心肌营养素对阿尔茨海默病转基因模型小鼠脑组织能量代谢的作用及机制研究

The consistent and progressive cerebral glucose hypometabolism occurs during the very early stages of AD and the extent of hypometabolism correlates with symptom severity. On the other hand,the amelioration of energy metabolism is an effective way of preventing or reversing the cognitive deficits and attenuating the atrophy that is observed in AD. Cardiotrophin-1 (CTF1) has been reported to act as a trophic factor for a few neurons, such as sensory, cholinergic, dopaminergic, motor and cortical neurons. Studies have indicated that CTF1 delays degenerative disease progression in motor neuron disease. However, little is known about the effects of CTF1 on degenerative disease in the brain. We have shown that expression of CTF1 is strongly down-regulated in the brain of the APPswe/PS1dE9 transgenic mouse model of Alzheimer’s disease (AD). Transgenic mice with brain tissue-specific CTF1 expression alone or in combination with APPswe/PS1dE9 transgenic mice were produced to study the effects of CTF1 on AD. Our previous study indicated that CTF1×APPswe/PS1dE9 transgenic mice exhibited improvements in learning and memory and ameliorated disturbances of brain energy metabolism compared to APPswe/PS1dE9 transgenic mice.To explore the underlying mechanisms of CTF1 on brain enegy metabolism and cognitive improvement, the brain glucose metabolism and mitochondrial function are emamined.This study will support an important concept that onset of neurodegenerative disease may be delayed or mitigated with improvement of energy metabolism deficits.

脑糖代谢减退及能量紊乱早于阿尔茨海默病认知障碍的发生，并且减退程度与记忆损伤程度呈正相关；提高能量代谢的治疗策略和方法可以改善海马神经元突触可塑性，促进神经元发生，从而改善认知障碍,延缓甚至逆转神经退行性病变的进程。心肌营养素-1（cardiotrophin-1,CTF1）改善心肌能量代谢，延缓运动神经系统退行性病变，参与维持大脑神经元的存活，发挥营养和保护作用。将CTF1转基因小鼠与APPswe/PS1dE9阿尔茨海默病模型小鼠杂交，前期研究发现CTF1可以增加脑组织葡萄糖摄取，改善模型小鼠学习记忆能力。本项目系统检测双转小鼠脑糖代谢（包括糖摄取、糖利用以及糖转化）以及神经元线粒体功能活性，探讨CTF1对能量代谢作用的分子机制，阐明CTF1改善模型小鼠行为认知障碍的机制，为CTF1应用于临床治疗阿尔茨海默病提供实验依据，为临床阿尔茨海默病的治疗提供新思路和新方法。

本项目研究了CT-1对阿尔茨海默病（AD）模型小鼠脑组织能量代谢的作用及分子机制，阐明CT-1改善AD小鼠行为认知障碍的机制，探索AD早期行为认知障碍的发生机制，并对今后临床AD早期临床预防治疗提供新的策略。.首次构建神经元特异性CT-1过表达小鼠，与APPswe/PS1dE9阿尔茨海默病转基因小鼠杂交制备4种小鼠：野生型小鼠（WT）、AD小鼠（APPswe/PS1dE9）、神经元特异性CT-1过表达小鼠（CT-1）、CT-1×APPswe/PS1dE9小鼠。首先通过包括被动回避实验、新物体识别实验以及Morris水迷宫的多项行为学实验检测4和8月龄CT-1对AD小鼠行为认知能力的作用，结果显示4月龄AD小鼠出现行为认知损伤，损伤程度与月龄呈正相关。无论是在疾病早期还是中晚期，CT-1均可以改善AD小鼠的认知障碍。分离4和8月龄AD小鼠突触和非突触线粒体，发现AD小鼠无论是早期还是晚期突触线粒体均显示出显著性损伤，而非突触线粒体只有在疾病晚期检测到。为了探讨CT-1对AD疾病早期病变的保护性效应以及机制，我们选取4月龄小鼠，分离并检测突触线粒体功能，发现CT-1能够显著改善AD模型小鼠突触线粒体失功能。考虑到突触线粒体对突触功能维持的关键性作用，通过免疫荧光双染急性脑片，发现CT-1增加AD模型小鼠海马CA1区的突触密度。进一步研究发现，CT-1通过增加AD模型小鼠海马AMPK激酶活性，上调GluA1的表达。为了确认CT-1对突触线粒体以及突触的保护性效应是否是通过增加AMPK激酶活性，构建携带AMPKα shRNA的慢病毒沉默AMPKα，感染经Aβ处理的原代海马神经元，发现CT-1可以改善外源性Aβ引起的海马神经元突触线粒体失功能以及突触损伤，而AMPKα shRNA可以逆转CT-1对突触线粒体以及突触的阳性效应。.上述结果表明，CT-1通过增加AMPK活性，缓解早期AD突触线粒体功能损伤和突触失功能，从而改善AD行为认知障碍。本项目明确了CT-1对早期AD突触线粒体和认知功能的作用，从改善能量代谢的途径寻找新的营养因子，延缓甚至逆转神经退行性疾病进程的提供新方法和新思路。. 迄今，项目研究期间，以第一完成人发表SCI论文6篇，其余成果正在数据处理和论文撰写，预计还能2~3篇高水平SCI论文。