非酒精性脂肪肝脾阳虚证与易感基因、危险因素的相关性研究

Non-alcoholic fatty liver disease (NAFLD) is the most predominant hepatic disease, takes about 15% of our population. It has continuous disease progression and effects on the metabolism, is becoming one of the major health problems in China. Previous studies have confirmed NAFLD Spleen deficiency subgroup with specific genotype. This proposal proposes a scientific hypothesis that "Gene-Environment-Symptoms interaction plays an important role in NAFLD development ". We use prospective cohort study with random sampling to recruited normal subjects, NAFLD subjects (Spleen deficiency syndrome as case and syndrome of phlegm and blood stasis as control population). Demographic data, lifestyle, TCM symptoms, blood and urine samples were collected. 30 SNP from previous studies are selected and genotyped in our subjects’ DNA samples. Multi-dimensional analysis including genetic association, gene by gene and gene-environment-TCM symptom will be applied in our proposal. This study aim to understand NAFLD susceptibility genes, gene-environment interaction in the pathogenicity, and develop multi-dimensional model of TCM factors-gene-environment-typical syndromes. Further cellular function experiment are applied to test the model results. We first applied gene-environment-TCM symptoms analysis into NAFLD disease and spleen deficiency subtype. Combining genetics with tradition Chinese symptoms/characters will explore a new area of understanding chronic disease.

非酒精性脂肪肝(NAFLD)约占我国人口的15%，会导致持续的肝脏病变及影响全身代谢，是关切国民健康的重要问题。课题组既往研究证实NAFLD脾阳虚证具有特异基因分型。本课题提出“基因-环境-证候交互作用在NAFLD及脾阳虚证候的发生发展起到重要作用”的科学假说，采用随机抽样、前瞻性队列，构建NAFLD脾阳虚证研究队列(正常人群为疾病对照，NAFLD痰瘀互结证为证候对照人群)。采集人口学数据、证候表观数据、生活习惯调查及DNA样本；选取相关通路研究的30个SNP，对样品进行基因分型；通过对基因关联、基因-基因、基因-环境的多维分析研究NAFLD易感基因及基因与环境在致病中的机理，并在多维分析中引入脾阳虚证候因素，建立基因-环境-典型证候的关联研究模型；最终以功能试验验证模型分析结果。本课题首次将证候因素和环境因素引入NAFLD疾病的基因组学研究，探索应用中西医结合在慢性病研究中的新思路。

项目课题成功纳入NAFLD病例组341例（包括脾阳虚证组163例和非脾阳虚证组178例）、健康对照组341例和验证集332例进行资料收集与标本采集。选取源于GWAS或候选基因研究报道的来自5个基因的10个SNP多态性位点，采用Sequenom平台的MassARRAY技术在测试样品中进行基因分型研究。经过研究获得以下研究结论：1. GCKR的rs1260326、rs780094、rs780093及MBOAT7的rs641738与汉族人群NAFLD发生相关联。2. 基于GCKR的rs780094、MBOAT7的rs641738的相加效应和代谢危险因素结合的风险预测模型，对于NAFLD发生风险的预测能力较高。3. GCKR的rs1260326、rs780094、rs780093是NAFLD脾阳虚证的基因组生物标志物，与脾阳虚证的程度变化相关。项目超额完成预期目标，已发表论文6篇（其中SCI收录4篇），累计影响因子10.39分；获高等学校科学研究优秀成果奖（科学技术）一等奖一项；获得软件著作权一项；参加学术会议3次，并作了口头交流；培养研究生3名。本课题组深入研究了与NAFLD致病相关通路的基因，在队列研究中寻找到更贴近中国人群的非酒精脂肪肝易感基因位点。分别进行了NAFLD病证关键差异基因分析、基因共表达网络分析结合致病通路分析，关键基因组生物标志物筛选；联合临床病证信息，建立机器学习模型对NAFLD病证风险进行建模，改进和优化了NAFLD风险预测系统；为疾病的演化和转归提供预警管理，可进一步指导中医药在非酒精性脂肪肝及脾阳虚证的精准治疗。