Deciphering the enigma of the function of alpha-tocopherol as a vitamin.

α-Tocopherol (α-T) is a vitamin, but the reasons for the α-T requirement are controversial. Given that α-T deficiency was first identified in embryos, we studied to the premier model of vertebrate embryo development, the zebrafish embryo. We developed an α-T-deficient diet for zebrafish and used fish consuming this diet to produce α-T deficient (E-) embryos. We showed that α-T deficiency causes increased lipid peroxidation, leading to metabolic dysregulation that impacts both biochemical and morphological changes at very early stages in development. These changes occur at an early developmental window, which takes place prior to an analogous time to when a human knows she is pregnant. We found that α-T limits the chain reaction of lipid peroxidation and protects metabolic pathways and integrated gene expression networks that control embryonic development. Importantly, not only is α-T critical during early development, but the neurodevelopmental process is highly dependent on α-T trafficking by the α-T transfer protein (TTPa). Data from both gene expression and evaluation of the metabolome in E- embryos suggest that the activity of the mechanistic Target of Rapamycin (mTOR) signaling pathway is dysregulated-mTOR is a master regulatory mechanism, which controls both metabolism and neurodevelopment. Our findings suggest that TTPa is needed not only for regulation of plasma α-T in adults but is a key regulator during embryogenesis.

Chronic Vitamin E Deficiency Dysregulates Purine, Phospholipid, and Amino Acid Metabolism in Aging Zebrafish Skeletal Muscle.

Muscle wasting occurs with aging and may be a result of oxidative stress damage and potentially inadequate protection by lipophilic antioxidants, such as vitamin E. Previous studies have shown muscular abnormalities and behavioral defects in vitamin E-deficient adult zebrafish. To test the hypothesis that there is an interaction between muscle degeneration caused by aging and oxidative damage caused by vitamin E deficiency, we evaluated long-term vitamin E deficiency in the skeletal muscle of aging zebrafish using metabolomics. Zebrafish (55 days old) were fed E+ and E- diets for 12 or 18 months. Then, skeletal muscle samples were analyzed using UPLC-MS/MS. Data were analyzed to highlight metabolite and pathway changes seen with either aging or vitamin E status or both. We found that aging altered purines, various amino acids, and DHA-containing phospholipids. Vitamin E deficiency at 18 months was associated with changes in amino acid metabolism, specifically tryptophan pathways, systemic changes in the regulation of purine metabolism, and DHA-containing phospholipids. In sum, while both aging and induced vitamin E deficiency did have some overlap in altered and potentially dysregulated metabolic pathways, each factor also presented unique alterations, which require further study with more confirmatory approaches.

Alpha-Tocopherol from People to Plants Is an Essential Cog in the Metabolic Machinery.

Significance: Protection from oxygen, a diradical, became a necessity with the evolution of photosynthetic organisms about 2.7 billion years. α-Tocopherol plays an essential, protective role in organisms from plants to people. An overview of human conditions that result in severe vitamin E (α-tocopherol) deficiency is provided. Recent Advances: α-Tocopherol has a critical role in the oxygen protection system by stopping lipid peroxidation, its induced damage, and cellular death by ferroptosis. Recent findings in bacteria and plants support the concept of why lipid peroxidation is so dangerous to life and why the family of tocochromanols are essential for aerobic organisms and for plants. Critical Issues: The hypothesis that prevention of the propagation of lipid peroxidation is the basis for the α-tocopherol requirement in vertebrates is proposed and further that its absence dysregulates energy metabolism, one-carbon metabolism, and thiol homeostasis. By recruiting intermediate metabolites from adjacent pathways to sustain effective lipid hydroperoxide elimination, α-tocopherol function is linked not only to NADPH metabolism and its formation through the pentose phosphate pathway via glucose metabolism but also to sulfur-containing amino acid metabolism and to one-carbon metabolism. Future Directions: Evidence from humans, animals, and plants supports the hypothesis, but future studies are needed to assess the genetic sensors that detect lipid peroxidation and cause the ensuing metabolic dysregulation. Antioxid. Redox Signal. 38, 775-791.

Guidance for the diagnosis and treatment of hypolipidemia disorders.

The Abetalipoproteinemia and Related Disorders Foundation was established in 2019 to provide guidance and support for the life-long management of inherited hypocholesterolemia disorders. Our mission is "to improve the lives of individuals and families affected by abetalipoproteinemia and related disorders". This review explains the molecular mechanisms behind the monogenic hypobetalipoproteinemia disorders and details their specific pathophysiology, clinical presentation and management throughout the lifespan. In this review, we focus on abetalipoproteinemia, homozygous hypobetalipoproteinemia and chylomicron retention disease; rare genetic conditions that manifest early in life and cause severe complications without appropriate treatment. Absent to low plasma lipid levels, in particular cholesterol and triglyceride, along with malabsorption of fat and fat-soluble vitamins are characteristic features of these diseases. We summarize the genetic basis of these disorders, provide guidance in their diagnosis and suggest treatment regimens including high dose fat-soluble vitamins as therapeutics. A section on preconception counseling and other special considerations pertaining to pregnancy is included. This information may be useful for patients, caregivers, physicians and insurance agencies involved in the management and support of affected individuals.

α-Tocopherol Pharmacokinetics in Adults with Cystic Fibrosis: Benefits of Supplemental Vitamin C Administration.

BACKGROUND: Numerous abnormalities in cystic fibrosis (CF) could influence tocopherol absorption, transportation, storage, metabolism and excretion. We hypothesized that the oxidative distress due to inflammation in CF increases vitamin E utilization, which could be positively influenced by supplemental vitamin C administration. METHODS: Immediately before and after receiving vitamin C (500 mg) twice daily for 3.5 weeks, adult CF patients (n = 6) with moderately advanced respiratory tract (RT) disease consumed a standardized breakfast with 30% fat and a capsule containing 50 mg each hexadeuterium (d6)-α- and dideuterium (d2)-γ-tocopheryl acetates. Blood samples were taken frequently up to 72 h; plasma tocopherol pharmacokinetics were determined. During both trials, d6-α- and d2-γ-tocopherols were similarly absorbed and reached similar maximal plasma concentrations ~18-20 h. As predicted, during vitamin C supplementation, the rates of plasma d6-α-tocopherol decline were significantly slower. CONCLUSIONS: The vitamin C-induced decrease in the plasma disappearance rate of α-tocopherol suggests that vitamin C recycled α-tocopherol, thereby augmenting its concentrations. We conclude that some attention should be paid to plasma ascorbic acid concentrations in CF patients, particularly to those individuals with more advanced RT inflammatory disease and including those with severe exacerbations.

Aerosolized vitamin E acetate causes oxidative injury in mice and in alveolar macrophages.

Although vitamin E acetate (VEA) is suspected to play a causal role in the development of electronic-cigarette, or vaping, product use-associated lung injury (EVALI), the underlying biological mechanisms of pulmonary injury are yet to be determined. In addition, no study has replicated the systemic inflammation observed in humans in a murine EVALI model, nor investigated potential additive toxicity of viral infection in the setting of exposure to vaping products. To identify the mechanisms driving VEA-related lung injury and test the hypothesis that viral infection causes additive lung injury in the presence of aerosolized VEA, we exposed mice to aerosolized VEA for extended times, followed by influenza infection in some experiments. We used mass spectrometry to evaluate the composition of aerosolized VEA condensate and the VEA deposition in murine or human alveolar macrophages. Extended vaping for 28 days versus 15 days did not worsen lung injury but caused systemic inflammation in the murine EVALI model. Vaping plus influenza increased lung water compared with virus alone. Murine alveolar macrophages exposed to vaped VEA hydrolyzed the VEA to vitamin E with evidence of oxidative stress in the alveolar space and systemic circulation. Aerosolized VEA also induced cell death and chemokine release and reduced efferocytotic function in human alveolar macrophages in vitro. These findings provide new insights into the biological mechanisms of VEA toxicity.

Vitamin E: How much is enough, too much and why!

α-Tocopherol (α-T) is a required dietary nutrient for humans and thus is a vitamin. This narrative review focuses on vitamin E structures, functions, biological determinants and its deficiency symptoms in humans. The mechanisms for the preferential α-T tissue enrichment in the human body include the α-T transfer protein (TTPA) and the preferential metabolism of non-α-T forms. Potential new α-T biomarkers, pharmacokinetic data, and whether there are better approaches to evaluate and set the α-T dietary requirement are discussed. Finally, the possible role of α-T supplements in delay of chronic diseases and the evaluation of vitamin E safety are considered.

Expanding role of vitamin E in protection against metabolic dysregulation: Insights gained from model systems, especially the developing nervous system of zebrafish embryos.

This review discusses why the embryo requires vitamin E (VitE) and shows that its lack causes metabolic dysregulation and impacts morphological changes at very early stages in development, which occur prior to when a woman knows she is pregnant. VitE halts the chain reactions of lipid peroxidation (LPO). Metabolomic analyses indicate that thiols become depleted in E- embryos because LPO generates products that require compensation using limited amino acids and methyl donors that are also developmentally relevant. Thus, VitE protects metabolic networks and the integrated gene expression networks that control development. VitE is critical especially for neurodevelopment, which is dependent on trafficking by the α-tocopherol transfer protein (TTPa). VitE-deficient (E-) zebrafish embryos initially appear normal, but by 12 and 24 h post-fertilization (hpf) E- embryos are developmentally abnormal with expression of pax2a and sox10 mis-localized in the midbrain-hindbrain boundary, neural crest cells and throughout the spinal neurons. These patterning defects indicate cells that are especially in need of VitE-protection. They precede obvious morphological abnormalities (cranial-facial malformation, pericardial edema, yolksac edema, skewed body-axis) and impaired behavioral responses to locomotor activity tests. The TTPA gene (ttpa) is expressed at the leading edges of the brain ventricle border. Ttpa knockdown using morpholinos is 100% lethal by 24 hpf, while E- embryo brains are often over- or under-inflated at 24 hpf. Further, E- embryos prior to 24 hpf have increased expression of genes involved in glycolysis and the pentose phosphate pathway, and decreased expression of genes involved in anabolic pathways and transcription. Combined data from both gene expression and the metabolome in E- embryos at 24 hpf suggest that the activity of the mechanistic Target of Rapamycin (mTOR) signaling pathway is decreased, which may impact both metabolism and neurodevelopment. Further evaluation of VitE deficiency in neurogenesis and its subsequent impact on learning and behavior is needed.

RedEfish: Generation of the Polycistronic mScarlet: GSG-T2A: Ttpa Zebrafish Line.

The vitamin E regulatory protein, the alpha-tocopherol transfer protein (Ttpa), is necessary for zebrafish embryo development. To evaluate zebrafish embryo Ttpa function, we generated a fluorescent-tagged zebrafish transgenic line using CRISPR-Cas9 technology. One-cell stage embryos (from Casper (colorless) zebrafish adults) were injected the mScarlet coding sequence in combination with cas9 protein complexed to single guide RNA molecule targeting 5' of the ttpa genomic region. Embryos were genotyped for proper insertion of the mScarlet coding sequence, raised to adulthood and successively in-crossed to produce the homozygote RedEfish (mScarlet: GSG-T2A: Ttpa). RedEfish were characterized by in vivo fluorescence detection at 1, 7 and 14 days post-fertilization (dpf). Fluorescent color was detectable in RedEfish embryos at 1 dpf; it was distributed throughout the developing brain, posterior tailbud and yolk sac. At 7 dpf, the RedEfish was identifiable by fluorescence in olfactory pits, gill arches, pectoral fins, posterior tail region and residual yolk sac. Subsequently (14 dpf), the mScarlet protein was found in olfactory pits, distributed throughout the digestive tract, along the lateral line and especially in caudal vertebrae. No adverse morphological outcomes or developmental delays were observed. The RedEfish will be a powerful model to study Ttpa function during embryo development.

Gene Expression of CRAL_TRIO Family Proteins modulated by Vitamin E Deficiency in Zebrafish (Danio Rerio).

An evaluation of the impact of vitamin E deficiency on expression of the alpha-tocopherol transfer protein (α-TTP) and related CRAL_TRIO genes was undertaken using livers from adult zebrafish based on the hypothesis that increased lipid peroxidation would modulate gene expression. Zebrafish were fed either a vitamin E sufficient (E+) or deficient (E-) diet for 9 months, then fish were euthanized, and livers were harvested. Livers from the E+ relative to E- fish contained 40-times more α-tocopherol (P <0.0001) and one fourth the malondialdehyde (P = 0.0153). RNA was extracted from E+ and E- livers, then subject to evaluation of gene expression of ttpa and other genes of the CRAL_TRIO family, genes of antioxidant markers, and genes related to lipid metabolism. Ttpa expression was not altered by vitamin E status. However, one member of the CRAL_TRIO family, tyrosine-protein phosphatase non-receptor type 9 gene (ptpn9a), showed a 2.4-fold increase (P = 0.029) in E- relative to E+ livers. Further, we identified that the gene for choline kinase alpha (chka) showed a 3.0-fold increase (P = 0.010) in E- livers. These outcomes are consistent with our previous findings that show vitamin E deficiency increased lipid peroxidation causing increases in phospholipid turnover.

Vitamin A and Vitamin E: Will the Real Antioxidant Please Stand Up?

Vitamin A, acting through its metabolite, all-trans-retinoic acid, is a potent transcriptional regulator affecting expression levels of hundreds of genes through retinoic acid response elements present within these genes. However, the literature is replete with claims that consider vitamin A to be an antioxidant vitamin, like vitamins C and E. This apparent contradiction in the understanding of how vitamin A acts mechanistically within the body is a major focus of this review. Vitamin E, which is generally understood to act as a lipophilic antioxidant protecting polyunsaturated fatty acids present in membranes, is often proposed to be a transcriptional regulator. The evaluation of this claim is another focus of the review. We conclude that vitamin A is an indirect antioxidant, whose indirect function is to transcriptionally regulate a number of genes involved in mediating the body's canonical antioxidant responses. Vitamin E, in addition to being a direct antioxidant, prevents the increase of peroxidized lipids that alter both metabolic pathways and gene expression profiles within tissues and cells. However, there is little compelling evidence that vitamin E has a direct transcriptional mechanism like that of vitamin A. Thus, we propose that the term antioxidant not be applied to vitamin A, and we discourage the use of the term transcriptional mediator when discussing vitamin E.

Vitamin E: necessary nutrient for neural development and cognitive function.

Vitamin E, discovered in 1922, is essential for pregnant rats to carry their babies to term. However, 100 years later, the molecular mechanisms for the vitamin E requirement during embryogenesis remain unknown. Vitamin E's role during pregnancy has been difficult to study and thus, a vitamin E-deficient (E-) zebrafish embryo model was developed. Vitamin E deficiency in zebrafish embryos initiates lipid peroxidation, depletes a specific phospholipid (DHA-phosphatidyl choline), causes secondary deficiencies of choline, betaine and critical thiols (such as glutathione), and dysregulates energy metabolism. Vitamin E deficiency not only distorts the carefully programmed development of the nervous system, but it leads to defects in several developing organs. Both the α-tocopherol transfer protein and vitamin E are necessary for embryonic development, neurogenesis and cognition in this model and likely in human embryos. Elucidation of the control mechanisms for the cellular and metabolic pathways involved in the molecular dysregulation caused by vitamin E deficiency will lead to important insights into abnormal neurogenesis and embryonic malformations.

Moderate Red Wine Consumption Increases the Expression of Longevity-Associated Genes in Controlled Human Populations and Extends Lifespan in Drosophila melanogaster.

The beneficial effects of moderate red wine consumption on cardiovascular health are well known. The composition of red wine includes several compounds, such as the phytoestrogen resveratrol, that exert these beneficial effects, although not all the mechanisms by which they act are known. Our aim was to study the effect of red wine consumption on longevity-related genes in controlled human populations, such as cloistered nuns. We found that the expression of catalase, manganese-superoxide dismutase, Sirt1, and p53 was increased in peripheral blood mononuclear cells after 14 days of moderate red wine consumption. This increase was accompanied by an enhanced metabolic wellness: fatty acids, cholesterol, branched chain amino acids (isoleucine and leucine), ketone bodies (acetoacetate), bacterial co-metabolites (trimethylamine), and cellular antioxidants (taurine) contributed to a change in metabolic profile after moderate red wine consumption by the nuns. No serious unwanted side effects were observed. Finally, we tested the effect of moderate red wine consumption on longevity in a controlled animal population, such as D. melanogaster, and found that it increased average life span by 7%. In conclusion, moderate red wine consumption increases the expression of key longevity-related genes and improves metabolic health in humans and increases longevity in flies.

Vitamin E Deficiency Disrupts Gene Expression Networks during Zebrafish Development.

Vitamin E (VitE) is essential for vertebrate embryogenesis, but the mechanisms involved remain unknown. To study embryonic development, we fed zebrafish adults (>55 days) either VitE sufficient (E+) or deficient (E-) diets for >80 days, then the fish were spawned to generate E+ and E- embryos. To evaluate the transcriptional basis of the metabolic and phenotypic outcomes, E+ and E- embryos at 12, 18 and 24 h post-fertilization (hpf) were subjected to gene expression profiling by RNASeq. Hierarchical clustering, over-representation analyses and gene set enrichment analyses were performed with differentially expressed genes. E- embryos experienced overall disruption to gene expression associated with gene transcription, carbohydrate and energy metabolism, intracellular signaling and the formation of embryonic structures. mTOR was apparently a major controller of these changes. Thus, embryonic VitE deficiency results in genetic and transcriptional dysregulation as early as 12 hpf, leading to metabolic dysfunction and ultimately lethal outcomes.

Vitamin E deficiency dysregulates thiols, amino acids and related molecules during zebrafish embryogenesis.

Vitamin E (α-tocopherol, VitE) was discovered as a nutrient essential to protect fetuses, but its molecular role in embryogenesis remains undefined. We hypothesize that the increased lipid peroxidation due to VitE deficiency drives a complex mechanism of overlapping biochemical pathways needed to maintain glutathione (GSH) homeostasis that is dependent on betaine and its methyl group donation. We assess amino acids and thiol changes that occur during embryogenesis [12, 24 and 48 h post fertilization (hpf)] in VitE-sufficient (E+) and deficient (E-) embryos using two separate, novel protocols to quantitate changes using UPLC-MS/MS. Using partial least squares discriminant analysis, we found that betaine is a critical feature separating embryos by VitE status and is higher in E- embryos at all time points. Other important features include: glutamic acid, increased in E- embryos at 12 hpf; choline, decreased in E- embryos at 24 hpf; GSH, decreased in E- embryos at 48 hpf. By 48 hpf, GSH was significantly lower in E- embryos (P < 0.01), as were both S-adenosylmethionine (SAM, P < 0.05) and S-adenosylhomocysteine (SAH, P < 0.05), while glutamic acid was increased (P < 0.01). Since GSH synthesis requires cysteine (which was unchanged), these data suggest that both the conversion of homocysteine and the uptake of cystine via the Xc- exchanger are dysregulated. Our data clearly demonstrates the highly inter-related dependence of methyl donors (choline, betaine, SAM) and the methionine cycle for maintenance of thiol homeostasis. Additional quantitative flux studies are needed to clarify the quantitative importance of these routes.

Vitamin E catabolism in women, as modulated by food and by fat, studied using 2 deuterium-labeled α-tocopherols in a 3-phase, nonrandomized crossover study.

BACKGROUND: Human vitamin E (α-tocopherol) catabolism is a mechanism for regulating whole-body α-tocopherol. OBJECTIVES: To determine the roles of the intestine and liver on α-tocopherol catabolism as affected by fat or fasting, 2 deuterium-labeled (intravenous d6- and oral d3-) forms of α-tocopherol were used. METHODS: Healthy women received intravenous d6-α-tocopherol and consumed d3-α-tocopherol with a 600-kcal defined liquid meal (DLM; 40% or 0% fat, n = 10) followed by controlled meals; or the 0% fat DLM (n = 7) followed by a 12-h fast (0% fat-fast), then controlled meals ≤72 h. The order of the 3-phase crossover design was not randomized and there was no blinding. Samples were analyzed by LC/MS to determine the α-tocopherol catabolites and α-carboxyethyl hydroxychromanol (α-CEHC) in urine, feces, and plasma that were catabolized from administered oral d3- and intravenous d6-α-tocopherols. RESULTS: Urinary and plasma d3- and d6-α-CEHC concentrations varied differently with the interventions. Mean ± SEM cumulative urinary d6-α-CEHC derived from the intravenous dose excreted over 72 h during the 40% fat (2.50 ± 0.37 µmol/g creatinine) and 0% fat (2.37 ± 0.37 µmol/g creatinine) interventions were similar, but a ∼50% decrease was observed during the 0% fat-fast (1.05 ± 0.39 µmol/g creatinine) intervention (compared with 0% fat, P = 0.0005). Cumulative urinary d3-α-CEHC excretion was not significantly changed by any intervention. Total urinary and fecal excretion of catabolites accounted for <5% of each of the administered doses. CONCLUSIONS: Differential catabolism of the intravenous d6-α-tocopherol and oral d3-α-tocopherol doses shows both liver and intestine have roles in α-tocopherol catabolism. During the 40% fat intervention, >90% of urinary d3-α-CEHC excretion was estimated to be liver-derived, whereas during fasting <50% was from the liver with the remainder from the intestine, suggesting that there was increased intestinal α-tocopherol catabolism while d3-α-tocopherol was retained in the intestine in the absence of adequate fat/food for α-tocopherol absorption.This trial was registered at clinicaltrials.gov as NCT00862433.

Vitamin E is necessary for zebrafish nervous system development.

Vitamin E (VitE) deficiency results in embryonic lethality. Knockdown of the gene ttpa encoding for the VitE regulatory protein [α-tocopherol transfer protein (α-TTP)] in zebrafish embryos causes death within 24 h post-fertilization (hpf). To test the hypothesis that VitE, not just α-TTP, is necessary for nervous system development, adult 5D strain zebrafish, fed either VitE sufficient (E+) or deficient (E-) diets, were spawned to obtain E+ and E- embryos, which were subjected to RNA in situ hybridization and RT-qPCR. Ttpa was expressed ubiquitously in embryos up to 12 hpf. Early gastrulation (6 hpf) assessed by goosecoid expression was unaffected by VitE status. By 24 hpf, embryos expressed ttpa in brain ventricle borders, which showed abnormal closure in E- embryos. They also displayed disrupted patterns of paired box 2a (pax2a) and SRY-box transcription factor 10 (sox10) expression in the midbrain-hindbrain boundary, spinal cord and dorsal root ganglia. In E- embryos, the collagen sheath notochord markers (col2a1a and col9a2) appeared bent. Severe developmental errors in E- embryos were characterized by improper nervous system patterning of the usually carefully programmed transcriptional signals. Histological analysis also showed developmental defects in the formation of the fore-, mid- and hindbrain and somites of E- embryos at 24 hpf. Ttpa expression profile was not altered by the VitE status demonstrating that VitE itself, and not ttpa, is required for development of the brain and peripheral nervous system in this vertebrate embryo model.