Emmanuelle Reboul
email: emmanuelle.reboul@univmed.fr

Patrick Borel
email: patrick.borel@univmed.fr

PROTEINS INVOLVED IN UPTAKE, INTRACELLULAR TRANSPORT AND BASOLATERAL SECRETION OF FAT-SOLUBLE VITAMINS AND CAROTENOIDS BY MAMMALIAN ENTEROCYTES

Keywords: bioavailability, brush border membrane, transporters, uptake, absorption, intestine

The present review provides an interesting and timely summary on the implication of active multifunctional transport systems in the uptake and cellular distribution of lipophilic vitamins and carotenoids. The topic addresses a rapidly developing field of research with strong impact on nutrition, nutrition-related diseases and nutrient deficiency. The paper summarizes modern molecular biological approaches in lipid/nutrition research to resolve long known phenomena (interindividual differences in absorption of lipids, interaction of lipids in the absorption process) and critically discuss the state of the research.

Abstract

Our understanding of the molecular mechanisms responsible for fat-soluble vitamin uptake and transport at the intestinal level has advanced considerably over the past decade. On one hand, it has long been considered that vitamin D and E as well as β-carotene (the main provitamin A carotenoid in human diet) were absorbed by a passive diffusion process, although this could not explain the broad inter-individual variability in the absorption efficiency of these molecules. On the other hand, it was assumed that preformed vitamin A (retinol) and vitamin K1 (phylloquinone) absorption occurred via energy-dependent processes, but the transporters involved have not yet been identified. The recent discovery of intestinal proteins able to facilitate vitamin E and carotenoid uptake and secretion by the enterocyte has spurred renewed interest in studying the fundamental mechanisms involved in the absorption of these micronutrients. The proteins identified so far are cholesterol transporters such as SR-BI (scavenger receptor class B type I), CD36 (cluster determinant 36), NPC1L1 (Niemann Pick C1-like 1) or ABCA1 (ATP-binding cassette A1) displaying a broad substrate specificity, but it is likely that other membrane proteins are also involved. After overviewing the metabolism of fat-soluble vitamins and carotenoids in the human upper gastrointestinal lumen, we will focus on the putative or identified proteins participating in the intestinal uptake, intracellular transport and basolateral secretion of these fat-soluble vitamins and carotenoids, and outline the uncertainties that need to be explored in the future. Identifying the proteins involved in intestinal uptake and transport of fat-soluble vitamins and carotenoids across the enterocyte is of great importance, especially as some of them are already targets for the development of drugs able to slow cholesterol absorption. Indeed, these drugs may also interfere with lipid vitamin uptake. A better understanding of the molecular mechanisms involved in fat-soluble vitamin and carotenoid absorption is a priority to better optimize their bioavailability.

INTRODUCTION

Vitamins are organic compounds with a relatively low molecular weight and no energetic value that are found in the diet in small quantities (< 1 g per d) yet are essential to ensure body growth, reproduction and functioning. As the body cannot synthesize its own vitamins (or at least not in sufficient amounts), adequate amounts of vitamins have to be provided by diet. Historically, the 13 vitamins have been divided into two classes based on their solubility: water-soluble vitamins (the Bs and C) and fat-soluble vitamins. Fat-soluble vitamins are further classed into 4 groups of compounds: vitamin A (preformed vitamin A and provitamin A carotenoids), D (cholecalciferol), E (tocopherols and tocotrienols) and K (phylloquinone and menaquinones) (see Table 1).

Nonprovitamin A carotenoids, such as lutein and lycopene (Table 2), are dietary lipid compounds. They cannot be considered true vitamins as their essentiality for humans has not yet been proven. However, they are assumed to have beneficial effects on human health [START_REF] Rao | Carotenoids and human health[END_REF][START_REF] Olson | Carotenoids and human health[END_REF], which in some cases may prove to be specific biological effects in humans, such as lutein and eye biology [START_REF] Stahl | Macular carotenoids: lutein and zeaxanthin[END_REF][START_REF] Krinsky | Biologic mechanisms of the protective role of lutein and zeaxanthin in the eye[END_REF][START_REF] Yagi | The effect of lutein supplementation on visual fatigue: A psychophysiological analysis[END_REF][START_REF] Ma | A 12-week lutein supplementation improves visual function in Chinese people with long-term computer display light exposure[END_REF]. We will thus include in this review the proteins involved in carotenoid uptake and metabolism in the enterocyte.

The fundamental mechanisms involved in the absorption of fat-soluble vitamins and carotenoids (FSV&C) were almost exclusively studied by Hollander's teams in the seventies [START_REF] Hollander | Vitamin K1 absorption by everted intestinal sacs of the rat[END_REF][START_REF] Hollander | Mechanism and site of vitamin K-3 small intestinal transport[END_REF][START_REF] Hollander | Mechanism and site of small intestinal absorption of alpha-tocopherol in the rat[END_REF][START_REF] Hollander | Vitamin K2 absorption by rat everted small intestinal sacs[END_REF][START_REF] Hollander | Factors affecting the absorption of vitamin K-1 in vitro[END_REF][START_REF] Hollander | Mechanism and site of small intestinal uptake of vitamin D3 in pharmacological concentrations[END_REF][START_REF] Hollander | Vitamin A1 intestinal absorption in vivo: influence of luminal factors on transport[END_REF][START_REF] Hollander | Vitamin K1 intestinal absorption in vivo: influence of luminal contents on transport[END_REF][START_REF] Hollander | Vitamin D-3 intestinal absorption in vivo: influence of fatty acids, bile salts, and perfusate pH on absorption[END_REF][START_REF] Hollander | Effect of luminal constituents on vitamin K1 absorption into thoracic duct lymph[END_REF][START_REF] Hollander | beta-carotene intestinal absorption: bile, fatty acid, pH, and flow rate effects on transport[END_REF][START_REF] Hollander | Intestinal absorption of vitamin A, E, D, and K[END_REF]. Thus, most of the assumptions regarding the absorption mechanisms of FSV&C come from these studies that, for the most part, were performed in rat everted intestinal sacs. Although well performed, these studies were obviously limited by the technologies available at the time. Nevertheless, they have suggested that some FSV&C) were absorbed by passive diffusion while others (preformed A and K) were absorbed via carrier-dependent proteins. Recent studies performed by several independent research groups over the past decade have revisited these assumptions, and have shown that these mechanisms of absorption are actually more complex than previously assumed: passive diffusion probably occurs at high, non-physiological concentrations of these compounds, while protein-mediated transport occurs at dietary doses.

Because the absorption mechanisms of these lipid molecules probably depend on the vehicle(s) in which they are provided to the apical membrane of the enterocyte, our first aim was to review current state-of-the-art on the metabolism of FSV&C in the human upper gastrointestinal (GI) lumen. Surprisingly, very little in known about the distribution of FSV&C between the different vehicles able to solubilize them in the human GI lumen, which include micelles, vesicles and, perhaps, proteins.

Our review of the literature also highlights that virtually nothing is known on the intracellular traffic of these molecules across the enterocyte, despite that fact that numerous candidate proteins are proposed with regard to the transport of these molecules in other cell types or tissues.

Finally, it is clear that numerous proteins assumed to transport cholesterol and other major lipids (fatty acids and phospholipids) are also involved in the transport of FSV&C. This review raises numerous questions, for which we propose numerous studies highlighting key points on the metabolism in the human enterocyte of these very important molecules.

STATE OF THE ART ON THE FATE OF FAT-SOLUBLE VITAMINS AND CAROTENOIDS IN THE HUMAN GI TRACT

FSV&C are assumed to follow the fate of lipids in the upper gastrointestinal tract [START_REF] Borel | Factors affecting intestinal absorption of highly lipophilic food microconstituents (fat-soluble vitamins, carotenoids and phytosterols)[END_REF].

However, some key issues on the fate of FSV&C in the GI lumen still need further investigation. For example, the main intestinal site of absorption of most of these compounds is not precisely localized, and their absorption efficiency remains a rough estimate given the wide range of data obtained in different studies. Finally, although it is assumed that these compounds are relatively labile, no significant chemical modification of carotenoids or vitamins A and E has been reported at the stomach level [START_REF] Tyssandier | Processing of vegetable-born carotenoids in the human stomach and duodenum[END_REF][START_REF] Borel | Processing of vitamin A and E in the human gastrointestinal tract[END_REF], and vitamin D and K are thought to follow the same pattern, although there is no clear data on this topic.

HYDROLYSIS OF FAT-SOLUBLE VITAMIN AND CAROTENOID ESTERS

It is assumed that only the free forms of the FSV&C can be absorbed, which suggests that hydrolysis should occur in the GI lumen when FSV&C esters are ingested. The main studies dedicated to this topic concern retinol, cholecalciferol and tocopherol esters. The hydrolysis may begin in the stomach, where gastric lipase can hydrolyze up to 17.5% of the triacyglycerols [START_REF] Carriere | Secretion and contribution to lipolysis of gastric and pancreatic lipases during a test meal in humans[END_REF]. However, this lipase has been shown to be relatively inefficient on retinyl palmitate hydrolysis [START_REF] Borel | Processing of vitamin A and E in the human gastrointestinal tract[END_REF]. The hydrolysis of FSV&C esters thus occurs mainly in the duodenum. Contrary to previous assumptions, this hydrolysis is mainly performed by pancreatic lipase and, at least for retinyl-palmitate, by pancreatic lipase-related protein 2 [START_REF] Reboul | Pancreatic lipase and pancreatic lipase-related protein 2, but not pancreatic lipase-related protein 1, hydrolyze retinyl palmitate in physiological conditions[END_REF]. Indeed, the initial involvement of cholesterol ester hydrolase [START_REF] Lombardo | Studies on the substrate specificity of a carboxyl ester hydrolase from human pancreatic juice. II. Action on cholesterol esters and lipid-soluble vitamin esters[END_REF][START_REF] Zahalka | Hydrolysis of stereoisomeric alphatocopheryl acetates catalyzed by bovine cholesterol esterase[END_REF][START_REF] Lauridsen | Hydrolysis of tocopheryl and retinyl esters by porcine carboxyl ester hydrolase is affected by their carboxylate moiety and bile acids[END_REF][START_REF] Breithaupt | Carotenol fatty acid esters: easy substrates for digestive enzymes?[END_REF] was not confirmed in vivo using knock-out mice [START_REF] Van Bennekum | Carboxyl ester lipase overexpression in rat hepatoma cells and CEL deficiency in mice have no impact on hepatic uptake or metabolism of chylomicron-retinyl ester[END_REF][START_REF] Weng | Intestinal absorption of dietary cholesteryl ester is decreased but retinyl ester absorption is normal in carboxyl ester lipase knockout mice[END_REF]. Other candidates could be a brush border esterase and a mucosal esterase localized to the endoplasmic reticulum of the enterocyte. Indeed, analytical subcellular fractionation studies on homogenates of isolated jejunal rat enterocytes showed an esterase activity against tocopheryl acetate localized to this organelle [START_REF] Mathias | Studies on the in vivo absorption of micellar solutions of tocopherol and tocopheryl acetate in the rat: demonstration and partial characterization of a mucosal esterase localized to the endoplasmic reticulum of the enterocyte[END_REF]. The exact proportion of FSV&C hydrolyzed by these enzymes is unknown, but it is assumed that the bulk of FSV&C esters are hydrolyzed in the intestinal lumen by pancreatic lipases. The remaining esters may be hydrolyzed intraluminally by mucosal enzymes or taken up intact by the intestinal cell and hydrolyzed intracellularly. However, as stated above, the consensus is that vitamin esters cannot be absorbed by the enterocyte.

DISTRIBUTION OF FAT-SOLUBLE VITAMINS AND CAROTENOIDS

BETWEEN VEHICLES ABLE TO SOLUBILIZE THEM IN THE GI LUMEN DURING DIGESTION, i.e. MICELLES, VESICLES AND PROTEINS

It is assumed that during digestion, FSV&C are transferred from the food matrix in which they are embedded to the mixed micelles generated by lipolysis of dietary triacylglycerols [START_REF] Borel | Factors affecting intestinal absorption of highly lipophilic food microconstituents (fat-soluble vitamins, carotenoids and phytosterols)[END_REF]. Mixed micelles are a mixture of phospholipids, cholesterol, lipid digestion products (such as free fatty acids, monoacylglycerols and lysophospholipids) and bile salts. It has been shown that a fraction of some FSV&C first transfers to the fat phase of the meal in the gastric environment [START_REF] Tyssandier | Processing of vegetable-born carotenoids in the human stomach and duodenum[END_REF][START_REF] Rich | Low pH enhances the transfer of carotene from carrot juice to olive oil[END_REF]. This transfer may only happen in the stomach, as mixed micelles present in the intestine apparently inhibit the carotenoid transfer from the food matrix to the fat phase [START_REF] Rich | Solubilization of carotenoids from carrot juice and spinach in lipid phases: II. Modeling the duodenal environment[END_REF]. FSV&C are then assumed to transfer to mixed micelles during dietary lipid lipolysis by the gut lipases, a phenomenon affected by several factors, including pH and bile lipid concentration [START_REF] Tyssandier | Main factors governing the transfer of carotenoids from emulsion lipid droplets to micelles[END_REF][START_REF] Borel | Digestion and absorption of tube-feeding emulsions with different droplet sizes and compositions in the rat[END_REF]. However, it should be underlined that direct transfer from the food matrix to the mixed micelles within the intestinal lumen may also occur [START_REF] Rich | Solubilization of carotenoids from carrot juice and spinach in lipid phases: II. Modeling the duodenal environment[END_REF]. The solubilization efficiency of FSV&C in micelles depends on a wide array of factors, including FSV&C species [START_REF] Borel | Factors affecting intestinal absorption of highly lipophilic food microconstituents (fat-soluble vitamins, carotenoids and phytosterols)[END_REF]. Indeed, the micellar solubility of different carotenoids, and even the solubility of different isomers of the same carotenoid, can be quite different [START_REF] Levin | Incorporation of all-trans-or 9-cis-beta-carotene into mixed micelles in vitro[END_REF]. It is assumed that the higher the percentage of FSV&C incorporation in micelles (a percentage called "bioaccessibility"), the higher their absorption efficiency.

Although an important step, FSV&C solubility in micelles is not the only factor to affect FSV&C absorption. Indeed, the nature of the lipids that constitute these micelles also plays a key role in this process. As expected, the amount of a liposoluble vitamin (phylloquinone) solubilized in mixed micelles dramatically increased with increasing amounts of phospholipids [START_REF] Nagata | Solubilization of vitamin K1 by bile salts and phosphatidylcholine-bile salts mixed micelles[END_REF]. However, it has been clearly shown by several teams that phospholipids can impair the absorption of cholesterol [START_REF] Homan | Phospholipase A2 relieves phosphatidylcholine inhibition of micellar cholesterol absorption and transport by human intestinal cell line Caco-2[END_REF], carotenoids [START_REF] Sugawara | Lysophosphatidylcholine enhances carotenoid uptake from mixed micelles by Caco-2 human intestinal cells[END_REF] and vitamin E [START_REF] Koo | Phosphatidylcholine inhibits and lysophosphatidylcholine enhances the lymphatic absorption of alpha-tocopherol in adult rats[END_REF]. The hypothesis to explain this inhibition is that because of their high hydrophobicity, cholesterol and FSV&C are associated with the long-chain acyl moieties of phospholipids in mixed micelles, resulting in reduced uptake by intestinal cells. The inhibitory effect of phospholipids on the absorption of cholesterol, αtocopherol and carotenoids is abolished when they are substituted by lysophospholipids [START_REF] Homan | Phospholipase A2 relieves phosphatidylcholine inhibition of micellar cholesterol absorption and transport by human intestinal cell line Caco-2[END_REF][START_REF] Sugawara | Lysophosphatidylcholine enhances carotenoid uptake from mixed micelles by Caco-2 human intestinal cells[END_REF][START_REF] Koo | Phosphatidylcholine inhibits and lysophosphatidylcholine enhances the lymphatic absorption of alpha-tocopherol in adult rats[END_REF][START_REF] Raju | Influence of phospholipids on beta-carotene absorption and conversion into vitamin A in rats[END_REF]. As lysophospholipid micelles are smaller than phospholipid micelles, micelle size may be one of the factors that determine the absorption of micellar lipophilic components. However, a comparison of cholesterol uptake from lysophospholipid and phospholipid micelles, which were similar in size, showed that uptake was still lower in phospholipid micelles [START_REF] Reynier | Intestinal cholesterol uptake: comparison between mixed micelles containing lecithin or lysolecithin[END_REF]. Given that lysophospholipid uptake by intestinal cells is much greater than phospholipid uptake [START_REF] Homan | Phospholipase A2 relieves phosphatidylcholine inhibition of micellar cholesterol absorption and transport by human intestinal cell line Caco-2[END_REF][START_REF] Koo | Phosphatidylcholine inhibits and lysophosphatidylcholine enhances the lymphatic absorption of alpha-tocopherol in adult rats[END_REF], it has been suggested that the increased cellular level of lipids they induce may shift the balance of FSV&C distribution from the mixed micelles to the cells [START_REF] Sugawara | Lysophosphatidylcholine enhances carotenoid uptake from mixed micelles by Caco-2 human intestinal cells[END_REF].

Although it is assumed that, in the GI lumen, the FSV&C recovered in the mixed micellerich aqueous fraction are localized into the mixed micelles, it cannot be excluded that a part of these molecules is incorporated in other lipid structures present in this aqueous fraction. Indeed, different lipid structures do coexist in the GI lumen during digestion: lipid droplets, vesicles (monolamellar and multilamellar liposomes), micelles and, if ingested, milk fat globules. Conversely to mixed micelles which are a mixture of lipids and bile salts, vesicles are assumed to be liposome-like structures composed of either a single bilayer (unilamellar vesicles) or multiple bilayers (multilamellar vesicles) of phospholipids. The hypothesis that FSV&C can be solubilized in vesicles during absorption is supported by the observation that some FSV&C (vitamin A, E and D) are incorporated in phospholipid bilayers [START_REF] Kirilenko | Fat soluble vitamins in liposomes: studies on incorporation efficiency and bile salt induced vesicle disintegration[END_REF][START_REF] Noy | Interactions of retinol with lipid bilayers: Studies with vesicles of different radii[END_REF].

Moreover, some FSV&C are even able to promote vesicle formation. Indeed, α-tocopherol promoted the assembly of bilayer structures and stabilized these structures in systems composed of phospholipids, whereas in the absence of α-tocopherol, phospholipids produced micellar structures [START_REF] Salgado | Alpha-tocopherol interacts with natural micelle-forming single-chain phospholipids stabilizing the bilayer phase[END_REF]. Moreover, vesicle stability to a bile salt (sodium deoxycholate) was enhanced by the presence of FSV&C (vitamin A, E and D) [START_REF] Kirilenko | Fat soluble vitamins in liposomes: studies on incorporation efficiency and bile salt induced vesicle disintegration[END_REF]. Finally, note that FSV&C can also transfer between unilamellar vesicles [START_REF] Ho | Spontaneous transfer of retinoic acid, retinyl acetate, and retinyl palmitate between single unilamellar vesicles[END_REF].

The distribution of FSV&C between the lipid structures, i.e. emulsified lipid droplets, vesicles and micelles, and their localization within these structures, i.e. in the core or the surface of lipid droplets [START_REF] Borel | Carotenoids in biological emulsions: solubility, surface-to-core distribution, and release from lipid droplets[END_REF] or inside or across phospholipid bilayers, depends on their solubility and their ability to interact with the different lipid classes that constitute these structures [START_REF] Borel | Carotenoids in biological emulsions: solubility, surface-to-core distribution, and release from lipid droplets[END_REF][START_REF] Avellone | Spectrophotometric investigation of the binding of vitamin E to water-containing reversed micelles[END_REF][START_REF] Lancrajan | Carotenoid incorporation into natural membranes from artificial carriers: liposomes and beta-cyclodextrins[END_REF][START_REF] Villalain | Calorimetric and infrared spectroscopic studies of the interaction of alpha-tocopherol and alpha-tocopheryl acetate with phospholipid vesicles[END_REF][START_REF] Sujak | Organisation of xanthophyll pigments lutein and zeaxanthin in lipid membranes formed with dipalmitoylphosphatidylcholine[END_REF]. For example, Woodall et al. [START_REF] Woodall | Carotenoids and protection of phospholipids in solution or in liposomes against oxidation by peroxyl radicals: relationship between carotenoid structure and protective ability[END_REF] found a significant difference in orientation of β-carotene (an apolar carotenoid) compared to zeaxanthin (a dihydroxy carotenoid) with respect to phospholipid bilayers. More precisely, the long axis of zeaxanthin formed a narrow angle of about 45° with the axis normal to the plane of the bilayer, while β-carotene was distributed homogeneously within the bilayer without any preferred well-defined orientation. Another example is the very different distribution of βcarotene and zeaxanthin between core and surface of triacylglycerol droplets stabilized with phospholipids [START_REF] Borel | Carotenoids in biological emulsions: solubility, surface-to-core distribution, and release from lipid droplets[END_REF].

The presence of FSV&C in a mixed micelle-rich aqueous phase does not rule out that a fraction of FSV&C may be associated with proteins solubilised in this aqueous phase either. Indeed, β-lactoglobulin, a lipocalin recovered in cow milk, is able to bind retinol and βcarotene [START_REF] Perez | Interaction of b-lactoglobulin with retinol and fatty acids and its role as a possible biological function for this protein : a review[END_REF][START_REF] Said | Intestinal uptake of retinol: enhancement by bovine milk beta-lactoglobuline[END_REF][START_REF] Godovac-Zimmermann | The structural motif of -Lactoglobulin and Retinol-Binding Protein:a basic framework for binding and transport of small hydrophobic molecules[END_REF][START_REF] Dufour | Binding of retinoids and beta-carotene to beta-lactoglobulin. Influence of protein modifications[END_REF]. It is therefore possible that proteins present in the gut and originating either from diet or from pancreatic/biliary secretions can bind a fraction of FSV&C and transport them to the enterocyte.

In conclusion, it is reasonable to suggest that FSV&C are not only recovered in mixed micelles but might also be distributed between different vehicles (micelles, vesicles and perhaps proteins) prior to their absorption by the intestinal cell. It is also reasonable to suggest that the mechanisms of absorption depend on the vehicles with which the FSV&C are associated. Indeed, mixed micelles may interact with some membrane proteins while vesicles or proteins may have totally distinct specific transport pathways. Thus, better knowledge of the distribution of FSV&C between the different vehicles able to solubilise them in the intestinal lumen will likely allow a better understanding of the mechanisms governing their absorption.

MAIN SITE OF FAT-SOLUBLE VITAMIN & CAROTENOID ABSORPTION

The main site of absorption of FSV&C is assumed to be in the mid sections of the GI tract.

However, the exact localization of this site in humans remains unknown, as the only data on this topic have been obtained in rodents. More precisely, by using everted small bowel sacs, Hollander et al. identified the sites where maximal absorption of vitamin E, D and K takes place in rats. Surprisingly, there is no such data for vitamin A, whether for retinol or for provitamin A carotenoids. Concerning vitamin E, Hollander's group found that the highest rate of absorption takes place in the medial portion, assumed to be the jejunum, of the small bowel [START_REF] Hollander | Mechanism and site of small intestinal absorption of alpha-tocopherol in the rat[END_REF]. They also found that maximal vitamin D uptake occurs in both jejunum and ileum [START_REF] Hollander | Vitamin D-3 intestinal absorption in vivo: influence of fatty acids, bile salts, and perfusate pH on absorption[END_REF], which is in agreement with our recent data obtained in mice [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF]. Finally, concerning vitamin K, Hollander's team found a maximal uptake of phylloquinone (vitamin K1) in the proximal gut segments, assumed to be duodenum and jejunum, and a relatively poor uptake in distal segments, assumed to be ileum and colon [START_REF] Hollander | Vitamin K1 absorption by everted intestinal sacs of the rat[END_REF], while bacterial menaquinones are assumed to be preferentially absorbed in the colon.

An initial hypothesis to explain why FSV&C absorption efficiency is not identical along the gut and why different FSV&C share different main sites of absorption is that intestinal proteins involved in FSV&C absorption are not homogeneously localized along the gut. Indeed, it is likely that the absorption efficiency of a given FSV&C is maximal where the main intestinal transporters of this FSV&C are expressed. The in-gut distribution of transporters involved in the absorption of cholesterol and long-chain fatty acidstransporters also involved in absorption of some FSV&C (see paragraph 3.1) -has recently been studied in post-mortem intestinal samples from 11 subjects. Data showed bell-shape patterns, with the highest levels in ileum for ABCA1, ABCG8, NPC1L1 and CD36 [START_REF] Masson | Fatty acid-and cholesterol transporter protein expression along the human intestinal tract[END_REF].

Another important point that can affect FSV&C absorption is the distribution of membrane transporters between the apical and the basolateral membrane of the enterocyte. Indeed, this distribution can vary along the gut. SR-BI, which is involved in the absorption of several FSV&C, was present on the apical surface of the proximal intestine, while the presence of SR-BI on the basolateral surface of the distal intestine suggested its possible involvement in intestinal lipoprotein uptake or secretion [START_REF] Cai | Differentiation-dependent expression and localization of the class B type I scavenger receptor in intestine[END_REF]. Moreover, SR-BI mRNA expression appears to be regulated by the intestine-specific transcription factor Isx (Intestine Specific Homebox) [START_REF] Choi | A dynamic expression survey identifies transcription factors relevant in mouse digestive tract development[END_REF][START_REF] Seino | Isx participates in the maintenance of vitamin A metabolism by regulation of beta-carotene 15,15'monooxygenase (Bcmo1) expression[END_REF][61] It is thus possible that the preferred site of absorption of FSV&C vary within individuals depending on genetics and diet. Better knowledge of the distribution of proteins involved in FSV&C absorption along the intestinal tract, and within the enterocyte, would significantly enhance our understanding of FSV&C absorption.

A second hypothesis to explain the occurrence of major absorption sites for some FSV&C is that these sites may be those where the availability of FSV&C is highest, i.e. where FSV&C are recovered at their maximal concentrations in vehicles that allow their maximal absorption. These sites would therefore depend on the ability of each FSV&C class to be extracted from its food matrix and solubilized in its physiological vehicles (micelles, vesicles, and proteins that facilitate its absorption) [START_REF] Said | Intestinal uptake of retinol: enhancement by bovine milk beta-lactoglobuline[END_REF]. Nevertheless, these two hypotheses are probably linked, and we suggest that transporters are mostly expressed in the regions of the gut where FSV&C are mostly recovered in their optimal vehicles for absorption.

ABSORPTION EFFICIENCY (% OF A GIVEN DOSE)

Data on FSV&C absorption efficiency are scarce and have been obtained with very different models (animals, healthy humans or patients with intestinal malabsorption, ileostomy patients). They have also been obtained with very variable doses of FSV&C, which were incorporated into different matrices and ingested in different meals. Knowing that i) FSV&C absorption is affected by numerous factors, including FSV&C dose, food matrix and meal nutrients [START_REF] Borel | Factors affecting intestinal absorption of highly lipophilic food microconstituents (fat-soluble vitamins, carotenoids and phytosterols)[END_REF], ii) absorption is probably mediated by intestinal proteins at dietary doses but is probably passive at pharmacological doses, and iii) there is a huge interindividual variability in the absorption efficiency of most FSV&C [START_REF] Borel | Low and high responders to pharmacological doses of beta-carotene: proportion in the population, mechanisms involved and consequences on beta-carotene metabolism[END_REF][START_REF] Jeanes | Comparative (2)H-labelled alpha-tocopherol biokinetics in plasma, lipoproteins, erythrocytes, platelets and lymphocytes in normolipidaemic males[END_REF], all these factors likely explains the huge variability in published data.

Concerning vitamin A, it has been reported that for preformed vitamin A (retinol), absorption efficiency ranges between 75% [START_REF] Biesalski | Bioavailability of vitamin A[END_REF] and about 100% [START_REF] Sivakumar | Absorption of labelled vitamin A in children during infection[END_REF]. This range is less variable than the range reported for absorption efficiency of provitamin A carotenoids, i.e. from about 3% to 90% for β-carotene [START_REF] O'neill | Intestinal absorption of -carotene, lycopene and lutein in men and women following a standard meal: response curves in the triacylglycerol-rich lipoprotein fraction[END_REF][START_REF] Novotny | Compartmental analysis of the dynamics of beta-carotene metabolism in an adult volunteer[END_REF][START_REF] Van Vliet | Intestinal beta-carotene absorption and cleavage in men: response of beta-carotene and retinyl esters in the triglyceride-rich lipoprotein fraction after a single oral dose of beta-carotene[END_REF][START_REF] Faulks | Absorption of all-trans and 9-cis beta-carotene in human ileostomy volunteers[END_REF][START_REF] Van Lieshout | Isotopic tracer techniques for studying the bioavailability and bioefficacy of dietary carotenoids, particularly beta-carotene, in humans: a review[END_REF][START_REF] Van Loo-Bouwman | Vitamin A equivalency and apparent absorption of beta-carotene in ileostomy subjects using a dual-isotope dilution technique[END_REF]. Furthermore, it is assumed that preformed vitamin A has a higher absorption efficiency than provitamin A carotenoidsan assumption confirmed by data obtained in our laboratory using Caco-2 cells (unpublished data). This could be explained by the presence of an efficient specific transporter for retinol, while provitamin A carotenoids are absorbed via non-specific transporters (see paragraph

3.1).

Concerning vitamin E, different studies report absorption efficiency in ranges of 10-95% [START_REF] Drevon | absorption, transport and metabolism of vitamin E[END_REF][START_REF] Traber | Vitamin E in humans: demand and delivery[END_REF][START_REF] Cohn | Bioavailability of vitamin E[END_REF]. However when deuterium-labeled vitamin E was used to estimate absorption, this range was reduced to 10-33% [START_REF] Bruno | Human vitamin E requirements assessed with the use of apples fortified with deuterium-labeled alpha-tocopheryl acetate[END_REF]. These nevertheless huge ranges were probably due to the different methods used to estimate absorption, to the effect of food matrix, and to broad inter-individual variability in absorption efficiency.

The absorption efficiency of vitamin D3 given in peanut oil was found to vary between 55% and 99% (mean 78%) in patients with intestinal malabsorption and control subjects [START_REF] Thompson | Absorption of vitamin D3-3 H in control subjects and patients with intestinal malabsorption[END_REF].

Animal experiments have reported similar rates, percentages ranging between 66% and 75%.

There is little data available on the absorption efficiency of vitamin K. The absorption efficiency of vitamin K1 supplied in its natural matrix, i.e. vegetables, is assumed to be relatively low, because it was found that only about 13% was absorbed from spinach [START_REF] Shearer | Studies on the absorption and metabolism of phylloquinone (vitamin K 1 ) in man[END_REF].

Absorption efficiency is assumed to be higher when vitamin K is given as a supplement.

Indeed, one study found that healthy subjects excreted less than 20% of a 1 mg dose of radiolabeled phylloquinone, suggesting that 80% was absorbed.

On the whole, this review of the literature shows that it is difficult to provide a single narrow range for the absorption efficiency of each FSV&C in humans. This is likely because absorption of FSV&C is affected by numerous factors [START_REF] Borel | Factors affecting intestinal absorption of highly lipophilic food microconstituents (fat-soluble vitamins, carotenoids and phytosterols)[END_REF], but also because there is a huge inter-individual variability in the absorption efficiency of these molecules [START_REF] Borel | Low and high responders to pharmacological doses of beta-carotene: proportion in the population, mechanisms involved and consequences on beta-carotene metabolism[END_REF][START_REF] Jeanes | Comparative (2)H-labelled alpha-tocopherol biokinetics in plasma, lipoproteins, erythrocytes, platelets and lymphocytes in normolipidaemic males[END_REF].

MECHANISMS OF UPTAKE BY THE INTESTINAL CELLS: PASSIVE DIFFUSION AND/OR PROTEIN-MEDIATED TRANSPORT

ARGUMENTS FOR PASSIVE DIFFUSION

It has long been assumed that intestinal absorption of vitamin D, E and carotenoids occurs by passive diffusion. The first vitamin to be studied was vitamin E. The intestinal transport of α-tocopherol was described as non-ATP-dependent, as the addition of 2,4-dinitrophenol, sodium azide or potassium cyanide, all of which inhibit ATP synthesis, to the incubation medium of everted rat bowel sacs did not impair its absorption. Moreover, the fact that vitamin E uptake was linear up to 1.2 mM was another argument in favour of passive diffusion [START_REF] Hollander | Mechanism and site of small intestinal absorption of alpha-tocopherol in the rat[END_REF]. A few years later, the same team used living unanaesthetized rats to conclude that vitamin D absorption was also passive. Indeed, cholecalciferol uptake was increased by H+ ions, which are supposed to decrease cell membrane resistance to micelle diffusion, and increased by perfusate flow rate, which diminishes the thickness of the unstirred water layer [START_REF] Hollander | Vitamin D-3 intestinal absorption in vivo: influence of fatty acids, bile salts, and perfusate pH on absorption[END_REF]. Similar experiments performed with -carotene led to comparable conclusions, i.e. that carotenoids were absorbed by passive diffusion through the brush border membrane (BBM) [START_REF] Hollander | beta-carotene intestinal absorption: bile, fatty acid, pH, and flow rate effects on transport[END_REF].

Finally, retinol uptake by Caco-2 cells was shown to occur by passive diffusion at pharmacological doses [START_REF] Quick | Vitamin A metabolism in the human intestinal caco-2 cell line[END_REF], which confirmed the early data obtained in rat intestinal segments for retinol concentrations above 450 nM [START_REF] Hollander | Vitamin A1 intestinal absorption in vivo: influence of luminal factors on transport[END_REF][START_REF] Hollander | Intestinal absorption of vitamin A, E, D, and K[END_REF].

ARGUMENTS FOR PROTEIN-MEDIATED TRANSPORT

The hypothesis of a passive diffusion mechanism for carotenoid and vitamin E and D uptake cannot explain the higher absorption of vitamin E in the mid-intestine compared to other parts of the intestine [START_REF] Hollander | Mechanism and site of small intestinal absorption of alpha-tocopherol in the rat[END_REF]. It also cannot explain the huge inter-individual variability in absorption observed during postprandial studies [START_REF] Borel | Low and high responders to pharmacological doses of beta-carotene: proportion in the population, mechanisms involved and consequences on beta-carotene metabolism[END_REF][START_REF] Jeanes | Comparative (2)H-labelled alpha-tocopherol biokinetics in plasma, lipoproteins, erythrocytes, platelets and lymphocytes in normolipidaemic males[END_REF]. Further, it cannot explain the isomer selectivity and competition for absorption between carotenoids in Caco-2 cells [START_REF] During | Carotenoid uptake and secretion by CaCo-2 cells: beta-carotene isomer selectivity and carotenoid interactions[END_REF],

nor the competition between vitamin E and the carotenoid canthaxanthin observed in rats [START_REF] Hageman | Excess vitamin E decreases canthaxanthin absorption in the rat[END_REF]. Moreover, a close look at the data obtained for -carotene shows that a saturation of absorption in the distal part of the intestine cannot be ruled out [START_REF] Hollander | beta-carotene intestinal absorption: bile, fatty acid, pH, and flow rate effects on transport[END_REF]. Finally, the identification of the Drosophila gene ninaD encoding a class B Scavenger receptor essential for carotenoid distribution into cells is another pioneering finding in favour of a proteinmediated membrane transport of carotenoids [START_REF] Kiefer | A class B scavenger receptor mediates the cellular uptake of carotenoids in Drosophila[END_REF]. Thus, all these arguments are in favour of a protein-mediated transport, which is not totally contradictory with Hollander et al's results, as i) some membrane proteins involved in FSV&C uptake are not ATP-dependent (see paragraph 3.1), and ii) both protein-mediated transport and passive diffusion are observed depending on the FSV&C dose used in experiments [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF].

Besides these observations, retinol uptake at physiological doses by Caco-2 cells has been shown to occur by a saturable carrier-mediated process [START_REF] Quick | Vitamin A metabolism in the human intestinal caco-2 cell line[END_REF]. This concurs with experiments performed in rats showing a facilitated uptake at a concentration of 150 nM [START_REF] Hollander | Vitamin A1 intestinal absorption in vivo: influence of luminal factors on transport[END_REF][START_REF] Hollander | Intestinal absorption of vitamin A, E, D, and K[END_REF]. However, despite more recent in vivo investigations being performed [START_REF] Dew | Specificity of the retinol transporter of the rat small intestine brush border[END_REF], no protein has yet been identified as involved in retinol transport BBM level.

Similarly to retinol, phylloquinone is also thought to be transported by a protein in the intestine [START_REF] Hollander | Vitamin K1 intestinal absorption in vivo: influence of luminal contents on transport[END_REF]. Indeed, the use of a nitrogen atmosphere (to inhibit mitochondrial respiration) as well as the addition of 2,4-dinitrophenol (to inhibit ATP synthesis) decreased phylloquinone absorption in rat intestine. However, again, no membrane transporter has yet been identified.

As a final point, it is equally important to underline that the phenomena of protein-mediated transport and passive diffusion may be complementary, with protein-mediated transport occurring at dietary concentrations and passive diffusion taking over at pharmacological doses.

MEMBRANE PROTEINS INVOLVED IN FAT-SOLUBLE VITAMIN & CAROTENOID APICAL UPTAKE AND EFFLUX

The fact that membrane proteins are involved in the uptake of FSV&C by the enterocyte, at least at dietary concentrations, raises two main questions: "are these proteins specific for some FSV&C?", and "can different membrane proteins be involved in the uptake of the same FSV&C?". Furthermore, as some membrane proteins involved in cholesterol uptake demonstrate broad substrate specificity, "do they play a role in FSV&C uptake?".

The answer to the first question is probably "yes" for preformed vitamin A, i.e. retinol, because 1) a study in isolated rat intestinal segments suggested that retinol absorption is mediated by a saturable carrier-mediated process [START_REF] Hollander | Vitamin A1 intestinal absorption in vivo: influence of luminal factors on transport[END_REF], 2) it has a much higher absorption efficiency than any other FSV&C, suggesting a specific transporter (personal, non published data), and 3) its absorption does not involve a membrane transporter with broad substrate specificity identified as playing a role in other FSV&C (see paragraph 3.1.1), which again points to a very specific transporter. A good candidate as a protein specific for retinol uptake by intestinal cells would be STRA6 (STimulated by Retinoic Acid 6), which has been identified as a specific membrane receptor for RBP (Retinol-Binding Protein) [START_REF] Kawaguchi | A membrane receptor for retinol binding protein mediates cellular uptake of vitamin A[END_REF].

This protein is found in many -but not all -tissues, and is expressed in the intestine [START_REF] Bouillet | Developmental expression pattern of Stra6, a retinoic acid-responsive gene encoding a new type of membrane protein[END_REF].

Interestingly, there is a synergy between STRA6 and Lecithin Retinol Acyl Transferase (LRAT) expression [START_REF] Isken | RBP4 disrupts vitamin A uptake homeostasis in a STRA6-deficient animal model for Matthew-Wood syndrome[END_REF].

Cells expressing both these proteinswhich is the case of intestinal cells -can uptake more retinol than cells expressing each protein individually. This indicates that the conversion of retinol into retinyl ester by LRAT within the cell maintains the driving force for STRA6mediated retinol uptake. It has been shown that STRA6 acts as a bidirectional transporter of retinol, with intracellular retinol concentrations determining the polarity of transport [START_REF] Isken | RBP4 disrupts vitamin A uptake homeostasis in a STRA6-deficient animal model for Matthew-Wood syndrome[END_REF]. It is possible that STRA6 functions at a basal level to import retinol into the cell in order to maintain intracellular retinoid homeostasis. In times of surplus retinol within the cell, it can be upregulated and thus act to export retinol, protecting the cell from a potentially cytotoxic build-up of retinoid. STRA6 expression control is likely tightly regulated at both gene and protein levels, and would depend on the cellular context and retinoid status of the animal.

There is also probably a specific transporter for vitamin K, or at least for vitamin K1 (phylloquinone), which is the other fat-soluble vitamin that apparently required an energysaturable transport mechanism for its absorption according to Hollander's studies [START_REF] Hollander | Vitamin K1 absorption by everted intestinal sacs of the rat[END_REF][START_REF] Hollander | Vitamin K2 absorption by rat everted small intestinal sacs[END_REF][START_REF] Hollander | Vitamin K1 intestinal absorption in vivo: influence of luminal contents on transport[END_REF][START_REF] Hollander | Effect of luminal constituents on vitamin K1 absorption into thoracic duct lymph[END_REF][START_REF] Hollander | Intestinal absorption of vitamin A, E, D, and K[END_REF]. The fact that there is a specific transporter for vitamin K, which is supported by only one study [START_REF] Hollander | Vitamin K1 intestinal absorption in vivo: influence of luminal contents on transport[END_REF], should be confirmed with more recent methodologies. Moreover, it should be kept in mind that other forms of vitamin K, i.e. menaquinones, are also recovered in the diet. Menaquinones are assumed to be absorbed by passive diffusion [START_REF] Hollander | Vitamin K2 absorption by rat everted small intestinal sacs[END_REF], but further studies are required to assess whether the lipid transporters described below are also involved, at least in part, in phylloquinone and menaquinone uptake by intestinal cells.

The answers to the other two questions are given in the text that follows.

LIPID TRANSPORTERS INVOLVED IN FAT-SOLUBLE VITAMIN & CAROTENOID UPTAKE

In recent years, lipid transporters -and especially cholesterol transporters -have been identified as playing a role in FSV&C uptake by the intestinal cell.

Scavenger receptor class B type I (SR-BI)

SR-BI (also known as CLA1 for CD36 and LIMPII Analogous-1 in humans) is a singlechain transmembrane glycoprotein of 80 kDa. SR-BI is found in various cell types including liver, testis, ovaries and macrophages [START_REF] Terpstra | Hepatic and extrahepatic scavenger receptors: function in relation to disease[END_REF], as well as on the BBM of enterocytes from the duodenum to the colon [START_REF] Lobo | Localization of the lipid receptors CD36 and CLA-1/SR-BI in the human gastrointestinal tract: towards the identification of receptors mediating the intestinal absorption of dietary lipids[END_REF]. Its role in lipoprotein metabolism is very important because it is able to bind not just HDL (high density lipoprotein) but also LDL (low-density lipoproteins) and VLDL (very-low-density lipoproteins) [START_REF] Terpstra | Hepatic and extrahepatic scavenger receptors: function in relation to disease[END_REF]. SR-BI protein has the ability to facilitate the selective entry in the cell compartment of esterified cholesterol recovered in HDL [START_REF] Krieger | Charting the fate of the "good cholesterol": identification and characterization of the high-density lipoprotein receptor SR-BI[END_REF]. At the intestinal level, it has first been shown that SR-BI facilitates the uptake of free cholesterol but also esterified cholesterol, phospholipids and triacylglycerol hydrolysis products, thus displaying broad ligand specificity [START_REF] Hauser | Identification of a receptor mediating absorption of dietary cholesterol in the intestine[END_REF][START_REF] Bietrix | Accelerated lipid absorption in mice overexpressing intestinal SR-BI[END_REF]. However, the effective role of SR-BI compared to another transporterthe Niemann Pick C1-like 1 (NPC1L1)in terms of cholesterol transport is still a matter of debate [START_REF] Nguyen | Influence of class B scavenger receptors on cholesterol flux across the brush border membrane and intestinal absorption[END_REF]. We hypothesize that the main role of SR-BI in the intestine is to facilitate the uptake of other lipid molecules. Indeed, in 2005, a study using Caco-2 cells identified SR-BI as playing a role in the intestinal uptake of the carotenoid lutein [START_REF] Reboul | Lutein transport by Caco-2 TC-7 cells occurs partly by a facilitated process involving the scavenger receptor class B type I (SR-BI)[END_REF], and this involvement has since been extended to other carotenoids such as -carotene [START_REF] Van Bennekum | Class B scavenger receptor-mediated intestinal absorption of dietary beta-carotene and cholesterol[END_REF], zeaxanthin [START_REF] During | Xanthophylls are preferentially taken up compared with beta-carotene by retinal cells via a SRBI-dependent mechanism[END_REF] and lycopene [START_REF] Moussa | Lycopene absorption in human intestinal cells and in mice involves scavenger receptor class B type I but not Niemann-Pick C1-like 1[END_REF] in various other tissues.

Moreover, after being shown to be involved in vitamin E uptake in the liver [START_REF] Witt | Regulation by vitamin E of the scavenger receptor BI in rat liver and HepG2 cells[END_REF] and in the brain [START_REF] Goti | Scavenger receptor class B, type I is expressed in porcine brain capillary endothelial cells and contributes to selective uptake of HDL-associated vitamin E[END_REF], SR-BI was then shown to mediate tocopherol uptake across the BBM in both Caco-2 and mice models [START_REF] Reboul | Scavenger receptor class B type I (SR-BI) is involved in vitamin E transport across the enterocyte[END_REF]. Finally, SR-BI was recently shown to be involved in vitamin D uptake via to a combined approach using Caco-2 cells, HEK cells transfected with SR-BI, mouse intestinal explants, and mice overexpressing SR-BI in the intestine [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF]. SR-BI is thus involved in the uptake of a broad variety of molecules (cholesterol ester, carotenoids, vitamins E and D, among others), but not in the uptake of micellar retinol [START_REF] During | Mechanisms of provitamin A (carotenoid) and vitamin A (retinol) transport into and out of intestinal Caco-2 cells[END_REF]. This raises questions on the mechanisms of interaction between the transporter and its ligands. We hypothesize that SR-BI interacts with the mixed micelles and facilitates the specific uptake of certain lipid molecules solubilized within them. Obviously, dedicated experiments should be performed to verify this hypothesis. Using both mouse models and human cell lines, it has recently been shown that intestinal lipid absorption by SR-BI is subject to control by retinoid signaling [START_REF] Choi | A dynamic expression survey identifies transcription factors relevant in mouse digestive tract development[END_REF][START_REF] Seino | Isx participates in the maintenance of vitamin A metabolism by regulation of beta-carotene 15,15'monooxygenase (Bcmo1) expression[END_REF][START_REF] Lobo | ISX is a retinoic acidsensitive gatekeeper that controls intestinal beta,beta-carotene absorption and vitamin A production[END_REF]. Indeed, retinoic acid induces the expression of the intestinal transcription factor ISX which represses the expression of both SR-BI and BCMO1 (carotenoid-15,15'-oxygenase). BCMO1 acts downstream of SR-BI and converts absorbed β-carotene to the retinoic acid precursor, retinaldehyde. Thus, there is a diet-responsive regulatory network that potentially controls SR-BI-mediated FSV&C absorption via a negative feedback regulation. We say "potentially" because it has been shown that ISX null mice have normal blood cholesterol levels despite intestinal overexpression of SR-BI [START_REF] Seino | Isx participates in the maintenance of vitamin A metabolism by regulation of beta-carotene 15,15'monooxygenase (Bcmo1) expression[END_REF].

Cluster Determinant 36 (CD36)

Another scavenger receptor of interest is FAT (Fatty Acid Translocase, also known as CD36 in humans). This trans-membrane glycoprotein of 90 kDa is mainly expressed in epithelial cells, adipocytes, platelets and macrophages [START_REF] Terpstra | Hepatic and extrahepatic scavenger receptors: function in relation to disease[END_REF]. CD36 is also expressed at the BBM level of the duodenum and the jejunum [START_REF] Dew | Specificity of the retinol transporter of the rat small intestine brush border[END_REF]. Like SR-BI, it can interact with a broad variety of ligands such as HDL, LDL, VLDL, anionic phospholipids, fatty acids, free cholesterol, thrombospondin, collagen, erythrocytes infected by P falciparum, and apoptotic cells [START_REF] Terpstra | Hepatic and extrahepatic scavenger receptors: function in relation to disease[END_REF]. CD36 appears to be involved in numerous cellular functions such as cellular adhesion [START_REF] Tandon | Identification of glycoprotein IV (CD36) as a primary receptor for platelet-collagen adhesion[END_REF], angiogenesis [START_REF] Dawson | CD36 mediates the In vitro inhibitory effects of thrombospondin-1 on endothelial cells[END_REF], lipoprotein ligation/endocytosis [START_REF] Endemann | CD36 is a receptor for oxidized low density lipoprotein[END_REF] or fat sensing in lingual papilla [START_REF] Laugerette | CD36 involvement in orosensory detection of dietary lipids, spontaneous fat preference, and digestive secretions[END_REF]. It is assumed to play a key role in fatty acid uptake in muscle and adipose tissue [START_REF] Coburn | Defective uptake and utilization of long chain fatty acids in muscle and adipose tissues of CD36 knockout mice[END_REF], as well as in the intestine [START_REF] Lobo | Localization of the lipid receptors CD36 and CLA-1/SR-BI in the human gastrointestinal tract: towards the identification of receptors mediating the intestinal absorption of dietary lipids[END_REF]. The first result on knock-out mice suggested that CD36 did not play a role in intestinal lipid absorption after a lipid load [START_REF] Goudriaan | Intestinal lipid absorption is not affected in CD36 deficient mice[END_REF].

Nevertheless, a following study showed that lipid secretion in the lymph was decreased in CD36-deficient mice, but this phenomenon was actually hidden by the fact that CD36 can also regulate chylomicron catabolism [START_REF] Drover | CD36 deficiency impairs intestinal lipid secretion and clearance of chylomicrons from the blood[END_REF]. It was shown that in the enterocyte, CD36 probably allows the routing of the long-chain fatty acids to the endoplasmic reticulum for chylomicron assembly. The underlying mechanism is unknown, but it may be linked to the intracellular traffic of the protein between the plasma membrane and the organites.

Interestingly, CD36 colocalizes with other proteins involved in cholesterol transport, such as caveolin-1 in lipid rafts [START_REF] Ring | Caveolin-1 is required for fatty acid translocase (FAT/CD36) localization and function at the plasma membrane of mouse embryonic fibroblasts[END_REF] which has been identified as playing a role in lipid endocytosis [START_REF] Field | Caveolin is present in intestinal cells: role in cholesterol trafficking?[END_REF]. It is thus possible that the two proteins cooperate for lipid transport.

Regarding FSV&C, CD36 was first shown to be involved in -carotene uptake using transfected COS-7 cells and mouse BBM vesicles [START_REF] Van Bennekum | Class B scavenger receptor-mediated intestinal absorption of dietary beta-carotene and cholesterol[END_REF]. This result was further supported by the fact that a CD36-related protein is involved in selective carotenoid transport in Bombyx mori [START_REF] Sakudoh | A CD36related transmembrane protein is coordinated with an intracellular lipid-binding protein in selective carotenoid transport for cocoon coloration[END_REF]. Moreover, CD36 has been shown to be involved in both lycopene and lutein uptake in mouse 3T3-L1 adipocytes and in mouse adipose tissue cultures [START_REF] Moussa | CD36 is involved in lycopene and lutein uptake by adipocytes and adipose tissue cultures[END_REF]. Recently, CD36 was also shown to mediate cholecalciferol uptake in human transfected HEK cells as well as in mouse internal explants [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF].

As discussed for SR-BI, the fact that CD36 is involved in the uptake of very different substrates raises the question of the mechanism involved. Because CD36 colocalizes with other proteins, such as caveolin-1, in lipid rafts, it is possible that the modulation of its activity/expression indirectly modulates the activity of these proteins. If these proteins are involved in FSV&C uptake, FSV&C absorption may subsequently be affected.

Niemann-Pick C1-like 1 (NPC1L1)

Among the cholesterol membrane transporters, one of the last to be identified was NPC1L1.

NPC1L1 is a 135 kDa protein widely expressed in human tissues, especially in the liver and at the plasma membrane of the intestinal cell [START_REF] Davies | Evidence for a Niemann-Pick C (NPC) gene family: identification and characterization of NPC1L1[END_REF][START_REF] Altmann | Niemann-Pick C1 Like 1 protein is critical for intestinal cholesterol absorption[END_REF][START_REF] Davies | Inactivation of NPC1L1 causes multiple lipid transport defects and protects against diet-induced hypercholesterolemia[END_REF][START_REF] Garcia-Calvo | The target of ezetimibe is Niemann-Pick C1-Like 1 (NPC1L1)[END_REF], as well as in Caco-2 cells [START_REF] Duval | Niemann-Pick C1 like 1 gene expression is down-regulated by LXR activators in the intestine[END_REF].

NPC1L1 has been described as the main cholesterol and phytosterol transporter in the intestine [START_REF] Davis | Niemann-Pick C1 Like 1 (NPC1L1) is the intestinal phytosterol and cholesterol transporter and a key modulator of whole-body cholesterol homeostasis[END_REF]. Indeed, it is thought to transport about 60% of dietary cholesterol according to results obtained in knock-out mice [START_REF] Altmann | Niemann-Pick C1 Like 1 protein is critical for intestinal cholesterol absorption[END_REF][START_REF] Davies | Inactivation of NPC1L1 causes multiple lipid transport defects and protects against diet-induced hypercholesterolemia[END_REF][START_REF] Davis | Niemann-Pick C1 Like 1 (NPC1L1) is the intestinal phytosterol and cholesterol transporter and a key modulator of whole-body cholesterol homeostasis[END_REF]. The phenotype of these mice corresponds to the phenotype of mice treated with ezetimibean inhibitor of cholesterol absorption [START_REF] Altmann | Niemann-Pick C1 Like 1 protein is critical for intestinal cholesterol absorption[END_REF][START_REF] Davis | Niemann-Pick C1 Like 1 (NPC1L1) is the intestinal phytosterol and cholesterol transporter and a key modulator of whole-body cholesterol homeostasis[END_REF]. It was first suggested that the target of ezetimibe was the annexin-2/caveolin-1 complex [START_REF] Smart | Annexin 2-caveolin 1 complex is a target of ezetimibe and regulates intestinal cholesterol transport[END_REF] or CD13 [START_REF] Kramer | Aminopeptidase N (CD13) is a molecular target of the cholesterol absorption inhibitor ezetimibe in the enterocyte brush border membrane[END_REF]. However, caveolin-1-deficient mice display no special phenotype for cholesterol absorption [START_REF] Valasek | Caveolin-1 is not required for murine intestinal cholesterol transport[END_REF], and there have been no further investigations on CD13. Another study elegantly showed that ezetimibe binds specifically to NPC1L1 [START_REF] Garcia-Calvo | The target of ezetimibe is Niemann-Pick C1-Like 1 (NPC1L1)[END_REF]. Recently, it has been demonstrated that NPC1L1 was recycled between the plasma membrane and the intracellular compartments via an endocytotic pathway regulated by cellular cholesterol. Cholesterol depletion would induce a relocalization of the protein at the cell surface and thus increase cholesterol uptake [START_REF] Yu | Cholesterol-regulated translocation of NPC1L1 to the cell surface facilitates free cholesterol uptake[END_REF]. NPC1L1 was further shown to be involved in both -tocopherol [START_REF] Narushima | Niemann-pick C1-like 1 mediates alpha-tocopherol transport[END_REF] and -tocotrienol [START_REF] Abuasal | Intestinal absorption of gamma-tocotrienol is mediated by Niemann-Pick C1-like 1: in situ rat intestinal perfusion studies[END_REF] intestinal absorption using Caco-2 cells and in situ perfusions in rats. Results were less clear on carotenoids [START_REF] Moussa | Lycopene absorption in human intestinal cells and in mice involves scavenger receptor class B type I but not Niemann-Pick C1-like 1[END_REF][START_REF] During | Carotenoid transport is decreased and expression of the lipid transporters SR-BI, NPC1L1, and ABCA1 is downregulated in Caco-2 cells treated with ezetimibe[END_REF]. In the first study, ezetimibe was shown to inhibit and -carotene uptake by 50%, -cryptoxanthin and lycopene uptake by 20%, and lutein and zeaxanthin uptake by 7% in Caco-2 cells. In the second study focusing on lycopene, neither ezetimibe nor a blocking antibody raised against NPC1L1 impaired its absorption in these cells.

Additional experiments are thus needed to clarify these discrepancies. Finally, it was recently shown using in vitro, ex vivo and in vivo mouse models that NPC1L1 is also involved, in a moderate way, in cholecalciferol intestinal uptake [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF].

LIPID TRANSPORTERS AND FAT-SOLUBLE VITAMIN & CAROTENOID APICAL EFFLUX

The enterocyte is not a simple entrance for nutrients into the body. Numerous proteins located at the apical side of the cells can efflux molecules back to the lumen to regulate or inhibit their absorption. For example, SR-BI's role is not limited to uptake, as it can function in both directions in the intestine [START_REF] Cai | Scavenger receptor class B type I reduces cholesterol absorption in cultured enterocyte CaCo-2 cells[END_REF] and other tissues [START_REF] Krieger | Charting the fate of the "good cholesterol": identification and characterization of the high-density lipoprotein receptor SR-BI[END_REF]. In particular, it was shown to be involved in both vitamin D [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF] and E [START_REF] Reboul | Scavenger receptor class B type I (SR-BI) is involved in vitamin E transport across the enterocyte[END_REF] efflux across the BBM using Caco-2 cells. Pre-charged FSV&C cells were able to efflux the vitamin back to the apical medium, especially when mixed micelles were added as acceptors, and this efflux was significantly inhibited in presence of SR-BI inhibitors (the chemical inhibitor Block-Lipid Transport 1 and/or blocking antibody).

It is very likely that other transporters can act as efflux pumps of FSV&C at the BBM level.

For example, the ATP-Binding Cassette (ABC) transporters are a superfamily of transmembrane proteins able to transport a wide variety of substrates, including drugs, lipids, bile salts, amino acids, peptides, proteins and carbohydrates. Most of these transporters are efflux proteins [START_REF] Dean | The human ATP-binding cassette (ABC) transporter superfamily[END_REF] requiring energy in the form of ATP to translocate their substrates [START_REF] Moody | Cooperative, ATP-dependent association of the nucleotide binding cassettes during the catalytic cycle of ATP-binding cassette transporters[END_REF][START_REF] Higgins | The ATP switch model for ABC transporters[END_REF]. ABC transporters are classified into 7 subfamilies (A to G) depending on their structure and homology, and on whether they are "complete" transporters such as ABCA1 or half-transporters associated in homo-or heterodimers, such as ABCG1 or ABCG5/G8 [START_REF] Dean | The human ATP-binding cassette (ABC) transporter superfamily[END_REF].

In particular, the ABCG5/G8 heterodimer is critical to sterol homeostasis. Mutations in the genes encoding these proteins cause -sitosterolemia [START_REF] Lee | Genetic basis of sitosterolemia[END_REF], a disease characterized by abnormally high levels of phytosterols in blood and tissues due to increased intestinal absorption and decreased secretion through the bile. The ABCG5/G8 complex acts like an efflux pump in the enterocyte where it limits mainly phytosterol absorption. Interestingly, the ABCG5/G8 complex can also transport cholesterol [START_REF] Yu | Overexpression of ABCG5 and ABCG8 promotes biliary cholesterol secretion and reduces fractional absorption of dietary cholesterol[END_REF]. However, there is only one study suggesting a role of these transporters in the absorption of a FSV&C: lutein [START_REF] Herron | The ABCG5 polymorphism contributes to individual responses to dietary cholesterol and carotenoids in eggs[END_REF].

This study suggested that ABCG5 plays a role in the plasma response to dietary carotenoids.

Finally, it has been suggested that ABCA1 is expressed at the apical surface of the enterocyte [START_REF] Repa | Regulation of absorption and ABC1-mediated efflux of cholesterol by RXR heterodimers[END_REF] and plays a role in cholesterol and phytosterol apical efflux [START_REF] Plat | Increased intestinal ABCA1 expression contributes to the decrease in cholesterol absorption after plant stanol consumption[END_REF][START_REF] Tachibana | Cholesterol and plant sterol efflux from cultured intestinal epithelial cells is mediated by ATP-binding cassette transporters[END_REF].

However, there is no data on the potential involvement of this transporter in apical efflux of FSV&C, and numerous studies describe ABCA1 as expressed at the basolateral side of the intestinal cell [START_REF] Murthy | LXR/RXR activation enhances basolateral efflux of cholesterol in CaCo-2 cells[END_REF] where it is implicated in lipid secretion via the portal route [START_REF] Murthy | LXR/RXR activation enhances basolateral efflux of cholesterol in CaCo-2 cells[END_REF][START_REF] Brunham | Intestinal ABCA1 directly contributes to HDL biogenesis in vivo[END_REF][START_REF] Field | LXR/RXR ligand activation enhances basolateral efflux of beta-sitosterol in CaCo-2 cells[END_REF] (see section 6.1).

In conclusion, it appears that several lipid transporters are involved in FSV&C uptake/efflux at the apical membrane of the enterocyte. As these lipid transporters are involved in the uptake of both cholesterol and different FSV&C, this likely explains the competition between these molecules for apical uptake (e.g. between lutein and carotenoids or vitamin E, and vice versa [START_REF] Reboul | Effect of the main dietary antioxidants (carotenoids, gamma-tocopherol, polyphenols, and vitamin C) on alphatocopherol absorption[END_REF][START_REF] Reboul | Differential effect of dietary antioxidant classes (carotenoids, polyphenols, vitamin C and vitamin E) on lutein absorption[END_REF], between and -tocopherol or cholesterol [START_REF] Reboul | Scavenger receptor class B type I (SR-BI) is involved in vitamin E transport across the enterocyte[END_REF], or between vitamin D and vitamin E or cholesterol [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF]). The relative contribution of the lipid transporters to the net uptake of each FSV&C is impossible to clarify at the present time due to the incomplete data available. Furthermore, it is likely that not all the transporters involved have yet been identified.

TRANSPORT OF FAT-SOLUBLE VITAMINS & CAROTENOIDS ACROSS THE ENTEROCYTE

FSV&C are metabolized differently by the enterocyte. Indeed, while tocopherols and cholecalciferol are theoretically incorporated into the chylomicrons without any chemical modification [START_REF] Drevon | absorption, transport and metabolism of vitamin E[END_REF][START_REF] Haddad | Human plasma transport of vitamin D after its endogenous synthesis[END_REF], retinol is first esterified in retinyl esters by LRAT and ARAT [START_REF] Harrison | Mechanisms of digestion and absorption of dietary vitamin A[END_REF] and carotenoids are partly cleaved in retinal and apocarotenoids by BCMO1 and BCDO2 [START_REF] Lobo | Carotenoid-oxygenases: Key players for carotenoid function and homeostasis in mammalian biology[END_REF], respectively. Finally, concerning phylloquinone, some authors have suggested that menadione could be produced by the intestine after an oral dose of phylloquinone [START_REF] Thijssen | Menadione is a metabolite of oral vitamin K[END_REF][START_REF] Okano | In vivo metabolism of vitamin K: in relation to the conversion of vitamin K1 to MK-4[END_REF].

However, this hypothesis remains to be further investigated. The intracellular transport of most of these molecules in the enterocyte is a black box. It is assumed that, being nonsoluble in water, these compounds require either intracellular proteins to cross the water compartments of the cells or incorporation in intracellular cell membranes to be transferred to the basolateral side of the enterocyte. The first candidates for this intracellular transport are the apical membrane transporters displaying an intracellular cycle between the apical membrane and the cellular organelles. For example, NPC1L1, which is implicated in vitamin E and D absorption, has been observed in endosomes, perinuclear regions, lysosomes and mitochondria of the human intestinal cell [START_REF] Yu | Cholesterol-regulated translocation of NPC1L1 to the cell surface facilitates free cholesterol uptake[END_REF][START_REF] Sane | Localization and role of NPC1L1 in cholesterol absorption in human intestine[END_REF]. Another candidate is CD36, which has been suggested to play a role in apical uptake of β-carotene and vitamin D. Indeed, this scavenger receptor has been detected in both the apical membrane and the Golgi apparatus [START_REF] Pohl | FAT/CD36-mediated long-chain fatty acid uptake in adipocytes requires plasma membrane rafts[END_REF]. Finally, SR-BI, which is involved in apical uptake of several FSV&C, has been found in apical and basolateral membranes of enterocytes, in cytoplasmic lipid droplets and in tubulovesicular membranes. Furthermore, SR-BI is mainly localized in the microvillus membrane in the fasting state and seems to be endocytosed during absorption of dietary fat [START_REF] Hansen | Scavenger receptor class B type I (SR-BI) in pig enterocytes: trafficking from the brush border to lipid droplets during fat absorption[END_REF]. We hypothesize that some FSV&C might stick to these transporters, or to membrane microdomains containing these transporters, at the apical side of the enterocyte and follow them within the cell to be transferred to either other intracellular transporters or to intracellular membranes.

The fact that apical membrane transporters can participate in the intracellular transport of FSV&C obviously does not exclude the possibility that proteins dedicated to the intracellular transport of these compounds are also involved. This is supported by the fact that proteins involved in intracellular transport of some FSV&C have been identified in several cell types, as well as in enterocytes. When such proteins from other tissues are also expressed in the enterocyte, it is tempting to hypothesize that they play the same role in the enterocyte, but this must be confirmed in dedicated studies.

SPECIFIC INTRACELLULAR TRANSPORTERS

Vitamin A

Intracellular transporters of vitamin A (retinol and its metabolites: retinal and retinoic acid) were discovered many years ago. These proteins are called retinoid-binding proteins [START_REF] Noy | Retinoid-binding proteins: mediators of retinoid action[END_REF][START_REF] Napoli | Interactions of retinoid binding proteins and enzymes in retinoid metabolism[END_REF]. They are expressed in numerous tissues and are involved in both intracellular transport and targeting of the essential vitamin A metabolite to the appropriate site in the cell, e.g. retinoic acid to the nucleus or retinal to opsin. A retinoid-binding protein of special interest for vitamin A transport within the enterocyte is CRBPII (Cellular Retinol-Binding Protein II). CRBPII is primarily expressed in the intestinal mucosa, and it is one of the most abundant soluble proteins in the jejunum, representing 0.4-1.0% of the total enterocyte cytosolic proteins [START_REF] Crow | Cell-specific immunohistochemical localization of a cellular retinolbinding protein (type two) in the small intestine of rat[END_REF]. The hypothesis that this protein is important for the intracellular transport of retinol in the enterocyte was first prompted by the finding that its mRNA expression increased in the small intestine of rats under a retinoid-deficient diet [START_REF] Suruga | Unsaturated fatty acids regulate gene expression of cellular retinol-binding protein, type II in rat jejunum[END_REF].

Further investigations, using CRBPII-deficient mice, have definitively demonstrated that CRBPII plays an important role in vitamin A metabolism in the intestine [START_REF] Zhang | Increased neonatal mortality in mice lacking cellular retinol-binding protein II[END_REF][START_REF] Wongsiriroj | The molecular basis of retinoid absorption: a genetic dissection[END_REF]. The involvement of another CRBP, CRBPI [START_REF] Uchio | Cellular retinol-binding protein-1 expression and modulation during in vivo and in vitro myofibroblastic differentiation of rat hepatic stellate cells and portal fibroblasts[END_REF], cannot be ruled out as this protein is also found in the enterocyte, but there is no evidence that it plays an important role in vitamin A metabolism in this cell type. Concerning carotenoids, a carotenoid-binding protein (CBP) assumed to transport carotenoids in the cytosol has been described in the midgut of the silkworm Bombyx mori [START_REF] Sakudoh | A CD36related transmembrane protein is coordinated with an intracellular lipid-binding protein in selective carotenoid transport for cocoon coloration[END_REF][START_REF] Tabunoki | Isolation, characterization, and cDNA sequence of a carotenoid binding protein from the silk gland of Bombyx mori larvae[END_REF]. Furthermore, two lutein-binding proteins have been described in human retina: GSTP1 (Glutathione S-Transferase Pi 1) [START_REF] Bhosale | Identification and characterization of a Pi isoform of glutathione S-transferase (GSTP1) as a zeaxanthin-binding protein in the macula of the human eye[END_REF] and HR-LBP (Human Retinal Lutein-Binding Protein) [START_REF] Bhosale | Purification and partial characterization of a lutein-binding protein from human retina[END_REF] that shows good cross-reactivity with antibodies raised against Bombyx mori silkworm CBP. If expressed in human enterocytes, LBP would be a good candidate as intracellular transporter of xanthophylls.

Vitamin E

In humans, a specific intracellular protein that binds tocopherol, the α-tocopherol-transfer protein (α-TTP), is mainly expressed in the liver. Mutations in the gene encoding this protein can disrupt tocopherol transfer from the liver into plasma lipoproteins, resulting in a clinical syndrome called Ataxia with isolated Vitamin E deficiency. Although a specific binding protein is needed for α-tocopherol transfer onto nascent hepatic VLDL [START_REF] Traber | Vitamin E trafficking[END_REF], it is assumed that other vitamin E-binding proteins are involved in the intracellular transport of vitamin E in various tissues [START_REF] Stocker | Tocopherol-binding proteins: their function and physiological significance[END_REF]. A vitamin E-binding protein called TAP (tocopherolassociated protein) has been identified in both bovine [START_REF] Stocker | Identification of a novel cytosolic tocopherolbinding protein: Structure, specificity, and tissue distribution[END_REF] and human tissues [START_REF] Zimmer | A novel human tocopherol-associated protein -Cloning, in vitro expression, and characterization[END_REF]. This protein belongs to a family of hydrophobic ligand-binding proteins that share the same cisretinal binding motif sequence. Northern analyses showed that TAP mRNA is ubiquitously expressed. Thus, TAP could be a good candidate for intracellular transport of vitamin E within the enterocyte, although its expression in intestinal cells has apparently not been measured [START_REF] Stocker | Identification of a novel cytosolic tocopherolbinding protein: Structure, specificity, and tissue distribution[END_REF]. Other candidates could be the sec14p-like proteins (hTAP1, 2 and 3), which are ubiquitously detectable in several tissues and which enhance the in vitro transport of α-tocopherol to mitochondria at the same rate as α-TTP [START_REF] Zingg | Characterization of three human sec14p-like proteins: alpha-tocopherol transport activity and expression pattern in tissues[END_REF]. Since there is no discrimination between the various forms of vitamin E, e.g. α, β, γ, δ tocopherols and tocotrienols, in terms of their absorption, we suggest that if a vitamin E-binding protein is implicated in intracellular transport of vitamin E within the enterocyte, it is a protein that has no specificity for RRR-α-tocopherol, conversely to α-TTP.

Finally, the fact that the basolateral membrane transporter ABCA1 is implicated in basolateral efflux of a fraction of vitamin E in apo-AI lipoparticles (see section 5) prompts the question "Is there one specific intracellular vitamin E-binding protein that channels vitamin E to the site where it is incorporated into chylomicrons, and another that channels vitamin E to the site where it is secreted in intestinal HDL via the ABCA1 transporter"?

Vitamin D

Apart from VDR, the nuclear receptor involved in the well-known effect of vitamin D on gene expression, the first intracellular vitamin D-binding protein (CDBPcytosol vitamin D-binding protein) immunologically related to DBP that is responsible for vitamin D transport in the blood was found in rat kidney [START_REF] Imawari | Synthesis of serum and cytosol vitamin D-binding proteins by rat liver and kidney[END_REF]. Fifteen years later, an intracellular vitamin D binding protein (called IDBP) was identified in new-world primate cell extracts [START_REF] Gacad | Proteins in the heat shock-70 family specifically bind 25hydroxyvitamin D3 and 17beta-estradiol[END_REF][START_REF] Gacad | Functional characterization and purification of an intracellular vitamin D-binding protein in vitamin D-resistant new world primate cells. Amino acid sequence homology with proteins in the hsp-70 family[END_REF]. It is not known whether CDBP and IDBP are the same protein. IDBP bind s25OHD and is putatively responsible for translocation of 25OHD to the vitamin D 1-and 24-hydroxylase enzymes located in the inner mitochondrial membrane. In humans, two IDBP (IDBP-1 and 2), which were related to the IDBP identified in new-world primates, have been cloned and expressed. [START_REF] Wu | Intracellular vitamin D binding proteins: novel facilitators of vitamin D-directed transactivation[END_REF]. Finally, in rat enterocytes, an intracellular binding protein with distinct characteristics from primate IDBP has been characterized [START_REF] Teegarden | Characterization of 25-hydroxyvitamin D binding protein from intestinal cells[END_REF]. This protein is named cytosolic vitamin D metabolite binding protein (cDBP), and because it preferentially binds 25OHD, it has been suggested that it would transport 25OHD across the enterocyte via the chylomicron-independent route [START_REF] Sitrin | Comparison of vitamin D and 25hydroxyvitamin D absorption in the rat[END_REF]. However, vitamin D is mainly ingested as cholecalciferol which is incorporated into chylomicrons, and there is no data on the expression of cDBP or other IDBP in human enterocytes or on their putative role in cholecalciferol transport across the enterocyte.

Vitamin K

Only one study has suggested the existence of an intracellular-specific vitamin K-binding protein, which was described in the nucleus of osteoblasts. It binds MK-4 (menatetrenone), which is produced via conversion of phylloquinone in the body [START_REF] Ronden | Intestinal flora is not an intermediate in the phylloquinone-menaquinone-4 conversion in the rat[END_REF], and phylloquinone and its molecular structure is close to human GAPDH [START_REF] Hoshi | Nuclear vitamin K2 binding protein in human osteoblasts: homologue to glyceraldehyde-3-phosphate dehydrogenase[END_REF]. There is too little relevant data to allow any identification of specific candidate protein(s) for the intracellular transport of vitamin K across the human enterocyte. In summary, although several candidate binding proteins exist for most FSV&C, only retinol presents an identified specific intracellular protein (CRBPII) which has been involved in its transport across the human enterocyte. Dedicated studies are therefore required to investigate this important aspect of FSV&C metabolism.

NON-SPECIFIC INTRACELLULAR TRANSPORTERS: THE FATTY ACID-BINDING PROTEINS

The hypothesis that fatty acid-binding proteins (FABPs) can participate in the intracellular transport of FSV&C comes from studies by Hollander's group. This team was the first to suggest, from results showing interactions between phylloquinone and linoleic acid, that a FABP might also be a carrier for phylloquinone [START_REF] Hollander | Vitamin K1 intestinal absorption in vivo: influence of luminal contents on transport[END_REF]. The fact that FABPs are also able to bind different hydrophobic molecules [START_REF] Velkov | Characterization of lipophilic drug binding to rat intestinal fatty acid binding protein[END_REF][START_REF] Chuang | Characterization of the drug binding specificity of rat liver fatty acid binding protein[END_REF] supports this hypothesis, and suggests that enterocyte FABPs might be involved in intracellular transport of some FSV&C. Two FABPs are co-expressed in the human enterocyte: intestinal FABP (IFABP) and liver FABP (LFABP) [START_REF] Besnard | New insights into the fatty acidbinding protein (FABP) family in the small intestine[END_REF]. Both exhibit high-affinity binding of long-chain fatty acids [START_REF] Hanhoff | Insights into binding of fatty acids by fatty acid binding proteins[END_REF]. Utilizing mice genetically lacking either IFABP or LFABP, it has been demonstrated that each of the enterocyte FABPs participates in specific pathways of intestinal lipid metabolism [START_REF] Lagakos | Different functions of intestinal-and liver-type fatty acid-binding proteins in intestine and in whole body energy homeostasis[END_REF]. In particular, LFABP appears to target fatty acids toward oxidative pathways and dietary monoacylglycerols toward anabolic pathways, while IFABP targets dietary fatty acids toward triacylglycerol synthesis. It is thus hypothesized that the enterocyte FABPs allow a specific trafficking of their ligands to their respective metabolic fates. Although dedicated studies should be performed to verify whether IFABP and/or LFABP are involved in FSV&C metabolism, the fact that a genetic association study found that a genetic variant in IFABP was associated to fasting plasma lycopene concentrations [START_REF] Borel | Human fasting plasma concentrations of vitamin E and carotenoids, and their association with genetic variants in apo C-III, cholesteryl ester transfer protein, hepatic lipase, intestinal fatty acid binding protein and microsomal triacylglycerol transfer protein[END_REF] can be considered as a first argument that the FABPs could well be involved in intracellular transport of some FSV&C.

BASOLATERAL SECRETION OF FAT-SOLUBLE VITAMINS &

CAROTENOIDS

It is assumed that most of the newly-absorbed FSV&C are incorporated into chylomicrons that are secreted into the lymph via an apoB-dependent route [START_REF] Hussain | Intestinal lipoprotein assembly[END_REF]. Vitamin A is assumed to be recovered as retinyl esters [START_REF] Harrison | Mechanisms of digestion and absorption of dietary vitamin A[END_REF][START_REF] Sauvant | Amounts and types of fatty acids in meals affect the pattern of retinoids secreted in human chylomicrons after a high-dose preformed vitamin A intake[END_REF] and as non-cleaved provitamin A carotenoids.

The other fat-soluble vitamins and the non-provitamin-A carotenoids are assumed to be recovered as non-esterified molecules. However, a growing body of evidence now suggests that the intestine secretes important amounts of HDL during the postprandial period. Indeed, it has been shown in differentiated Caco-2 cells and in rat enterocytes that free cholesterol was secreted at the basoteral side by two distinct routes: an apolipoprotein B-dependent pathway (chylomicrons) and a non-apo-B-dependent pathway. Conversely, cholesterol esters were exclusively secreted into chylomicrons. As this non-apo-B pathway was i) inhibited by glyburide and ii) increased by agonists of the nuclear receptors LXR/RXR (liver X receptor/retinoid X receptor), it was assumed to be an ABCA1/HDL-dependent pathway [START_REF] Hussain | Intestinal lipoprotein assembly[END_REF]. Results from animal studies led to the conclusion that intestinal ABCA1 would contribute to approximately 30% of steady-state plasma cholesterol levels [START_REF] Attie | ABCA1: at the nexus of cholesterol, HDL and atherosclerosis[END_REF].

ABCA1 is a protein of 240 kDa, and its expression during embryonic development has been correlated to apoptosis areas. Defects in the ABCA1 gene were subsequently shown to be the cause of Tangier disease, which is characterized by a lack, or abnormally low levels of HDL and a markedly increased risk of coronary artery diseases. Studies using knock-out mice have confirmed that ABCA1 plays a pivotal role in reverse cholesterol transport.

Increased expression of ABCA1 in a number of cell types results in elevated net efflux of cellular cholesterol and phospholipids. This efflux is dependent on the presence of an acceptor such as lipid-poor HDL or apolipoprotein AI (Apo AI) [START_REF] Oram | ATP-binding cassette transporter A1 and cholesterol trafficking[END_REF]. The molecular functioning of ABCA1 remains unclear, but it has been suggested that the ligation/hydrolysis of ATP on the protein induces a change in the conformation of the transporter, which then binds to a lipid-poor Apo AI. Phospholipids then transfer from the cell membrane to Apo AI, which in turn induces cholesterol transfer [START_REF] Borst | Mammalian ABC transporters in health and disease[END_REF]. Proteins other than Apo AI may also act as acceptors, although their physiological relevance is presently unclear. In addition to cholesterol and phospholipids, ABCA1 has also been reported to promote secretion of -tocopherol, apoE and interleukin-1 at tissue level [START_REF] Oram | ATP-binding cassette transporter A1: a cell cholesterol exporter that protects against cardiovascular disease[END_REF]. ABCA1 is highly expressed in the liver and the intestine [START_REF] Baloch | Ontogenesis of the murine hepatic extracellular matrix -an immunohistochemical study[END_REF], especially at the basolateral side of the cell [START_REF] Murthy | LXR/RXR activation enhances basolateral efflux of cholesterol in CaCo-2 cells[END_REF], where it can play a role in intestinal HDL production and thus in basolateral cholesterol and phytosterol secretion [START_REF] Murthy | LXR/RXR activation enhances basolateral efflux of cholesterol in CaCo-2 cells[END_REF][START_REF] Brunham | Intestinal ABCA1 directly contributes to HDL biogenesis in vivo[END_REF][START_REF] Field | LXR/RXR ligand activation enhances basolateral efflux of beta-sitosterol in CaCo-2 cells[END_REF]. As ABCA1 controls HDL formation by the cellular efflux of a broad range of lipids [START_REF] Field | LXR/RXR ligand activation enhances basolateral efflux of beta-sitosterol in CaCo-2 cells[END_REF][START_REF] Wang | ATP-binding cassette transporter A1 (ABCA1) functions as a cholesterol efflux regulatory protein[END_REF][START_REF] Tam | ABCA1 mediates high-affinity uptake of 25-hydroxycholesterol by membrane vesicles and rapid efflux of oxysterol by intact cells[END_REF][START_REF] Panzenbock | Oxidation of methionine residues to methionine sulfoxides does not decrease potential antiatherogenic properties of apolipoprotein A-I[END_REF] and is associated with α-tocopherol secretion in macrophages [START_REF] Oram | ATP-binding cassette transporter A1 mediates cellular secretion of alpha-tocopherol[END_REF], it was not surprising that in vitro models showed that a fraction of α-tocopherol was also secreted with intestinal HDL by an ABC transporter [START_REF] Anwar | Transport of vitamin E by differentiated Caco-2 cells[END_REF][START_REF] Anwar | Mechanisms involved in vitamin E transport by primary enterocytes and in vivo absorption[END_REF]. The use of ABCA1-deficient mice definitively confirmed that intestinal ABCA1 is significantly involved in the intestinal absorption of vitamin E (α-and γ-tocopherol) but not vitamin A (retinyl palmitate) [START_REF] Reboul | ATP-binding cassette transporter A1 is significantly involved in the intestinal absorption of alpha-and gamma-tocopherol but not in that of retinyl palmitate in mice[END_REF]. It has also been suggested that free retinol efflux from intestinal cells was probably facilitated by ABCA1 [START_REF] During | Mechanisms of provitamin A (carotenoid) and vitamin A (retinol) transport into and out of intestinal Caco-2 cells[END_REF], but this result needs to be confirmed.

Additionally to ABCA1, other transporters may play a role in FSV&C basolateral efflux.

The first candidate would be ABCG1, which is also expressed in the intestinal cell [START_REF] Landrier | Vitamin E decreases endogenous cholesterol synthesis and apo-AI-mediated cholesterol secretion in Caco-2 cells[END_REF]. Indeed, its activity is sometimes difficult to differentiate from ABCA1 activity, although it is thought to efflux lipid to mature HDL and not to lipid-free apo AI [START_REF] Terasaka | ABCG1 and HDL protect against endothelial dysfunction in mice fed a high-cholesterol diet[END_REF]. Another example would be SR-BI, as it is also partly expressed at the basolateral membrane of the intestinal cell [START_REF] Cai | Differentiation-dependent expression and localization of the class B type I scavenger receptor in intestine[END_REF][START_REF] Reboul | Lutein transport by Caco-2 TC-7 cells occurs partly by a facilitated process involving the scavenger receptor class B type I (SR-BI)[END_REF] and is clearly involved in lipid and FSV&C efflux [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF][START_REF] Krieger | Charting the fate of the "good cholesterol": identification and characterization of the high-density lipoprotein receptor SR-BI[END_REF][START_REF] Reboul | Scavenger receptor class B type I (SR-BI) is involved in vitamin E transport across the enterocyte[END_REF][START_REF] Cai | Scavenger receptor class B type I reduces cholesterol absorption in cultured enterocyte CaCo-2 cells[END_REF][START_REF] Jian | Scavenger receptor class B type I as a mediator of cellular cholesterol efflux to lipoproteins and phospholipid acceptors[END_REF]. However, further studies are required to confirm the contributions of these transporters to intestinal FSV&C basoteral secretion.

HYPOTHESIS ON THE MODULATION OF FAT-SOLUBLE VITAMIN & CAROTENOIDS ABSORPTION VIA MODULATION OF THE EXPRESSION OR ACTIVITY OF PROTEINS INVOLVED IN FAT-SOLUBLE VITAMIN & CAROTENOID METABOLISM WITHIN THE ENTEROCYTE

The fact that intestinal proteins are involved in FSV&C absorption raises the question of the consequences of modulations in their expression or activity on FSV&C bioavailability. Two main factors modulate the expression and activity of intestinal proteins involved in nutrient absorption: 1) the nutrients absorbed via these proteins themselves (negative feedback regulation) and 2) genetic variations in the genes encoding these proteins.

The first nutrients that can likely modulate the expression of proteins involved in absorption of FSV&C are FSV&C themselves. To support this hypothesis, the expression of SR-BI, the transporter involved in apical uptake of several FSV&C, is modulated by both vitamin A [START_REF] Lobo | ISX is a retinoic acidsensitive gatekeeper that controls intestinal beta,beta-carotene absorption and vitamin A production[END_REF] and vitamin E [START_REF] Landes | Vitamin E activates gene expression via the pregnane X receptor[END_REF][START_REF] Sporstol | Pregnane X receptor-agonists down-regulate hepatic ATP-binding cassette transporter A1 and scavenger receptor class B type I[END_REF]. Moreover, vitamin E is also able to downregulate the expression of SR-BI post-transcriptionally [START_REF] Witt | Regulation by vitamin E of the scavenger receptor BI in rat liver and HepG2 cells[END_REF]. Vitamin E also downregulates CD36 [START_REF] Venugopal | RRR-alpha-tocopherol decreases the expression of the major scavenger receptor, CD36, in human macrophages via inhibition of tyrosine kinase (Tyk2)[END_REF][START_REF] Devaraj | Alpha-tocopherol decreases CD36 expression in human monocyte-derived macrophages[END_REF][START_REF] Ozer | Vitamin E inhibits CD36 scavenger receptor expression in hypercholesterolemic rabbits[END_REF], a transporter involved in intestinal uptake of β-carotene [START_REF] Van Bennekum | Class B scavenger receptor-mediated intestinal absorption of dietary beta-carotene and cholesterol[END_REF], and ABCA1 [START_REF] Landrier | Vitamin E decreases endogenous cholesterol synthesis and apo-AI-mediated cholesterol secretion in Caco-2 cells[END_REF][START_REF] Landes | Vitamin E activates gene expression via the pregnane X receptor[END_REF][START_REF] Sporstol | Pregnane X receptor-agonists down-regulate hepatic ATP-binding cassette transporter A1 and scavenger receptor class B type I[END_REF], a transporter involved in its basolateral secretion. These observations suggest that some FSV&C can modulate their own absorption by a negative feedback regulation, which further supports a key role of the intestinal proteins regulated by FSV&C in the absorption of these compounds. Because we have recently found that SR-BI, CD36 and NPC1-L1 are involved in intestinal uptake of vitamin D [START_REF] Reboul | Vitamin D intestinal absorption is not a simple passive diffusion: Evidences for involvement of cholesterol transporters[END_REF], and because this vitamin has a well-known effect on gene expression, it would be interesting to assess whether this vitamin modulates the expression of proteins involved in its own absorption.

The fact that several intestinal proteins involved in FSV&C absorption are also involved in fatty acid and cholesterol absorption makes it is reasonable to hypothesize that dietary fatty acids, or cholesterol, modulate the expression of these proteins. This is supported by the fact that NPC1L1, as well as the ATP-binding cassette proteins ABCA1, ABCG5 and ABCG8 are downregulated by a cholesterol-free high-fat diet [START_REF] De Vogel-Van Den Bosch | A cholesterol-free, high-fat diet suppresses gene expression of cholesterol transporters in murine small intestine[END_REF]. For other transporters, there is little data at the intestinal level, but it has been shown that heart CD36 and hepatic SR-BI expressions are regulated by dietary fat [START_REF] Greenwalt | Heart CD36 expression is increased in murine models of diabetes and in mice fed a high fat diet[END_REF][START_REF] Spady | Polyunsaturated fatty acids up-regulate hepatic scavenger receptor B1 (SR-BI) expression and HDL cholesteryl ester uptake in the hamster[END_REF]. We can thus suggest that the fat and cholesterol content in the diet may affect the absorption efficiency of FSV&C by modulating the expression of lipid transporters involved in their absorption. Finally, the expression of an intestinal protein assumed to be specific for the transport of a FSV&C, i.e. CRBPII that transport retinol, is modulated by diets containing long-chain fatty acids [START_REF] Suruga | Unsaturated fatty acids regulate gene expression of cellular retinol-binding protein, type II in rat jejunum[END_REF].

The hypothesis that genetic variants in genes encoding proteins involved in FSV&C absorption may affect FSV&C absorption comes from the observation that there is a huge inter-individual variability in FSV&C assimilation [START_REF] Borel | Low and high responders to pharmacological doses of beta-carotene: proportion in the population, mechanisms involved and consequences on beta-carotene metabolism[END_REF][START_REF] Jeanes | Comparative (2)H-labelled alpha-tocopherol biokinetics in plasma, lipoproteins, erythrocytes, platelets and lymphocytes in normolipidaemic males[END_REF], that the "responder characteristic" is apparently an intrinsic characteristic of the subjects, and that proteins are involved in FSV&C absorption. Indeed, the finding that intestinal proteins are involved in absorption of FSV&C leads to the hypothesis that genetic variations in their related genes may affect their expression (promoter region of the gene) or their activity (modification of amino acid sequence) and, in turn, their ability to absorb/transport these compounds. This exciting hypothesis needs to be verified in specific studies, but data on the associations between genetic variants SR-BI and CD36 and blood concentrations of carotenoids [START_REF] Borel | Human plasma levels of vitamin E and carotenoids are associated with genetic polymorphisms in genes involved in lipid metabolism[END_REF][START_REF] Borel | Genetic variants in BCMO1 and CD36 are associated with plasma lutein concentrations and macular pigment optical density in humans[END_REF] and between CD36 and vitamin E [START_REF] Lecompte | Polymorphisms in the CD36/FAT gene are associated with plasma vitamin E levels in humans[END_REF] support the proposal. Nevertheless, it should be stressed that these associations may be due to the effect of genetic variants on the expression or activity of the proteins in tissues other than the intestine. Indeed, these scavenger receptors are expressed in several tissues, and it is possible that the association with fasting blood concentration of FSV&C is also due to an effect of the genetic variants on uptake or efflux of carotenoids and vitamin E in other tissues.

If the hypothesis that genetic variants affect FSV&C absorption efficiency is verified, it would prompt the following questions: "do groups of subjects with low responder genotypes need higher dietary intakes of FSV&C?", and conversely "do groups of subjects with high responder genotypes need to avoid high intakes of some FSV&C due to the risk of toxic effects [START_REF] Rautalahti | Betacarotene did not work: Aftermath of the ATBC study[END_REF]?". Finally, because some intestinal proteins are involved in absorption of different FSV&C (e.g. SR-BI is involved in absorption of vitamin E), carotenoids and vitamin D, we can ask whether genetic variants can simultaneously affect the absorption efficiency of these different FSV&Cor, in other words, whether certain subjects could be at risk of micronutrient multideficiency.

PERSPECTIVES

Numerous gaps need to be filled before we can gain a full understanding of the uptake, intracellular transport and basolateral secretion of FSV&C by the enterocyte. Although some specific proteins such as CRBPII, and several non-specific proteins such as SR-BI, NPC1-L1, and ABCA1 have been identified, other transporters, (e.g. those involved in apical uptake of retinol and phylloquinone) remain to be identified. Another important point to unravel is how some proteins can be involved in the metabolism of FSV&C but display strikingly different chemical structures (SR-BI and CD36 are implicated in the metabolism of carotenoids, vitamin E and vitamin D). Finally, the discovery that proteins are involved in FSV&C absorption has not only opened a new field of research on the topic, but has also allowed the proposal of an explanation for the huge inter-individual variability observed in FSV&C absorption. Moreover, this discovery has led to the hypothesis that genetic variants in genes encoding these proteins may affect the absorption efficiency of these micronutrients, leading to suggestions that future Recommended Dietary Allowances of FSV&C may need to take these genetic variants into account. This would constitute a first step toward personalized nutrition based on the genetic characteristics of individuals.

Tables Table 1: Molecules belonging to the different fat-soluble vitamin groups

Vitamin group

Structures of the main molecules provided by the diet Main food sources (µg/ 100 g) [START_REF] Martin | Apports nutritionnels conseillés pour la population française[END_REF][START_REF] Reboul | Bioaccessibility of carotenoids and vitamin E from their main dietary sources[END_REF]

Table 3 :

 3 Proteins identified as involved in fat-soluble vitamin and carotenoid transport in the enterocyte Passive diffusion ; E = Unidentified basolateral efflux transporter ; ? = Putative pathway. Vitamins D and E, as well as carotenoids, are captured from mixed micelles by apical membrane transporters: SR-BI (Scavenger Receptor class B type I), CD36 (Cluster Determinant 36) and NPC1L1 (Niemann-Pick C1-Like 1). Apical membrane proteins involved in apical uptake of retinol and phylloquinone have not been identified yet. A fraction of the vitamins and carotenoids might then be effluxed back to the intestinal lumen via apical membrane transporters (SR-BI and possibly other transporters). Another fraction is transported to the site where they are incorporated into chylomicrons. It is hypothesized that proteins are involved in intracellular transport of fat-soluble vitamins and carotenoids, although only CRBPII (cellular retinol binding protein II) has clearly been identified. Non-metabolized vitamins and carotenoids are secreted in the lymph into chylomicrons, while a part of the more polar vitamins and carotenoid metabolites may be secreted via the portal route. It has been shown that vitamin E can also be secreted at the basolateral side via ABCA1 (apoAI pathway).