VLA Proteins Research Articles

Upregulated Expression of VLA Proteins and CD29 in Peripheral Blood Lymphocytes of Chronic Alcoholics Without Ethanol‐Related Diseases

To analyze adhesion molecule expression on peripheral blood mononuclear cells (PBMCs) and on different lymphocyte subpopulations (CD2+, CD8+, CD19+, and CD56+ subsets) in chronic alcoholism, 30 well-nourished chronic alcoholics without ethanol-related diseases and 30 matched controls were included in the study. Adhesion molecules that mediate adhesion to other cells and to extracellular matrix proteins, and whose cellular expression is modified during lymphocyte activation, were selected for study. A detailed clinical evaluation, laboratory analysis, nutritional assessment, and study of adhesion molecule expression was performed. A significant higher expression of CD29 (beta1-integrin) (p = 0.001), VLA-3 (p = 0.002), VLA-4 (p = 0.03), and VLA-5 (p = 0.001) were observed on PBMCs of chronic alcoholics, compared with control subjects, whereas no changes were observed in CD18 (beta2-integrin) and CD50 (ICAM-3) expression. The upregulation of CD29 and VLA proteins only affected T lymphocytes (CD2+/CD8+/CD4+ cells). These data confirm that T cells of chronic alcoholics are basally activated and that changes in adhesion molecule expression on PBMCs may be responsible of disturbances of adhesion processes in chronic alcoholics without ethanol-related diseases.

Upregulated Expression of VLA Proteins and CD29 in Peripheral Blood Lymphocytes of Chronic Alcoholics Without Ethanol-Related Diseases

Upregulated Expression of VLA Proteins and CD29 in Peripheral Blood Lymphocytes of Chronic Alcoholics Without Ethanol-Related Diseases

IL-15 is chemotactic for natural killer cells and stimulates their adhesion to vascular endothelium.

Interleukin-15 (IL-15) is a recently described cytokine with IL-2-like stimulating activities on T lymphocytes and natural killer (NK) cells. IL-15 mediates its function through the beta- and gamma-chains of the IL-2 receptor. In this work, we have investigated the effect of IL-15 on the directional migration of NK cells in chemotaxis assays and on the ability of NK cells to bind to vascular endothelium. IL-15 (10-20 ng/mL) had chemotactic effects on freshly isolated resting NK cells as well as on long-termed IL-2-cultured NK cells. A checkerboard experiment demonstrated that migration in response to IL-15 was observed only in the presence of a positive gradient (chemotaxis). Overnight treatment of freshly isolated NK cells with IL-15 (10-20 ng/mL) augmented their binding to cultured endothelial cells (EC) in vitro, especially to resting EC. IL-15-activated NK cells bound to resting and tumor necrosis factor-activated EC by use of LFA-1/ICAM-1 and VLA-4/VCAM-1 adhesion pathways, essentially as untreated NK cells do. The fact that IL-15 increased NK cell binding to ICAM-1-transfected NIH-3T3 fibroblasts, together with the finding that IL-15 did not increase binding to extracellular matrix proteins, where the major molecules involved are VLA proteins, indicated that IL-15 primarily stimulates LFA-1-dependent adhesion. By increasing NK cell adhesion to vascular endothelium and migratory response, IL-15 is an important determinant of NK cell recruitment in tissues.

Lymphocyte Traffic and Positioning in Vivo: An Expande Role for the ECM, the VLA Proteins and the Cytokines

The continuous circulation of lymphocytes between blood and lymph, constitutes, with the specific recognition of foreign antigens and with memory, one of the hallmarks of the immunological system. In the routine process of recirculating between blood and lymph in adult life, lymphocytes display the unique capacity of recognizing "self within self". The physiological recognition of "self within self" is expressed at three levels: (1) an overall ability to distinguish between peripheral lymphoid and non-lymphoid organs, (2) the ability of lymphocyte subpopulations to discriminate between peripheral lymphoid organs, (3) a fine recognition of distinct microenvironments within the peripheral lymphoid organs. Historically, interest in regulation of entry into the lymph dominated this field, focusing primarily on the interaction of lymphocytes with post-capillary venules with high endothelium found in some higher vertebrate lymph nodes. Lymphocytes, however, enter the lymph in species without lymph nodes, and lymphocytes recirculation is well established in fetal life. Regardless of route of entry, lymphocytes in birds, fish, rodents, and humans enter the peripheral lymphoid organs and display the capacity to segregate and arrange themselves in distinct territories, a phenomenon called "ecotaxis". This paper reviews the evidence for the relative contribution of so called specialized lymphocyte endothelium interactions and other interactions, to the physiological regulation of lymphocyte traffic and positioning. Of the latter, interactions of lymphocytes with the ECM appear of some significance for two reasons: there is a selective distribution of some ECM components in peripheral lymphoid organ areas, as recently shown for tenascin in the thymus-dependent zones (Chilosi et al., Am J Pathol 143: 1348-1355, 1993). This selective distribution may serve as a basis for preferential T cell migration through those zones. The abnormal expression of tenascin in non-lymphoid organs with autoimmune lesions may serve as an identical basis for abnormal T lymphocyte migration in autoimmunity (Chilosi, personal communication). Additional evidence for abnormal cell-ECM interactions playing a role in autoimmune-like lesions comes from the recent observations in TGF-beta 1 knockout mice, in which mononuclear cell infiltration of the heart and lungs has been corrected by the systemic administration of synthetic FN peptides (Hines at al., PNAS 91:5187-5191, 1994). Changes in expression of ECM components have aso being described in rat heart allografts preceding lymphocyte accumulation in the process of allograft rejection (Coito et atl. Transplantation 57:599-605, 1994).(ABSTRACT TRUNCATED AT 400 WORDS)

Adhesion molecules: Cellular recognition mechanisms

Integrin Expression and Epithelial Cell Differentiation V. Quaranta. Fibronectin Mutations in Mice E.N. Georges, et al. Extracellular and Intracellular Functions of VLA Proteins M.E. Hemler, et al. Signal Transduction from Leukocyte Integrins E.J. Brown. Coordinate Role for Proteoglycans and Integrins in Cell Adhesion J.B. McCarthy, et al. Adhesion Molecules at Endothelial Cell to Cell Junctions M.G. Lampugnani, et al. Activation of LFA1 C.G. Figdor, Y. van Kooyk. Interaction of Cytotoxic T Lymphocytes with Autologous Melanoma A. Anichini, et al. Tumor Cell-Endothelial Cell Interactions during Blood Borne Metastasis G.L. Nicolson, et al. Adhesion Mechanisms in Lymphoma and Carcinoma Metastasis G. La Riviere, et al. CD44 and Splice Variants of CD44 in Normal Differentiation and Tumor Progression P. Herrlich, et al. 6 additional articles. Index.

Functional role of alpha 2/beta 1 and alpha 4/beta 1 integrins in leukocyte intercellular adhesion induced through the common beta 1 subunit.

Whereas all of the integrins in the VLA protein subfamily are involved in cell-extracellular matrix interactions, only VLA-4 (through the alpha 4 subunit) has been implicated in the triggering of intercellular adhesion. Here we describe that the VLA protein beta 1 subunit (CD29) is also involved in the induction of homotypic cell aggregation. We have obtained three novel anti-beta 1 monoclonal antibodies (mAb) with the ability to induce cell aggregation on different leukocyte cell types. These mAb recognize an antigenic site on the common beta 1 chain of VLA proteins which is topographically and/or functionally distinct from other epitopes previously defined by several prototype anti-beta 1 mAb. Induction of cell aggregation by anti-beta 1 mAb is epitope specific, isotype and Fc independent, and displays kinetics similar to alpha 4-mediated aggregation. This cell aggregation requires an intact cellular metabolism, the presence of divalent cations in the extracellular medium, and the integrity of the cytoskeleton. We also have found that the Na+/H+ antiporter may be essential for this process. For Ramos cells, which bear only the VLA alpha 4/beta 1 heterodimer, intercellular adhesion induced through the VLA-beta 1 chain could be selectively inhibited by other anti-beta 1 mAb as well as by anti-alpha 4 mAb. Interestingly, anti-beta 1 mAb which induced strong aggregation of VLA-alpha 2- or VLA-alpha 4-transfected K562 cells, had minimal effect on the alpha 2- alpha 4- alpha 5+ K562 cell line. Furthermore, the beta 1-mediated induction of cell aggregation on alpha 2-K562- and alpha 4-K562-transfected cells was blocked by preincubation with either anti-alpha 2 or anti-alpha 4 mAb, respectively, as well as by other anti-beta 1 mAb. Interestingly, parental K562 cells were able to interact with both alpha 2- and alpha 4-transfected K562 cells, thus suggesting that counter-receptors for both integrins (VLA-2 and VLA-4) might exist on these cells. Together these results provide strong evidence supporting the involvement of alpha 2/beta 1 and alpha 4/beta 1 heterodimers in intercellular interactions and underline the pivotal role of the common beta 1 chain of VLA proteins in the integrin-mediated induction of cell aggregation.

Two integrin-binding peptides abrogate T cell-mediated immune responses in vivo.

Two VLA proteins (or beta 1 integrins; originally called very late activation antigens) that bind to distinct determinants on fibronectin (FN) are increased on activated immune or memory T cells. VLA-4 binds to the peptide sequence Gly-Pro-Glu-Ile-Leu-Asp-Val-Pro-Ser-Thr (GPEILDVPST in single-letter code) on the alternatively spliced CS-1 form of FN, whereas VLA-5 binds to an Arg-Gly-Asp sequence found on all forms of FN. It has been proposed that the migration of immune T cells out of blood vessels and through connective tissue to a site of antigenic challenge is facilitated by the interaction of such integrins with matrix protein molecules. We have examined directly the role of T-cell integrins in vivo by using the well-characterized, T-cell-mediated contact hypersensitivity (CHS) response to the hapten trinitrochlorobenzene (TNCB). We demonstrate that the cells that transfer CHS to TNCB adhere to FN in the presence of Ca2+/Mg2+, and T-cell populations depleted of FN-adherent cells do not transfer immunity. We further show that TNCB-immune T cells treated with the synthetic peptides GPEILDVPST or Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP in single-letter code), ligands for VLA-4 and VLA-5, respectively, lose their ability to mediate this immune response in a murine model, whereas the control peptides Val-Ile-Pro-Asp-Leu-Thr-Glu-Ser-Pro-Gly and Gly-Arg-Gly-Glu-Ser-Pro have no effect. Neither GPEILDVPST nor GRGDSP significantly inhibited the proliferative response of TNCB-immune T cells in vitro. These data suggest that FN-binding integrins on T cells play a role in the localization of T cells to sites of antigenic challenge in tissue.

T cell receptor-dependent, antigen-specific stimulation of a murine T cell clone induces a transient, VLA protein-mediated binding to extracellular matrix.

Upon Ag stimulation, an arsonate-specific murine T cell clone exhibited a rapid but transient increase in cell adhesion to collagen, fibronectin, and laminin. This increase in cell adhesion was not observed when a mutant T cell clone lacking TCR expression was utilized. However, upon stimulation by phorbol esters, both parent and mutant T cell clones exhibited a similar transient increase in adhesion to the three matrix proteins. The observed cell adhesion was extensively inhibited by antibodies to the integrin beta 1 subunit, indicating the involvement of VLA proteins. Despite changes in the adhesive properties, there was essentially no difference in the expression of VLA-1, -3, -4, -5, and -6 between resting and stimulated T cells. Together these results suggest that Ag stimulation transmits signals via the TCR complex resulting in a rapid, but transient, up-regulation of matrix protein binding by VLA proteins already present at the cell surface. Because the appropriate reagents that recognize individual mouse VLA proteins were not available, we used the human T cell line Jurkat to demonstrate that T cell binding to collagen, laminin, and fibronectin is mediated largely by VLA-2, VLA-6, and a combination of VLA-5 and VLA-4, respectively.

Human Epidermal Langerhans Cells Express Integrins of the β1 Subfamily

The expression of VLA integrins on human epidermal Langerhans cells (LC) was investigated by indirect immunogold labeling on transmission electron microscopy, followed by a quantitative analysis. Labelings on suspensions enriched in freshly isolated LC were carried out with an antibody recognizing the beta 1 subunit, common to all members of the VLA family, and with antibodies specific for the six different alpha subunits, alpha 1 to alpha 6. Normal human epidermal LC were all beta 1 positive, of which 60% were highly positive. By labelings with different VLA-alpha-chain MoAb, there appeared two subpopulations of LC: one positive and one negative. On average 40% of LC bore small amounts of VLA-1 and VLA-3, 53% and 77% of LC expressed moderate amounts of VLA-2 and VLA-5, respectively, and VLA-4 and VLA-6 were expressed by 67% and 90% of LC, respectively. VLA proteins are mainly extracellular matrix protein receptors. VLA-6 (laminin receptor) and VLA-5 (fibronectin receptor) are expressed mainly by LC, and in this way could subserve LC to leave the epidermal compartment through the basement membrane, to migrate throughout the fibronectin network of the dermis before migrating via the afferent lymphatics to the regional lymph nodes, where they present antigen to T cells. VLA proteins such as VLA-4, VLA-3, or VLA-2, which are found involved in cell-cell contacts, could contribute to the promotion of T-cell activation by facilitating adherence between LC and T cells.

VLA protein expression on epidermal cells (keratinocytes, Langerhans cells, melanocytes): a light and electron microscopic immunohistochemical study

The very late antigens, or VLA proteins, are a family of cell surface heterodimers (alpha 1-b beta 1) that mediate cell adhesion to specific components of the extracellular matrix (collagens, fibronectin, laminin). In normal human epidermis, the common VLA beta 1 subunit is expressed on basal keratinocytes (BK). Langerhans cells (LC) and melanocytes. By means of light and electron microscopic immunostaining procedures, we have investigated the distribution of VLA alpha 1,2,3,4,6 subunits on normal human adult and foetal epidermal cells. alpha 1 antigen expression was not observed on any epidermal cell type. Both during foetal development and in adult epidermis, alpha 2 and alpha 3 were strongly expressed on the cell membrane BK, while alpha 6 was mainly expressed at their dermal pole. These different patterns of distribution suggest that the alpha 6 subunit may mediate BK anchorage to the basement membrane zone, while the alpha 2 and alpha 3 subunits may also be involved in intracellular adhesion. Moreover, with immunoelectron microscopy, LC were seen to be weakly alpha 5 and alpha 6 positive and melanocytes were alpha 3 and alpha 6 positive. Thus, VLA proteins are expressed by epidermal cells in a cell-type-specific pattern that could be related to particular functional roles of these proteins.

Identification of specific human epithelial cell integrin receptors as VLA proteins

Cell adhesion to extracellular matrix is mediated by a set of heterodimeric cell surface receptors called integrins. We have examined the expression of the very late antigens or αβ1 group of integrins in human epithelial cells. The six known members of this group share a common β1 subunit but have distinct α subunits that confer selective affinity toward collagen, fibronectin, and laminin essentially. Using a panel of specific antibodies we showed that freshly harvested human epidermal basal cells express VLA-2 and VLA-3 receptors, a low amount of VLA-5, but fail to express VLA-4. The findings reveal that these receptors are characterized by the α subunits which associate with a β subunit different in weight ( M r 110,000 reduced) from that normally seen ( M r 130,000). Moreover, immunoprecipitates of VLA-2 contained additional proteins of M r 80,000 and M r 40,000 and immunoprecipitates of VLA-3 contained an additional protein of M r 90,000. Experiments carried out to investigate the functional roles of these receptors in mediating cell adhesion to extracellular matrix revealed that cell attachment to type IV collagen was completely inhibited by antibodies to VLA-2 α chain, that antibody to VLA-3 α chain significantly blocked attachment to fibronectin while antibodies to both VLA-2 and VLA-3 partially inhibited attachment to type I collagen. Cell attachment to types I and IV collagen and to fibronectin was not affected by antibodies to VLA-4 and VLA-6. These results show that multiple VLA receptors function in combination to mediate epidermal basal cell adhesion to extracellular matrix. This cooperation function of multiple VLA receptors and their differential expression could be considered to be one of the controlling points in the localization of epithelial basal cells in the epidermis.

VLA Proteins in the Integrin Family: Structures, Functions, and Their Role on Leukocytes

Article de synthese sur les roles des integrines β1 (proteines VLA) comme recepteurs dans l'adherence des leucocytes a la matrice extracellulaire et comme recepteurs dans l'adherence cellule a cellule

VLA Proteins In The Integrin Family: Structures, Functions, And Their Role On Leukocytes

VLA Proteins In The Integrin Family: Structures, Functions, And Their Role On Leukocytes

Isolated human follicular dendritic cells display a unique antigenic phenotype.

In the present study, follicular dendritic cells (FDCs) were purified to homogeneity in order to define the lineage and function of these cells. FDCs were identified by their characteristic morphology and by their expression of receptors for the third complement component, the myeloid-restricted antigen CD14, and the FDC antigen DRC-1. Unclustered FDCs displayed a unique antigenic phenotype since they expressed several B- and myeloid lineage-restricted antigens, but lacked T and NK cell antigens as well as the leukocyte common antigen. FDCs expressed adhesion molecules, including most of the VLA proteins, intercellular adhesion molecule 1 (ICAM-1), and CD11b. FDCs could be isolated to homogeneity by their intense staining with anti-CD14 using flow cytometric cell sorting. These highly purified FDCs expressed CD14 and CD21 but lacked CD20. This antigen pattern and characteristic morphology confirmed that these cells were, in fact, homogeneous FDC preparations. Analysis of polymerase chain reaction-amplified cDNA from highly purified FDCs showed no transcripts for IL-6. The isolation of homogeneous FDC populations will be important for the analysis of the functional role of FDCs within the lymphoid follicle.

Mapping of human fibronectin receptor beta subunit gene to chromosome 10.

Human-mouse hybrid cells were examined by indirect immunofluorescence with Mab DH12, a monoclonal antibody that recognizes the beta subunit of the human fibronectin receptor. Cells that expressed the antigen at their surface were sorted by FACS and karyotyped. Immunoaffinity chromatography on Mab DH12 was used to confirm the presence of the human antigen. The chromosome assignment was strengthened by isozyme analysis of markers for chromosomes 9 and 10. The results are suggestive for a 10p mapping of this beta subunit of the fibronectin receptor. Since the gene coding for the beta subunit of the VLA proteins was previously assigned to the same chromosome, our result could provide further evidence for the relationship between the beta subunit of the human fibronectin receptor and the VLA protein family.

Fibronectin receptor structures in the VLA family of heterodimers.

Multiple cell surface proteins of relative molecular mass 115,000-155,000 (Mr 115K-155K) have been implicated as receptors mediating adhesion to extracellular matrix proteins. But the organization and relatedness of these peptides has remained unclear. In separate studies, the 'very late antigens' VLA-1 (Mr 210K/130K) and VLA-2 (Mr 160K/130K) were initially characterized as surface heterodimers appearing 2-4 weeks after in vitro stimulation of human T cells. Three more VLA heterodimers have since been discovered, which, like VLA-1 and VLA-2, are each composed of unique alpha-subunits in association with a common 130K beta subunit. This paper shows that the common VLA beta-subunit is equivalent to subunits found in structures with known fibronectin and laminin receptor activity, and that VLA-3 and VLA-5 are similar or identical to these previously defined receptors for adhesion molecules. Antibody blocking studies confirmed that at least some of the widely distributed VLA proteins of previously unknown function are involved in cell adhesion to fibronectin and laminin. We suggest that the VLA family of receptors may provide cells with multiple independent substrate adhesion capabilities.

The VLA protein family. Characterization of five distinct cell surface heterodimers each with a common 130,000 molecular weight beta subunit.

The family of human cell surface heterodimers which includes VLA-1 and VLA-2 is now shown to include three additional heterodimers, here called VLA-3, VLA-4, and VLA-5. Each of these separate VLA structures is composed of a distinct alpha subunit (Mr 110,000-200,000 nonreduced) noncovalently associated with a common beta subunit (Mr 110,000 nonreduced). Chemical cross-linking experiments provided evidence that each VLA complex exists predominantly as a 1:1 alpha beta heterodimer. The VLA proteins are widely distributed, with one or more of the heterodimers present on nearly all cell types tested. Evidence for five distinct VLA alpha subunits was obtained from differences observed in antibody recognition, cell distribution patterns, two-dimensional gel analyses, and V8 protease cleavage patterns. On the other hand, the beta subunit present in each heterodimer was immunochemically and electrophoretically indistinguishable, and yielded identical V8 cleavage fragments. Immunoblotting experiments revealed that besides the Mr 110,000 beta normally seen, another beta protein was present that is smaller in size (Mr 90,000 nonreduced), altered or deficient in glycosylation, and not available for cell surface radiolabeling.

Use of the monoclonal antibody 12F1 to characterize the differentiation antigen VLA-2.

A monoclonal antibody, 12F1, has been produced that specifically immunoprecipitates the human cell surface structure VLA-2 from platelets and long-term activated T cells, as well as from fibroblast and neuroblastoma cell lines. Cross-linking studies indicate that the VLA-2 structure exists on the cell surface as a 165,000 Mr heavy chain (alpha 2) in noncovalent 1:1 association with a 130,000 Mr light chain (beta). The monoclonal antibody A-1A5, which reacts with the beta subunit common to all VLA structures, was able to completely preclear VLA-2, indicating that all of the alpha 2 subunit was associated with VLA beta-chain. The specificity of 12F1 for VLA-2 allowed independent immunoprecipitation and flow cytometry analysis of this alpha 2 beta structure separate from any other VLA structures that may have been present such as VLA-1 or free beta-subunit. Subunit dissociation studies were used to demonstrate that 12F1 recognizes an epitope on the alpha 2 chain on VLA-2, which is consistent with the 12F1 specificity for VLA-2 alone among the VLA proteins. Analysis of activated T cells indicated that VLA-2, like VLA-1, is another "very late" appearing T cell activation antigen that arises concurrently with VLA-1 starting at day 7 and increasing through 2 wk. VLA-2 was found on many of the same cells as VLA-1 (inactivated T cells, T cell leukemia cells, fibroblasts, SK-N-SH neuroblastoma cells), but VLA-1 and VLA-2 can be expressed independently, because VLA-2 was also present on VLA-1-negative cells such as HSB and platelets, and VLA-1 was present on VLA-2-negative C8215 cells.