Nesfatin-1通过肝-脑轴固有巨噬细胞miRNA-34a-TREM2/DAP12-Wnt/β-catenin信号通路调控NAFLD大鼠的脂质代谢紊乱及神经精神行为异常

Non-alcoholic fatty liver disease (NAFLD) is one of the most common cause of chronic liver disease, resulting in not only the liver dysfunction, glucose and lipid metabolism disorder, but also the neuropsychiatric injuries. Results of our previous studies suggested that consuming of a high-fat diet for 4 consecutive weeks could successfully induce a NAFLD rat model, as indicated by obesity, hyperlipemia, hyperglycemia, decreased plasma levels of leptin and insulin, and inflammation and mild hepatocyte steatosis in the liver. Moreover, the plasma nesfatin-1 concentration was increased in the NAFLD rats and negatively correlated with the learning and memory ability. In the present study, a NAFLD rat model will be established by fed with high-fat diet. The behavior performance will be observed dynamically via the open field test (OFT), the elevated plus maze (EPM), the sucrose preference test (SPT), the forced swimming test (FST), and the Morris water maze (MWM) 4, 8, 12, and 16 weeks later. Meanwhile, the plasma concentrations of alanine aminotransferase (ALT), glucose, free fatty acid (FFA), total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), nesfatin-1, soluble triggering receptors expressed on myeloid cells 2 (sTREM2) will be detected , and the expressions of micro-RNA 34a, TREM2, DNAX-activating protein12 (DAP12), p-glycogen synthase kinase-3β (GSK-3β), GSK-3β, p-β-catenin, β-catenin, and cyclin D1 in the liver, hypothalamus, hippocampus and PFC will be detected. Moreover, these parameters will be measured after changing the nesfatin-1 abundance via over-expression or gene knockout. Through the combination of experiments in vivo and in vitro, the aim of the present study is to prove our hypothesis: nesfatin-1 plays an important role in the dysfunction of lipid metabolism and neuropsychiatric behaviors in NAFLD rats, the mechanism of which might be associated with the imbalance of the miRNA-34a-TREM2/DAP12-Wnt/β-catenin signaling pathway in the resident macrophages of the liver-brain axis.

本课题组前期研究表明高脂饮食可诱导大鼠表现出与摄食抑制因子nesfatin-1血浆浓度上升相关的脂代谢紊乱和神经行为学异常。本项目拟采用高脂饮食诱导大鼠非酒精性脂肪性肝病（nonalcoholic fatty liver disease, NAFLD）模型，在4、8、12和16周，通过生物样品检测和神经行为学实验，动态观察nesfatin-1丰度改变在NAFLD大鼠脂质代谢、肝功能、情绪及学习记忆功能改变中的作用，着重探讨肝-脑轴固有巨噬细胞miRNA-34a-TREM2/DAP12-Wnt/β-catenin信号通路的功能。在此基础上采用基因干预手段，结合细胞培养及分子生物学实验方法，从整体、细胞和分子水平验证理论假设：Nesfatin-1通过肝-脑轴固有巨噬细胞miRNA-34a-TREM2/DAP12-Wnt/β-catenin信号通路调控NAFLD大鼠脂质代谢紊乱及神经精神行为异常

非酒精性脂肪肝（NAFLD ）当前全球最重要的公共健康问题之一，其表现出的脂质代谢紊乱、肝脏形态和功能异常以及神经精神损伤严重影响人类健康。本项目通过建立整体动物模型及细胞模型，系统观察了非酒精性脂肪肝（NAFLD）状态下nesfatin-1丰度改变及其与糖脂代谢及神经行为学改变之间的关联，并探索性研究了Resveratrol及Quecetin的治疗作用及可能机制，为NAFLD的干预治疗提供了新的可能靶标。通过临床观察、整体动物模型及细胞模型，系统观察了2型糖尿病（T2DM）神经精神损伤的表现并探索性研究了其发生机制，在验证了糖化血红蛋白（HbA1c）对T2DM病情判定的重要性基础上，发现Nesfatin-1、VitD及TREM1/2丰度改变与T2DM的神经精神损伤密切相关，原创性提出nesfatin-1及25(OH)D3水平可作为辨识T2DM认知功能损伤的潜在标志物；初步揭示了炎症及免疫功能功能失调、昼夜节律紊乱在NAFLD、T2DM等代谢性疾病神经精神损伤机制中的作用；进一步建立多种整体动物模型，系统观察了情绪及学习记忆受损动物海马及前额叶皮层的突触可塑性改变及BNDF/TrkB/Akt/ERK/CREB信号通路关键分子表达的变化，明确了以海马小胶质细胞激活及炎症因子分泌增加为主要表现的炎症及免疫反应激活起重要作用，探索性发现侧脑室注射骨髓间充质干细胞来源的外泌体及RES灌胃给药均可改善海马立体定位注射STZ诱导的小鼠神经精神损伤；Melatonin可通过调节昼夜节律及炎症反应改善实验动物的糖脂代谢紊乱及神经精神行为异常。整体动物模型及临床观察结果提示，外周血炎症因子水平升高与抑郁症的发生密切相关，同时伴有外泌体丰度及miRNA表达谱的改变，其中IL-6有望成为诊断及疗效评定的潜在标志物。IL-6受体拮抗剂Tocilizumab可通过调节炎症及免疫反应改善实验动物的昼夜节律紊乱及神经精神行为异常。本项目的实施有助于科学解释抑郁症及NAFLD等代谢性疾病神经精神损伤的发生机制，并为治疗药物开发提供实验依据。