硫喷妥钠通过MyD88/NF-κB信号通路抑制巨噬细胞在脂多糖诱导下的免疫反应  

Thiopental inhibits macrophage immune response induced by lipopolysaccharide through MyD88/NF-κB signaling pathway

作　　者：陈刚[1] 何爱萍[1] CHEN Gang;HE Ai-Ping(Department of Anesthesiology,the First Affiliated Hospital of Xi′an Medical College,Xi′an 710077,China)

机构地区：[1]西安医学院第一附属医院麻醉科,西安710077

出　　处：《中国免疫学杂志》2021年第2期149-154,共6页Chinese Journal of Immunology

摘　　要：目的:研究硫喷妥钠对巨噬细胞在脂多糖(LPS)刺激诱导下的免疫反应,并探讨其作用机制。方法:利用佛波酯(PMA)诱导人单核细胞系THP-1细胞分化为巨噬细胞,使用不同浓度的硫喷妥钠(0、7.5、15、30μmol/L)进行处理后,LPS刺激并活化巨噬细胞;RT-PCR及Western blot检测硫喷妥钠对活化后的巨噬细胞中MyD88 mRNA及蛋白表达的影响;将抑制MyD88表达的si-MyD88转染入巨噬细胞,流式细胞术及ELISA分别检测硫喷妥钠对转染前后的巨噬细胞表面CD80及其对炎症因子IL-6、TNF-α、IL-10表达的影响;Western blot检测硫喷妥钠对巨噬细胞NF-κB信号通路中p65蛋白磷酸化的影响。结果:PMA成功诱导人单核细胞系THP-1细胞分化成为巨噬细胞;流式细胞术及ELISA实验结果显示,硫喷妥钠能够明显抑制LPS刺激活化的THP-1巨噬细胞表面CD80的表达及其对炎症因子IL-6、TNF-α、IL-10的释放;RT-PCR及Western blot实验结果显示,硫喷妥钠能够下调LPS刺激诱导下的THP-1巨噬细胞中MyD88 mRNA及蛋白质的表达;THP-1巨噬细胞转染si-MyD88后,RT-PCR及Western blot结果显示,细胞中MyD88 mRNA及蛋白质的表达均明显下降,流式细胞术及ELISA实验结果显示,LPS刺激诱导下的THP-1巨噬细胞表面CD80的表达及IL-6、TNF-α、IL-10的释放被显著抑制;Western blot实验结果显示,硫喷妥钠能够显著抑制NF-κB信号通路中p65蛋白磷酸化。结论:硫喷妥钠能够抑制巨噬细胞在LPS诱导下的免疫反应,其作用机制可能与降低细胞中MyD88/NF-κB信号通路激活水平有关。Objective:To study the immune response of thiopental sodium to macrophages stimulated by lipopolysaccharide(LPS),and to explore the mechanism.Methods:Human monocyte line THP-1 cells were induced to differentiate into macrophages by PMA.After being treated with different concentrations of thiopental sodium(0,7.5,15,30μmol/L),macrophages were stimulated and activated by LPS;RT-PCR and Western blot were used to detect the mRNA and protein expression of MyD88 in activated macrophages;si-MyD88 was transfected into macrophages.The effects of thiopental on the expression of CD80 and the release of inflammatory factors IL-6,TNF-αand IL-10 of activated macrophages were detected by flow cytometry and ELISA.Western blot was used to detect the effect of thiopental on the phosphorylation of p65 protein in NF-κB signaling pathway.Results:Human monocyte THP-1 cells were successfully induced to differentiate into macrophages by PMA.The results of flow cytometry and ELISA showed that thiopental significantly inhibited the expression of CD80 and the release of IL-6,TNF-α,IL-10 on the surface of LPS stimulated THP-1 macrophages.RT-PCR and Western blot results showed that thiopental sodium could down-regulate the expressions of MyD88 mRNA and protein in THP-1 macrophages stimulated by LPS.After transfection of si-MyD88 into THP-1 macrophages,RT-PCR and Western blot showed that the expressions of MyD88 mRNA and protein in THP-1 macrophages were decreased significantly.Flow cytometry and ELISA showed that the expression of CD80 and the release of IL-6,TNF-αand IL-10 on THP-1 macrophages stimulated by LPS were significantly inhibited.Western blot results showed that thiopental sodium could significantly inhibit the phosphorylation of p65 protein in the NF-κB signaling pathway.Conclusion:Thiopental sodium can inhibit the immune response of macrophages induced by LPS,and its mechanism may be related to the reduction of the activation level of MyD88/NF-κB signaling pathway in cells.

关 键 词：硫喷妥钠 脂多糖 巨噬细胞 MYD88 

分 类 号：R392.5[医药卫生—免疫学]