BLR&D Research Career Scientist Award Application for Teresita Bellido, PhD

骨生物学的最新进展，我的实验室的研究部分得到了VA的支持， 骨细胞（最丰富的骨细胞）在骨中起着关键作用， 通过调节成骨细胞和破骨细胞的产生和活性来维持体内平衡， 分别溶解骨。然而，对骨细胞在骨病理生理学中的功能知之甚少。 我们的工作表明，骨细胞是甲状旁腺激素（PTH）作用的关键靶细胞， 骨细胞中PTH 1受体（PTH 1 R）的信号传导增加骨形成并增强骨重建， PTH骨骼作用的公认特征。骨细胞PTH 1 R信号转导降低了 Sost/sclerostin，一种骨形成的骨细胞源性抑制剂，并增加RANKL的表达， 破骨细胞分化的主要诱导剂。我们还发现，骨细胞中PTH 1 R缺乏的小鼠表现出 吸收减少和对PTH的合成代谢反应缺陷。在最近的研究中，我们发现， 糖尿病（DM）小鼠表现出的骨量和较差的机械和材料性能， 伴有形成减少、吸收增加和骨髓脂肪细胞（BMAT）增加， 沿着骨细胞凋亡增加和Sost/sclerostin高表达。此外，DM小鼠的治疗 通过PTH 1 R起作用的PTH相关蛋白（PTHrP）衍生肽（1-37）纠正了这些问题， 改变，并激活生存信号，防止骨细胞凋亡。这项研究的长期目标是 目的是确定骨细胞及其产物治疗骨病的潜力。具体 本研究的目的是揭示PTH 1 R保护骨骼退化的潜在机制 DM中的信号。我们的假设是激活骨细胞中的PTH 1 R信号通路， （FDA批准的骨合成代谢剂）通过调节 骨细胞衍生因子，从而维持骨量和强度，保持骨细胞活力， 降低BMAT。该假设将使用已建立的1型和2型DM的鼠模型进行检验， 分别与低胰岛素血症和高胰岛素血症相关，并使用药理学和遗传学工具激活 或抑制PTH 1 R信号传导，并干扰骨细胞基因产物。我们将努力实现以下目标： 目的1将检测PTH或abaloparavine对PTH 1 R信号传导的药理学激活是否恢复 在1型和2型DM小鼠模型中（在近交系C57 BL/6和远交系Swiss韦伯斯特中）的骨量和强度 菌株）;并揭示潜在的细胞和分子机制。目的2将检查骨细胞对 通过研究PTH或abaloparthritis DM的作用，PTH 1 R信号传导对DM骨病的保护作用 小鼠和骨细胞中PTH 1 R缺失的对照小鼠（DMP 1 - 8 kb-Cre）。目标3将检查 通过研究骨细胞衍生的Wnt/βcatenin拮抗剂在DM诱导的骨骼退化中的作用， 无论是骨细胞中缺乏Sost、Dkk 1或两者的小鼠（Sostf/f; Dkk 1f/f; DMP 1 - 8 kb-Cre）还是表达 LRP 5高骨量突变pG 171 V（对Sost和Dkk 1介导的Wnt-β连环蛋白抑制具有抗性 信号传导）免受DM在骨中的损伤作用。

Recent advances in bone biology, to which research of my laboratory in part supported by the VA have significantly contributed, demonstrate that osteocytes (the most abundant bone cells) play a critical role in bone homeostasis by regulating the production and activity of osteoblasts and osteoclasts, the cells that form or dissolve bone, respectively. However, less is known about the function of osteocytes in bone pathophysiology. Our work showed that osteocytes are crucial target cells of parathyroid hormone (PTH) action and that activation of PTH 1 receptor (PTH1R) signaling in osteocytes increases bone formation and enhances bone remodeling, recognized features of PTH skeletal action. Osteocytic PTH1R signaling decreases the expression of Sost/sclerostin, an osteocyte-derived inhibitor of bone formation, and increases the expression of RANKL, the master inducer of osteoclast differentiation. We also showed that mice lacking the PTH1R in osteocytes exhibit decreased resorption and defective anabolic response to PTH. In more recent work, we established that the low bone mass and inferior mechanical and material properties exhibited by mice with diabetes mellitus (DM) is accompanied by decreased formation, increased resorption, and increased bone marrow adipocytes (BMAT), along with increased osteocyte apoptosis and high expression of Sost/sclerostin. Further, treatment of 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. Our hypothesis is that activating PTH1R signaling in osteocytes 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 murine models of established type 1 and type 2 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 type 1 and type 2 DM mouse models (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 DM mice 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 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 the damaging effects of DM in bone.