**2.2. Intracellular trafficking of vitamin E**

E deficiency has been linked to several disease states like ataxia with vitamin E deficiency (AVED) [2, 3] and Alzheimer's disease [4, 5], indicating a role in the preservation of human health. AVED has severe neurological consequences and is caused by a defect in the α-TOH transfer protein (α-TTP); the protein responsible for the discrimination of α-TOH from the other vitamin E forms in the liver [2, 3]. This emphasizes the role of the liver as a central organ in human vitamin E handling. The liver further distributes vitamin E in the body [6] and metabolizes excess vitamin E in order to form products for excretion [6] or presumably to produce activated metabolites of vitamin E as known for other lipophilic vitamins [7]. Given the crucial role of the liver for vitamin E handling, this review aims to summarize the knowledge on the physiological hepatic handling of vitamin E as well as on factors influencing hepatic

The liver is the central organ of vitamin E handling. While intestinal absorption efficiency is similar for all forms of vitamin E [8], the plasma concentrations of vitamin E forms differ a lot (e.g., 22.1 μM for α-TOH vs. 2.2 μM for γ-TOH [9]). The preference of α-TOH in the human

Vitamin E is absorbed in the intestine along with lipids (for details, see [8]) and is packed into lipoproteins. These are transported via lymph or blood toward the liver (via chylomicron remnants, low density lipoproteins (LDL), and high density lipoproteins (HDL) [10, 11]). Different mechanisms facilitate the cellular uptake of vitamin E: (i) via lipid transfer proteins or lipases, (ii) receptor-mediated lipoprotein endocytosis, and (iii) selective lipid uptake [12]. The degradation of chylomicrons to chylomicron remnants by lipoprotein lipase (LPL) seems to be highly important for vitamin E uptake in the liver; when lipolysis of triglyceride-rich chylomicrons by LPL is inhibited, the α-TOH uptake in the liver is diminished [13]. The phospholipid transfer protein (PLTP) mediates the exchange of phospholipids between lipoproteins [14] and is also able to bind α-TOH *in vitro* [15]. PLTP-null mice have lower hepatic levels of vitamin E than the wild-type mice [16]; hence, the transfer of vitamin E between the lipoproteins seems to be important for its effective hepatic uptake. The chylomicron remnants and LDL are taken up by the liver via endocytosis, mainly mediated through the LDL receptor (LDLR) or LDLR-related proteins [6, 17]. In addition, the cholesterol transporter Niemann-Pick C1-like 1 (NPC1L1) is involved in hepatic vitamin E uptake; α-TOH binds to the N-terminal domain of NPC1L1, which mediates α-TOH uptake via endocytosis (mechanism similar to intestinal cholesterol uptake) [18]. The scavenger receptor B type I (SR-BI) is known to mediate the uptake of vitamin E in several tissues (e.g., intestine [19], epithelium [20], and hepatocytes [21]) by channeling the molecules into the cells (shown for cholesterol or triglycerides [22]). Furthermore, the scavenger receptor cluster of differentiation 36 (CD36)

body is mediated by several complex and interacting hepatic mechanisms.

handling of vitamin E.

2 Vitamin E in Health and Disease

**2. Physiological hepatic handling of vitamin E**

**2.1. Hepatocellular uptake of vitamin E**

is likely involved in hepatic uptake of vitamin E [23].

Following its lipophilic nature, vitamin E is transported by intracellular carrier proteins [24]. The intestinally absorbed vitamin E is taken up via endocytosis [25] and follows endosomal fate. Here, the hepatic sorting of vitamin E forms starts as a specific protein, called α-TTP selectively recognizes and preferentially binds α-TOH, which is then extracted from endosomes and transported to the inner leaflet of the plasma membrane [26]. α-TTP is therefore considered to be a "gatekeeper", which discriminates non-α-TOH forms [27] and regulates the plasma concentrations of α-TOH [1]. The affinity of α-TTP to the different forms of vitamin E differs greatly: it is defined as 100% for α-TOH, whereas β-TOH has 38%, γ-TOH 9%, δ-TOH 2%, and α-tocotrienol (T3) 12% affinity to α-TTP [28]. The regular function of α-TTP is crucial, since missense mutations lead to the disruption of α-TOH distribution and the development of a severe degenerative disease, termed AVED [29]. The transfer of α-TOH from endosomes to the plasma membrane is a multi-step process. First, it is speculated whether the ATP-binding cassette transporter A1 (ABCA1) enriches the outer layer of endosomes with α-TOH [30]. The cholesterol transporter NPC1 may also be involved, as a genetic missense mutation of the *NPC1* gene leads to an accumulation of α-TOH in late endosomes [31]. Second, α-TTP extracts the α-TOH from endosomes, and third, α-TTP mediates its transport to the plasma membrane [24]. This process seems to depend on phosphatidylinositol phosphates (PIPs; preferentially PI(4,5)P<sup>2</sup> and PI(3,4)P<sup>2</sup> ) in the plasma membrane, as α-TTP binds to them, in turn targeting α-TOH to the plasma membrane and stimulating its release [32]. Chung et al. analyzed the localization of α-TTP depending on the cellular α-TOH concentration [33]. They found (i) perinuclear localization for α-TOH-depleted cells, (ii) a directional transport of α-TOH/α-TTP toward the plasma membrane, when depleted cells were pulsed with a low dose of α-TOH, and (iii) a homogenous cytosolic pattern under long-term and high-dose treatment of cells with α-TOH, which was suggested to be the picture of several α-TOH transport cycles [33]. Furthermore, the authors also postulated a bi-phasic concentration-dependent circulation of α-TTP: the PI(4,5)P<sup>2</sup> gradient (low in endosomes and high in plasma membrane) forces the α-TTP-mediated transport of α-TOH toward the plasma membrane, whereas the α-TOH gradient (low in plasma membrane and high in endosomes) triggers the recycling of α-TTP toward the endosomes [33]. It has been proposed that once α-TOH is incorporated into the plasma membrane, it is mediated toward the outer leaflet of the membrane by a flippase, maybe ABCA1, and is then available for the uptake via very low density lipoproteins (VLDL) [34]. For more details on the process, please see Section 2.5 "Release of vitamin E".

#### **2.3. Intracellular storage of vitamin E**

Intracellular storage of vitamin E is limited to the lipophilic sites of the cell, which are membranes and lipid droplets [33]. Not much is known about a specific localization of vitamin E accumulation in liver cells, apart from the observation that lysosomal membranes of rat livers seemed to have the highest concentration of all membranes [35–37]. However, it is known that one-third of the total body vitamin E is stored in the liver [38]. Within membranes, vitamin E is thought to stabilize the membrane bilayers due to colocalization with phosphatidylcholine [39] and cholesterol (leading to an association to lipid rafts) [40]. It was further hypothesized that vitamin E also colocalizes with poly-unsaturated fatty acids (PUFAs) in nonraft domains in order to provide protection from lipid peroxidation [41]. Newly added α-TOH in cell culture enriches in the same organelles as the endogenous α-TOH pool [42]. Hereby, the subcellular content of α-TOH was directly proportional to the lipid content [43].

chain. In line with this assumption is the identification of the respective unsaturated metabolites from 13′-carboxytrienol down to carboxymethylbutadienylhydroxychromanol (CMBenHC) in human and mouse samples [49]. According to these findings, the side chain of the T3 metabolites needs a saturation step before the shortening of the chain. Enzymes involved in the degradation of unsaturated fatty acids like 2,4-dienoyl-CoA reductase and 3,2-enoyl-CoA isomerase were

The Hepatic Fate of Vitamin E

5

http://dx.doi.org/10.5772/intechopen.79445

Following the nature of the lipoprotein metabolism, hepatic release of vitamin E is mostly realized via VLDL. Thus, this section will focus on the packaging of vitamin E into VLDL particles, notwithstanding that the mechanism is not well understood. However, hepatic transfer of vitamin E to HDL has also been suggested [56]. Since it was shown that the expression of α-TTP is crucial for the maintenance of plasma α-TOH levels [57, 58] and that the liver is controlling plasma α-TOH levels [59], hepatic α-TTP is likely involved in the incorporation of vitamin E into lipoproteins. This concept is supported by the observation that nascent VLDL particles are preferentially enriched with *RRR*-α-TOH after oral administration of vitamin E ([60, 61]. In contrast, in the liver, no preferential retention of *RRR*-α-TOH was found, indicating that α-TTP is not involved in the delivery of vitamin E to the liver, but in the release from the liver [62]. Hence, efforts have been made to identify the intracellular location of VLDL enrichment with α-TOH mediated by α-TTP [30]. According to the assembly of VLDL, either the rough ER or the Golgi apparatus were assumed. However, the action of α-TTP in these compartments was not confirmed as the nascent VLDL particles contained equal amounts of SRR and RRR α-TOH forms [30]. Further, the inhibition of ER/Golgi action in cells overexpressing α-TTP did not prevent α-TOH secretion [63]. In conclusion, α-TTP is necessary for the hepatic release of

vitamin E, but the enrichment of VLDL with *RRR*-α-TOH occurs after exocytosis.

Based on this, the hypothesis of α-TOH uptake by VLDL directly from the plasma membrane was developed. This idea was inspired by the proposed mechanism of the incorporation of free cholesterol into nascent VLDL [64], that is, the spontaneous transfer from membranes to lipoproteins [65]. The hypothesis involves also the α-TTP-mediated trafficking of vitamin E from late endosomes (where vitamin E occurs after cellular uptake and large parts of α-TTP are located [66]) to the plasma membrane. This process might involve ABCA1, which has been shown to transport α-TOH [67] and could thus present vitamin E to α-TTP at the outer leaflet of the endosomal membrane. After the transport to the plasma membrane, a yet unidentified flippase is required to transfer α-TOH to the appropriate site of the membrane for uptake by nascent VLDL [30]. This hypothesis is supported by findings of Chung et al. [33], which provided a model of α-TTP-facilitated trafficking of vitamin E from endosomes to the plasma membrane (the reader is referred to Section 2.2 "Intracellular trafficking of vitamin E"). Taken together, the release of α-TOH from hepatocytes depends on vesicular transport [21, 31, 63, 68, 69], but is independent from ER or Golgi [63]. Hence, lipoproteins are not loaded with TOH during their intracellular assembly, but rather after exocytosis, a mechanism is required for the presentation of α-TOH at the plasma membrane. Evidence has been provided that the trafficking of α-TOH to the plasma membrane is realized via α-TTP which is located at recycling endosomes in hepatocytes [33]. However, the mechanism of

suggested to contribute to the degradation of T3s [55].

**2.5. Release of vitamin E**

Our knowledge about the storage of vitamin E in lipid droplets is also limited. It was recently reported that newly endocytosed vitamin E was also found in lipid droplets, thus indicating endosome-lipid-droplet interactions [33].
