In vitro action of 1,25-dihydroxycholecalciferol and 24,25-dihydroxycholecalciferol on matrix organization and...

Growth plates of 18-day-old rabbits were incubated in a protein-free synthetic medium, either without any additive, with 1,25-dihydroxycholecalciferol [1,25-(OH)2D3] (10(-10) M), with 24,25-dihydroxycholecalciferol [24,25-(OH)2D3] (10(-10) M and 10(-9) M), with both metabolites, or with the ethanol solvent alone. Cartilages, before and after 5 days of incubation, were studied by light and electron microscopy. The intracellular calcium distribution was analyzed by the potassium pyroantimonate method, and the calcium content was verified by x-ray microprobe analysis. When compared to nonincubated samples the cartilages incubated for 5 days without any additive as well as the cartilages incubated with the solvent alone showed excessive hydratation and hypertrophy of the chondrocytes, which had lost their columnar arrangement. The matrix and the cells were devoid of mineral. The ultrastructure of the cells was well preserved. These changes were largely prevented by the presence of both vitamin D3 metabolites. With regard to calcium distribution, 1,25-(OH)2D3 maintained calcium in mitochondria and crystals in matrix vesicles, whereas 24,25-(OH)2D3 only partly maintained mitochondrial mineral. In the chondrocytes incubated with this latter metabolite, small calcium granules were seen in the cytoplasm; most vesicles were devoid of crystals, and amorphous precipitates were seen in the matrix. These data demonstrate the in vitro influence of vitamin D3 metabolites on the organization and mineralization of the cartilage matrix and on the distribution of intracellular calcium in chondrocytes. Furthermore, they support the hypothesis that the in vitro action of 1,25-(OH)2D3 is different from that of 24,25-(OH)2D3 in that 1,25-(OH)2D3 may influence calcium storage in mitochondria and matrix vesicles, whereas 24,25-(OH)2D3 is likely to be involved in calcium transport and release.