Age-associated decline in effective immune synapse formation of CD4(+) T cells is reversed by vitamin E supplementation

Aging is associated with reduced IL-2 production and T cell proliferation. Vitamin E supplementation, in aged animals and humans, increases cell division and IL-2 production by naive T cells. The immune synapse forms at the site of contact between a T cell and an APC and participates in T cell activation. We evaluated whether vitamin E affects the redistribution of signaling proteins to the immune synapse. Purified CD4(+) T cells, from the spleens of young and old mice, were treated with vitamin E before stimulation with a surrogate APC expressing anti-CD3. Using confocal fluorescent microscopy, we observed that CD4(+) T cells from old mice were significantly less likely to recruit signaling proteins to the immune synapse than cells from young mice. Vitamin E increased the percentage of old CD4(+) T cells capable of forming an effective immune synapse. Similar results were found following in vivo supplementation with vitamin E. When compared with memory cells, naive T cells from aged mice were more defective in immune synapse formation and were more responsive to vitamin E supplementation. These data show, for the first time, that vitamin E significantly improves age-related early T cell signaling events in naive CD4(+) T cells.     

Impact of vitamin E or selenium deficiency on nematode-induced alterations in murine intestinal function

The effects of deficiencies in the antioxidant nutrients, vitamin E and selenium, on the host response to gastrointestinal nematode infection are unknown. The aim of the study was to determine the effect of antioxidant deficiencies on nematode-induced alterations in intestinal function in mice. BALB/c mice were fed control diets or diets deficient in selenium or vitamin E and the response to a secondary challenge inoculation with Heligmosomoides polygyrus was determined. Egg and worm counts were assessed to determine host resistance. Sections of jejunum were mounted in Ussing chambers to measure changes in permeability, absorption, and secretion, or suspended in organ baths to determine smooth muscle contraction. Both selenium and vitamin E deficient diets reduced resistance to helminth infection. Vitamin E, but not selenium, deficiency prevented nematode-induced decreases in glucose absorption and hyper-contractility of smooth muscle. Thus, vitamin E status is an important factor in the physiological response to intestinal nematode infection and may contribute to antioxidant-dependent protective mechanisms in the small intestine.     

Influence of vitamin E and C supplementation on lipoprotein oxidation in patients with Alzheimer's disease.

Because increased oxidation is an important feature of Alzheimer's disease (AD) and low concentrations of antioxidant vitamins C and E have been observed in cerebrospinal fluid (CSF) of AD patients, supplementation with these antioxidants might delay the development of AD. Major targets for oxidation in brain are lipids and lipoproteins. We studied whether supplementation with antioxidative vitamins E and C can increase their concentrations not only in plasma but also in CSF, and as a consequence decrease the susceptibility of lipoproteins to in vitro oxidation. Two groups, each consisting of 10 patients with AD, were for 1 month supplemented daily with either a combination of 400 IU vitamin E and 1000 mg vitamin C, or 400 IU vitamin E alone. We found that supplementation with vitamin E and C significantly increased the concentrations of both vitamins in plasma and CSF. Importantly, the abnormally low concentrations of vitamin C were returned to normal level following treatment. As a consequence, susceptibility of CSF and plasma lipoproteins to in vitro oxidation was significantly decreased. In contrast, the supplementation with vitamin E alone significantly increased its CSF and plasma concentrations, but was unable to decrease the lipoprotein oxidizability. These findings document a superiority of a combined vitamin E + C supplementation over a vitamin E supplementation alone in AD and provide a biochemical basis for its use.     

Inhibition of immunoglobulin E production in allergic model mice by supplementation with vitamin E and beta-carotene

A diet containing different amounts of vitamin E (alpha-tocopherol; 0.5 mg, 5 mg, 10 mg or 50 mg per 100 g diet) was supplemented to BALB/c mice for 6 weeks. These mice were subcutaneously immunized twice with ovalbumin (OVA). A passive cutaneous anaphylaxis (PCA) analysis demonstrated that the mice fed on the diet containing 5 mg of vitamin E produced the highest level of the OVA-specific immunoglobulin E (IgE) antibody. A lower level of serum IgE was found in the mice supplemented with 0.5 mg, 10 mg and 50 mg of vitamin E. A sandwich ELISA analysis showed that the pattern of the total IgE antibody level among these four groups was the same as that of the allergen-specific IgE. In a separate experiment, 5 mg of vitamin E and/or 50 mg of beta-carotene was supplemented to the basal diet containing vitamin E as alpha-tocopherol acetate (5 mg) in order to evaluate the effect of their combination on OVA-specific and total IgE production in the mice. The supplementation with beta-carotene alone had no effect on OVA-specific or total IgE production. In contrast, supplementation with vitamin E plus beta-carotene effectively suppressed both the antigen-specific and total IgE antibodies. The serum vitamin E and beta-carotene levels were increased by supplementation with the respective compounds. These results strongly suggest that the combination of dietary vitamin E and beta-carotene suppressed IgE production and would therefore help to prevent the type-I allergic reaction.     

Effect of vitamin C and E supplementation in modulating the peripheral nerve conduction following cold exposure in humans

Exposure to an extremely cold environment without proper protection leading to hypothermia is an emergency, one of the several complications of which is impairment in nerve conduction. Our previous work in the rat model has shown the beneficial effect of vitamin C in modulating the effect of hypothermia on nerve conduction. The present study aimed to evaluate the effect of vitamins C and E, administered alone or in combination, in modulating the effect of mild hypothermia on human ulnar nerve conduction. The study was carried out on 26 volunteers divided into three groups: group I received vitamin C supplementation (2000 mg/day in a single dose and 1,000 mg/day for the next 6 days), group II received vitamins C and E in combination (1,000 mg and 800 mg respectively in a single dose and 500 mg and 400 mg respectively for the next 6 days) and group III received vitamin E (800 mg in a single dose and the same for the next 6 days). The recordings were carried out before and after single and weekly supplementation in each group. There was a fall in ulnar nerve conduction velocity with a reduction in the oral temperature of 2–2.5 °C. Vitamin C administered alone and in combination with vitamin E reduced the fall in ulnar nerve conduction velocity. Prior supplementation with vitamin C and E could help ameliorate the impairment in human ulnar nerve conduction due to hypothermia.     

Dietary supplementation with vitamin E and C attenuates dexamethasone-induced glucose intolerance in rats

Glucocorticoid excess induces marked insulin resistance and glucose intolerance. A recent study has shown that antioxidants prevent dexamethasone (DEX)-induced insulin resistance in cultured adipocytes. The purpose of this investigation was to examine the effects of dietary vitamin E and C (Vit E/C) supplementation on DEX-induced glucose intolerance in rats. We hypothesized that feeding rats a diet supplemented with Vit E/C would improve glucose tolerance and restore insulin signaling in skeletal muscle, adipose, and liver and prevent alterations in AMPK signaling in these tissues. Male Wistar rats received either a control or Vit E/C-supplemented diet (0.5 g/kg diet each of L-ascorbate and DL-all rac-alpha-tocopherol) for 9 days prior to, and during, 5 days of daily DEX treatment (subcutaneous injections 0.8 mg/g body wt). DEX treatment resulted in increases in the glucose and insulin area under the curve (AUC) during an intraperitoneal glucose tolerance test. The glucose, but not insulin, AUC was lowered with Vit E/C supplementation. Improvements in glucose tolerance occurred independent of a restoration of PKB phosphorylation in tissues of rats stimulated with an intraperitoneal injection of insulin but were associated with increases in AMPK signaling in muscle and reductions in AMPK signaling and the expression of fatty acid oxidation enzymes in liver. There were no differences in mitochondrial enzymes in triceps muscles between groups. This study is the first to report that dietary Vit E/C supplementation can partially prevent DEX-induced glucose intolerance in rats.     

Exercise-induced oxidant stress in the lung tissue: role of dietary supplementation of vitamin E and selenium.

Strenuous physical exercise in the form of swimming in female albino rats increased the oxidative reactions, probably leading to the generation of oxy-free radicals in the lung tissue. Free radical-mediated lipid peroxidation measured in the form of lipid peroxides increased in the pulmonary tissue in response to exhaustive exercise, indicating such a possibility. Dietary supplementation of vitamin E (Vit.E) and selenium (Se) for a period of 12 weeks reduced the oxidative reactions and the ensuing lipid peroxidation in the pulmonary tissue. Physical exercise in control animals induced the activity of superoxide dismutase (SOD), the superoxide anion radical (O2-.) quencher. However, the SOD levels in nutrient-fed animals at rest and after exercise remained well below the control levels, indicating the decreased generation of oxy-free radicals in them. Similarly, selenium-dependent glutathione peroxidase (Se-GSH Px), the enzyme involved in the reduction of organic and inorganic peroxides, and glutathione S- transferase (GST), the multifunctional protein involved in the detoxification of a number of xenobiotics, were increased in response to exercise in control animals, but were significantly decreased in nutrient-fed animals upon exercise. The induction of GST seems to be more towards the peroxidase activity of GST, i.e., non-selenium glutathione peroxidase (Non-Se-GSH Px), which is primarily involved in the reduction of endoperoxides. The studies thus indicate the induction of oxidative stress in the pulmonary tissue upon exhaustive physical exercise and the effectiveness of vit.E and Se independently and more so in combination in combating the exercise-induced oxidant stress.     

Effects of vitamin E and C supplementation either alone or in combination on exercise-induced lipid peroxidation in trained cyclists

Seven trained male cyclists (ate 22.3 +/- 2 years) participated in 4 separate supplementation phases. They ingested 2 capsules per day containing the following treatments: placebo (placebo plus placebo); vitamin C (1 g per day vitamin C plus placebo); vitamin C and E (1 g per day vitamin C plus 200 IU per kg vitamin E); and vitamin E (400 IU per kg vitamin E plus placebo). The treatment order (placebo, vitamin C, vitamin C and E, and vitamin E) was the same for all subjects. Performance trials consisting of a 60-minute steady state ride (SSR) and a 30-minute performance ride (PR) on Cybex 100 Metabolic cycles were performed after each trial. Workloads of 70% of the VO2max were set for the SSR and PR rides, with pedal rate maintained at 90 rpm (SSR) or self determined (PR). Blood samples (5 ml) were drawn pre- and postexercise and analyzed for malonaldehyde (MDA) and lactic acid. The results indicate that vitamin E treatment was more effective than vitamin C alone or vitamin C and E. Pre-exercise plasma levels of MDA in the vitamin E trial was 39% below the pre-exercise MDA levels of the placebo: 2.94 +/- 0.54 and 4.81 +/- 0.65 micromol per ml, respectively. Plasma MDA following exercise in the vitamin E group was also lower than teh placebo: 4.32 +/- 0.37 vs 7.89 +/- 1.0 micromol per ml, respectively. Vitamin C supplementation, on the other hand, elevated both the resting and exercise plasma levels of MDA. None of th supplemental phases had any significant effect on performance. In conclusion, the results indicate that 400 IU/day of vitamin E reduces membrane damage more effectively than vitamin C but does not enhance performance. Athletes are encouraged to include antioxidants, such as vitamin E and C, in their diet to counteract these detrimental effects of exercise. The data presented here suggests that 400 IU/day of vitamin E will provide adequate protection but supplementing the diet with 1 g per day of vitamin C may promote cellular damage. However neither of these vitamins, either alone or in combination, will enhance exercise performance.     

Deoxycytidine glyoxal: lesion induction and evidence of repair following vitamin C supplementation in vivo

Oxidative DNA damage is postulated to be involved in carcinogenesis, and as a consequence, dietary antioxidants have received much interest. A recent report indicates that vitamin C facilitates the decomposition of hydroperoxides in vitro, generating reactive aldehydes. We present evidence for the in vivo generation of glyoxal, an established product of lipid peroxidation, glucose/ascorbate autoxidation, or free radical attack of deoxyribose, following supplementation of volunteers with 400 mg/d vitamin C. Utilizing a monoclonal antibody to a deoxycytidine-glyoxal adduct (gdC), we measured DNA lesion levels in peripheral blood mononuclear cells. Supplementation resulted in significant (p =.001) increases in gdC levels at weeks 11, 16, and 21, with corresponding increases in plasma malondialdehyde levels and, coupled with previous findings, is strongly suggestive of a pro-oxidative effect. However, continued supplementation revealed a highly significant (p =.0001) reduction in gdC levels. Simultaneous analysis of cyclobutane thymine dimers revealed no increase upon supplementation but, as with gdC, levels decreased. Although no single mechanism is identified, our data demonstrate a pro-oxidant event in the generation of reactive aldehydes following vitamin C supplementation in vivo. These results are also consistent with our hypothesis for a role of vitamin C in an adaptive/repair response and indicate that nucleotide excision repair specifically may be affected.     

The influence of mRNA primary and secondary structure on human IFN-gamma gene expression in E. coli.

Parameters influencing the efficiency of expression of the human immune interferon (IFN-gamma) gene in E. coli were studied by comparing a series of eight in vitro-derived gene variants. These contained all possible combinations of silent mutations in the first three codons of the mature IFN-gamma polypeptide coding sequence. Expression levels varied up to 50-fold among the different constructions. Comparison of messenger RNA secondary structure models for each variant suggested that the presence of stem-loop structures blocking the translation initiation signals could drastically decrease the efficiency of IFN-gamma synthesis. With variants displaying no stable mRNA secondary structure in the region, a C----U transition at position +11 after the AUG resulted in a 5-fold increase in expression indicating that RNA primary structure also plays an important role in expression. In addition we demonstrate that, in this system, a spacing of 8 nucleotides between the Shine-Dalgarno region and AUG was optimal for gene expression and that the steady-state production level of IFN-gamma rose exponentially with increasing rate of synthesis.     

The NADPH oxidase Nox3 constitutively produces superoxide in a p22phox-dependent manner: its regulation by oxidase organizers and activators

Nox3, a member of the superoxide-producing NADPH oxidase (Nox) family, participates in otoconia formation in mouse inner ears, which is required for perception of balance and gravity. The activity of other Nox enzymes such as gp91(phox)/Nox2 and Nox1 is known to absolutely require both an organizer protein (p47(phox) or Noxo1) andanactivatorprotein (p67(phox) or Noxa1); for the p47(phox)-dependent activation of these oxidases, treatment of cells with stimulants such as phorbol 12-myristate 13-acetate is also indispensable. Here we show that ectopic expression of Nox3 in various types of cells leads to phorbol 12-myristate 13-acetate-independent constitutive production of a substantial amount of superoxide under the conditions where gp91(phox) and Nox1 fail to generate superoxide, i.e. in the absence of the oxidase organizers and activators. Nox3 likely forms a functional complex with p22(phox); Nox3 physically interacts with and stabilizes p22(phox), and the Nox3-dependent superoxide production is totally dependent on p22(phox). The organizers p47(phox) and Noxo1 are capable of enhancing the superoxide production by Nox3 in the absence of the activators, and the enhancement requires the interaction of the organizers with p22(phox), further indicating a link between Nox3 and p22(phox). The p47(phox)-enhanced Nox3 activity is further facilitated by p67(phox) or Noxa1, whereas the activators cancel the Noxo1-induced enhancement. On the other hand, the small GTPase Rac, essential for the gp91(phox) activity, is likely dispensable to the Nox3 system. Thus Nox3 functions together with p22(phox) as an enzyme constitutively producing superoxide, which can be distinctly regulated by combinatorial use of the organizers and activators.     

Developmental regulation of hepatic testosterone hydroxylases: simultaneous activation and repression of constitutively expressed cytochromes P450 in senescent rats

The aging process is generally associated with marked decreases in the activities of numerous enzymes as well as lower levels of sex hormones such as testosterone. We therefore examined testosterone metabolism in liver microsomes from individual 3- and 24-month-old male rats. Although the old rats exhibited lower 16 alpha-, 6 beta-, and 2 alpha-hydroxylase activities than the young rats, the old rats had a higher 7 alpha-hydroxylase activity. Immunoquantitation of P450a, a known 7 alpha-hydroxylase, showed that the level of this protein was elevated in the old rats, and was correlated with 7 alpha-hydroxylase activity. The mRNA for P450a was measured with a cDNA probe and its level was fivefold higher in the old rats, whereas levels of mRNA coding for a 6 beta-hydroxylase P450 were markedly decreased. The increased expression of cytochrome P450a demonstrates that the observed common decrease in cytochrome P450-catalyzed activities with senescence is not a universal phenomenon. Thus, constitutive expression of specific cytochrome P450 genes is repressed or activated in senescent rats.     

Dietary fatty acid composition rather than vitamin E supplementation influence ex vivo cytokine and eicosanoid response of porcine alveolar macrophages

This study examined the influence of different dietary fat sources (animal fat, sunflower oil, and fish oil) and supplementation of vitamin E (85, 150 and 300 mg all-rac-alpha-tocopheryl acetate/kg diet) on the ex vivo synthesis of eicosanoids and cytokines by porcine alveolar macrophages. Supplementation of vitamin E provoked an increase in the concentration of alpha-tocopherol of the macrophages irrespective of fat sources. Fish oil increased the macrophage n-3 content with 100% and 40%, and reduced the n-6 with 60% and 53% in comparison with sunflower oil and animal fat, respectively. Fish oil decreased the production of TNF-alpha, IL-8, LTB4, and PGE2 (but not IL-6) relative to the other dietary fat sources, and no difference was observed between sunflower oil and animal fat. Positive correlations were found between the n-6 fatty acid content and the production of PGE2, and the PGE2 production was positively correlated with TNF-alpha and IL-8. Negative correlations were found between the n-3 PUFA content and the concentration of PGE2, TNF-alpha and IL-8. In conclusion, dietary fish oil supplemented at a level of 5%, but not supplemental vitamin E, influenced the inflammatory responses of alveolar macrophages isolated from weaned pigs relatively to animal fat and sunflower oil.     

The influence of dietary vitamin C and E supplementation on the physiological response of pirarucu, Arapaima gigas, in net culture

This study evaluated the efficacy of dietary vitamin C (ascorbic acid or AA), vitamin E (alpha-tocopherol or alpha-T), and C+E supplementation on the blood parameters of Arapaima gigas grown in net cages for 45 days. Four treatments were tested: control (commercial feed); C800; E500 and C+E (800+500) with supplementation of 800 mg AA kg(-1), 500 mg alpha-T kg(-1) and 800+500 mg AA+alpha-T kg(-1), respectively. Hematocrit (Ht), red blood cells (RBC), and hemoglobin concentration (Hb) (oxidative status indicators), thrombocytes and leukocytes (immunological indicators), plasma protein and glucose were evaluated. Fish fed vitamin C and C+E supplemented diets showed greater weight gain and survival. Dietary vitamin C and C+E diet supplementation resulted in increased Ht, Hb, RBC, MCHC, total leukocytes, total proteins, thrombocytes and eosinophils compared to the control and alpha-T. The alpha-tocopherol-supplemented diet reduced the number of total thrombocytes, lymphocytes and neutrophils and increased glucose and eosinophils relatively to the control. In general, leukocytes and thrombocytes were good indicators of the efficiency of vitamin on the defense mechanism of the A. gigas reared in cages. Results indicate that high alpha-T diet supplementation provides no benefit for the maintenance of the oxidative or the immunological status of A. gigas. However, it was demonstrated that high dietary AA improves A. gigas immunological status. Red blood cell indices and immune system indicators showed no synergistic effect between the vitamins after supplementing the A. gigas diet with alpha-T+AA.     

Role of vitamin C and E supplementation on IL-6 in response to training

Vitamin C and E supplementation has been shown to attenuate the acute exercise-induced increase in plasma interleukin-6 (IL-6) concentration. Here, we studied the effect of antioxidant vitamins on the regulation of IL-6 expression in muscle and the circulation in response to acute exercise before and after high-intensity endurance exercise training. Twenty-one young healthy men were allocated into either a vitamin (VT; vitamin C and E, n = 11) or a placebo (PL, n = 10) group. A 1-h acute bicycling exercise trial at 65% of maximal power output was performed before and after 12 wk of progressive endurance exercise training. In response to training, the acute exercise-induced IL-6 response was attenuated in PL (P < 0.02), but not in VT (P = 0.82). However, no clear difference between groups was observed (group × training: P = 0.13). Endurance exercise training also attenuated the acute exercise-induced increase in muscle-IL-6 mRNA in both groups. Oxidative stress, assessed by plasma protein carbonyls concentration, was overall higher in the VT compared with the PL group (group effect: P < 0.005). This was accompanied by a general increase in skeletal muscle mRNA expression of antioxidative enzymes, including catalase, copper-zinc superoxide dismutase, and glutathione peroxidase 1 mRNA expression in the VT group. However, skeletal muscle protein content of catalase, copper-zinc superoxide dismutase, or glutathione peroxidase 1 was not affected by training or supplementation. In conclusion, our results indicate that, although vitamin C and E supplementation may attenuate exercise-induced increases in plasma IL-6 there is no clear additive effect when combined with endurance training.     

Natural vitamin E alpha-tocotrienol: retention in vital organs in response to long-term oral supplementation and withdrawal

The natural vitamin E tocotrienol (TCT) possesses biological properties not shared by tocopherols (TCP). Nanomolar alpha-TCT, not alpha-TCP, is potently neuroprotective (JBC 275:13049; 278:43508; Stroke 36:2258). The report that the affinity of TTP to bind (alpha-TCT is an order of magnitude lower than that for alpha-TCP questions the bioavailability of orally taken TCT to tissues. Oral supplementation of TCT for 3 years in nine generations of female and male rat was studied. Ten vital organs were examined. To gain insight into the turnover of alpha-TCT in tissues, a subset of supplemented rats was moved to vitamin E deficient diet for 7 weeks. Orally supplemented alpha-TCT was delivered to all vital organs including the brain and spinal cord in significant amounts. In organs such as the skin, adipose and gonads the maximum level of alpha-TCT achieved in response to supplementation was folds higher than baseline values of alpha-TCP in rats maintained on laboratory chow. Females had higher levels of alpha-TCT compared to matched tissues of corresponding males. To gain insight into how quickly alpha-TCT is metabolized in the tissues, washout of alpha-TCT from vital organs was examined. alpha-TCT accumulated in vital organs over more than 2 years was almost completely lost in less than 2 months when the supplementation was stopped. This is in sharp contrast with findings related to alpha-TCP retention. The ability of long-term oral supplementation to maintain and elevate alpha-TCT levels in vital organs together with the rapid elimination of the intact vitamin from all organs studied underscores the need for continuous oral supplementation of TCT.     

The absence of adaptive changes in tissue activities of glutathione-S-transferase, superoxide dismutase and catalase following selenium and vitamin E supplementation in subjects with low selenium status

Three groups of New Zealand women were given daily in a double blind randomised study, 200 micrograms Se as sodium selenite, 170 mg alpha-tocopherol or a placebo for 4 wk. Activities of glutathione-S-transferase, superoxide dismutase and catalase were assayed in erythrocytes, plasma and platelets and in liver and muscle biopsy tissues. No changes in activities of any of these tissue enzymes were observed in any of the three groups. There were also no changes in non-selenium dependent glutathione peroxidase activities in liver or plasma. The lack of changes in any of these enzymes following selenium supplementation suggests that adaptive changes to the low selenium status of these subjects had not occurred through these lipid peroxidation defense mechanisms.