下肢动脉粥样硬化斑块内miRNA-199a-3p对外周血单核细胞的影响

Effects of miRNA-199a-3p in atherosclerotic plaque on peripheral blood mononuclear cells

目的探讨下肢动脉粥样硬化斑块内miRNA-199a-3p对外周血单核细胞的影响。方法选取37例下肢动脉粥样硬化闭塞症(AS)患者外周血为AS组。另外选取同期37名健康体检者外周血为对照组。新鲜血管组织取自1例下肢AS患者截肢远端血管为实验组,以其截肢近端血管为对照。实时荧光定量PCR检测新鲜血管组织,分离得到的外周血中单核细胞和微泡中miRNA-199a-3p表达水平。以人单核细胞TTHP-1为载体,miRNA-199a-3p mimics组和miRNA-199a-3p inhibitor组分别转染miRNA-199a-3p mimics和miRNA-199a-3p inhibitor,同时设置空白对照组。实时荧光定量PCR检测各组miRNA-199a-3p、趋化因子单核细胞趋化蛋白(MCP)-1、调解活化正常T细胞表达和分泌的趋化因子(RANTES)和Fractalkine表达水平。结果下肢动脉粥样硬化斑块内miRNA-199a-3p表达水平显著高于正常组织(P<0.05)。AS组外周血单核细胞和微泡中miRNA-199a-3p表达水平均显著高于对照组(P<0.05)。与空白对照组相比,miRNA-199a-3p mimics组miRNA-199a-3p表达水平显著上升(P<0.05),miRNA-199a-3p inhibitor组miRNA-199a-3p表达水平显著下降(P<0.05)。与空白对照组相比,miRNA-199a-3p mimics组MCP-1、RANTES和Fractalkine的mRNA表达水平显著降低(P<0.05),miRNA-199a-3p inhibitor组MCP-1、RANTES和Fractalkine的mRNA表达水平显著上升(P<0.05)。结论动脉粥样硬化斑块内巨噬细胞高表达miRNA-199a-3p,并以微泡形式分泌到外周血中,抑制单核细胞中MCP-1、RANTES和Fractalkine等趋化因子的表达,是一种动脉粥样硬化保护因素。

Objective To investigate the effect of miRNA-199a-3p in atherosclerotic（ AS） plaque on peripheral blood mononuclear cells. Methods Twenty patients with arterial atherosclerosis were treated as AS group. In addition,37 healthy subjects were selected as the control group. Fresh blood vessels were taken from 1 patient with AS who had been treated. The expression of miRNA-199a-3p in blood vessels,peripheral blood mononuclear cells and microbubbles were detected by real-time fluorescent quantitative PCR. miRNA-199a-3p mimics group and miRNA-199a-3p inhibitor group were transfected with miRNA-199a-3p mimics and miRNA-199a-3p inhibitor,and the blank control group was set up with human monocyte TTHP-1 as the vector. The levels of miRNA-199a-3p,chemokine monocyte chemoattractant protein-1（ MCP-1）,RANTES and Fractalkine were detected by real-time fluorescent quantitative PCR. Results The expression of miRNA-199a-3p in AS plaques was significantly higher than that in normal tissues（ P〈0.05）. The expression of miRNA-199a-3p in peripheral blood mononuclear cells and microbubbles of coronary heart disease group were significantly higher than those of control group（ P〈0. 05）. Compared with that of blank control group,the expression level of miRNA-199a-3p in miRNA-199a-3p mimics group was significantly increased（ P〈0.05）,miRNA-199a-3p expression level in miRNA-199a-3p inhibitor group was significantly decreased（ P〈0.05）. Compared with those of blank control group,the mRNA expressions of MCP-1,RANTES and Fractalkine in miRNA-199a-3p mimics group were significantly decreased（ P〈0.05）,mRNA expression of MCP-1,RANTES and Fractalkine in miRNA-199a-3p inhibitor group were significantly increased（ P〈0.05）. Conclusions miRNA-199a-3p is highly expressed in macrophages of AS plaques and secreted into peripheral blood in microbubbles to inhibit the expressions of chemokines such as MCP-1,RANTES and Fractalkine in monocytes. miRNA-199a-3p is a protective factor for atherosclerosis.