血清外泌体miR-144与巨噬细胞脂质堆积：OSAHS促进动脉粥样硬化的新机制

Obstructive sleep apnea/hypopnea syndrome (OSAHS) is an independent risk factor of atherosclerosis. However, the mechanism underlying the pathological process is still not fully unrevealed. And there is still no effective intervention for OSAHS-related atherosclerosis. Serum exosome (SExo) exerts its function of cell-to-cell communication through the mediation of miRNA cargos. However, the involvement of SExo in OSAHS-related atherosclerosis is hardly unknown. The preliminary experiments demonstrated that SExos derived from chronic intermittent hypoxia (CIH) mice aggravated atheroslcerosis development in apoE-/- mice after in vivo treatment and caused the abnormal neutral lipid accumulation in macrophage cells. miRNA chip assay and qPCR validation presented that the expression of miR-144 from CIH mouse or OSAHS patient SExo was significantly increased compared to normal groups, implying that serum exosomal miR-144 possibily mediates the lipid aggregation in macrophage and then promotes atherosclerosis under CIH status. This study will utilize gene intervention combinded with molecular biology techniques to confirm the cause-effect relationship and the regulation pathways between the upregulated SExo miR-144 and OSAHS-related atherosclerosis, and reveal the mechanism through which CIH status increases SExo miR-144 expression. And more importantly, this study will determine the role of the expression and regulation intervention of miR-144 in OSAHS-related atherosclerosis by carrying out in vivo and in vitro experiments. This research will provide the theory basis for the clinical prevailing OSAHS-related atherosclerosis, and may promote new directions in future research concerning the etiology and treatment of OSAHS-related atherosclerosis.

阻塞性睡眠呼吸暂停低通气综合征（OSAHS）是动脉粥样硬化的独立危险因素，但分子机制不清，缺乏有效干预措施。血清外泌体（SExo）通过miRNA调节细胞功能，但是否参与OSAHS促进动脉粥样硬化尚不明了。预实验发现慢性间歇性缺氧（CIH）小鼠SExo促进巨噬细胞脂质积累，加剧粥样斑块发展；芯片筛选发现CIH小鼠和OSAHS患者SExo中miR-144表达显著增加。本研究将应用基因干预与分子生物学技术，结合在体与细胞研究手段，确认SExo中miR-144表达增加与动脉粥样硬化巨噬细胞脂代谢异常的因果关系及调节通路；揭示CIH导致外泌体miR-144表达增加的分子机制；确定干预miR-144表达与调节通路的措施抑制动脉粥样硬化的保护效应。本研究不但试图证明SExo中miR-144表达上升可能是OSAHS促进动脉粥样硬化的分子机制的科学假设，也有助于为OSAHS相关动脉粥样硬化提供新的治疗靶点。

细胞外囊泡是由不同类型细胞分泌的纳米颗粒，包被各种功能蛋白、脂质和核酸，细胞外囊泡作为细胞间通讯的重要媒介，参与了多种生理病理状态的调控，循环细胞外囊泡可被内皮细胞吸收，从而影响血管生成、血管通透性、血管张力的调节。红细胞是血液中最丰富的细胞，是循环细胞外囊泡的重要来源之一，间歇性缺氧条件下红细胞外囊泡的脱落可能通过诱导异常的血管活性因子的产生而影响内皮功能，通过囊泡提取，囊泡鉴定，组学筛选，血管功能研究及临床相关性研究等关键技术对红细胞来源的细胞外囊泡在间歇性低氧相关内皮功能障碍中的作用及机制研究，得出红细胞细胞外囊泡在间歇缺氧状态下可通过抑制内皮细胞 PI3K/Akt/eNOS 通路、促进Erk/ET-1 通路，及改变miR-144/Nrf2通路导致内皮功能障碍，从而诱导内皮依赖性血管舒张功能损伤，参与了OSA相关高血压的发生发展。此外研究了心肌细胞细胞外囊泡在间歇性缺氧状态的MicroRNA差异表达谱及其对内皮功能的影响，通过qPCR分析进行验证，最终证明11个miRNA具有与鉴定的miRNA芯片结果相同的趋势；KEGG功能通路分析表明，差异表达的miRNA的主要富集在PI3K/Akt信号传导途径。血管张力实验检测系统检测发现间歇缺氧条件下心肌细胞分泌的细胞外囊泡损伤血管内皮细胞舒张功能，并且抑制血管内皮细胞中AKT/p-eNOS通路表达。这些研究首次阐明了红细胞和心肌细胞细胞外囊泡在间歇缺氧环境下对血管功能的影响及机制，为相关疾病的临床诊治提供新的线索。我们还进行了糖尿病细胞外囊泡生物标志物研究，首次阐明血清以及血清细胞外囊泡中的髓过氧化物酶水平增加2型糖尿病独立相关，并不受其他混杂因素的影响。这些研究探索了细胞外囊泡包括血清细胞外囊泡、红细胞细胞外囊泡和心肌细胞外囊泡在糖尿病及间歇缺氧环境下对血管功能的影响和机制，为糖尿病及间歇缺氧状态血管内皮功能障碍提供了新诊疗线索和靶点。该项目支持下，完成四篇SCI文章，合计影响因子25分。