基于BP人工神经网络探讨PPARs 基因多态性与团头鲂营养性脂肪肝病易感性的关系

Nutritional fatty liver has become common diseases in aquaculture. Peroxisome proliferator-activated receptors (PPARs) is very important in the glucose and lipid metabolism. Its polymorphic sites are closely related to the susceptibility of nutritional fatty liver according to the reports of human and rodents. Therefore, in this study, we will describe the association between PPARs polymorphic sites and genetic susceptibility of nutritional fatty liver in Megalobrama amblycephala. For this purpose, according to full-length cDNA of PPARs, their polymorphic sites will be determined by PCR-SSCP and sequencing.We will develop Megalobrama amblycephala’s susceptible group and unsusceptible group of nutritional fatty liver,then, a correlation between genetic susceptibility of nutritional fatty liver disease and different genotypes of these polymorphic sites will be developed by back propagation artificial neural network model, logistic regression and multifactor dimensionality reduction method according to the genotype of each Megalobrama amblycephala, their pathologic diagnosis of nutritional fatty liver and other related data. It is also studied that effects of the interactions among genes or gene-environment on the risk of nutritional fatty liver disease.Then, by real time PCR analysis and Western blot analysis, we will detect the expression levels of mRNA and protein, which are the key enzymes related to fatty liver formation and target genes of PPARs. In addition, we will also detect biochemical indexes in blood and liver tissues. At last, we will expound the mode by which different genotypes of PPARs how to regulate lipid and glucose metabolism. Thereby this project will pave the way for clarifying the causation of nutritional fatty liver which was caused by genetic variants and screening dominant genes which can help Megalobrama amblycephala to resist high-carbohydrate and high-fat diets.

营养性脂肪肝病是水产养殖业中的常见疾病。PPARs在糖脂代谢中具有重要作用，人类和啮齿动物的研究发现PPARs基因多态性与营养性脂肪肝病关系密切。因此，本项目以团头鲂为研究对象，据已获得的cDNA序列，利用PCR-SSCP、测序等技术确定它的多态性位点；建立团头鲂营养性脂肪肝病的易感和不易感群体，然后根据团头鲂个体的基因型和对应的诊断结果及相关指标，利用BP人工神经网络，并结合Logistic回归分析和多因子降维法研究PPARs多态性与营养性脂肪肝病易感性的关系，以及基因-基因、基因-环境间的联合作用对营养性脂肪肝发病风险的影响；然后，通过实时定量PCR和蛋白免疫印迹杂交技术检测脂肪肝形成的关键酶和PPARs靶基因的表达情况，并检测血液和肝组织中相关生化指标，探讨PPARs不同基因型实现其不同调节功能的方式，为阐明营养性脂肪肝病的分子病因，筛选耐高糖高脂饲料的团头鲂优势基因型提供理论依据。

高糖高脂饲料的大量应用使营养性脂肪肝成为养殖业中的常见病。PPARs在糖脂代谢中具有重要作用，其基因多态性与营养性脂肪肝病关系密切。本项目以团头鲂为研究对象，从营养和遗传相结合的角度出发，通过PCR-SSCP、测序、转录组分析获得了PPARs基因的19个多态性位点，并利用BP人工神经网络、Logistic回归及多因子降维法分析了团头鲂易感和不易感群体中不同基因型对营养性脂肪肝病易感性的影响，通过回归方程确认其中5个多态性位点与营养性脂肪肝病密切相关，尤其是PPARβa的3个连续多态性位点（GG和CT型）和PPARr 的1个多态性位点（TT型），它们均可分别作为独立因素影响营养性脂肪肝病的易感性；同时，通过多组学分析和实时定量PCR技术探讨了营养性脂肪肝病的形成机制，发现高糖高脂日粮会通过影响NAFLD、碳代谢及氨基酸转化等代谢路径中的基因表达和代谢产物含量来导致团头鲂肝脏代谢的紊乱，进而诱导营养性脂肪肝的形成，相关研究结果已在《Metabolomics》和《BMC Genomic》等杂志发表，这为预防生产中营养性脂肪肝病的发生、筛选耐高糖高脂饲料的团头鲂优势基因型提供理论依据。