SIRT1 Overexpression in Cellular Mitochondrial Metabolism and Function

描述（由申请人提供）：氧化应激和线粒体功能障碍与广泛的神经退行性疾病和代谢紊乱（如糖尿病）相关。一个主要的公共卫生问题是与肥胖有关的疾病，如糖尿病及其并发症的发病率增加，以及老年人口中帕金森病和阿尔茨海默病等神经退行性疾病的发病率增加。迫切需要治疗和预防策略，以减少糖尿病的并发症和治疗神经退行性疾病。一些证据表明，这些疾病的共同联系是线粒体氧化磷酸化减少和对氧化损伤的反应。核呼吸共激活因子是线粒体功能的关键调节因子，有助于调节线粒体氧化磷酸化，防止细胞和神经元损伤。SIRT1是NAD+依赖性去乙酰化酶sirtuin家族的一员，它被认为是热量限制提供的健康益处的原因。此外，在红葡萄皮中发现的白藜芦醇可以增加SIRT1的活性，延长小鼠的寿命，并可能预防神经变性。SIRT1介导的保护性反应的一个关键组成部分是转录因子PGC-11的去乙酰化和激活，导致线粒体再生增加和细胞氧化能代谢改善。SIRT1的作用及其在细胞水平上的作用机制尚不确定；然而，具有广泛研究焦点的研究人员对了解SIRT1在不同组织中的生物学作用感兴趣。为了研究这一点，需要产生有条件表达SIRT1的转基因小鼠。为了响应NCRR （PA-07-336）的PA，开发适用于两个或多个NIH分类研究所/中心的人类疾病动物模型，我们将开发一种转基因小鼠，在四环素应答元件（trer -SIRT1/ mitto - eyfp）的控制下表达小鼠SIRT1和线粒体靶向增强黄色荧光蛋白。mito-eYFP与SIRT1的共表达将用于鉴定、分离和研究SIRT1表达对线粒体功能的影响。然后，通过与CamKII-1 tTA小鼠杂交，将SIRT1表达靶向中枢和外周神经元。本研究开发的转基因小鼠将帮助NINDS、NIDDK、NIA等机构的研究人员检测疾病机制，开发SIRT1介导的治疗方法。具体来说，我们将研究SIRT1保护中枢和周围神经元免受糖尿病诱导的神经元损伤的机制。该提案还描述了其他研究人员如何开发动物模型来研究非神经元细胞中的SIRT1生物学。我们有两个目的：(1)建立在四环素应答元件（TRE;TRE-SIRT1/ mitto - eyfp）控制下表达小鼠SIRT1的转基因小鼠。(2)对双基因SIRT1神经元特异性小鼠模型进行表型分析。概要描述：肥胖、糖尿病和神经退行性疾病影响着大量的人。SIRT1蛋白被认为是人体防御疾病的主要调节器，被红葡萄中的白藜芦醇激活。我们的目标是揭示S1RT1保护神经元免受糖尿病诱导的神经系统并发症的机制。

DESCRIPTION (provided by applicant): Oxidative stress and mitochondrial dysfunction have been associated with a wide range of neurodegenerative diseases and metabolic disorders such as diabetes. A major public health problem is the increase in the incidence of obesity-related diseases, such as diabetes and its complications and the increased incidence of neurodegenerative diseases, for example Parkinson's and Alzheimer's diseases in the aging population. Therapeutic and preventive strategies to reduce the complications of diabetes and to treat neurodegenerative diseases are urgently needed. Several lines of evidence indicate that a common link in these diseases is diminished mitochondrial oxidative phosphorylation and response to oxidative injury. Key regulators of mitochondrial function, the nuclear respiratory coactivators help to regulate mitochondrial oxidative phosphorylation and prevent cellular and neuronal injury. SIRT1 is a member of the sirtuin family of NAD+dependent deacetylases, which is proposed to be responsible for health benefits provided by caloric restriction. Furthermore, resveratrol found in the skin of red grapes increases the activity SIRT1, prolongs life-span in mice, and may prevent neurodegeneration. A key component of the protective response mediated by SIRT1 is deacetylation and activation of the transcription factor PGC-11 leading to increased mitochondrial regeneration and improved cellular oxidative energy metabolism. The role of SIRT1 and its mechanism/s of action at cellular level are uncertain; however investigators with a wide spectrum of research foci have an interest in understanding the biological actions of SIRT1 in different tissues. To study this, generation of transgenic mice that conditionally expresses SIRT1 is needed. In response to an PA from NCRR (PA-07-336) to develop animal models of human disease that are applicable to the research interests of two or more categorical NIH Institutes/Centers, we will develop a transgenic mouse that expresses mouse SIRT1 and mitochondrial targeted enhanced yellow fluorescent protein under the control of tetracycline responsive element (TRE-SIRT1/mito-eYFP). Co-expression of mito-eYFP with SIRT1 will be used to identify, isolate and study the influence of SIRT1 expression on mitochondrial function. Then, SIRT1 expression will be targeted to central and peripheral neurons by crossing it with CamKII-1 tTA mice. The transgenic mice developed in this study will aid investigators from NINDS, NIDDK, NIA and other institutes to test disease mechanisms and develop SIRT1 mediated therapies. Specifically in this proposal, we will investigate the mechanism by which SIRT1 protects central and peripheral neurons against diabetes-induced neuronal injury. The proposal also describes how the animal models produced can be developed by other investigators to study SIRT1 biology in non-neuronal cells. We have two aims: (1) To develop a transgenic mouse that expresses mouse SIRT1 under the control of tetracycline responsive element (TRE;TRE-SIRT1/mito-eYFP). (2) To phenotype the bigenic SIRT1 neuron specific mouse model. Lay description: Obesity, diabetes, and neurodegenerative diseases affect large numbers of people. The SIRT1 protein is considered to be a master regulator of the body's defense against disease and is activated by resveratrol found in red grapes. We aim to uncover the mechanism by which S1RT1 protects neurons from diabetes induced neurological complications. PUBLIC HEALTH RELEVANCE: This proposal is submitted in response to a PA from NCRR PA-07-336 "Development of Animal models and Related Biological Materials for Research". The research objective of this PA is to "develop, characterize or improve animal models for human disease and that models to be considered must be applicable to the research interests of two or more categorical NIH Institutes/Centers". A major public health problem is the increase in the incidence of obesity-related diseases, such as diabetes and its complications and the increased incidence of neurodegenerative diseases, for example Parkinson's and Alzheimer's diseases in the aging population. We will develop a transgenic mouse that expresses mouse SIRT1 and mitochondrial targeted enhanced yellow fluorescent protein under the control of tetracycline responsive element (TRE-SIRT1/mito-eYFP). Co-expression of mito-eYFP with SIRT1 will be used to identify, isolate and study the influence of SIRT1 expression on mitochondrial function. Then, SIRT1 expression will be targeted to central and peripheral neurons by crossing it with CamKII-1 tTA mice. The transgenic mice developed in this study will aid investigators from NINDS, NIDDK, NIA and other institutes to test disease mechanisms and develop SIRT1 mediated therapies. Our overall hypothesis is that activation of SIRT1 in the central (CNS) and peripheral nervous system (PNS) would reduce oxidative stress and improve regulation of Mt function in neurons and other tissues that may be important in neurodegenerative diseases, diabetes and its complications, and in delaying or reducing the effect of aging in the nervous system. Activation of the sirtuins offers the potential for a novel treatment of several human diseases that are related to oxidative injury and defects of mitochondrial function.