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P Moldeus

Publications and source records attributed to P Moldeus.

At least 19 recordsLinked to original sources
PubMed1996

Age-related differences in the metabolism of sulphite to sulphate and in the identification of sulphur trioxide radical in human polymorphonuclear leukocytes.

Sulphite oxidation and sulphur trioxide radical formation were studied in polymorphonuclear leukocytes (PMNs) isolated from healthy young, old and centenarian donors and from patients with Down's syndrome. The sulphur radical formation measured by electron spin resonance spectroscopy-spin trapping (EPR-ST) was correlated with the activity of sulphite oxidase and with the rate of sulphite oxidation to sulphate by PMNs. Sulphite metabolism was studied both in resting, and phorbol myristate acetate (PMA) stimulated freshly isolated cells. The rate of sulphur trioxide radical formation was demonstrated by use of the spin trapping agent 5,5-dimethyl-1-pyroline-1-oxide (DMPO) with subsequent formation of an adduct. The intensity of adduct formation was most intense in cells with low sulphite oxidase activity, while a mixture of the adduct and of DMPO hydroxyl radical was mainly observed in cells with high sulphite oxidase activity. Furthermore, experiments carried out on purified sulphite oxidase showed that in the presence of sulphite the enzyme could also give rise to a DMPO-OH adduct. Sulphite oxidase activity in cells isolated from healthy young and old donors was positive correlated with both rates of sulphur trioxide radical formation and sulphite oxidation to sulphate, respectively. However, sulphite oxidase activity in cells isolated from centenarians and patients with Down's syndrome seems to loose partly its rate of oxidising sulphite to sulphate. The intensity of the sulphur centred radical adduct increased in the two latter groups of population and the radical observed was predominantly sulphur trioxide radical.

Aging↗
PubMed1996

Inverse relation between the concentration of low-density-lipoprotein vitamin E and severity of coronary artery disease.

Oxidation of low-density lipoprotein (LDL) is believed to play an important role in atherogenesis, and antioxidant vitamins are thought to protect against coronary artery disease (CAD). We investigated whether the vitamin E concentrations in serum and LDL were associated with the severity of CAD as assessed by a semiquantitative scoring system in which coronary angiograms are analyzed for the number and size of distinct stenotic lesions (global stenosis score). The study group consisted of 64 consecutive male survivors of myocardial infarction aged < 45 y. Lipid-adjusted serum and LDL vitamin E concentrations were significantly lower in the patients than in 35 age-matched male control subjects, whereas the absolute serum and LDL vitamin E concentrations did not differ significantly. No associations were found between the serum concentration or lipid-adjusted serum values of vitamin E and the stenosis score. In contrast, significant inverse correlation was found between the LDL vitamin E concentration, whether adjusted to the lipid (r=-0.477,P<0.001) or protein (r=-0.375, P<0.01) content of LDL, and the global coronary stenosis score. We conclude that a low LDL vitamin E concentration might play a role in the development of stenoses in coronary arteries and may contribute to clinically manifest CAD.

Adult↗
PubMed1995

Analysis of the reactivity of [14C]toluene diisocyanate (TDI) in an isolated, perfused lung model.

An isolated, perfused, guinea pig lung model was used to investigate the molecular events which occur when a 14C-labeled TDI vapor reaches the airways. Exposure concentrations of 0.2 and 0.7 ppm were tested. Perfusate composition included: Krebs Ringer buffer only, as well as buffer containing either guinea pig serum albumin, human serum albumin, or diluted guinea pig plasma. Radioactivity was detected in the perfusate within minutes of exposure, and following a delay, increased linearly. The rate of uptake was dependent on TDI concentration and the composition of the perfusate. Biochemical characterization of the state of the 14C-labeled material in the perfusate was performed. The distribution between low and high molecular weight reaction products was determined by molecular sieve fractionation and varied as a function of perfusate composition but no variability was observed as a function of time during the 45 min of exposure. An increase in nucleophile concentration in the perfusate was associated with both a higher percentage of conjugated products (from 15% with buffer only to 45% with diluted guinea pig plasma) and an increase in the rate of TDX uptake (from 0.5 microns Eq/min with buffer alone to 0.1 micrograms Eq/min with diluted GPSA as perfusate at 0.7 ppm). GC-MS analysis of the samples for free TDA, before and after acid hydrolysis, showed that the low molecular weight product(s), which represented from 55-85% of the circulating radioactivity, was composed of hydrolyzable and non-hydrolyzable conjugates and metabolites with approximately 4% of the label associated with free TDA. Although the distribution between high and low molecular weight species varies, this result is analogous to the findings from in vivo studies and suggests that the isolated, perfused lung (IVPL) system may be a useful tool in investigating the molecular mechanisms of isocyanate-induced disease and metabolic activity of the lung.

Animals↗
PubMed1993

Zonation of acetaminophen metabolism and cytochrome P450 2E1-mediated toxicity studied in isolated periportal and perivenous hepatocytes.

To study the mechanism of centrilobular damage developing in the centrilobular region after high doses of acetaminophen (APAP), its metabolism and toxicity were compared in periportal and perivenous hepatocytes isolated by digitonin/collagenase perfusion. Contrary to earlier reports, based on perfusions, no evidence for a periportal dominance of APAP sulfation could be observed. Glucuronidation, the dominant pathway of conjugation at high (5 mM) APAP concentration, was faster in perivenous cells. During primary culture, prolonged exposure (> or = 24 hr) to 5 mM APAP damaged perivenous cells, with a higher P450 2E1 level than periportal cells. When cells were isolated from ethanol-pretreated rats, to induce P450 2E1 levels specifically in the perivenous region, perivenous hepatocytes exhibited enhanced APAP vulnerability and extensive glutathione depletion. In contrast, corresponding periportal cells retained good viability. Isoniazid, an inhibitor of cytochrome P450 2E1, protected cells against APAP toxicity and prevented glutathione depletion. Induction of P450 2E1 also caused a 3-fold increase in the covalent binding of reactive intermediates from [14C]APAP, and this increase was mainly confined to perivenous cells. These results indicate that in rat liver there is only slight perivenous zonation of APAP conjugation and suggest that zone-specific APAP activation, mediated by the regional expression of ethanol-inducible cytochrome P450 2E1, is responsible for the characteristic centrilobular liver damage elicited by APAP.

Acetaminophen↗
PubMed1993

Thiols as peroxidase substrates.

The abilities of haem peroxidases to catalyse the oxidation of various thiols were studied using the spin-trapping electron spin resonance (ESR) technique. Myeloperoxidase, a neutrophil and monocyte enzyme, catalysed the oxidation of cysteamine, cysteine methyl, and ethyl ester and to some extent 2-mercaptoethanol and thioglycollic acid. This peroxidase poorly catalysed the oxidation of cysteine, N-acetylcysteine, penicillamine, and glutathione under the same conditions. The dependence on pH of peroxidase-catalysed thiol oxidation may indicate that the thiolate anion form is the actual peroxidase substrate. Another leucocyte peroxidase, eosinophil peroxidase, had similar catalytic properties toward thiols as myeloperoxidase. Lactoperoxidase (found in milk, saliva, and tears) and the plant horseradish peroxidase were, however, different from the aforementioned leucocyte peroxidases in their abilities to catalyse the oxidation of thiols.

Catalysis↗
PubMed1993

Analysis of lipoprotein diene formation in human serum exposed to copper.

The susceptibility of low density lipoprotein (LDL) to oxidative modification can be determined by analyzing the lag phase for initiation of diene formation in isolated LDL exposed to Cu2+. However, the applicability of this assay for clinical studies is limited by the requirement of a preparative ultracentrifugation of LDL and that the influence of water soluble antioxidants and other lipoproteins is not accounted for. The present paper describes a modification of this assay allowing determination of lag phase for lipoprotein diene formation in serum. The formation of dienes in serum exposed to Cu2+ begins following the consumption of serum alpha-tocopherol, correlates to the formation of thiobarbituric acid reactive substances (r = 0.987, n = 8), is inhibited by the addition of ascorbic acid and is absent in lipoprotein-deficient serum. It is also accompanied by an increased mobility of serum lipoproteins on agarose gel electrophoresis and with an ability of serum to displace isolated copper-oxidized LDL from binding sites mediating degradation in mouse peritoneal macrophages. The coefficient of variance of the analysis is below 3%. It is concluded that this technique allows analysis of lipoprotein oxidation susceptibility in serum samples and may prove to be useful in clinical analysis of the lipoprotein oxidation susceptibility.

Animals↗
PubMed1992

Vasoconstriction and bronchoconstriction induced by 2,5-di-(tert-butyl)1,4-benzohydroquinone, an endoplasmic reticular Ca2+-ATPase inhibitor, in isolated and perfused rat lung.

The microsomal Ca(2+)-ATPase inhibitor 2,5-di-(tert-butyl)-1,4-benzohydroquinone (tBuBHQ) induced bronchoconstriction and vasoconstriction in the isolated perfused and ventilated rat lung. These effects were accompanied by increased levels of thromboxane and prostacyclin in the effluent perfusate. The effect of tBuBHQ was inhibited by L-655,240, a thromboxane receptor antagonist, indicating thromboxane-A2-mediated bronchoconstriction and vasoconstriction. Accordingly, the cyclooxygenase inhibitor indomethacin largely blocked the effects of tBuBHQ. The involvement of a phospholipase in the generation of thromboxane A2(TXA2) was supported by dibucaine protection on tBuBHQ effects. The results from this study indicate that tBuBHQ, probably by inhibiting the microsomal Ca(2+)-ATPase, can trigger the arachidonic acid cascade leading to the formation of TXA2, which in turn causes bronchoconstriction and vasoconstriction in rat lung.

Animals↗
PubMed1991

Sodium nitrite-stimulated metabolic activation of benzo[a]pyrene 7,8-dihydrodiol in human polymorphonuclear leukocytes.

Sodium nitrite was shown to enhance the metabolism of trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (BP-7,8-diol) to 7/8,9,10- and 7,10/8,9-tetrahydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (tetraols) in phorbol myristate acetate (PMA)-stimulated polymorphonuclear leukocytes (PMNs). The production of these tetraols implicates the intermediate formation of the corresponding trans-7,8-dihydroxy-9,10-epoxy-7,8-9,10-tetrahydrobenzo[a]pyrene (anti-BPDE). A 2- to 3-fold increase in the tetraol yield was observed in the presence of nitrite in excess of 1 mM. Sodium azide, an inhibitor of myeloperoxidase and catalase, reduced the nitrite-stimulated metabolism of BP-7,8-diol in PMA-activated leukocytes. Diphenylene iodonium sulphate, a NADPH-oxidase inhibitor, lowered the production of tetraols in PMA-stimulated leukocytes both in the absence and presence of nitrite. Additionally, nitrite markedly enhanced the covalent binding of metabolites derived from [3H](-)-BP-7,8-diol to leukocyte proteins as well as to DNA present extracellularly. The nitrite-stimulated covalent binding to both proteins and DNA was inhibited by the presence of sodium azide. The mechanism underlying the effect of nitrite on the metabolism of BP-7,8-diol to reactive intermediates in PMA-activated human polymorphonuclear leukocytes is not known. However, the results are compatible with a peroxidase-dependent mechanism although other possible pathways may contribute to the enhanced rate of metabolism.

Biotransformation↗
PubMed1990

Quinone imine-induced Ca2+ release from isolated rat liver mitochondria.

Incubation of Ca2(+)-loaded rat liver mitochondria with N-acetyl-p-benzoquinone imine (NAPQI) or its two dimethylated analogues resulted in a concentration dependent Ca2+ release, with the following order of potency: 2,6-(Me)2-NAPQI greater than NAPQI greater than 3,5-(Me)2-NAPQI. The quinone imine-induced Ca2+ release was associated with NAD(P)H oxidation and was prevented when NAD(P)+ reduction was stimulated by the addition of 3-hydroxybutyrate. Mitochondrial glutathione was completely depleted within 0.5 min by all three quinone imines, even at low concentrations that did not result in Ca2+ release. Depletion of mitochondrial GSH by pretreatment with 1-chloro-2,4-dinitrobenzene enhanced quinone imine-induced NAD(P)H oxidation and Ca2+ release. However, 3-hydroxybutyrate protected from quinone imine-induced Ca2+ release in GSH-depleted mitochondria. Mitochondrial membrane potential was lost after the addition of quinone imines at concentrations that caused rapid Ca2+ release; however, subsequent addition of EGTA led to the complete recovery of the transmembrane potential. In the absence of Ca2+, the quinone imines caused only a small and transient loss of the transmembrane potential. Taken together, our results suggests that the quinone imine-induced Ca2+ release from mitochondria is a consequence of NAD(P)H oxidation rather than GSH depletion, GSSG formation, or mitochondrial inner membrane damage.

Animals↗
PubMed1989

Effects of the new nitrosourea derivative, fotemustine, on the glutathione reductase activity in rat tissues in vivo and in isolated rat hepatocytes.

Fotemustine, a new clinically active nitrosourea, is demonstrated herein to be a poor inhibitor of glutathione reductase activity from rat liver, lung and kidney cytosols. In order to show that an intracellular step of activation does not lead to a toxic intermediary metabolite, rat hepatocytes were incubated with fotemustine. Their glutathione-related pathways were checked and shown not to be altered, while under similar experimental conditions BCNU was shown to be dramatically harmful. Furthermore, association of fotemustine with a H2O2 production leading drug, diquat, was shown to be inefficient--while BCNU is efficient--in potentiating the diquat toxicity. Considering the role of glutathione level in the detoxification of mutagens and carcinogens, the advantage of fotemustine over BCNU in therapeutic use seems substantiated.

Animals↗
PubMed1989

Detection of oxygen radicals during reperfusion of intestinal cells in vitro.

The nitroxide OXANO. (2-Ethyl-2,5,5-trimethyl-3-oxazolidinoxyl) which in its reduced form, OXANOH (2-Ethyl-1-hydroxy-2,5,5-trimethyl-3-oxazolidine), is capable of reacting with short-lived radicals, forming a secondary stable radical, was used for ESR-detection of radical production in isolated cells. The properties of OXANO. and OXANOH in terms of stability in cellular and subcellular systems, membrane permeability and effects on cellular viability were evaluated. Ischemia and reperfusion was simulated in vitro in a preparation of cells from rat intestinal mucosa by incubation at high density (4 X 10(8) cells/ml) under an atmosphere of nitrogen for 25 min and resuspended with fresh oxygenated buffer containing 5 mM OXANOH. A significant increase in radical formation during the 15 min reperfusion period studied was obtained in cells exposed to ischemia compared to control cells incubated at normal density under an atmosphere of oxygen. The addition of 5 microM of the scavenging enzyme superoxide dismutase reduced the radical formation by 50%. The time sequence of the superoxide formation was calculated as the difference in radical production in the presence and absence of superoxide dismutase.

Animals↗
PubMed1988

Cellular metabolism of arsenocholine.

The biotransformation of arsenocholine and arsenobetaine, which are organic arsenic compounds present in certain aquatic organisms, has been studied in vitro using synthetic reference substances. Incubation of arsenocholine with different liver cell fractions showed arsenocholine to be biotransformed only in presence of the mitochondrial fraction. The biotransformation products were arsenobetaine aldehyde, arsenobetaine, trimethylarsine oxide and trimethylarsine. Arsenobetaine was the major metabolite and it was formed via arsenobetaine aldehyde. Trimethylarsine oxide was formed via a side reaction from arsenobetaine aldehyde. Further reduction of trimethylarsine oxide, produced trimethylarsine. In vitro studies of arsenobetaine, did not show any formation of trimethylarsine oxide or trimethylarsine. Furthermore, cytotoxicity of arsenobetaine or arsenocholine in isolated hepatocytes was not observed.

Alcohol Oxidoreductases↗
PubMed1987

Cytotoxicity of the redox cycling compound diquat in isolated hepatocytes: involvement of hydrogen peroxide and transition metals.

Diquat is a hepatotoxin whose toxicity in vivo and in vitro is mediated by redox cycling and greatly enhanced by pretreatment with 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase. The mechanism by which redox cycling mediates diquat cytotoxicity is unclear, however. Here, we have attempted to examine the roles of three potential products of redox cycling, namely superoxide anion radical (O2-.), hydrogen peroxide (H2O2), and hydroxyl radical (.OH), in the toxicity of diquat to BCNU-treated isolated hepatocytes. Addition of high concentrations of catalase, but not superoxide dismutase, to the incubations provided some protection against the toxic effect of diquat, but much better protection was observed when catalase was added in combination with the iron chelator desferrioxamine. Addition of desferrioxamine alone also provided considerable protection, whereas the addition of copper ions enhanced diquat cytotoxicity. Taken together, these results indicate that both H2O2 and the transition metals iron and copper could play major roles in the cytotoxicity of diquat. The role of O2-. remains less clear, however, but studies with diethylenetriaminepentaacetic acid indicate that O2-. is unlikely to significantly contribute to the reduction of Fe3+ to Fe2+. The hydroxyl radical or a related species seems the most likely ultimate toxic product of the H2O2/Fe2+ interaction, but hydroxyl radical scavengers afforded only minimal protection.

Animals↗
PubMed1986

Role of redox cycling and lipid peroxidation in bipyridyl herbicide cytotoxicity. Studies with a compromised isolated hepatocyte model system.

The role of active oxygen species and lipid peroxidation in the toxic effects of diquat, paraquat and other bipyridyl herbicides remains controversial. In vitro studies have shown that these compounds are potent generators of active oxygen species by redox cycling and that they stimulate lipid peroxidation. In vivo studies have failed, however, to show clear evidence of lipid peroxidation resulting from toxic exposures to these compounds. We have directly compared the abilities of three bipyridyl herbicides, diquat (DQ), paraquat (PQ) and benzyl viologen (BV), to generate superoxide anion radical (O2-.) in rat liver microsomes and H2O2 in hepatocytes and correlated this with their relative toxicities to a compromised isolated hepatocyte system. DQ was the most potent generator of O2-. and H2O2, being slightly more potent than BV and much better than PQ. This ability of the bipyridyls to generate active oxygen was positively correlated with the ability to induce toxicity in hepatocytes pretreated with 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) to inhibit their glutathione reductase activity, i.e. DQ greater than BV greater than PQ. DQ caused a rapid depletion of cellular GSH and a concomitant increase in GSSG in this system. Toxicity, measured as loss of plasma membrane integrity, was pronounced after only 30-60 min of incubation and was accompanied by a significant increase in lipid peroxidation. The onset of lipid peroxidation could not be separated temporally from the expression of toxicity. However, the total inhibition of lipid peroxidation by the antioxidants Trolox C, promethazine and N,N'-diphenyl-p-phenylenediamine only delayed toxicity, indicating that, even though lipid peroxidation may play some role in enhancing bipyridyl herbicide toxicity, it is not essential for the toxicity to manifest itself.

Animals↗
PubMed1986

Mechanism and relevance of glutathione mutagenicity.

The ubiquitous tripeptide glutathione (GSH) has previously been shown to be mutagenic to Salmonella typhimurium TA100 when incubated with kidney subcellular fractions at physiological concentrations (Glatt et al., 1983). Here we report that the mutagenic effect of GSH can be inhibited by the use of the gamma-glutamyl-transpeptidase (gamma-GT) inhibitor anthglutin and by the metal chelators bathocuproine, EDTA and diethyldithiocarbamate. As the chelating agents did not inhibit gamma-GT activity this suggested that the mechanism underlying the mutagenic effect of GSH was at least a two-step process, dependent upon the cleavage of GSH by gamma-GT and the presence of either free transition metals or those contained in enzymes such as glutathione oxidase. As gamma-GT is located on the outer surface of kidney tubule cells and is therefore exposed to relatively low concentrations of GSH, and the precise physiological control of levels of transition metals, this mechanism is unlikely to occur in vivo.

Animals↗