Effects of dietary vitamin E on the biosynthesis of 5-lipoxygenase products by rat polymorphonuclear leukocytes (PMNL)

Activation of polymorphonuclear neutrophils (PMNL) leads to the release of arachidonate from cellular phospholipids via a phospholipase A2, and conversion of products of the 5-lipoxygenase pathway. Evidence to date indicates the dietary vitamin E ((R,R,R)-alpha-tocopherol) can influence both cyclooxygenase and phospholipase A2 activities and that the effect of this vitamin is cell/tissue specific. The present study was undertaken in order to examine the effects of varying dietary tocopherol on PMNL tocopherol content and 5-lipoxygenase product profile using the ionophore A23187 as stimulant in the presence and absence of exogenous arachidonate. Feeding semi-purified diets containing 0, 30 or 3000 ppm of (R,R,R)-alpha-tocopherol acetate to weanling rats for 17 weeks resulted in a dose-related enrichment of PMNL tocopherol. Stimulation of PMNL elicited a significant and rapid loss of tocopherol. When PMNL were stimulated with A23187 alone, the synthesis of 5-HETE, LTB4 and 19-hydroxy-LTB4 was decreased in proportion to increasing dietary tocopherol concentrations. However, when exogenous arachidonate was provided with A23187, intermediate amounts of dietary tocopherol (30 ppm) still suppressed the formation of 5-lipoxygenase products, but high doses (3000 ppm) did not have any additional inhibitory effect. This differential response to high concentrations of vitamin E in the presence and absence of exogenous arachidonate highly suggest that at these concentrations, tocopherol may act principally at the level of substrate release whereas at lower concentrations, 5-lipoxygenase is inhibited. Data from this study demonstrated that attenuation of the formation of 5-lipoxygenase products in PMNL can be achieved by dietary vitamin E enrichment.     

Kinetics of tissue RRR-alpha-tocopherol depletion and repletion. Effect of cold exposure

Vitamin E was estimated in plasma and tissues of rats kept for three months on a low vitamin E diet or a high vitamin E diet. Some of the animals from each group were switched to the opposite diet, and the kinetics of uptake and depletion of vitamin E were followed 3, 8, and 15 days after the diet change. Some rats were also submitted to cold exposure (6 degrees C) for three days. During repletion plasma, red blood cells, liver, spleen, and adrenal gland were the only tissues that responded rapidly to the diet change; after three days, their vitamin E levels corresponded to that of the new diet. Heart, brain, lung, muscle, and thymus were slow in reacting to diet change. Fifteen days after the change in diet, white adipose tissue did not respond. The rate of repletion for all tissues was more rapid than the rate of depletion, but liver was the only tissue that after three days had vitamin E levels corresponding to the low-vitamin diet. Cold exposure for three days did not produce any significant change in the vitamin E content of any tissue, indicating that despite high oxygen consumption by the animal, vitamin E was not consumed or mobilized.     

植物维生素E生物强化研究进展

维生素E是一种只能在光合组织中合成的脂溶性小分子有机化合物,是人体和动物营养不可缺少的重要维生素。由于植物中维生素E含量较低,人类大多处于慢性缺乏维生素E--“隐性饥饿”的状态,而动物饲料中则需要添加外源合成的维生素E以满足其营养需求。因此,提高植物中维生素E的含量是改善维生素E缺乏的重要途径之一。从维生素E的合成途径入手,详细地综述了维生素E合成关键酶基因的表达变化以及前体物质的含量变化对维生素E合成的影响,发现三烯生育酚和α-生育酚的生物强化效果较好,而生育酚总量提高受限;进而从遗传的角度探讨了维生素E合成受限的原因以及遗传上可能影响维生素E合成的其他代谢途径;最后结合可能影响维生素E合成的调控因子以及其前体物质的转运等方面为今后维生素E的生物强化提出了新的思路。     

Distribution of tocotrienols in rats fed a rice bran tocotrienol concentrate

To examine the distribution of rice bran tocotrienol (T3), we gave rice bran T3 to rats after considering an acceptable daily intake of vitamin E for humans. Male SD rats (5 weeks of age) were fed for 3 weeks on a commercial diet containing 6.4 mg of vitamin E per 100 g wt and additively received vitamin E or the vehicle (vitamin E-free corn oil) by oral intubation. The animals were randomly divided into 4 groups depending on the type of test diet: control (vehicle), non-T3 (no T3 + 4.3 mg of tocopherol (TOC)/kg body weight (b.w.)/day), low-T3 (0.8 mg T3 + 3.5 mg TOC/kg b.w./day), and high-T3 (3.2 mg T3 + 1.1 mg TOC/kg b.w./day). The control rats and rats in the non-T3, low-T3, and high-T3 groups took 4.3 and 8.6 mg of vitamin E/kg b.w./day, respectively. Rice bran gamma-T3 was significantly distributed to the adipose tissue and increased from 1.1 to 10.2 nmol/g of adipose tissue according to the rice bran T3 intake.     

Distribution of Tocotrienols in Rats Fed a Rice Bran Tocotrienol Concentrate

To examine the distribution of rice bran tocotrienol (T3), we gave rice bran T3 to rats after considering an acceptable daily intake of vitamin E for humans. Male SD rats (5 weeks of age) were fed for 3 weeks on a commercial diet containing 6.4 mg of vitamin E per 100 g wt and additively received vitamin E or the vehicle (vitamin E-free corn oil) by oral intubation. The animals were randomly divided into 4 groups depending on the type of test diet: control (vehicle), non-T3 (no T3 + 4.3 mg of tocopherol (TOC)/kg body weight (b.w.)/day), low-T3 (0.8 mg T3 + 3.5 mg TOC/kg b.w./day), and high-T3 (3.2 mg T3 + 1.1 mg TOC/kg b.w./day). The control rats and rats in the non-T3, low-T3, and high-T3 groups took 4.3 and 8.6 mg of vitamin E/kg b.w./day, respectively. Rice bran γ-T3 was significantly distributed to the adipose tissue and increased from 1.1 to 10.2 nmol/g of adipose tissue according to the rice bran T3 intake.     

In vitro oxidation of alpha-tocopherol (vitamin E) in human platelets upon incubation with unsaturated fatty acids, diamide and superoxide

Incubation of human blood platelets in vitro in Tyrode solution with unsaturated fatty acids, diamide or superoxide (generated in situ) resulted in the oxidation of tocopherol in the platelets. Arachidonate concentrations of (3-5).10(-4) M caused a 50% decrease in platelet alpha-tocopherol. The addition of saturated fatty acids or platelet-active substances such as ADP, dibutyryl cyclic AMP, and some prostaglandins, or peroxidizing agents such as hydrogen peroxide and tert-butylhydroperoxide to the incubation medium did not cause any change in platelet tocopherol content. During incubations of platelets with arachidonate, malonaldehyde as well as alpha-tocopherolquinone were produced. The latter was also produced during incubations with diamide or superoxide. The oxidation of tocopherol induced by unsaturated fatty acids may be one factor responsible for the well-known increase in dietary vitamin E requirements induced by polyunsaturated fatty acids. The oxidative consumption of tocopherol in the membranes could be expected to take place during localized release of oxidants such as superoxide and polyunsaturated fatty acids during normal biological function (e.g., phagocytosis) or pathological processes (e.g., ischemia). Tocopherol utilization is kept low probably by the regeneration of the compound by vitamin C and/or the preferential utilization of the other biological antioxidants.     

Serum Retinol,alpha‐tocopherol,and lipids in four species of adult captive pinnipeds

The sera of adult aquarium‐held pinnipeds from four species (family Phocidae: harbor seals (Phoca vitulina) and gray seals (Halichoerus grypus); family Otariidae: northern fur seals (Callorhinus ursinus) and California sea lions (Zalophus californianus)) were analyzed for vitamin A (retinol), vitamin E (α‐tocopherol), total cholesterol, triglycerides, phospholipids, and fatty acids. Each subject animal was healthy at the time of blood collection, was fasted for at least 12 hr prior to sampling, and was maintained on a constant diet and supplement regime throughout the study. Retinol values for the four species ranged from 0.16 to 0.92 μg/mL, with the lowest concentrations seen in the harbor seals and the highest in the northern fur seals. Vitamin E values ranged from 10.55 to 43.58 μg/mL, with northern fur seals showing the highest and gray seals the lowest levels. Vitamin E/lipid ratios (cholesterol, triglyceride, phospholipid, and total lipids) were also examined. A significant correlation was seen between vitamin E and total lipids (P<0.05) and phospholipid (P<0.01). Statistical analysis of the retinol, tocopherol, triglyceride, and phospholipid levels showed significant differences between phocid and otariid seals. Otariids had significantly lower tocopherol and phospholipid values (19.36 μg/mL, 4.29 mg/mL) and the phocids had significantly lower retinol and triglyceride levels (0.29 μg/mL, 124 mg/dL). There was no significant difference in serum cholesterol. Zoo Biol 22:83–96, 2003. © 2003 Wiley‐Liss, Inc.     

维生素E对妊娠中期高糖环境大鼠皮下脂肪组织中asprosin的表达影响

摘要 目的：探讨维生素E对妊娠中期高糖环境大鼠皮下脂肪组织中asprosin的表达影响。方法：将妊娠中期高糖环境大鼠(n=21)随机平分为三组-模型组、吡格列酮与维生素E组。格列酮与维生素E组分别灌胃80 mg/kg的吡格列酮和5 mg/kg的维生素E,模型组灌胃等剂量的0.9 % NaCl,1次/d,检测皮下脂肪组织中asprosin表达情况。结果：吡格列酮组与维生素E组给药第3 d、第7 d的血糖、体重低于模型组(P<0.05),维生素E组低于吡格列酮组(P<0.05)。吡格列酮组与维生素E组给药第7 d的皮肤组织超氧化物歧化酶(Superoxide dismutase,SOD)含量高于模型组(P<0.05),丙二醛(Malondialdehyde,MDA)含量低于模型组(P<0.05),吡格列酮组与维生素E组对比差异也都有统计学意义(P<0.05)。吡格列酮组与维生素E组给药第7 d的皮肤组织asprosin蛋白相对表达水平低于模型组(P<0.05),维生素E组低于吡格列酮组(P<0.05)。结论：维生素E在妊娠中期高糖环境大鼠的应用能抑制皮下脂肪组织中asprosin的表达,提高SOD活性,降低MDA的表达,从而降低大鼠的体重与血糖水平。     

Simultaneous expression of <Emphasis Type="Italic">Arabidopsis</Emphasis> ρ-hydroxyphenylpyruvate dioxygenase and MPBQ methyltransferase in transgenic corn kernels triples the tocopherol content

The quantity and composition of tocopherols (compounds with vitamin E activity) vary widely among different plant species reflecting the expression, activity and substrate specificity of enzymes in the corresponding metabolic pathway. Two Arabidopsis cDNA clones corresponding to ρ-hydroxyphenylpyruvate dioxygenase (HPPD) and 2-methyl-6-phytylplastoquinol methyltransferase (MPBQ MT) were constitutively expressed in corn to further characterize the pathway and increase the kernel tocopherol content. Transgenic kernels contained up to 3 times as much γ-tocopherol as their wild type counterparts whereas other tocopherol isomers remained undetectable. Biofortification by metabolic engineering offers a sustainable alternative to vitamin E supplementation for the improvement of human health.     

Role of vitamin E in the protection of the resident macrophage membrane against oxidative damage

The variation in tocopherol content of resident peritoneal rat macrophages was investigated during an oxidative stress provided by superoxide anions. Fluorometric measurements showed that phagocytic cells contain 298 +/- 18 ng vit.E/mg prot. The vitamin E level remains nearly constant during 1 h of incubation: 266 +/- 46 ng vit.E/mg prot. HPLC control at 37 degrees C validates our fluorometric measurement. Superoxide anions (O2-.) synthesis was activated by phorbol myristate acetate (PMA) (0.5 microgram/ml), after 1 h of incubation a decrease of 40% of the macrophage tocopherol level was observed and assessed by HLPC control. No tocopherolquinone (TQ) was detected. To clarify this point, tocopherol oxidation was followed spectrophotometrically. Results did not show any appearance of TQ at 265 nm but appearance of a peak at 307 mm. This our results show for the first time that macrophages possess vitamin E which plays a partial role in the protection of their plasma membrane. The lack of detection of TQ is of interest and the study of this unidentified product of oxidation should help us to understand the exact metabolism of vitamin E.     

Cytochrome P450 omega-hydroxylase pathway of tocopherol catabolism. Novel mechanism of regulation of vitamin E status

Postabsorptive elimination of the various forms of vitamin E appears to play a key role in regulation of tissue tocopherol concentrations, but mechanisms of tocopherol metabolism have not been elucidated. Here we describe a pathway involving cytochrome P450-mediated omega-hydroxylation of the tocopherol phytyl side chain followed by stepwise removal of two- or three-carbon moieties, ultimately yielding the 3'-carboxychromanol metabolite that is excreted in urine. All key intermediates of gamma-tocopherol metabolism via this pathway were identified in hepatocyte cultures using gas chromatography-mass spectrometry. NADPH-dependent synthesis of the initial gamma- and alpha-tocopherol 13'-hydroxy and -carboxy metabolites was demonstrated in rat and human liver microsomes. Functional analysis of several recombinant human liver P450 enzymes revealed that tocopherol-omega-hydroxylase activity was associated only with CYP4F2, which also catalyzes omega-hydroxylation of leukotriene B(4) and arachidonic acid. Tocopherol-omega-hydroxylase exhibited similar binding affinities but markedly higher catalytic activities for gamma-tocopherol than alpha-tocopherol, suggesting a role for this pathway in the preferential physiological retention of alpha-tocopherol and elimination of gamma-tocopherol. Sesamin potently inhibited tocopherol-omega-hydroxylase activity exhibited by CYP4F2 and rat or human liver microsomes. Since dietary sesamin also results in elevated tocopherol levels in vivo, this pathway appears to represent a functionally significant means of regulating vitamin E status.     

Effect of dietary vitamin E on platelet tocopherol values and 12-lipoxygenase activity

This study was designed to evaluate the effects of different amounts of dietary vitamin E on platelet tocopherol levels and 12-lipoxygenase activity when exogenous arachidonic acid was used as substrate. Weanling male Sprague-Dawley rats were fed diets containing 0, 50, and 5000 ppm of D-alpha-tocopherol acetate for 4 months. Platelet tocopherol was increased with increasing concentrations of dietary vitamin E; however, the conversion of exogenously added arachidonate by platelet to 12-HETE (12-hydroxyeicosatetraenoic acid) and thromboxane B2 from these three dietary groups was essentially the same. This study provides direct evidence that platelet 12-lipoxygenase activity is independent of its vitamin E content when exogenously added arachidonate was used as substrate.     

Biochemical consequences of heritable mutations in the alpha-tocopherol transfer protein

Tocopherol transfer protein (TTP) regulates vitamin E status by facilitating the secretion of tocopherol from liver to circulating lipoproteins. Heritable mutations in the ttpA gene, encoding for TTP, result in ataxia with vitamin E deficiency (AVED) syndrome, typified by low vitamin E levels and a plethora of neurological disorders. The molecular mechanisms by which TTP facilitates tocopherol secretion are presently unknown. We recently showed that vitamin E is taken up by hepatocytes through an endocytic process and that, shortly following uptake, the vitamin is found primarily in lysosomes. We showed further that TTP is localized to late endocytic vesicles and that it facilitates the intracellular trafficking of tocopherol from lysosomes to the plasma membrane. To gain insight into the molecular mechanisms that underlie TTP actions, we studied the physiological impact of three naturally occurring heritable mutations in the ttpA gene (the R59W, R221W, and A120T substitutions). We found that these mutations impair the ability of TTP to facilitate the secretion of vitamin E from cells. Furthermore, the degree of impairment corresponded to the severity of the AVED pathology associated with each mutation. In cells that express mutated TTP proteins, vitamin E did not traffic to the plasma membrane and remained "trapped" in lysosomes. In addition, we observed that substitution mutations that cause the AVED syndrome impart a marked instability on the TTP protein. These observations suggest that the physiological role of TTP is anchored in its ability to direct vitamin E trafficking from the endocytic compartment to transport vesicles that deliver the vitamin to the site of secretion at the plasma membrane.     

Differential Indicators of Diabetes-Induced Oxidative Stress in New Zealand White Rabbits: Role of Dietary Vitamin E Supplementation

Determination of reliable bioindicators of diabetes-induced oxidative stress and the role of dietary vitamin E supplementation were investigated. Blood (plasma) chemistries, lipid peroxidation (LPO), and antioxidant enzyme activities were measured over 12 weeks in New Zealand White rabbits (control, diabetic, and diabetic + vitamin E). Cholesterol and triglyceride levels did not correlate with diabetic state. PlasmaLPOwas influenced by diabetes and positively correlated with glucose concentration only, not cholesterol or triglycerides. Liver glutathione peroxidase (GPX) activity negatively correlated with glucose and triglyceride levels. Plasma and erythrocyte GPX activities positively correlated with glucose, cholesterol, and triglyceride concentrations. Liver superoxide dismutase activity positively correlated with glucose and cholesterol concentration. Vitamin E reduced plasma LPO, but did not affect the diabetic state. Thus, plasmaLPOwas the most reliable indicator of diabetes-induced oxidative stress. Antioxidant enzyme activities and types of reactive oxygen species generated were tissue dependent. Diabetes-induced oxidative stress is diminished by vitamin E supplementation.     

Synthetic byproducts of tocopherol oxidation as inhibitors of platelet function

Vitamin E and its fully oxidized form tocopherol quinone are known inhibitors of platelet aggregation. Previous results from our laboratory have shown that the quinone was approximately equal in effectiveness to vitamin E. A recent report of a far greater inhibitory activity of the quinone produced by nitric acid oxidation of vitamin E prompted this investigation. Our studies show that the unusually high inhibition of platelet aggregation, release and cyclooxygenase activity associated with nitric acid oxidized vitamin E is due to byproducts of the oxidation process and not due to tocopherol quinone. Treatment of vitamin E acetate, a substance of mild effect on aggregation and arachidonate metabolism of platelets, with nitric acid did not produce tocopherol quinone but exerted as potent an inhibition as oxidized vitamin E. We conclude that nitric acid oxidation is unsuitable for preparation of tocopherol quinone unless the latter is carefully isolated. Oxidation with permanganate proved to be an alternate method without these difficulties.     

Vitamin E concentrations in different regions of the spinal cord and sciatic nerve of the rat

Since the spinal cord and peripheral nerves are vulnerable to the effects of vitamin E deficiency, vitamin E concentrations in various discrete regions of these parts of the nervous system of the rat were determined. Furthermore, as acrylamide toxicity and vitamin E deficiency share some neuropathological features, tissue vitamin E concentrations in acrylamide-treated rats were also studied. Male Sprague Dawley rats (200 to 250 g body weight) were fed normal rat chow with or without 0.03% acrylamide in their drinking water. After 24 days, the animals were sacrificed and the tissues assayed for vitamin E by a liquid chromatographic method. Vitamin E concentrations decreased from cerebral cortex to spinal cord with no concentration gradients between different levels of the spinal cord. Sciatic nerve concentration of alpha tocopherol was as high as that of cerebral cortex, and the former also contained measurable amounts of gamma tocopherol. Vitamin E concentrations in the majority of nervous tissue samples remained unchanged with acrylamide treatment.Presented in part at the Sixteenth Annual Meeting of the American Society for Neurochemistry, March 1985.     

Fasting and lactation effect fat-soluble vitamin A and E levels in blood and their distribution in tissue of grey seals (Halichoerus grypus)

Grey seals among other phacoids represent a good model to study the mobilisation, transfer and deposition of fat-soluble components such as vitamins in lactating females and suckling pups because during the lactation period mothers may fast completely while secreting large quantities of high fat milks, and pups deposit large amounts of fat as blubber. The level of vitamins A and E in different tissues (liver, adipose tissue, kidney, heart, skeletal muscle, testis) and blood plasma of adult grey seal females and males changed as a result of fasting and lactation; changes were also observed in pups. The most obvious effects were a significant increase of retinol and a decrease of vitamin E levels in plasma of females with the onset of lactation as well as a substantial decrease in liver vitamin E. In suckling pups both retinol and vitamin E levels in plasma increased with the onset of suckling; after weaning no changes in retinol but a significant decrease in plasma vitamin E was observed. While liver vitamin A levels tended to be unaffected by suckling or post-weaning fast, liver vitamin E levels increased with the uptake of milk substantially (P<0.01) and returned at weaning to low levels similar to that in fetuses. Adipose tissue levels of vitamin A and E in both females and pups were only marginally affected by lactation, suckling or post-weaning fast. Results indicate that both plasma and liver levels of vitamin A and E are affected by the mobilisation, absorption and deposition of these components during lactation in seals to a much greater extent than adipose tissue, from which fat-soluble vitamins are mobilized at rates similar to that of lipids.     

Homogentisate phytyltransferase activity is limiting for tocopherol biosynthesis in Arabidopsis

Tocopherols are essential components of the human diet and are synthesized exclusively by photosynthetic organisms. These lipophilic antioxidants consist of a chromanol ring and a 15-carbon tail derived from homogentisate (HGA) and phytyl diphosphate, respectively. Condensation of HGA and phytyl diphosphate, the committed step in tocopherol biosynthesis, is catalyzed by HGA phytyltransferase (HPT). To investigate whether HPT activity is limiting for tocopherol synthesis in plants, the gene encoding Arabidopsis HPT, HPT1, was constitutively overexpressed in Arabidopsis. In leaves, HPT1 overexpression resulted in a 10-fold increase in HPT specific activity and a 4.4-fold increase in total tocopherol content relative to wild type. In seeds, HPT1 overexpression resulted in a 4-fold increase in HPT specific activity and a total seed tocopherol content that was 40% higher than wild type, primarily because of an increase in gamma-tocopherol content. This enlarged pool of gamma-tocopherol was almost entirely converted to alpha-tocopherol by crossing HPT1 overexpressing plants with lines constitutively overexpressing gamma-tocopherol methyltransferase. Seed of the resulting double overexpressing lines had a 12-fold increase in vitamin E activity relative to wild type. These results indicate that HPT activity is limiting in various Arabidopsis tissues and that total tocopherol levels and vitamin E activity can be elevated in leaves and seeds by combined overexpression of the HPT1 and gamma-tocopherol methyltransferase genes.     

Chloroplast membrane photostability in chlP transgenic tobacco plants deficient in tocopherols

The phototolerance of three chlP transgenic tobacco (Nicotiana tabacum) lines, affected in geranylgeranyl reductase and, hence, deficient in tocopherols (vitamin E), was estimated by in vivo luminescence and fluorescence measurements and was compared with that of the wild type (WT). Exposure of leaf discs to high light (1 mmol photon m(-2) s(-1)) and low temperature (10 degrees C) led to a rapid inhibition of photosystem II (PSII) photochemistry that showed little dependence on the tocopherol level. PSII photo-inhibition was followed by lipid peroxidation with a time delay of about 4 h, and this phenomenon was exacerbated in the tocopherol-deficient leaves. A linear correlation was observed in these short-term experiments between resistance to photooxidation and tocopherol content. When whole plants were exposed to the same treatment, PSII was severely photo-inhibited in mature leaves of all genotypes. Lipid peroxidation was also observed in all plants, but it occurred much more rapidly in tocopherol-deficient transgenic plants relative to WT plants. The time at which extensive lipid peroxidation occurred was correlated with the tocopherol content of the leaves. The present results show that tocopherols protect thylakoid membranes against photodestruction through lipid peroxidation. However, tocopherol deficiency was compensated in young, developing leaves that were able to photo-acclimate in the long term and did not suffer from photooxidative damage. Soluble antioxidants (glutathione and ascorbate) did not accumulate in photo-acclimated chlP transgenic leaves relative to WT leaves. In contrast, a selective accumulation of xanthophyll cycle pigments was observed in young transgenic leaves, and this could represent a compensatory mechanism for tocopherol deficiency.     

Vitamin E and its structural analogs in tuberculosis]

Efficiency of tocopherol, its analog with a shortened side chain as well as their quinons for tuberculosis was determined. All the studied compounds inhibited peroxide-formation processes in the liver homogenate and mitochondria. The vitamin E amount in the blood is considerably decreased in case of tuberculosis. The studied analogs increased its content and possessed a weak tuberculostatic but expressed antiedemic and antiinflammatory action. Administration of vitamin E in combination with isoniazid to sick animals had a favourable effect on the processes of tissue respiration and oxidative phosphorylation. If taking a sum of characters, tocopherol acetate with a shortened side chain is the most efficient of all the analogs under study for the tuberculosis therapy.