Dissecting the Molecular Mechanisms that Govern Notch Mediated Skeletal Stem Cell Maintenance Throughout Adulthood/Aging.

剖析控制 Notch 介导的整个成年/衰老过程中骨骼干细胞维持的分子机制。

• 批准号: 10387942
• 负责人: Lindsey Hope Remark
• 金额: $ 5.18万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-28 至 2026-02-27
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10387942
• 关键词: Adult; Affect; Age; Age-Related Bone Loss; Aging; American; Animals; Biological Assay; Biology; Bone Regeneration; Bromodeoxyuridine; Cell Count; Cell Cycle Progression; Cell Differentiation process; Cell Maintenance; Cell Proliferation; Cell physiology; Cell surface; Cells; Chronic; Data; Disease; Economic Burden; Elements; Exhibits; Femur; Flow Cytometry; Fracture; Frequencies; Genes; Homeostasis; Human; Impairment; In Vitro; Individual; Injury; Knock-out; Knockout Mice; Label; Light; Link; Literature; Longevity; Maintenance; Mediating; Mediator of activation protein; Modeling; Molecular; Morbidity - disease rate; Mus; Orthopedics; Osteoblasts; Osteoclasts; Osteogenesis; Osteoporosis; Patients; Persons; Phenotype; Physiologic pulse; Play; Publishing; Quality of life; Regenerative Medicine; Role; Signal Transduction; Skeletal bone; Skeleton; Small Interfering RNA; Stains; Testing; Therapeutic; Time; Training; Undifferentiated; Unemployment; Work; age related; aging population; base; bone; bone fracture repair; bone fragility; bone healing; bone loss; bone mass; cell behavior; combat; conditional knockout; disability; experimental study; fracture risk; global health; health economics; high risk; human old age (65+); in vivo; insight; middle age; mouse model; nicastrin protein; notch protein; novel; osteoblast differentiation; osteogenic; overexpression; prevent; self-renewal; skeletal; skeletal stem cell; stem; stem cell biology; stem cell function; stem cell homeostasis; stem cells; stemness; substantia spongiosa; therapeutic target; tibia; transcriptome sequencing

PROJECT SUMMARY Orthopedic injuries represent one of the most significant health and economic burdens in our nation. Bone fragility with age due to diseases such as osteoporosis increases the risk for fracture, while at the same time leading to delayed bone healing resulting in a higher risk for non-union. With a growing aging population, the demand for therapeutics to combat age-related bone loss is one of the pillars of the American Orthopedic Association’s “Own-the-Bone” initiative. Current treatments mainly focus on preventing further bone loss rather than increasing bone mass in a substantial manner. Skeletal stem and progenitor cells (SSPCs) are essential for proper bone mass and bone healing. SSPCs are known to decline in frequency and function with age. Therefore, SSPCs represent the most direct target to increase bone mass. Previous studies and our own data suggest that Notch signaling plays a pivotal role in SSPC homeostasis and self-renewal. We recently identified a conditional Notch knockout mouse model which exhibits a striking phenotype of increasing bone formation in all skeletal elements in aging mice. This mouse model, the Ncstnf/f;LepR-cre mouse, is deficient in Nicastrin (Ncstn), an essential component of the Notch signal transduction machinery of all 4 Notch receptors in LepR+ SSPCs. This model has exciting therapeutic potential to combat age-related bone loss. Based on this cKO model and our preliminary data I hypothesize that Notch signaling is essential in maintenance of the skeletogenic stem cell pool during adulthood and aging. In order to investigate the molecular mechanism by which Notch signal inhibition leads to increased trabecular bone mass with age, we performed bulk RNA sequencing on LepR+ SSPCs from middle-aged WT and cKO mice. This unbiased sequencing approach identified Ebf3 as a significantly downregulated gene in cKO cells and therefore potential downstream mediator of Notch signaling in promoting SSPC differentiation into osteoblasts. Previous work has linked Ebf3 to the inhibition of osteogenesis, however, the upstream regulators of Ebf3 are not yet known. Also, while others have looked into the role of Notch in SSPCs, the downstream mechanisms by which Notch is controlling SSPC function are still largely unknown. Therefore, this proposal aims to i) define the precise role of Notch signaling in SSPCs during aging/adulthood on proliferation, self-renewal and differentiation and ii) uncover the molecular mechanism by which Notch signaling is having the effects on proliferation, self-renewal and differentiation. These aims will provide novel specific mechanistic insights into the mechanism by which a Notch-Ebf3 signaling axis maintains skeletal stem cells in their primitive state throughout the lifespan of an animal.

项目总结 骨科伤害是我国最严重的健康和经济负担之一。骨 随着年龄的增长，骨质疏松症等疾病导致的脆弱性增加了骨折的风险，同时 导致骨愈合延迟，导致骨不连的风险更高。随着人口老龄化的不断加剧， 对对抗年龄相关性骨丢失的治疗药物的需求是美国骨科的支柱之一 该协会的“骨子里”倡议。目前的治疗方法主要集中在防止进一步的骨丢失上，而不是 而不是以实质性的方式增加骨量。骨骼干细胞和祖细胞(SSPC)是必不可少的 用于适当的骨量和骨愈合。众所周知，SSPC的频率和功能随着年龄的增长而下降。 因此，SSPC是增加骨量的最直接靶点。以前的研究和我们自己的数据 提示Notch信号在SSPC的动态平衡和自我更新中起着关键作用。我们最近确认了 一种条件性Notch基因敲除小鼠模型，该模型显示出显著的骨形成增加表型 衰老小鼠体内的所有骨骼元素。这种小鼠模型Ncstnf/f；Lepr-cre小鼠缺乏尼古丁。 (Ncstn)，Lepr+中所有4种Notch受体的Notch信号转导机制的重要组成部分 SSPC。这种模式在对抗年龄相关性骨丢失方面具有令人兴奋的治疗潜力。基于这一CKO模型 我们的初步数据假设，Notch信号在维持骨骼发育的干细胞中是必不可少的 成年期和衰老期间的细胞池。为了研究Notch信号传递的分子机制 抑制导致骨小梁随着年龄的增长而增加，我们对Lepr+进行了批量RNA测序 WT和CKO中年小鼠的SSPC。这种无偏见的测序方法将EBF3识别为 CKO细胞中显著下调的基因，因此可能是Notch信号的下游介体 促进SSPC向成骨细胞分化。以前的工作已经将EBF3与抑制 然而，EBF3的上游调控因子尚不清楚。另外，虽然其他人也在调查 Notch在SSPC中的作用，Notch控制SSPC功能的下游机制仍然是 很大程度上是未知的。因此，本提案旨在：i)定义在SSPC过程中Notch信令的确切作用 衰老/成年期对增殖、自我更新和分化的影响以及II)通过以下方式揭示分子机制 哪些Notch信号对细胞的增殖、自我更新和分化有影响。这些目标将 提供了对Notch-EBF3信号轴维持机制的新的具体机制的见解 骨骼干细胞在动物的整个生命周期中处于原始状态。