cAMP信号在热量限制延缓机体衰老中的作用研究

Calorie restriction (CR) is the most robust intervention demonstrated to extend life span and delay the physiological deterioration associated with aging. Because CR involves a number of overlapping and interconnected signaling pathways, it is difficult to identify with certainty the mechanisms underlying the beneficial effects of CR. The positive health effects of CR and sirtuin activity in animal models have provoked intense interest in the development of small-molecule to prevent or delay agingrelated diseases..cAMP signaling is highly complex, and its outcomes vary depending on the effector activated by cAMP and on other factors including the cell type, the cellular compartment of cAMP action, and the duration and intensity of cAMP signaling. cAMP is a key mediator of metabolic regulation, and the identification of PDEs as resveratrol targets might explain how resveratrol mimics some aspects of CR.cAMP-dependent protein kinase A (PKA) pathway plays a key role in the regulation of metabolism and multiple cellular functions. However, the comprehensive effects of cAMP sigaling on aging mammalian systems and the underlying molecular mechanisms are still unclear. .Here the present study aimed to investigate physiological changes and variational expression of age-related genes with possible alterative cAMP signaling during the aging process induced by metabolic disorder. By daily subcutaneous injection of D-galactose, we prepared presenile model mice that were kept in oxidative stress due to glycometabolism dysfunction. We treated the model mice with administration of exogenous cAMP and its antagonists, with the control mice of high-fat diet model and dietary restriction model. .Our investigation indicates that activation of PKA could improve behavioral performance, including Morris Water Maze test and Locomotor Activity in presenile mice. It shows much thiner subcutaneous fat-pads,without loss of bodyweight in presenile mice treated with cAMP (lifespan under observation), accompanying with significant increase in blood SOD activity, compared to the untreated group. In vitro study shows that polyphenols extracted from green tea, which has inhibitory effects on ROS generation, may alter the expression level of AMPK and be a regulator of PKA. These findings indicated activation of PKA could decrease subcutaneous fat aggradation without alteration of bodyweight and improve the clearance of ROS, suggesting the natural products altering PKA activity may have pharmacological implications on obesity and age-related diseases.

本课题旨在研究热量限制对小鼠寿命延长作用的机理，探讨热量限制对衰老小鼠代谢功能的改善作用与cAMP信号水平的内在联系。通过对热量限制小鼠、自然衰老小鼠和高脂饮食小鼠的cAMP信号及其上下游关键蛋白水平的测定，确定热量限制延寿作用与cAMP信号水平之间的关系。通过考察外源性引发cAMP水平变化对衰老相关信号转导通路和衰老相关疾病产生的影响，阐明cAMP对代谢紊乱所导致的衰老表征及相关退行性病变的干预作用和可能的分子机制。本课题主要从动物整体和重要器官水平考察cAMP信号水平增高对热量限制延寿作用的贡献，同时为了进一步揭示cAMP的作用机制，我们对小鼠的多种组织器官进行关键蛋白表达水平检测，并利用离体细胞考察cAMP信号通路及其上下游关键靶点蛋白对衰老过程的干预作用。课题方案基于我们已有的研究成果并结合相关文献制定，具有坚实的理论依据和前期工作基础。

热量限制是除基因干预以外最有效的延寿措施，但是热量限制发挥作用的前提是坚持减少30%~40%的热量摄入，这是大多数人无法办到的，因此研究者们一直试图寻找模拟热量限制的药物。环磷酸腺苷（cAMP）作为第二信使在许多生物过程中发挥重要作用，已有报道称白藜芦醇通过cAMP发挥抗衰老作用。我们的研究表明，外源性cAMP能通过激活Sirtuin和AMPK模拟热量限制改善老年小鼠衰老相关表型，改善高脂饮食小鼠的代谢紊乱。细胞和体外实验证明，cAMP能与Sirtuin家族蛋白（SIRT1和SIRT3）发生直接相互作用并提高其活性。外源性cAMP最终会被细胞内的PDE降解成为AMP后升高AMP/ATP比值促进AMPK（Thr172）磷酸化。这些发现表明cAMP通过调节代谢与衰老的关键蛋白改善代谢功能并延缓衰老进程，因此cAMP是一个很好的热量限制模拟候选药物。作为细胞内第二信使，cAMP在许多生物过程中都很重要，同时衰老的过程也是复杂的，无法通过单一信号通路来解释，我们认为cAMP抗衰老的作用可能以来信号转导的网络，而不仅仅是瞄准单一的目标或针对单一的衰老相关疾病。我们的工作属于衰老相关的基础研究，旨在探索有效健康的干预手段，缓解了我国因人口老龄化带来的医疗保障体系的巨大压力，为干预代谢紊乱诱导的衰老相关疾病提供了有前景的治疗对策。