DNA Methylation Profiles in a Polygenic Mouse Model of Diet-Induced Obesity

描述（由申请人提供）：肥胖是冠心病（CHD）和非胰岛素依赖型糖尿病（NIDDM）的主要危险因素。虽然双胞胎研究和收养研究的结果为遗传对体重的影响提供了强有力的支持，但导致肥胖的突变在人群中很少见，导致普通肥胖的主要基因尚未被确定。这导致了这样的建议，即个体之间肥胖的变化可能是由多个遗传基因座决定的，其中任何单个基因的变化仅产生中等影响。根据这一公式，肥胖已被报道与差异基因表达，这些改变的复合效应可能至少部分负责NIDDM和CHD的易感性。这项提案的长期目标是评估在比较肥胖和瘦个体时，是否可以检测到可能导致基因激活或沉默的DNA甲基化模式的修饰。实现这一长期目标的第一步是在实验环境中使用动物模型进行概念验证研究。向C57 BL/6小鼠提供高脂肪饮食导致体重增加、高胰岛素血症和胰岛素抵抗，并且与喂食正常小鼠食物的同窝小鼠相比，主动脉脂肪条纹病变的出现加速。这些小鼠的使用将允许营养暴露变化，同时保持遗传背景不变。基于先前的报道，即DNA甲基化的变化可能先于载脂蛋白E基因突变导致血浆胆固醇升高的小鼠动脉粥样硬化病变的发展，该建议的目的是检验以下假设：（a）野生型成人肥胖和动脉粥样硬化的发病机制，喂食致动脉粥样硬化饮食的A型小鼠可能与治疗动物和对照动物之间DNA甲基化的功能差异相关，以及（B）DNA甲基化的获得性变异可能与正常衰老有关，并可能通过追求以下特定目标而促进复杂疾病的发展：1）评估C57 BL/6小鼠中饮食诱导的肥胖和动脉粥样硬化是否与肥胖相关。6小鼠动脉粥样硬化病变和其他靶器官的DNA甲基化模式的改变，通过全基因组微阵列分析。2)鉴定和克隆甲基化序列，并验证通过微阵列分析检测到的具有改变的甲基化模式的基因在体外和体内受到甲基化的调节。3)评估C57 BL/6小鼠衰老过程中多个组织中的整体和基因特异性DNA甲基化。这些实验可能揭示DNA甲基化变化在响应营养和其他环境影响的同时疾病过程中的作用和序列，并且还可能产生一组新的候选基因，这些基因随后可以在人类队列的关联研究中进行测试。高脂肪饮食是一种环境暴露，可能会改变DNA甲基化和基因表达。这将首先在小鼠中进行研究，以了解这种类型的变化是否有助于解释非胰岛素依赖型糖尿病（NIDDM）和冠心病（CHD）的风险增加。

DESCRIPTION (provided by applicant): Obesity is a major risk factor for the development of coronary heart disease (CHD) and non-insulin dependent diabetes mellitus (NIDDM). Although the results of twin studies and adoption studies provide strong support for a genetic influence on body weight, mutations causing obesity are rare in the population and major genes contributing to common obesity have not been identified. This has led to the suggestion that the variation in adiposity between individuals may be determined by multiple genetic loci with variations in any single gene producing only a moderate effect. In accordance with this formulation, obesity has been reported to be associated with differential gene expression, and the composite effect of these alterations may be at least partly responsible for the susceptibility to NIDDM and CHD. It is the long-term goal of this proposal to evaluate whether modifications in DNA methylation patterns that may result in either activation or silencing of genes can be detected when comparing obese and lean individuals. The first step in approaching this long-term goal is a proof-of-concept study using an animal model in an experimental setting. Provision of a high-fat diet to C57BL/6 mice results in increased body weight, hyperinsulinemia and insulin-resistance, and the accelerated appearance of aortic fatty streak lesions when compared to littermates fed normal mouse chow. The use of these mice will allow nutrient exposure to be varied while keeping genetic background constant. Based on previous reports that changes in DNA methylation may precede the development of atherosclerotic lesions in mice with a mutation in the apolipoprotein E gene causing elevated plasma cholesterol, the goal of this proposal is to test the hypotheses that (a) the pathogenesis of adult-onset obesity and atherosclerosis in wild- type mice fed an atherogenic diet may be correlated with functional differences in DNA methylation between treated and control animals and (b) that acquired variability in DNA methylation may be involved in normal aging and could contribute to the development of complex disease by pursuing the following specific aims: 1) to evaluate whether diet-induced obesity and atherosclerosis in C57BL/6 mice entails alterations in DNA methylation patterns in atherosclerotic lesions and other target organs by genome-wide microarray analysis. 2) to identify and clone methylated sequences, and validate that genes with altered methylation patterns detected by microarray analysis are regulated by methylation in vitro and in vivo. 3) to assess global and gene-specific DNA methylation in multiple tissues during aging in C57BL/6 mice. These experiments may reveal the role and sequence of DNA methylation changes in simultaneous disease processes in response to nutrition and other environmental influences, and may also yield a set of novel candidate genes that can later be tested in association studies in human cohorts. Consuming a high-fat diet is an environmental exposure that may alter DNA methylation and change gene expression. This will be studied first in mice to see whether this type of change could help to explain the increased risk for non-insulin dependent diabetes mellitus (NIDDM) and coronary heart disease (CHD) with obesity.