Parathyroid Hormone and Osteoblast Mitogenesis

甲状旁腺激素和成骨细胞有丝分裂

• 批准号: 6654915
• 负责人: LOUIS M LUTTRELL
• 金额: $ 3.23万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-15 至 2003-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6654915
• 关键词: G protein coupled receptor kinase; SDS polyacrylamide gel electrophoresis; arrestins; autocrine; bone metabolism; cell differentiation; cell growth regulation; cell proliferation; confocal scanning microscopy; epidermal growth factor; genetically modified animals; hormone receptor; hormone regulation /control mechanism; immunoaffinity chromatography; laboratory mouse; metalloendopeptidases; mitogen activated protein kinase; osteoblasts; paracrine; parathyroid hormones; phosphorylation; physiologic bone resorption; polymerase chain reaction; prostaglandin receptor; receptor coupling; western blottings

DESCRIPTION (provided by applicant): Osteoblasts regulate the deposition of bone matrix protein and its subsequent mineralization. In situ, regulation of osteoblast proliferation, differentiation and function occurs via the complex interplay of extracellular signals mediated by steroid hormone and vitamin D receptors, receptor tyrosine kinases, and G protein-coupled receptors (GPCRs), such as those for parathyroid hormone (PTH) and prostaglandins. While it is clear that GPCRs transmit signals that are critical for the regulation of osteoblast metabolism, their significance as potential regulators of osteoblast growth and differentiation has only recently been appreciated. The broad goal of this research proposal is to determine how GPCRs regulate the growth and differentiation of osteoblasts through activation of the ERK mitogen-activated protein kinase (MAP) cascade, a key regulatory pathway in terms of both cell proliferation and differentiation. We provide preliminary data that demonstrate that GPCRs employ two novel mechanisms to direct the temporal and spatial activation of MAP kinases. First, matrix metalloprotease-mediated ectodomain shedding causes the release of autocrine ligands that induce "transactivation" of epidermal growth factor receptors (EGFRs). Second, beta-arrestins, proteins which bind to agonist-occupied GPCRs and uncouple them from their cognate G proteins, function as scaffolds for the component kinases of the ERK cascade and lead to the targeted activation of MAP kinase within specific cellular compartments. The first Aim of this proposal is to characterize the molecular mechanisms whereby PTH and prostaglandin receptors regulate the activity of MAP kinase cascades in osteoblasts. The second Aim is to determine the role of EGFR and beta-arrestin-dependent signals in regulating osteoblast proliferation, differentiation, and matrix production in vitro. These studies will employ osteoblastic cell lines and primary cultures of osteoblasts from mice in which EGFR or beta-arrestin function has been selectively inhibited. The third Aim of the proposal is to determine the role of EGFR and beta-arrestin-dependent signals in the control of anabolic bone metabolism by PTH in vivo. These studies will employ transgenic mouse models, in which beta-arrestins have been knocked out by homologous recombination or EGFR function has been impaired through osteoblast-specific expression of a dominant inhibitory mutant EGFR. By increasing our understanding of the mechanisms of GPCR signaling in bone, these studies may permit the rational development of safe strategies for employing PTH analogues to modulate osteoblast number and/or function.

描述（由申请人提供）：成骨细胞调节 骨基质蛋白及其随后的矿化。在现场， 成骨细胞的增殖、分化和功能通过复合物发生， 类固醇激素和维生素D介导细胞外信号相互作用 受体、受体酪氨酸激酶和G蛋白偶联受体（GPCR）， 如甲状旁腺激素（PTH）和甲状旁腺素。虽然 清楚地表明，GPCR传递的信号对调节 成骨细胞代谢及其作为成骨细胞潜在调节因子的意义 生长和分化只是最近才受到重视。广泛的目标 这项研究的目的是确定GPCR如何调节生长， 成骨细胞的分化通过激活ERK丝裂原活化 蛋白激酶（MAP）级联反应，一个关键的调节途径，在这两个细胞 增殖和分化。我们提供的初步数据表明， GPCR采用两种新的机制来指导时间和空间 MAP激酶的激活。第一，基质金属蛋白酶介导的胞外域 脱落引起诱导“反式激活”的自分泌配体的释放， 表皮生长因子受体（EGFR）。第二，β-抑制蛋白，蛋白质 其与激动剂占据的GPCR结合并将它们与其同源的G 蛋白质，作为ERK级联的组分激酶的支架起作用 并导致特定细胞内MAP激酶的靶向激活， 隔间本建议的第一个目的是表征分子 PTH和前列腺素受体调节成骨细胞中MAP激酶级联活性的机制。第二个目的是确定EGFR的作用， 和β-抑制蛋白依赖性信号在调节成骨细胞增殖中的作用， 分化和体外基质生产。这些研究将采用 成骨细胞系和来自小鼠的成骨细胞的原代培养物，其中 EGFR或β-抑制蛋白功能已被选择性抑制。第三个目标 该提案旨在确定EGFR和β-arrestin依赖性 在体内通过PTH控制合成代谢骨代谢的信号。这些 研究将采用转基因小鼠模型，其中β-抑制蛋白已被 被同源重组敲除或EGFR功能受损 通过成骨细胞特异性表达显性抑制突变EGFR。通过 增加我们对骨中GPCR信号传导机制的理解，这些 研究可能允许合理开发安全策略， PTH类似物调节成骨细胞数量和/或功能。