Biphasic oxidation of mitochondrial NAD(P)H
The redox state of mitochondrial pyridine nucleotides is known to be important for structural integrity of mitochondria. In this work, we observed a biphasic oxidation of endogenous NAD(P)H in rat liver mitochondria induced by tert-butylhydroperoxide. Nearly 85% of mitochondrial NAD(P)H was rapidly oxidized during the first phase. The second phase of NAD(P)H oxidation was retarded for several minutes, appearing after the inner membrane potential collapse and mitochondria swelling. It was characterized by disturbance of ATP synthesis and dramatic permeabilization of the inner membrane to pyridine nucleotides. The second phase was completely prevented by 0.5 microM cyclosporin A or 0.2 mM EGTA or was significantly delayed by 25 microM butylhydroxytoluene or trifluoperazine. The obtained data suggest that the second phase resulted from oxidation of the remaining NADH via the outer membrane electron transport system of permeabilized mitochondria, leading to further oxidation of the remaining NADPH in a transhydrogenase reaction.
Available from: Jan Škrha jr.
- "When the MPT was allowed to proceed, we observed that the first rapid decrease in NAD(P)H was followed by a second decline of the autofluorescence that paralleled the course of swelling and was a consequence of the MPT rather than a cause (Fig. 6). Lemeshko  observed a similar biphasic course of NAD(P)H oxidation during the MPT of rat liver mitochondria challenged by tert-butyl hydroperoxide and suggested that the second phase of NAD(P)H oxidation could be catalyzed by the outer membrane NADH-dependent oxidoreductase system in conjunction with transhydrogenase. However, we noted that any changes of light scatter associated with mitochondrial swelling can result in artifacts in the measurement of NAD(P)H autofluorescence (Fig. 6D). "
[Show abstract] [Hide abstract]
ABSTRACT: The mitochondrial permeability transition (MPT) plays an important role in cell death. The MPT is triggered by calcium and promoted by oxidative stress, which is often catalyzed by iron. We investigated the induction of the MPT by physiological concentrations of iron. Isolated rat liver mitochondria were initially stabilized with EDTA and bovine serum albumin and energized by succinate or malate/pyruvate. The MPT was induced by 20μM calcium or ferrous chloride. We measured mitochondrial swelling, the inner membrane potential, NAD(P)H oxidation, iron and calcium in the recording medium. Iron effectively triggered the MPT; this effect differed from non-specific oxidative damage and required some residual EDTA in the recording medium. Evidence in the literature suggested two mechanisms of action for the iron: NAD(P)H oxidation due to loading of the mitochondrial antioxidant defense systems and uptake of iron to the mitochondrial matrix via a calcium uniporter. Both of these events occurred in our experiments but were only marginally involved in the MPT induced by iron. The primary mechanism observed in our experiments was the displacement of adventitious/endogenous calcium from the residual EDTA by iron. Although artificially created, this interplay between iron and calcium can well reflect conditions in vivo and could be considered as an important mechanism of iron toxicity in the cells.
[Show abstract] [Hide abstract]
ABSTRACT: The lignan otobaphenol, (8R,8'R,7R)-4'-hydroxy-5'-methoxy-3,4-methylenedioxy-2',7,8,8'-neolignan, extracted from Virola Aff. Pavonis leaves, completely inhibits at a concentration of 2.5 micro M the Fe(3+)-ascorbate-induced lipoperoxidation of rat liver mitochondria that was determined by oxygen consumption and accumulation of thiobarbituric acid-reactive species. At 25 micro M, it delays the mitochondrial permeability transition induced by tert-butyl hydroperoxide or Ca(2+), substantially inhibits the state 3 respiration, does not affect the state 4 respiration and the ADP/O ratio (with succinate), diminishes the rate of Ca(2+) uptake by mitochondria, and delays the ruthenium red-insensitive uncoupler-induced release of the loaded Ca(2+). Dose-dependent delaying of the calcium-induced swelling of mitochondria in the presence of otobaphenol nonlinearly correlates with its 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activity. At 75 micro M and higher, this lignan causes mitochondrial aggregation and is able to aggregate itself, without mitochondria. The formed aggregates of otobaphenol do not cause an aggregation of subsequently added mitochondria. Thus, otobaphenol seems to be a promising target to prevent the oxidative stress death of cells.
Available from: jbc.org
[Show abstract] [Hide abstract]
ABSTRACT: Mitochondrial permeability transition (MPT), which contributes substantially to the regulation of normal mitochondrial metabolism, also plays a crucial role in the initiation of cell death. It is known that MPT is regulated in a tissue-specific manner. The importance of MPT in the pancreatic beta-cell is heightened by the fact that mitochondrial bioenergetics serve as the main glucose-sensing regulator and energy source for insulin secretion. In the present study, using MIN6 and INS-1 beta-cells, we revealed that both Ca(2+)-phosphate- and oxidant-induced MPT is remarkably different from other tissues. Ca(2+)-phosphate-induced transition is accompanied by a decline in mitochondrial reactive oxygen species production related to a significant potential dependence of reactive oxygen species formation in beta-cell mitochondria. Hydroperoxides, which are indirect MPT co-inducers active in liver and heart mitochondria, are inefficient in beta-cell mitochondria, due to the low mitochondrial ability to metabolize them. Direct cross-linking of mitochondrial thiols in pancreatic beta-cells induces the opening of a low conductance ion permeability of the mitochondrial membrane instead of the full scale MPT opening typical for liver mitochondria. Low conductance MPT is independent of both endogenous and exogenous Ca(2+), suggesting a novel type of nonclassical MPT in beta-cells. It results in the conversion of electrical transmembrane potential into DeltapH instead of a decrease in total protonmotive force, thus mitochondrial respiration remains in a controlled state. Both Ca(2+)- and oxidant-induced MPTs are phosphate-dependent and, through the "phosphate flush" (associated with stimulation of insulin secretion), are expected to participate in the regulation in beta-cell glucose-sensing and secretory activity.
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.