectin and vitronectin was similar in the three cell types. The integrin alpha5 beta1 mediated fibronectin adhesion, demonstrating for the first time the existence of a functionally active beta1 integrin on mouse PMN. Vitronectin binding was mediated by alpha(v) beta3, as demonstrated by the ability of alpha(v)-specific cyclic L-Arg-L-Gly-L-Asp-D-Phe-L-Val (RGDfV) peptide (EMD66203), and anti-beta3 antibody to inhibit cell adhesion. 32DC13 adhesion to laminin-1 was via the alpha6 beta1 integrin. None of the three cell types tested bound to the basement membrane proteins collagen type IV and perlecan, or to the interstitial stromal constituents tenascin, collagen types I, V and VI. Interestingly, perlecan and collagen type IV were found to repel all three cell types. The relative inability of PMN, WEHI 78/24, and 32DC13 to bind to extracellular matrix proteins characteristic of basement membranes and their ability to bind inflammatory markers of the interstitium is discussed with respect to leukocyte extravasation processes.

Animals↗

PubMed1997

Cytotoxic T lymphocytes express a beta3 integrin which can induce the phosphorylation of focal adhesion kinase and the related PYK-2.

Fibronectin has been shown to stimulate tyrosine phosphorylation of a number of proteins in the 115-125 kDa range and facilitate degranulation by alloantigen-specific cytotoxic T lymphocyte (CTL) clones in response to substimulatory amounts of anti-CD3 or anti-T cell receptor (TCR). The current study was initiated to further characterize integrin expression and usage by these CTL clones. We demonstrate that vitronectin and fibrinogen, but not laminin or collagen, are also able to both facilitate degranulation in the presence of substimulatory anti-CD3 and stimulate tyrosine phosphorylation of these 115-125-kDa proteins, with a 115-kDa protein being the most prominently phosphorylated. These results implicate the expression and usage of the vitronectin receptor, alpha beta3 integrin, by these CTL clones. We demonstrate by both flow cytometry and immunoprecipitation that CTL clones do in fact express beta3 integrin. Immobilized antibody to beta3 stimulates the phosphorylation of the 115-125-kDa proteins, suggesting that engagement of beta3 transmits the same signal into these cells as fibronectin or vitronectin. The fibronectin and vitronectin-induced phosphorylation as well as adhesion to either fibronectin or vitronectin can be significantly inhibited with antibodies to beta3 integrins. Finally, we are able to immunoprecipitate 115-kDa proteins with antiserum to focal adhesion kinase and a related kinase, called PYK-2, that becomes phosphorylated in response to vitronectin or immobilized anti-beta3. Taken together, these results demonstrate that CTL express and use beta3-integrins as signaling molecules which can augment TCR-mediated stimulation.

Animals↗

PubMed1995

Distribution of extracellular matrix receptors in various forms of glomerulonephritis.

Integrins are heterodimeric transmembrane receptor glycoproteins consisting of alpha and beta subunits that mediate adhesion and interactions between cells and extracellular matrix. Such interactions may be perturbed in various pathologic states, resulting in the altered phenotypic expressions of the integrins in affected tissues. To ascertain the alterations in integrins in various renal diseases, their distribution was investigated in different forms of glomerulonephritis by indirect immunofluorescence and immunoelectron microscopy using specific antibodies directed against beta 1 integrins and integrin alpha v beta 3 (vitronectin receptor). In addition, the distribution of certain extracellular matrix components (ie, fibronectin, vitronectin, and type IV collagen) was examined. Integrin beta 1 and alpha v beta 3 were highly expressed in proliferating mesangial cells in immunoglobulin A nephropathy, membranoproliferative glomerulonephritis type I and diffuse proliferative lupus nephritis. Their putative ligands (ie, fibronectin, vitronectin, and type IV collagen) also were increased in the expanded mesangial regions. In immunoglobulin A nephropathy, integrin beta 1 and alpha v beta 3 were seen by immunoelectron microscopy to be localized to the mesangial cell membranes in close proximity to the immune complex deposits; however, fibronectin and vitronectin immunoreactivities were observed in the mesangial immune complex deposits. Similarly, vitronectin also was detected in the immune complex deposits of other forms of proliferative nephritis, ie, membranoproliferative glomerulonephritis type I and diffuse proliferative lupus nephritis. In diffuse proliferative lupus nephritis, the cellular crescents displayed immunoreactivity toward integrin alpha v beta 3 and vitronectin. In nonimmune complex glomerular disease associated with nephrotic syndrome (ie, minimal change nephrotic syndrome), integrin alpha 3 beta 1, which normally has a linear capillary distribution, was decreased.(ABSTRACT TRUNCATED AT 250 WORDS)

Collagen↗