Genetic Analysis of Second Messengers in PTH Signaling in Bone

骨 PTH 信号传导第二信使的遗传分析

• 批准号: 7627067
• 负责人: HENRY M. KRONENBERG
• 金额: $ 39.62万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-12-01 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7627067
• 关键词: Affect; Age; Anabolic Agents; Apoptosis; Apoptotic; Birth; Bone Resorption; Calcium; Cell Survival; Cells; DEXA; Data; Development; Diet; Disease; Dissection; Doxycycline; Excision; Fibroblast Growth Factor; Flow Cytometry; Fracture; Gene Expression; Generations; Genes; Genetic Models; Goals; Heterotrimeric GTP-Binding Proteins; Hormonal; In Situ Hybridization; In Vitro; Infusion procedures; Injection of therapeutic agent; Kidney; Knock-in Mouse; Knock-out; Link; Measurement; Mediating; Mediator of activation protein; Modeling; Molecular; Mus; Mutate; Organ; Osteoblasts; Osteoclasts; Osteocytes; Osteogenesis; Osteoporosis; Parathyroid Hormone Receptor; Partner in relationship; Pathway interactions; Phospholipase C; Physiology; Receptor Activation; Role; Second Messenger Systems; Serological; Signal Transduction; Staging; Stromal Cells; Sum; TRANCE protein; Tetanus Helper Peptide; Tetracyclines; Time; Withdrawal; bone; bone mass; calcium metabolism; genetic analysis; in vivo; in vivo Model; mutant; osteoblast differentiation; parathyroid hormone-related protein; promoter; receptor; recombinase; response; second messenger

Understanding the mechanisms of the anabolic action of PTH on bone is important both because PTH represents the first anabolic agent useful to treat osteoporosis and because a molecular understanding of PTH's anabolic action will allow the development of even more effective anabolic agents. Our goal here is to use in vivo models to establish the roles of specific "second messenger" pathways responsible for the various actions of PTH that, in sum, result in an increase in bone mass. Understanding of the multiple, specific pathways leading in varying ways to increased bone will make possible the next rational generation of anabolic agents. We also want to understand these mechanisms to understand better the roles of PTH and PTHrP in normal physiology and disease. PTH and PTHrP stimulate both bone formation and bone resorption through activation of the PTH/PTHrP receptor (PTHR1) in cells of the osteoblast lineage. The PTHR1 activates several heterotrimeric G proteins, with Gs and Gq/11 being the best characterized. We will use two genetic models that allow the separation of the actions of these downstream mediators in vivo. One model uses a "knock-in" mouse in which the normal PTHR1 gene has been mutated to selectively disrupt the activation of phospholipase C by the Gq/11 pathway without affecting activation of Gs. The other model is a conditional knockout in which the osterix promoter, active in early osteoblasts, drives the expression of ere recombinase in a way that can be suppressed by tetracycline derivatives ("tet-off"). This ere is used to ablate expression of Gsa postnatally in early cells of the osteoblast lineage through mating with a Gsa floxed mouse and administration of doxycycline for various times. Three models of receptor activation (continuous elevation of PTH through minipump infusion or low calcium diet, and once daily injection of PTH) will be used to determine the roles of distinct pathways in the variety of actions of the PTHR1 on cells of the osteoblast lineage. Aim 1. Role of PLC signaling in actions of the PTHR1 in bone. Models of intermittent and continuous PTH administration will be used to determine the role of PTHR1 activation of PLC in bone.Aim 2. Role of Gs signaling in cells of the osteoblast lineage. Comparison of mice missing Gsa in osteoblastic cells with the same mice exposed to continuous or intermittent elevation of PTH levels should allow identification of the responses to activation of the PTHR1 that require Gsalpha

了解PTH对骨骼的合成代谢作用的机制很重要，因为PTH 代表第一个用于治疗骨质疏松症的合成代谢剂，并且是因为分子理解 PTH的合成代谢作用将允许开发更有效的合成代谢剂。我们的目标是 使用体内模型来确定负责特定“第二使者”途径的作用 总之，PTH的各种作用会导致骨骼量的增加。对倍数的理解， 以不同方式导致骨骼增加的特定途径将使下一代成为可能 合成代谢剂。我们也想了解这些机制，以更好地理解PTH的作用 正常生理和疾病中的PTHRP。 PTH和PTHRP刺激骨形成和骨骼 通过在成骨细胞谱系细胞中激活PTH/PTHRP受体（PTHR1）的吸收。这 PTHR1激活了几种异三聚体G蛋白，GS和GQ/11的特征最佳。我们将 使用两个遗传模型，可以将这些下游介体在体内的作用分离。一 模型使用一种“敲入”小鼠，其中正常的PTHR1基因已突变以选择性破坏 通过GQ/11途径激活磷脂酶C，而不会影响GS的激活。另一个模型是 有条件的敲除，其中的Osterix启动子（活性在早期成骨细胞中）驱动ERE的表达 重组酶以可以被四环素衍生物抑制的方式（“ Tet-Off”）抑制。这个习惯了 与GSA Floxed交配，在成骨细胞谱系的早期细胞中的GSA表达在成骨细胞谱系的早期细胞中 小鼠和多西环素的不同时间。受体激活的三种模型（连续 PTH通过微小注输注或低钙饮食升高，一旦每天注射PTH）将被使用） 确定不同途径在PTHR1在成骨细胞细胞中的各种作用中的作用 血统。 AIM 1。PLC信号在PTHR1在骨骼中的作用中的作用。间歇性和 连续的PTH给药将用于确定PLC在骨中的PTHR1激活的作用。IAM2。 GS信号在成骨细胞谱系细胞中的作用。比较成骨细胞中缺少GSA的小鼠 相同的小鼠暴露于PTH水平的连续或间歇性升高应允许识别 需要GSALPHA的PTHR1激活的响应