BONE RESORPTION IN TISSUE CULTURE 631 



In view of the controversy which still exists concerning a direct 

 action of vitamin D on the skeleton, it was deemed worth while to 

 test the effect of this vitamin in our tissue culture system (50 per 

 cent O2). Dihydrotachysterol, a member of the vitamin D group 

 which is closely related to calciferol and has calcemic activity in 

 vivo, caused a clear-cut enhancement of resorption when added to 

 our system. The extent of bone resorption was directlv related to 

 the concentration of dihydrotachysterol in the medium. The micro- 

 scopic appearance of living cultures exposed to 0.001 mg, 0.01 mg, 

 and 0.1 mg of dihydrotachysterol per ml of medium for 1 week may 

 be seen in Figs. 32 to 35. Preliminarv experiments with crystalline 

 vitamin Di- or Da in our system ( 50 per cent O- ) have demonstrated 

 enhancement of bone resorption when the vitamins were added at 

 the level of 0.1 mg (4000 units) per ml to the medium. The extent 

 of resorption, which was similar to that found with 0.1 mg per ml 

 of dihydrotachysterol, was not so marked as that obtained with 0.5 

 unit of parathyroid extract per ml of medium. Further experiments 

 are continuing with the D vitamins to determine optimum dosages 

 and the influence of oxygen tension on their ability to induce bone 

 resorption. 



Discussion 



Several points about these findings seem worth emphasizing and 

 discussing briefly. 



It should be stressed that variation in culturing techniques or 

 media can make a vast difference in the response obtained to a given 



Fig. 30. Gross appearance of 12-day roller-tube cultures containing 24 units 

 vitamin A per ml and exposed to 10 per cent and 20 per cent Oo. Note the 

 enhanced resorption induced by the presence of vitamin A as compared with 

 controls, as well as the influence of 20 per cent O2 as compared with 10 per 

 cent Oo. 



Fig. 31. Gross appearance of 12-day roller-tube cultures containing 24 units 

 vitamin A per ml and exposed to 30 per cent and 50 per cent Oo. Note that 

 the best resorption in the experiment occurs with the combination of 24 units 

 of vitamin A per ml and 30 per cent Oo. Although the vitamin A cultures gassed 

 with 50 per cent Oo show less of an effect at this stage, it should be noted that 

 this group demonstrated the best resorption microscopically during the first 6 

 days. 



