慢性间歇低氧小鼠内皮细胞micro-RNA表达谱及分子机制研究

Obstructive sleep apnea syndrome is a common sleep-related repiratory disease. Much attention has been paid to endothelial dysfunction, which is regarded as the contributors to the development of vascular conditions such as hypertension, ischemic stroke, and myocardial ischemia in patients with OSAHS. Chronic intermittent hypoxemia (oxygen desaturation and re-oxygenation) may be responsible for triggering endothelial dysfunction in OSAHS． MicroRNAs, a class of 20-23 nucleotides, noncoding single-stranded RNAs that regulate gene function,leading to a significant negative regulation of relevant mRNAs to be translated into proteins. Nuclear factor-κB and hypoxia-inducible factor 1 are central transcriptional factor that regulates a battery of genes that are critical to chronic intermittent hypoxia-induced vascular endothelial impairment,，however, the role of miRNAs in chronic intermittent hypoxia-induced vascular endothelial impairment remains less defined. We hypothesized that microRNA may also play a role in the pathological process. The aim of this study is to investigate the expression profile and function of microRNAs which are expressed in vascular endothelial cells in CIH models, using the methods of microRNA array analysis and quantitative real-time PCR. We also investigate the function of microRNAs using specific mimic or inhibitor transfect the endothelium cell to induce overexpression or deficiency of the target microRNA．The results microRNAs are potential targets for novel therapeutics to treat endothelial dysfunction associated OSAHS .

OSAHS是一种常见的睡眠呼吸障碍性疾病。内皮功能紊乱是OSAHS心血管并发症的主要机制。慢性间歇低氧是OSAHS内皮功能紊乱的始动因素。核因子-κB和缺氧诱导因子-l作为重要的转录因子调控下游靶基因的表达介导慢性间歇低氧导致的内皮损伤。微小RNA（microRNA）是一种长度只有20-23nt 的内源性非编码单链RNA，在转录后水平发挥广泛而重要的负调控作用。然而目前microRNA在慢性间歇缺氧细胞组织模型的调控作用尚不明确。基于既往研究microRNA参与了核因子-κB和缺氧诱导因子-l的表达调控，建立慢性间歇缺氧的内皮细胞模型，利用microRNA芯片分析筛选出特异性microRNA，通过microRNA模拟物和抑制物转染内皮细胞以及实验动物模型以研究其对HIF-1、NF-κB及其靶基因的表达调控功能。本研究期望能够为microRNA成为OSAHS内皮损伤防治的新靶点提供研究基础。

本项目通过microRNA芯片筛选CIH状态下小鼠主动脉内皮细胞差异性表达的miRNA。芯片结果mmu-miR-30b, mmu-miR-193, mmu-miR-1249、mmu-miR-218-1表达增加, 而mmu-miR-718 和 mmu-miR-16 表达下降，并进行RT-PCR验证，根据mirbase 和 Tarbase 基因网络分析预测特定microRNA调控的靶基因。研究选取microRNA218及microRNA193进行对靶基因调控功能验证，通过下调慢性间歇低氧状态下小鼠内皮细胞microRNA218及microRNA193表达可以减轻慢性间歇低氧下小鼠内皮细胞的凋亡，因此microRNA218，microRNA193有望作为慢性间歇低氧状态下内皮损伤的重要干预靶点。本项目通过确定特异性MicroRNA以调控OSAHS内皮功能相关基因，将为OSAHS心血管并发症的防治提供新的思路和途径。