Simulated acidosis does not impair 1,25-dihydroxyvitamin D₃ production by cultured kidney cells

Summary

Cultured mouse kidney cells grown in serum-free medium were used to assess the metabolism of 25-hydroxyvitamin D₃ in the presence of simulated metabolic acidosis. Kidney epithelial cells isolated from 4–6 week old mice were grown to confluence in a defined serum-free medium at pH 7.4. The confluent monolayers were incubated with tritiated 25-hydroxyvitamin D₃ for 6 hours, the samples were extracted, and vitamin D metabolites were separated and quantitated by high pressure liquid chromatography (HPLC). The pH of the incubation medium was set at 6.9, 7.1, 7.4, or 7.7 by adjusting the bicarbonate concentration, using chloride as the balancing anion at constant Pco₂. When pH was altered at the beginning of the 6 hour assay, production of 1,25-dihydroxyvitamin D₃ was the same at each pH. More prolonged pH perturbation for a total of 30 hours likewise had no influence on 1,25-dihydroxyvitamin D₃ production. These results confirm that intact mammalian kidney cells in serum-free culture possess an active 25-hydroxyvitamin D₃-1-hydroxylase and that the activity of the enzyme is unaffected by pH over the range 6.8–7.7. In experiments where acidosis has been shown to alter 1,25-dihydroxyvitamin D₃ production, the mechanism was probably indirect.