Synthesis of 25-hydroxyvitamin D3-26,23-lactone but not 24,25-dihydroxyvitamin D3 from 25-hydroxyvitamin D3 in opossum kidney cells treated with 1 alpha, 25-dihydroxyvitamin D3.

The pathway from 25-hydroxyvitamin D3(25[OH]D3) to 25-hydroxyvitamin D3-26,23-lactone (25[OH]D3-26,23-lactone) is important in the renal degradation of vitamin D metabolites. Our objective was to investigate the regulation of 25(OH)D3 metabolism in the opossum kidney (OK) cell line which has been used for studying the actions of the calcium-regulating hormones. While the untreated OK cells did not produce 25(OH)D3 metabolites, the synthesis of the lactone was increased dose-dependently by 20h pretreatment with 1 alpha,25-dihydroxyvitamin D3 (1,25[OH]2D3), peaked at 10(-7) M 1,25(OH)2D3, and decreased with higher concentrations. Little production of 24,25-dihydroxyvitamin D3 (24,25[OH]2D3) was observed after any dose of 1,25(OH)2D3. The increase in the production of lactone induced by 10(-7) M 1,25(OH)2D3 was detectable at 2 h, peaked between 4 and 24 h, and decreased gradually until 48 h. Its induction was inhibited by actinomycin D and cycloheximide, suggesting that de novo protein synthesis is required for such induction. After treatment with 10(-7) M 1,25(OH)2D3 for 20 h, 24-hydroxylase (24-OHase) mRNA was not detected in OK cells, but was found evident in COS-7 cells (a renal cell line). Moreover, the COS-7 cells treated with 10(-7) M 1,25(OH)2D3 metabolized 25(OH)D3 to 24,25(OH)2D3. These results indicate that 1,25(OH)2D3 stimulates the synthesis of 25(OH)D3-26,23-lactone in a dose- and time-dependent manner, but does not induce the gene expression or activity of 24-OHase in OK cells.