麻花艽6个成分对类风湿性关节炎滑膜成纤维样细胞的影响及机制

Effects and its mechanism of six components of Gentiana straminea Maxim.in rheumatoid arthritis fibroblast-like synoviocytes

目的:探讨麻花艽6个成分对类风湿性关节炎滑膜成纤维样细胞(RA-FLS)的影响及机制。方法:采用CCK-8法筛选麻花艽6个成分的最佳给药浓度;流式细胞术测定最佳给药浓度干预下的细胞凋亡水平,ELISA检测细胞凋亡诱导因子(AIF)及基质金属蛋白酶(MMPs)表达水平;Western Blot法检测沉默信息调节因子1(SIRT1)、核因子-κB p65(NF-κB p65)、Bcl-2、Bax蛋白表达情况。结果:麻花艽6个成分最佳给药浓度:龙胆苦苷为150μmol/L、马钱苷酸为150μmol/L、獐牙菜苦苷为30μmol/L、獐牙菜苷为150μmol/L、异荭草素为200μmol/L、异牡荆素为200μmol/L。与空白对照组比较,龙胆苦苷的促细胞凋亡作用最强(P<0.01),马钱苷酸、獐牙菜苦苷、异荭草素次之(P<0.01),獐牙菜苷、异牡荆素组最次(P<0.05);各给药组能显著增加RA-FLS中AIF表达(P<0.01,P<0.05),显著降低RA-FLS中MMPs表达(P<0.01,P<0.05);各给药组能显著降低RA-FLS中NF-κB p65、Bcl-2蛋白表达(P<0.05),显著增加SIRT1、Bax蛋白表达(P<0.05)。结论:麻花艽6个成分具有一定的抑制RA-FLS增殖活性和促凋亡作用,并能调控AIF和MMPs的表达,其潜在机制可能是通过抑制NF-κB信号通路炎症因子的释放和促进线粒体凋亡通路中诱导凋亡因子、促凋亡蛋白水平来促进RA-FLS凋亡。

Objective:To investigate the effects and mechanism of six components of Gentiana straminea Maxim.on rheumatoid arthritis fibroblast-like synoviocytes(RA-FLS).Methods:The optimal concentrations of Gentiana straminea Maxim.were selected by CCK-8 method.Flow cytometry(FCM)was used to determine the expression levels of pro-inflammatory factors AIF and MMPs were detected by ELISA.The expressions of SIRT1,NF-κB p65,Bcl-2 and Bax were detected by Western Blot.Results:Optimal concentrations of six components of Gentiana straminea Maxim.:gentimacroside was 150μmol/L,loganic acid was 150μmol/L,swertiamarin was 30μmol/L,sweroside was 150μmol/L,isoorientin was 200μmol/L,isovitexin was 200μmol/L.Compared with the blank control group,the apoptotic effect of gentimacroside was the strongest(P<0.01),followed by loganic acid,swertiamarin,isoorientin(P<0.01),sweroside and isovitexin were the weakest(P<0.05),all the treatment groups significantly increased the expression of AIF(P<0.01,P<0.05),and significantly decreased the expression of MMPs in RA-FLS(P<0.01,P<0.05),all the treatment groups could significantly reduce the expression of NF-xB p65 and Bcl-2(P<0.05)and increase the expression of SIRT1 and Bax(P<0.05).Conclusion:The six components of Gentiana straminea Maxim.can inhibit the proliferation activity of RA-FLS and promote apoptosis,and regulate the expression of AIF and MMPs.The potential mechanism may be to promote the apoptosis of RA-FLS by inhibiting the release of inflammatory factors in NF-κB signaling pathway and promoting the levels of apoptosis-inducing factors and pro-apoptotic proteins in mitochondrial apoptosis pathway.