摘要
目的探讨长链非编码RNA AW112010(lncRNA AW112010)是否可能通过miR-204/POU类同源盒2(POU2F2)信号通路改善衰老脂肪细胞的胰岛素抵抗。方法体内实验:将C57BL/6小鼠按体重分成年轻对照组(4月龄)、衰老模型组(18月龄);实时荧光定量PCR及Western印迹法检测附睾脂肪组织中lncRNA AW112010、miR-204-5p、POU2F2、衰老相关指标(p16、p21)及胰岛素信号通路基因[胰岛素受体(INSR)、胰岛素受体底物1(IRS1)、磷脂酰肌醇激酶(PI3K)、蛋白激酶B(AKT)]的表达量。体外实验:用阿霉素诱导3T3-L1衰老脂肪细胞模型,β-gal染色观察细胞衰老,成功构建miR-204抑制剂及miR-204模拟物的小干扰RNA,转染3T3-L1脂肪细胞。结果实时荧光PCR与Western印迹结果表明,与年轻小鼠比较,衰老小鼠附睾脂肪组织中AW112010的表达水平增高,而miR-204-5p的表达水平降低;衰老小鼠附睾脂肪组织中POU2F2、p16、p21的表达水平增高,而INSR、IRS1、PI3K、GLUT4 mRNA及蛋白的表达水平降低。β-gal染色结果表明,阿霉素诱导的3T3-L1衰老脂肪细胞数量明显增多,且与对照组相比,阿霉素诱导的衰老脂肪细胞中AW112010、POU2F2、p16、p21的表达量增高,而miR-204-5p、INSR、IRS1、PI3K、GLUT4的表达量降低,培养基中葡萄糖的剩余量增多。与对照组相比,miR-204过表达后,衰老指标p16、p21及靶基因POU2F2的表达量降低;INSR、GLUT4的表达量增高。结论衰老小鼠脂肪细胞中上调的lncRNA AW112010可能通过靶向miR-204-5p/POU2F2/IRS1导致胰岛素抵抗。
Objective To investigate whether long non-coding RNA(lncRNA)AW112010 can improve insulin resistance in aging adipocytes through the miR-204/POU2F2 signaling pathway.Methods In vivo experiment:C57BL/6 mice were divided into young control group(4 months old)and aging model group(18 months old)based on body weight.The expression levels of AW112010,miR-204-5p,POU2F2,aging related indicators(p16,p21),and insulin signaling pathway genes[insulin receptor(INSR),insulin receptor substrate 1(IRS1),phosphatidylinositol kinase(PI3K),protein kinase B(AKT)]in epididymal adipose tissue were detected using real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting.In vitro experiment:Using adriamycin(ADR)to induce 3T3-L1 aging adipocyte model,β-gal staining was used to observe cellular senescence,and miR-204 inhibitor and miR-204 mimic small interfering RNA were successfully constructed and transfected into 3T3-L1 adipocytes.Results RT-qPCR and Western blot results showed that compared with the young group,the expression of AW112010 in the adipose tissue of aging mice was increased,while the expression of miR-204-5p was decreased.The expressions of POU2F2,p16,and p21 in the adipose tissue of aging mice were increased,while the expressions of INSR,IRS1,PI3K,GLUT4 mRNA and protein were decreased.Theβ-gal stainging results showed that the number of 3T3-L1 senescent adipocytes induced by ADR was significantly increased,and the expression levels of AW112010,POU2F2,p16,and p21 in ADR-induced senescent adipocytes were increased compared with the control group,while the expression levels of miR-204-5p,INSR,IRS1,PI3K,GLUT4 were decreased,and remaining glucose in the culture medium was increased.Compared with control,overexpression of miR-204 resulted in decreased expressions of aging indicators p16,p21,and target gene POU2F2 while the expressions of INSR and GLUT4 were increased.Conclusion Upregulation of lncRNA AW112010 in adipocytes of aging mice may induce insulin resistance by targeting miR-204-5p/POU2F2/IRS1.
作者
王锐
王淑文
张一凡
胡雅琪
原琪
温媛
陈晓玲
卢婷
郑颖
林志勇
薛梦珍
王雅琦
夏芳奇
朱蕾奇
袁成福
Wang Rui;Wang Shuwen;Zhang Yifan;Hu Yaqi;Yuan Qi;Wen Yuan;Chen Xiaoling;Lu Ting;Zheng Ying;Lin Zhiyong;Xue Mengzhen;Wang Yaqi;Xia Fangqi;Zhu Leiqi;Yuan Chengfu(Third-grade Pharmacological Laboratory on Traditional Chinese Medicine Approved by State Administration of Traditional Chinese Medicine,College of Basic Medical Sciences,Three Gorges University,Yichang 443002,China)
出处
《中华内分泌代谢杂志》
CAS
CSCD
北大核心
2024年第1期44-52,共9页
Chinese Journal of Endocrinology and Metabolism
基金
国家自然科学基金项目(81974528、82174035、81773959)
湖北省自然科学基金创新群项目(2021CFA015)
湖北省中央引导地方科技发展专项(2020ZYYD016)。
