基于差异蛋白质组学的罗非鱼脂肪肝发病机制及药物干预研究

Fatty liver disease seriously endangers the health of farmed fish as common disease in tilapia farming process. It is complicated by a variety of diseases and causes mass mortality in fish. However, pathogenesis of fatty liver diseases has not yet been completely elucidated. The functional expression for tilapia fat metabolism-related enzyme genes was founded in lesion process. Moreover, it proved that tilapia fatty liver lesion has changed according to differential proteomics. On these bases, this topic intends to replicate and establish tilapia fatty liver disease models and drug intervention by feeding high-energy high-fat diet. It will reveal genome-related protein species as well as its amount in a liver fat condition expression and existence according to proteomics, in order to filter and establish differential protein profiles library for tilapia liver lesions. This study aims at the analysis of identified the main metabolic and signal transduction pathways involved in (differential) proteins. And it uses RT-qPCR technology quantitative analysis the functional expression in the fatty liver lesions and drug intervention processing on fat metabolism-related enzyme gene and (difference) protein genes. From the perspective of molecular biology, it confirms differential proteomics experimental results, elucidates tilapia fatty liver disease and drug protection mechanisms at the molecular level. In the mean time, it provides new ideas and new targets for prevention and treatment of fatty liver disease.

脂肪肝是严重危害养殖鱼健康的疾病，也是罗非鱼养殖过程常见的疾病，易并发多种疾病引起鱼类大量死亡，机制尚未彻底阐明。在我们发现罗非鱼脂肪代谢相关酶基因在脂肪肝病变过程中的功能性表达，并证明罗非鱼脂肪肝病变中差异蛋白质组学存在变化的基础上，本课题拟用高脂高能饲料喂养复制建立罗非鱼脂肪肝病模型及药物干预，通过蛋白质组学技术揭示基因组相关蛋白质种类和数目在脂肪肝病理状况下的表达和存在方式，筛选并建立罗非鱼肝脏组织病变的差异蛋白质图谱库，针对分析鉴定到的（差异）蛋白参与的主要的代谢和信号转导途径；并利用RT-qPCR技术定量分析脂肪肝病变及药物干预过程中脂肪代谢相关酶基因及（差异）蛋白基因的mRNA功能性表达，从分子生物学角度印证差异蛋白质组学实验结果，从分子水平阐明罗非鱼脂肪肝发病及药物保护作用机制，为脂肪肝病的防治提供新思路、新靶点和新药物。

由饲料营养失衡、养殖环境等引发的脂肪肝病严重危害养殖鱼类的健康，也是罗非鱼养殖过程常见的疾病。脂肪肝病易并发多种疾病引起鱼类大量死亡，机制尚未彻底阐明。项目首先通过喂食高脂饲料稳定诱导罗非鱼脂肪肝病模型，并对模型进行了生长发育、组织、生理生化多层次的评估。一方面，通过蛋白质组学解析脂肪肝病发生的分子基础，进一步构建了高脂诱导、垂盆草提取物干预的蛋白质组谱图，并系统性分析了致病及干预涉及的差异蛋白所参与的功能类别、信号和代谢通路、蛋白-蛋白相互作用网络。另一方面，通过转录组测序，关联分析了致病及干预涉及的基因表达差异，筛选分析了相关的基因功能。对关键蛋白进行了靶向蛋白质组学验证、基因转录表达水平的验证。研究表明，PPAR信号通路及其调控的脂肪消化及转运等代谢过程能够受长期高脂喂食抑制，而饲料中垂盆草提取物的适量添加能够有效维持鱼体脂肪代谢、氧化还原以及免疫反应的平衡。该项目研究阐明罗非鱼脂肪肝发病及药物保护的潜在作用机制，为脂肪肝病的防治提供新思路、新靶点和新药物。