Genetic Analysis of Second Messengers in PTH Signaling in Bone

了解甲状旁腺素对骨的合成代谢作用的机制很重要，因为 代表了第一种用于治疗骨质疏松症的合成代谢剂，因为对 PTH的合成代谢作用将使开发更有效的合成代谢制剂成为可能。我们的目标是 利用活体模型建立特定的“第二信使”通路的作用 甲状旁腺激素的各种作用，总而言之，导致骨量增加。理解多个， 以不同方式通向骨骼增加的特定途径将使下一代理性成为可能 合成代谢药剂。我们也想了解这些机制，以便更好地了解甲状旁腺素的作用 和PTHrP在正常生理和疾病中的作用。甲状旁腺素和甲状旁腺激素受体促进骨形成和骨形成 成骨细胞系通过激活甲状旁腺激素/甲状旁腺素受体(PTHR1)进行吸收。这个 PTHR1可激活多种异源三聚体G蛋白，其中Gs和GQ/11的活性最高。我们会 使用两种遗传模型，允许在体内分离这些下游介体的作用。一 该模型使用了一种“敲入”小鼠，在该小鼠中，正常的PTHR1基因发生了突变，选择性地扰乱了 通过GQ/11途径激活磷脂酶C，而不影响Gs的激活。另一种模式是 在早期成骨细胞中活跃的Osterix启动子驱动ERE表达的条件性基因敲除 重组酶以可被四环素衍生物抑制的方式(tet-off)。这个地方是用来 GSA与成骨细胞系交配后早期细胞中GSA的表达 给小鼠灌胃多西环素不同时间。受体激活的三种模式(连续 通过小泵输注或低钙饮食升高甲状旁腺素，每天注射一次甲状旁腺素) 确定不同途径在PTHR1对成骨细胞作用的不同中的作用 血统。目的1.PLC信号在PTHR1在骨中的作用。断断续续的模型和 持续给予甲状旁腺素将被用来确定PTHR1在骨内PLC激活中的作用。 Gs信号在成骨细胞系细胞中的作用。小鼠成骨细胞缺失GSA的比较 同样的小鼠持续或间歇性地升高甲状旁腺素水平应该可以识别 对PTHR1激活的反应中需要Gsalpha

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