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miR-122 promotes hepatic lipogenesis via inhibiting the LKB1/AMPK pathway by targeting Sirt1 in non-alcoholic fatty liver disease
Molecular Medicine ( IF 5.7 ) Pub Date : 2019-06-13 , DOI: 10.1186/s10020-019-0085-2
Jun-Ke Long 1 , Wen Dai 1 , Ya-Wen Zheng 2 , Shui-Ping Zhao 1
Affiliation  

BackgroundNon-alcoholic fatty liver disease (NAFLD) is a common hepatic disease with an increasing prevalence but an unclear aetiology. This study aimed to investigate the functional implications of microRNA-122 (miR-122) in the pathogenesis of NAFLD and the possible molecular mechanisms.MethodsBoth in vitro and in vivo models of NAFLD were generated by treating HepG2 and Huh-7 cells with free fatty acids (FFA) and by feeding mice a high-fat diet (HFD), respectively. HE and Oil Red O staining were used to examine liver tissue morphology and lipid deposition, respectively. Immunohistochemical (IHC) staining was used to examine Sirt1 expression in liver tissues. qRT-PCR and Western blotting were employed to measure the expression of miR-122, Sirt1, and proteins involved in lipogenesis and the AMPK pathway. Enzyme-linked immunosorbent assay (ELISA) was used to quantify triglyceride (TG) levels in HepG2 and Huh-7 cells and in liver tissues. The interaction between miR-122 and the Sirt1 gene was further examined by a dual luciferase reporter assay and RNA-immunoprecipitation (RIP).ResultsNAFLD hepatic tissues and FFA-treated HepG2 and Huh-7 cells presented excess lipid production and TG secretion, accompanied by miR-122 upregulation, Sirt1 downregulation, and potentiated lipogenesis-related genes. miR-122 suppressed Sirt1 expression via binding to its 3′-untranslated region (UTR). Knockdown of miR-122 effectively mitigated excessive lipid production and suppressed the expression of lipogenic genes in FFA-treated HepG2 and Huh-7 cells via upregulating Sirt1. Furthermore, miR-122 knockdown activated the LKB1/AMPK signalling pathway.ConclusionThe inhibition of miR-122 protects hepatocytes from lipid metabolic disorders such as NAFLD and suppresses lipogenesis via elevating Sirt1 and activating the AMPK pathway. These data support miR-122 as a promising biomarker and drug target for NAFLD.

中文翻译:

非酒精性脂肪肝中miR-122通过靶向Sirt1抑制LKB1/AMPK通路促进肝脏脂肪生成

背景非酒精性脂肪性肝病(NAFLD)是一种常见的肝脏疾病,发病率不断增加,但病因尚不明确。本研究旨在探讨微小RNA-122(miR-122)在NAFLD发病机制中的功能意义和可能的分子机制。方法通过用游离脂肪处理HepG2和Huh-7细胞产生NAFLD的体外和体内模型。酸(FFA)和分别给小鼠喂食高脂肪饮食(HFD)。HE和油红O染色分别用于检查肝组织形态和脂质沉积。免疫组织化学 (IHC) 染色用于检查肝组织中的 Sirt1 表达。qRT-PCR 和蛋白质印迹法用于测量 miR-122、Sirt1 和参与脂肪生成和 AMPK 途径的蛋白质的表达。酶联免疫吸附测定 (ELISA) 用于量化 HepG2 和 Huh-7 细胞以及肝组织中的甘油三酯 (TG) 水平。通过双荧光素酶报告基因检测和 RNA 免疫沉淀 (RIP) 进一步检查了 miR-122 和 Sirt1 基因之间的相互作用。 结果 NAFLD 肝组织和 FFA 处理的 HepG2 和 Huh-7 细胞呈现出过多的脂质产生和 TG 分泌,伴随着miR-122 上调、Sirt1 下调和增强的脂肪生成相关基因。miR-122 通过与其 3'-非翻译区 (UTR) 结合来抑制 Sirt1 的表达。敲除 miR-122 可通过上调 Sirt1 有效减轻过度的脂质产生并抑制 FFA 处理的 HepG2 和 Huh-7 细胞中脂肪生成基因的表达。此外,miR-122 敲低激活了 LKB1/AMPK 信号通路。结论抑制miR-122可保护肝细胞免受NAFLD等脂质代谢紊乱的影响,并通过上调Sirt1和激活AMPK通路抑制脂肪生成。这些数据支持 miR-122 作为 NAFLD 的有希望的生物标志物和药物靶点。
更新日期:2019-06-13
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