GROWTH-PLATE MATRIX VESICLE BIOGENESIS - THE ROLE OF INTRACELLULAR CALCIUM

Isolated bovine growth plate chondrocytes suspended in a synthetic car tilage buffer containing 10(-7) to 10(-3) M concentrations of calcium were treated with 1 mu M ionomycin to induce changes in intracellular ionized calcium as measured by Fura-2 fluroscence. An increase in intr acellular calcium of 10(-6) to 10(-4) M calcium resulted in the secret ion of vesicles that contain alkaline phosphatase. An increase in intr acellular calcium above 10(-4) M resulted in nonspecific cell fragment ation because of cell damage. Electron microscopy of the secreted vesi cles demonstrated that their structure resembled matrix vesicles. Isol ated cells treated with ionomycin (synthetic cartilage buffer with 10( -6) M calcium) demonstrated intact cell membranes, normal intracellula r architecture, and numerous plasma membrane processes undergoing vesi culation. The plasma membrane of these isolated cells and the isolated vesicles demonstrated a positive stain for alkaline phosphatase. Fres h growth plate tissue demonstrated plasma membrane staining for alkali ne phosphatase at the lower proliferative and upper hypertrophic cell zone, and an increasing number of alkaline phosphatase stained matrix vesicles in the matrix of the lower hypertrophic cell zone. The data i ndicate that an increase in intracellular calcium in the lower prolife rative and hypertrophic cell zones of the growth plate induces the sec retion of plasma membrane derived matrix vesicles.