雷公藤多苷抗巨噬细胞炎症及对TLR4/NF-κB调控炎症作用的研究

Study on effects of Tripterygium wilfordii polycoride in resisting macrophage inflammation and regulating inflammation via TLR4/NF-κB

为研究雷公藤多苷(Tripterygium wilfordii polycoride,TWP)对LPS诱导巨噬细胞炎症反应时,对炎症因子TNF-α,IL-1β的抑制作用,对TLR4/NF-κB调控炎症作用的干预研究,同时观察其对TLR4/NF-κB调控炎症作用的影响。采用噻唑蓝(MTT)法检测受试药物TWP、地塞米松(DXM)及硫唑嘌呤(AZA)对细胞生长的影响,选择合适的药物浓度。通过LPS诱导小鼠RAW264.7细胞株产生炎症反应,ELISA法检测TNF-α及IL-1β的释放,Westen blotting法检测TLR4及NF-κB p65蛋白表达水平,荧光定量PCR(RT-PCR)测定TLR4及NF-κB的表达。实验结果显示TWP剂量依赖性抑制巨噬细胞炎性细胞因子TNF-α,IL-1β的释放,TWP各剂量组作用均逊于DXM,但TWP高剂量组与AZA组作用比较,对TNF-α作用其优于AZA组;对IL-1β作用其相当于AZA组。对TLR4/NF-κB调控炎症作用的影响,TWP剂量依赖性下调TLR4及NF-κB p65的表达,其作用相当于或优于DXM及AZA。最终得出结论:该项研究显示TWP具有抑制TNF-α及IL-1β的作用,下调TLR4,NF-κB p65受体的表达是其作用的机制之一。

To investigate the effect of Tripterygium wilfordii polycoride （TWP） on LPS-induced macrophage inflammatory response, particularly the inhibitory effect on inflammatory factors TNF-α and IL-1β and the regulatory effect on inflammation via TLR4/NF-κB. The MTT method was adopted to test the effects of tested drugs, TWP, dexamethasone （DXM） and azathioprine （AZA） on cell growth to define the appropriate concentration. LPS was used to induce the inflammatory reaction in mouse RAW264. 7 cell lines. The Elisa kit was adopted to test the release level of TNF-α and IL-1. The Western blotting was applied to test the protein expressions of TNF-α and IL-1β. The RT-PCR was adopted to test the expressions of TLR4 and NF-κB. According to the results, TWP could inhibit the release of macrophage inflammatory factors TNF-α and IL-1β in a dose dependent manner. All of TWP groups showed a weaker efficacy than that of the DXM group. But the TWP high dose group revealed a better effect on TNF-α and equal effect on IL-1β compared with the AZA group. TWP show an equal or better effect in down-regulating TLR4 and NF-κB p65 expressions in a dose dependent manner than DXM and AZA. In conclusion, TWP could inhibit TLR4 and NF-κB p65, which may be related to the down-regulation of TLR4 and NF-κB p65 receptor expressions.