| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 2000: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1999: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
We obtained following three findings on the function of bone sialoprotein(BSP).
1. We investigated molecular interaction between BSP and Type I collagen (COL1) molecules by observing the modification of the time course of the COL1 fibrillogenesis in vitro, BSP dramatically promoted the COL1 fibrillogensis at slightly acidic (pH6.55) condition, dose depehdently. However, in near neutral (pH7.25) or slightly alkaline (pH7.67) condition, BSP did not affect the kinetics. Decorin, plasma fibronectin, and chondroitin sulfate did not affect the kinetics, at any pH conditions. This findings suggested that BSP performed some functions in bone turnover at the initiation of bone formation in the resorption lacuna of acidic milieu just after osteoclasts dettached in vivo.
2. The cell attachment assay by osteoblastic cell line (MC-3T3 E1 cell) was performed, on the plastic surface coated by only BSP, and on the surface coated by at first by BSP, thereafter, coated by plasma fibronectin(PFN) secondly. The surface coated by pFN after BSP collected significantly more cells than the surface coated only by BSP. This cell behavior suggested that pFN bound BSP that had already bound on the surface. This result suggested that in vivo BSP functions as an anchor for the other ECM (extracellular matrix) molecules, such as FN, etc. on hydroxyapatite crystal.
3. The existence of BSP in blood mede us looked for the serum components which interacted with BSP. We found at least one molecule which bound to BSP fixed on agarose gel. This substance was determined to thrombospondin (TSP1) by Western blot. This results, TSP1 binds BSP in vitro, suggests that BSP is functioning as an anchor in bone ECM for TSP1 derived from blood circulation or secreted by osteoblasts. BSP may provides TSP1 with the milieu where TSP interacts with such bioactive ECM proteins.
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