丹参酮ⅡA通过抑制HDAC3影响巨噬细胞极化的作用研究

Effect of Tanshinone ⅡA on macrophage polarization by inhibiting HDAC3

目的探讨丹参酮ⅡA(TanⅡA)通过抑制组蛋白去乙酰化酶3(HDAC3)影响巨噬细胞极化的作用。方法应用中药系统药理学数据库与分析平台(TCMSP)筛选TanⅡA药效靶点和动脉粥样硬化作用靶点,并将二者的交集基因进行KEGG通路分析,采用Cytoscape 3.7.1软件对主要交集基因-信号通路进行可视化分析。选用THP-1单核细胞株经佛波醇12-十四酸酯13-乙酸酯(PMA)诱导为贴壁的巨噬细胞后,分为M0组、氧化型低密度脂蛋白(ox-LDL)组、TanⅡA+ox-LDL组及ox-LDL+HDAC3 siRNA组。采用流式细胞术、免疫荧光实验及qRT-PCR技术检测TanⅡA对ox-LDL诱导的巨噬细胞极化方向的改变及HDAC3 mRNA表达水平的变化。结果生物信息学分析得到TanⅡA药效靶点和动脉粥样硬化作用靶点有23个交集基因,选取排序靠前的20条信号通路进行可视化分析,发现主要富集于动脉粥样硬化、流体剪切力和TNF信号通路等。其中,JUN、FOS、RELA、NFKBIA四个交集基因参与巨噬细胞极化的信号通路调控。流式细胞术、免疫荧光实验及qRT-PCR结果显示ox-LDL可诱导M0巨噬细胞CCR7和CCL2表达显著升高;与ox-LDL组相比,TanⅡA预处理的M0巨噬细胞经ox-LDL诱导后HDAC3、CCR7及CCL2 mRNA表达均下降,和ox-LDL+HDAC3 siRNA组结果一致。结论TanⅡA能有效地阻止ox-LDL诱导巨噬细胞往M1方向极化,其机制可能是通过调控巨噬细胞HDAC3表达介导的。

Aim To explore the effect of Tanshinone ⅡA(Tan ⅡA)on macrophage polarization by inhibiting histone deacetylase 3(HDAC3).Methods Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP)was applied to screen drug targets of Tan ⅡA and therapeutic targets of atherosclerosis(As).The intersection genes were enriched and visualized by KEGG pathway analysis and Cytoscape 3.7.1 software,respectively.THP-1 monocytes were induced as adherent macrophages by phorbol 12-myristate-13-acetate(PMA),and divided into four groups:M0 group,oxidized low density lipoprotein(ox-LDL)group,Tan ⅡA+ox-LDL group and ox-LDL+HDAC3 siRNA group.Flow cytometry,immunofluorescence and qRT-PCR were used to detect the change of polarization and expression of HDAC3 mRNA in macrophage.Results Twenty-three intersection genes were obtained of Tan ⅡA drug targets and As therapeutic targets by Bioinformatics analysis.The top twenty signal pathways were selected for visual analysis and displayed that these genes concentrated in signaling pathways such as atherosclerosis,fluid shear force and TNF signaling pathway,among which JUN,FOS,RELA and NFKBIA,were involved in the regulation of macrophage polarization.Results of Flow cytometry,immuno fluorescence and qRT-PCR showed that ox-LDL could significantly increased the expression of CCR7 and CCL2 in M0 macrophages.Compared with ox-LDL group,HDAC3,CCR7 and CCL2 expression decreased in Tan ⅡA+ox-LDL group,consistent with ox-LDL+HDAC3 siRNA group.Conclusion Tan ⅡA could effectively prevent ox-LDL-induced macrophage polarization toward M1 direction,the mecha-nism may be mediated by regulating HDAC3 expression.