Function of Regulator of G protein signaling in aging skeleton

描述（申请人提供）：骨质疏松症是一种主要的健康问题，其特征是随着年龄的增长，破骨细胞生成和活性增加，导致骨重建和代谢失衡。目前的治疗选择受到反应率较低或副作用的限制，无法有效解决骨质疏松症的负担和异质性。建立成功的治疗方法的进一步进展迫切需要更清楚地了解破骨细胞（OC）分化和骨重建随年龄的机制。有证据表明，RANKL诱发的Ca 2+振荡在PLC β和NFATc 1的激活以及OC分化中起着开关作用。进一步的研究表明，FcR β和DAP 12调节PLC β的磷酸化，PLC β是RANKL诱导的Ca 2+振荡-NFATc 1通路中的关键组分。尽管有这些新的见解，但目前仍不清楚RANKL如何引起触发NFATc 1激活和OC分化的基本Ca 2+振荡，也不清楚这种机制是否以及如何参与年龄相关的骨丢失。我们最近的研究表明，G蛋白信号传导蛋白12（RGS 12）的调节器在OC分化过程中显着表达，并且RGS 12的表达随着年龄的增加而显着增加。RGS 12表达的敲低抑制了Ca 2+振荡、NFATc 1表达和OC分化。从造血/早期OC谱系细胞（RGS 12/ /cre）中缺失RGS 12导致骨量显著增加，并且这种骨量增加在RGS 12/ /cre小鼠中免受年龄相关的骨丢失。我们还发现，小鼠中RGS 12的缺失会随着年龄的增长而降低OC数量和骨髓脂肪生成水平。基于这些结果，我们假设与年龄相关的骨重建和代谢需要RGS 12的表达和功能，并且RGS 12是控制衰老过程中Ca 2+振荡和OC分化的关键调节剂。我们将通过以下两个具体目标来检验这一假设。目的1：通过对RGS 12基因敲除小鼠的表型和病理机制的研究，揭示RGS 12在OC分化和衰老过程中的作用。我们将使用Mx 1-cre和Cathepsin K-Cre小鼠分析年轻（6个月（m））、中年（14个月）和老年（24个月）小鼠的骨表型，以确定早期和晚期OC谱系细胞中RGS 12的缺失如何影响出生后年龄相关的骨代谢。我们将进一步在体外表征来自老化RGS 12/ /cre小鼠的OC，并确定间充质谱系细胞对老化RGS 12/ /cre小鼠中OC和骨骼表型的贡献。在目标2中，我们将阐明RGS 12相互作用的机制，赋予特异性的Ca 2+振荡和OC与年龄的特征病理机制和途径，RGS 12结构域功能和其异源二聚化的合作伙伴。本项目的目的是研究RGS 12在病理性年龄条件下OC分化和活化中的作用和机制，为年龄相关性骨质疏松症和其他骨疾病提供新的、更有效的治疗靶点。

DESCRIPTION (provided by applicant): Age-associated osteoporosis is a major health problem, which is characterized by an imbalance in bone remodeling and metabolism due to the increase of osteoclastogenesis and activity with age. The current treatment options have been constrained with lower response rates or side-effects and ineffectively tackle the burden and heterogeneity of osteoporosis. Further progress in establishing successful treatment urgently requires a clearer understanding of the mechanisms of osteoclast (OC) differentiation and bone remodeling with age. Evidences have documented that RANKL -evoked Ca2+ oscillations play a switch-on role in the activation of PLC� and NFATc1, and OC differentiation. Further study showed that FcR� and DAP 12 regulate the phosphorylation of PLC�, which is the critical component involved in the RANKL-induced Ca2+ oscillations- NFATc1 pathway. Despite these new insights, it remains unclear how RANKL evokes the essential Ca2+ oscillations that triggers NFATc1 activation and OC differentiation, and it is unknown whether and how this mechanism is involved in age-associated bone loss. Our recent research demonstrated that Regulator of G protein signaling protein 12 (RGS12) was prominently expressed during OC differentiation and that RGS12 expression was significantly increased with increasing age. Knockdown of RGS12 expression inhibited Ca2+ oscillations, NFATc1 expression and OC differentiation. Deletion of RGS12 from hematopoietic/early OC lineage cells (RGS12 / /cre) led to a significantly increase of bone mass, and this increase of bone mass was protected from age-associated bone loss in RGS12 / /cre mice. We also found that loss of RGS12 in mice decreased OC number and levels of marrow adipogenesis with age. Based on these results, we hypothesize that age-associated bone remodeling and metabolism requires RGS12 expression and function, and RGS12 is a critical regulator in controlling Ca2+ oscillation and OC differentiation during aging. We will test the hypothesis through the following two specific aims. In Aim 1, we will reveal the function of RGS12 in OC differentiation and function during aging through characterization of the phenotypes and pathomechanism of RGS12 conditional knockout mice. We will analyze bone phenotypes in young (6 months (m)), mid-aged (14m) and aged (24m) mice to determine how deletion of RGS12 in early and late OC lineage cells affect postnatal age-associated bone metabolism by using Mx1-cre and Cathepsin K-Cre mice. We will further characterize in vitro of OCs derived from aging RGS12 / /cre mice and determine the contribution of mesenchymal lineage cells to the OCs and skeletal phenotype in aging RGS12 / /cre mice. In Aim 2, we will elucidate the mechanism of RGS12 interactions that confer specificity of Ca2+ oscillation and OCs with age by characterizing pathomechanism and pathways, RGS12 domain functions and its heterodimerization partners. The goal of this project is to discover the role and mechanism of RGS12 in OC differentiation and activation in pathologic age condition, and provide new and more effective therapeutic targets to age-associated osteoporosis and other bone diseases.