牛蒡甙元在人类风湿性关节炎成纤维样滑膜细胞生长与凋亡中的影响

The effects of arctigenin on human rheumatoid arthritis fibroblast-like synoviocytes

目的人类风湿性关节炎成纤维样滑膜细胞(RAFLS)已被证实在类风湿性关节炎发生和发展过程中具有至关重要的作用。本实验目的在于研究牛蒡甙元对RAFLS增殖和凋亡的影响。方法 RAFLS经过牛蒡甙元处理后,通过噻唑蓝和膜联蛋白V/碘化丙啶分析细胞的活性和凋亡,用免疫印迹分析检测细胞凋亡相关基因的变化情况。结果 RAFLS在经过10、20、30μM牛蒡甙(RAFLS)已被证实在类风湿性关节炎发生和发展过程中具有至关重要的作用。本实验元处理后细胞活性分别降低了23%、30%、38%,半数抑制浓度为38μM;同时诱导细胞凋亡分别为9%、15%、21%,凋亡原因为线粒体膜电位减低,细胞色素C释放至胞质,促凋亡蛋白Bax上调,以及抗凋亡蛋白Bcl-2下调。在经过牛蒡甙元处理的RAFLS中,线粒体通路的激活诱导了胱解酶9、胱解酶3和ADP核糖聚合酶(PARP)的分解。此外,牛蒡甙元还能抑制P65的核转位、减少IκBα的降解、降低Akt的磷酸化。结论研究结果显示,牛蒡甙元在RAFLS中通过调节NF-κB和Akt信号通路,能够抑制细胞增殖,并诱导线粒体凋亡。

Objective Rheumatoid arthritis fibroblast-like synoviocytes（ RAFLSs） play a key role in the initiation and progression of RA,which are resistant to proliferate and apoptosis in an anchorage-independent way.The effects of arctigenin on the proliferation and apoptosis of RAFLSs were explored.Me thods RAFLSs was treated for 48 h by arctigenin（ 0-160μM）.Cell viability and apoptosis were assessed by 3-（ 4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazoliumbromide assay and annexin V /propidium iodide staining.Western blot analysis was performed to detect the changes of apoptosis-related genes.Re sults Arctigenin decreased cell viability by 23,30,and 38% at the dose of 10,20 and 30 μM,respectively.The half maximal inhibitory concentration（ IC50） of arctignein on RAFLSs was about 38 μM.Moreover,9,15,and 21% of RAFLSs are induced apoptosis by10,20 and 30 μMof arctigenin.The apoptotic response was due to the loss of mitochondrial membrane potential,coupled with the release of cytochrome c into cytoplasm,the down-regulation of antiapoptotic protein,B cell lymphoma 2（ Bcl-2）,and up-regulation of pro-apoptotic protein,Bax.The activation of mitochondrial pathway in arctigenin-treated RAFLSs induced the cleavage of caspase-9,caspase-3 and poly（ ADP-ribose） polymerase（ PARP）.Additionally,arctigenin inhibited the nuclear translocation of p65,decreased the degradation of inhibitor of kappa B alpha（ IκBα）,and attenuated the phosphorylation of Akt.Conlusions Our results reveal that arctigenin inhibits cell proliferation and induces mitochondrial apoptosis of RAFLSs,which is associated with the modulation of NF-κB and Akt signaling pathways.