Orally ingested (13)C(2)-retinol is incorporated into hepatic retinyl esters in a nonhuman primate (Macaca mulatta) model of hypervitaminosis A - PubMed (original) (raw)

Orally ingested (13)C(2)-retinol is incorporated into hepatic retinyl esters in a nonhuman primate (Macaca mulatta) model of hypervitaminosis A

Anne L Escaron et al. Comp Med. 2010 Feb.

Abstract

The mechanism responsible for the metabolism of vitamin A during hypervitaminosis is largely unknown. This study investigated hepatic (13)C-retinol uptake in hypervitaminotic A rhesus monkeys. We hypothesized that individual retinyl esters would be enriched in (13)C after a physiologic dose of (13)C(2)-retinyl acetate, thus suggesting de novo in vivo hepatic retinol esterification. Male rhesus macaques (n = 16; 11.8 +/- 2.9 y) each received 3.5 micromol 14, 15-(13)C(2)-retinyl acetate. Blood was drawn at baseline and 5 h and 2, 4, 7, 14, 21, and 28 d after administration. Liver biopsies were collected 7 d before and 2 d after dose administration (n = 4) and at 7, 14, and 28 d after dose administration (n = 4 per time point). (13)C enrichments of retinol and retinyl esters HPLC-purified from liver samples were measured by using gas chromatography-combustion-isotope ratio mass spectrometry. (13)C enrichment of total vitamin A and individual retinyl esters were significantly greater 2 d after dose administration compared with baseline levels. In contrast, the concentration of isolated retinyl esters did not always increase 2 d after treatment. Given that the liver biopsy site differed between monkeys, these data suggest that the accumulation of hepatic retinyl esters is a dynamic process that is better represented by combining analytical techniques. This sensitive methodology can be used to characterize vitamin A trafficking after physiologic doses of (13)C-retinol. In this nonhuman primate model of hypervitaminosis A, hepatic retinyl esters continued to accumulate with high liver stores.

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Figures

Figure 1.

Figure 1.

(A) Mass chromatogram for labeled 13C2-retinol standard (50 ng/µL; 3% 13C2-retinol). (B) Mass chromatogram of retinol from saponified retinyl oleate and palmitate isolated from baseline rhesus monkey liver, indicating that the retinol saponified from hepatic retinyl esters at baseline has the same retention time as the labeled retinol standard. For both panels A and B, the top image indicates the ratio of CO2 molecules, whereas the bottom image indicates the signal strength (intensity of the peaks in volts). The dotted line in the top images represents the mass 45/44 trace—the variation of mass 45 (predominantly 13CO2) to mass 44 (12CO2). The solid line in the top panels represent the mass 46/44 trace —the variation of mass 46 (predominantly 12C18O16O) to mass 44 (12CO2). The first 3 peaks in both the top and bottom panel are the reference gas (CO2) pulses.

Figure 2.

Figure 2.

Semilog plot of serum retinol enrichment over time in captive rhesus monkeys given an oral dose of 3.5 µmol 13C2-retinyl acetate. Serum retinol enrichment is shown as serum atom percentage excess (adjusted for baseline). Repeated measures using Tukey adjustment for multiple comparisons demonstrate that serum retinol enrichments at 5 h (*) and 2 d (#) after dose administration differ significantly (P < 0.05) from those at all other times. Values are presented as mean ± 1 SD (n = 16 per time point, except for 5 h and 28 d, for which n = 15).

Figure 3.

Figure 3.

Pre- and postdose At%13C for hepatic retinyl esters that were HPLC-purified and saponified to retinol: total retinyl esters (TOT), retinyl linoleate (RL), retinyl oleate and palmitate (RO + RP), and retinyl stearate (RS). Baseline (white bars) and postdose (black bars) values are given as mean ± 1 SD; values for baseline and day 2 after dose administration reflect the same 4 monkeys at each point. *, 1-way ANOVA by day revealed significant difference relative to baseline value (TOT, P = 0.0065; RL, P = 0.0003; RO + RP, P = 0.0022; and RS, P = 0.0027).

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