New tools for investigating mitochondrial dynamics in stem cells during aging

 描述(申请人提供)：成体干细胞支持组织内稳态，并在个体一生中充当修复受损组织的细胞库。然而，皮肤、肝脏、血液和肌肉等组织的维持和再生随着年龄的增长而急剧减少。干细胞不能充分替代老化和受损的组织可能是与年龄相关的组织动态平衡下降和疾病发生率增加的重要因素。因此，在老龄化的背景下，识别和表征导致干细胞功能丧失的机制是至关重要的，因为延缓或对抗老年人干细胞功能丧失的策略很可能在未来几年成为再生医学的重要组成部分。生物体衰老在许多层面上与新陈代谢的改变有关--从外在的、系统的因素(例如。激素、胰岛素/胰岛素样生长因子)导致细胞器效率降低， 比如线粒体。我们实验室公布的数据表明，增强黑腹果蝇肠道干细胞(ISCs)的线粒体生物发生足以延长寿命，并延缓肠道和雄性生殖系中与年龄相关的组织稳态的下降。然而，这些研究提出了许多基本问题。例如，如果干细胞中增强的线粒体生物生成足以延缓组织衰老，干细胞使用什么策略来维持健康的线粒体池？在哺乳动物中解决新陈代谢、干细胞行为、组织动态平衡和衰老之间的联系是困难和复杂的，而在果蝇中则更直接，因为它们的寿命相对较短，在哺乳动物系统中调节衰老和新陈代谢的保守信号通路，以及几个离散的成年干细胞群体。我们的初步数据表明，成年果蝇睾丸中生殖系干细胞(GSCs)的维持依赖于这些细胞内足够的线粒体分裂和融合。此外，我们有证据表明，GSCs将线粒体运输到邻近的、有丝分裂后的体细胞壁龛细胞(中枢细胞)。转移的频率似乎随着年龄的增长而增加；因此，我们假设转移是GSCs处理受损线粒体的一种机制。在这里，我建议将尖端的电子显微镜(EM)成像技术与传统的遗传方法相结合，以提供对完整组织中干细胞中线粒体和线粒体蛋白质的组织、分离和结构的前所未有的见解。除了解决有关线粒体转移的问题外，这些工具还将有助于更详细地分析许多线粒体蛋白质的位置和功能。我们的发现将对以细胞为基础的疗法在年龄和代谢性疾病治疗中的使用产生重大影响，特别是在老年人中。

 DESCRIPTION (provided by applicant): Adult stem cells support tissue homeostasis and serve as a cellular reservoir for repair of damaged tissue throughout the life of an individual. However, maintenance and regeneration of tissues such as skin, liver, blood, and muscle decrease dramatically with age. The inability of stem cells to adequately replace aging and damaged tissues is likely a significant contributing factor to age-related decreases in tissue homeostasis and increased incidence of disease. Therefore, it is essential to identify and characterize mechanisms leading to loss of stem cell function, in the context of aging, as strategies to delay or counter loss of stem cell function in older individuals will likely become a important component of regenerative medicine in years to come. Organismal aging is linked to altered metabolism at many levels- from changes in the quantity of extrinsic, systemic factors (ex. hormones, insulin/Insulin-like growth factors) to decreased efficiency of cellular organelles, such as mitochondria. Published data from our lab suggested that enhanced mitochondrial biogenesis in Drosophila melanogaster intestinal stem cells (ISCs) is sufficient to increase lifespan and delay the age-related decline in tissue homeostasis, in both the intestine and male germ line. However, many fundamental questions were raised by these studies. For example, if enhanced mitochondrial biogenesis in stem cells is sufficient to delay tissue aging, what strategies are used by stem cells to maintain a healthy pool of mitochondria? Addressing the links between metabolism, stem cell behavior, tissue homeostasis, and aging is difficult and complex in mammals yet more straightforward in Drosophila, given a relatively short lifespan, conserved signaling pathways that regulate aging and metabolism in mammalian systems, and several discrete populations of adult stem cells. Our preliminary data suggest that maintenance of germline stem cells (GSCs) in the testis of adult Drosophila melanogaster is dependent upon adequate mitochondrial fission and fusion within these cells. Furthermore, we have evidence that GSCs traffic mitochondria to adjacent, post- mitotic, somatic niche cells (hub cells). The frequency of transfer appears to increase with age; therefore, we hypothesize that transfer occurs as one mechanism for GSCs to dispose of damaged mitochondria. Here, I propose to combine cutting-edge electron microscopy (EM) imaging techniques with traditional genetic approaches in Drosophila melanogaster to provide unprecedented insight into the organization, segregation, and structure of mitochondria and mitochondrial proteins in stem cells in intact tissues. In addition to addressing questions regarding mitochondrial transfer, these tools will facilitate analyzing the location and function of many mitochondrial proteins in more detail. Our findings will have major implications for the use of cell-based therapies in the treatment of age-onset and metabolic diseases, particularly in older individuals.