MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in Type 2 Diabetes mellitus

Dysregulation in expression of microRNAs (miRNAs) in various tissues has been linked to a wide spectrum of diseases, including Type 2 Diabetes mellitus (T2D). In this study, we compared the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients with tissues from T2D rat models. Healthy adult males with no past history of T2D (n=158) and with desirable cholesterol and blood pressure profiles were enrolled in this study. They were then classified according to fasting glucose levels to have T2D, IFG or as healthy controls (CTL), for comparison of miRNA expression profiles. Employing miRNA microarray, we identified ‘signature miRNAs’ in peripheral blood samples that distinguished IFG and T2D. Eight selected miRNAs were further validated using stem-loop real-time RT-PCR. miR-144 expression was found to be dysregulated in Type 2 Diabetes, wherein its expression was significantly higher than in healthy controls. Insulin receptor substrate 1 (IRS1) has been predicted to be a potential target of miR-144. Consistent with this observation, IRS1 mRNA and protein levels, verified by quantitative real-time PCR and western blotting respectively, were found to be down-regulated. Using luciferase assay, we further demonstrated that miR-144 directly targets IRS1 and showed its effects on protein expression via immunocytochemistry. From this cross-sectional study in humans, we have identified signature miRNAs which could explain the pathogenesis of T2D. Whether miRNAs like miR-144 could be potential therapeutic targets for management of T2D will need to be explored by further mechanistic and functional studies.多种组织中微小RNA（microRNAs, miRNAs）的表达失调与包括2型糖尿病（Type 2 Diabetes mellitus, T2D）在内的多种疾病密切相关。本研究对比了空腹血糖受损（Impaired Fasting Glucose, IFG）男性患者、2型糖尿病男性患者的血液样本miRNA表达谱，以及2型糖尿病大鼠模型的组织miRNA表达谱。本研究纳入158名无2型糖尿病病史、胆固醇及血压水平均处于理想范围的健康成年男性，随后根据空腹血糖水平将其分为2型糖尿病组、空腹血糖受损组及健康对照组（CTL），以比较各组的miRNA表达谱。通过miRNA微阵列技术，我们在外周血样本中筛选出了可区分空腹血糖受损与2型糖尿病的"特征性miRNAs"。随后选取8种miRNAs，采用茎环实时逆转录聚合酶链反应进行了验证。研究发现，miR-144在2型糖尿病患者体内表达失调，其表达水平显著高于健康对照组。胰岛素受体底物1（Insulin receptor substrate 1, IRS1）被预测为miR-144的潜在靶基因。与此发现一致的是，通过实时定量PCR及蛋白质印迹法分别验证的IRS1 mRNA及蛋白水平均呈下调趋势。进一步通过荧光素酶报告基因实验证实，miR-144可直接靶向IRS1，并通过免疫细胞化学实验验证了其对蛋白表达的调控作用。本项人类横断面研究筛选出了可阐释2型糖尿病发病机制的特征性miRNAs。诸如miR-144这类miRNA是否可作为2型糖尿病治疗的潜在靶点，仍需后续的机制与功能研究进一步探索。