A role for nitric oxide in the regulated expression of the 25-hydroxy-vitamin D-1-hydroxylation reaction in the chick myelomonocytic cell line HD-11.
We have recently described the existence of a cytochrome P450-associated, mitochondrial-based 25-hydroxyvitamin D (25-OHD)-1-hydroxylation reaction in the chick macrophage-like cell line HD-11. Considering that this reaction is regulated by the same set of factors (ie. interferon-gamma, lipopolysaccharide, and glucocorticoids) that modulate expression of the macrophage nitric oxide synthase (mac NOS), we investigated the possibility that endogenous nitric oxide (NO) production may be linked to 1,25-dihydroxyvitamin D3 (1,25-(OH)2D) synthesis by HD-11 cells in vitro. To test this hypothesis we investigated the effects excluding from the extracellular medium the essential amino acid L-arginine, substrate for endogenous NO production, on the basal and stimulated expression of the HD-11 cell 25-OHD-1-hydroxylation reaction. Depletion of L-arginine from the extracellular medium for as little as 6 h resulted in a significant decrease (p < 0.02) in basal 1,25-(OH)2D synthesis; after 15 h in an L-arginine-free environment hormone production was reduced to < 10% of basal levels without any adverse affect on cell viability. Reintroduction of L-arginine, but not D-arginine, into the extracellular medium restored 1,25-(OH)2D3 synthetic capacity fully if done after < or = 6 h of incubation in the absence of L-arginine. Competitive inhibition of NOS with Nw-nitro-L-arginine methyl ester (p < 0.002) and Nw-nitro-L-arginine (p < 0.02) significantly inhibited 1,25-(OH)2D synthesis, indicating that macrophage NO generating capacity is functionally linked to endogenous synthesis of the active vitamin D metabolite.
Adams
J S
JS
Division of Endocrinology, Cedars-Sinai Medical Center, UCLA School of Medicine 90048.
Ren
S Y
SY
Arbelle
J E
JE
Clemens
T L
TL
Shany
S
S
D000818:Animals; D000831:Animals, Newborn; D001120:Arginine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D002117:Calcitriol / Q000378:metabolism; D002460:Cell Line; D002645:Chickens; D016895:Culture Media, Serum-Free; D005110:Extracellular Space / Q000378:metabolism; D006900:Hydroxylation; D008264:Macrophages / Q000378:metabolism*; D009569:Nitric Oxide / Q000037:antagonists & inhibitors / Q000502:physiology*
D016895:Culture Media, Serum-Free; D009569:Nitric Oxide; D001120:Arginine; D002117:Calcitriol