Supplement to: EX VIVO REJUVENATION AND EXPANSION OF MUSCLE STEM CELLS FROM AGED MICE

“来自老龄小鼠的肌肉干细胞的离体返老还童和扩增” 项目总结/摘要 肌肉干细胞（MuSC），也被称为卫星细胞，在整个生命过程中对肌肉再生至关重要 [1]的文件。在衰老过程中，骨骼肌质量和损伤后的再生能力逐渐下降，导致 老年人的生活质量下降[2]。对老年骨骼肌功能障碍的解释 组织的研究集中在限制MuSC功能的组织微环境因素中与衰老相关的变化[3]。 在我的博士后研究中，我已经证明，使用组织再生的非侵入性成像分析， 从老年小鼠中前瞻性分离的MuSC的再生能力相对于 年轻的MuSC，揭示了以前未检测到的老年MuSC中的内在干细胞缺陷。此外，我 确定了一种新的体外策略来克服老年MuSC的再生功能障碍;老年MuSC的治疗 MuSC维持在具有p38促分裂原的小分子抑制剂的软仿生水凝胶平台上[4]。 相关的蛋白激酶产生功能性干细胞绝对数量的扩增，并恢复其功能。 与年轻MuSC的再生功能相似。这种方法为恢复和增加 MuSC的数量，可以使局部自体干细胞治疗老年人肌肉萎缩 个体，在临床上没有药物治疗。 我建议将先前的生物工程和干细胞生物学训练与新的肌肉训练相结合 生理学和系统生物学，以进一步研究衰老中MuSC的再生功能障碍及其 体外拯救。在目标1（K99阶段），我将评估体外处理的旧MuSC是否可以拯救缺陷的MuSC。 肌肉再生和增加老年小鼠的肌肉力量，并能够长期恢复活力 响应连续的再生需求而运行。在目标2（连接K99/R 00阶段）中，我将阐明 在老年MuSC中，缺陷的再生功能是否是同质或异质表型， 结合多参数质谱细胞术（CyTOF）[5]和SPADE算法[6]分析， 移植测定以鉴定和比较分离的MuSC亚群的再生功能 年轻和年老的老鼠。在目标3（R 00阶段），我将阐明失调的信号网络机制 作为旧MuSC的干细胞功能障碍的基础，用于使用信号传导网络改善治疗性治疗- 系统生物学方法[7]。 这一向独立过渡的建议描述了研究和职业发展活动， 包括参加会议和课程培训，这将使我成为一个有竞争力的候选人， 独立的教师职位，并帮助我在一个创新的，成功的研究计划的发展， 干细胞衰老的生物学和治疗。这些活动将由Helen Blau博士（主要），Scott Delp（共同导师）和Garry Nolan（共同导师）在斯坦福大学，这是一个世界级的干细胞生物学 研究机构。

"Ex vivo Rejuvenation and Expansion of Muscle Stem Cells from Aged Mice" Project Summary/Abstract Muscle stem cells (MuSCs), also known as satellite cells, are essential to muscle regeneration throughout life [1]. In aging, skeletal muscle mass and regenerative capacity after injury progressively decline, leading to diminished quality of life in aged individuals [2]. Efforts to explain the dysfunction of aged skeletal muscle tissue have focused on aging-related changes in tissue microenvironment factors restricting MuSC function [3]. In my postdoctoral research, I have demonstrated, using non-invasive imaging assays of tissue regeneration, that MuSCs prospectively isolated from old mice have a marked reduction in regenerative capacity relative to young MuSCs, revealing a previously undetected intrinsic stem cell defect in old MuSCs. Further, I have identified a novel ex vivo strategy to overcome the regenerative dysfunction of old MuSCs; treatment of old MuSCs maintained on a soft biomimetic hydrogel platform [4] with a small molecule inhibitor of p38 mitogen- associated protein kinase yields an expansion in absolute numbers of functional stem cells and restores their function in regeneration to that of young MuSCs. This approach offers promise for rejuvenating and increasing the numbers of MuSCs and could enable localized autologous stem cell therapy for muscle wasting in aged individuals, for which there are no pharmacologic treatments in clinical use. I propose to merge prior training in bioengineering and stem cell biology with new training in muscle physiology and systems biology to further investigate the regenerative dysfunction of MuSCs in aging and its rescue ex vivo. In Aim 1 (K99 phase), I will evaluate whether ex vivo-treated old MuSCs can rescue defective muscle regeneration and increase muscle strength in old mice and are capable of long-term rejuvenated function in response to successive regenerative demands. In Aim 2 (bridging K99/R00 phases), I will elucidate whether defective regenerative function is a homogeneous or heterogeneous phenotype in old MuSCs by combining multi-parameter mass cytometry (CyTOF) [5] and SPADE algorithm [6] analysis with sensitive transplantation assays to identify and compare the regenerative functions of MuSC sub-populations isolated from young and old mice. In Aim 3 (R00 phase), I will elucidate dysregulated signaling network mechanisms underlying the stem cell dysfunction of old MuSCs for improved therapeutic treatment using signaling network- level systems biology approaches [7]. This Transition to Independence proposal describes research and career development activities, including conference attendance and course training, that will establish me as a competitive candidate for an independent faculty position and aid my development of an innovative, successful research program in the biology and treatment of stem cell aging. These activities will be mentored by Drs. Helen Blau (primary), Scott Delp (co-mentor), and Garry Nolan (co-mentor) at Stanford University, which is a world-class stem cell biology research institution.