艾灸对APP/PS1双转基因AD小鼠线粒体能量代谢和凋亡通路的影响及作用因素研究

Moxibustion as one of the three main traditional Chinese medicine therapies could reinforce original Qi, tonify the kidney and secure the origin. While Alzheimer’s disease (AD) is one of the most common neurodegenerative diseases in central nervous system closely related to aging, in which disease mitochondrial dysfunction plays an important role. The dysfunction of the mitochondrial could result in the rise of oxidative stress state, metabolic disturbance of energy and apoptosis of the nervous cells, thus increasing the generation of pathological products Aβ in the cerebral cortex and hippocampus that eventually causes progressive cognitive impairment and loss of memory. Our previous study found that moxibustion could reduce abnormal deposition of neurotoxic Aβ in the brain of AD mice model, strengthen excitation of CNS and enhance learning capability of aging mice. According to these, we hypothesize that moxibustion may exhibit neuroprotective effect by means of protecting mitochondria, lowering the oxidative stress state and improving metabolic disturbance of energy mitochondria. Our research will observe the effect of moxibustion on mitochondrial oxidative stress state, energy metabolism and apotosis pathyway in APP/PS1 transgenic Alzheimer disease mouse model, and aimed at clarifying the mechanism of moxibustion on delaying the onset or slowing the progression of AD and making a further study on different effects of moxibustion heat and moxa smoke in this disease.

艾灸疗法是祖国医学三大主流疗法之一，具有补益元气、培肾固本的功效。阿尔茨海默病(AD)是一种与年龄增长密切相关的中枢神经系统退行性疾病。线粒体功能障碍是该病发生发展的重要原因之一，其主要通过诱导机体氧化应激水平升高、能量代谢紊乱及神经细胞凋亡，导致大脑皮质和海马内AD病理产物Aβ生成增多，引发渐进性认知障碍和记忆散失。前期研究发现艾灸具有减少AD模型小鼠脑内神经毒性Aβ异常沉积，增强中枢神经系统兴奋性，改善小鼠学习记忆能力的作用。据此，我们认为艾灸可能是通过保护线粒体功能，降低机体氧化应激水平，改善能量代谢紊乱，来保护神经细胞。本课题拟通过观察艾灸对APP/PS1双转基因AD小鼠氧化应激状态、线粒体能量代谢及凋亡通路产生的影响，深入阐明艾灸延缓AD发生发展的作用机制，并进一步探讨艾燃烧生成物及温热刺激在其中发挥的作用异同。

本课题观察艾灸不同作用因素对阿茨海默病的影响，从学习记忆行为、脑组织线粒体功能、氧化应激状态、尿液代谢物变化、细胞凋亡线粒体途径角度探索艾灸、艾烟、无烟灸的作用机制。. 将5月龄C57BL/6小鼠作为空白对照组（20只/组），同龄80只APP/PS1小鼠随机分为模型组、艾灸组、无烟灸组、艾烟组（20只/组）。艾灸与无烟灸组灸治小鼠关元穴。艾烟组小鼠束缚暴露于10～15mg/m3烟气环境，空白与模型组常规抓取、固定。所有干预20min/天，6天/周，共8周。检测小鼠脑组织线粒体功能、尿液代谢物变化、细胞凋亡线粒体途径等指标。. 结果显示：. 1.与空白组比，模型组多项行为学成绩降低。艾灸组在自主行为测试和避暗实验中成绩较模型组升高。. 2.与空白组比，模型组前额叶、海马ROS、8-OhdG、4-HNE、3-NT含量升高，ATP含量、线粒体呼吸链复合物活性下降，海马淀粉样斑块较多；与模型组比，艾灸不同作用因素干预后小鼠脑组织ROS、8-OhdG、4-HNE、3-NT含量显著降低，ATP含量升高、线粒体呼吸链复合物活性增强，海马淀粉样斑块减少。. 3.模型组与艾灸组尿液差异代谢产物有15种，主要涉及碳水化合物、氨基酸的合成与分解等代谢途径。. 4.模型组脑组织Bax、Caspase-3、Caspase-9表达升高，Bcl-2表达下降；与模型组比，艾灸不同作用因素干预后小鼠Bax、Caspase-3、Caspase-9表达减少，Bcl-2表达升高。. 结论：. 1.艾灸、艾烟、无烟灸均可提高AD模型小鼠中枢神经系统兴奋性，增强运动能力，恢复记忆功能，且艾灸比单纯艾烟干预或温热刺激效应更强。. 2.艾灸、艾烟、无烟灸可以通过调节线粒体呼吸链功能，减轻氧化应激损伤；抑制细胞凋亡线粒体途径相关蛋白表达，以保护神经元，进而延缓AD病理进程。艾烟干预对线粒体功能的调节作用比温热刺激更强，艾烟干预与温热刺激对细胞凋亡线粒体途径上游蛋白作用更明显。. 3.艾灸、艾烟、无烟灸能够调节AD模型小鼠体内氨基酸、淀粉蔗糖代谢、戊糖和葡萄糖醛酸酯相互转化、氮代谢以及氨酰生物合成等代谢途径中相关生物标志物含量。