恒定菌群移植抗焦虑效应及其调控苯二氮卓结合抑制蛋白DBI的机制

Fecal Microbiota Transplantation (FMT) provided new therapeutic option for some diseases, though the mechanism remains unclear. Based on our clinical research on FMT for more than 600 patients, we demonstrated that FMT treatment improves sleeping and relieves/cures anxiety after the effective remodeling of gut microbiota. Importantly, our exploring through urine proteomics displayed the higher level of diazepam-binding inhibitor (DBI) in patients with improvement of intestinal disease and anxiety. This difference was confirmed by blood ELISA analysis of DBI and larger sample size. Since previous studies have proved that the highly expression of TLR2, TLR4 and DBI in hippocampus are related to anxiety, we had enough evidence to have hypothesis that higher level of endogenous DBI could be induced by FMT and TLR2/TLR4 in hippocampus may critically involved with the regulation of DBI. Therefore the present study will base on clinical trials, animal models of mice with colitis related pain and anxiety and cultured microglia cells. The key methodologies include 16S rRNA analysis for intestinal microbiota, stable FMT, injection of TLR2/TLR4 shRNA lentivirus into hippocampus through stereotaxis, tests for anxiety-like behavior, tests for pain, patch clamp recording technique and et al. This study will evaluate the antianxiety effects of FMT and its mechanism of regulation on endogenous DBI. The findings should be theoretical evidence for clinical FMT and brain-gut-microbiota axis.

粪菌移植（FMT)为部分疾病提供了新的治疗机会，但是机制不清。本研究组前期在超过600例的FMT临床研究中意外发现：FMT有效重建肠道菌群之后，睡眠和焦虑得以有效治疗，尿液蛋白组学筛查和血液ELISA证实FMT增加了苯二氮卓结合抑制剂DBI水平。由于DBI和细菌模式识别受体TLR2与TLR4均在海马区影响焦虑样行为。遂提出假设：FMT通过“脑-肠”轴增加“安定”样神经肽DBI高表达，DBI结合γ-氨基丁酸受体产生抗焦虑效应，其中，海马区TLR2和TLR4可能调节DBI表达。拟在临床试验、肠炎合并焦虑动物模型以及体外培养小胶质细胞水平，通过16S rRNA菌群分析、恒定菌群移植、TLR2/TLR4 shRNA慢病毒颗粒海马区立体定向注射、焦虑行为学实验、疼痛反应试验、全细胞膜片钳技术等方法，研究FMT抗焦虑效应及其调控内源性DBI的机制，为临床FMT及“脑-肠-菌群”轴奠定理论基础。

接受本项目资助后，依据粪菌移植（fecal microbiota transplantation, FMT）治疗脑肠轴疾病的临床发现，以及肠道菌群研究领域内发生的变化，本研究组对研究计划进行了适当的调整，以确保在领域内的领先性。本研究组主要开展了以下研究：1、探索人源FMT对树鼩熬夜应激状态动物模型的影响，本研究表明FMT可以有效逆转应激后树鼩生理活动和脑区结构的变化，揭示了FMT是治疗神经精神疾病的潜在治疗方法。2、评估FMT治疗难治性癫痫的疗效与安全性，本研究表明FMT可以有效治疗难治性癫痫，且安全性较高。FMT未来将是难治性癫痫治疗的一个重要选择。3、首次通过结肠植管途径实现健康人肠道深部菌群的动态取样来揭示健康人肠道菌群的昼夜节律，为人类肠道菌群的变化提供了新的见解。