耐力训练及饮食限制影响心肌Pink1/Parkin线粒体自噬信号并提高自噬水平
Endurance Training and Dietary Restriction Influence the Myocardial Pink1/Parkin Mediated Mitophagy Signals and Promote Autophagy

摘要：目的：观察耐力训练和饮食限制对小鼠心肌Pink1、Parkin及Drp-1的影响，探讨运动和饮食干预心肌Pink1/Parkin介导的线粒体自噬分子机制。方法：8周雄性C57BL/6小鼠随机分为安静对照组（C组）、耐力训练组（T组）、饮食限制组（D组）及耐力训练加饮食限制组（TD组），8只/组。C组不做运动，自由饮食；T组进行10周游泳训练，1次/d，5 d/周，第1周20 min/次，每周增加10 min, 第8周维持在90 min/次直至实验结束；D组进行40%饮食削减；TD组进行耐力训练加饮食限制，方案分别与T、D组同。实验结束后取心肌，RT- PCR和Western blot技术检测Pink1、Parkin和Drp-1的mRNA及蛋白表达，透射电镜观察心肌细胞自噬发生情况。结果：1)与C组比较，T组体重下降幅度较小，心脏重量无显著变化(P>0.05)，心系数显著升高(P<0.01)；而D、TD组体重和心脏重量均显著下降 (P<0.01)。2)T组心肌自噬体增多，仅有Pink1 mRNA和蛋白表达增高(P<0.05)；D组心肌自噬体增加，仅有Drp-1蛋白表达增加(P<0.05)；而TD组心肌自噬体增加最明显，同时线粒体受损严重，Parkin mRNA显著下降(P<0.05)，但其蛋白无显著变化(P>0.05)，而Pink1、Drp-1 mRNA及蛋白表达显著增加(P<0.05)。结论：耐力训练或饮食限制均能提高小鼠心肌自噬水平，耐力训练提高Pink1表达来增强Pink1/Parkin介导的线粒体自噬信号；饮食限制提高Drp-1蛋白表达，协助心肌线粒体自噬发生；耐力训练加饮食限制导致心肌线粒体等结构病变严重，Pink1和Drp-1蛋白表达提高是高水平线粒体自噬发生的机制。
Abstract: Objective: This study investigated effects of endurance training and dietary restriction on myocardial Pink1, Parkin and Drp-1 expressions, and discussed molecular mechanism of Pink1/Parkin mediated mitophagy regulated by exercise and diet. Methods: Eight weeks male C57BL/6 mice were randomly assigned to 4 groups (n=8): control group (C group), endurance training group (T group), dietary restriction group (D group) and endurance Training+dietary restriction group (TD group). T group undertook 10 weeks increased load swimming training, 1 time/day, 5 days/week. They swam 20 min in the 1st week, and increased 10 min per week and finished 90 min in the last 3 weeks. D group was suffered 40% food reduction. TD group undertook the same endurance training and dietary restriction. Cardiac muscle was collected after experiment, the mRNA and protein expressions of Pink1, Parkin and Drp-1 were tested by RT-PCR and Western blot, and the autophagy in myocardial cells was observed by transmission electron microscope. Results: 1) Compared with C group, T group’ body weight decreased less, heart weight had no difference (P > 0.05), heart coefficient increased significantly (P < 0.01); D and TD groups’ body weight and heart weight decreased (P < 0.01). 2) Cardiac autophagosome of T group increased, and the mRNA and protein expressions of Pink1 increased (P < 0.05); cardiac autophagosome of D group increased, and the protein expression of Drp-1 increased (P < 0.05); cardiac autophagosome of TD group increased most obviously, mitochondria was damaged seriously, and the mRNA expression of Parkin increased (P < 0.05), but Parkin protein expression didn’t change (P > 0.05), the mRNA and protein expressions of Pink1 and Drp-1 increased (P < 0.05). Conclusions: Endurance