醛固酮通过抑制血管平滑肌细胞自噬促进血管钙化的机制

Vascular calcification (VC) is an important risk factor of increased cardiovascular events in hypertension. Our previous studies demonstrated that abnormal activation of renin-angiotensin-aldosterone system played an important role in this process. Our recently published paper on Hypertension indicated that excess aldosterone is closely associated with prevalent abdominal aortic calcification. Autophagy and transdifferentiation of vascular smooth muscle cells (VSMCs) are the key elements in VC. We found that aldosterone promoted VC, which was accompanied by inhibition of VSMC autophagy. However, the underlying mechanism is unclear. Based on RNA sequencing results, we raise the hypothesis that aldosterone inhibits autophagy through upregulation of lncRNA H19 and downregulation of chromatin remodeling factor SMARCA4/5, resulting in acceleration of VC. In this proposal, we intervened lncRNA H19 and its downstream signaling targets in vivo model of VC (wild-type mice fed with diet containing 0.75% adenine and 0.9% phosphorus) and in vitro model of VC (VSMCs induced by 2.6 mM phosphate) combined treatment with aldosterone. With the methods of RNA silencing and overexpression, dual-luciferase reporter gene analysis, EMSA and CHIP, we aim to investigate the detailed mechanisms of aldosterone-autophagy-VSMC transdifferentiation-VC in vivo and in vitro level. This project will provide a new therapeutic strategy for VC.

血管钙化是高血压发生心脑血管事件的重要危险因素之一。我们的系列研究证实肾素－血管紧张素－醛固酮系统的异常激活在其中扮演着重要作用。最近我们发表于《Hypertension》杂志的工作提示，高醛固酮血症与腹主动脉钙化的发生率密切相关。自噬和血管平滑肌细胞转分化是血管钙化的关键环节，我们发现醛固酮能促进血管钙化，同时抑制血管平滑肌细胞自噬，然而其潜在机制亟待阐明。结合RNA测序的结果，我们提出醛固酮通过上调长链非编码RNA H19、下调染色质重塑因子SMARCA4/5抑制自噬，加速血管钙化的假说。本研究分别采用高腺嘌呤高磷饮食诱导及高磷刺激建立动物及细胞钙化模型，并予醛固酮处理，同时干预长链非编码RNA H19以及下游的信号分子，结合RNA干扰与过表达、双荧光素酶报告基因、EMSA及CHIP等方法深入剖析醛固酮－自噬－血管平滑肌细胞表型转化－血管钙化的作用规律，为血管钙化的防治提供新策略。

血管钙化是老龄化、慢性肾脏病、高血压等普遍存在的共同的病理生理变化，是未来发生心脑血管事件重要的预测因素。本项目结合前期预实验结果，系统探讨了醛固酮在促进血管钙化过程中的作用及其分子机制。首先，我们在动物模型中证实，醛固酮促血管钙化的过程中需要高磷环境，而单独醛固酮无法诱导钙化形成。其次，我们证实醛固酮通过激活盐皮质激素受体（MR）、抑制血管平滑肌细胞（VSMCs）自噬，促进VSMCs钙化及成骨表型转分化。第三，结合前期RNA测序及生信预测的结果，我们进一步证实醛固酮通过与MR结合，上调长链非编码RNA H19（H19）启动子活性，竞争性抑制miR-106a-5p与Runx2结合，解除其对Runx2抑制效应，从而加速血管钙化。本项目通过离体、在体实验，运用RNA干扰、双荧光素酶报告基因分析、EMSA及CHIP等多种手段深入验证了H19、自噬在醛固酮促血管钙化中的作用规律，为高醛固酮血症所致的血管钙化的精准防治提供新策略。