The Effects And Mechanisms Of Wasp Venom On Rheumatoid Arthritis Fibroblast-like Synoviocytes

Objective:The study of functional mechanism of wasp venom extract on apoptosis and inflammation of rheumatoid arthritis fibroblast synovium cell MH7 A induced by tumor necrosis factor TNF-α,and further explored the best effect of wasp venom extract on the proliferation and inflammation of MH7 A cells,and preliminarily explored the possible mechanism of inducing apoptosis and reducing inflammation of MH7 A cells.Methods:In this experiment,TNF-α was used to induce inflammatory reaction and excessive proliferation of MH7 A cells,resulting in RA model.After successful establishment of the model,wasp crude venom,<3 k Da wasp venom,3～10 k Da wasp venom and >10 k Da wasp venom were named as wasp crude venom,wasp venom Ⅰ,wasp venom Ⅱ and wasp venom Ⅲ.1.Screening of anti-rheumatoid arthritis activity of wasp crude venom,wasp venom Ⅰ and wasp venom Ⅱ in vitro.According to the molecular weight of wasp venom(< 3 k Da,3～10 k Da,> 10 k Da),the crude wasp venom was divided into wasp venom Ⅰ,wasp venom Ⅱ and wasp venom Ⅲ by ultracentrifugation tube method.Because wasp venom III contains a large number of allergic media,it is not used as an active screening object.MTT method was used to detect cell proliferation /inhibition and the best time for wasp venom to inhibit the proliferation of MH7 A cells.Enzyme-linked immunosorbent assay was used to detect the expression of inflammatory factors IL-1β and IL-6,and to investigate the best effective parts of wasp crude venom,wasp venom Ⅰ and wasp venomⅡ on RA.2.Preliminary study on the mechanism of anti-rheumatoid arthritis effect of wasp venom Ⅱ in vitro.The apoptosis rate was detected by Annexin V-FITC/PI double staining and the expression of p-JAK2,p-STAT3,Bax and Bcl-2 in JAK2/STAT3 and Bax/Bcl-2 signal pathway was detected by Western blotting method.Results:1.The half inhibitory rate of MH7 A cells treated with wasp crude venom for24 h was not significantly different from that at 48 h and 72 h;wasp crude venom,wasp venom Ⅰ and wasp venom Ⅱ could inhibit the proliferation of MH7 A cells and reduce the expression of IL-1β and IL-6 in MH7 A cells induced by TNF-α in a dose-dependent manner.However,the activity of wasp venom Ⅱ is better than that of wasp venom and wasp venom Ⅰ.2.Wasp venom Ⅱ could increase the apoptosis rate of MH7 A cells induced by TNF-α,and wasp venom Ⅱinhibit the expression of phosphorylated proteins of JAK2 and STAT3 in JAK2/STAT3 signal pathway,down-regulate the expression of Bcl-2protein,and up-regulate the expression of Bax protein.It had a significant differences(p < 0.01 or p < 0.05)and showed a dose-dependent manner.Conclusion:The results showed that wasp venom could down-regulate the expression of inflammatory cytokines(IL-1β,IL-6)in TNF-α-induced MH7 A cells through inhibiting the activation of JAK2/STAT3 signal pathway,and regulate the apoptosis of TNF-α-induced MH7 A cells via Bax and Bcl-2 proteins in mitochondria.It was found that wasp venom Ⅱ was an effective part of anti-RA,which provided a certain experimental basis for the further development of active monomers of wasp venom.