膳食氧化固醇通过调节肠道微生态诱导的阿尔茨海默发生发展的作用及机制

Dietary oxysterols, mainly 27-hydroxycholesterol (27-OHC), may increase the risk of developing Alzheimer’s disease (AD), a neurodegenerative disease characterized by cognitive disorder. However, the underlying mechanisms remain largely unclear. Our previous studies have shown that elevated level of 27-OHC in blood were associated with increased β-amyloid (Aβ) in brain and risk of memory impairment, alteration of intestinal bacteria density and variety, which is the most important. In this project, a nested case-control study on the basis of the prospective cohort established in 2014 will be used to evaluate the association among 27-OHC, diversity and DNA methylation of fecal bacteria, structure and function of intestinal mucosa, target protein for Aβ synthesis and clearance in brain and risk of cognitive disorders. Besides, to explore whether excessive 27-OHC affects the balance of intestinal microecology, disturbs Aβ metabolism through miRNA/TREM2-TLR2-APP signaling system, and eventually accelerates learning and memory loss, APP/PS1 transgenic mice and germ-free mice will be used to investigate the alteration of density and diversity of fecal bacteria, translocation of intestinal bacteria, permeability of intestinal mucosa barrier, Aβ metabolism in brain and neurobehavioral manifestation. The hypothesis that excessive oxysterols may induce Aβ metabolic disturbance in brain and promote AD through intestinal microecology will be investigated through population-based research and animal studies as well as high-throughput and bioinformatic technology. The results will provide more scientific evidence for dietary preventive strategies of AD.

膳食氧化固醇（主要是27-羟基胆固醇）与以认知障碍为特征的阿尔茨海默病（AD）有关，但机制不清。课题组前期研究发现过量27-羟基胆固醇（27-OHC）除可增加β-淀粉样肽（Aβ）水平和记忆障碍发生风险外，重要的是还与肠道细菌数量和种类有关。本项目在已有队列基础上，采用巢式病例对照研究，检测并分析27-OHC与粪便细菌多样性和DNA甲基化、肠黏膜屏障结构和功能、脑Aβ合成与清除相关靶因子及认知障碍的关系；用APP/PS1转基因鼠和无菌鼠，检测粪便细菌组成、肠道菌移位和定植、肠黏膜通透性、脑Aβ代谢及神经行为等，探讨过量27-OHC通过调节肠道微生态，干扰脑miRNA/TREM2-TLR2-APP信号系统，影响Aβ代谢，导致记忆减退的机制。通过人群及动物研究，采用高通量检测及生物信息学技术，验证过量氧化固醇通过调节肠道微生态，诱发脑Aβ代谢紊乱，促进AD发病的假说。为膳食预防AD提供科学数据。

膳食氧化固醇及肠道菌群与以认知功能障碍为特征的阿尔茨海默病（AD）/轻度认知障碍（MCI）发病有关，但研究结果尚有争议且作用机制不清。本项目通过巢式病例对照研究和转基因动物实验，探究了膳食氧化固醇及肠道菌群在阿尔茨海默病发病中的作用及其机制。主要研究结果：（1）人群巢式病例对照研究：与对照组相比，MCI组血清27-羟基胆固醇（27-OHC）等14种氧化固醇水平存在差异；肠道菌群α多样性和β多样性明显降低，肠道菌群构成在门水平和属水平存在显著差异；血清短链脂肪酸（丙酸、异丁酸、丁酸和戊酸）在MCI组显著升高，17种炎症因子在MCI组显著升高，27-OHC与多种炎症因子呈正相关；基因功能和代谢通路富集分析发现，JAK-STAT通路显著发生改变。基于差异氧化固醇、SCFAs和炎症因子的相关性网络分析显示，氧化固醇与肠道菌群代谢物和炎症反应在MCI中的重要作用。较高膳食指南指数（CDGI-2018）（RR: 0.75, 95%CI: 0.58, 0.98）和健康生活方式指数（HLS）（RR: 0.54, 95%CI: 0.34, 0.84）可降低MCI的发生风险，较高膳食炎性指数（EDII）可增加MCI发生风险（RR:1.46, 95%CI: 1.14, 1.87）。MCI组4种血清miRNA表达发生改变，血清let-7g-5p、miR-107和miR-186-3p的相对表达下调均可增加MCI的发生风险，肠道菌群α多样性指数Shannon和Simpson升高可降低MCI的发生风险。基于膳食质量、肠道菌群和miRNA相关指标建立MCI预测模型，可较好的预测MCI的发生，其灵敏度为0.67，特异度为0.95，曲线下面积AUC=0.88 (95%CI:0.82-0.94)。（2）动物研究：为揭示氧化固醇通过肠道菌群在认知功能减退中的作用及机制，分别采用混合氧化固醇饲料、27-OHC及其合成酶抑制剂（阿那曲唑）处理C57BL/6J野生型小鼠和APP/PS1双转基因小鼠，验证了氧化固醇是通过调节肠道菌群及其代谢物、血脑屏障和肠黏膜屏障结构与功能、促进炎症反应等作用机制最终导致学习记忆能力损伤。