摘要
目的探讨微小RNA(miRNA)Let-7f-5p在糖尿病大鼠心肌微血管病变中的作用及机制。方法将25只SD雄性大鼠随机分为正常对照组5只和模型组20只,正常对照组给予普通饲料喂养;模型组给予高脂高糖饲料喂养4周后,一次性腹腔注射链脲佐菌素制作糖尿病模型。取造模成功的15只大鼠,随机分为空病毒组、Let-7f-5p过表达组、Let-7f-5p抑表达组各5只,Let-7f-5p过表达组、Let-7f-5p抑表达组分别尾静脉一次性注射Let-7f-5p过表达病毒原液50μL、Let-7f-5p抑制表达病毒原液50μL;空病毒组尾静脉一次性注射等体积空病毒原液。继续高脂高糖饲料喂养12周,诱导糖尿病心肌微血管病变。所有大鼠麻醉,使用彩色多普勒超声显像仪记录左心室舒张末期内径(LVIDd)、左心室收缩末内径(LVIDs)、左心室射血分数(LVEF)、左心室缩短分数(FS)。取心肌组织,免疫组化标记心肌微血管数目,计算毛细血管数量/心肌细胞核比值(C/M)用于评估心肌微血管密度;RT-PCR法检测Let-7f-5p及Argonaute蛋白1(AGO1)、血管内皮生长因子(VEGF)mRNA表达,Western blotting法检测AGO1、VEGF蛋白表达。结果与对照组比较,空病毒组LVIDd、LVIDs、LVEF、FS均降低(P均<0.05);C/M降低(P均<0.05);心肌组织Let-7f-5p mRNA表达降低,AGO1 mRNA及蛋白表达增加,VEGF mRNA和蛋白表达降低(P均<0.05)。与空病毒组比较,Let-7f-5p过表达组LVIDd、LVIDs、LVEF、FS均升高(P均<0.05);C/M升高(P均<0.05);心肌组织Let-7f-5p mRNA表达升高,AGO1 mRNA及蛋白表达降低,VEGF mRNA和蛋白表达升高(P均<0.05)。与空病毒组比较,Let-7f-5p抑表达组LVIDd、LVIDs、LVEF、FS均降低(P均<0.05);C/M降低(P均<0.05);心肌组织Let-7f-5p mRNA表达减少,AGO1 mRNA及蛋白表达增加,VEGF mRNA和蛋白表达降低(P均<0.05)。结论糖尿病大鼠心肌微血管密度降低,心脏收缩功能受损。上调Let-7f-5p表达可能通过Let-7/AGO1/VEGF信号通路增加心肌微血管密度,改善心脏功能;
Objective To explore the role of miRNA Let-7f-5p in myocardial microvascular disease of diabetic rats and the mechanism.Methods Twenty-five male Sprague Dawley(SD)rats were randomly divided into the normal con⁃trol group(n=5)and model group(n=20);the rats in the model group were fed with high-fat and high-sugar diet for 4 weeks,and then streptozotocin was injected intraperitoneally to create diabetic rat models.After successful establishment of diabetic rat models,fifteen diabetic rats were selected and randomly divided into the empty virus group,Let-7f-5p overexpression group and Let-7f-5p down-expression group,with 5 rats in each group.Rats in the Let-7f-5p over-expression group were injected with 50μL Let-7f-5p over-expression virus through tail vein,and rats in Let-7f-5p down-expression group were injected with 50μL Let-7f-5p expression inhibition virus through tail vein.In the empty virus group,the same volume of empty virus was injected into the tail vein at one time.Diabetic myocardial microvascular lesions were induced by continuous high-fat and high-glucose diet for 12 weeks.The left ventricular internal diameter at end-diastole(LVIDd),left ventricular internal diameter at end-systole(LVIDs),left ventricular ejection fraction(LVEF),and left ventricular shortening fraction(FS)were recorded by color Doppler ultrasound after anesthesia.After cardiac ultrasound examina⁃tion,the myocardial tissue was removed rapidly.We used immunohistochemistry to label the number of myocardial mi⁃crovessels,and calculated the ratio of capillary number/cardiomyocyte nucleus(C/M)to evaluate myocardial microvessel density.RT-PCR was used to detect the mRNA expression levels of Let-7f-5p,Argonaute protein 1(AGO1)and vascular endothelial growth factor(VEGF),and Western blotting was used to detect the protein expression levels of AGO1 and VEGF.Results Compared with the control group,LVIDd,LVIDs,LVEF and FS decreased(all P<0.05),C/M ratio decreased(P<0.05),myocardial Let-7f-5p mRNA expression decreased(P<0.05),AGO1 mRNA and
作者
黄河
周瑞
刘星
刘应才
HUANG He;ZHOU Rui;LIU Xing;LIU Yingcai(The Affiliated Hospital of Southwest Medical University,Luzhou 646000,China)
出处
《山东医药》
CAS
2021年第14期14-18,共5页
Shandong Medical Journal
基金
四川省泸州市科技厅—西南医科大学联合课题项目(2016LZXNYD-J02)。
