| 3577 Roles of biglycan on collagen matrix maturation and organization | ||
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D. PARISUTHIMAN1, Y. MOCHIDA2, W.R. DUARTE2, S. PORNPRASERTSUK3, and M. YAMAUCHI2, 1Curriculum in Oral Biology, The University of North Carolina at Chapel Hill, USA, School of dentistry, Thammasat University, Thailand, 2University of North Carolina, Chapel Hill, USA, 3Curriculum in Oral Biology, The University of North Carolina at Chapel Hill, USA, Faculty of dentistry, Mahidol University, Bangkok, Thailand Fibrillar type I collagen provides a stable template for mineral deposition and growth in most mineralized tissues. Therefore, the organization and stability of collagen matrix are crucial for subsequent collagen mineralization. Through its ability to interact with collagen, biglycan (BGN), a member of the Small Leucine Rich Proteoglycans (SLRPs), has been implicated in collagen organization/maturation and, therefore, is potentially involved in collagen mineralization. OBJECTIVE: To investigate the effects of biglycan overexpression on collagen organization and maturation. METHODS: MC3T3-E1 cells were transfected with a pcDNA3.1/V5-His TOPO vector containing the full-length mouse Bgn cDNA and stable cell clones were generated. To establish clones expressing higher levels of BGN (S clones), the levels of BGN-V5 fusion protein were evaluated by immunoprecipitation followed by Western Blot analysis using anti-V5 antibody. Proliferation rates of S clones and MC3T3-E1 cells were examined by cell counting at days 3, 5 and 8. To assess the collagen organization and maturation, S clones and MC3T3-E1 cells were cultured in the presence of 2 mM b-glycerophosphate and 50 mg/ml ascorbic acid for 20 days, stained with picosirius red and examined under a polarized light microscope. RESULTS: Several S clones expressing higher levels of BGN were established. The proliferation rates of S clones were similar to those of MC3T3-E1 cells. Picosirius red staining demonstrated that the collagen fibrils in MC3T3-E1 cultures were overall mature and well organized with relatively distinct direction. However, in S cultures there were significant amounts of immature collagen matrix and the direction was not as distinct as that of MC3T3E1 cells. The presence of these immature/disorganized collagen fibrils in S clones was proportional to the level of BGN expression. CONCLUSION: These results indicate that BGN may regulate collagen matrix organization and maturation in vitro. Supported by NIH grant DE10489 NASA NAG2-1596. | ||
| Seq #365 - Bone and Tooth Matrix Biology 10:15 AM-11:30 AM, Saturday, 13 March 2004 Hawaii Convention Center Exhibit Hall 1-2 | ||
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