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.

项目摘要 骨科伤害代表着我们国家最重要的健康和经济伯恩斯之一。骨 由于骨质疏松症（例如骨质疏松症）而导致的年龄随着年龄的增长而脆弱，同时又增加了骨折的风险 导致骨骼愈合延迟，导致非工会风险更高。随着人口衰老的增长， 对与年龄相关的骨质损失打击治疗的需求是美国骨科的支柱之一 协会的“骨头”倡议。目前的治疗主要集中于防止进一步的骨质流失 比以实质性的方式增加骨质量。骨骼干和祖细胞（SSPC）是必不可少的 适当的骨骼质量和骨骼愈合。已知SSPC的频率和功能随着年龄的增长而下降。 因此，SSPC是增加骨骼质量的最直接目标。以前的研究和我们自己的数据 表明Notch信号传导在SSPC稳态和自我更新中起关键作用。我们最近确定了 有条件的缺口敲除小鼠模型，该模型表现出增加骨形成的打击表型 老化小鼠的所有骨骼元素。该鼠标模型，NCSTNF/F; LEPR-CRE鼠标，缺乏尼卡斯特林 （NCSTN），LEPR+中所有4个Notch受体的Notch信号传输机械的重要组成部分 SSPC。该模型具有令人兴奋的治疗潜力，以打击与年龄相关的骨质流失。基于此CKO模型 我们的初步数据我假设Notch信号对于维持骨骼发明茎是必不可少的 成年和衰老期间的细胞池。为了研究Notch信号的分子机制 抑制会导致小梁骨量随着年龄的增长而增加，我们在LEPR+上进行了大量RNA测序 来自中年WT和CKO小鼠的SSPC。这种公正的测序方法将EBF3确定为 CKO细胞中显着下调基因，因此潜在的下游介体的Notch信号传导 在将SSPC分化为成骨细胞时。以前的工作已将EBF3与抑制联系起来 然而，成骨，EBF3的上游调节剂尚不清楚。另外，其他人也调查了 Notch在SSPC中的作用，Notch控制SSPC函数的下游机制仍然是 在很大程度上未知。因此，该提案的目的是i）定义Notch信号在SSPC中的确切作用 在增殖，自我更新和分化以及ii）中衰老/成年，ii）通过 Notch信号传导对增殖，自我更新和分化产生影响。这些目标会 提供新的特定机械见解，以了解Notch-EBF3信号轴维持的机制 在动物的整个生命周期中，骨骼干细胞处于原始状态。