β-细辛醚逆转miR-30a表达进而激活MEF2D/PINK1/Parkin通路发挥抗帕金森病的分子机制

Parkinson's disease (PD) is the third biggest threat to the health of the elderly. The damage of dopaminergic neurons is the main cause of PD. And to find drugs for nutrition and repair of dopaminergic neurons are important to reverse or delay the progression of PD. Our previous studies have confirmed that β-asarone acts well on anti-PD activity and its mechanism is closely related to the key protein of mitophagy: MEF2D. Recent studies show that mitophagy dysfunction mediating MEF2D/PINK1/Parkin is the main cause of PD, which can induce the death of dopaminergic neurons. And down-regulated miR-30a expression can promote the expression of MEF2D. We found that miR-30a can target mitophagy receptor MEF2D. Therefore, we speculated that β-asarone may regulate mitophagy to improve the clinical symptoms of PD through miR-30a/MEF2D/PINK1/Parkin signal transduction. This study intends to carry out in vivo and in vitro experiment by applying miR-30a gene overexpression and silencing technology, transmission electron microscope, confocal microscope, flow cytometry, molecular biology and other evaluation methods. We will further investigate the influence of β-asarone on miR-30a as well as on MEF2D/PINK1/Parkin mediated mitophagy, and clarify the mechanism of β-asarone on repairing dopaminergic neurons damage in its anti-PD effects. This study not only provides experimental and theoretical basis for the development of new anti-PD drugs by β-asarone, but also provides new ideas for treating PD.

帕金森病（PD）是威胁老年人健康的第三大杀手，DA能神经元受损是其主要病因，营养和修复DA能神经元药物对逆转或延缓PD病程具有重要意义。前期研究证实β-细辛醚具有显著的抗PD活性，其作用与线粒体自噬受体关键蛋白MEF2D密切相关。最新研究表明，MEF2D/PINK1/Parkin介导的线粒体自噬出现异常，导致DA能神经元死亡，而下调miR-30a会促进MEF2D表达。因此我们推测：β-细辛醚可能通过miR-30a调控MEF2D/PINK1/Parkin信号转导调节线粒体自噬进而改善PD临床症状。本课题拟建立PD体内外模型，采用基因超表达和沉默技术、电镜、激光共聚焦等多种评价方法，深入探讨β-细辛醚与miR-30a及MEF2D/PINK1/Parkin线粒体自噬通路的关系，阐明其修复受损DA能神经元的分子机制。本研究不仅为β-细辛醚开发成抗PD新药提供实验和理论依据，也为防治PD提供新思路。

帕金森病（PD）作为一种典型的老年慢性疾病，是严重影响人类健康的第二大神经退行性疾病。临床上对PD采用对症治疗，只能缓解临床症状并不能阻止神经元退化，以DA类药物替代治疗为主，尚无有效的治疗手段。因此，亟需深入开展新型、高效、低毒副作用的天然抗PD药物的开发研究。传统中药石菖蒲的主要活性成分β-细辛醚在抗PD药物研发中，能够明显改善多种PD模型动物的运动功能障碍，然而β-细辛醚对线粒体自噬作用机制不清，作用靶标尚未明确。本研究采用6-OHDA所诱导的PD大鼠模型，明确了β-细辛醚对线粒体和miR-30a的影响，β-细辛醚可减轻氧化应激因子水平，上调神经营养因子水平从而减轻线粒体损伤，相关机制可能与其调控了miR-30a和MEF2D/PINK/Parkin通路相关。在此基础上，采用6-OHDA诱导的SN4741细胞，通过蛋白印迹、PCR、激光共聚焦和荧光素酶报告基因检测鉴定miR-30a与MEF2D的关系等，揭示MEF2D是miR-30a的靶蛋白之一，miR-30a的激活会抑制MEF2D的表达。调控机制上，β-细辛醚提高细胞中mfn1、mfn2、opa1、P62和p-MEF2D水平，降低PINK，drp1、VDAC1、Parkin mRNA和PINK mRNA水平。综上所述，本项目发现了β-细辛醚通过调控miR-30a的表达对MEF2D/PINK1/Parkin 通路进行调节，激活线粒体自噬，改善DA能神经元的线粒体功能，从而发挥抗PD作用。