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目的:研究青藤碱和乌头原碱单独或配伍使用对活化的T淋巴细胞RANKL表达和两药合用对破骨细胞生成的影响。方法:采用anti-CD3激活小鼠CD4~+T细胞,用流式细胞仪检测青藤碱、乌头原碱对小鼠辅助性T细胞RANKL表达的影响;同时用TRAP染色法观察两药合用对RANKL诱导破骨细胞生成的影响。结果:0.25~1mmol/L青藤碱能够剂量依赖性地抑制小鼠CD4~+T细胞RANKL表达;0.25、0.5mmol/L乌头原碱也能显著地降低CD4~+T细胞RANKL表达,而另一毒性较大的活性成分苯甲酰乌头原碱只在高剂量0.5mmol/L时才能下调RANKL表达。乌头原碱与低浓度青藤碱合用后能使CD4~+T细胞RANKL表达进一步降低,而苯甲酰乌头原碱与低浓度青藤碱合用后对RANKL表达无影响。研究发现,青藤碱与乌头原碱合用后对RANKL诱导的RAW264.7细胞分化成破骨细胞的抑制作用较两药单独使用明显增强。结论:青风藤和附子活性成分均可能通过下调活化的T细胞RANKL表达间接抑制RANKL诱导的破骨细胞生成和骨吸收,改善类风湿性关节炎引起的骨破坏;附子久煎后的无毒活性成分乌头原碱与青藤碱配伍使用后对T细胞RANKL表达和RANKL相关破骨细胞生成的作用均优于单用,提示中医临床使用附子时久煎不仅能够减毒,还可能更有利于配伍其它药物治疗类风湿性关节炎骨破坏。
Objective: To investigate the effect of sinomenine and aconitine alone or in combination on the expression of RANKL in activated T lymphocytes and the effect of two drugs on osteoclastogenesis. Methods: CD4 + T cells were activated by anti-CD3 and the effects of sinomenine and aconitine on the expression of RANKL in mouse T helper cells were detected by flow cytometry. Meanwhile, TRAP staining was used to observe the effects of both drugs On RANKL-induced osteoclastogenesis. Results: Sinomenine 0.25 ~ 1 mmol / L inhibited the expression of RANKL in CD4 ~ + T cells in a dose-dependent manner. 0.25 and 0.5 mmol / L aconitine also significantly reduced the RANKL expression in CD4 ~ + T cells Another more toxic active ingredient benzoyl aconitine alkali only at high doses of 0.5mmol / L can be reduced RANKL expression. Aconitum alkaloids combined with low concentrations of sinomenine can make RANKL expression of CD4 ~ + T cells further reduced, while the combination of benzoyl aconitine and low concentrations of sinomenine had no effect on RANKL expression. The study found that sinomenine combined with aconitine alkali RANKL induced RAW264.7 cells differentiated into osteoclasts compared with the two drugs alone significantly enhanced. CONCLUSION: Both the active ingredients of Gynura divaricata and A. officinalis may indirectly inhibit RANKL-induced osteoclastogenesis and bone resorption by decreasing the expression of RANKL in activated T cells and improve the bone destruction caused by rheumatoid arthritis. Components of aconitine and sinomenine compatibility with the use of T cells after RANKL expression and RANKL related osteoclast formation are better than single use, suggesting that clinical use of aconite in the long time Jianfu not only can be attenuated, but also may be more conducive to Compatibility with other drugs to treat rheumatoid arthritis bone destruction.