肠道菌群代谢物氧化三甲胺对血管老化的影响及其作用机制

Circulating level of the gut microbe-derived metabolite trimethylamine-N-oxide (TMAO) has recently been linked to many diseases associated with aging, especially atherosclerosis. An increasing body of evidence suggests that dietary supplementation of mice with TMAO can promote atherosclerosis . But the effects of TMAO on vascular aging have not yet been studied. We previously found that TMAO could promote the cell senescence and reduce the expression of SIRT1 in human umbilical vein endothelial cells(HUVECs) . To determine whether TMAO plays a direct contributory role in the development of vascular aging, the following of studies will be performed in the project. Firstly, the levels of the fasting plasma TMAO in different weeks of age of health rats and in health young and postadolescent people and in the eldly will be measured with metabolomics and the relationship between TMAO and aging will be analyzed. Secondly,the senescence-accelerated mice prone 8, the knock out FMO3 mice and the overexpression SIRT1 mice established by SIRT1 adenovirus will be fed with dietary TMAO supplementation for 16 weeks. The aging and its relevant factors in aorta of mice will be studied. Finally, HUVECs, overexpression SIRT1 HUVECs established by SIRT1 adenovirus and transiently knockdown p53 and p21 HUVECs established by p53 and p21 siRNA will be treated with TMAO. The senescence and its relevant factors of cells will be investigated. The aim of the study is to test the hypothesis that TMAO can suppress SIRT1 and induce p53/p21 protein expression to inhibit the cell cycle and induce vascular endothelial senescence which will promote vascular aging. The study will provide both the new theories and therapeutic targets for the delay and treatment of vascular aging and will have great significance in clinical medicine.

肠道菌群代谢物循环氧化三甲胺（TMAO）水平与许多衰老相关疾病如动脉粥样硬化（AS）有关，动物研究表明TMAO能加重AS，但能否促进血管老化尚未知。在我们前期实验发现TMAO促进培养人脐静脉内皮细胞（HUVECs）衰老并下调SIRT1表达基础上，本项目拟用代谢组学动态测定不同周龄健康大鼠和健康青壮年和老年人血浆TMAO水平；对快速老化小鼠、构建的FMO3基因敲除小鼠、SIRT1过表达的C57bl/6J小鼠分别喂食TMAO16周后，研究小鼠主动脉老化及其相关因子的变化；对HUVECs、构建的SIRT1过表达HUVECs和siRNA沉默p53、p21基因的HUVECs分别进行TMAO干预后，研究细胞衰老及其相关因子变化，以证明TMAO通过抑制SIRT1和上调p53和p21表达，使细胞周期停滞和血管内皮细胞衰老，而促进血管老化的假说。为延缓血管老化的研究提供新理论和治疗靶点，有重要意义。

肠道菌群与衰老关系研究是当前热点与重点，氧化三甲胺（TMAO）是重要肠道菌群代谢物，与许多心血管疾病相关，但与衰老关系尚未知。本项目探讨TMAO对血管衰老、脑老化作用及其机制，取得以下重要结果：（1）老年小鼠和老年健康人血浆TMAO水平分别高于年轻小鼠和健康青壮年和中年人。（2）动物实验：喂食 TMAO可促进正常老化小鼠SAMR1、加重和加快快速老化小鼠SAMP8主动脉衰老，其机制与TMAO引起主动脉氧化应激水平增加、抑制SIRT1表达而激活p53/p21/Rb信号通路有关； SIRT1激活剂白藜芦醇可减轻TMAO引起的小鼠主动脉衰老。（3）细胞水平： TMAO促进培养的人脐静脉内皮细胞（HUVECs）衰老，其机制与TMAO导致HUVECs ROS水平增加、SIRT1表达下调而激活p53/p21/Rb信号通路有关；白藜芦醇可上调HUVECs SIRT1、抑制p53/p21表达而减轻TMAO引起HUVECs衰老。（4）动物实验：喂食TMAO 可引起小鼠海马区神经元细胞衰老，机制可能是TMAO诱导小鼠脑组织氧化应激水平升高、破坏突触可塑性、损伤神经元超微结构，抑制mTOR/4EBP2/p70S6K 信号通路蛋白磷酸化，加快SAMR1小鼠、加重和加快 SAMP8 小鼠脑老化和认知功能障碍进程。本项目发现小鼠和健康人血浆TMAO随增龄衰老而升高；TMAO通过SIRT1/p53/p21信号通路促进血管衰老和血管内皮细胞衰老；SIRT1激活剂白藜芦醇可减轻TMAO引起的血管衰老；TMAO可能通过抑制mTOR/4EBP2/p70S6K 信号通路蛋白磷酸化诱导小鼠海马区神经元细胞衰老而促进小鼠脑老化。本项目已发表论文5篇（SCI 3篇），影响因子15.662，参加会议论文8篇。2019年《福建省血管衰老重点实验室》被福建省科技厅认定为30个省重点实验室之一。本研究为血管衰老及其相关疾病防治提供新思路和新靶点。