Studies in Mice Lacking the aTocopherol Transfer Protein

Although studies in humans with defects in the TTP gene have suggested that TTP was required for maintenance of plasma a-tocopherol concentrations (17), only limited information is available on these vitamin E-deficient subjects' tissue a-tocopherol concentrations (18). Mice in which the gene for TTP was deleted have extraordinarily low vitamin E plasma and tissue concentrations (19-21) and express vitamin E deficiency symptoms (7,19). To determine whether TTP knockout mice were unable to discriminate between natural and synthetic vitamin E (21), adult TTP knockout (Ttpa-~, n = 5), heterozygous (Ttpa+/~, n = 7), and control (Ttpa+I+, n = 3) mice consumed equimolar d6 RRR- and d^ all-rac-a-tocopheryl acetates (30 mg each/kg diet) for 3 mo; labeled and unlabeled a-tocopherols in plasma and 17 tissues were then measured by liquid chromatography/mass spectrometry (LC/MS) (21). Although deuteriumlabeled a-tocopherols represented >85% of the plasma a-tocopherol in all groups, Ttpa-- mice had plasma total a-tocopherol concentrations that were only 5.4% of Ttpa+/+ and 7.7% of Ttpa+mice. Ttpa-- tissue (except liver) total a-tocopherol concentrations were 2-20% of those in Ttpa+/+ mice. These data are consistent with the concept that vitamin E is absorbed and transported to the liver, and only if TTP is expressed is a-tocopherol exported from the liver into the plasma for tissue delivery. Clearly, TTP is required not only to maintain plasma a-tocopherol, but also tissue a-tocopherol.

The other proposed function of TTP is the preferential secretion of RRR-a-tocopherol from the liver into the plasma. In mice fed 1:1 d6 RRR- and d^ all-rac-a-tocopheryl acetates for 3 mo, the d6:d3 ratios in plasma and 16 tissues from

Ttpa+/+ and Ttpa+/- mice were double those of Ttpa-/- mice (Fig. 11.1) (21). In TTP-expressing mice, tissue enrichment of natural over synthetic a-tocopherol appears to be due to nonspecific uptake of a-tocopherol from the plasma, which contained 2:1 d6:d3 a-tocopherols. Ttpa+/- mice that expressed half the amount of hepatic TTP (20) also had d6^ ratios of nearly 2. These data suggest that plasma a-tocopherol concentrations are highly dependent upon the function of TTP and that this protein preferentially selects only the 2R-a-tocopherol forms from all-rac-a-tocopherol for secretion into plasma. Importantly, patients who were a-TTP heterozygotes (expressing two different mutations) were also able to discriminate between RRR- and SRR-a-tocopherols (17,22). These 2:1 ratios are also consistent with findings in normal humans administered labeled RRR- and all-rac-a-toco-pheryl acetates (23-25).

Fig. 11.1. a-Tocopherol transfer protein (TTP)-knockout mice do not discriminate between natural and synthetic vitamin E. Shown are the d6:d3 a-tocopherol ratios in adult a-TTP knockout (Ttpa~f~, n = 5), heterozygous (Ttpa+/~, n = 7), and wild-type (Ttpa+/+, n = 3) mice that consumed a 1:1 d6-RRR- and d3-all-rac-a-tocopheryl acetate-containing diet (30 mg each/kg diet) for 3 mo. Labeled and unlabeled a-toco-pherols in plasma and 17 tissues were measured. The d6:d3 a-tocopherol ratios in plasma and 16 tissues (excluding liver) from Ttpa+/+, Ttpa+/- and Ttpa-/- mice were 1.8 ± 0.2, 1.9 ± 0.2, and 1.1 ± 0.1, respectively (P < 0.0001, Ttpa-/- vs. Ttpa+U or Ttpa+/-). Source: Adapted from (21).

Fig. 11.1. a-Tocopherol transfer protein (TTP)-knockout mice do not discriminate between natural and synthetic vitamin E. Shown are the d6:d3 a-tocopherol ratios in adult a-TTP knockout (Ttpa~f~, n = 5), heterozygous (Ttpa+/~, n = 7), and wild-type (Ttpa+/+, n = 3) mice that consumed a 1:1 d6-RRR- and d3-all-rac-a-tocopheryl acetate-containing diet (30 mg each/kg diet) for 3 mo. Labeled and unlabeled a-toco-pherols in plasma and 17 tissues were measured. The d6:d3 a-tocopherol ratios in plasma and 16 tissues (excluding liver) from Ttpa+/+, Ttpa+/- and Ttpa-/- mice were 1.8 ± 0.2, 1.9 ± 0.2, and 1.1 ± 0.1, respectively (P < 0.0001, Ttpa-/- vs. Ttpa+U or Ttpa+/-). Source: Adapted from (21).

Vitamin E Kinetics in Pigs

Vitamin E supplements are often provided to farm animals; therefore, the relative bioavailabilities of RRR- and all-rac-a-tocopherols were evaluated in swine (26). Deuterium-labeled vitamin E (150 mg each of d3-RRR- and d6-all rac-a-tocopheryl acetates) was administered orally to female pigs (n = 3) with the morning feed. Blood samples were obtained at 0, 3, 6, 9, 12, 24, 36, 48, and 72 h. The maximum observed plasma concentration of d3-RRR-a-tocopherol was achieved at 12 h (1.12 ^mol/L), whereas d6-a-tocopherol peaked earlier (at 9 h) at a lower concentration (0.66 ^mol/L, P < 0.05, Fig. 11.2). The d3/d6 ratios were 1.35 ± 0.73 at 3 h and increased to 2.00 ± 0.14 at 72 h (P < 0.03). The d3-a- and d6-a-tocopherol disappearance rates were similar for both tocopherols and estimated to be 0.029 ^mol/L per hour. The AUC for d3-a- and d6-a-tocopherol were 38.2 and 18.5, respectively. These studies suggest that pigs rapidly discriminate between forms of vitamin E and selectively retain 2R-forms in the plasma.

Fig. 11.2. Discrimination between natural and synthetic vitamin E in swine. The plasma d3- and d6-a-tocopherol concentrations after deuterium-labeled vitamin E (150 mg each d3-RRR- and d6-all-rac-a-tocopheryl acetates) was administered orally to adult female pigs (n = 3) with the morning feed are shown. The ratio of d3-/d6-a-tocopherols increased from 1.35 ± 0.73 at h 3 to 2.0 ± 0.14 at h 72 (P< 0.03). Source: Adapted from (26).

Fig. 11.2. Discrimination between natural and synthetic vitamin E in swine. The plasma d3- and d6-a-tocopherol concentrations after deuterium-labeled vitamin E (150 mg each d3-RRR- and d6-all-rac-a-tocopheryl acetates) was administered orally to adult female pigs (n = 3) with the morning feed are shown. The ratio of d3-/d6-a-tocopherols increased from 1.35 ± 0.73 at h 3 to 2.0 ± 0.14 at h 72 (P< 0.03). Source: Adapted from (26).

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