摘要
目的探究独活寄生汤含药血清对肿瘤坏死因子-α(TNF-α)诱导的类风湿性关节炎(RA)成纤维样滑膜细胞(FLS)增殖、凋亡和炎症的影响及作用机制。方法将MH7A细胞分为正常组(10%空白血清)、模型组(10μg·L^(-1) TNF-α+10%空白血清)、独活寄生汤低(10μg·L^(-1) TNF-α+2.5%含药血清+7.5%空白血清)、中(10μg·L^(-1) TNF-α+5%含药血清+5%空白血清)、高(10μg·L^(-1) TNF-α+10%含药血清)剂量组共5组;MTT法筛选独活寄生汤含药血清给药浓度;EdU染色检测细胞增殖情况;免疫荧光染色检测增殖标志物Ki67表达情况;流式细胞仪检测细胞凋亡率;ELISA法检测细胞中IFN-γ、IL-4、IL-6和IL-10的含量;RT-qPCR和Western blot检测Bax、Bcl-2、caspase-3及TLR4/MyD88/NF-κB信号通路mRNA和蛋白的表达情况。结果与正常组比较,模型组细胞增殖水平明显上升,凋亡水平明显减弱,IFN-γ和IL-6的含量升高,IL-4和IL-10的含量减少,TLR4/MyD88/NF-κB信号通路被激活;经独活寄生汤含药血清低、中、高剂量组干预后,均能有效改善细胞异常增殖情况,增强细胞凋亡,抑制炎性反应和TLR4/MyD88/NF-κB信号通路的激活。结论独活寄生汤含药血清能够抑制TNF-α诱导的RA-FLS异常增殖和促炎因子的分泌,增强细胞凋亡和抗炎水平,其作用机制可能与调控TLR4/MyD88/NF-κB信号通路有关。
Aim To explore the effect of serum containing Duhuo Jisheng Decoction on the proliferation,apoptosis and inflammation of fibroblast-like synoviocytes(FLS)induced by tumor necrosis factor-α(TNF-α)in rheumatoid arthritis(RA)and to reveal the underlying mechanism.Methods The MH7A cells were divided into five groups:normal group(10%blank serum),model group(10μg·L^(-1) TNF-α+10%blank serum),Duhuo Jisheng Decoction low(10μg·L^(-1) TNF-α+2.5%drug-containing serum+7.5%blank serum),medium(10μg·L^(-1) TNF-α+5%drug-containing serum+5%blank serum),high(10μg·L^(-1) TNF-α+10%drug-containing serum)dose group.The concentration of serum containing Duhuo Jisheng Decoction was screened by MTT method.Cell proliferation was detected using EdU staining.The expression of proliferation marker Ki67 was detected using immunofluorescence staining.The apoptosis rate was measured by flow cytometry.The contents of IFN-γ,IL-4,IL-6 and IL-10 in each group were detected by ELISA.The mRNA and protein expression of Bax,Bcl-2,caspase-3 and TLR4/MyD88/NF-κB signaling pathway were detected by RT-qPCR and Western blot.Results Compared with the normal group,the cell proliferation activity of the model group significantly increased,and the level of apoptosis decreased.The content of IFN-γand IL-6 increased,and the content of IL-4 and IL-10 decreased.The TLR4/MyD88/NF-κB signaling pathway was activated.After the intervention of low,medium and high dose groups of serum containing Duhuo Jisheng Decoction,it could effectively improve the abnormal proliferation of cells and enhance apoptosis.At the same time,it inhibited the inflammatory response and the activation of TLR4/MyD88/NF-κB signaling pathway.Conclusions The serum containing Duhuo Jisheng Decoction can inhibit the abnormal proliferation of RA-FLS induced by TNF-αand the secretion of pro-inflammatory factors,and enhance apoptosis and anti-inflammatory levels.The mechanism may be related to the regulation of TLR4/MyD88/NF-κB signaling pathway.
作者
吴婧若
步亚男
岳进茹
陈佳
王桂宇
张育敏
WU Jing-ruo;BU Ya-nan;YUE Jin-ru;CHEN Jia;WANG Gui-yu;ZHANG Yu-min(College of Basic Medical science,Shanxi University of Chinese Medicine,Jinzhong Shanxi 030619,China)
出处
《中国药理学通报》
CAS
CSCD
北大核心
2024年第9期1665-1673,共9页
Chinese Pharmacological Bulletin
基金
国家自然科学基金资助项目(No 82274365)
山西省自然科学基金资助项目(No 20210302123301)
山西省中医药科研基金资助项目(No 2023ZYYA031)
山西中医药大学“青年科学家培育专题”项目(No 2021PY-QN-01)。
