miR-221对糖尿病肾病小鼠胰岛β细胞功能的保护作用及其机制

Protective effect of miR-221 on β cells in mice with diabetic nephropathy and its mechanism

目的:研究miR-221对糖尿病肾病(DN)小鼠胰岛β细胞功能的影响,阐明miR-221在DN中的保护作用机制。方法:8周龄野生型雄性C57BL/6J小鼠20只随机分成对照组和DN组,每组10只,对照组小鼠给予正常饮食,DN组小鼠给予高脂饮食,并注射100mg·kg^(-1)链脲佐菌素(STZ)诱导部分胰岛素缺陷,对照组小鼠注射同体积柠檬酸缓冲液。之后DN组继续给予高脂饮食,对照组给予正常饮食,饲养10周后收集小鼠血液和尿液,检测血糖、血肌酐、血尿素氮浓度和24h尿白蛋白排泄率等生理参数。在DN组小鼠中分离得到胰岛细胞。利用Real-time PCR检测胰岛细胞中SOCS3mRNA表达水平,采用MTT法检测胰岛细胞增殖情况,ELISA法检测胰岛细胞中胰岛素含量和胰岛素的释放水平,荧光素酶报告基因法检测SOCS3活性荧光酶素报告基因活性。结果:成功构建DN小鼠模型。DN组小鼠的血糖、血肌酐、血尿素氮浓度和24h尿白蛋白排泄率均高于对照组(P<0.05)。过表达miR-221后,DN组小鼠胰岛细胞中SOCS3mRNA表达水平低于正常胰岛细胞(P<0.05),过表达miR-221后胰岛细胞的增殖能力低于正常胰岛细胞(P<0.05),荧光素酶报告基因法确定SOCS3为miR-221下游靶基因。过表达miR-221后,胰岛细胞中胰岛素质量分数和胰岛素释放水平均高于正常胰岛细胞(P<0.05)。结论:miR-221通过下调SOCS3水平促进胰岛细胞合成和分泌胰岛素的功能,改善DN小鼠中胰岛细胞的功能障碍。

Objective：To study the influence of miR-221 in theβcells of mice with diabetic nephropathy （DN）, and to clarify the protective effect of miR-221 on DN.Methods：Twenty wild type C57BL/6J mice aged 8 weeks were divided into control group and DN group （n=10）.The DN mice models were constructed with 14 weeks of high fat diet,and 100 mg·kg-1 streptozotocin （STZ）was used to induce the partial insulin deficiency.10 weeks later the blood glucose,serum creatinine,blood urea nitrogen content and 24h-urinary albumin excretion rate were detected.Theβcells of islet were isolated from the DN mice.The expression level of SOCS3 mRNA inβ cells of islet was valued by Real-time PCR.The proliferation of pancreaticβcells of islet was examined by MTT assay.The contents of insulin and insulin release levels in pancreaticβ cells of islet were detected by ELISA assay.Luciferase reporter gene assay was used to detect the luciferase reporter gene activity of SOCS3.Results：The DN mouse models were constructed successfully.The blood glucose,serum creatinine,blood urea nitrogen and 24h-urinary＆amp;nbsp;albumin excretion rate of the mice in DNA group were higher than those in control group （P 〈 0.05 ）.After overexpression of miR-221,the expression level of SOCS3 mRNA in pancreaticβcells of islet and the proliferation ability ofβcells of islet of the mice in DN group were lower than those of normal islet cells （P 〈 0.05）.Then luciferase reporter gene method found SOCS3 as one of the target genes of miR-221.After overexpression of miR-221,the insulin release level and insulin content in pancreaticβcells of islet were higher than those of normal islet cells （P 〈0.05）.Conclusion：miR-221 can promopt the synthesis and secretion ofβ cells of islet and inhibit the dysfunction ofβcells of islet in the DN mice by down-regulating the SOCS3 level.