Regulation by vitamin E of phosphatidylcholine metabolism in rat heart.

Lysophosphatidylcholine is the major lysophospholipid in mammalian tissues and has been shown to be cytolytic at high concentrations. In the present study we demonstrated that the level of lysophosphatidylcholine was significantly increased in the heart of rats fed with a vitamin E-deficient diet. Moreover, the cardiac lysophosphatidylcholine level was decreased in rats fed with a high vitamin E diet. The alterations in cardiac lysophosphatidylcholine level by dietary vitamin E were attributed to the changes in the activity of cardiac phospholipase A. Dietary vitamin E affected both phospholipase A1 and A2 in the same manner, but had no effect on the other major enzymes which are responsible for the metabolism of lysophosphatidylcholine. Kinetic studies revealed that the inhibition of enzyme activity by vitamin E was essentially non-competitive. The accumulation of lysophosphatidylcholine in the rat heart may be one of the underlying biochemical causes of the observed cardiac dysfunctions produced during vitamin E deficiency.     

Biophysical models of the heart electrical activity

Based on the fundamental knowledge of the space-temporal organization of extracellular electrical fields of the myocardium, a system for 3-D computer modeling of the cardiac electrical activity at different structural levels of the object is being developed at the Institute of Theoretical and Experimental Biophysics. The system is based on the earlier proposed and modified biophysical model of the electrocardiosignal genesis represented by a double electrical layer along the surface of the electrically active myocardium. The system combines the model for activation and repolarization of the heart ventricles; the advanced model for the evaluation of parameters of the cardiac electric field, which makes it possible to derive model electrocardiosignals both in the direct regime of calculation of the potentials and in the regime of calculation of electrocardiosignals from preliminarily determined components of the multipole equivalent electrical heart generator; a database for the model parameters and their combinations in the form of cards of simulated "patients", and a database of modeled electrocardiosignals. In the present paper (first from three within the framework of the problem), simulation methods in electrocardiology are briefly described and a biophysical model of the heart electrical activity is presented, which has made up the basis of the system for computer modeling of forward and inverse problems of the cardiac electric field. The parameters of the model are electrophysiological, anatomical, and biophysical characteristics of the heart.     

Effects of vitamin E on prostacyclin release and lipid composition of the ischemic rat heart

Free radical-mediated reperfusion injury has been established as an important mechanism leading to post-ischemic reperfusion myocardial damage. The present study was undertaken to determine the protective role of vitamin E, a membrane-bound free-radical scavenger, on ischemia-reperfusion myocardial injury. After 4 months of feeding a semipurified diet containing 0, 30, and 3000 ppm of R,R,R,-alpha-tocopherol acetate, rat hearts were subjected to Langendorff perfusion. Myocardial damage was judged by the release of creatine phosphokinase (CPK) after 45 min of global ischemia followed by 20 min of reperfusion. Effluent CPK was significantly lowered in the two tocopherol-supplemented groups, although increasing dietary vitamin E by 100-fold above requirement did not confer further protection. However, effluent prostacyclin, detected as the stable metabolite 6-keto-PGF1 alpha by radioimmunoassay, was potentiated by dietary vitamin E in a dose-dependent manner. Analysis of lipids in cardiac subcellular fractions showed considerable enrichment of tocopherol in these membranes by diets, but the levels of polyunsaturated fatty acids, phospholipids, and cholesterol were essentially unchanged by dietary treatment or ischemia-reperfusion. These data demonstrated that requirement level of tocopherol (30 ppm) in the diet is sufficient to protect against reperfusion injury of the myocardium and suggests that tocopherol is important in maintaining cardiac prostacyclin synthesis under conditions of oxygen stress.     

Phosphatidylcholine metabolism in isolated rat heart: modulation by ethanol and vitamin E

Prolonged ethanol administration has been reported to cause defects in cardiac performance and abnormal cardiac lipid contents. However, little is known regarding the short-term administration of ethanol to the perfused heart and its effect on cardiac phospholipid metabolism. In this study, the isolated Langendorff heart perfusion was used as a model to study the effects of ethanol and a combination of ethanol and vitamin E (DL-alpha-tocopherol) on phospholipid metabolism. When perfused with 1% ethanol for 4 h, the major cardiac phospholipids were not altered but a 60% increase in lysophosphatidylcholine level was observed. Studies on the lysophosphatidylcholine metabolic enzymes revealed that phospholipase A (both phospholipase A1 and A2) activity was enhanced in the ethanol-perfused heart, but lysophospholipase and acyltransferase activities were unaffected by ethanol treatment. When the heart was perfused with 1% ethanol in the presence of 50-100 microM vitamin E, the ethanol-induced lysophosphatidylcholine accumulation was completely abolished. This was largely attributed to the attenuation of phospholipase A activities by vitamin E. In order to delineate the opposing effects of ethanol and vitamin E on phospholipid metabolism in the heart, phospholipase A activities in the subcellular fractions were determined in the presence of 0.5-2.0% ethanol or a combination of 1% ethanol and 0-100 microM vitamin E. Ethanol alone exhibited a biphasic effect on phospholipase A activity with maximum stimulation of enzyme activities at 1% concentration. When phospholipase A was assayed in 1% ethanol and vitamin E (25-100 microM), its activity was inhibited by vitamin E in a dose-dependent manner. The mechanism by which ethanol enhanced phospholipase A activities was further investigated with a partially purified enzyme from the rat heart cytosol. Kinetic studies with different concentrations of phosphatidylcholine revealed that at low substrate concentrations, ethanol was inhibitory to the reaction, whereas at high substrate concentrations, the reaction was enhanced by ethanol. Vitamin E (50 microM) completely abolished the ethanol-induced enhancement of enzyme activity in a noncompetitive manner. Since lysophosphatidylcholine is cytolytic at high concentration and its accumulation in the heart has been postulated as a biochemical cause of cardiac dysfunction, the level of the lysolipid in the heart must be under rigid control. Our result suggest that the modulation of cardiac phospholipase A activity is an important mechanism for the the regulation of lysophosphatidylcholine levels in the rat heart.     

Adenosine triphosphatase activity of streptozotocin-induced diabetic rat brain microsomes. Effect of vitamin E

Hyperglycemia causes protein glycosylation, oxidation and alterations in enzyme activities, which are the underlying causes of diabetic complications. This study was undertaken to test the role of vitamin E treatment on Ca2+-ATPase activity, protein glycosylation and lipid peroxidation in the brain of streptozotocin (STZ)-induced diabetic rats. Male rats weighing about 250-300 g were rendered diabetic by a single STZ injection of 50 mg/kg via the tail vein. Both the diabetic and non-diabetic rats were fed a vitamin E supplemented diet (500 IU/kg/day). Ca2+-ATPase activity was significantly reduced at week 10 of diabetes compared to the control group (p < 0.05), with 0.225+/-0.021 U/I (mean +/- S.E.M.) in the control group and 0.072 +/- 0.008 U/l (mean +/- S.E.M.) in the diabetic group. Vitamin E treatment prevented the enzyme activity from decreasing. The activities observed were 0.226 +/- 0.020 U/l and 0.172 +/- 0.011 U/I (mean +/- S.E.M.) in the vitamin E-treated control and diabetic group, respectively. STZ-induced diabetes resulted in an increased protein glycosylation and lipid peroxidation. Vitamin E treatment led to a significant inhibition in blood glucose, protein glycosylation and lipid peroxidation, which in turn prevented abnormal activity of the enzyme in the brain. This study indicates that vitamin E supplementation may reduce complications of diabetes in the brain.     

Ontogenesis of the vitamin D receptor in rat heart

1,25-Dihydroxyvitamin D(3) through its receptor (vitamin D receptor; VDR) has important physiological effects such as calcium transport and cell growth and differentiation. Although the VDR is present in a variety of cell lines as well as in numerous tissues, including rat and human heart, no data are available about the presence of VDR in heart at different steps of rat life. In this study we evaluated the VDR expression using RT-PCR and immunohistochemical techniques in fetal (17, 18 and 20 gestational days), neonatal (4 and 8 days) and adult rat heart. Immunohistochemical techniques showed the VDR protein localisation in the nuclei of cardiac muscle fibres. Also, we demonstrated that VDR mRNA expression is changing over these different periods of development, showing significant differences in 20 days versus 18 days of fetal age. These changes in VDR expression may be related to other parameters associated with the development of the cardiac muscle and/or intracellular cardiac cell calcium homeostasis.     

Genetic vitamin E deficiency does not affect MPTP susceptibility in the mouse brain

Oxidative stress is involved in the degeneration of the nigrostriatal dopaminergic system in Parkinson's disease (PD). Vitamin E (alpha-tocopherol) is a potent antioxidant in the cell membrane that can trap free radicals and prohibit lipid peroxidation. The retention and secretion of vitamin E are regulated by alpha-tocopherol transfer protein (TTP) in the brain and liver. Dysfunction of TTP results in systemic deficiency of vitamin E in humans and mice, and increased oxidative stress in mouse brain. In this study, we investigated the effect of vitamin E deficiency in PD development by generating an 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD using TTP knockout (TTP-/-) mice. Vitamin E concentration in the brains of TTP+/- mice was half that in TTP+/+ mice, and in TTP-/- mice, was undetectable. MPTP treatment tended to decrease striatal dopamine, but the effect was comparable and not significant in any of the three genotypes. Furthermore, the extent of loss of dopaminergic cell bodies in the substantia nigra did not differ among the groups. One the other hand, oral administration of vitamin E resulted in the partial protection of striatal dopaminergic terminals against MPTP toxicity. Our results suggest that vitamin E does not play a major protective role in MPTP-induced nigrostriatal dopaminergic neurodegeneration in the brain.     

The extraordinary antioxidant activity of vitamin E phosphate

The antioxidant activities of RRR-vitamin E (VE), all-rac-vitamin E (all-rac-VE), trolox, RRR-vitamin E acetate (VEA), all-rac-vitamin E phosphate (VEP) and RRR-vitamin E succinate (VES) were compared. In this study, the rank order in the inhibition of lipid peroxidation (LPO) of VE and its derivatives was trolox>VE approximately all-rac-VE>VEA>VES. VE and trolox inhibited LPO in non-heated and heated rat liver microsomes. It has generally been accepted that this is due to scavenging of free radicals by these antioxidants, and during this protection the antioxidants are oxidized. VEA and VES have to be converted into VE by esterases to obtain antioxidant activity against LPO. VEP, however, had a potent antioxidant effect of its own without conversion to VE. In contrast to VE, VEP is not consumed during this protection. Of the compounds tested, VEP is the most potent in induction of hemolysis of erythrocytes. EPR experiments using the spin label 16-doxylstearic acid showed that VEP reduces membrane fluidity, in contrast to VE. This indicates that VEP acts as a detergent and forms a barrier that might inhibit the transfer of radicals from one polyunsaturated fatty acid to another. This new mechanism may form the basis for a new class of antioxidants.     

Dose-dependent modulation of systemic lipid peroxidation and activity of anti-oxidant enzymes by vitamin E in the rat

The objective of this work was to examine the time-dependent pro-oxidant versus antioxidant effect of various doses of vitamin E used commonly in experimental studies. Erythrocyte activity of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT) and plasma lipid peroxidation levels were investigated following biweekly intramuscular administration of 100, 300 and 600 mg/kg of vitamin E at a baseline time point, and additionally at 2, 4 and 6 weeks after initiating treatment. Vitamin E had an antioxidant effect when administered at low doses over short time periods, and increased the activity of antioxidant enzymes. At higher doses and over longer time periods, it increased the level of lipid peroxidation, and attenuated the activity of antioxidant enzymes. These results suggest that time-dependent variations in vitamin E effects should be considered in design and interpretation of experimental antioxidant studies, as well as during clinical trials.     

Dose-dependent modulation of systemic lipid peroxidation and activity of anti-oxidant enzymes by vitamin E in the rat

Abstract

The objective of this work was to examine the time-dependent pro-oxidant versus antioxidant effect of various doses of vitamin E used commonly in experimental studies. Erythrocyte activity of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT) and plasma lipid peroxidation levels were investigated following biweekly intramuscular administration of 100, 300 and 600 mg/kg of vitamin E at a baseline time point, and additionally at 2, 4 and 6 weeks after initiating treatment. Vitamin E had an antioxidant effect when administered at low doses over short time periods, and increased the activity of antioxidant enzymes. At higher doses and over longer time periods, it increased the level of lipid peroxidation, and attenuated the activity of antioxidant enzymes. These results suggest that time-dependent variations in vitamin E effects should be considered in design and interpretation of experimental antioxidant studies, as well as during clinical trials.     

The effects of diazinon on lipid peroxidation and antioxidant enzymes in rat heart and ameliorating role of vitamin E and vitamin C

Diazinon is one of the most widely used organophosphate insecticides (OPIs) in agriculture and public health programs. Reactive oxygen species (ROS) caused by OPIs may be involved in the toxicity of various pesticides. The aim of this study was to investigate how diazinon affects lipid peroxidation (LPO) and the antioxidant defense system in vivo and the possible ameliorating role of vitamins E and C. For this purpose, experiments were done to study the effects of DI on LPO and the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in adult rat heart. Experimental groups were: (1) control group, (2) diazinon treated (DI) group, (3) DI+vitamins E and C-treated (DI+Vit) group. The levels of malondialdehyde (MDA) and the activities of SOD and CAT increased significantly in the DI group compared with the control group. The activity of SOD and the levels of MDA decreased significantly in the DI+Vit group compared with the DI group. The differences between the DI+Vit and control groups according to the MDA levels and the activities of both SOD and CAT were statistically significant. These results suggest that treating rats with a single dose of diazinon increases LPO and some antioxidant enzyme activities in the rat myocardium and, in addition, that single-dose treatment with a combination of vitamins E and C after the administration of diazinon can reduce LPO caused by diazinon, though this treatment was not sufficiently effective to reduce the values to those in control group.     

Increased ultra weak chemiluminescence emission from rat heart at postischemic reoxygenation: protective role of vitamin E

Aim of this study was to confirm an increased free radical generation rate during ischemia-reoxygenation, by ultra-weak chemiluminescence detection at the surface of perfused rat heart. We observed that reoxygenation following 30 min global ischemia, induces an increase of ultraweak chemiluminescence emission in isolated perfused heart only if partial depletion of vitamin E is induced by dietary manipulation. Moreover, in normal diet fed rats, vitamin E is partially consumed during global ischemia, but not during reoxygenation. Since chemiluminescence increases during post-ischemic reperfusion, when vitamin E myocardial content is lowered, the most probable free radicals involved are the hydroperoxyl radical derivatives of lipids. These radicals, indeed, are known both to produce photoemission by disproportion and to react with vitamin E. On the other hand, the nature of the reaction that consumes vitamin E during ischemia is still obscure. Accordingly, the basal level of vitamin E myocardial content seems to be a key factor for protecting the heart against reoxygenation injury and its consumption during ischemia could be a determinant of myocardial sensitivity to oxidative stress during reperfusion.     

In vitro activity of essential oils extracted from plants used as spices against fluconazole-resistant and fluconazole-susceptible Candida spp

In the present study, the antifungal activity of selected essential oils obtained from plants used as spices was evaluated against both fluconazole-resistant and fluconazole-susceptible Candida spp. The Candida species studied were Candida albicans, Candida dubliniensis, Candida tropicalis, Candida glabrata, and Candida krusei. For comparison purposes, they were arranged in groups as C. albicans, C. dubliniensis, and Candida non-albicans. The essential oils were obtained from Cinnamomum zeylanicum Breyn, Lippia graveolens HBK, Ocimum basilicum L., Origanum vulgare L., Rosmarinus officinalis L., Salvia officinalis L., Thymus vulgaris L., and Zingiber officinale. The susceptibility tests were based on the M27-A2 methodology. The chemical composition of the essential oils was obtained by gas chromatography-mass spectroscopy and by retention indices. The results showed that cinnamon, Mexican oregano, oregano, thyme, and ginger essential oils have different levels of antifungal activity. Oregano and ginger essential oils were found to be the most and the least efficient, respectively. The main finding was that the susceptibilities of fluconazole-resistant C. albicans, C. dubliniensis, and Candida non-albicans to Mexican oregano, oregano, thyme, and ginger essential oils were higher than those of the fluconazole-susceptible yeasts (P<0.05). In contrast, fluconazole-resistant C. albicans and Candida non-albicans were less susceptible to cinnamon essential oil than their fluconazole-susceptible counterparts (P<0.05). A relationship between the yeasts' susceptibilities and the chemical composition of the essential oils studied was apparent when these 2 parameters were compared. Finally, basil, rosemary, and sage essential oils did not show antifungal activity against Candida isolates at the tested concentrations.     

Utility of a fluorescent vitamin E analogue as a probe for tocopherol transfer protein activity

The tocopherol transfer protein (TTP) is a member of the CRAL-TRIO family of lipid binding proteins that facilitates vitamin E transfer between membrane vesicles in vitro. In cultured hepatocytes, TTP enhances the secretion of tocopherol to the media; presumably, tocopherol transfer is at the basis of this biological activity. The mechanism underlying ligand transfer by TTP is presently unknown, and available tools for monitoring this activity suffer from complicated assay procedure and poor sensitivity. We report the characterization of a fluorescent vitamin E analogue, (R)-2,5,7,8-tetramethylchroman-2-[9-(7-nitrobenz[1,2,5]oxadiazol-4-ylamino)nonyl]chroman-6-ol (NBD-TOH), as a sensitive and convenient probe for the ligand binding and transfer activities of TTP. Upon binding to TTP, NBD-TOH fluorescence is blue shifted, and its intensity is greatly enhanced. We used these properties to accurately determine the affinity of NBD-TOH to TTP. The analogue binds to TTP reversibly and with high affinity (K(d) = 8.5 +/- 6 nM). We determined the affinity of NBD-TOH to a TTP protein in which lysine 59 is replaced with a tryptophan. When occurring in humans, this heritable mutation causes the ataxia with vitamin E deficiency (AVED) disorder. We find that the affinity of NBD-TOH to this mutant TTP is greatly diminished (K(d) = 71 +/- 19 nM). NBD-TOH functioned as a sensitive fluorophore in fluorescent resonance energy transfer (FRET) experiments. Using the fluorescent lipids TRITC-DHPE or Marina Blue-DHPE as a donor or an acceptor for NBD-TOH fluorescence, we obtained high-resolution kinetic data for tocopherol movement out of lipid bilayers, a key step in the TTP-facilitated ligand transfer reaction.