miR-133b在Huh7肝细胞中对糖异生的作用研究

Effect of miR-133b on gluconeogenesis in Huh7 hepatocytes

目的探讨在不同小鼠模型肝脏中微小非编码RNA-133b(miR-133b)的表达水平及其在Huh7肝细胞中对糖异生的作用。方法利用实时荧光定量-聚合酶链反应(RT-PCR)分别检测了miR-133b在高脂饮食喂养野生型C57BL/6小鼠、饥饿24 h的野生型C57BL/6小鼠以及糖尿病动物模型db/db小鼠肝脏组织中的表达水平,并在人源肝癌细胞系Huh7肝细胞中转染miR-133b类似物(mimic)或抑制剂(inhibitor),检测细胞的葡萄糖产出量,磷酸烯醇式丙酮酸羧激酶(PEPCK)和葡萄糖-6-磷酸酶(G6Pase)的信使RNA(mRNA)水平及蛋白水平,并分析其对糖异生的影响。结果miR-133b在高脂饮食喂养野生型C57BL/6小鼠肝脏组织中表达显著升高,而在饥饿24 h的野生型C57BL/6小鼠及db/db小鼠的肝脏组织中表达水平却明显下降。Huh7肝细胞中过表达miR-133b明显降低了Huh7肝细胞的糖异生水平,而抑制miR-133b表达后可以显著升高糖异生水平。结论miR-133b的表达水平与肝脏的营养状态密切相关,并可通过调节Pepck基因的表达水平调节肝细胞的糖异生。

ObjectiveTo determine the expression level of micro non-coding RNA-133b(miR-133b)in the liver of different mouse model and its effect of miR-133b on gluconeogenesis in Huh7 hepatocytes.Methods The expression of miR-133b in the livers of wild type C57BL/6 mice fed with high fat diet,wild type C57BL/6 mice fasted for 24 h and diabetes animal model db/db mice were detected by real-time polymerase chain reaction(RT-PCR).After miR-133b mimic or inhibitor was transfected into Huh7 hepatocytes,glucose production experiment was conducted and mRNA and protein levels of phosphoenolpyruvate carboxykinase(PEPCK)and glucose-6-phosphatase(G6Pase)were measured.ResultsThe expression of miR-133b in the liver of wild-type C57BL/6 mice fed with high-fat diet was significantly increased,while that in the liver of wild-type C57BL/6 mice and db/db mice fasted for 24 h was significantly decreased.Overexpression of miR-133b in Huh7 hepatocytes significantly reduced the level of gluconeogenesis in Huh7 hepatocytes,while inhibition of miR-133b expression significantly increased the level of gluconeogenesis in Huh7 hepatocytes.ConclusionsmiR-133b is closely related to liver nutrient metabolism and it can regulate the glycogenesis of hepatocytes by regulating the expression level of Pepck gene.