AMPK-mTOR通路介导细胞自噬在高尿酸诱导心室重构中的作用及机制研究

Epidemiologic data suggests an association between hyperuricemia(HUA) and heart failure(HF). However, the precise functions of HUA in HF are incompletely understood. Compared to normal rat, 3 weeks after 2% potassium oxazine salt administration caused apoptosis and interstitial fibrosis of left ventricle and the myocyte showed reduced cell contraction and relaxation in our previous study. Meanwhile autophagy was observed significantly increased. AMPK-mTOR signal pathway may play an important role in HUA-induced myocyte autophagy. Therefore, we hypothesize that AMPK-mTOR signaling pathway induced autophagy may be a novel important molecular mechanism in the happen and development of HUA induced ventricular remodeling. This project was aimed to elucidate the relationship among AMPK-mTOR signaling pathway, autophagy and HUA related ventricular remodeling by the following two experimental parts. First of all, clarify the relationship between changes of autophagy and ventricular remodeling in HUA. Secondly, uncover the mechanisms of AMPK-mTOR signaling pathway regulating autophagy in HUA. Functional analysis, morphology and molecular biology are employed both in vivo and in vitro to reveal a novel mechanism and target for the prevention and cure of ventricular remodeling induced by HUA from the new perspective of autophagy.

流行病学数据显示高尿酸血症与心衰关系密切，但高尿酸血症在心衰发生、发展中确切作用及分子机制尚不清楚。我们前期研究发现：高尿酸血症大鼠模型心室凋亡细胞增多，纤维化程度增加，心肌细胞收缩和舒张功能降低。我们还发现高尿酸可激活心肌细胞自噬，AMPK-mTOR信号通路可能在高尿酸诱导心肌细胞自噬中发挥重要调控作用。因此，我们提出研究假设：高尿酸可通过AMPK-mTOR信号通路激活心肌细胞自噬，并在高尿酸引起的心室重构中发挥重要调控作用。为验证上述假设，本研究将：(1)阐明高尿酸诱导的心肌细胞自噬在心室重构中发挥的确切作用；(2)证实AMPK-mTOR信号通路是高尿酸激活心肌细胞自噬的主要机制。本项目拟从在体和细胞水平，采用功能学、形态学和分子生物学等技术，深入揭示高尿酸对心室结构和功能的确切影响及具体分子机制，为临床有效防治高尿酸血症引起的心室重构提供重要理论依据。

高尿酸血症与心衰关系密切，但高尿酸血症在心衰发生、发展中的确切作用及分子机制尚未明确，单纯高尿酸血症是否具有直接致心衰作用尚不清楚。自噬作为细胞自身质控的重要机制在心脏生理和病理状态下均发挥重要调节作用。本项目拟通过细胞水平和在体实验证实高尿酸血症可通过AMPK-mTOR信号通路激活心肌细胞自噬进而参与舒张性心衰的发生、发展。主要研究内容：1. 随时间延长，高尿酸大鼠心脏逐渐出现舒张功能减低，模型时间越长，舒张功能损害越明显，但直至延长至6个月，仍未发现明显收缩功能异常，即我们构建的动物模型为尿酸致舒张功能减低动物模型；2. 组织形态学发现尿酸组心室组织中凋亡细胞增多，心室纤维化程度明显，透视电镜发现自噬小体增多；3. 分离并测定单个心肌细胞舒缩功能，与正常大鼠心肌细胞相比，高尿酸组心肌细胞舒张功能明显减低，收缩功能未见明显异常；4. 与正常心肌组织相比，高尿酸大鼠心肌组织LC3BⅡ蛋白表达增加，且LC3BⅡ/LC3BⅠ比值增加；5. 培养大鼠原代心肌细胞，尿酸预处理，在此基础上针对AMPK-mTOR自噬信号通路的关键环节进行干预，结果发现高尿酸干预组心肌细胞，AMPK信号通路激活，抑制mTORC1磷酸化，磷酸化ULK1，启动自噬。科学意义：研究证实高尿酸可导致心脏出现舒张功能障碍，与尿酸诱导激活的细胞自噬关系密切，AMPK-mTOR信号通路是高尿酸激活细胞自噬的主要机制。本研究为临床有效防治高尿酸血症引起的心脏损害提供重要理论依据。