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E Lenartowicz

Publications and source records attributed to E Lenartowicz.

15 recordsLinked to original sources
PubMed1999

Zinc as an inducer of the membrane permeability transition in rat liver mitochondria.

It is shown that 2-10 microM Zn2+ induces swelling of rat liver mitochondria incubated in a buffered sucrose medium either with valinomycin or with FCCP, Ca2+, ionophore A23187, oligomycin, and nigericin. This swelling was associated with the release of GSH from mitochondria. Both processes were sensitive to known inhibitors of the mitochondrial permeability transition (MPT), cyclosporin A, and Mg2+. Mitochondrial swelling induced by Zn2+ was also inhibited by rotenone, antymycin A, N-ethylmaleimide, butylhydroxytoluene, and spermine, whereas it was stimulated by tert-butyl hydroperoxide, diamide, and monobromobimane. It did not require the addition of phosphate. The same sensitivity to pH of the mitochondrial swelling induced by Zn2+ and by phenylarsine oxide suggests the same site of the interaction, namely, thiol groups. The ability of Zn2+ to induce mitochondrial swelling gradually decreased along with its increasing concentration above 10 microM. It is concluded that micromolar Zn2+ induces the MPT presumably by the interaction with cysteinyl residues. This process is independent of the mitochondrial membrane potential.

Animals↗
PubMed1996

Relation between the activities reducing disulfides and the protection against membrane permeability transition in rat liver mitochondria.

The oxidation of some mitochondrial sulfhydryl groups acts as an inducer of the mitochondrial membrane permeability transition. This membrane damage can be reversed by regeneration of the sulfhydryl groups. Hence the mitochondrial activities reducing disulfides are especially important for the defense against oxidative injury. The ability of isolated rat liver mitochondria to reduce disulfides was examined by the reduction of 5,5'-dithiobis-(2 nitro-benzoic acid) (DTNB), the reaction catalyzed by thioredoxin reductase and glutathione reductase. The incubation of mitochondria with DTNB induced their swelling which was prevented by EGTA, cyclosporin A, Mg2+, and pyrophosphate. The rate of DTNB reduction by mitochondria was compared with the changes in their volume and in the total content of their sulfhydryl groups. The reduction of DTNB was stimulated by cation chelators, both unphysiological and physiological, and was suppressed by up to 65% during mitochondrial swelling. In mitochondria treated with tert-butylhydroperoxide the total content of mitochondrial sulfhydryl groups decreased in the correlation with the suppression of DTNB reduction and with mitochondrial swelling. The content of nonprotein sulfhydryl groups was significantly lowered under all conditions applied. However, the protein sulfhydryl groups were preserved in the presence of EGTA or Mg2+ when the rate of DTNB reduction was relatively high and this was associated with the strong inhibition of mitochondrial swelling. It is suggested that during oxidative stress mitochondrial activities reducing disulfides are involved in the maintenance of sulfhydryl groups in mitochondria thus protecting their membranes against permeabilization.

Animals↗
PubMed1995

Enzymatic reduction of 5,5'-dithiobis-(2-nitrobenzoic acid) by lysate of rat liver mitochondria.

The oxidation of mitochondrial sulphydryl groups is known to increase the permeability of mitochondrial membranes and to be a key event in oxidative stress. Resistance to this damage is thought to involve thioredoxin reductase. In this study, the reduction of 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) by a lysate of rat liver mitochondria was used to assay the mitochondrial disulphide reducing capacity. NADPH-dependent reduction of DTNB was used to distinguish enzymatic reduction from the non-enzymatic reduction. Enzymatic reduction by the mitochondrial lysate was suppressed by DTNB at concentrations exceeding 0.25 mM and by pH above 7.0. It was strongly inhibited by Zn2+ and Mn2+ (IC50 about 2.5 and 20 microM, respectively) and was more weakly inhibited by Mg2+ and Ca2+ (IC50 about 1.8 and 2.1 mM, respectively). As a consequence of inhibition by divalent cations, the reaction was stimulated by both physiological (ATP, ADP, pyrophosphate and citrate) and non-physiological (EDTA and EGTA) chelators. Reduction of insulin disulphides by the mitochondrial lysate was dependent on the presence of a divalent cation chelator during the isolation of mitochondria and during the enzyme assay. Our results suggest that stimulation of mitochondrial disulphide reducing activity by lowered pH, as well as by increased levels of ATP, ADP and citrate, has the potential to contribute to the maintenance of mitochondrial sulphydryl groups under oxidative conditions.

Adenosine Diphosphate↗
PubMed1992

Ca(2+)-sensitive reduction of 5,5'-dithiobis-(2-nitrobenzoic acid) by rat liver mitochondria.

Energized rat liver mitochondria in the presence of EGTA reduced linearly 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) at the rate of 7 nmol SH/min per mg protein within more than 1 hour at 20 degrees C. The Km for DTNB, 1.4 mM, was decreased by Mg2+ and spermine to 0.5 and 0.7 mM, respectively. The reaction was suppressed under conditions of decreasing mitochondrial content of NADPH, was blocked by 1,3-bis-(2-chloroethyl)-1-nitrosourea, the inhibitor of disulfide reductases, and was sensitive to external free Ca2+ in the micromolar range. After lysis of mitochondria the reduction of DTNB required the addition of NADPH and EGTA and was inhibited by 1 mM sodium arsenite. These observations suggest that the reduction of DTNB by mitochondria is catalyzed by Ca(2+)-sensitive thioredoxin reductase (EC 1.6.4.5).

Animals↗
PubMed1991

Phenylarsine oxide induces the cyclosporin A-sensitive membrane permeability transition in rat liver mitochondria.

This paper reports an investigation on the effects of the hydrophobic, bifunctional SH group reagent phenylarsine oxide (PhAsO) on mitochondrial membrane permeability. We show that PhAsO is a potent inducer of the mitochondrial permeability transition in a process which is sensitive to both the oxygen radical scavanger BHT and to cyclosporin A. The PhAsO-induced permeability transition is stimulated by Ca2+ but takes place also in the presence of EGTA in a process that maintains its sensitivity to BHT and cyclosporin A. Our findings suggest that, at variance from other known inducers of the permeability transition, PhAsO reacts directly with functional SH groups that are inaccessible to hydrophilic reagents in the absence of Ca2+.

Animals↗
PubMed1990

A complex effect of arsenite on the formation of alpha-ketoglutarate in rat liver mitochondria.

This investigation presents disturbances of the mitochondrial metabolism by arsenite, a hydrophilic dithiol reagent known as an inhibitor of mitochondrial alpha-keto acid dehydrogenases. Arsenite at concentrations of 0.1-1.0 mM was shown to induce a considerable oxidation of intramitochondrial NADPH, NADH, and glutathione without decreasing the mitochondrial membrane potential. The oxidation of NAD(P)H required the presence of phosphate and was sensitive to ruthenium red, but occurred without the addition of calcium salts. Mitochondrial reactions producing alpha-ketoglutarate from glutamate and isocitrate were modulated by arsenite through various mechanisms: (i) both glutamate transaminations, with oxaloacetate and with pyruvate, were inhibited by accumulating alpha-ketoglutarate; however, at low concentrations of alpha-ketoglutarate the aspartate aminotransferase reaction was stimulated due to the increase of NAD+ content; (ii) the oxidation of isocitrate was stimulated at its low concentration only, due to the oxidation of NADPH and NADH; this oxidation was prevented by concentrations of citrate or isocitrate greater than 1 mM; (iii) the conversion of isocitrate to citrate was suppressed, presumably as a result of the decrease of Mg2+ concentration in mitochondria. Thus the depletion of mitochondrial vicinal thiol groups in hydrophilic domains disturbs the mitochondrial metabolism not only by the inhibition of alpha-keto acid dehydrogenases but also by the oxidation of NAD(P)H and, possibly, by the change in the ion concentrations.

Animals↗
PubMed1988

Significance of the alanine aminotransferase reaction in the formation of alpha-ketoglutarate in rat liver mitochondria.

The total production of alpha-ketoglutarate from glutamate and isocitrate was estimated in isolated rat liver mitochondria. Mitochondrial alanine aminotransferase converts glutamate to alpha-ketoglutarate [A.K. Groen et al. (1982) Eur. J. Biochem. 122, 87-93], thus participating in the net formation of the tricarboxylic acid cycle intermediates from glutamate. The present investigation indicates a significant contribution of the alanine aminotransferase reaction to glutamate oxidation by isolated rat liver mitochondria in the presence of bicarbonate. It amounted to 41-74 and 7-31% of the total utilization of glutamate in States 4 and 3, respectively, in various conditions in vitro, at pyruvate concentrations in the range of 0.1-10 mM. The participation of glutamate in the total production of alpha-ketoglutarate at physiological concentrations of glutamate, citrate, and isocitrate varied in the range of 72-82%. It was calculated that alpha-ketoglutarate formation by the reaction of alanine aminotransferase amounted to 30 and 5% of the total mitochondrial alpha-ketoglutarate production in States 4 and 3, respectively, at physiological concentrations of its precursors and in the presence of 0.5 mM malate and 0.1 mM pyruvate. It constituted 77-97% of the net production of the tricarboxylic acid cycle intermediates from glutamate in rat liver mitochondria. The importance of alpha-ketoglutarate production via the alanine aminotransferase reaction under various physiological conditions is discussed.

Alanine↗
PubMed1984

Intramitochondrial reductive carboxylation of 2-oxoglutarate in adipose tissue and its contribution to fatty acid synthesis.

The reductive carboxylation of 2-oxoglutarate was found to proceed in mitochondria of rat epididymal fat pads and rabbit perirenal adipose tissue at a rate similar to that in liver mitochondria. In rat fat pads the incorporation of 14C from [5-14C]2-oxoglutarate into fatty acids via the carboxylation was suppressed by butylmalonate by 30%. 2-Oxoglutarate and glutamate stimulated the incorporation into fatty acids of 14C from [2-14C]acetate in rat fat pads with the simultaneous reduction of tissue NADP. These effects persisted after inhibition of succinate dehydrogenase by malonate. It is concluded that in adipose tissue 2-oxoglutarate carboxylation proceeds in both the cytoplasm and mitochondria. Therefore, it can supply carbon atoms as well as NADPH for fatty acid synthesis.

Acetates↗
PubMed1976

The inhibition of isocitrate oxidation by palmitoyl-l-carnitine and palmitoyl-C0 A in rat liver mitochondria.

Palmitoyl-L carnitine decreases the oxidation of isocitrate in rat liver mitochondria in state 3 by 25-30%. Palmitoyl-L-carnitine acts as an additional substrate raising the rate of oxidative phosphorylation, NAD reduction and ATP/ADP ratio in mitochondria. Palmitoyl-CoA added to mitochondria oxidizing isocitrate in state 3 causes a strong inhibition of isocitrate oxidation and of oxidative phosphorylation and a considerable elevation of intramitochondrial NADH/NAD and ATP/ADP ratios. The effect of palmitoyl-CoA is dependent on its concentration and is competitive with ADP. Carnitine restores only oxidative phosphorylation, but the oxidation of isocitrate remains inhibited. Evidence is presented that the transport of isocitrate is not affected by palmitoyl-CoA is due to the inhibition of adenine nucleotide translocation. The kinetic studies of NAD-dependent isocitrate dehydrogenase in the soluble fraction of sonicated mitochondria revealed that the enzyme is very sensitive towards the inhibition by NADH and only very slightly affected by ATP (Ki for NADH and ATP are 0.017 and 3.6 mM respectively). On the basis of the kinetic data the relative contribution of NADH and ATP in the inhibition of isocitrate oxidation by fatty acids was calculated. It is concluded that the inhibition of isocitrate oxidation caused by palmitoyl-L-carnitine and palmitoyl-CoA is primarily due to the increased reduction of NAD, whereas the increase of ATP/ADP ratio is much less important.

Adenosine Diphosphate↗