过表达miR-128a抑制大鼠前体脂肪细胞分化  

Inhibition of the Pre-adipocyte Differentiation in Rat(Rattus norvegicus) Caused by Over-expression of miR-128a

作　　者：仇杨[1] 刘帅[1] 宁小敏[1] 董培越[1] 程佳[1] 何晶晶[1] 李艳杰[1] 杨公社[1] 孙世铎[1] 

机构地区：[1]西北农林科技大学动物脂肪沉积与肌肉发育实验室,杨凌712100

出　　处：《农业生物技术学报》2013年第4期388-395,共8页Journal of Agricultural Biotechnology

基　　金：国家自然基因项目(No.31072014);陕西省农业厅陕西生猪产业技术体系资助项目(No.K332021105)

摘　　要：为阐明miR-128a在大鼠前体脂肪细胞分化过程中的功能,本实验检测了大鼠(Rattus norvegicus)前体脂肪细胞成脂分化过程中miR-128a的表达,明确其在分化过程中的表达趋势;构建miR-128a的腺病毒过表达载体并侵染大鼠前体脂肪细胞,随后采用Real-time PCR和Western blot分别在mRNA和蛋白水平检测了成脂标记基因过氧化物酶体增生物激活受体γ(peroxisome proliferator-activated receptor gamma,PPARγ)和脂肪酸结合蛋白2(adipocyte protein 2,aP2)的表达;过表达miR-128a的大鼠前体脂肪细胞在第10天进行油红O染色,从形态学上观察成脂分化情况。结果显示,miR-128a的表达量在脂肪细胞诱导分化第二天降到最低点,随后维持在第0天的水平。在过表达miR-128a后,大鼠前体脂肪细胞成脂标记基因PPARγ和aP2的蛋白表达量与对照相比显著下降,脂滴明显少于对照组。实验结果表明:在大鼠前体脂肪细胞中过表达miR-128a能够抑制前体脂肪细胞的分化;并且通过PPARγ mRNA和蛋白水平的差异性变化,结合miRNA的作用机理,推测PPARγ有可能是miR-128a的靶基因。本实验也为研究miR-128a在脂肪细胞分化中发挥作用的机理提供了理论基础。To clarify the function of miR-128a during the differentiation of rat （Rattus norvegicus） preadipocytes, we detected the expression level of miR-128a during adipogenesis and clarified its expression tendency during differentiation; then we constructed an over-expression adenovirus vetor of pre-miR-128a and infected rat preadipocytes. Subsequently, the expression of adipogenesis marker peroxisome proliferator- activated receptor gamma（PPART） and adipocyte protein 2（aP2） were analysed by Real-time PCR and Western blot in mRNA and protein level, respectively. After the adenovirus vector infecting rat preadipocytes, we detected lipids accumulation in 10th day by Oil-red O staining, observing the adipogenesis in morphology. Results showed that the expression level of miR- 128a was at the minimum point in the next day of the adipocyte differentiation, and then maintained at the level of day 0. After the over-expression of miR-128a, the expression levels of adipocyte marker genes PPARγ, aP2 and the number of lipid droplets had obviously declined, compared with that in control. The results showed that miR- 128a could inhibit the differentiation of pre-adipocytes in rat. We suspected that PPARγ is a potential target gene of miR- 128a according to the changes of mRNA and protein expressing levels observed after the treatment of adenovirus infection in rat pre-adipocytes. Also, this study supplies the basic theory for the further research about the function of miR-128a during the differentiation of adipocytes.

关 键 词：miR-128a 大鼠前体脂肪细胞 过表达 

分 类 号：S852.2[农业科学—基础兽医学]