ABSTRACT
OBJECTIVE: To evaluate the modulation of RhoA/Rho kinase (ROCK), a small Rho GTPase, on migration, invasion and proliferation of fibroblast like synoviocytes (FLS) from rheumatoid arthritis (RA) patients. METHODS: RA FLS were collected from active RA patients. And 10% fetal bovine serum (FBS) and interleukin-1ß (IL-1ß) were used as stimuli in migration and proliferation experiments respectively. RhoA activity was measured by pull down assay while ROCK activity by Western blot. FLS migration and invasion in vitro were measured by the Transwell chamber method. And thiazolyl blue tetrazolium bromide (MTT) test was used to detect cell proliferation. RESULTS: There were increased activities of RhoA and ROCK in ex vivo FLS from RA versus OA patients and healthy control. The migrated cell number of FBS-induced, C3-treated and Y27632-treated groups was 85 ± 14, 51 ± 15 and 42 ± 11 respectively. The Matrigel invading cell number of 3 groups was 64 ± 13, 39 ± 12 and 26 ± 9 respectively. Statistical differences existed in cell number between FBS-induced, C3-treated or Y27632-treated group (P < 0.05) in above migration and invasion experiments. Inhibition of RhoA and ROCK activity also suppressed the cytoskeletal reorganization and proliferation of RA FLS. CONCLUSION: Increased RhoA/ROCK activity may contribute to abnormal migration, invasion and proliferation of RA FLS. Thus inhibition of ROCK activity may be a new therapeutic target for RA.
Subject(s)
Arthritis, Rheumatoid/metabolism , rho-Associated Kinases/metabolism , rhoA GTP-Binding Protein/metabolism , Arthritis, Rheumatoid/pathology , Cell Movement , Cell Proliferation , Cells, Cultured , Female , Fibroblasts/cytology , Humans , Male , Synovial Membrane/cytologyABSTRACT
OBJECTIVE: To evaluate the modulation of RhoA/Rho kinase (ROCK) signaling pathway, a small Rho GTPase that is considered as an important modulator in inflammatory responses, on Toll-like receptor-2 mediated chemokine secretion in fibroblast-like synoviocytes (FLS) from rheumatoid arthritis (RA) patients. METHODS: The RhoA activity was measured by a pull-down assay. And the ROCK activity was assessed by Western blot. The secretion of chemokines was measured by ELISA (enzyme-linked immunosorbent assay). MTT test was used to detect the cellular viability. RESULTS: The stimulation of peptidoglycan (PG, 5 mg/L) increased the levels of IL-8 (interleukin-8), RANTES (regulated upon activation normal T cell expressed & secreted) and MCP-2 (monocyte chemotactic protein-2) and boosted the activities of RhoA and ROCK versus the unstimulated RA FLS. And these effects of PG were suppressed by anti-TLR-2 monoclonal antibody. Inhibition of RhoA and ROCK with a specific inhibitor inhibited the secretion of IL-8, RANTES and MCP-2 in PG-induced RA FLS. CONCLUSION: The present study provides novel evidence that the RhoA/ROCK signal pathway modulates the TLR-2-mediated secretion of chemokines in RA FLS. It suggests that the inhibition of RhoA/ROCK may be a new therapeutic approach for RA.
