摘要
目的:探讨枫杨总黄酮(PHSTF)通过Fas/FasL介导的线粒体凋亡治疗类风湿关节炎(RA)的可能机制。方法:体外培养胶原诱导型关节炎(CIA)-成纤维细胞样滑膜细胞(FLS),CCK-8检测PHSTF对细胞存活率的影响;流式细胞术检测PHSTF对细胞凋亡的影响;Western Blot检测Fas、FasL、Bax、Bcl-2、t-Bid、C-Caspase8、C-Caspase9、C-Caspase3蛋白表达,加入Fas抑制剂KR-33493后进一步检测PHSTF对相关蛋白表达的影响。体内实验采用Ⅱ型胶原诱导法构建CIA大鼠复制RA模型,设立Control组、Model组、PHSTF组、TG组,每组8只,评估大鼠关节炎指数(AI)和足趾肿胀度;ELISA检测血清肿瘤坏死因子α(TNF-α)、白细胞介素(IL)-1β、IL-6含量;HE染色观察膝关节病理学变化;Western Blot检测滑膜组织Fas/FasL介导的线粒体凋亡相关蛋白的表达。结果:体外实验显示,PHSTF能够诱导CIA-FLS细胞凋亡(P<0.01);与Control组比较,PHSTF各浓度组Fas、FasL、Bax、t-Bid、C-Caspase8、C-Caspase9、C-Caspase3蛋白表达均显著增加、抗凋亡蛋白Bcl-2表达显著降低(P<0.05,P<0.01);加入Fas抑制剂KR-33493干扰Fas信号通路后,逆转了Fas信号通路相关蛋白的表达。体内实验显示,与Model组比较,PHSTF、TG组AI评分和足趾肿胀度显著降低(P<0.05),膝关节病理变化得到改善,血清中TNF-α、IL-1β、IL-6含量显著降低(P<0.01);Fas、FasL、Bax、t-Bid、C-Caspase8、C-Caspase9、C-Caspase3蛋白表达均增加、Bcl-2蛋白表达显著降低(P<0.01)。结论:PHSTF能够通过激活Fas/FasL介导的线粒体凋亡,抑制CIA-FLS细胞增殖、促进凋亡,改善CIA大鼠关节炎表型,起到治疗RA的作用。
Objective:To explore the possible mechanism of total flavonoids of Pterocarya Hupehensis Skan(PHSTF)in the treatment of rheumatoid arthritis(RA)through Fas/FasL mediated mitochondrial apoptosis.Methods:Collagen induced arthritis(CIA)-fibroblast synovial cells(FLS)cells were cultured in vitro,and the effect of PHSTF on cell survival rate was detected by CCK-8;The effect of PHSTF on cell apoptosis was detected by flow cytometry;Fas,FasL,Bax,Bcl-2,t-Bid,C-Caspase8,C-Caspase9,C-Caspase3 protein expression was detected by Western Blot.The effect of PHSTF on the expression of related proteins was further tested after adding the Fas inhibitor KR-33493.In vivo experiment using typeⅡcollagen induction method to construct CIA rat replicated RA model.Establish control group,model group,PHSTF group,and TG group with 8 rats in each group.The AI index and toe swelling degree of each group were evaluated;and the TNF-α,IL-1β,IL-6 in serum was detected by ELISA.The pathological changes of the knee joint were observed through HE staining.The expression of mitochondrial apoptosis related proteins mediated by Fas/FasL in synovial tissue was detected through Western Blot.Results:In vitro experiments had shown that PHSTF could significantly induce apoptosis in CIA-FLS cells(P<0.01).Compared with the control group,the expression of Fas,FasL,Bax,t-Bid,C-Caspase8,C-Caspase9,and C-Caspase3 proteins was increased in all concentration groups of PHSTF,while the expression of anti apoptotic protein Bcl-2 was significantly reduced(P<0.05,P<0.01).After adding the Fas inhibitor KR-33493 to interfere with the Fas signaling pathway,it reversed the expression of Fas signaling pathway related proteins.In vivo experiments showed that compared with the model group,the AI score and toe swelling of the PHSTF and TG groups were significantly reduced(P<0.05),the pathological changes of the knee joint were improved,and the the level of TNF-α,IL-1β,IL-6 in serum was significantly reduced(P<0.01).The expression of Fas,FasL,Bax,t-Bid,C-Caspase8,C-Caspas
作者
侯孜明
向阳
吴昊
冯佳
陶柏楠
田梦杰
袁林
HOU Ziming;XIANG Yang;WU Hao;FENG Jia;TAO Bonan;TIAN Mengjie;YUAN Lin(Hubei Provincial Key Laboratory of Occurrence and Intervention of Rheumatic Diseases,Hubei Minzu University,Enshi 445000,China;Department of Medicine,Hubei Minzu University,Enshi 445000,China;Hubei Enshi College,Enshi 445000,China)
出处
《中华中医药杂志》
CAS
CSCD
北大核心
2024年第10期5493-5498,共6页
China Journal of Traditional Chinese Medicine and Pharmacy
基金
国家自然科学基金项目(No.81960776)
湖北民族大学风湿性疾病发生与干预湖北省重点实验室开放基金项目(No.PT022206)。
关键词
枫杨总黄酮
类风湿关节炎
成纤维细胞样滑膜细胞
胶原诱导型关节炎
FAS/FASL
Total flavonoids of Pterocarya Hupehensis Skan(PHSTF)
Rheumatoid arthritis(RA)
Fibroblast synovial cells(FLS)
Collagen induced arthritis(CIA)
Fas/FasL
