蠲痹强骨方含药血清通过FGFR1信号诱导人类风湿关节炎滑膜成纤维细胞凋亡的机制研究

Mechanism of apoptosis of synovial fibroblasts in human rheumatoid arthritis induced by FGFR1 signaling from medicated serum of Juan Bi Qiang Gu Formula

目的基于成纤维细胞生长因子受体1信号通路探讨蠲痹强骨方含药血清对人类风湿关节炎滑膜成纤维细胞增殖和凋亡的影响及其作用机制。方法以不同浓度的蠲痹强骨方含药血清处理人类风湿关节炎滑膜成纤维细胞(RA-FLS),通过CCK8法检测细胞增殖情况,筛选蠲痹强骨方抑制增殖的浓度。利用LPS诱导RA-FLS炎症反应,加入蠲痹强骨方含药血清干预,通过流式细胞仪检测蠲痹强骨方对RA-FLS细胞的凋亡情况。RT-PCR检测mRNA相对表达水平;Western blotting检测蛋白相对表达水平。结果通过CCK8实验发现,不同浓度的蠲痹强骨方含药血清48h和72h可以显著抑制RA-FLS细胞的增殖。流式细胞分析显示,在LPS诱导的炎症环境下,蠲痹强骨方含药血清可以促进RA-FLS细胞的早期和晚期凋亡。RT-PCR结果显示,蠲痹强骨方可以下调LPS诱导下RA-FLS细胞中抗凋亡基因AKT和Bcl-2的mRNA表达。Western blot结果表明,蠲痹强骨方可以抑制LPS诱导下RA-FLS细胞中FGFR1的磷酸化水平和抗凋亡蛋白Bcl-2的表达,促进促凋亡蛋白BAX的表达。结论蠲痹强骨方含药血清对LPS诱导RA-FLS细胞增殖有显著的抑制作用,并可促使其凋亡,其机制可能与抑制FGFR1信号通路转导有关。

Objective This study aims to explore the effects of Juan Bi Qiang Gu Formula-containing serum on the proliferation and apoptosis of human rheumatoid arthritis synovial fibroblasts(RA-FLS)and its mechanism of action,focusing on the fibroblast growth factor receptor 1 signaling pathway.Methods Human RA-FLS were treated with various concentrations of Juan Bi Qiang Gu Formula-containing serum.Cell proliferation was assessed using the CCK8 assay to determine the concentration at which the formula inhibits proliferation.LPS was used to induce an inflammatory response in RA-FLS,followed by intervention with the formula-containing serum.Flow cytometry was utilized to examine the apoptosis of RA-FLS cells.RT-PCR was conducted to measure the relative mRNA expression levels,and Western blotting was employed to analyze the relative protein expression levels.Results The CCK8 assay revealed that the Juan Bi Qiang Gu Formula-containing serum significantly inhibited the proliferation of RA-FLS cells at 48h and 72h across various concentrations.Flow cytometry analysis indicated that,under LPS-induced inflammatory conditions,the formula-containing serum promoted both early and late apoptosis of RA-FLS cells.RT-PCR results showed that the formula downregulated the mRNA expression of anti-apoptotic genes AKT and Bcl-2 in RA-FLS cells induced by LPS.Western blot results demonstrated that the formula suppressed the phosphorylation levels of FGFR1 and the expression of the anti-apoptotic protein Bcl-2,while promoting the expression of the pro-apoptotic protein BAX in RA-FLS cells under LPS induction.Conclusion The Juan Bi Qiang Gu Formula-containing serum significantly inhibits the proliferation of LPS-induced RA-FLS cells and induces their apoptosis.The mechanism may be associated with the suppression of FGFR1 signaling pathway transmission.