摘要
目的研究miR-17对冠状动脉疾病(CAD)中血管平滑肌细胞(VSMCs)的潜在作用机制。方法RT-qPCR检测CAD患者血清和VSMCs中miR-17和胰岛素样生长因子1(IGF-1)的mRNA表达,通过CCK-8和克隆形成实验测定VSMCs的增殖情况;通过生物信息学和荧光素酶报告基因检测系统分析miR-17的潜在靶标。结果RT-qPCR结果显示,与对照组相比,CAD患者血清和VSMCs中miR-17的mRNA表达均显著上调(P<0.01)。CCK-8和克隆形成测定结果显示,与对照组相比,miR-17过表达组VSMCs细胞增殖和克隆形成数均显著升高(P<0.01);miR-17低表达组均显著降低(P<0.01)。生物信息学分析结果显示,IGF-1的3′-UTR中具有miR-17的结合位点。荧光素酶报告基因测定结果显示,与野生型IGF-1质粒和miR-17模拟物共转染的VSMCs的荧光素酶活性升高(P<0.001),而转染突变型IGF-1质粒和miR-17模拟物的VSMCs的荧光素酶活性无改变。与对照组相比,过表达miR-17组VSMCs中的IGF-1表达上调(P<0.01);与对照组相比,IGF-1过表达组VSMCs细胞增殖和克隆形成数均显著升高(P<0.05)。结论miR-17通过靶向IGF-1促进VSMCs增殖,表明miR-17可作为CAD的预测生物标志物,IGF-1可能是潜在的治疗靶点。
Objective To investigate the potential mechanism of miR-17 in vascular smooth muscle cells in coronary artery disease(CAD).Methods mRNA expression of miR-17 and insulin growth factor 1(IGF-1)in serum and VSMCs of CAD patients were detected by RT-qPCR.Potential targets of miR-17 were detected by bioinformatics and luciferase reporter assay;CCK-8 and cloning formation assay was performed to measure the proliferation of VSMCs.Results RT-qPCR results showed that compared with those in control group,the miR-17 mRNA expression in VSMCs and serum of CAD patients were significantly upregulated(P<0.01).The results of CCK-8 and clone formation assay showed that compared with those in control group,the number of VSMCs proliferation and cloning formation in the miR-17 overexpression group were significantly increased(P<0.01);those in the miR-17 low expression group were significantly reduced(P<0.01).Bioinformatics analysis showed that the 3’-UTR of IGF-1 had an miR-17 binding site.The luciferase reporter assay showed that the luciferase activity of VSMCs co-transfected with wild-type IGF-1 plasmid and miR-17 mimic was increased(P<0.001).However,the luciferase activity of VSMCs transfected with mutant IGF-1 plasmid and miR-17 mimics remained unchanged.Compared with that in control group,the expression of IGF-1 in VSMCs was upregulated after miR-17 overexpression(P<0.01).And the number of VSMCs proliferation and clone formation in the IGF-1 overexpressiongroup was significantly increased(P<0.05).Conclusion miR-17 promotes the proliferation of VSMCs by targeting IGF-1.This indicates that miR-17 can be used as a predictive biomarker of CAD,and IGF-1 may be a potential therapeutic target.
作者
金爱萍
李书琳
张倩榕
李冰
JIN Aiping;LI Shulin;ZHANG Qianrong;LI Bing(Department of Elderly Cardiovascular Medicine, The Second Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710004, China)
出处
《西安交通大学学报（医学版）》
CAS
CSCD
北大核心
2021年第4期497-501,514,共6页
Journal of Xi’an Jiaotong University（Medical Sciences）
基金
陕西省重点研发计划项目(No.2019SF-101)。
