Avian medullary bone in organ culture: effects of vitamin D metabolites on collagen synthesis.

A new organ culture system for the study of bone metabolism has been developed using chicken medullary bone. The presence of viable bone cells in culture was demonstrated by histological and histochemical techniques. Incorporation of 3H-proline into collagenase-digestible protein (CDP) and noncollagen protein (NCP) was determined using purified bacterial collagenase. Collagen accounted for approximately 10-15% of the total protein labeled. The addition of 1,25-dihydroxycholecalciferol (1,25(OH)2D3) resulted in a dose-dependent inhibition of 3H-proline incorporation into CDP at doses from 10(-10)M to 10(-7)M, with maximal suppression reaching 30% of control. The effect was specific for collagen, since 3H-proline incorporation into NCP was unaffected. Hydroxyproline analysis of bone explants and culture medium revealed a 1,25(OH)2D3-induced decrease in the 3H-hydroxyproline content of the system (bone + medium), suggesting that the effect of 1,25(OH)2D3 is due to inhibition of collagen synthesis rather than enhanced collagen degradation, impaired incorporation of collagen into bone matrix, or bone resorption. Medullary bone collagen synthesis was not affected by 24,25(OH)2D3, either alone or in combination with 1,25(OH)2D3. Structure-activity studies of vitamin D metabolites showed that 1,25(OH)2D3 and 1,24,25(OH)3D3 were the most potent metabolites tested, followed by 1-alpha(OH)D3. 25(OH)D3 and 24,25(OH)2D3 had no effect at concentrations as high as 10(-7)M. These results indicate a possible role for vitamin D in the regulation of medullary bone formation during the reproductive cycle of the egg-laying hen, and suggest the potential utility of medullary bone as an in vitro model for the study of bone formation.