PTH INDUCES RGS2, NURR1 & NFIL3/E4BP4 EXPRESSION IN BONE

描述（改编自研究者摘要）：骨的数量和质量是牙科手术后临床结果的主要决定因素。改善骨代谢和促进骨细胞适应治疗要求的合成代谢药物可改善牙科治疗和预后。甲状旁腺激素（PTH）是血清钙的主要调节剂，对骨代谢具有合成代谢和分解代谢作用，这取决于其间歇或连续给药。了解PTH这些相反作用的机制可能会导致更有效的骨合成代谢治疗。甲状旁腺激素激活的信号传导迅速诱导几种成骨基因的转录。这些主要反应基因是第一个受PTH治疗影响的基因。主要研究者已经确定了三个主要的反应基因，快速和短暂诱导的PTH在成骨细胞。它们是G蛋白信号传导的调节因子、RGS 2、孤儿核受体CR 1 R1和转录因子NFIL 3/E4 BP 4。他假设这些基因介导PTH对骨代谢的影响。为了解决这一假设，主要研究者提出了三个具体目标：（1）确定介导PTH诱导的RGS 2、IGFR 1和NFIL 3/E4 BP 4基因表达的信号通路。成骨细胞和小鼠颅骨将用PTH激活的信号传导通路的激动剂和抑制剂处理。将分别通过北方印迹和/或RT-PCR、核运行试验和蛋白质印迹分析测量RGS 2、RGSR 1和NFIL 3/E4 BP 4 mRNA基因转录和蛋白质水平。(2)确定RGS 2、CRISPR 1和NFIL 3/E4 BP 4蛋白在成骨细胞功能和PTH调控成骨细胞基因表达中的重要性。RGS 2、IGFR 1和NFIL 3/E4 BP 4蛋白将通过反义策略过表达或阻断，并且将研究对成骨细胞增殖和分化的影响。还将测试PTH在过表达或缺乏RGS 2、CRISPR 1和NFIL 3/E4 BP 4蛋白的细胞中调节基因表达的能力。(3)检测PTH在体内诱导的RGS 2、RGS 1 R1和NFIL 3/E4 BP 4基因表达。将用间歇或连续PTH（已知分别为合成代谢和分解代谢）处理小鼠，并将在骨和其他PTH靶组织中进行原位杂交。这些实验将有助于理解PTH诱导RGS 2、NF 1 R1和NFIL 3/E4 BP 4基因表达的机制，并揭示这些基因在PTH调控成骨细胞基因表达中的作用。

DESCRIPTION (adapted from the Investigator's abstract): Bone quantity and quality are major determinants of clinical outcome following dental surgery. Anabolic agents that improve bone metabolism and promote bone-cell adaptation to treatment requirements improve dental treatment and prognosis. Parathyroid hormone (PTH), the major regulator of serum calcium, has anabolic and catabolic effects on bone metabolism depending on its intermittent or continuous administration. Understanding the mechanisms that underlie these opposing actions of PTH might lead to more effective anabolic treatments for bone. PTH-activated signaling rapidly induces transcription of several osteoblastic genes. These primary response genes are the first genes to be affected by PTH treatment. The Principal Investigator has identified three primary response genes that are rapidly and transiently induced by PTH in osteoblasts. They are the regulator of G-protein signaling, RGS2, the orphan nuclear receptor NURR1, and the transcription factor NFIL3/E4BP4. He hypothesizes that these genes mediate PTH effects on bone metabolism. To address this hypothesis, the Principal Investigator proposes three specific aims: (1) to identify the signaling pathway(s) that mediate PTH-induced RGS2, NURR1 and NFIL3/E4BP4 gene expression. Osteoblastic cells and mouse calvariae will be treated with agonists and inhibitors of the signaling pathways activated by PTH. RGS2, NURR1 and NFIL3/E4BP4 mRNA gene transcription and protein levels will be measured by Northern blot and/or RT-PCR, nuclear run-on assays and Western blot analysis, respectively. (2) To identify the importance of RGS2, NURR1 and NFIL3/E4BP4 proteins in osteoblastic function and PTH regulation of osteoblastic gene expression. RGS2, NURR1 and NFIL3/E4BP4 proteins will be overexpressed or blocked by antisense strategies and the effects on osteoblastic proliferation and differentiation will be studied. The ability of PTH to regulate gene expression in cells overexpressing or lacking RGS2, NURR1 and NFIL3/E4BP4 proteins will also be tested. (3) To examine PTH-induced RGS2, NURR1 and NFIL3/E4BP4 gene expression in vivo. Mice will be treated with intermittent or continuous PTH, known to be anabolic and catabolic, respectively and in situ hybridization will be performed in bone and other PTH-target tissues. The experiments proposed will help understand the mechanisms of PTH-induction of RGS2, NURR1 and NFIL3/E4BP4 gene expression and unveil the role of these genes in the PTH regulation of osteoblastic gene expression.