DNA Repair Deficiency Associated with Obesity and the Metabolic Syndrome

描述（由申请人提供）：暴露于氧化应激条件和过量活性氧（ROS）的产生通常被假设为几种人类疾病病因学中的常见致病因素，包括但不限于脂肪肝疾病、血脂异常、胰岛素抵抗型2型糖尿病、心血管疾病/高血压和肥胖症（统称为代谢综合征）。尽管已经确定脂质、蛋白质和核酸是内源性和外源性产生的ROS的关键细胞靶标，但直到最近，ROS诱导的DNA碱基损伤的修复缺陷还没有被认为是上述疾病的关键，而被认为是致癌的核心。然而，已经建立了两种敲除小鼠模型（neil 1和ogg 1），其中氧化损伤DNA的碱基切除修复的启动是有缺陷的，并且在这两种模型中，小鼠都出现了与代谢综合征一致的症状子集。neil 1基因敲除小鼠的疾病表现可能包括肥胖、脂肪肝、血脂异常和高胰岛素血症，雄性敲除小鼠比雌性小鼠受到的影响更严重。此外，从这些小鼠中分离的核DNA的分析揭示了高水平的ROS损伤的碱基的积累，并且线粒体DNA（mtDNA）显示相对于对照同窝出生的小鼠增加的稳态碱基损伤和大的缺失。由于已知过度的氧化应激可在修复能力强的生物体中诱导代谢综合征的症状，因此假设NEIL 1或OGG 1的缺失降低了氧化应激诱导的疾病表现的阈值。在缺乏修复的情况下，受损mtDNA基因组的逐渐积累导致能量产生不足，以及游离脂肪酸和脂质代谢的改变。为了检验这一假设，将评价neil 1-/-、ogg 1-/-和neil 1-/-ogg 1-/-小鼠及其野生型同窝仔在促氧化剂激发与对照条件下的变化的多个生理参数。这些数据将与病理变化率以及通过GC/MS和定量PCR测量的线粒体和核DNA损伤累积相关。这些分析将通过研究NEIL 1在调节生存，诱变和线粒体功能中的作用来补充，以应对氧化或一氧化氮应激条件。此外，由于假设mtDNA的修复在维持代谢稳态中是关键的，因此提出了实验设计来建立各种NEIL 1同种型的细胞内分布，并确定在neil 1-/-细胞中表达这些酶的核靶向形式或髓靶向形式的生物学后果。公共卫生相关性：目前估计，人类肥胖症的日益流行影响超过6000万成年美国人，其继发后果包括但不限于脂肪肝疾病、心血管疾病和胰岛素抵抗/2型糖尿病，统称为代谢综合征。对DNA修复缺陷小鼠模型的研究表明，这些疾病的潜在潜在分子机制缺乏氧化DNA碱基损伤修复，并表现出代谢综合征的许多定义特征：严重肥胖，脂肪肝疾病，血脂异常和胰岛素抵抗。这些研究将检查DNA修复减少在这些疾病的发生和发展中的作用。

DESCRIPTION (provided by applicant): Exposure to oxidative stress conditions and the generation of excessive reactive oxygen species (ROS) are generally hypothesized to be a common causative factor in the etiology of several human diseases including, but not limited to, fatty liver disease, dyslipidemia, insulin-resistant type 2 diabetes, cardiovascular disease/hypertension, and obesity (collectively known as Metabolic Syndrome). Although it is well established that lipids, proteins and nucleic acids are critical cellular targets for endogenously and exogenously produced ROS, until recently, deficiencies in repair of ROS-induced DNA base damage had not been considered to be key in the aforementioned diseases, while being considered central to carcinogenesis. However, two knockout mouse models (neil1 and ogg1) have been created in which the initiation of base excision repair of oxidatively-damaged DNA is defective, and in both models, mice develop subsets of symptoms consistent with Metabolic Syndrome. Disease manifestations in the neil1 knockout mice may include obesity, fatty liver disease, dyslipidemia and hyperinsulinemia, with male knockouts much more severely affected than females. In addition, analyses of nuclear DNAs isolated from these mice reveal the accumulation of high levels of ROS-damaged bases and mitochondrial DNAs (mtDNA) show both increased steady-state base damage and large deletions relative to control littermates. Since it known that excessive oxidative stress can induce symptoms of Metabolic Syndrome in repair-proficient organisms, it is hypothesized that the loss of NEIL1 or OGG1 lowers the threshold at which oxidatively stress-induced disease is manifested. In the absence of repair, the progressive accumulation of compromised mtDNA genomes leads to deficiencies in energy production, as well as alterations in free fatty acid and lipid metabolism. In order to test this hypothesis, multiple physiological parameters will be evaluated for changes in neil1-/-, ogg1-/- and neil1-/-ogg1-/- mice and their wild-type littermates during pro-oxidant challenges versus control conditions. These data will be correlated with rates of pathological changes and mitochondrial and nuclear DNA damage accumulation as measured by GC/MS and quantitative PCR. These analyses will be complemented by examining the role of NEIL1 in the modulation of survival, mutagenesis, and mitochondrial function in response to oxidative or nitric oxide stress conditions. Further, since repair of mtDNA is hypothesized to be critical in maintaining metabolic homeostasis, experimental designs are proposed to establish intracellular distribution of various NEIL1 isoforms and determine the biological consequences of expressing nuclear or mitochondrially-targeted forms of these enzymes in neil1-/- cells. PUBLIC HEALTH RELEVANCE: The growing epidemic of human obesity is currently estimated to affect over 60 million adult Americans, with secondary consequences including, but not limited to, fatty liver disease, cardiovascular disease, and insulin resistance/type 2 diabetes, collectively known as the Metabolic Syndrome. A potential underlying molecular mechanism for these diseases is suggested by the study of DNA repair-deficient mouse models that lack oxidative DNA base damage repair and exhibit many of the defining hallmarks of the Metabolic Syndrome: severe obesity, fatty liver disease, dyslipidemia, and insulin resistance. These investigations will examine the role of reduced DNA repair in the onset and progression of these diseases.