Biomedical subjects

N Haramaki

Publications and source records attributed to N Haramaki.

35 records · Page 2Linked to original sources

PubMed1995

Role of ascorbate in protection by nitecapone against cardiac ischemia-reperfusion injury.

The antioxidant properties of nitecapone, a catechol derivative and an inhibitor of catechol-O-methyltransferase, were reported recently. In the present study, the influence of nitecapone on isolated rat heart ischemia-reperfusion injury was investigated to elucidate its cardioprotective role. Nitecapone, administered in the perfusion buffer from the beginning of the pre-ischemic phase, significantly improved recovery of cardiac mechanical function, suppressed enzyme leakage in the coronary effluent, and minimized loss of ascorbate, compared with the control group. In rats fed a diet containing 4% ascorbate, myocardial ascorbate content in ascorbate-fed rats after ischemia-reperfusion was higher than that in control rats fed a normal diet without ischemia. However, supplemented rats did not show any beneficial effects on cardiac mechanical recovery or enzyme leakage, suggesting that maintenance of tissue ascorbate level is not the cause, but the result of the protective effects of nitecapone against cardiac ischemia-reperfusion injury. The iron-chelating effect of nitecapone was also tested. It was confirmed, using electron spin resonance, that 50 microM nitecapone chelates the same concentration of iron released from the heart into the coronary effluent. Hence, the iron-chelating ability of nitecapone may be responsible, at least in part, for its cardioprotective effects in ischemia-reperfusion injury.

Animals↗

PubMed1995

Reduction and transport of lipoic acid by human erythrocytes.

Reduction of exogenous lipoic acid to dihydrolipoate is known to occur in several mammalian cells and tissues. Dihydrolipoate is a potent radical scavenger, and may provide significant antioxidant protection. Because lipoic acid appears in the bloodstream after oral administration, we have examined the reduction of exogenous lipoate by human erythrocytes. Normal human erythrocytes reduced lipoate to dihydrolipoate only in the presence of glucose; deoxyglucose did not substitute for glucose, indicating that the reduction of lipoate requires glucose metabolism. Furthermore, the reduction was shown to be NADPH dependent. Erythrocytes isolated from a human subject with a genetic deficiency of glucose-6-phosphate dehydrogenase (and, therefore, deficient in the formation of NADPH) did not reduce lipoate. Dehydroepiandrosterone, a specific inhibitor of glucose-6-phosphate dehydrogenase, inhibited lipoate reduction. Our findings imply that some of the reduction of exogenous lipoic acid is catalysed by glutathione reductase, a flavoprotein dehydrogenase; mitomycin C, an inhibitor of FAD-dependent reductases, inhibited lipoate reduction by erythrocytes, and glutathione reductase purified from human erythrocytes was observed to reduce lipoic acid in a cell-free system. We further explored these findings with erythrocyte ghosts and liposomes. Our results indicate that a transport system exists for alpha-lipoic acid and dihydrolipoate; resealed erythrocyte ghosts, containing trapped lipoamide dehydrogenase and pyridine nucleotides, reduced externally added lipoate. By contrast, liposomes prepared with enzyme and pyridine nucleotides did not catalyze reduction of lipoate. This work indicates that uptake of exogenous lipoate and reduction to dihydrolipoate by normal human erythrocytes may contribute to oxidant protection in the human bloodstream.

Biological Transport↗

PubMed1995

Glutathione reductase and lipoamide dehydrogenase have opposite stereospecificities for alpha-lipoic acid enantiomers.

The reduction of exogenous alpha-lipoic acid to dihydrolipoate by mammalian cells and tissues confers additional antioxidant protection to the cell. Both (R+) and (S-) isomers of alpha-lipoic acid were analyzed as substrates with glutathione reductase from several sources and with mammalian lipoamide dehydrogenase. Mammalian glutathione reductase catalyzed faster reduction of (S)-lipoic acid (1.4-2.4-fold greater activity) than of (R)-lipoic acid, whereas lipoamide dehydrogenase had a very marked preference for (R)-lipoic acid (18-fold greater activity) over (S)-lipoic acid. Mammalian glutathione reductase showed better affinity for (S)-lipoic acid substrate; Km values were 3.5 mM for (S)-lipoic acid and and 7 mM for (R)-lipoic acid. Glutathione reductase from yeast reduced lipoic acid less efficiently than the mammalian enymes, had a Km for both stereoisomers of about 10 mM, and showed little stereospecificity. Although (S)-lipoic acid is not formed in nature, these findings indicate that exogenous (S)-lipoic acid may have a useful role as an antioxidant for mammalian systems.

Animals↗

PubMed1995

Bio-Catalyzer alpha . rho No. 11 (Bio-Normalizer) supplementation: effect on oxidative stress to isolated rat hearts.

Bio-Catalyzer alpha . rho No. 11 (Bio-Normalizer), a natural health food product prepared by yeast fermentation of medicinal plants, has been recently reported to possess antioxidant properties. To better define its antioxidant action, we investigated the effects of orally supplemented Bio-Normalizer on oxidative damage in the rat heart. Hearts were isolated from control or Bio-Normalizer supplemented animals and 1) exposed to ischemia-reperfusion using the Langendorff technique, or 2) homogenized and exposed to peroxyl radicals generated from (2,2'-azobis (2,4'-dimethylvaleronitrile) (AMVN). During reperfusion following 40 minutes of ischemia, leakage of lactate dehydrogenase from hearts isolated from Bio-Normalizer supplemented rats was significantly lower than from hearts of control animals. Furthermore, lower levels of AMVN-induced accumulation of thiobarbituric acid reactive substances and of protein carbonyl derivatives were measured in homogenates prepared from hearts isolated from Bio-Normalizer supplemented rats than in samples from control animals. Our findings confirm an antioxidant action of Bio-Normalizer and show that it protects the heart against ischemia-reperfusion induced damage.

Analysis of Variance↗

PubMed1995

The influence of vitamin E and dihydrolipoic acid on cardiac energy and glutathione status under hypoxia-reoxygenation.

The combination of vitamin E supplementation with dihydrolipoic acid perfusion synergistically improves cardiac functional recovery during post-ischemic reperfusion or post-hypoxic reoxygenation of the rat heart. To elucidate the mechanism of this effect, isolated rat hearts were perfused using a working heart system. In hearts perfused with a buffer containing dihydrolipoic acid, ATP levels were significantly higher than those of hearts perfused without addition of dihydrolipoic acid during 90 min of reoxygenation following 30 min of hypoxia. Cardiac tissue glutathione status measured in hearts after perfusion experiments showed significant elevation of reduced glutathione in vitamin E supplemented normoxic rat hearts without hypoxia. Significant elevation of oxidized glutathione was observed in dihydrolipoic acid perfused heart after hypoxia-reoxygenation. It is concluded that vitamin E and dihydrolipoic acid exert separate and synergistic effects in the protection of the hypoxic-reoxygenated heart.

Animals↗

PubMed1994

Effects of natural antioxidant ginkgo biloba extract (EGB 761) on myocardial ischemia-reperfusion injury.

Recently, it was reported that Ginkgo biloba extract (EGb 761), which is known to have antioxidant properties, also has antiarrhythmic effects on cardiac reperfusion-induced arrhythmias. In the present study, effects of EGb 761 on cardiac ischemia-reperfusion injury were investigated from the point of view of recovery of mechanical function as well as the endogenous antioxidant status of ascorbate. Isolated rat hearts were perfused using the Langendorff technique, and 40 min of global ischemia were followed by 20 min of reperfusion. EGb 761 improved cardiac mechanical recovery and suppressed the leakage of lactate dehydrogenase (LDH) during reperfusion. Furthermore, EGb 761 diminished the decrease of myocardial ascorbate content after 40 min of ischemia and 20 min of reperfusion. Interestingly, EGb 761 also suppressed the increase of dehydroascorbate. These results indicate that EGb 761 protects against cardiac ischemia-reperfusion injury and suggest that the protective effects of EGb 761 depend on its antioxidant properties.

Analysis of Variance↗

PubMed1994

DNA single-strand breaks and cytotoxicity induced by chromate(VI), cadmium(II), and mercury(II) in hydrogen peroxide-resistant cell lines.

The induction of cytotoxicity and DNA single-strand breaks by chromium(VI), cadmium(II), and mercury(II) were compared in H2O2-resistant Chinese hamster ovary (CHO(R)) cells and parental (CHO(P)) cells. Using a colony-forming assay, CHO(R) cells were found to be significantly more resistant than CHO(P) cells to the cytotoxicity caused by CdCl2 and HgCl2, but not to that caused by Na2CrO4. However, the DNA single-strand breaks produced by each of these metals were significantly lower in the CHO(R) cells. With respect to chromium reduction, the level of chromium(V) in CHO(R) cells was decreased. The role of intracellular active oxygen in the heavy metal-induced DNA damage and cytotoxicity is discussed.

Animals↗

PubMed1994

Protective effects of riboflavin and its derivatives against ischemic reperfused damage of rat heart.

The effects of riboflavin and its derivatives such as FAD, FMN and lumichrome on the levels of high energy phosphate compounds (ATP and creatine phosphate) and intracellular pH in ischemic reperfused rat hearts were investigated using a Langendorff perfusion technique. 31P-NMR study showed a decrease in the levels of high energy phosphate compounds and pH values in myocardium after 30 min global ischemia and a slight recovery of these levels after a 30 min reperfusion following ischemia. However, in all the hearts perfused with riboflavin and its derivatives during ischemia-reperfusion, a marked recovery of high energy phosphate compounds and pH values were observed. In addition, the cardiac mitochondrial respiratory function was protected from ischemia-reperfusion injury. These results suggest that riboflavin, FAD, FMN, and lumichrome have a protective effect against ischemia-reperfusion injury to rat myocardium in vitro. It is assumed that these substances exert their effect directly in the extracellular space.

Adenosine Triphosphate↗

PubMed1993

Cardiac recovery during post-ischemic reperfusion is improved by combination of vitamin E with dihydrolipoic acid.

Effects of dietary vitamin E supplementation in rats were studied to determine whether or not they have a higher tolerance against cardiac ischemia-reperfusion injury using the working or Langendorff heart systems. Also, dihydrolipoic acid, recently reported to have potent antioxidant properties and accelerate vitamin E recycling of membrane in vitro, was perfused into the heart model systems to investigate its in vivo relationship with vitamin E. Tissue vitamin E content was increased by vitamin E feeding, but heart preparations did not show any improved functional recovery. Control hearts perfused with dihydrolipoic acid also did not show any improvement. However, a synergistic response is observed with the combination of dihydrolipoic acid perfusion and high dietary vitamin E using both perfusion systems in improvement of cardiac recovery. These results indicate that a high concentration of myocardial vitamin E does not increase tolerance to ischemia-reperfusion injury by itself, but, the combination of exogenous dihydrolipoic acid and high endogenous vitamin E can produce synergistic protective effects on recovery from ischemia during reperfusion.

Analysis of Variance↗

PubMed1993

Influence of o-phenanthroline on DNA single-strand breaks, alkali-labile sites, glutathione reductase, and formation of chromium(V) in Chinese hamster V-79 cells treated with sodium chromate (VI).

The effect of the cell-permeable metal chelator o-phenanthroline (OP) on the formation of chromium(V), DNA breaks, alkali-labile sites, and enzyme inhibition of chromium(VI) was studied using Chinese hamster V-79 cells. Alkaline elution assays demonstrated that treatment with OP (100-500 microM) resulted in a dose-dependent decrease in cellular levels of either alkali-labile sites or the combination of alkali-labile sites plus DNA single-strand breaks caused by Na2CrO4. Cellular treatment with OP also attenuated the inhibition of glutathione reductase attributed to Na2CrO4. Under the same experimental conditions, the cellular uptake of chromate was not affected by OP. ESR studies revealed that cellular treatment with OP (100-500 microM) resulted in a concentration-dependent decrease in the level of chromium(V) intermediate in cells treated with Na2CrO4. Furthermore, OP inhibited the chromium(V) complex that formed during the reaction of Na2CrO4 with glutathione in vitro, resulting in the decrease of chromium(V)-mediated hydroxyl radical formation while neither the reduction of chromium(VI) nor the formation of the chromium(VI)-glutathione complex was affected, indicating that OP may react with chromium(V) but not with chromium(VI). These results suggest that the metal chelator OP decreases chromate-induced alkali-labile sites or the combination of alkali-labile sites plus DNA strand breaks, as well as chromium inhibition of glutathione reductase, possibly through its ability to directly decrease chromium(V) complex in cells.

Animals↗

PubMed1993

Effect of alpha-tocopherol on high energy phosphate metabolite levels in rat heart by 31P-NMR using a Langendorff perfusion technique.

To examine the action of alpha-tocopherol on high energy phosphate compounds, a 31P-NMR technique was applied to perfused Langendorff rat hearts. Rats were treated with tocopherol acetate (25 mg/kg body wt i.p.) for 7 consecutive days. On the 7th day, the rat hearts were isolated for the Langendorff experiment. After 30 min of global ischemia the NMR signals of creatine phosphate and ATP in myocardium disappeared, and then recovered slightly in the reperfusion following ischemia. However, in the tocopherol-treated rat hearts, the restoration of high energy phosphate compounds occurred quickly after the beginning of reperfusion, although there was no significant difference in the destruction of high energy phosphate compounds during the ischemia. The alpha-tocopherol level in the myocardium was severely depleted by ischemia-reperfusion. In the alpha-tocopherol-treated animals, the alpha-tocopherol level in myocardium was still significantly higher than the control level at the end of 30 min of global ischemia. The heart mitochondrial respiratory function was simultaneously protected against ischemia-reperfusion injury. The role of alpha-tocopherol was discussed as a radical scavenger and membrane stabilizer against oxygen stress.

Adenine Nucleotides↗

PubMed1993

DNA single-strand breaks and cytotoxicity induced by sodium chromate(VI) in hydrogen peroxide-resistant cell lines.

Hydrogen peroxide-resistant Chinese hamster ovary (CHOR) cells were developed by exposing parental (CHO(P)) cells to sequential increases in H2O2 concentration. Cytotoxicity as well as DNA single-strand breaks induced by Na2CrO4 were then compared in CHOR and CHO(P) cell lines. Using the colony-forming assay, it was found that the cytotoxicity caused by Na2CrO4 did not differ in the parent and resistant cells. However, alkaline elution studies showed that the production of DNA single-strand breaks in CHOR cells treated with Na2CrO4 was reduced by about 50% as compared with that in CHO(P) cells. Similarly, electron spin resonance (ESR) studies revealed that the level of chromium(V) in CHOR cells during treatment with Na2CrO4 was about 50% that in CHO(P) cells. CHOR cells were also found to be cross-resistant to the cytotoxicity and DNA breaks caused by other toxic metals such as CdCl2 and HgCl2. Catalase activity in resistant cells was 2-fold and the cellular content of glutathione was 3-fold that in parental cells. However, no obvious differences were seen in superoxide dismutase and glutathione reductase activity, although the contents of ascorbic acid or alpha-tocopherol were slightly decreased in CHOR cells, suggesting that the resistance in CHOR cells may be associated with the increase in both catalase activity and glutathione contents in cells. These results indicate that chromate-induced DNA breaks appear to be mediated by a different mechanism than that for the cytotoxicity of this metal, and also suggest that the formation of active oxygen species and/or chromium(V) during reduction of chromium(VI) inside cells might be associated with the induction of the DNA strand breaks caused by the metal.

Animals↗

PubMed1992

Studies on the hepatotoxicity induced by bis (tributyltin) oxide.

The toxic effects of bis (tributyltin) oxide (TBTO) on the rat liver were studied with an electron microscope and the accumulation sites of tin were determined with an X-ray microanalyzer. The activities of serum enzymes and the concentration of serum bilirubin were also analyzed. Male Wistar rats received an intramuscular injection of 0.5 ml/kg of TBTO. Marked swelling of the mitochondria appeared in the hepatocytes 4 h after injection of TBTO. Cytoplasmic vacuoles, which contained degenerated mitochondria, gradually increased in number in these hepatocytes. This in turn may have caused a decrease in the volume of hepatic cell cords and an enlargement of sinusoids in the entire hepatic lobule. However, fine structures of intrahepatic bile ducts were not altered. By X-ray microanalysis, tin peaks were preferentially obtained from swollen mitochondria of the hepatocytes. By polarographic analysis of the respiratory responses of mitochondria, it was demonstrated that rates of state 4 respiration and respiratory control ratio were significantly disturbed in TBTO-treated rats in comparison with those of controls. The activities of AST (aspartate aminotransferase) and ALT (alanine aminotransferase) were significantly increased after TBTO treatment, but those of ALP (alkaline phosphatase), LAP (leucine aminopeptidase) and total bilirubin were not changed. These results indicated that parenterally administered TBTO accumulated in the liver cell mitochondria and disturbed oxidative phosphorylation. Mitochondrial dysfunction might induce severe damage of the hepatocytes. Four days after injection of TBTO, hepatic structures and chemical indices were almost restored by the regeneration of hepatocytes.

Alanine Transaminase↗

PubMed1991

Electron spin resonance studies on the mechanism of adriamycin-induced heart mitochondrial damages.

Rats were given i.p. injections of Adriamycin (4 mg/kg body weight) for 6 consecutive days. Electron spin resonance spectrometry with spin labeling and a trapping technique was applied to heart mitochondria obtained from treated and control animals, in order to examine the physical response of heart mitochondrial membrane affected by Adriamycin. The Adriamycin treatment resulted in a decrease of membrane fluidity (and an increase in order parameter S), with concomitant dysfunction of respiratory responses. The generation of hydroxyl radicals from mitochondria was enhanced in the Adriamycin-treated group. In addition, superoxide dismutase activity in the mitochondrial matrix was found to decrease. The heart mitochondria of Adriamycin-treated animals contained a large amount of lipid peroxide. These results suggest that the enhancement of hydroxyl radical formation in mitochondria affected by Adriamycin is one of the factors involved in Adriamycin-induced cardiotoxicity.

Animals↗

PubMed1991

Corneal edema induced by bis (tributyltin) oxide.

Corneal edema induced by bis (tributyltin) oxide (TBTO) was studied with an electron microscope and the accumulation sites of tin were determined with an X-ray microanalyzer. Male Wistar rats received an intramuscular injection of 0.5 ml/kg TBTO. After time intervals of 2, 4, 6, 8, 10, 12 h after injection, corneas were isolated and provided for electron microscopy. Corneas from untreated rats served as controls. Marked swelling of mitochondria in the corneal endothelial cells occurred at 4 h after TBTO injection. The corneal edema appeared in the endothelial layer and the stroma at 6 h after injection. By X-ray microanalysis, Sn peaks were obtained from swollen mitochondria in the endothelial cells. At 12 h after TBTO injection, edematous swelling of the corneal tissue became more advanced. These results indicated that parenterally administered TBTO accumulated in the mitochondria of corneal endothelial cells. The direct toxic effects of TBTO on the mitochondria might cause the interference with active pump function of endothelial cells and induced the corneal edema.

Animals↗

PubMed1991

Mechanism of lipid radical formation following exposure of epidermal homogenate to ultraviolet light.

It has been suggested that oxygen free radicals are important mediators of lipid peroxidation in the epidermis exposed to ultraviolet (UV) light. However, it is not clear whether it is the superoxide anion radical (O2-.) or the hydroxyl radical (.OH) that plays the major role in producing the lipid radicals (L.) following UV exposure. In this study, we used electron spin resonance (ESR) technique with the spin trap (5,5-dimethyl-1-pyrroline-N-oxide [DMPO]) to determine which active oxygen species is involved in the UV-induced lipid radical formation (DMPO-L.: aN = 15.5 G, aH = 22.7 G). In the presence of superoxide dismutase or the metal-chelating agent, the DMPO-spin adduct spectrum of lipid radicals was reduced remarkably. The lipid radicals were formed by the hydroxyl radical generation system, not the superoxide anion generation system. The hydroxyl radical was found to be the direct active oxygen species that can generate lipid radicals as a result of .OH-mediated hydrogen atom abstraction. Superoxide anion radical stimulated the generation of hydroxyl radical via the iron-catalyzed reaction.

Animals↗

PubMedDate not supplied

Platelet-derived 12-hydroxyeicosatetraenoic acid plays an important role in mediating canine coronary thrombosis by regulating platelet glycoprotein IIb/IIIa activation.

BACKGROUND: In the thrombotic process of acute coronary syndromes, the pathophysiological role of thromboxane A2 via cyclooxygenase is well established; however, the role of 12-HETE via 12-lipoxygenase is little known. Therefore, we used OPC-29030, a novel specific inhibitor of 12-HETE synthesis, to test whether platelet-derived 12-HETE is involved in mediating cyclic flow variations (CFVs) and platelet aggregation in stenosed and endothelium-injured canine coronary arteries. METHODS AND RESULTS: After developing CFVs, dogs received a vehicle or OPC-29030 intravenously. Plasma and intraplatelet 12-HETE levels increased after CFVs. OPC-29030 but not vehicle reduced CFVs, which was associated with decreases in plasma and intraplatelet 12-HETE levels. Cessation of OPC-29030 restored CFVs in association with increases in plasma and intraplatelet 12-HETE levels. ADP and U46619 induced ex vivo platelet 12-HETE production and aggregation. After OPC-29030 administration, the ADP- and U46619-induced increases in ex vivo platelet 12-HETE production and aggregation were inhibited significantly. Platelet aggregation was linearly correlated with platelet 12-HETE production. OPC-29030 suppressed activation of human platelet glycoprotein IIb/IIIa. CONCLUSIONS: OPC-29030 reduced intraplatelet 12-HETE levels, resulting in the inhibition of coronary thrombosis in vivo in dogs. OPC-29030 inhibited human platelet glycoprotein IIb/IIIa activation in vitro. Thus, platelet-derived 12-HETE may play an important role in mediating thrombotic process.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗