The Effect Of Oleanolic Acid On Tumor Necrosis Factor Alpha-induced Inflammatory Cytokines Expression And The Related Mechanism In Synovial Cells

Objective: to investigate the effect of oleanolic acid on tumor necrosis factor alpha-induced inflammatory cytokines production and the related mechanism in synovial cells,which will lay a theoretical basis for the application of oleanolic acid in rheumatoid arthritis.Methods: 1.Cultured and passaged the fibroblasts synovial cells in vitro.2.Experimental methods: the activity of synovial cells was determined by 4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT).RT-PCR was used to detect the mRNA expression of Interleukin-6(Interleukin-6,IL-6)and Interleukin-1?(Interleukin-1?,IL-1?).Western blot was used to detect the protein expression of p38 MAPK and NF-?B.ELISA was used to detect the levels of IL-6 and IL-1? in cell supernatant.3.Experimental groups: according to the research purpose,the experiment is divided into three parts:(1)to detect the effect of oleanolic acid on the basic condition of synovial cells,the cells can be divided into six groups:control group and different concentration of oleanolic acid group(5 ?mol/L,10?mol/L,20?mol/L,40?mol/L,80?mol/L);(2)to detect the effect of oleanolic acid on TNF-?-induced inflammatory cytokines release,the cells were divided into five groups: control group,the TNF-? group and TNF-? +different concentrations oleanolic acid group(5?mol/L,10?mol/L,20?mol/L);(3)to test p38 MAPK and NF-?B protein expression,the cells were divided intofive groups: control group,the TNF-? group,TNF-?+ different concentrations oleanolic acid group(5?mol/L,10?mol/L,20?mol/L);(4)to detect the effect of the inhibitors of p38 MAPK and NF-?B on the levels of inflammatory cytokines,the cells were divided into 5 groups: control group,the TNF-? group,TNF-?+oleanolic acid(20?mol/L)group,TNF-?+ the inhibitor of p38 MAPK group,TNF-?+ the inhibitor of NF-?B group.Results: 1.5?mol/L-20?mol/L Oleanolic acid has no effect on the activity of synovial cells(P > 0.05).However,when the concentration is more than 40?mol/L,Oleanolic acid can inhibit the activity of synovial cells(P < 0.05).2.Compared with control group,TNF-? significantly promote the secretion of IL-6 and IL-1?,statistically significant difference(P < 0.05).3.Compared with the TNF-? group,oleanolic acid can inhibit TNF-?-induced mRNA expression of IL-6 and IL-1? in concentration dependence(P < 0.05).4.Compared with control group,TNF-?can promote the phosphorylated p38 MAPK,statistically significant difference(P < 0.05);Compared with the TNF-? group,oleanolic acid can inhibit p38 MAPK phosphorylation induced by TNF-? in concentration dependence(P< 0.05).5.Compared with control group,the TNF-? can promote the protein expression of NF-?B in nucleus,statistically significant difference(P < 0.05);Compared with TNF-? group,oleanolic acid can inhibit the nucleoprotein expression of NF-?B induced by TNF-?(P < 0.05).6.When added the p38 MAPK blocker SB203580 or NF-?B inhibitors Bay11-7082,the secretion of IL-6 and IL-1? decreased significantly compared with control group(P <0.05).Conclusions: 1.TNF-? can promote the expression of IL-6 and IL-1?,and promote the phosphorylated p38 MAPK and NF-?B nuclear transfer in synovial cells.2.Oleanolic acid can inhibit TNF-?-induced expression of inflammatory cytokines(IL-6 and IL-1?),and suppress TNF-?-induced phosphorylation of p38 MAPK and NF-?B nuclear transfer in synovial cells.3.Oleanolic acid suppressed the expression of inflammatory cytokines by inhibiting p38 MAPK and NF-?B signaling pathway.