The Role of miR-219a-5p in Bone Metabolism

项目摘要/摘要 骨质疏松症是一种常见的临床症状，其特征是骨量减少，增加了脆性的风险。 老年人口中的骨折。由于骨质疏松患者的骨形成明显受损，因此更多的 完全了解调节骨骼新陈代谢的基本分子机制可能会 导致了新疗法的发展。因此，新的分子途径的鉴定 影响骨形成对抗击骨质疏松症的临床治疗的发展至关重要。我们的 以前的工作已经明确地证明，抗骨转录因子，维甲酸受体- 相关孤儿受体-β(Ror-β)在成骨细胞分化中起抑制作用。不断增加的表达 生理性衰老过程中的RORβ水平有助于骨丢失，因为缺乏RORβ的小鼠表现出更高的骨量 通过激活骨骼合成代谢的Wnt途径。这导致了一种概念，即抑制RoRβ可能 代表了一种在整个衰老过程中增加骨量的新范例。我们假设 MicroRNAs(MiR)可以控制RoRβ的水平，并发现一种特定的miR，miR-219a-5p(miR-219a)，顶部 预测的RoRβ-调节miR，表现出与RoRβ相反的表达模式，提示有潜在的 RoRβ/MiR-219a在骨骼中的调节轴。我们证明了miR-219a直接调节RORβ水平 成骨细胞，miR-219a的输送模拟促进成骨细胞的分化，miR-219a 拮抗剂抑制分化。我们进一步表明miR-219a增加了骨合成代谢的活性。 WNT途径。因此，我们认为miR-219a是一种骨合成代谢蛋白，可用于保存骨量。 随着年龄的增长，可能通过增加Wnt途径的活性。使用一种新的鼠标模型，在那里我们可以激活 MiR-219a以组织特异性的方式表达，在目标1中，我们将确定miR-219a在RORβ中的作用。 在成人骨量或骨折后表达细胞，假设RoRβ下调 通过miR-219a可以增加骨量，加速骨折愈合。在目标2中，我们将探索更多 MiR-219a通过激活不同骨细胞系中的miR-219a在骨稳态中的一般作用，与 假设RoRβ和/或这些谱系中的其他miR-219a靶点下调将增加骨骼 随年龄增长的质量。最后，在目标3中，我们将进行体外实验，以探索其分子机制。 MiR-219a如何调节Wnt通路并确定miR-219a直接靶向Rorβ是否影响 WNT活性和骨细胞功能。这些关键研究的完成不仅将提供更完整的 了解miR-219a在骨骼中的功能，但如果呈阳性，也将为继续研究提供强有力的理由 MIR-219a模拟作为一种新的治疗策略，在各种临床条件下刺激骨形成。

PROJECT SUMMARY/ABSTRACT Osteoporosis is a common clinical condition characterized by low bone mass that increases the risk of fragility fractures in the elderly population. Since bone formation is clearly impaired in osteoporotic patients, a more complete understanding of the fundamental molecular mechanisms that regulate bone metabolism is likely to lead to the development of novel therapies. Therefore, identification of novel molecular pathways which influence bone formation is crucial to the development of clinical treatments to combat osteoporosis. Our previous work has definitively established that the anti-osteogenic transcription factor, retinoic acid receptor- related orphan receptor-beta (Rorβ) plays an inhibitory role in osteoblast differentiation. Increasing expression levels of Rorβ during physiological aging contributes to bone loss, as mice lacking Rorβ exhibit higher bone mass through activation of the bone anabolic Wnt pathway. This leads to the notion that inhibition of Rorβ may represent a novel paradigm to increase bone mass throughout the aging process. We hypothesized that microRNAs (miRs) may control Rorβ levels, and found that a specific miR, miR-219a-5p (miR-219a), the top predicted Rorβ-regulatory miR, exhibits an inverse expression pattern to Rorβ, suggesting a potential Rorβ/miR-219a regulatory axis in bone. We demonstrate that miR-219a directly regulates Rorβ levels in osteoblasts, that delivery of miR-219a mimics enhances osteoblast differentiation, and that miR-219a antagonists suppress differentiation. We further show that miR-219a increases the activity of the bone anabolic Wnt pathway. Therefore, we propose that miR-219a is bone anabolic and may be used to preserve bone mass with age, possibly through increasing Wnt pathway activity. Using a novel mouse model where we can activate miR-219a expression in a tissue-specific manner, in Aim 1 we will determine the effects of miR-219a in Rorβ- expressing cells on adult bone mass or following a fracture, with the hypothesis that downregulation of Rorβ through miR-219a will increase bone mass and accelerate fracture healing. In Aim 2, we will explore the more general effects of miR-219a in bone homeostasis by activating miR-219a in various bone cell lineages, with the hypothesis that downregulation of Rorβ, and/or other miR-219a targets in these lineages, will increase bone mass with age. Finally, in Aim 3 we will perform in vitro experiments to explore the molecular mechanism of how miR-219a regulates the Wnt pathway and determine whether direct targeting of Rorβ by miR-219a affects Wnt activity and bone cell function. Completion of these pivotal studies will not only provide a more complete understanding of miR-219a function in bone, but if positive will also provide a strong justification for pursuing miR-219a mimics as a novel therapeutic strategy to stimulate bone formation in various clinical conditions.