| 2984 Phosphophoryn -exon5 of DMP3- regulates osteoblast gene expression and mineralization | ||
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J.A. JADLOWIEC1, H. KOCH2, M. SEYEDAIN3, P.G. CAMPBELL1, and C. SFEIR3, 1Carnegie Mellon University, Pittsburgh, PA, USA, 2University of Greifswald, Griefswald, Germany, 3University of Pittsburgh, PA, USA Background: The ECM is a dynamic microenvironment consisting of signals that regulate tissue morphogenesis and remodeling. ECM proteins serve as both a structural support for cells and as signals for cell behavior. We hypothesize that non-collagenous bone/dentin ECM proteins regulate bone marker gene expresssion and stimulate mineralization. Objectives: To determine the role of bone/dentin extracellular matrix proteins such as Phosphophoryn (PP) in cell signaling and differentiation by: 1) Quantify gene expression of osteoblast marker genes in response to PP. 2) Measure phenotypic response to PP. 3) Determine the mechanism of action for PP. Methods: We exploited quantitative PCR (qPCR) to quantify gene expression levels of Runx2, Osterix (Osx), bone/liver/kidney alkaline phosphatase (Alp), Osteocalcin (Ocn) and bone sialoprotein (Bsp) in response to PP. To measure phenotypic outcome, we measured OCN secretion by ELISA, ALP activity and calcium deposition by alizarin red staining. The mechanism of action was determined by inhibiting signaling pathways during PP treatment. Significant differences among treatment groups were determined by a one-way ANOVA with Fisher's LSD post hoc test at a 95% confidence level. Results: PP up-regulated Runx2, Osx, Alp and Ocn in primary human adult mesenchymal stem cells (hMSC), a mouse osteoblastic cell line (MC3T3-E1) and a mouse fibroblastic cell line (NIH3T3) over untreated controls. PP increased OCN secretion and ALP activity in hMSC and MC3T3-E1 and calcium deposition in hMSC. The mechanism by which PP exerts its effect was identified and will be presented. Conclusions: We reason that PP functions in osteogenesis/dentinogenesis in two primary roles. First, PP binds to cell surface receptors activating intracellular signaling pathways that culminate in expression of osteoblast marker genes such as Runx2, Osx, Alp and Ocn. Second, PP is localized within the ECM of osteoblasts and pre-osteoblasts and functions as a regulator of calcium phosphate deposition and crystal growth. | ||
| Seq #324 - Senior Category 10:15 AM-11:30 AM, Saturday, 13 March 2004 Hawaii Convention Center Exhibit Hall 1-2 | ||
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