肥胖儿童“非酒精性脂肪肝病”发病遗传机制的探索性研究

Non-alcoholic fatty liver disease (NAFLD) in obese children has become a global health problem. Although the specific pathogenesis of NAFLD is still not clear, hereditary susceptibility is believed to be the basis for the development of NAFLD. Current studies on predisposing genes of NAFLD mainly focus on adults. Although studies have found that some common variants are associated with pediatric NAFLD, there are still few reports about low-frequency/rare variants with stronger pathogenic effects. The present study takes predisposing genes screening as the breakthrough of genetic mechanism in pediatric NAFLD. Firstly, a case-control study with relatively small sample will be used to identify related candidate genes for NAFLD by whole exome sequencing (Discovery Research). Secondly, we will take another case-control study with large sample to confirm the candidate genes by genotyping (Validation Study). Finally, on the basis of the above studies and environmental influencing factors of NAFLD in obese children, gene-environment interactions will be detected with the methods of crossover analysis, multifactor dimensionality reduction, etc. The purpose of this research is to provide new clues and targets for further research on genetic mechanism of NAFLD in obese children and offer scientific evidence for the precise intervention of environmental risk factors.

肥胖儿童非酒精性脂肪肝病（Non-alcoholic fatty liver disease，NAFLD）已成为全球儿童健康问题。尽管发病机制仍未明确，但遗传易感性被认为是NAFLD发病的基础。目前国内外NAFLD易感基因的研究主要集中在成人，儿童中虽有一些常见变异研究，但致病效应较强的低频/罕见变异，国内外尚无报道。本研究将易感基因筛查作为遗传机制探索的突破口，首先，拟采用小样本病例对照研究，通过全外显子组测序，大范围筛选NAFLD相关候选基因（发现研究）；其次，采用大样本病例对照研究，通过基因分型技术，验证筛选发现的候选基因（验证研究）。最后，在上述研究基础上，结合肥胖儿童NAFLD环境影响因素，采用叉生分析、多因子降维法等方法，解析基因-环境因素之间的交互作用。本研究旨在为肥胖儿童NAFLD发病遗传机制的深入研究提供新线索及靶点，也为环境危险因素的精准干预提供科学依据。

本研究对62例NAFLD和31例单纯性肥胖儿童进行全外显子测序。首次在中国肥胖儿童人群中进行NAFLD的罕见变异探索研究，共检出641225个罕见变异。通过生物信息学方法、基因负荷检验和ACMG有害性分类等方法筛选致病或可能致病变异，得到7个致病或可能致病变异基因位点。同时，研究首次在中国肥胖儿童人群中进行常见变异与NAFLD的全外显子关联分析，共检出232467个常见变异。进一步通过生物信息学方法、NAFLD关联分析和KEGG通路富集分析得到8条与NAFLD相关的信号通路。本研究通过选取低频罕见变异与常见变异（胰岛素抵抗通路、炎性信号通路和脂质代谢通路等信号通路）共41个相关基因的53个SNPs进行大样本验证。在大样本验证研究中，通过基因分型技术，采用4种传统遗传模型511例NAFLD例和511例单纯性肥胖儿童进行分析，包括共显性遗传模型、显性遗传模型、隐性遗传模型和加性遗传模型。总体而言，本研究通过小样本探索及大样本验证技术发现了来自脂代谢通路的ECI2基因rs2326408位点、ECI2，C6orf201基因rs659305位点和CALML5基因rs10904516位点、炎性通路的TLR5基因rs851139位点、胰岛素抵抗通路的INS基因rs3842748位点、SREBF1基因rs2297508位点多态性与肥胖儿童NAFLD有关。利用多因素降维的方法检测基因-基因的相互作用，发现来自脂代谢通路的ECI2基因rs2326408位点、CALML5基因rs10904516位点和来自炎性通路的TLR5基因rs851139位点多态性与肥胖儿童NAFLD有关且存在交互作用。这些基因位点有望为肥胖儿童NAFLD发病遗传机制的深入研究提供新的线索及靶点。