有氧运动通过miR-21/AMPK/Sirt1通路调控自噬改善衰老小鼠心肌纤维化的研究

关键词：衰老  心肌纤维化  自噬  有氧运动

Keywords:aging  myocardial fibrosis  autophagy  aerobic exercise

摘要：

      摘要 目的：探讨有氧运动对衰老诱导的心肌纤维化的改善作用及miR-21在其中的参与机制。 方法：将2月龄及19月龄雄性C57BL/6J小鼠随机分为青年对照组（Y）、青年运动组（YE）、老年对照组（A）和老年运动组（AE）。运动组小鼠接受12周的有氧跑台训练。运动干预结束24h后处死小鼠，收集其心脏组织。测量小鼠体重、心脏重量及心脏器官指数；HE染色观察心肌组织病理变化；Masson染色检测心肌纤维化水平；qPCR检测心肌纤维化及自噬标志蛋白、miR-21及其下游信号通路的mRNA表达水平，Western Blot检测其蛋白表达水平。 结果：①与Y组小鼠比，A组小鼠心脏重量、心脏器官指数显著增加（P<0.05），有氧运动干预后未出现明显变化。②衰老小鼠心肌细胞体积增大、肿胀、排列无序，而接受12周有氧运动干预的衰老小鼠心肌细胞肿胀得到改善，细胞排列整齐。③在心肌纤维化方面，衰老小鼠心肌组织出现损伤，蓝色胶原纤维沉积增多，纤维化标志物α-SMA、collagen-I表达明显增加（P<0.05），经过运动干预后心肌纤维化得到明显改善。④在心肌细胞自噬方面，衰老组小鼠心肌自噬蛋白ATG5、ULK1、Beclin1、LC3Ⅱ的表达水平均有所下降（P<0.05），运动干预后均出现显著升高（P<0.05）。⑤与Y组小鼠相比，A组小鼠miR-21的表达显著上升，下游基因AMPK、Sirt1、LKB1的含量明显降低（P<0.05），而运动干预后，AE组小鼠心肌组织miR-21的含量显著下降，AMPK、Sirt1、LKB1水平均显著增加（P<0.05）。 结论：有氧运动可以改善衰老诱导的心肌纤维化，其机制可能与抑制miR-21的表达，激活下游AMPK/Sirt1自噬信号通路有关。

Abstract:

      Abstract Objective： To investigate the beneficial effects of aerobic exercise on age-related myocardial fibrosis and to explore the involvement mechanism of the miR-21/AMPK/Sirt1 signaling pathway. Method： Male C57BL/6J mice aged 2-month-old (young) and 19-month-old (aged) were randomly divided into four groups: young control group （Y）， young exercise group （YE）， aged control group （A） and aged exercise group （AE）. Mice in the exercise group underwent 12 weeks of aerobic treadmill training. Mice were euthanized 24 hours after the exercise intervention， and heart tissues were collected. Measurements included body weight， myocardial weight， and myocardial organ index. Histopathological changes were assessed by hematoxylin-eosin (HE) staining, and myocardial fibrosis was evaluated using Masson staining. mRNA expression levels of fibrosis-related markers, autophagy-related genes, miR-21, and downstream signaling molecules were measured by quantitative PCR, and corresponding protein expression levels were assessed by Western Blot. Result： ①Compared to the Y group mice，aged mice showed significantly increased heart weight and heart weight index（P<0.05）， with no significant changes after aerobic exercise intervention. ②Aged mice showed cardiomyocyte hypertrophy, swelling, and disorganized arrangement， while these morphological abnormalities were improved after 12 weeks of aerobic exercise.③Myocardial fibrosis was significantly increased in aged mice, with increased deposition of blue collagen fibers and significant increase in the expression of fibrosis markers α-SMA and collagen-I （P<0.05）， but myocardial fibrosis was significantly improved after exercise intervention. ④The expression of myocardial autophagy proteins ATG5， ULK1， Beclin1， and LC3II was decreased in the aged group of mice （P<0.05）， but was significantly increased after exercise intervention （P<0.05）. ⑤The level of miR-21 in the A group significantly increased， while the expression of downstream genes AMPK， Sirt1， LKB1 significantly decreased （P<0.05）. After exercise intervention， the content of miR-21 in the AE group significantly decreased， while the content of AMPK， Sirt1， LKB1 significantly increased （P<0.05）. Conclusion： Aerobic exercise can improve age-related myocardial fibrosis， and the mechanism may be related to the inhibition of miR-21 expression and activating the AMPK/Sirt1 autophagy signaling pathway.