SEQUENTIAL EXPRESSION OF BONE-MATRIX PROTEINS DURING RAT CALVARIA OSTEOBLAST DIFFERENTIATION AND BONE NODULE FORMATION IN-VITRO

We investigated the expression of osteocalcin (OC), bone sialoprotein (BSP), osteonectin (ON), and alkaline phosphatase (ALP) during cell di fferentiation and bone nodule formation by fetal rat calvaria cells, u sing immunofluorescent and immunogold techniques at light and electron microscopic levels. Six hours after plating all proteins were express ed in calvaria cells. However, expression was not detected during the proliferation phase after plating. Cell morphological modifications we re observed in osteoblastic cells expressing ALP, OC, and BSP, but not ON. During the matrix formation phase, all proteins were expressed wi th various intensities and OC was limited to differentiated osteoblast ic cells. EM observations demonstrated that BSP was selectively associ ated with clusters of needle-like crystals, but not with collagen fibe rs, in mineralization foci and in the mineralized matrix. OC was local ized intracellularly and in all the extracellular compartments, and wa s concentrated at the mineralization front. ON was distributed uniform ly throughout the osteoid and mineralized matrix, which was intensely labeled. The results show that the expression of bone matrix proteins during differentiation of calvaria cells and nodule formation in vitro duplicate what is observed during osteogenesis in vivo.