miRNA-153-5p通过靶向TFRC调控巨噬细胞极化在病毒性心肌炎中的作用

miRNA-153-5p regulates macrophage polarization in viral myocarditis by targeting TFRC

目的:探讨miRNA-153-5p调控巨噬细胞极化在病毒性心肌炎(VM)中的作用。方法:建立VM小鼠模型,提取心脏,RT-qPCR检测心肌组织miRNA-153-5p的表达,HE染色观察心脏炎症程度,ELISA检测小鼠血清肌钙蛋白cTnI的水平,流式细胞术检测心脏浸润巨噬细胞表型。体外分离BMDMs诱导为M1/M2型巨噬细胞,过表达和抑制miR-153-5p,RT-qPCR检测M1/M2标志物iNOS、IL-12、TNF-α、Arg1、YM-1、FIZZ1的表达。生物信息学软件预测结合双荧光素酶实验验证miR-153-5p与转铁蛋白受体(TFRC)的靶向关系。结果:miR-153-5p在CVB3感染7 d的VM小鼠心脏组织中表达明显增加(P<0.05)。体内实验中抑制miR-153-5p表达能减轻VM小鼠心脏的病变程度,改善心脏功能(P<0.01)。抑制miR-153-5p表达能够促进小鼠心脏浸润的巨噬细胞向M2极化(P<0.01)。体外实验中,较之M2,miR-153-5p在M1巨噬细胞中表达升高(P<0.01)。过表达miR-153-5p促进巨噬细胞向M1极化,抑制miR-153-5p表达则促进巨噬细胞向M2极化(P<0.01)。TFRC是miR-153-5p的靶基因,抑制miR-153-5p表达可上调TFRC的mRNA和蛋白水平(P<0.01)。结论:miRNA-153-5p通过靶向TFRC调控VM小鼠心脏浸润巨噬细胞的极化。

Objective:To investigate the role of miRNA-153-5p in regulating macrophage polarization in viral myocarditis.Methods:A mouse model of viral myocarditis was established,and from which the hearts were obtained.RT-qPCR was performed to detect the expression of miRNA-153-5p in myocardial tissue,and heart tissues were stained with HE to examine the degree of inflammation.ELISA was applied to measure the serum troponin cTnI level in mice,and flow cytometry was used to detect the phenotype of infiltrating macrophages in the heart.Bone marrow-derived macrophages(BMDMs)were isolated in vitro,and then induced as M1/M2 type macrophages,overexpressed and suppressed miR-153-5p.RT-qPCR was used to determine the expression of M1/M2 markers iNOS,IL-12,TNF-α,Arg1,YM-1,and FIZZ1.Finally,bioinformatics software prediction combined with dual luciferase assay were used to verify the targeting relationship of miR-153-5p with TFRC.Results:miR-153-5p was highly expressed in the heart tissues of mice with viral myocarditis at day 7 of CVB3 infection.Inhibition of miR-153-5p expression in in vivo experiments reduced the extent of lesions and improved cardiac function in viral myocarditis mouse hearts.Inhibition of miR-153-5p expression generated polarization of mouse heart-infiltrating macrophages toward M2.In in vitro experiments,miR-153-5p expression was elevated in M1 macrophages compared to M2.Overexpression of miR-153-5p resulted in macrophage polarization toward M1,whereas inhibition of miR-153-5p expression stimulated macrophage polarization toward M2.The transferrin receptor TFRC functioned a target gene of miR-153-5p,and inhibition of miR-153-5p expression upregulated the mRNA and protein levels of TFRC.Conclusion:miRNA-153-5p can regulate the polarization of cardiac infiltrating macrophages in viral myocarditis mice by targeting TFRC.