cs1-peptide and Arthritis--Rheumatoid

VLA-4 is a critical adhesion molecule that regulates mononuclear cell trafficking to sites of inflammation. VCAM-1 is a primary ligand of VLA-4, although alternatively spliced fibronectin (FN) containing the CS1 region (CS1 FN) also binds to VLA-4. CS1 FN is expressed by rheumatoid arthritis (RA) synovial endothelial cells, but the factors that regulate CS1 FN expression are not known. We incubated human umbilical vein endothelial cells (HUVEC) with IL-1 (0.1-10 ng/ml) for 8-48 h and determined total FN and CS1 FN mRNA by Northern blot analysis. Both were constitutively expressed by HUVEC, and IL-1 increased total FN mRNA and the CS1-containing isoform (P < 0.05). IL-1 also increased CS1 FN protein expression on HUVEC as determined by Western blot analysis. An adhesion assay using (51)Cr-labeled Jurkat cells and IL-1-stimulated HUVEC was used to determine if IL-1-induced CS1 FN mediates cell binding. Cyclic CS1 peptide (10 microg/ml) blocked 49 +/- 5% of IL-1-induced Jurkat cell adhesion to HUVEC (P < 0.01), whereas anti-VCAM-1 antibody inhibited binding by only 35 +/- 5% (P < 0.01). CS1 peptide and anti-VCAM antibody treatment were not additive (50 +/- 7% inhibition), and 38 +/- 6% of new VLA-4-mediated adhesion to IL-1-treated HUVEC was due to an increase in CS1 FN. These data show that IL-1 increases CS1 FN expression by HUVEC and increases CS1-mediated cell adhesion. CS1 mimetics might have therapeutic efficacy by blocking recruitment of VLA-4-bearing cells.

Topics: Alternative Splicing; Arthritis, Rheumatoid; Cell Adhesion; Cells, Cultured; Dose-Response Relationship, Drug; Endothelium, Vascular; Fibronectins; Gene Expression Regulation; Humans; Intercellular Signaling Peptides and Proteins; Interleukin-1; Jurkat Cells; Peptides; Peptides, Cyclic; Protein Isoforms; Synovial Membrane; Vascular Cell Adhesion Molecule-1

It has previously been shown that the adhesion of lymphocytes to microvascular endothelium mediates lymphocyte extravasation within inflamed synovium. After passing the endothelial barrier, binding of lymphocytes to matrix proteins and synovial lining cells may further lead to synovial membrane hyperplasia and subsequent cartilage destruction. Thus, we have explored the molecular basis of T cell-synovial lining cell interaction in the synovial membrane of patients with rheumatoid arthritis (RA).. Using an immunohistochemical staining technique and an in vitro frozen section assay we studied the expression and the role of several adhesion molecules in T lymphocyte-synovial lining cell interaction in the inflamed synovial membrane.. In RA the macrophage-like (type A) synovial lining cells express high levels of intercellular adhesion molecule 1 [ICAM-1 (CD54)], whereas the fibroblast-like (type B) synovial lining cells predominantly express vascular cell adhesion molecule 1 (VCAM-1), in addition to moderate levels of ICAM-1. Both cell types express low levels of fibronectin. Unstimulated and anti-CD3 stimulated peripheral blood T cells bear the respective ligands lymphocyte function associated antigen 1 [LFA-1 (CD18/11a)], and very late antigen 4 and 5 [VLA-4 (CD29/49d) and VLA-5 (CD29/49e)]. T lymphocytes predominantly bound to type B synovial lining cells. Inhibition studies with monoclonal antibodies revealed that this binding involves the VLA-4/VCAM-1 and VLA-5/fibronectin (FN), but not the VLA-4/CS1 pathway. LFA-1 is also involved in this interaction via its ligand ICAM-1.. These results show that the molecular basis of T lymphocyte binding to rheumatoid synovial lining cells is different from that described for T lymphocyte binding to synovial membrane vascular endothelium which involves the VLA-4/VCAM-1 and VLA-4/CS-1 pathways, but not the LFA-1/ICAM-1 pathway.

Topics: Antibodies, Monoclonal; Arthritis, Rheumatoid; Cell Adhesion; Cell Adhesion Molecules; Cell Line; Endothelium, Vascular; Humans; Immunohistochemistry; Intercellular Adhesion Molecule-1; Intercellular Signaling Peptides and Proteins; Lymphocyte Function-Associated Antigen-1; Peptides; Receptors, Very Late Antigen; Synovial Membrane; T-Lymphocytes; Vascular Cell Adhesion Molecule-1

It has previously been shown that the very late antigen-4/vascular cell adhesion molecule-1 (VLA-4/VCAM-1) pathway functions as a receptor/ligand interaction system mediating the recruitment of activated lymphocytes to inflamed synovium of patients with rheumatoid arthritis. This study was performed to determine whether VLA-4 also affects lymphocyte adhesion to inflamed synovium through interaction with the alternatively spliced CS1 domain of fibronectin.. The effect of the synthetic peptide CS1 on lymphocyte binding to human synovial and peripheral lymph node high endothelial venules (HEVs) was measured in an in vitro frozen section assay.. In the presence of the CS1 peptide or antibody to fibronectin, significant inhibition of binding was observed (54 and 51% respectively). Blocking with antibody to VCAM-1 yielded inhibition of binding to 46% of the control value. Maximum inhibition of binding was obtained with a combination of antibody to VCAM-1 and CS1 (65%) and with antibody to VLA-4 alpha (68%). Blocking the classical fibronectin receptor with antibody to VLA-5 alpha gave a slightly lower inhibition at 42%. In normal peripheral lymph nodes, the synthetic peptide CS1 and antibodies to fibronectin and VLA-5 also partially inhibited T cell binding to HEVs (45, 47, and 52% respectively).. These results show that fibronectin mediates lymphocyte-HEV interactions not only through its classical VLA-5 receptor, but also through its CS1 adhesion motif in inflamed synovium and peripheral lymph nodes.

Topics: Arthritis, Rheumatoid; Binding, Competitive; Cell Adhesion; Culture Techniques; Endothelium, Lymphatic; Fibronectins; Humans; Immunoenzyme Techniques; Intercellular Signaling Peptides and Proteins; Peptides; Synovial Membrane; T-Lymphocytes