Age-related changes in the neural stem cells of the olfactory epithelium

描述（由申请人提供）：成体干细胞是组织稳态和再生的基础。由于所有的成人组织都随着年龄的增长而退化，因此了解成人组织干细胞的功能如何随着年龄的增长而下降是一个重要的问题。我的研究的总体目标是整合一系列遗传学，基因组学，生物化学和生物物理学方法，以阐明组织发育，维持和再生以及最终退化过程中干细胞和祖细胞功能的基因调控和控制变化的分子机制;也就是说，控制结构化的、可塑的生物形式如何形成、维持和改造以及最终降解的分子基础是什么。我们不了解神经干细胞年龄相关性衰退的机制。我的职业发展计划这个K 01奖涉及课程，研讨会，并与实验室和导师的直接互动，总而言之，将为我提供高通量测序（HT-Seq）和计算基因组学的专业知识，以及在衰老的分子机制的强大背景。然后，我想联合收割机和计算基因组学方法相结合，以补充我已经采用的遗传、分子和生物化学方法，以研究嗅上皮中干细胞功能与年龄相关的下降的潜在机制。我建议的研究重点是了解发生在嗅觉上皮神经干细胞中的与年龄相关的变化，这些变化是神经发生和组织再生与年龄相关的下降的基础。嗅上皮（OE）是一种感觉神经上皮，支持成年神经发生和组织再生损伤后。在中枢神经系统的神经发生的网站，其更新和再生能力随着年龄的增长而下降。OE的特征在于两个干细胞群体：水平基底细胞（HBC）大多是静止的，并作为储备干细胞在损伤后再生OE，而球状基底细胞（GBC）是干细胞和祖细胞的异质群体，其通过不断更新神经元群体而成为正常组织稳态的基础。OE为研究体内神经发生过程中介导储备和活性干细胞功能年龄相关变化的分子机制提供了一个很好的模型。我们计划测试的假设，即年龄相关的减少，在OE的神经发生和再生能力是由于耗尽的GBC的承诺祖细胞亚群和HBC和GBC干细胞的增殖和分化的能力。使用转基因报告细胞系的特定组合，免疫组化特定的干细胞和祖细胞群体表达的基因，和有丝分裂标记保留实验，我们将表征细胞动力学和功能的不同的干细胞和祖细胞的OE随年龄的变化。然后，我们将进行RNA-Seq，以彻底表征年轻和老年神经干细胞亚群的转录组。总之，这将使我们能够以高度定量的方式辨别哪些基因差异表达，哪些剪接变体用于年轻与老年储备和活性干细胞。这一系列的调查有可能确定干细胞中与年龄相关的变化，挑战将是确定近因，并确定它们是否可以被修改或逆转。从这些努力中获得的知识也可以为细胞替代疗法和年龄相关疾病的建模提供信息。

DESCRIPTION (provided by applicant): Adult stem cells underlie tissue homeostasis and regeneration. Because all adult tissues degrade with age, understanding how adult tissue stem cells decline in function with age is an important problem. The overall goal of my research is to integrate a range of genetic, genomic, biochemical, and biophysical approaches to elucidate the molecular mechanisms that underlie gene regulation and control changes in stem and progenitor cell function during tissue development, maintenance and regeneration, and ultimately, degeneration; i.e. what are the molecular underpinnings that control how structured, plastic biological form is made, maintained and reformed, and finally degraded. We do not understand the mechanisms responsible for the age-related decline in neural stem cells. My career development plan for this K01 award involves coursework, seminars, and direct interaction with labs and mentors that in sum will provide me with expertise in high throughput sequencing (HT-Seq) and computational genomics and a strong background in the molecular mechanisms of aging. I want to then combine the HT-Seq and computational genomic methods to complement the genetic, molecular, and biochemical approaches I have already been employing to investigate the mechanisms underlying age- related decline in stem cell function in the olfactory epithelium. My proposed research focuses on understanding the age-related changes that occur in the olfactory epithelial neural stem cells that underlie the age-related decline in neurogenesis and tissue regeneration. The olfactory epithelium (OE) is a sensory neuroepithelium that supports adult neurogenesis and tissue regeneration following injury. As in the sites of neurogenesis in the central nervous system, its renewal and regenerative capacity decreases with age. The OE is characterized by two stem cell populations: the horizontal basal cells (HBCs) are mostly quiescent and function as reserve stem cells to regenerate the OE following injury, and the globose basal cells (GBCs) are a heterogeneous population of stem and progenitor cells that underlie normal tissue homeostasis by continually renewing the neuronal population. The OE provides an excellent model for investigating the molecular mechanisms that mediate age-related changes in reserve and active stem cell function during neurogenesis in vivo. We plan to test the hypothesis that the age-associated reduction in neurogenesis and regenerative capacity in the OE is due to a depletion of the committed progenitor subpopulation of GBCs and the inability of the HBC and GBC stem cells to proliferate and differentiate. Using specific combinations of transgenic reporter lines, immunohistochemistry to genes expressed by specific stem and progenitor cell populations, and mitotic label retention experiments, we will characterize how the cellular dynamics and function of the different stem and progenitor cells of the OE change with age. We will then perform RNA-Seq to thoroughly characterize the transcriptome of young and aged subpopulations of neural stem cells. Together, this will allow us to discern in a highly quantitative manner which genes are differentially expressed and which splice-variants utilized in young versus aged reserve and active stem cells. This line of inquiry has the potential to identify age-related changes in stem cells, and the challenge will be to define the proximate causes and determine if they can be modified or reversed. The knowledge gained from such endeavors may also inform approaches to cell replacement therapies and modeling of age-related disease.