Hypervitaminosis D in the chick embryo: Comparative study on the activity of various vitamin D₃ metabolites

Summary

Chick embryos were injected in the yolk sac at various ages with various doses of different vitamin D₃ metabolites. Serum concentrations of total calcium and inorganic phosphate were determined 24 h after the injection and histological and electron microscopic studies of the tibiae were conducted 3–6 days after. Confirming previous results, the injection of 1,25(OH)₂D₃ was found to produce significant hypercalcemia and hypophosphatemia. The dose required to produce these effects decreased with age: 100 ng on the 9th day, 50 ng on the 11th, and 10 ng on the 15th. This finding is interpreted as resulting from the fact that the specialized cells in the chorionic epithelium which are considered to be involved in mineral resorption from the shell differentiate between the 11th and 13th days. Although no bone changes were observed in embryos injected before the 11th day, a rim of unmineralized trabeculae (osteoid) was observed at the periphery of the cortex of the tibial diaphysis in the embryos which had been injected after that age. Thus, in embryos injected on the 11th day with 100 ng 1,25(OH)₂D₃, the trabeculae formed between the 11th and 14th day remained unmineralized until the 15th or 16th day at which time they completed their mineralization. In the embryos injected on the 14th day, the alterations were more severe and could be produced with doses 10 times smaller than those required when the injections were made on the 11th day. At all ages, the doses that produced an osteoid rim also induced hypercalcemia and hypophosphatemia. The electron microscopical study of the osteoid trabeculae showed that osteoblasts and osteocytes had normal cytological characteristics and that the bone matrix did not present changes other than the reduction in mineral deposition. While the above findings do not exclude a direct action of 1,25(OH)₂D₃ on bone cells as the mechanism of osteoid formation, they do underline the importance of the humoral changes at least as partial determinants of this phenomenon. The activities of various vitamin D metabolites were compared using as parameter the threshold-dose required to produce a rim of unmineralized trabeculae in the tibia of 14–15 days embryos (T-D). The most active metabolite appeared to be 1,25(OH)₂D₃ (T-D: 10 ng); it was followed by 1,24,25(OH)₃D₃ (T-D: 100 ng) and 1,25,26(OH)₃D₃ (T-D: 100 ng). Vitamin D₃ itself (T-D: 100 µg), 25(OH)D₃ (T-D: 2.5µg) and 24,25(OH)₂D₃ (T-D: 5 µg) produced similar responses but only when administered in much larger doses.