MAMMALIAN SIR2 FUNCTION IN PANCREATIC BETA CELLS

描述（由申请人提供）：我们在该提案中的长期目标是了解由系统性NAD生物合成作为驱动器和Sirt 1作为调节哺乳动物代谢和衰老的介质组成的新型代谢网络的生理意义。我们特别关注Sirt 1在胰腺B细胞中的作用，并通过分析胰腺β细胞特异性Sirt 1过表达（BESTO）转基因小鼠，证明Sirt 1促进葡萄糖刺激的B细胞胰岛素分泌。有趣的是，这种Sirt 1介导的B细胞功能增强在老年BESTO小鼠中减弱，部分原因是与年龄相关的NAD生物合成下降。此外，我们在BESTO和热量限制小鼠的胰岛中进行的广泛表达谱表明，CR增强了B细胞中的Sirt 1活性，这可能是由于在系统水平上增强了NAD的生物合成。这些发现为关于NAD生物合成和Sirt 1在代谢和衰老调节中的联系的令人兴奋的研究开辟了新的途径。我们以前已经证明，烟酰胺磷酸核糖转移酶（Nampt），从烟酰胺（维生素B3的一种形式）的NAD生物合成途径中的限速酶，在调节Sirt 1活性和葡萄糖刺激的2细胞胰岛素分泌中起着重要作用。总之，这些发现使我们假设，在衰老和热量限制（CR）中Nampt介导的NAD生物合成的动态变化导致全身水平Sirt 1活性的显著变化，从而诱导多种组织（如2细胞和肝细胞）中年龄相关和CR响应性代谢变化。为了解决这一假设，我们提出以下具体目标：1）为了进一步确定Nampt介导的NAD生物合成在B细胞和肝细胞中Sirt 1活性调节中的生理学意义，将通过遗传和生物学操作Nampt和Sirt 1活性，在原代胰岛和肝细胞中检查NAD含量、Sirt 1靶基因表达和生理学功能，2）研究B细胞和肝细胞中Nampt介导的NAD生物合成和Sirt 1活性是否随年龄而下降，在衰老过程中检查Nampt蛋白水平、Nampt和Sirt 1活性以及B细胞和肝功能，和3）分析CR是否增强B细胞和肝细胞中Nampt介导的NAD生物合成和Sirt 1活性，CR对Nampt介导的NAD生物合成和Sirt 1活性的代谢作用将在各种遗传和药理学模型中进行分析。这些研究应该提供新的见解的生理意义和治疗应用的这种新的代谢网络组成的系统NAD生物合成和Sirt 1的年龄相关的并发症在人类。 公共卫生相关性：这项研究将提供以下重要成果：首先，这项研究将确定一个以前未被认识到的代谢网络，它在哺乳动物的代谢和衰老调节中起着关键作用。其次，这项研究将提供深入了解组织功能，特别是在胰腺B细胞和肝脏中的年龄相关变化的机制，以及热量限制的抗衰老作用。最后，本研究的预期结果将为开发新的预防/治疗干预措施提供重要的见解，这些干预措施用于重要的年龄相关代谢并发症，如人类葡萄糖耐量受损和2型糖尿病。

DESCRIPTION (provided by applicant): Our long-term objectives in this proposal are to understand the physiological significance of a novel metabolic network comprised of systemic NAD biosynthesis as a driver and Sirt1 as a mediator in the regulation of metabolism and aging in mammals. We have focused particularly on the role of Sirt1 in pancreatic b cells and demonstrated that Sirt1 promotes glucose-stimulated insulin secretion in b cells by analyzing pancreatic beta cell-specific Sirt1-overexpressing (BESTO) transgenic mice. Interestingly, this Sirt1-mediated enhancement of b cell function is blunted in aged BESTO mice, partly due to an age-associated decline in NAD biosynthesis. Furthermore, our extensive expression profiling performed in islets from BESTO and calorically restricted mice suggests that CR enhances Sirt1 activity in b cells, possibly due to augmented NAD biosynthesis at a systemic level. These findings set the stage for a novel avenue of exciting research concerning the connection between NAD biosynthesis and Sirt1 in the regulation of metabolism and aging. We have previously demonstrated that nicotinamide phosphoribosyltransferase (Nampt), the rate-limiting enzyme in the NAD biosynthetic pathway from nicotinamide (a form of vitamin B3), plays an important role in the regulation of Sirt1 activity and glucose-stimulated insulin secretion in 2 cells. Together, these findings led us to the hypothesis that a dynamic alteration in Nampt-mediated NAD biosynthesis in aging and caloric restriction (CR) causes significant changes in Sirt1 activity at a systemic level and thereby induces age-associated and CR-responsive metabolic changes in multiple tissues, such as 2 cells and hepatocytes. To address this hypothesis, we propose the following specific aims: 1) To further establish the physiological significance of Nampt-mediated NAD biosynthesis in the regulation of Sirt1 activity in b cells and hepatocytes, NAD content, Sirt1 target gene expression, and physiological functions will be examined in primary islets and hepatocytes by manipulating Nampt and Sirt1 activities genetically and pharmacologically, 2) to investigate whether Nampt-mediated NAD biosynthesis and Sirt1 activity decline with age in b cells and hepatocytes, Nampt protein levels, Nampt and Sirt1 activities, and b cell and hepatic functions will be examined in the process of aging, and 3) to analyze whether CR augments Nampt-mediated NAD biosynthesis and Sirt1 activity in b cells and hepatocytes, the metabolic effects of CR on Nampt-mediated NAD biosynthesis and Sirt1 activity will be analyzed in various genetic and pharmacological models. These studies should provide new insight into the physiological significance and therapeutic applications of this novel metabolic network comprised of systemic NAD biosynthesis and Sirt1 for age-associated complications in humans. Public Health Relevance: The proposed study should provide the following important outcomes: First, this study will identify a previously unrecognized metabolic network that plays a critical role in the regulation of metabolism and aging in mammals. Second, this study will provide insights into the mechanisms for age-associated changes in tissue function, especially in pancreatic b cells and liver, and anti-aging effects of caloric restriction. Lastly, the anticipated outcome of this study will provide important insight into the development of new preventive/therapeutic interventions for important age-associated metabolic complications, such as impaired glucose tolerance and type 2 diabetes in humans.