The role of Piezo1 in bone homeostasis and mechanotransduction

项目摘要/摘要 机械刺激促进骨骼生长，对成年后骨骼的动态平衡至关重要。损失 机械信号会降低骨量，增加骨折风险。骨细胞，这是埋在骨膜中的细胞 骨基质来源于成骨细胞，能够感知机械负荷的变化并协调骨骼 改建。多条证据表明钙通道参与机械感觉。 由骨细胞加载。例如，钙内流是骨细胞对机械反应最早的反应之一。 体外和体内刺激物。与钙信号在机械反应中的功能作用一致 力，骨细胞对机械刺激的反应可以通过阻断钙通道来抑制 化学阻滞剂。此外，钙显著抑制负荷诱导的大鼠尺骨的骨形成。 通道阻滞剂。然而，机械力激活的钙通道的同一性和它们的 在骨骼中作为机械传感器的功能作用尚不清楚。我们发现Piezo1钙通道是 在骨细胞中高表达，并通过机械刺激增加其表达和活性。 骨细胞。此外，在成骨细胞和骨细胞中Piezo1的缺失会同时减少骨量和骨量 小鼠的力量，与骨骼对机械刺激的反应性丧失一致。此外， 在成骨细胞和骨细胞中缺乏Piezo1的小鼠，骨骼对合成代谢负荷的反应迟钝。WNT1，a Wnt信号的配体，已知由机械信号上调并刺激骨形成，是 在Piezo1条件性基因敲除小鼠中表达下调。重要的是，Piezo1被其化学激动剂激活， Yoda1，模拟液体流动对骨细胞的影响，并增加小鼠的骨量。在此基础上 证据，我们假设骨细胞通过Piezo1感知机械信号的变化，从而 促进骨形成的部分原因是通过激活信号通路增加WNT1的表达。为了测试 在这个假设下，我们将确定骨细胞表达Piezo1是否是机械感觉所必需的 在小鼠的骨骼里。我们将产生Piezo1从骨细胞中删除的小鼠，但不会从成骨细胞中删除， 并将它们的骨骼表型与成骨细胞和骨细胞中缺乏Piezo1的小鼠进行比较。 我们还将在成年小鼠出生后删除Piezo1，并研究它们对胫骨机械载荷的反应 压缩(目标1)。此外，为了了解Piezo1是如何促进骨形成的，我们将确定 用小鼠遗传学方法研究Wnt1在Piezo1介导的体内骨形成中的作用(目标2)。在目标3中，我们 将确定Piezo1是否负责使用尾部悬吊卸载时的骨骼反应 模特。最后，我们将确定Piezo1的药理激活是否能防止相关的骨丢失 使老年小鼠的骨量减少或增加。这项工作的圆满完成将建立一个新的 了解骨骼对合成代谢机械负荷反应的模型，并可能提出新的策略 开发与废用或老化相关的骨丢失的合成代谢疗法。

PROJECT SUMMARY/ABSTRACT Mechanical stimuli promote bone growth and are critical for skeletal homeostasis during adulthood. Loss of mechanical signals decreases bone mass and increases fracture risk. Osteocytes, which are cells buried in the bone matrix and derived from osteoblasts, are able to sense changes in mechanical load and orchestrate bone remodeling. Several lines of evidence suggest that calcium channels are involved in the sensing of mechanical load by osteocytes. For example, calcium influx is one of the earliest responses of osteocytes to mechanical stimuli in vitro and in vivo. Consistent with a functional role for calcium signaling in the response to mechanical forces, the response of osteocytes to mechanical stimuli can be inhibited by blocking calcium channels using chemical blockers. Moreover, load-induced bone formation in the rat ulna is significantly blunted by calcium channel inhibitors. However, the identity of the calcium channels activated by mechanical forces and their functional role as mechanosensors in bone remain unclear. We have found that Piezo1 calcium channel is highly expressed in osteocytes, and that its expression and activity are increased by mechanical stimulation in osteocytes. In addition, deletion of Piezo1 in osteoblasts and osteocytes decreases both bone mass and bone strength in mice, consistent with loss of skeletal responsiveness to mechanical stimulation. Moreover, the skeletal response to anabolic loading is blunted in mice lacking Piezo1 in osteoblasts and osteocytes. Wnt1, a ligand for Wnt signaling that is known to be upregulated by mechanical signals and stimulate bone formation, is downregulated in Piezo1 conditional knockout mice. Importantly, activation of Piezo1 by its chemical agonist, Yoda1, mimics the effects of fluid flow on osteocytes and increases bone mass in mice. Based on this evidence, we hypothesize that osteocytes sense changes in mechanical signals through Piezo1 and thereby promote bone formation in part by activating signaling pathways that increase the expression of Wnt1. To test this hypothesis, we will determine whether Piezo1 expression by osteocytes is required for mechanical sensing in the murine skeleton. We will generate mice in which Piezo1 is deleted from osteocytes, but not osteoblasts, and compare their skeletal phenotype to that observed in mice lacking Piezo1 in osteoblasts and osteocytes. We also will delete Piezo1 postnatally in adult mice and investigate their response to mechanical loads by tibia compression (Aim 1). In addition, to understand how Piezo1 promotes bone formation, we will determine the role of Wnt1 in Piezo1-mediated bone formation in vivo using a mouse genetic approach (Aim 2). In Aim 3, we will determine whether Piezo1 is responsible for the skeletal response to unloading using a tail-suspension model. Lastly, we will determine whether pharmacological activation of Piezo1 prevents bone loss associated with unloading or increases bone mass in old mice. Successful completion of this work should establish a new model for understanding the skeletal response to anabolic mechanical loading and may suggest new strategies to develop anabolic therapies for bone loss related to disuse or aging.