In Vitro Metabolism of 25-Hydroxycholecalciferol by Chick Intestinal and Renal Cell Preparations

Abstract

Abstract Free cell suspensions of intestinal epithelial and renal tubule cells were prepared by enzymatic digestion of tissue from rachitic chickens. The cells were demonstrated to remain metabolically functional during prolonged incubation at 37°. Both preparations were demonstrated to be capable of the in vitro metabolism of 25-hydroxycholecalciferol to products chromatographically more polar than 25-hydroxycholecalciferol. One product of the in vitro renal tubule metabolism of 25-hydroxycholecalciferol was shown to be identical to 1,25-dihydroxycholecalciferol by Sephadex LH-20 column chromatography, countercurrent distribution in two different solvent systems, thin layer chromatography, and by its insensitivity to periodic acid oxidation. Additionally it was demonstrated that 25,26-dihydroxycholecalciferol is not and that 21,25-dihydroxycholecalciferol, if produced at all, is a very minor product of the in vitro renal tubule metabolism of cholecalciferol. 1,25-Dihydroxycholecalciferol was not a product of the in vitro intestinal epithelial cell metabolism of 25-hydroxycholecalciferol. More importantly, 1,25-dihydroxycholecalciferol also was not significantly metabolized in vitro by the epithelial cell preparation. This observation provides a unique demonstration that 1,25-dihydroxycholecalciferol and not some further metabolite of 1,25-dihydroxycholecalciferol is the cholecalciferol metabolite responsible for the initiation and maintenance of intestinal calcium absorption.