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目的探讨断藤益母汤(DYD)含药血清对体外培养的类风湿关节炎(rheumatoid arthritis,RA)患者滑膜成纤维细胞(fibrolast-like synoviocytes,FLS)增殖以及TLR4(Toll like receptor 4)mRNA表达的影响和作用机制。方法体外培养RA-FLS,取第3~5代细胞,分别加入来氟米特和DYD高、中、低剂量家兔含药血清,空白组加入正常家兔血清,MTT法检测24 h、48 h、72 h后各组含药血清对RA-FLS的影响。取RA-FLS,空白组不加脂多糖(LPS),阳性对照组加LPS20μg.mL-1,实验组在加入LPS后分两组,分别加入来氟米特和细胞抑制率最高的DYD含药血清。Realtime-PCR法检测以上4组细胞TLR4 mRNA的表达。结果来氟米特和DYD高剂量组含药血清作用48 h、72 h,二者均能显著抑制RA-FLS增殖(Ρ<0.05);对LPS诱导的TLR4的mRNA表达也有显著抑制作用(Ρ<0.05);来氟米特和DYD药效相当,两者比较差异无统计学意义。结论断藤益母汤高、中剂量含药血清作用48 h、72 h后能显著抑制RA-FLS增殖,且高剂量血清可下调LPS诱导的TLR4mRNA表达,这可能是该方治疗RA的机制之一。
Objective To investigate the effects of DYT-containing serum on the proliferation of fibrolast-like synoviocytes (FLS) and the expression of TLR4 (Toll like receptor 4) in rheumatoid arthritis (RA) Effect of mRNA expression and its mechanism of action. Methods RA-FLS were cultured in vitro. Cells from passage 3 to passage 5 were harvested, and serums from high, medium, and low doses of leflunomide and DYD were respectively added into the rabbits. h, 72 h after the drug-containing serum on RA-FLS. The rats in the experimental group were divided into two groups with LPS supplemented with leflunomide and the highest cytotoxicity of DYD-containing drug serum. Realtime-PCR method was used to detect the expression of TLR4 mRNA in the above four groups of cells. Results Both leflunomide and DYD high-dose groups could significantly inhibit the proliferation of RA-FLS (P <0.05) at 48 h and 72 h, and also inhibit LPS-induced TLR4 mRNA expression (Ρ <0.05). The efficacy of leflunomide and DYD was comparable, with no significant difference between the two. Conclusions Serum of Tengtong Yimu Decoction can significantly inhibit the proliferation of RA-FLS at 48 h and 72 h, and high-dose serum can down-regulate the expression of TLR4 mRNA induced by LPS, which may be the mechanism of RA one.