title: Constitutive expression of a vitamin D 1-hydroxylase in a myelomonocytic cell line: a model for studying 1,25-dihydroxyvitamin D production in vitro.
abstract: The capacity of the v-myc-transformed, chicken myelomonocytic cell line HD-11 to metabolize 25-hydroxyvitamin D3 (25-OHD3) was examined. HD-11 cells produced and secreted a metabolite of 25-OHD3 that was bound with high affinity by receptor for 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. On normal-phase HPLC, this metabolite cochromatographed with authentic 1,25-(OH)2D3 in both hexane- and methylene chloride-based solvent systems. The 25-OHD3 1-hydroxylation reaction was substrate saturable with a Km of 73 nM 25-OHD3 and a maximal velocity of 167 fmol per 10(6) cells per h. This reaction was inhibited by ketoconazole, a recognized inhibitor of cytochrome P450 mixed-function oxidases including the authentic, renal 25-OHD3 1-hydroxylase. On the other hand, HD-11 cell 1,25-(OH)2D3 production was not affected by the antioxidant DPPD, a known inhibitor of free radical-generated 1,25-(OH)2D3. In addition to synthesizing 1,25-(OH)2D3, this monocyte-macrophage cell line also has the potential to be a target for the hormone; HD-11 cells express high-affinity receptor for 1,25-(OH)2D3 (Kin = 0.06 nM).
authors.affiliation: Division of Endocrinology and Metabolism, Cedars-Sinai Medical Center-UCLA School of Medicine 90048.
authors.forename: J S
authors.initials: JS
authors.lastname: Adams
authors.forename: T G
authors.initials: TG
authors.lastname: Beeker
authors.forename: T
authors.initials: T
authors.lastname: Hongo
authors.forename: T L
authors.initials: TL
authors.lastname: Clemens
mesh_terms: D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000378:metabolism*; D000818:Animals; D002117:Calcitriol / Q000096:biosynthesis*; D002461:Cell Line, Transformed; D002645:Chickens; D016166:Free Radical Scavengers; D005609:Free Radicals; D007654:Ketoconazole / Q000494:pharmacology; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism*; D008954:Models, Biological; D009000:Monocytes / Q000378:metabolism*; D010655:Phenylenediamines / Q000494:pharmacology; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid / Q000378:metabolism
chemical_list: D016166:Free Radical Scavengers; D005609:Free Radicals; D010655:Phenylenediamines; D018167:Receptors, Calcitriol; D011987:Receptors, Steroid; C004845:N,N'-diphenyl-4-phenylenediamine; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D002117:Calcitriol; D007654:Ketoconazole