肠道菌群代谢物氧化三甲胺通过SIRT3调控 PINK1-Parkin依赖性线粒体自噬诱导脑老化

The intestinal flora plays an important role in the metabolic regulation of the body, but the specific mechanism is not clear. Studies have confirmed that the intestinal flora metabolite trimethylamine (TMAO) is closely related to the development of many diseases. However, whether it is related to the body aging is still poorly understood. In our previous study, we have found that TMAO levels of healthy people in plasma were raising with ages, and exogenous TMAO could destroy the mitochondrial structure in hippocampus and induce neuronal senescence and cognitive dysfunction in mice. These results suggest that TMAO plays an important role in brain aging. Until recently, there are still no reports of TMAO promoting brain aging mechanisms. Based on these discoveries, we hypothesized that TMAO will regulate PINK1-Parkin-mediated mitochondrial autophagy, and further interfere with the cleaning of damaged mitochondria, and induce neuronal senescence finally through down-regulating the SIRT3/FOXO3/PINK1-Parkin signaling pathway. To prove this hypothesis, this research will clarify the relationship between circulating TMAO levels and cognitive function, and whether TMAO can inhibit SIRT3/FOXO3/PINK1-Parkin signaling pathway and PINK1-Parkin-mediated mitochondrial autophagy afterwards, at molecular, cellular, and tissue levels and through animal experiments, which will induce neuronal senescence and cognitive dysfunction and promote brain aging. This study is of great significance because of exploring the mechanism of brain aging and related cognitive dysfunction from the totally new viewpoint of TMAO, providing a potential target for delaying brain aging.

肠道菌群在机体代谢调控中具有重要作用，但具体机制暂不明确。研究证实肠道菌群代谢物氧化三甲胺（TMAO）与许多疾病发生发展密切相关，但是否与机体衰老相关仍知之甚少。我们前期研究证实，健康人血浆TMAO水平随增龄而增加，外源性TMAO能损伤线粒体，诱导小鼠脑老化和认知功能障碍。结果提示TMAO在机体脑老化中起重要作用。目前未见TMAO促进脑老化机制的报道。据此我们提出假说：TMAO下调SIRT3/FOXO3a，调控PINK1-Parkin依赖性线粒体自噬，干扰损伤线粒体清除，诱导神经元衰老。为验证这一假说，本课题将从分子、细胞，组织和动物水平等多层次阐明血浆TMAO与认知功能相关关系；揭示TMAO通过SIRT3/FOXO3，调控PINK1-Parkin依赖性线粒体自噬，诱导神经元衰老，促进脑老化和认知障碍。本课题从TMAO这个新视点探讨脑老化的发生机制，为延缓脑老化提供新靶点，具有重要意义。

预防脑老化，延缓认知功能障碍是当下临床关注的热点。肠道菌群代谢物氧化三甲胺（TMAO）可诱导小鼠脑老化及认知障碍，但具体机制尚不清楚，本项目主要从临床、细胞和动物等方面进行探讨。.研究内容一：分析老年人血浆TMAO与认知功能障碍关系：收集福建医科大学附属协和医院志愿加入研究的老年MCI患者和健康老年人空腹血浆，行血浆TMAO含量测定。共纳入138例志愿者，健康老年人76例，老年MCI62例。结果：老年MCI组血浆TMAO水平（12.82±0.4369μmol/L）高于健康老年人组（10.78±0.4628μmol/L）（p<0.001）。 .研究内容二：体外观察TMAO通过SIRT3调控线粒体自噬诱导人脑微血管内皮细胞（HBMECs）衰老的作用机制：以体外培养的HBMECs为研究对象，通过重组腺病毒感染构建SIRT3过表达细胞。分组：①对照组；②TMAO组；③SIRT3过表达+TMAO组：AdSIRT3+TMAO组。TMAO组和AdSIRT3+TMAO组都分别予以TMAO（80nmoL/L，7d）干预。结果显示：（1）TMAO干预后，衰老的HBMECs明显增加，SIRT3表达下降，P21、P53表达均明显增加，细胞内的正常线粒体数量明显减少，线粒体结构被破坏，双层嵴不明显甚至消失。（2）构建SIRT3过表达细胞，再给予TMAO（80nmoL/L，7d）干预，衰老的HBMECs明显减少，P53、P21和P16表达均明显低于TMAO组；细胞免疫荧光染色和透射电镜下，均观察到自噬小体明显增多，线粒体结构较TMAO组清晰；线粒体自噬相关蛋白Pink1、Parkin和LC3表达均明显高于TMAO组，但P62表达无显著变化。.研究内容三：在 SIRT3 过表达小鼠模型中验证 TMAO 通过 SIRT3 调控线粒体自噬诱导脑老化和认知障碍的作用机制：快速老化小鼠（SAMP8），6月龄，随机分为对照组、TMAO干预组和TMAO+RSV组，干预16周。结果显示：（1）随周龄增加,各组小鼠体质量无显著性差异，但干预结束时，Con组小鼠的体质量明显高于其他两组（p<0.01），但TMAO组和RSV组无显著性差异。（2）RSV可改善小鼠因TMAO诱导的学习认知障碍，减少海马区的衰老细胞。.本项目研究成果将为脑老化的研究提供新的靶点和理论依据，对脑老化的防治提供新的策略，具有重要的意义。