Patch clamp physiological techniques were used to characterize the voltage-activated calcium currents (VACC) expressed in the plasma membrane of osteoblastic cells as a function of time in culture and proliferative state of the cell. Osteoblast-enriched preparations were isolated by collagenase digestions of newborn rat calvaria and cultured under different conditions which affected cell proliferation (i.e., low serum in the media to arrest proliferation). VACC were isolated by replacing the intracellular potassium with cesium, and adding 1 microM tetrodotoxin to the bath. Under conditions that favored cell proliferation, low cell density, and media supplemented with 10% fetal calf serum (FCS), a transient calcium current was not expressed until day 3 in culture. There was a statistically significant relationship between the percentage of cells expressing this current and the time in culture. The magnitude of the current significantly increased as days in culture increased. Under the same conditions, the sustained VACC was detected after 7 or 8 days in culture. However, arresting cell proliferation after 2 days in culture by reducing the FCS concentration to 0.01% induced the expression of the sustained VACC the next day. The data suggest that the expression of VACC in the plasma membrane of rat calvarial osteoblasts depends on the time in culture and the state of proliferation of the cells. These results should prove to be valuable in studying the functional significance of VACC in osteoblastic cells and their regulation by various bone regulatory agents.
These results suggest that PDGF-BB is rapidly adsorbed then slowly released from the anorganic bovine bone-collagen matrix. PDGF-BB adsorbed to this material is able to stimulate proliferation of the attached osteoblastic cells. These data suggest that it may be clinically feasible to adsorb PDGF to this bone-collagen matrix and that this combination of bone growth factor and mineral-collagen matrix has the potential for clinical applications.
These results suggest that PDGF-BB can be adsorbed to the anorganic bovine bone mineral matrix and that this growth factor subsequently enhances the osteogenic properties of this bone graft material. IGF-I also adsorbed to the graft material; however, it was not readily released and it did not produce significant effects in the biologic assay. It appears that it may be clinically feasible to adsorb PDGF to anorganic bovine bone and that this combination of bone growth factor and mineral matrix has the potential for clinical applications.
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