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Retinol circulates in rat plasma bound to a specific protein, retinol-binding protein (RBP). A radioimmunoassay for rat RBP was developed with a double antibody precipitation technique. The immunoassay accurately measures RBP in amounts of 0.5 to 3 ng per assay tube. There was no significant difference in the immunoreactivity of apo-RBP as compared to holo-RBP. Using this assay, a study was conducted to examine the effects of vitamin A depletion and deficiency, and of repletion, on the level of serum RBP, in order to explore the role of nutritional vitamin A status in the regulation of RBP metabolism. Weanling rats were divided into five groups, each fed a vitamin A-deficient diet with or without supplements as follows: Group 1, control: supplemented with vitamin A; Group 2, pair-fed control: supplemented with vitamin A but with food intake matched to that of the deficient rats; Group 3, deficient: no vitamin A supplementation; Groups 4 and 5, retinoic acid: supplemented, after the initial depletion period, with a modest daily dose (14 or 28 µg) of retinoic acid. The rats were studied for 75 days. In the deficient group the serum vitamin A levels decreased gradually during the first 25 to 30 days of the study, to levels of about 2 µg/100 ml. Serum RBP levels also declined during the induction of vitamin A deficiency, with a time course similar to that seen for vitamin A, but with a lag of about 3 days (from 50 ± 4 µg per ml on Day 3 to 20 ± 2 µg per ml at Day 27, and then more slowly to 13 ± 2 µg per ml at Day 75). After 3 to 4 weeks most of the circulating RBP was present as the apoprotein, not containing a molecule of bound retinol. The results obtained with the retinoic acid-fed rats, who continued to grow normally, were identical with those of the deficient rats. In contrast both the pair-fed and ad libitum control groups exhibited no major changes in either serum vitamin A or RBP, throughout the entire study.
Liver homogenates were immunoreactive, and generated immunoassay curves which were indistinguishable from those obtained with pure rat RBP. The level of immunoreactive RBP in the livers of deficient rats was 4 times (p < 0.001) that in the livers of control rats. When vitamin A was administered orally to deficient rats on Day 53, a very rapid increase in serum RBP level, from a mean of 14 to 56 µg per ml, was seen within 5 hours (the first time interval sampled). These findings suggest that vitamin A deficiency primarily interferes in some way with the secretion, rather than with the synthesis, of RBP by the liver, and that the deficient liver contains a pool of previously formed apo-RBP which can be released rapidly into the serum, as holo-RBP, when vitamin A becomes available.
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Published online: April 25, 1972
Received:
March 24,
1977
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© 1972 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology.
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