Abstract
The vitamin D metabolite, 24,25(OH) 2 D 3 has been reported to have hormonal activity. Catalase has been reported to be a binding protein for 24,25(OH) 2 D 3 , based on sequence analysis of the protein isolated on the basis of specific binding of the metabolite. In the current work, we report that 24R,25(OH) 2 D 3 , not 24S,25(OH)2D3 is the effective metabolite for catalase redistribution as judged by confocal microscopy. We have used male chick intestinal cells treated with either vehicle, 24S,25(OH) 2 D 3 , 24R,25(OH) 2 D 3 or 1,25(OH) 2 D 3 to determine the localization of catalase. Confocal microscopy analyses showed punctate staining, on the cell surface and in the cytoplasm of cells treated with vehicle, 24R,25(OH) 2 D 3 , 24S,25(OH) 2 D 3 or 1,25(OH) 2 D 3 for all time points tested. Cells treated with 24R,25(OH) 2 D 3 showed punctuate staining of catalase inside the nucleus. Western analysis confirmed that the punctuate staining in the nucleus arose from the redistribution of cell surface catalase. Western analysis also indicated 24S,25(OH) 2 D 3 treatment resulted in redistribution of catalase to the nucleus, but to a lesser extent than treatment with 24R,25(OH)2D3. By understanding the molecular and cellular actions of 24,25(OH)2D3 in chick intestine, progress will be made in enhancing phosphate and calcium absorption in animals to supply the minerals for adequate bone growth, and phosphate in manure of production animals could be diminished.
Highlights
Vitamin D was discovered in 1922 and has been categorized as a pre-hormone ever since, based on the fact that the utilization of most vitamin D in higher animals undergoes a photochemical process
Activation of vitamin D starts in the liver with the production of major circulating metabolite 25-hydroxyvitamin D3 [25(OH)D3], followed by hydroxylation in the kidneys to yield two metabolites, either 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] made when phosphate and calcium levels are low or 24,25-dihydroxyvitamin D3 [24,25(OH)2D3] made when phosphate and calcium levels are high
While 1,25-dihydroxyvitamin D3 stimulates the rapid transport of calcium and phosphate in both perfused chick duodenal loops and isolated enterocytes, 24,25-dihydroxyvitamin D3 inhibits such stimulation [3,4]
Summary
Vitamin D was discovered in 1922 and has been categorized as a pre-hormone ever since, based on the fact that the utilization of most vitamin D in higher animals undergoes a photochemical process. Chick intestinal cells were cultured in petri dishes, treated with 0.01% ethanol, 6.5 nM 24R,25(OH)2D3, 200 pM 24S,25(OH)2D3, or 300 pM 1,25(OH)2D3, concentrations that have been shown to be equivalent to circulating levels [19], for selected times (15 sec, 30 sec, 7 min, 10min, 15 min, 25 min, 30 min, 40 min, 50 min, 60 min), and fixed for confocal microscopy.
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