Expression And Significance Of CCR3 And Its Ligand Eotaxin In Rheumatoid Arthritis

Rheumatoid arthritis (rheumatoid arthritis, RA) is a systemic autoimmune diseases characterized with chronic multiple arthritis. The release of inflammatory medium and inflammatory cytokines and the following synovitis and vasculitis is the primary pathological change. Based in many studies with RA patients and rheumatoid arthritis animal model, we have known that the chemokines family and their receptors are involved in the process to recruit inflammatory cells in the iinflammatory articulus and synovial neovascularization. The role of inflammatory chemokines is powerful than the effect of anti-inflammatory chemokines, thus accelerating the progress of the synovial tissue inflammation and played an important role in the RA pathological progress. But the expression of chemokine receptor on RA synovial fibroblast-like synoviocytes and its role in joint inflammation is not clear.CCR3,a seven-transmembrane domain G-protein-coupled receptor, is founded on the surface of the eosinophils, is a member of the CC chemokine receptor family. It’s also be found expressed on the basophile,Th2cell,dendritic cell,mast cell,respiratory epithelial cells and endothelial cells. The CCR3ligand including CCL5(RANTES), CCL7(MCP-2), CCL8(MCP-3), EOTAXIN/CCL11(Eotaxin), CCL13(MCP-4), CCL15(HCC-2), CCL24(Eotaxin-2), CCL26(Eotaxin-3), CCL28(MEC),but the particular one is the Eotaxin/CCL11and CCR3is the only signal recognition receptor of Eotaxin.. With combination, Eotaxin and CCR3activated the MAP kinase, phospholipase A2, phosphatidyl inositol3kinase, protein kinase C, signal transduction and activation of transcription factor (STAT), intracellular Ca+inflow, actin polymerization, of protease activation and active release oxygen. The serum Eotaxin/CCL11level is relatively higher in Juvenile rheumatoid arthritis and early rheumatoid arthritis (RA), and in peripheral blood CD4+T lymphocytes and monocytes,the CCR3expression is obviously decreased. The pro-inflammatory cytokines can stimulate arthritic cartilage cells release/CCL13MCP-4, and Eotaxin/CCL11can promote the MMP3expression and secretion of arthritic cartilage cells.The research results of animal experiments confirmed that CCR3is involved in the recruiting white blood cells into the articular. CCR3and its ligands involved in joint inflammation, and may play an important role in the development of joint inflammation. But the role of CCR3and its ligands, especially the Eotaxin, in inflammation articular is still unclear. As the important cell in arthritis, whether CCR3express on RAFLS or not is not known. To study the CCR3and its ligands can help us to understand their role in joint inflammation well, and provide some references for further clinical study. PartⅠ The expression of CCR3in peripheral blood and synovialtissue of RA patients BACKGROUND:The expression of CCR3in peripheral blood and synovial tissue of RA patients has been reported. However, the distribution and expression level of CCR3in different cell populations of T cells and synovial fibroblasts surface are still unclear. METHODS:Flow cytometry analysis of the distribution and expression of CCR3on CD4+, CD8+-positive cells in synovial tissue from RA (n=16) and OA (n=18) patients, and peripheral blood from RA (n=8) and OA (n=8) patients. The expression of CCR3on synovial fibroblast surface of synovial tissue was also analysed. The possible role and significance of CCR3in the development of RA were explored. RESULTS:Compared with patients with OA, the percentage of CD4+CCR3+ cells in the single-cell suspensions from synovial tissue and the percentage of CD8+CCR3+cells from peripheral blood of RA decreased, which had a significant difference (p<0.05). The percentage of CD4+CD25+FoxP3+CCR3+cells from peripheral blood in RA was lower than that in OA (p<0.05), However, the percentage of CCR3on CD19+CD3-CD40cells (synovial fibroblast) in RA was higher than that in OA, which also had a significant difference (p<0.05). CONCLUSION:The percentage of CD4+CCR3+cells from synovial tissue and CD8+CCR3+cells from peripheral blood of RA is decreased. CCR3is high expressed on RAFLS in RA. CCR3and chemokine ligand may participate in the improvement of the RAFLS migration and invasion ability, and in RA joints play an important role in the process of pathological damage. PART ⅡEotaxin and CCR3promotes migration and invasive ability of rheumatoid arthritis (RA) synovial fibroblastsBACKGROUND:Articular synovial is the main target tissure in rheumatoid arthritis (RA). Fibroblast like synoviocyte(FLS) plays the most important role in RA pathology. Eotaxin and CCR3not only play an important role in the inflammatory cell recruitment, also participated in the cancer cells and endothelial cells migration and invasion. We known that the CCR3expression on CD19+CD3-CD40cells (synovial fibroblast) in RA was higher, This research expounds the pro-inflammation cytokine promote RAFLS production of Eotaxin. Eotaxin interaction with CCR3increased the MMP2and MMP9secretion, and improved the migration and invasion ability of RAFLS. It’s partly illuminated the role of Eotaxin and CCR3in the inflammation and bone erosion of RA. METHODS:Culturing primary synovial fibroblasts and stimulating with different concentration of TNF-alpha,ILl-beta respectively, collected at different time points. The expression of CCR3,Eotaxin mRNAwere assayed with Real-time PCR detection;The concentrations of Eotaxin/CCL11,eotaxin-2/CCL24and eotaxin-3/CCL26in supernatant preparations of RAFLS were determined by Enzyme-Linked Immunosorbent Assay (ELISA). Immunofluorescence double stain observation CCR3expression on FLS surface. FLS were cultured in the presence or absence of Eotaxin, CCR3inhibitor SB328437, or controls, then studied for migration,invasion and MMP-2-9production.RESULTS:TNF-α, IL-1β can significantly improve the RAFLS CCR3mRNA expression (P<0.05), and RAFLS produced higher levels of Eotaxin secretion (P <0.05). IL-1β significantly stimulated CCL24production, but TNF-a decreased the CCL24secretion of RAFLS.Eotaxin with different concentrations increased RAFLS CCR3mRNA expression(P<0.05). Eotaxin treatment increased RAFLS migration,invasion, and MMP-2-9mRNA expression.Th increase of migration and invasion was decreased by SB328437as much as42% and64% separately.CONCLUSION:We describe and characterize a new autocrine/paracrine role for Eotaxin-CCR3in the regulation of RAFLS migration,invasion, and MMPs expression. These observations suggest that the Eotaxin-CCR3interaction is a potential new target for therapies aimed at reducing FLS migration,invasion and its associated cartilage damage,bone destruction and carticular disability in RA.