Hu Mengdie, Huang Xiuting, Han Xueyao, Ji Linong
Department of Endocrinology and Metabolism, Peking University People's Hospital, Peking University Diabetes Center, Beijing 100044, People's Republic of China.
Diabetes Metab Syndr Obes. 2020 Mar 2;13:627-639. doi: 10.2147/DMSO.S236915. eCollection 2020.
Mutations in hepatocyte nuclear factor 1α (HNF1α) are the cause of maturity-onset diabetes of the young type 3 (MODY3) and involved in the development of hepatocellular adenoma and abnormal lipid metabolism. Previously, we have found that the serum microRNA (miR)-122 levels in MODY3 patients were lower than those in type 2 diabetes mellitus and healthy controls. This study aimed to investigate the mechanism of decreased miR-122 levels in patients with MODY3 and whether low levels of miR-122 mediate tumorigenesis and abnormal lipid metabolism associated with HNF1α deficiency in human hepatocytes.
The expression of miR-122 was examined by real-time PCR. Dual-luciferase reporter assay was performed to confirm the transcriptional regulation of miR-122 by HNF1α. HepG2 cells were transfected with siRNA or miRNA mimic to downregulate or upregulate the expression of HNF1α or miR-122, respectively. CCK-8 and colony formation assay were used to determine cell proliferation. Lipid accumulation was examined by Oil Red O staining and intracellular triglyceride and cholesterol quantification assays.
HNF1α regulated the expression of miR-122 by directly binding to its promoter. Knockdown of HNF1α in HepG2 cells reduced the expression of miR-122, increased proliferation and promoted intracellular cholesterol accumulation. Overexpression of miR-122 partially rescued the phenotypes associated with HNF1α deficiency in human hepatocytes. Mechanistically, HNF1α modulated cholesterol homeostasis via miR-122-dependent activation of sterol regulatory element-binding protein-2 (SREBP-2) and regulation of proprotein convertase subtilisin/kexin type 9 (PCSK9). Moreover, circulating miR-122 levels were associated with serum cholesterol levels.
Loss of HNF1α function led to hepatocyte proliferation and abnormal cholesterol metabolism by downregulating miR-122. Our findings revealed a novel mechanism that low levels of miR-122 mediate tumorigenesis and abnormal lipid metabolism associated with MODY3. MiR-122 may be a potential therapeutic target for the treatment of MODY3.
肝细胞核因子1α（HNF1α）突变是青年发病的成年型糖尿病3型（MODY3）的病因，且与肝细胞腺瘤的发生及脂质代谢异常有关。此前，我们发现MODY3患者血清微小RNA（miR）-122水平低于2型糖尿病患者和健康对照者。本研究旨在探究MODY3患者miR-122水平降低的机制，以及低水平的miR-122是否介导了与人类肝细胞中HNF1α缺乏相关的肿瘤发生和脂质代谢异常。
通过实时聚合酶链反应检测miR-122的表达。进行双荧光素酶报告基因检测以证实HNF1α对miR-122的转录调控。分别用小干扰RNA（siRNA）或miRNA模拟物转染HepG2细胞，以下调或上调HNF1α或miR-122的表达。采用细胞计数试剂盒-8（CCK-8）法和集落形成试验测定细胞增殖。通过油红O染色及细胞内甘油三酯和胆固醇定量检测来检查脂质蓄积情况。
HNF1α通过直接结合其启动子来调控miR-122的表达。在HepG2细胞中敲低HNF1α可降低miR-122的表达，增加细胞增殖并促进细胞内胆固醇蓄积。miR-122的过表达部分挽救了人类肝细胞中与HNF1α缺乏相关的表型。机制上，HNF1α通过miR-122依赖的方式激活固醇调节元件结合蛋白2（SREBP-2）并调节前蛋白转化酶枯草溶菌素/kexin 9型（PCSK9）来调节胆固醇稳态。此外，循环miR-122水平与血清胆固醇水平相关。
HNF1α功能丧失通过下调miR-122导致肝细胞增殖和胆固醇代谢异常。我们的研究结果揭示了一种新机制，即低水平的miR-122介导了与MODY3相关的肿瘤发生和脂质代谢异常。miR-122可能是治疗MODY3的潜在治疗靶点。
