The AMPK/ULK1/p27Kip1 axis regulates autophagy and cell survival in aged satellite cells

A.项目摘要/摘要： 骨质疏松症是与年龄相关的骨骼肌质量和力量的丧失；它会导致一系列的共病。 包括身体功能的丧失。石棺减少症患者的一种这样的不安是能力减弱 在受伤后再生肌肉。肌肉干细胞，被称为卫星细胞，需要激活， 增殖和分化，以再生肌肉和恢复身体功能。老化的卫星细胞速度较慢 在受伤时激活；易发生细胞凋亡；修复受损肌肉的效率较低。AMPK- P27Kip通路似乎是成功从静止状态进入细胞周期的关键。我们的 初步数据表明，随着年龄的增长，AMPK-p27Kip通路出现了扰动。这段获奖期 将研究AMPK-p27Kip通路在人类和人类卫星细胞衰老表型中的作用 老鼠模型。在Aim1中，我们将检验AMPK及其下游靶点p27Kip激活的假设 调节衰老卫星细胞的自噬/凋亡决策。我们将使用分子检测来拯救 衰老细胞中这一途径的功能丧失，并恢复增殖能力。在目标2中，我们将测试 假设运动是AMPK和自噬的生理诱导者，刺激AMPK-p27Kip 途径，从而增强老化的小鼠和人类卫星细胞的增殖和代谢功能。在……里面 目的3，我们将验证一种假设，即在老化的卫星细胞中拯救AMPK-p27Kip通路的活性将 通过幼龄和老年小鼠损伤肌肉移植实验，提高体内再生能力。 总之，这个方案中的实验将检验AMPK-p27Kip通路受损的假设 在老化的卫星细胞中；假设这种损伤导致了延迟的增殖率， 易发生细胞凋亡，损伤后肌肉再生能力降低。怀特博士的关键方面 职业提升将是：学习如何协调临床运动试验；培训卫星方法 细胞分离和新陈代谢分析，特别是在衰老有机体的背景下。培训计划将 需要专门的内部和外部科学报告；干细胞方面的相关课程工作/研讨会 生物学和老龄化；以及密集的职业指导，以确保在走向独立的过程中取得进展。这项研究 本提案中详细介绍的职业发展计划将由一支优秀的导师团队进行。 威廉·E·克劳斯博士是杜克医学院的教授，是临床锻炼方面的知名专家 研究和肌肉/卫星细胞生物学；他将担任主要导师。怀特博士也将有医生。 布鲁斯·斯皮格曼、艾米·韦杰斯和安娜·玛丽亚·库尔沃作为共同导师；他们将分别提高 细胞新陈代谢训练；衰老干细胞生物学；自噬。杜克大学医学院 环境是本提案中概述的研究和培训活动的理想环境。该奖项将提供 怀特博士接受了最佳的培训，以确保他作为独立调查员的职业生涯有了一个出色的开端。

a. Project summary/abstract: Sarcopenia is the age-related loss in skeletal muscle mass and strength; it leads to a host of co-morbidities including loss of physical function. One such perturbation in persons with sarcopenia is the diminished ability to regenerate muscle after injury. Muscle stem cells, referred to as satellite cells, are required to activate, proliferate and differentiate to regenerate muscle and restore physical function. Aged satellite cells are slower to activate upon injury; susceptible to apoptosis; and less efficient in repairing injured muscle. The AMPK- p27Kip pathway appears critical for successful transition from quiescence to entry into the cell cycle. Our preliminary data identify perturbations in the AMPK-p27Kip pathway with advanced age. This award period will investigate the role of the AMPK-p27Kip pathway in the phenotype of satellite cell aging in both human and mouse models. In Aim1, we will test the hypothesis that activation of AMPK and its downstream target p27Kip regulates the autophagy/apoptosis decision in aged satellite cells. We will use molecular assays to rescue the functional loss of this pathway in aged cells and return proliferative capacity. In Aim 2, we will test the hypothesis that exercise, a physiological inducer of AMPK and autophagy, stimulates the AMPK-p27Kip pathway, thereby enhancing proliferation and metabolic function in aging murine and human satellite cells. In Aim 3, we will test the hypothesis that rescuing activity of the AMPK-p27Kip pathway in aged satellite cells will improve in vivo regenerative capacity by transplantation experiments in injured muscle of young and old mice. Together, the experiments in this proposal will test the hypothesis that the AMPK- p27Kip pathway is impaired in aging satellite cells; and the hypothesis that this impairment contributes to the delayed proliferation rate, susceptibility to apoptosis, and reduced ability to regenerate muscle after injury. Key aspects of Dr. White’s career enhancement will be: to learn how to coordinate clinical exercise trials; to train in methods of satellite cell isolation and metabolic analysis, especially in the context of the aging organism. The training program will entail dedicated internal and external scientific presentations; pertinent course work/workshops in stem cell biology and aging; and intensive career mentorship to ensure progress toward independence. The research and career development plan detailed in this proposal will be conducted with a team of outstanding mentors. Dr. William E. Kraus, a professor at the Duke Medical School is an established expert in clinical exercise studies and muscle/satellite cell biology; he will serve as the primary mentor. Dr. White will also have Drs. Bruce Spiegelman, Amy Wagers and Ana Maria Cuervo as co-mentors; they will respectively enhance training in cell metabolism; aging stem cell biology; and autophagy. The Duke School of Medicine environment is ideal for the research and training activities outlined in this proposal. This award will provide Dr. White with optimal training to ensure an outstanding start to his career as an independent investigator.