Genetic Factors Governing Inter-individual Variation to Oxidative Stress Response

描述(申请人提供)：我建议测量氧化应激反应在个人和人群中的变异，并确定与这种变异相关的遗传位点。氧化应激与年龄相关疾病的病因学密切相关，并被认为是长寿的决定因素。介导氧化应激反应的途径已经得到了广泛的研究，但我们对这些途径的功能个体差异仍然知之甚少。了解控制氧化应激反应个体差异的因素可能会提高我们对年龄相关疾病和生理性衰退的理解。在这里，我提出了一系列实验和统计分析，以研究氧化应激反应变异的遗传基础。具体地说，在目标1中，我将从70名高加索人和70名非裔美国人中提纯CD4+T细胞。我将使这些细胞暴露在氧化应激下72小时，方法是将它们培养在较高的(20%)氧气水平，同时保持相同个体的对照细胞处于生理(5%)氧气水平。在此期间的特定时间点，我将测量氧化的谷胱甘肽与还原的谷胱甘肽的比率(氧化应激的指标)；氧化的鸟嘌呤与未修饰的鸟嘌呤的比率 鸟嘌呤(氧化应激诱导的DNA损伤的指标)；以及全基因组mRNA的丰度，在处理细胞和对照细胞中。在目标2中，我将收集目标1中使用的所有样本的基因类型，并绘制数量性状基因座(QTL)，包括解释氧化应激反应变化的表达QTL。然后，将在AIM 3中使用报告基因分析来验证大约30个假定的eQTL。

DESCRIPTION (provided by applicant): I propose to measure variation in oxidative stress response across individuals and populations, and to identify the genetic loci that are associated with such variation. Oxidative stress has been strongly implicated in the etiology of age-related disease and as a determinant of longevity. Pathways that mediate oxidative stress response have been extensively studied, but we still know relatively little about individual variation in th function of these pathways. Understanding the factors that govern variation in oxidative stress response across individuals may improve our understanding of age-related disease and physiological decline. Here, I propose a series of experiments and statistical analyses to study the genetic basis of variation in oxidative stress response. Specifically, in Aim 1, I will purify CD4+ T cells from 70 Caucasians and 70 African Americans. I will expose these cells to oxidative stress for 72 hours by culturing them in elevated (20%) oxygen levels, while maintaining control cells from the same individuals in physiological (5%) oxygen levels. At specified time points during this period, I will measure the ratio of oxidized glutathione to reduced glutathione (an indicator of oxidative stress); the ratio of oxidized guanine to unmodified guanine (an indicator of oxidative stress-induced DNA damage); and genome-wide mRNA abundance, in both treated and control cells. In Aim 2, I will collect genotypes from all samples used in aim 1 and map quantitative trait loci (QTLs), including expression QTLs, that explain variation in oxidative stress response. Reporter gene assays will then be used in Aim 3 to validate approximately 30 of the putative eQTLs.