Role of 25-hydroxyvitamin D3 dose in determining rat 1,25-dihydroxyvitamin D3 production.
To understand the relationships among 1) the dose of 25-hydroxyvitamin D [25(OH)D] in vivo, 2) the activity of 1-hydroxylase in renal mitochondria, and 3) the production of 1,25-dihydroxyvitamin D [1,25(OH)2D] in vivo, we gave rats different chronic or acute doses of 25-hydroxyvitamin D3 [25(OH)D3]. We followed the metabolism of intracardially administered [25-hydroxy-26,27-methyl-3H]cholecalciferol [25(OH)[3H]D3] for 24 h before killing by measuring extracts of serum by chromatography. Specific activity of 1-hydroxylase in kidney was measured at death. In rats given 0-2,000 pmol 25(OH)D3 chronically by mouth, there was a dose-dependent decline in the percent of serum radioactivity made up of 1,25-dihydroxy-[26,27-methyl-3H]cholecalciferol [1,25(OH)2[3H]D3] as well as a decline in mitochondrial 1-hydroxylase, and these correlated significantly (r = 0.83, P less than 0.001). Serum %1,25(OH)2[3H]D3 in this experiment ranged from 0.8 to 42%. A small part of this range could be accounted for by a faster metabolic clearance rate (MCR) of 1,25(OH)2D3 from rats supplemented with 25(OH)D3 (MCR, 2.12 +/- 0.10 ml/min) compared with rats restricted in vitamin D (MCR, 0.94 +/- 0.06 ml/min, P less than 0.001). The activity of 1-hydroxylase was by far the major factor determining serum %1,25(OH)2[3H]D3. When different acute doses of 25(OH)D3 were given to rats with identical specific activities of 1-hydroxylase, the resulting 1,25(OH)2D3 concentrations in serum correlated with the 25(OH)D3 dose (r = 0.99, P less than 0.001). We conclude that the behavior of 1-hydroxylase in vivo is analogous to the classic behavior in vitro of an enzyme functioning below its Michaelis constant (Km). The amount of 1-hydroxylase present in renal mitochondria determines the fraction (not simply the quantity) of 25(OH)D metabolized to 1,25(OH)2D3 in vivo.
Department of Clinical Biochemistry, University of Toronto, Ontario, Canada.
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D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000378:metabolism; D000818:Animals; D001711:Biotransformation; D002112:Calcifediol / Q000097:blood / Q000378:metabolism*; D002117:Calcitriol / Q000096:biosynthesis / Q000097:blood / Q000378:metabolism*; D007668:Kidney / Q000201:enzymology; D008297:Male; D008433:Mathematics; D008657:Metabolic Clearance Rate; D008928:Mitochondria / Q000201:enzymology; D008954:Models, Biological; D011865:Radioisotope Dilution Technique; D051381:Rats; D011919:Rats, Inbred Strains; D014316:Tritium
D014316:Tritium; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D002117:Calcitriol; D002112:Calcifediol