电针基于AMPK/mTOR通路调节颅脑损伤后自噬与康复新机制

Traumatic brain injury (TBI) has a high rate of morbidity and mortality, which brings a large burden of economic to families and society. The key of treatment is to prevent excessive autophagy and apoptosis of brain nerve cells, and to promote the rehabilitation of cerebral nerve function. The clinical curative effect of electroacupuncture treatment is exact, but the mechanism is unknown. In the previous study,our research group mainly from the mitochondrial apoptotic pathway to clarify electroacupuncture adjustable part factors that can affect the recovery of brain neurons，but autophagy and its key signaling pathway AMPK/mTOR are closely related to the recovery of neuronal function after craniocerebral injury, and after consulting the literature, there is no study on mechanisms of treatment about acupuncture on TBI from the AMPK/mTOR pathway and autophagy at present. We believe that “electroacupuncture regulate autophagy of brain neurons and promote TBI rehabilitation after TBI via AMPK/mTOR pathway” is a scientific hypothesis. TBI model of rats were used as the research object, selecting "Quchi,Hegu, Baihui,Guanyuan，Zusanli, Yongquan "as the acupoints of acupuncture therapy, using behavioral evaluation methods, electron microscopy and molecular biology etc. Dynamically observe the regulating effects of electroacupuncture on autophagy related proteins and the key proteins of downstream of AMPK/mTOR pathway and their genes of TBI rats.It is expected to elucidate the mechanism of electroacupuncture for promoting rehabilitation of TBI, provide scientific experimental basis for extensive clinical application and studies of the treatment of electroacupuncture on TBI.

颅脑损伤（TBI）具有很高的致残率和致死率，给家庭及社会带来了沉重的经济负担，治疗的关键是阻止脑神经细胞过度自噬及凋亡，促使脑神经功能康复。电针治疗临床疗效确切，但机制不明。课题组前期研究主要从线粒体凋亡途径阐明电针可调节影响脑神经元康复的部分因素，而细胞自噬及其关键信号通路AMPK/mTOR与颅脑损伤后神经元功能康复密切相关，经查文献，目前尚无从本通路及自噬研究电针治疗TBI的机制。本课题以“电针可能通过AMPK/mTOR通路调节脑神经细胞自噬，促使TBI康复”为科学假说。以TBI大鼠模型为研究对象，采用“曲池、合谷、百会、关元、足三里、涌泉”穴电针治疗，利用行为学评价方法，电子显微镜、分子生物学等现代检测技术，动态观测电针对TBI大鼠AMPK/mTOR通路关键蛋白、自噬相关蛋白及其基因的调节作用。有望阐明电针促TBI康复的作用机制，为临床应用和推广针灸治疗TBI提供科学的实验依据。

颅脑损伤（traumatic brain injury，TBI）具有很高的致残率和致死率，给家庭及社会带来了沉重的经济负担，治疗的关键是阻止脑神经细胞过度自噬及凋亡，促使神经功能康复。电针治疗临床疗效确切，但机制不明，而细胞自噬及其关键信号通路AMPK/mTOR与颅脑损伤后神经元功能康复密切相关，因此，我们假设电针对TBI的治疗作用与其抑制AMPK/mTOR介导的自噬有关。SD大鼠被随机分为三组：假手术组、TBI组和TBI+EA组。使用eCCI制备TBI大鼠模型，大鼠在造模后12小时接受电针治疗，每天15分钟，连续14天。电针选取曲池、合谷、百会、关元、足三里和涌泉穴位，采用疏密波，频率为1Hz，振幅为1mA。造模后3d、7d、14d，mNSS、转棒疲劳、MWM试验表明电针治疗减轻了TBI大鼠脑水肿、病理损伤和神经元凋亡，促进了神经功能恢复。TEM观察电针改善了TBI大鼠异常形态的线粒体超微结构和增加的自噬体以及ATP、ADP、AMP的浓度。Western blot和IHC检测IL-10、自噬相关蛋白和AMPK/mTOR通路关键蛋白。研究表明：电针治疗诱导IL-10的产生，抑制了AMPK/mTOR信号通路转导和过度自噬。此外，AMPK抑制剂的效果与电针相似。AMPK激动剂和IL-10中和抗体均逆转了电针对自噬相关蛋白和AMPK/mTOR信号通路的影响。综上所述，电针通过诱导IL-10的产生和阻断AMPK/mTOR通路抑制过度自噬，促进了TBI大鼠的神经功能恢复，减轻了病理损伤和神经元凋亡。阐明了电针促TBI康复的部分机制，为临床应用和推广针灸治疗TBI提供科学的实验依据。