雷公藤甲素对RA患者T-bet/GATA3及CXCL10/CXCR3表达的影响

Effect of triptolide on the expression of T-bet/GATA3 and CXCL10/CXCR3 in rheumatoid arthritis patients

关键词:  雷公藤甲素  类风湿关节炎  T-bet  GATA3  趋化因子CXCL10  趋化因子受体CXCR3

Key words:  triptolide  rheumatoid arthritis  T-bet  GATA3  CXCL10  CXCR3

摘要:

[目的] 考察雷公藤甲素（TP）对类风湿关节炎（RA）患者外周血单个核细胞（PBMC）中转录因子T-bet/GATA3和趋化因子（CXCL）10及CXCL受体3（CXCR3）表达的影响。[方法] 使用L929细胞考察TP的细胞毒性，确定实验药物浓度。CCK-8法检测TP对RA患者PBMC细胞增殖活性的影响，流式细胞仪分析TP对PBMC中Th细胞亚群比例及CXCR3受体表达调节，Luminex技术检测RA患者PBMC分泌干扰素-γ（IFN-γ）、白细胞介素（IL）-17/IL-17A、肿瘤坏死因子-α（TNF-α）、IL-4、IL-6、IL-10及CXCL10的表达水平。实时荧光定量聚合酶链反应（RT-qPCR）法分析T-bet、GATA3、CXCL10及CXCR3的mRNA表达水平。[结果] 当TP浓度<25 nmol/L，培养时间为48 h时，TP对L929没有显著的细胞毒性（P>0.05）。在此浓度范围内，当TP浓度为5 nmol/L时即表现出对PBMC细胞增殖具有显著抑制作用（P<0.05）。Th、Th1细胞亚群比例以及CXCR3受体表达均受到TP抑制（P<0.05），但对Th2细胞亚群比例没有显著调节作用（P>0.05）。抗炎因子IL-4、IL-10，促炎因子IFN-γ、IL-6、TNF-α、IL-17/IL-17A，以及CXCL10表达均显著降低（P<0.05）。RT-qPCR结果显示，仅CXCL10表达显著降低，T-bet、GATA3以及CXCR3表达无显著变化（P>0.05）。[结论] TP对Th1细胞增殖及其相关细胞因子具有抑制作用，并且能显著降低CXCL10及CXCR3的表达，但未观察到对T-bet、GATA3表达的调节作用。

Abstract:

[Objective] To investigate the effects of triptolide(TP) on the expression of T-bet/GATA3, CXCL10 and CXCR3 in rheumatoid arthritis (RA) patients.[Methods] L929 cells were used to investigate the cytotoxicity of TP and determine the concentration of experimental drugs. The cell toxity of TP and the proliferation of PBMC were analyzed by CCK-8. The subpopulaions of Th, Th1 and Th2 from RA patients' peripheral mononuclear cells (PBMC) together with the expression of CXCR3 receptor were detected by flow cytometry. The expression level of IFN-γ, IL-17/IL-17A, TNF-α, IL-4, IL-6, IL-10 and CXCL10 secreted by PBMC were demonstrated by Luminex. mRNA expression of T-bet, GATA3, CXCL10 and CXCR3 were analyzed by RT-qPCR.[Results] When the concentration was less than 25 nmol/L and cultured for 48 h, TP had no significant cytotoxicity to L929. Within this concentration range, when the concentration of TP was only 5 nmol/L, it showed a significant effect on inhibition of PBMC proliferation(P<0.05). The subpopulations of Th, Th1 and CXCR3 expression were inhibited by TP with dose dependent manner (P<0.05), except Th2. The ratio of Th1/Th2 was lower after TP treatment. The expression levels of IL-4, IL-10 and IFN-γ, IL-6, TNF-α, IL-17/IL-17A, CXCL10 were all reduced (P<0.05). Additionally, only CXCL10 expression was decreased (P<0.05), while the expression levels of T-bet, GATA3 and CXCR3 had no significant difference(P>0.05).[Conclusion] TP might inhibit RA immune reaction through supressing mutiple cytokines from Th1 and Th2 cells with a non-spicific way, however TP could down regulate CXCL10 and CXCR3 expression significantly except T-bet or GATA3.