来氟米特及其活性代谢物对关节炎模型大鼠TNF-α分泌及mRNA表达的影响

目的观察免疫抑制剂来氟米特(leflunomide,LEF)及其活性代谢产物A771726(A77)对佐剂性关节炎(AA)模型大鼠TNF-α分泌活性及mRNA表达的影响。方法TNF-α活性及mRNA表达用ELISA法和RT-PCR方法。结果 AA大鼠腹腔巨噬细胞(PMφ)呈高度活化状态,TNF-α分泌水平明显升高。LEF明显抑制LPS诱导的大鼠PMφ TNF-α释放且呈剂量依赖关系。体外用A77后TNF-α mRNA表达量降低。结论LEF和A77对AA模型大鼠TNF-α分泌活性及mRNA表达有抑制作用,此可能为其抗炎和免疫抑制作用的机制之一。

EFFECTS OF LEFLUNOMIDE AND ITS ACTIVE METABOLITE ON THE PRODUCTION AND mRNA EXPRESSION OF TNF-α IN PERITONEAL MACROPHAGES AND SYNOVIAL CELLS WITH ADJUVANT ARTHRITIS IN RATS

AIM: To observe the effects of leflunomide (LEF), an isoxazole immunomodulatory agent and its active metabolite, A771726, on the production and mRNA expression of TNF-alpha in peritoneal macrophages and synovial cells with adjuvant arthritis rats and to further investigate the immunosuppression effects of leflunomide and its mechanisms. METHODS: ELISA methods were used for assaying the levels of TNF-alpha. The RT-PCR methods were used for measuring the expression of TNF-alpha. RESULTS: The production of TNF-alpha was increased in the supernatant of PM psi in adjuvant arthritis (AA) model rat. LEF (5, 10, 25 mg.kg-1, ig) was shown to inhibit the release of TNF-alpha from peritoneal macrophages induced by LPS and the inhibitory effects were in a dose-effect relevance manner. A parallel investigation of cytokine mRNA expression was undertaken using semi-quantitative reverse transcribed polymerase chain reaction (RT-PCR) to follow the kinetics of cytokine appearance in PM psi and synovial membrane tissue cells obtained from AA/normal rats treated with A771726. The results of RT-PCR from macrophages and synovial membrane tissue cells of AA rats at the peak of inflammatory phase showed that TNF-alpha mRNA expression levels were higher than those of normal rats, while the expression of TNF-alpha mRNA was reduced by treating with A771726 in vitro. On the other hand, the TNF-alpha mRNA expression showed kinetics very similar to those obtained by ELISA technique which measured protein expression. CONCLUSION: Leflunomide and its active metabolite, A771726, was found to inhibit the production and mRNA expression of TNF-alpha in peritoneal macrophages and synovial cells with adjuvant arthritis rats model. It might be involved in the mechanisms of its anti-inflammation and immunosupression.