运动训练经SIRT3增强OPA1活性促进脑缺血大鼠线粒体结构修复及功能恢复

After cerebral ischemia, mitochondrial dysfunction can induce serious neurological damage and postischemic exercise training can promote the recovery of the mitochondrial function, but the underlying molecular mechanism remains incompletely understood. Our previous study has found that in ischemic rats, exercise training-induced OPA1 expression increase facilitates the repair of the mitochondrial structure and its functional recovery. In stress responses, other research has indicated that the role of OPA1 in the recovery of mitochondrial function involves not only its expression level but also its activity and the latter can be enhanced by SIRT3-induced deacetylation. In our preliminary study, exercise training can upregulate the SIRT3 expression in ischemic rats. Therefore, we speculate that in the cerebral ischemic model, exercise training can facilitate the repair of the mitochondrial structure and its functional recovery in cerebral ischemia via SIRT3-enhanced OPA1 activity. In this study, the cortical neurons of SD fetal rats and healthy male SD rats will be used to establish the cerebral ischemic model. The conventional molecular biology techniques, overexpression of target protein, RNA interference, molecular imaging technology and electron microscopy are used to explore the novel role of SIRT3-mediated OPA1 activity and its potential mechanisms in the repair of the mitochondrial structure and the recovery of the mitochondrial function after ischemic injury, and to ascertain whether treadmill exercise can promote mitochondrial structural repair and functional recovery via SIRT3-enhanced OPA1 activity in ischemic rats. This study will provide a new theoretical foundation for the clinical application of treadmill exercise.

脑缺血后线粒体功能障碍会导致严重神经功能损害。运动训练能促进脑缺血后线粒体功能恢复，但分子机制未完全明了。我们前期研究证实运动训练通过提高OPA1表达促进脑缺血大鼠线粒体结构修复及功能恢复。相关应激研究发现OPA1的促线粒体功能恢复作用与其表达水平和活性有关，后者与SIRT3诱导的去乙酰化紧密相关。已有前期研究发现运动训练能提高脑缺血大鼠SIRT3表达，因此本课题推测运动训练能通过提高SIRT3表达增强OPA1活性促进脑缺血大鼠线粒体结构修复和功能恢复。本项目拟用SD胎鼠皮层神经元及健康雄性SD大鼠建立体外和体内脑缺血模型，应用常规分子生物学技术、病毒过表达、RNA干扰技术、分子成像技术、电镜等方法探讨OPA1活性调节在脑缺血后结构修复和功能恢复中的作用及其机制。明确运动训练可否通过SIRT3增强OPA1活性促进脑缺血后线粒体结构修复和功能恢复，这将为脑缺血后运动训练的临床应用提供新依据。

缺血性脑卒中可引起线粒体功能障碍，从而导致严重神经功能损害。运动训练能促进脑缺血后线粒体功能恢复，但分子机制尚未完全明了。我们前期研究：运动训练能通过提高OPA1表达促进脑缺血大鼠线粒体结构修复及功能恢复。相关研究发现：OPA1促进线粒体功能恢复的作用与其表达水平和活性均有关，后者与SIRT3介导的去乙酰化作用密切相关。既往研究：运动训练能提高脑缺血大鼠SIRT3表达，但目前仍不明确运动训练能否通过提高SIRT3表达增强OPA1活性，从而促进脑缺血大鼠线粒体结构修复和功能恢复。本项目在已有研究基础上进一步探究了SIRT3在脑缺血后，运动促进线粒体结构修复和功能恢复的作用及机制。本课题在构建了皮层神经元氧糖剥夺体外脑缺血模型(OGD)和大鼠短暂性脑缺血模型(tMCAO)后，采用神经功能评分、磁共振、运动训练、免疫荧光、Western-blot、qPCR、透射电镜等方法发现:在体外和体内脑缺血后，SIRT3表达降低。在大鼠tMCAO后，运动训练通过增加SIRT3的表达，促进线粒体生物发生因子的表达和线粒体结构修复。此外，我们分别使用SIRT3过表达腺相关病毒和SIRT3过表达慢病毒调节大鼠脑缺血周边区和皮质神经元OGD损伤后的SIRT3的表达量。我们发现，大鼠皮层神经元OGD后，SIRT3过表达能减轻凋亡，促进能量代谢，抑制过度氧化应激。同时，大鼠tMCAO后，SIRT3过表达明显缩小脑缺血大鼠梗死体积，抑制神经元凋亡，并修复了线粒体结构；此外，促进了缺血周边区线粒体完整性的恢复及生物发生因子、线粒体DNA（mtDNA）的表达水平。而分别抑制OPA1的表达和活性显著降低SIRT3的上述效应。上述研究充分说明：运动训练能通过增加SIRT3的表达，来提高OPA1的表达水平，并增强其活性，从而促进大鼠脑缺血后线粒体结构修复及功能恢复。这些研究结果为运动训练及SIRT3在脑缺血的临床应用提供新的理论和实践依据。本研究成果共以通讯作者/第一作者/并列第一作者发表SCI论文4篇，本研究成果共以通讯作者/第一作者/并列第一作者发表SCI论文4篇，其中中科院一区SCI论文1篇，二区SCI论文2篇，三区SCI论文1篇；并获得第十六届福建青年科技奖、福建省科学技术进步奖各1项。