Lutein transport by Caco-2 TC-7 cells occurs partly by a facilitated process involving the scavenger receptor class B type I (SR-BI)

Abstract

The carotenoid lutein is thought to play a role in the human eye and to protect against age-related macular degeneration. Lutein transport in the human intestine has not been characterized. We examined lutein transport processes using Caco-2 TC-7 monolayers as a model for human intestinal epithelium. Purified lutein was mixed with phospholipids, lysophospholipids, cholesterol, mono-olein, oleic acid and taurocholate to obtain lutein-rich mixed micelles that mimicked those found under physiological conditions. The micelles were added to the apical side of Caco-2 TC-7 cell monolayers for 30 min or 3 h at 37 degrees C. Absorbed lutein, i.e. the sum of lutein recovered in the scraped cells and in the basolateral chamber, was quantified by HPLC. Transport rate was measured (i) as a function of time (from 15 to 60 min), (ii) as a function of micellar lutein concentration (from 1.5 to 15 microM), (iii) at 4 degrees C, (iv) in the basolateral to apical direction, (v) after trypsin pretreatment, (vi) in the presence of beta-carotene and/or lycopene, (vii) in the presence of increasing concentrations of antibody against SR-BI (scavenger receptor class B type 1) and (viii) in the presence of increasing concentrations of a chemical inhibitor of the selective transfer of lipids mediated by SR-BI, i.e. BLT1 (blocks lipid transport 1). The rate of transport of lutein as a function of time and as a function of concentration was saturable. It was significantly lower at 4 degrees C than at 37 degrees C (approx. 50%), in the basal to apical direction than in the opposite direction (approx. 85%), and after trypsin pretreatment (up to 45%). Co-incubation with beta-carotene, but not lycopene, decreased the lutein absorption rate (approx. 20%) significantly. Anti-SR-BI antibody and BLT1 significantly impaired the absorption rate (approx. 30% and 57% respectively). Overall, these results indicate that lutein absorption is, at least partly, protein-mediated and that some lutein is taken up through SR-BI.

Effect of the main dietary antioxidants (carotenoids, γ-tocopherol, polyphenols, and vitamin C) on α-tocopherol absorption

“…The effect of lutein was lower and the few effect of b-carotene can be explained by its low micellar concentration. This inhibitory effect of carotenoids on a-tocopherol absorption, which has never been described before can be explained by competition between a-tocopherol and other carotenoids for uptake through the SR-BI receptor, which facilitates apical uptake of both classes of antioxidants (Reboul et al, 2005a, c). Concerning polyphenols, the second set of experiments showed that only naringenin impaired a-tocopherol absorption.…”

“…(R,R,R)-aand (R,R,R)-g-tocopherol, lutein, b-carotene and lycopene) to cells, mixed micelles rich in each of the microconstituents were separately prepared as described previously (Reboul et al, 2005a) to obtain the following final concentrations: 0.04 mM phosphatidylcholine, 0.16 mM lysophosphatidylcholine, 0.3 mM monoolein, 0.1 mM free cholesterol, 0.5 mM oleic acid, 5 mM taurocholate and 0.1-5 mM of the relevant microconstituent. Microconstituent concentrations in the micellar solutions were checked before each experiment.…”

“…Caco-2 clone TC7 cells (Salvini et al, 2002) were a gift from Dr M Rousset (U178 INSERM, Villejuif, France). Cells were cultured in the presence of DMEM supplemented with 20% heat-inactivated FBS, 1% nonessential amino acid and 1% antibiotics (complete medium), as described previously (Reboul et al, 2005a). For each experiment, cells were seeded and grown on semi-permeable filters as described previously (Reboul et al, 2005a) to obtain differentiated confluent cell monolayers.…”

“…Cells were cultured in the presence of DMEM supplemented with 20% heat-inactivated FBS, 1% nonessential amino acid and 1% antibiotics (complete medium), as described previously (Reboul et al, 2005a). For each experiment, cells were seeded and grown on semi-permeable filters as described previously (Reboul et al, 2005a) to obtain differentiated confluent cell monolayers. Twelve hours before each experiment, the medium of apical and basolateral chambers was changed to a serum-free complete medium.…”

Transport and Retinal Capture of Lutein and Zeaxanthin with Reference to Age-related Macular Degeneration

“…49,96 Recently, Reboul et al examined the transport of lutein using Caco-2 TC-7 monolayers as an in vitro model for human intestinal epithelium, in an attempt to characterize the transport of lutein across human enterocytes. 68 This in vitro model has previously been used by other investigators to evaluate the intestinal absorption of other carotenoids, 26,87 and it has been shown to correlate well with in vivo results. 3 Several interesting findings arose from these experiments: firstly, the rate of lutein uptake was saturable under physiological conditions; secondly, this rate of uptake was significantly impaired at 4 C; and thirdly, the rate of absorption was slower from the basolateral side of the cell monolayer to the apical side than in the opposite direction.…”