PTH receptor Signaling and Diabetes-induced Bone Disease

骨细胞通过调节成骨细胞的产生和活性，在骨稳态中发挥关键作用 破骨细胞；但对骨细胞在骨病理生理学中的作用知之甚少。由PI在工作期间完成 该奖项之前的资助期表明，甲状旁腺激素1受体(PTH1R)的激活 骨细胞内信号转导抑制骨细胞源性抑制因子Sost/skerostin的表达 并增加破骨细胞分化的主要诱导物RANKL的表达。 此外，RANKL启动子中PTH调节结构域的缺失取消了PTH1R驱动的吸收 骨细胞中的激活。此外，PTH1R成骨细胞缺失的小鼠表现出吸收减少和 甲状旁腺素的合成代谢反应有缺陷。导致这一应用的研究也证实了糖尿病小鼠 链脲佐菌素诱导的糖尿病(S-DM)表现为骨量低，力学和材料性能较差； 吸收增加，形成减少，骨髓脂肪细胞(BMAT)增加； 骨细胞凋亡率增加，Sost/skerostin高表达。此外，还对S-DM小鼠进行了治疗。 甲状旁腺素相关蛋白(PTHrP)衍生的多肽(1-37)通过PTH1R发挥作用，纠正了这些变化， 激活存活信号，阻止骨细胞凋亡。这项研究的长期目标是 确定靶向骨细胞及其产品治疗骨病的潜力。具体目标 这项建议的目的是揭示PTH1R信号保护骨骼退化的潜在机制 在DM中。我们假设甲状旁腺素或阿巴拉肽(FDA批准)激活的骨细胞中的PTH1R信号 骨合成代谢剂)通过调节骨细胞衍生因子来中和DM在骨骼中的损害作用， 从而保持骨量和强度，保持骨细胞活性，并减少BMAT。这一假设 将使用两种已建立的1型和2型糖尿病小鼠模型进行测试，即S-糖尿病和高脂饮食(HFD-DM)， 分别与低胰岛素血症和高胰岛素血症有关，并使用药理学和遗传工具激活 或抑制PTH1R信号转导，干扰骨细胞基因产物。我们将致力于实现以下目标： 目标1将检查PTH1R信号与甲状旁腺素或阿巴拉肽的药理激活是否恢复 S-DM或HFD-DM小鼠(近交系C57BL/6和近交系瑞士Webster品系)的骨量和强度；以及 揭示潜在的细胞和分子机制。Aim 2将检测骨细胞对PTH1R的贡献 信号对糖尿病骨病变的保护作用，通过观察甲状旁腺素和阿巴拉肽在S糖尿病中的作用， HFD-DM和对照组小鼠骨细胞中PTH1R缺失(DMP1-8KB-CRE)。《目标3》将审查 骨细胞源性WnT/β连接素拮抗剂在S糖尿病和高脂血症所致骨骼退化中的作用 DM，通过研究骨细胞(Sostf/f；Dkk1f/f；DMP1-8KB-Cre)中是否缺乏SOST、Dkk1或两者的小鼠 表达LRP5高骨量突变pG171V的小鼠(抵抗SOST和Dkk1介导的抑制 WNT-β连环蛋白信号转导系统)可防止S糖尿病和高脂糖尿病对骨骼的影响。

Osteocytes play a critical role in bone homeostasis by regulating the production and activity of osteoblasts and osteoclasts; but less is known about osteocyte function in bone pathophysiology. Work by the PI during the previous funding period of this Merit award showed that activation of parathyroid hormone 1 receptor (PTH1R) signaling in osteocytes inhibits the expression of Sost/sclerostin, the osteocyte-derived inhibitor of bone formation, and increases in osteocytes the expression of RANKL, the master inducer of osteoclast differentiation. Moreover, deletion of PTH-regulated domains in the RANKL promoter abolished resorption driven by PTH1R activation in osteocytes. Further, mice with osteocytic deletion of PTH1R exhibit decreased resorption and defective anabolic response to PTH. Work leading to this application also established that mice with diabetes mellitus induced by streptozotocin (S-DM) exhibit low bone mass and inferior mechanical and material properties; increased resorption, decreased formation, and increased bone marrow adipocytes (BMAT); as well as increased osteocyte apoptosis and high expression of Sost/sclerostin. Further, treatment of S-DM mice with a PTH related protein (PTHrP)-derived peptide (1-37), which acts through the PTH1R, corrected these changes, and activated survival signaling preventing osteocyte apoptosis. The long term goal of this research is to determine the potential of targeting osteocytes and their products for treating bone maladies. The specific goal of this proposal is to unveil the mechanisms underlying protection of skeletal deterioration by PTH1R signaling in DM. We hypothesize that PTH1R signaling in osteocytes activated by PTH or abaloparatide (FDA-approved bone anabolic agents) counteracts the damaging actions of DM in bone by regulating osteocyte-derived factors, thus maintaining bone mass and strength, preserving osteocyte viability, and reducing BMAT. This hypothesis will be tested using two murine models of established type 1 and type 2 DM, S-DM and high fat diet (HFD-DM), associated with low versus high insulinemia, respectively, and using pharmacologic and genetic tools to activate or inhibit PTH1R signaling, and to interfere with osteocytic gene products. We will pursue the following aims: Aim 1 will examine whether pharmacologic activation of PTH1R signaling with PTH or abaloparatide restores bone mass and strength in S-DM or HFD-DM mice (in inbred C57BL/6 and outbred Swiss Webster strains); and reveal underlying cellular and molecular mechanisms. Aim 2 will examine osteocyte contribution to PTH1R signaling protective action on DM bone disease, by investigating the effect of PTH or abaloparatide in S-DM, HFD-DM and control mice with deletion of the PTH1R in osteocytes (DMP1-8kb-Cre). And Aim 3 will examine the role of osteocyte-derived Wnt/βcatenin antagonists on the skeletal deterioration induced by S-DM or HFD- DM, by investigating whether mice lacking Sost, Dkk1, or both in osteocytes (Sostf/f; Dkk1f/f; DMP1-8kb-Cre) or mice expressing the LRP5 high bone mass mutation pG171V (resistant to Sost- and Dkk1-mediated inhibition of Wnt-βcatenin signaling) are protected from S-DM and HFD-DM induced effects on bone.