The Effect Of MTX On CRP Induced-inflammation Of Rheumatoid Arthritis Synovial Fibroblasts

Objective To investigate the effect of methotrexate on C reaction protein induced-inflammation of rheumatoid arthritis synovial fibroblasts.Methods Rheumatoid arthritis Fibroblast-like synoviocytes (RAFLS) and Normal Human synovial fibroblasts (HFLS) were cultured in vitro and were divided into the control group, CRP stimulation group, MTX combined with CRP group, respectively. The level of mRNA and protein expression of IL-6, MMP3 and TNFa in RAFLS and HFLS was determined by semi-quantitative real-time PCR (qPCR) and Enzyme-Linked Immunosorbent assays (ELISA), respectively. The level of the protein expression of phosphorylation of P65 (P-P65)and IκB in each group was also estimated by Western-blot method in RAFLS.Results Compared with control and HFLS, the levels of mRNA and protein expression of IL-6, MMP3, TNFa, and P-P65 in RAFLS induced by CRP was significantly higher, while the level of IkB expression was lower. It is suggested that CRP can active the NF-Kb pathway and induce inflammation in RAFLS. Compared with control and RAFLS induced by CRP group, MTX could significantly down-regulate the mRNA and protein expression level of CRP. Moreover, MTX could antagonize the inducing inflammatory role of CRP which up-regulated the mRNA and its protein expression level of IL-6, MMP3 and TNFa in RAFLS. CRP can promote phosphorylation of P65 and degradation IκB, meanwhile, MTX suppress the expression of CRP, leading to decreasing phosphorylation of P65 and degradation IκB.Conclusion The result of our study demonstrates that MTX could significantly down-regulate the expression of CRP in RAFLS, antagonistic the NF-κB pathway which activated by CRP, and inhibit the production of IL-6, MMP3 and TNFa. It suggested that suppression effect on induced-inflammation of CRP in RFLS may be one of the mechanisms of MTX treated RA.