Predominant activation of MAP kinases and pro-destructive/pro-inflammatory features by TNF α in early-passage synovial fibroblasts via TNF receptor-1:: failure of p38 inhibition to suppress matrix metalloproteinase-1 in rheumatoid arthritis

Objective: To examine the relative importance of tumour necrosis factor- receptor 1 (TNF-R1) and TNF-R2 and their signalling pathways for pro- inflammatory and pro- destructive features of early-passage synovial fibroblasts (SFB) from rheumatoid arthritis (RA) and osteoarthritis ( OA). Methods: Cells were stimulated with tumour necrosis factor (TNF) a or agonistic anti- TNF-R1/TNF-R2 monoclonal antibodies. Phosphorylation of p38, ERK and JNK kinases was assessed by western blot; proliferation by bromodesoxyuridine incorporation; interleukin (IL) 6, IL8, prostaglandin E-2 (PGE(2)) and matrix metalloproteinase (MMP)-1 secretion by ELISA; and MMP- 3 secretion by western blot. Functional assays were performed with or without inhibition of p38 (SB203580), ERK (U0126) or JNK (SP600125). Results: In RA- and OA- SFB, TNF alpha- induced phosphorylation of p38, ERK or JNK was exclusively mediated by TNF-R1. Reduction of proliferation and induction of IL6, IL8 and MMP-1 were solely mediated by TNF-R1, whereas PGE2 and MMP-3 secretion was mediated by both TNF-Rs. In general, inhibition of ERK or JNK did not significantly alter the TNF alpha influence on these effector molecules. In contrast, inhibition of p38 reversed TNFa effects on proliferation and IL6/ PGE2 secretion (but not on IL8 and MMP-3 secretion). The above effects were comparable in RA- and OA- SFB, except that TNFa- induced MMP-1 secretion was reversed by p38 inhibition only in OA- SFB. Conclusion: In early- passage RA/OA- SFB, activation of MAPK cascades and pro-inflammatory/ prodestructive features by TNFa is predominantly mediated by TNF-R1 and, for proliferation and IL6/PGE2 secretion, exclusively regulated by p38. Strikingly, RA-SFB are insensitive to p38 inhibition of MMP-1 secretion. This indicates a resistance of RA-SFB to the inhibition of pro-destructive functions and suggests underlying structural/ functional alterations of the p38 pathway, which may contribute to the pathogenesis or therapeutic sensitivity of RA, or both.