獐牙菜苦苷、龙胆苦苷和獐牙菜苷对类风湿关节炎成纤维样滑膜细胞促凋亡作用及Bcl-2表达情况的影响

Effect of swertiamarin,gentiopicrin and sweroside on cell apoptosis and expression of Bcl-2 in rheumatoid arthritis fibroblast-like synoviocytes

探讨獐牙菜苦苷、龙胆苦苷和獐牙菜苷对类风湿关节炎成纤维样滑膜细胞(rheumatoid arthritis fibroblast-like synoviocytes,RA-FLSs)及B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl-2)的影响并讨论其作用机制。通过ZINC数据库和RCSB PDB数据库查询獐牙菜苦苷、龙胆苦苷和獐牙菜苷的三维化学成分结构和Bcl-2的蛋白靶点三维结构;借助AutoDock Mgltools 1.5.6软件、AutoDock Vina 1.1.2软件和pyMOL 2.2.0软件将獐牙菜苦苷、龙胆苦苷和獐牙菜苷的三维化学成分结构与Bcl-2(1GJH)靶点蛋白三维结构进行对接并分析;借助Annexin V-FITC细胞凋亡试剂盒检测给药后各组RA-FLSs的凋亡情况;借助Western blot法检测给药后各组RA-FLSs的Bcl-2蛋白表达情况;借助RT-PCR技术检测给药后各组RA-FLSs的Bcl-2 mRNA表达情况。各给药组的成分结构与Bcl-2(1GJH)的结合活性较好,獐牙菜苦苷的结合能为-6.9 kcal·mol^(-1),龙胆苦苷的结合能为-6.7 kcal·mol^(-1),獐牙菜苷的结合能为-6.4 kcal·mol^(-1);龙胆苦苷组的促细胞凋亡作用最强(P<0.01),獐牙菜苷组次之,獐牙菜苦苷组最次;龙胆苦苷组的Bcl-2蛋白表达水平最低,抑制Bcl-2蛋白表达作用最强(P<0.01),獐牙菜苦苷组和獐牙菜苷组间比较无差异,作用相近;各给药组较空白组的Bcl-2 mRNA基因表达水平显示出不同程度降低(P<0.01,P<0.05),且作用相近。龙胆苦苷可以促进RA-FLSs凋亡,降低RA-FLSs的Bcl-2蛋白表达水平和Bcl-2 mRNA的基因表达水平,进而缓解因RA-FLSs的凋亡不足与异常增殖引起的病症。

The aim of this paper was to discuss the effect of swertiamarin,gentiopicrin and sweroside on rheumatoid arthritis fibroblast-like synoviocytes(RA-FLSs)and B-cell lymphoma-2(Bcl-2)and their mechanisms.ZINC database and RCSB PDB database were retrieved for 3 D chemical structures of swertiamarin,gentiopicrin and sweroside and 3 D target protein structures.AutoDock Mgltools 1.5.6,AutoDockVina 1.1.2 and pyMOL 2.2.0 were applied for molecular docking to analyze the relationship between Bcl-2(1 GJH)target protein and important ingredients.The cell apoptosis of RA-FLSs was tested by Annexin V-FITC.The Bcl-2 protein expression of RA-FLSs treated with different ingredients was tested by Western blot.The Bcl-2 mRNA expression of RA-FLSs treated with different ingredients was tested by RT-PCR.Swertiamarin,gentiopicrin and sweroside were docked well with Bcl-2(1 GJH).The binding energy of swertiamarin was-6.9 kcal·mol^(-1),the binding energy of gentiopicrin was-6.7 kcal·mol^(-1)and the binding energy of sweroside was-6.4 kcal·mol^(-1).Compared with the blank group,the Bcl-2 protein expression of each group were reduced,while that of the gentiopicrin group was the highest(P<0.01).Compared with the blank group,the Bcl-2 mRNA expression of each groups were reduced.Gentiopicrin can reduce the Bcl-2 protein expression and the Bcl-2 mRNA expression,so as to promote the RA-FLSs apoptosis.