The involvement of intracellular calcium ion concentration and calmodulin in the 25-hydroxylation of cholecalciferol in ovine and rat liver.
The effect of Ca2+ ion concentration on the 25 hydroxylation of tritiated cholecalciferol (3HD3) was investigated using homogenates of ovine liver from vitamin D replete sheep. A significant decrease in the production of 25 hydroxycholecalciferol (25OHD3) was observed when the concentration of Ca2+ in the homogenate was raised above 0.68 mmol/l by the addition of calcium gluconate. Similarly, a final concentration of 37 mumol EGTA/1 (equivalent to a Ca2+ concentration of 26.5 nmol/l) was associated with a 50% reduction of 25OHD3 production. That is, a broad bell-shaped relationship was observed between the production of 25OHD3 and the Ca2+ concentration in the homogenate. These changes in the rate of production of 25OHD3 were reproduced with hepatocytes from vitamin D replete rats, prepared by collagenase perfusion, using the drugs dantrolene sodium (DaNa) to reduce (ED50 = 57 mmol/l) and veratridine to increase (ED50 = 550 mmol/l) the intracellular Ca2+ concentration. Hepatocytes from vitamin D replete rats also showed a reduction in 25 hydroxylation of D3 (ED50 = 6 ng/ml) in response to the addition of 1-25 dihydroxycholecalciferol (1-25 (OH)2D3). The calmodulin antagonists; W7, compound 48/80, trifluoperazine (TFP) and calmidazolium (R24571) were all found to effect a dose response inhibition of the 25 hydroxylation of cholecalciferol by homogenates of ovine liver. R24571 had a similar inhibitory effect (ED50 = 70 mumol/l) upon the 25 hydroxylase enzyme of rat hepatocytes. It is concluded that the 25 hydroxylation of cholecalciferol in liver of vitamin D replete rats and sheep is calcium sensitive and is reduced in the presence of increased concentrations of 1,25(OH)2D3. Calmodulin may also be involved in the regulation of hepatocyte 25-hydroxylase activity by Ca2+.
Corlett
S C
SC
Department of Animal Physiology & Nutrition, University of Leeds, U.K.
Chaudhary
M S
MS
Tomlinson
S
S
Care
A D
AD
D000818:Animals; D002112:Calcifediol / Q000096:biosynthesis; D002118:Calcium / Q000502:physiology*; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000502:physiology*; D002413:Cations, Divalent; D002762:Cholecalciferol / Q000378:metabolism*; D053493:Cholestanetriol 26-Monooxygenase; D003620:Dantrolene / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D005260:Female; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D012756:Sheep; D013250:Steroid Hydroxylases / Q000378:metabolism*
D002147:Calmodulin; D002413:Cations, Divalent; D002762:Cholecalciferol; D004533:Egtazic Acid; D013250:Steroid Hydroxylases; D053493:Cholestanetriol 26-Monooxygenase; D003620:Dantrolene; D002112:Calcifediol; D002118:Calcium