KROX 20 AND BONE MASS

描述（由申请人提供）：Krox 20和骨量。我们建议描绘的作用，转录因子Krox 20成骨细胞的功能和骨量控制的调节。在该项目的上一个资助周期中，我们发现Krox 20的表达和转录活性受到糖皮质激素（GC）的强烈抑制，有助于抑制培养成骨细胞中的骨钙素转录。我们建议将该项目扩展到GC诱导的骨质疏松症（GIO）问题之外，并研究Krox 20在体内和体外基础成骨细胞生物学中的作用。这样的作用通过以下方式提示：（i）Krox 20-/-小鼠中骨小梁形成的发育停滞;（ii）成熟Krox 20小鼠中低的骨小梁质量和降低的骨形成速率;（iii）Krox 20和Krox 20-/-成骨细胞培养物中受损的矿化;（iv）Wnt信号通路刺激Krox 20表达和活性;（iv）Wnt信号通路刺激Krox 20表达和活性。和（v）与Krox 20在成骨细胞功能中的作用一致的体内和体外表达模式。我们将分析Krox 20不足和/或成骨细胞特异性过量小鼠的骨骼表型随年龄的变化。体内方法将包括显微计算机断层扫描（μ CT）以及组织学和血清学分析。Krox 20的细胞和分子靶点将通过分析成骨细胞培养物来确定，其中Krox 20的表达已通过遗传操作或病毒感染而改变。体外分析将在成骨细胞表型发育期间以短时间间隔进行，以鉴定Krox 20敏感性分化阶段。Krox 20的具体作用将在细胞周期进程、细胞凋亡、生化标志物的发展和促进成骨细胞表型的基因表达方面确定，包括可能由Krox 20直接调节和由Wnt信号传导途径间接调节的基因。最后，我们将回到新闻局的调查。我们将评估GCs对Krox 20在小鼠GIO发育的详细描述（包括mu CT和细胞凋亡）的背景下体内表达的影响;并且，我们将确定用Krox 20病毒或重组Wnt 3A治疗是否拮抗GCs在成骨细胞中的不良作用。这些研究将阐明在健康和疾病中控制成骨细胞功能和骨形成的新的调节机制，并为骨合成代谢开辟新的途径。

DESCRIPTION (provided by applicant): Krox20 and Bone Mass. We propose to delineate the role of the transcription factor Krox20 in the regulation of osteoblast function and bone mass control. During the previous funding cycle of this project, we discovered that Krox20 expression and transcriptional activity were strongly inhibited by glucocorticoids (GCs), contributing to the repression of osteocalcin transcription in cultured osteoblasts. We propose to expand the project beyond the problem of GC-induced osteoporosis (GIO) and study the role of Krox20 in basic osteoblast biology in vivo and in vitro. Such a role is suggested by: (i) developmental arrest of trabecular bone formation in Krox20-/- mice; (ii) low trabecular bone mass and decreased bone formation rate in mature Krox20 mice; (iii) impaired mineralization in Krox20 and Krox20-/- osteoblast cultures; (iv) stimulation of Krox20 expression and activity by the Wnt signaling pathway; and (v) expression pattern in vivo and in vitro consistent with a role for Krox20 in osteoblast function. We will analyze as a function of age the skeletal phenotype of mice with insufficient and/or osteoblast-specific excess of Krox20. In vivo approaches will include microcomputed tomography (mu CT) as well as histological and serological analyses. The cellular and molecular targets of Krox20 will be determined analyzing osteoblast cultures in which Krox20 expression has been altered by either genetic manipulations or viral infection. The in vitro analyses will be performed at short intervals during the development of the osteoblast phenotype to identify the Krox20-sensitive differentiation stage. The specific roles of Krox20 will be determined with respect to cell cycle progression, apoptosis, development of biochemical markers and expression of genes that promote the osteoblast phenotype, including genes potentially regulated directly by Krox20 and indirectly by the Wnt signaling pathway. Finally, we will return to the investigation of GIO. We will assess the effect of GCs on Krox20 expression in vivo in the context of a detailed description (including mu CT and apoptosis) of GIO development in the mouse; and, we will determine whether treatment with a Krox20 virus or recombinant Wnt3A antagonize adverse effects of GCs in osteoblasts. These studies will elucidate novel regulatory mechanisms that control osteoblast function and bone formation in health and disease, and open new avenues in the pursuit of bone anabolics.