原发性高血压的微小RNA表达谱及其机制的初步研究

MicroRNA expression profile and pathogenetic initial study in essential hypertension

目的 比较原发性高血压（EH）患者和健康人血浆中微小（microRNA）的表达,进一步理解EH发生、发展的分子机制.方法 收集15例EH患者和5例健康人的全血,3000 r/min离心10 min分离血浆,从血浆中提取RNA.使用Exiqon公司的microRNA芯片检测microRNA表达谱,并对差异表达的microRNA进行定量RT-PCR验证.另外选取独立验证样本（包含24例EH患者和22例健康人）进一步验证.结果 EH患者中的microRNA表达谱与健康人相比有显著不同,1700个microRNA中有46个存在差异表达,其中27个microRNA已被Sanger microRNA数据库收录.在这27个microRNA中有9个上调,18个下调.miR-296-5p（Fold change 0.47,P=0.013）和miR-133b（Fold change 0.57,P=0.033）在EH患者中下调;而let-7e（Fold change 1.62,P=0.009）和hcmv-miR-UL112（Fold change 2.72,P=0.004）表达上调.hcmv-miR-UL112是人巨细胞病毒编码的microRNA.这些差异表达microRNA在另一个独立验证样本中也得到证实.通过荧光报告检测法我们证实,MHC Ⅰ类多肽相关序列B（MICB）和干扰素调节因子1（IRF-1）是hcmv-miR-UL112的下游靶基因.IRF-1可通过作用于一氧化氮合酶和血管紧张素Ⅱ受体参与血压调节.结论 EH患者有着独特的microRNA表达谱,hcmv-miR-UL112对于EH的发病机制有一定的意义.

Objective To study the differential microRNAs expression between patients with essential hypertension and healthy controls.Methods Whole blood from 15 hypertensive patients and 5 controls healthies were separated into plasma at 3000 rpm for 10 minutes. MicroRNAs were harvested using kit, and stored at -80 ℃.MicroRNAs profiling were performed using Exiqon microRCURYTM LNA microRNAs array, and were quantitative RT-PCR for the differential microRNAs expression. In addition, we used a set of plasma samples from 24 hypertensive patients and 22 healthy donors to independently validate the expression of these signature microRNAs.Results MicroRNAs expression profile was found to be differentially in the essential hypertensive patients compared with the healthy donors. Of 1700 microRNAs detected on the microarray, 46 microRNAs were found to be differentially expressed in the essential hypertensive patient, 27 microRNAs were collected in Sanger microRNAs data-bank, the function of remaining 19 microRNAs were unknown. In the 27 microRNAs, 9 microRNAs were up-regulated in the hypertension patient samples, while 18 known microRNAs were down-regulated. MiR-296-5p（Fold change 0.47,P=0.013） and miR-133b （Fold change 0.57,P=0.033）were consistently down-regulated in the patient plasma, whereas let-7e （Fold change 1.62,P=0.009）and hcmv-miR-UL112（Fold change 2.72,P=0.004）, one human cytomegalovirus encoded microRNAs, were up-regulated in the patient samples. The microRNAs expression was independently validated using another sample. We showed that MHC class Ⅰ polypeptide-related chain B（MHC class Ⅰ polypeptide-related chain B,MICB）and Interferon regulatory factor 1（Interferon regulatory factor 1,IRF1） were functional targets of hcmv-miR-UL112 by fluorescent reporter assays.Conclusions The hypertensive patients have distinct microRNAs expression Profile. Hcmv-miR-UL112 may have important implications toward pathogenesis of essential hypertension.