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.
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