Wen Jian-Ting, Liu Jian, Wang Xin, Wang Jie
Anhui University of Traditional Chinese Medicine Hefei 230038, China.
Department of Rheumatology, First Affiliated Hospital of Anhui University of Traditional Chinese Medicine Hefei 230031, China.
Zhongguo Zhong Yao Za Zhi. 2021 Jan;46(2):436-443. doi: 10.19540/j.cnki.cjcmm.20200915.402.
The aim of this paper was to observe the effect of Xinfeng Capsules(XFC)-containing serum on the apoptosis and inflammation of fibroblast-like synoviocytes(FLS) in rheumatoid arthritis(RA) induced by tumor necrosis factor-α(TNF-α), so as to investigate the mechanism of XFC in the treatment of RA. RA-FLS immortalized cell line was established, and XFC drug-containing serum was prepared. CCK-8, ELISA, RT-qPCR, immunofluorescence and TUNEL were used to observe the effect of XFC-containing serum on RA-FLS apoptosis and inflammatory indexes. CCK-8 results showed that the optimal concentration and time of TNF-α on RA-FLS were 10 ng·mL(-1) and 48 h, respectively; and the optimal concentration and time of XFC on RA-FLS were 6.48 mg·g(-1) and 72 h, respectively. The results of ELISA showed that compared with RA-FLS group, the expressions of TNF-α, IL-1β, IL-6, IL-8 in TNF-α+RA-FLS group were significantly increased, while the expressions of IL-4 and IL-10 were significantly decreased(P<0.01); after intervention with XFC-containing serum, the expressions of TNF-α, IL-1β, IL-6, IL-8 were significantly decreased, whereas the expressions of IL-4 and IL-10 were significantly increased(P<0.01). The results of RT-qPCR showed that compared with RA-FLS group, the mRNA expressions of Fas, FasL, caspase-3, caspase-8, Bax, Bcl-X1 in TNF-α+RA-FLS group were significantly decreased, while the mRNA expression of Bcl-2 was significantly increased(P<0.001); after intervention with XFC-containing serum, the mRNA expressions of Fas, FasL, caspase-3, caspase-8, Bax, Bcl-X1 were significantly increased, whereas the mRNA expression of Bcl-2 was significantly decreased(P<0.01). The results of immunofluorescence showed that compared with RA-FLS group, the protein expressions of caspase-3 and Bax in TNF-α+RA-FLS group was significantly lower than those in RA-FLS group(P<0.05); after intervention with XFC-containing serum, the protein expressions of caspase-3 and Bax were significantly increased, whereas the protein expression of Bcl-2 was significantly decreased(P<0.05). TUNEL results showed that compared with RA-FLS group, the apoptosis of TNF-α+RA-FLS group was decreased(P<0.05); after intervention with XFC-containing serum, the apoptosis was significantly increased(P<0.05). One of the mechanisms of XFC in the treatment of RA is to promote the apoptosis of RA-FLS and inhibit its inflammatory reaction.
本文旨在观察含新风胶囊（XFC）血清对肿瘤坏死因子-α（TNF-α）诱导的类风湿关节炎（RA）成纤维样滑膜细胞（FLS）凋亡及炎症的影响，以探讨XFC治疗RA的作用机制。建立RA-FLS永生化细胞系，并制备含XFC药物血清。采用CCK-8、ELISA、RT-qPCR、免疫荧光及TUNEL法观察含XFC血清对RA-FLS凋亡及炎症指标的影响。CCK-8结果显示，TNF-α作用于RA-FLS的最佳浓度和时间分别为10 ng·mL⁻¹和48 h；XFC作用于RA-FLS的最佳浓度和时间分别为6.48 mg·g⁻¹和72 h。ELISA结果显示，与RA-FLS组相比，TNF-α+RA-FLS组TNF-α、IL-1β、IL-6、IL-8表达显著升高，而IL-4和IL-10表达显著降低（P<0.01）；经含XFC血清干预后，TNF-α、IL-1β、IL-6、IL-8表达显著降低，而IL-4和IL-10表达显著升高（P<0.01）。RT-qPCR结果显示，与RA-FLS组相比，TNF-α+RA-FLS组Fas、FasL、caspase-3、caspase-8、Bax、Bcl-X1的mRNA表达显著降低，而Bcl-2的mRNA表达显著升高（P<0.001）；经含XFC血清干预后，Fas、FasL、caspase-3、caspase-8、Bax、Bcl-X1的mRNA表达显著升高，而Bcl-2的mRNA表达显著降低（P<0.01）。免疫荧光结果显示，与RA-FLS组相比，TNF-α+RA-FLS组caspase-3和Bax蛋白表达显著低于RA-FLS组（P<0.05）；经含XFC血清干预后，caspase-3和Bax蛋白表达显著升高，而Bcl-2蛋白表达显著降低（P<0.05）。TUNEL结果显示，与RA-FLS组相比，TNF-α+RA-FLS组细胞凋亡减少（P<0.05）；经含XFC血清干预后，细胞凋亡显著增加（P<0.05）。XFC治疗RA的机制之一是促进RA-FLS凋亡并抑制其炎症反应。
