SIRT1 Overexpression in Cellular Mitochondrial Metabolism and Function

描述(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有关，如糖尿病。一个主要的公共卫生问题是肥胖相关疾病的发病率增加，例如糖尿病及其并发症，以及神经退行性疾病的发病率增加，例如老龄化人口中的帕金森氏症和阿尔茨海默氏症。迫切需要采取治疗和预防战略，以减少糖尿病并发症和治疗神经退行性疾病。一些证据表明，这些疾病的一个共同联系是线粒体氧化磷酸化和对氧化损伤的反应减弱。作为线粒体功能的关键调节因子，核呼吸辅活化子有助于调节线粒体的氧化磷酸化，防止细胞和神经元损伤。SIRT1是NAD依赖的脱乙酰酶中sirtuin家族的成员，被认为负责热量限制提供的健康益处。此外，在红葡萄的皮肤中发现的白藜芦醇可以增加SIRT1的活性，延长小鼠的寿命，并可能防止神经退化。SIRT1介导的保护性反应的一个关键成分是转录因子PGC-11的去乙酰化和激活，从而增加线粒体再生和改善细胞氧化能量代谢。SIRT1的作用及其在细胞水平的作用机制/S尚不确定；然而，具有广泛研究焦点的研究人员有兴趣了解SIRT1在不同组织中的生物学作用。为了研究这一点，需要有条件地表达SIRT1的转基因小鼠的产生。为了响应NCRR的PA(PA-07-336)建立适用于两个或多个分类NIH研究所/中心的人类疾病动物模型，我们将开发一种在四环素反应元件(tre-SIRT1/mito-EYFP)控制下表达小鼠SIRT1和线粒体靶向增强型黄色荧光蛋白的转基因小鼠。MITO-EYFP与SIRT1的共表达将被用于鉴定、分离和研究SIRT1表达对线粒体功能的影响。然后，通过将SIRT1与CaMKII-1 TTA小鼠杂交，SIRT1将被靶向中枢和外周神经元。本研究开发的转基因小鼠将帮助来自NINDS、NIDDK、NIA等机构的研究人员测试疾病机制并开发SIRT1介导的治疗方法。具体地说，在这项提案中，我们将研究SIRT1保护中枢和外周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所产生的动物模型来研究非神经细胞中的SIRT1生物学。我们的目标有两个：(1)建立在四环素反应元件(tre；tre-sirt1/mito-EYFP)调控下表达小鼠SIRT1的转基因小鼠。(2)双基因SIRT1神经元特异性小鼠模型的表型。外行描述：肥胖、糖尿病和神经退行性疾病影响着大量的人。SIRT1蛋白被认为是人体抵御疾病的主要调节器，并被红葡萄中发现的白藜芦醇激活。我们的目标是揭示S1RT1保护神经元免受糖尿病引起的神经并发症的机制。 公共卫生相关性：本提案是对NCRR PA-07-336“用于研究的动物模型和相关生物材料的发展”的PA的回应。这个PA的研究目标是“开发、表征或改进人类疾病的动物模型，所考虑的模型必须适用于两个或更多分类NIH研究所/中心的研究兴趣”。一个主要的公共卫生问题是肥胖相关疾病的发病率增加，例如糖尿病及其并发症，以及神经退行性疾病的发病率增加，例如老龄化人口中的帕金森氏症和阿尔茨海默氏症。我们将开发一种在四环素反应元件(tre-SIRT1/mito-EYFP)控制下表达小鼠SIRT1和线粒体靶向增强黄色荧光蛋白的转基因小鼠。MITO-EYFP与SIRT1的共表达将被用于鉴定、分离和研究SIRT1表达对线粒体功能的影响。然后，通过将SIRT1与CaMKII-1 TTA小鼠杂交，SIRT1将被靶向中枢和外周神经元。本研究开发的转基因小鼠将帮助来自NINDS、NIDDK、NIA等机构的研究人员测试疾病机制并开发SIRT1介导的治疗方法。我们的总体假设是，中枢(CNS)和外周神经系统(PNS)中SIRT1的激活将减少神经元和其他组织中的氧化应激并改善MT功能的调节，这可能在神经退行性疾病、糖尿病及其并发症以及在延缓或减少神经系统衰老的影响方面发挥重要作用。Sirtuins的激活为几种与氧化损伤和线粒体功能缺陷有关的人类疾病提供了一种新的治疗方法。

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.