Comparison of α-Tocopherol, α-Tocopherol Acetate, and α-Tocopheryl Polyethylene Glycol Succinate 1000 Absorption by Caco-2 TC7 Intestinal Cells

Abstract

(1) Background: vitamin E is often supplemented in the form of tocopherol acetate, but it has poor bioavailability and can fail to correct blood tocopherol concentrations in some patients with severe cholestasis. In this context, α-tocopheryl polyethylene glycol succinate 1000 (TPGS) has been of value, but very little is known about the mechanisms of its absorption. The aim of our work was to evaluate the mechanisms of absorption/secretion of TPGS compared to tocopherol acetate (TAC) and α-tocopherol by human enterocyte-like Caco-2 TC7 cells. (2) Methods: two weeks post-confluence Caco-2 cells were incubated with tocopherol- or TAC- or TPGS-rich mixed micelles up to 24 h and, following lipid extraction, TAC and tocopherol amounts were measured by high performance liquid chromatography (HPLC) in apical, cellular, and basolateral compartments. (3) Results: at equivalent concentrations of tocopherol in the apical side, the amounts of tocopherol secreted at the basolateral pole of Caco-2 cells are (i) significantly greater when the tocopherol is in the free form in the micelles; (ii) intermediate when it is in the TAC form in the micelles (p < 0.001); and (iii) significantly lower with the TPGS form (p < 0.0001). Interestingly, our results show, for the first time, that Caco-2 cells secrete one or more esterified forms of the vitamin contained in TPGS at the basolateral side.

Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias

“…The fact that we observed a lower α-tocopherol secretion in MTTP -KO clones than in SAR1B -KO clones is consistent with clinical observations, as FHBL-SD1 patients present more severe vitamin E deficiency than FHBL-SD3 patients [ 40 ]. Our results are also consistent with a previous cell-based study designed to characterize the absorption of free tocopherol, TAC and TPGS [ 13 ]. This previous study also reported a basolateral secretion of TAC in its intact form (202 pmol/well after 24 h of incubation).…”

“…We then characterized the secretion of α-tocopherol, TAC and TPGS by wild-type Caco-2 cells compared to KO clones. Alpha-tocopherol is one of the main dietary forms of vitamin E. In clinical nutrition, vitamin E deficiency is usually treated with oral α-tocopherol-acetate (TAC) to protect the phenol group against oxidation [ 13 , 39 ]. Due to the poor bioavailability of TAC and because of the interest in treating patients with Monogenic Hypobetalipoproteinemia Disorders with TPGS, a drinkable form of tocopherol, we characterized and compared the uptake and secretion of α-tocopherol, TAC and TPGS in the different clones.…”

“…The Caco-2/TC7 cell line, derived from a human colon adenocarcinoma, naturally differentiates into a cell monolayer expressing several morphological and functional characteristics of enterocytes [ 15 ]. Caco-2/TC7 cells were cultured on 25 cm 2 flasks in DMEM supplemented with 16% FBS, 1% of non-essential amino acids and 1% penicillin/streptomycin [ 13 ]. The percentage of FBS in the complete medium was then reduced from 16% (usual percentage) to 8% to ensure a correct growth of the KO clones.…”

“…A volume of 50–200 µL was used for HPLC analysis. TPGS was indirectly assayed by quantifying α-tocopherol released after saponification by potassium hydroxide (KOH 5.5% + pyrogallol 1%) for 10 min at room temperature and by subtracting α-tocopherol determined before saponification as previously described [ 13 ].…”

“…The mechanisms hampering vitamin E plasma level restoration in treated FHBL-SD1 and FHBL-SD3 patients are still unknown. We previously studied the mechanisms of absorption/secretion of α-tocopheryl polyethylene glycol succinate 1000 (TPGS, i.e., tocofersolan), a water-soluble derivative of α-tocopherol, compared to tocopherol acetate (TAC), the form usually used in clinical nutrition [ 13 ] by human wild-type Caco-2 TC7 cells, known to be a relevant model of the intestinal barrier [ 14 ]. In this work, we created a relevant knock-out (KO) cellular model of FHBL-SD1 and FHBL-SD3 by invalidating either MTTP or SAR1B in Caco-2/TC7 cells using the CRISPR/Cas9 genome-editing system.…”