Mechanism of citrinin-induced dysfunction of mitochondria. IV—Effect on Ca2+ transport

The effect of citrinin on Ca²⁺ transport was studied in isolated kidney cortex and liver mitochondria, and baby hamster kidney cultured cells. The mycotoxin significantly inhibited the activity of 2-oxoglutarate and pyruvate dehydrogenases in both kidney cortex and liver mitochondria. Citrinin promoted a decrease in the velocity and in the total capacity of Ca²⁺ uptake, in both mitochondria. Apparently, citrinin acts by a mechanism similar to ruthenium red. In intact cultured cells, citrinin also had a preferential effect on mitochondrial Ca²⁺ fluxes. Citrinin promoted a marked decrease in the Ca²⁺ level in the mitochondrial matrix, whereas that of the extramitochondiral fraction became less affected. All the observed effects were dependent on the citrinin concentration.     

Cubebin and derivatives as inhibitors of mitochondrial complex I. Proposed interaction with subunit B8

The effects on mitochondrial respiration and complex I NADH oxidase activity of cubebin and derivatives were evaluated. The compounds inhibited the state 3 glutamate/malate-supported respiration of hamster liver mitochondria with IC₅₀ values ranging from 12.16 to 83.96 μM. NADH oxidase reaction was evaluated in submitochondrial particles. The compounds also inhibited this activity, showing the same order of potency observed for effects on state 3 respiration, as well as a tendency towards a non-competitive type of inhibition (K_I values ranging from 0.62 to 16.1 μM). A potential binding mode of these compounds with complex I subunit B8, assessed by docking calculations, is proposed.     

Functional activity and ultrastructure of mitochondria isolated from myocardial apoptotic tissue

Apoptosis in myocardial tissue slices was induced by extended incubation under anoxic conditions. Mitochondria were isolated from the studied tissue. A new method of isolation of mitochondria in special conditions by differential centrifugation at 1700, 10,000, and 17,000g resulted in three fractions of mitochondria. According to the data of electron microscopy the heavy mitochondrial fraction (1700g) consisted of mitochondrial clusters only, the middle mitochondrial fraction (10,000g) consisted of mitochondria with typical for isolated mitochondria ultrastructure, and the light fraction consisted of small mitochondria (2 or 3 cristae) of various preservation. The heavy fraction contained unusual structural elements that we detected earlier in apoptotic myocardial tissue—small electron-dense mitochondria incorporated in bigger mitochondria. The structure of small mitochondria from the light fraction corresponded to that of the small mitochondria from these unusual elements—mitochondrion in mitochondrion. The most important functions of isolated mitochondria are strongly inhibited when apoptosis is induced in our model. The detailed study of the activities of the two fractions of the apoptotic mitochondria showed that the system of malate oxidation is completely altered, the activity of cytochrome c as electron carrier is partly inhibited, while succinate oxidase activity is completely preserved (complexes II, III, and IV of the respiration chain). Succinate oxidase activity was accompanied by high permeability of the internal membrane for protons: the addition of uncoupler did not stimulate respiration. ATP synthesis in mitochondria was inhibited. We demonstrated that in our model of apoptosis cytochrome c remains in the intermembrane space, and, consequently, is not involved in the cascade of activation of effector caspases. The possible mechanisms of induction of apoptosis during anoxia are discussed.     

Fasciola hepatica: cytochrome c oxidoreductases and effects of oxygen tension and inhibitors

Studies were made on the mechanism of respiration in Fasciola hepatica (Trematoda). Respiration was found to be dependent on the oxygen tension. The respiratory enzyme systems, NADH-cytochrome c oxidoreductase (EC 1.6.2.1), succinate-cytochrome c oxidoreductase (EC 1.3.99.1) NADH oxidase and cytochrome c-oxygen oxidoreductase (EC 1.9.3.1) were detected in a mitochondrial preparation, the NADH oxidase activity being markedly stimulated by addition of mammalian cytochrome c. Amytal and rotenone inhibited NADH oxidase activity. Antimycin A inhibited succinoxidase activity only at relatively high concentrations. Azide was inhibitory at high concentrations. However, cyanide was found to stimulate respiration. Hydrogen peroxide was found to be an end product of respiration in F. hepatica.     

Respiration of the Roots of Flood-Tolerant and Flood-Intolerant Senecio Species: Affinity for Oxygen and Resistance to Cyanide

The affinity of respiration for oxygen in the roots of six Senecio species studied was low compared with the affinity of cytochrome oxidase for oxygen. Half saturation values of approximately 22 μM oxygen were measured. Root respiration was to a large extent insensitive to cyanide in flood-tolerant as well as in flood-sensitive species. The evidence presented suggests that high activity of salicylhydroxamic acid (SHAM)-sensitive oxidase in Senecio roots was the basis for the low oxygen affinity and for the high cyanide-insensitivity of root respiration in the Senecio species. Methods are described to determine the in vivo activity of the SHAM-sensitive oxidase. It was estimated that it contributed 70% to the total root respiration. The presence of SHAM-sensitive oxidase activity could explain a higher efficiency of root growth respiration under a low oxygen tension if this alternate oxidase was inhibited at a low oxygen concentration in the root medium. However, the SHAM-sensitive oxidase was not specifically involved in either growth respiration or maintenance respiration. Its significance in regulation of the redox state of the cells is discussed.     

Mitochondrial sensitivity to AZT

The possibility of tissue-specific effects regarding mitochondrial sensitivity to AZT was evaluated in this study. When mitochondria isolated from liver, kidney, skeletal and cardiac muscle were oxidizing glutamate, a dose-dependent inhibition by AZT of state 3 respiration was observed; using succinate as substrate the inhibition occurred only in skeletal and cardiac muscle mitochondria. The same results were obtained with FCCP-uncoupled mitochondria. NADH oxidase of intact and disrupted mitochondria, isolated from all four tissues was strongly inhibited. Succinate oxidase activity was inhibited by AZT only in intact mitochondria from skeletal and cardiac muscles, suggesting the involvement of succinate transport systems. Similarly, inhibition by the drug of the hydrolytic activity of H⁺-ATPase was observed only in mitochondria of these tissues. These effects taken together, indicate a tissue/carrier-specific inhibition in vitro, although its precise mechanism requires further research. © 1998 John Wiley & Sons, Ltd.     

Respiratory Activity and Naphthoquinone Synthesis in the Fungus Fusarium decemcellulare Exposed to Oxidative Stress

The effect of oxidants (hydrogen peroxide and juglone) on the growth, respiration, and naphthoquinone synthesis in the fungus Fusarium decemcellulare was studied. The addition of the oxidants to the exponential-phase fungus inhibited cell respiration (either partially or completely, depending on the oxidant concentration), culture growth, and naphthoquinone synthesis. The treatment of fungal cells with nonlethal concentrations of H₂O₂ (below 0.25 mM) and juglone (below 0.1 mM) induced the resistance of cell respiration to cyanide. The residual respiration in the presence of cyanide could be inhibited by benzohydroxamic acid, indicating the occurrence of alternative oxidase. Increased concentrations of oxidants (0.25 mM juglone and 0.5 mM H₂O₂) rapidly and irreversibly inhibited cell respiration. These observations suggest that the mitochondrial respiratory chain of fungal cells exposed to oxidative stress is subject to the action of active oxygen species. The treatment of fungal cells with nonlethal concentrations of H₂O₂ and juglone activated cellular glutathione reductase and glucose-6-phosphate dehydrogenase, which are protective enzymes against oxidative stress.     

Inhibition of mitochondrial function in isolated rat liver mitochondria by azole antifungals

Ketoconazole is an imidazole oral antifungal agent with a broad spectrum of activity. Ketoconazole has been reported to cause liver damage, but the mechanism is unknown. However, ketoconazole and a related drug, miconazole, have been shown to have inhibitory effects on oxidative phosphorylation in fungi. Fluconazole, another orally administered antifungal azole, has also been reported to cause liver damage despite its supposedly low toxicity profile. The primary objective of this study was to evaluate the metabolic integrity of adult rat liver mitochondria after exposure to ketoconazole, miconazole, fluconazole, and the deacetylated metabolite of ketoconazole by measuring ADP-dependent oxygen uptake polarographically and succinate dehydrogenase activity spectrophotometrically. Ketoconazole, N-deacetyl ketoconazole, and miconazole inhibited glutamate-malate oxidation in a dose-dependent manner such that the 50% inhibitory concentration (I₅₀ was 32, 300, and 110 μM, respectively. In addition, the effect of ketoconazole, miconazole, and fluconazole on phosphorylation coupled to the oxidation of pyruvate/malate, ornithine/malate, arginine/malate, and succinate was evaluated. The results demonstrated that ketoconazole and miconazole produced a dose-dependent inhibition of NADH oxidase in which ketoconazole was the most potent inhibitor. Fluconazole had minimal inhibitory effects on NADH oxidase and succinate dehydrogenase, whereas higher concentrations of ketoconazole were required to inhibit the activity of succinate dehydrogenase. N-deacetylated ketoconazole inhibited succinate dehydrogenase with an I₅₀ of 350 μM. In addition, the reduction of ferricyanide by succinate catalyzed by succinate dehydrogenase demonstrated that ketoconazole caused a dose-dependent inhibition of succinate activity (I₅₀ of 74 μM). In summary, ketoconazole appears to be the more potent mitochondrial inhibitor of the azoles studied; complex I of the respiratory chain is the apparent target of the drug's action. © 1997 John Wiley & Sons, Inc.     

Enhanced cytochrome oxidase activity and modification of lipids in heart mitochondria from hyperthyroid rats

In order to further investigate the mechanism regulating the control of mitochondrial respiration by thyroid hormones, the effect of the hyperthyroidism on the kinetic characteristics of cytocrome c oxidase in rat heart mitochondria was studied. Mitochondrial preparations from both control and hyperthyroid rats had equivalent K_m values for cytochrome c, while the maximal activity of cytochrome oxidase was significantly increased (by around 30%) in mitochondrial rats. This enhanced activity of cytochrome oxidase was associated to a parallel increases in mitochondrial State 3 respiration. The hormone treatment resulted in a decrease in the flux control coefficient of the oxidase. The enhanced activity of cytochrome oxidase in hyperthyroid rats does not appear to be dependent on an increases in the mass of this enzyme complex in that the heme aa₃ content was equivalent in both hyperthyroid and control preparations. The Arrhenius plot characteristics differ for cytochrome oxidase activity in mitochondria from hyperthyroid rats as compared with control rats in the breakpoint of the biphasic plot is shifted to a lower temperature. Cardiolipin content was significantly increased in mitochondrial preparations from hyperthyroid rats, while there were no significant alterations in the fatty acid composition of cardiolipin of control and hyperthyroid preparations. The results support the conclusion that the enhanced cytochrome oxidase activity in heart mitochondrial preparations from hyperthyroid rats is due to a specific increase in the content of cardiolipin.     

Studies on the Residual Respiration not Inhibited by KCN Plus Hydroxamic Acid in Tobacco Callus Cultures

A residual respiration not inhibited by KCN plus hydroxamic acid had been observed in many plant organs and tissues. The relative O2 uptake of it was 20–30% of total respiration in tobacco callus cultures. However, there is no report concerning the nature of the residual respiration and its localization in cell. The object of this study is to elucidate the characteris- tics of this residual respiration and its localization in cell. The experimental results are as follows: 1. The additions of glycolate and glyoxylate cause a marked rise in residual respiration not inhibited by KCN (or NaN3) plus m-CLAM. 2. The O2 uptake induced by glycolate and the residual respiration is inhibited by the addition of α-hydroxy ethanesulfonate. 3. The mitochondrial respiration is completely inhibited by KCN plus m-CLAM, but no effect by adding of glycolate. 4. Oxidation reactions of glycolate and glyoxylate in supernatant are observed after mitochondria are removed. Based on the above results, it is suggested that the residual respiration not inhibited by KCN plus m-CLAM in tobacco callus cultures is primarily catalyzed by glycolic acid oxidase localized within microbodies.     

Spegazzinine,a new inhibitor of mitochondrial oxidative phosphorylation

The effects of spegazzinine, a dihydroindole alkaloid, on mitochondrial oxidative phosphorylation were studied.Spegazzinine inhibited coupled respiration and phosphorylation in rat liver mitochondria. The I₅₀ was 120 μM. Uncouplers released the inhibition of coupled respiration. Arsenate-stimulated mitochondrial respiration was partially inhibited by spegazzinine. The stimulation of mitochondrial respiration by Ca²⁺ and the proton ejection associated with the ATP-dependent Ca²⁺ uptake were not affected by the alkaloid.Oxidative phosphorylation and the P_i-ATP exchange reaction of phosphorylating beef heart submitochondrial particles were strongly inhibited by spegazzinine (I₅₀, 50 μM) while the ATP-dependent reactions, reduction of NAD⁺ by succinate and the pyridine nucleotides transhydrogenase were less sensitive (I₅₀, 125 μM). Oxygen uptake by submitochondrial particles was not affected.The 2,4-dinitrophenol-stimulated ATPase activity of rat liver mitochondria was not affected by 300 μM spegazzinine, a concentration of alkaloid that completely inhibited phosphorylation. However, higher concentrations of spegazzinine did partially inhibit it. The ATPase activities of submitochondrial particles, insoluble and soluble ATPases were also partially inhibited by high concentrations of spegazzinine.The inhibitory properties of spegazzinine on energy transfer reactions are compared with those of oligomycin, aurovertin and dicyclohexylcarbodiimide. It is concluded that spegazzinine effects are very similar to the effects of aurovertin and that its site of action may be the same or near the site of aurovertin.     

The effect of phenolic compounds on the activity of respiratory chain enzymes and on respiration and phosphorylation activities of potato tuber mitochondria

The effect of derivatives of benzoic and cinnamic acids, quereetin,p-benzoquinone, and 2,5-dimethylbenzoquinone on oxygen consumption mitoehondrial suspensions and on the activity of some respiratory chain enzymes was studied. Benzoquinone and 2,5-dimethylbenzoquinone highly significantly inhibited the respiration and phosphorylation rates and malate- and succinate dehydrogenase activities. Chlorogenic acid, similarly as the quinones, very significantly inhibited the activities of the studied dehydrogenases but did not affect cytochrome oxidase. Oxygen consumption by intact mitochondria was not inhibited, only the oxidativo phosphorylation was significantly uncoupled. Quereetin significantly enhanced dehydrogenase activities and completely inhibited cytochrome oxidase activity. The respiration and phosphorylation activities of the mitochondria were significantly inhibited by quereetin. The effect of the other phenolic compounds studied on respiration and phosphorylation activities was not significant. Succinate dehydrogenase activity was the most affected enzyme among the respiratory chain enzymes. It was significantly inhibited by all the above phenolic compounds at 1^-4M or 5 10^-5M concentrations with the exception of gallic acid.     

The use of diaminobenzidine (DAB) for the histochemical demonstration of cytochrome oxidase activity in unfixed plant tissues

Summary Cytochrome oxidase activity was demonstrated in unfixed root segments from Lupinus albus at the ultrastructural level using the osmiophilic reagent 3,3-diaminobenzidine (DAB). Precipitate, the formation of which was completely inhibited by 0.01 M KCN, and observed almost entirely on mitochondrial cristae, is considered to be produced by cytochrome oxidase activity. Heterogeneity of mitochondria as to the intensity of the reaction in the same cell could not be established with certainity. However, mitochondria of the root tip cells and cells belonging to the plerome consistently did not show histochemically demonstrable cytochrome oxidase activity.     

Respiratory Chain of a Pathogenic Fungus, Microsporum gypseum: Effect of the Antifungal Agent Pyrrolnitrin

Pyrrolnitrin has been reported to inhibit Bacillus megaterium primarily by forming complexes with phospholipids and to block electron transfer of Saccharomyces cerevisiae between succinate or reduced nicotinamide adenine dinucleotide (NADH) and coenzyme Q. We found that pyrrolnitrin inhibited respiration of conidia of Microsporum gypseum. In mitochondrial preparations, pyrrolnitrin strongly inhibited respiration and the rotenone-sensitive NADH-cytochrome c reductase. The rotenone-insensitive NADH-cytochrome c reductase, the succinate-cytochrome c reductase, and the reduction of dichlorophenolindophenol by either NADH or succinate were inhibited to a lesser extent. However, the activity of cytochrome oxidase was not affected by pyrrolnitrin. The extent of reduction of flavoproteins by NADH and succinate, measured at 465 - 510 nm, was unaltered; however, the reduction of cytochrome b, measured at 560 - 575 nm, was partially inhibited by pyrrolnitrin. The level of totally reduced cytochrome b was restored with antimycin A. We, therefore, concluded that the primary site of action of this antifungal antibiotic is to block electron transfer between the flavoprotein of the NADH-dehydrogenase and cytochrome b segment of the respiratory chain of M. gypseum.     

Inhibition and stimulation of root respiration in pisum and plantago by hydroxamate : its consequences for the assessment of alternative path activity

The contribution of the alternative pathway in root respiration of Pisum sativum L. cv Rondo, Plantago lanceolata L., and Plantago major L. ssp major was determined by titration with salicylhydroxamate (SHAM) in the absence and presence of cyanide. SHAM completely inhibited the cyanide-resistant component of root respiration at 5 to 10 millimolar with an apparent K_i of 600 micromolar. In contrast, SHAM enhanced pea root respiration by 30% at most, at concentrations below 15 millimolar. An unknown oxidase appeared to be responsible for this stimulation. Its maximum activity in the presence of low SHAM concentrations (1-5 millimolar) was 40% of control respiration rate in pea roots, since 25 millimolar SHAM resulted in 10% inhibition. In plantain roots, the maximum activity was found to be 15%. This hydroxamate-activated oxidase was distinct from the cytochrome path by its resistance to antimycin. The results of titrations with cyanide and antimycin indicated that high SHAM concentrations (up to 25 millimolar) block the hydroxamate-activated oxidase, but do not affect the cytochrome path and, therefore, are a reliable tool for estimating the activity of the alternative path in vivo. A considerable fraction of root respiration was mediated by the alternative path in plantain (45%) and pea (15%), in the latter because of the saturation of the cytochrome path.     

Existence of aa 3-Type Ubiquinol Oxidase as a Terminal Oxidase in Sulfite Oxidation of Acidithiobacillus thiooxidans

It was found that Acidithiobacillus thiooxidans has sulfite:ubiquinone oxidoreductase and ubiquinol oxidase activities in the cells. Ubiquinol oxidase was purified from plasma membranes of strain NB1-3 in a nearly homogeneous state. A purified enzyme showed absorption peaks at 419 and 595 nm in the oxidized form and at 442 and 605 nm in the reduced form. Pyridine ferrohaemochrome prepared from the enzyme showed an α-peak characteristic of haem a at 587 nm, indicating that the enzyme contains haem a as a component. The CO difference spectrum of ubiquinol oxidase showed two peaks at 428 nm and 595 nm, and a trough at 446 nm, suggesting the existence of an aa ₃-type cytochrome in the enzyme. Ubiquinol oxidase was composed of three subunits with apparent molecular masses of 57 kDa, 34 kDa, and 23 kDa. The optimum pH and temperature for ubiquinol oxidation were pH 6.0 and 30 °C. The activity was completely inhibited by sodium cyanide at 1.0 mM. In contrast, the activity was inhibited weakly by antimycin A₁ and myxothiazol, which are inhibitors of mitochondrial bc ₁ complex. Quinone analog 2-heptyl-4-hydoroxyquinoline N-oxide (HOQNO) strongly inhibited ubiquinol oxidase activity. Nickel and tungstate (0.1 mM), which are used as a bacteriostatic agent for A. thiooxidans-dependent concrete corrosion, inhibited ubiquinol oxidase activity 100 and 70% respectively.     

STUDIES ON CELL METABOLISM AND CELL DIVISION : V. CYTOCHROME OXIDASE ACTIVITY IN THE EGGS OF ARBACIA PUNCTULATA

1. An enzyme capable of oxidizing reduced cytochrome c (i.e. a cytochrome oxidase) has been obtained from Arbacia eggs. In 0.02 M hydroquinone, the cytochrome oxidase was half activated at a cytochrome c concentration of approximately 4 x 10^–6 M. The concentration of the cytochrome oxidase was found to be nearly the same in unfertilized and fertilized eggs, the amount of the enzyme—as measured by means of its activity toward cytochrome c as a representative substrate—being more than sufficient to account for the highest rate of oxygen utilization yet observed in the intact, living, fertilized eggs, and of the same order as that in certain rat tissues. 2. The Arbacia cytochrome oxidase was strongly inhibited by carbon monoxide in the dark, the inhibition being almost completely reversed by light. The inhibition constant was not greatly altered by variation in the concentration of cytochrome c or the concentration of hydroquinone used as reductant for the cytochrome c, having a value of 3 to 5 under the conditions used. The inhibition constant was about 2 with p-phenylenediamine as reductant for the cytochrome c, but apparently had the surprisingly low value of about 0.5 with 0.02 M cysteine as reductant. 3. The cytochrome oxidase was completely inhibited by sufficiently high concentrations of sodium cyanide, sodium azide, and sodium sulfide. It was also completely inhibited in 0.6 M sodium chloride. It was not inhibited by two inhibitors of copper containing enzymes, 8-hydroxyquinoline and sodium diethyldithiocarbamate. It was also not significantly inhibited by 2,4-dinitrothymol, 2,4-dinitro-o-cyclohexylphenol, phenylurethane, 5-isoamyl-5-ethylbarbituric acid, or iodoacetic acid. 4. Quantitative examination of the fertilized eggs showed that cytochrome c, if present at all, occurred in a concentration of less than 2 micrograms per gram of wet fertilized Arbacia eggs. On the basis of these data and those of Fig. 2, above, it seems safe to conclude that cytochrome c cannot carry a significant fraction of the oxygen consumption of fertilized Arbacia eggs. It was also found that, in contrast to similar preparations from certain other animal tissues, the Arbacia cytochrome oxidase preparation displayed no succinic dehydrogenase activity when tested manometrically in the presence of excess cytochrome c. 5. Extending previously reported (3) experiments with other inhibitors, the effects of sodium azide and sodium sulfide on the respiration and cell division of fertilized Arbacia eggs were determined, the eggs being initially exposed to the reagents 30 minutes after fertilization at 20°C. With either reagent cleavage was completely blocked by a concentration of reagent which reduced the respiration to approximately 50 per cent of the normal level. 6. On the basis of certain theoretical considerations regarding the possible mechanism of action of cyanide and other respiratory inhibitors it is suggested that a fraction of the respiration apparently concerned with supplying energy for division processes in the fertilized Arbacia egg may be keyed into the respiratory cycle through a carrier having a somewhat higher potential than those which carry the larger portion of the egg respiration. The theory is also employed in an effort to resolve a number of hitherto apparently paradoxical observations regarding the effects of cyanide, azide, and carbon monoxide on cell respiration.     

Action of halothane upon mitochondrial respiration

The inhibitory action of halothane upon respiration was studied with rat liver mitochondria (RLM³), beef heart mitochondria (HBHM), and electron-transport particles (ETP). With intact mitochondrial preparations the oxidation of NADH-linked substrates but not of succinate was markedly suppressed by low concentrations of halothane (<2 mm as determined by gas-liquid chromatography). This inhibitory action of halothane was completely reversible. In contrast, a number of other mitochondrial processes were found to be sensitive in an irreversible manner at higher concentrations of the anesthetic. Likewise, the oxidation of added NADH by HBHM, ETP, and detergent-disrupted RLM was found to be sensitive in a reversible manner to low concentrations of halothane. The energy-dependent transfer of electrons from succinate to NAD by ETP_H was also sensitive to halothane. On the other hand, the NADH-ferricyanide reductase and the succinic oxidase activities of ETP and the NADH-cytochrome c reductase activity of microsomes were all insensitive to halothane. The site of inhibition by halothane appears to be in the vicinity of the rotenone-sensitive site of complex I (NADH-CoQ reductase). A number of other general anesthetics inhibited respiration at or near the same site as halothane.     

Reduction of Hg2+ with Reduced Mammalian Cytochrome c by Cytochrome c Oxidase Purified from a Mercury-Resistant Acidithiobacillus ferrooxidans Strain,MON-1

Acidithiobacillus ferrooxidans AP19-3, ATCC 23270, and MON-1 are mercury-sensitive, moderately mercury-resistant, and highly mercury-resistant strains respectively. It is known that 2,3,5,6-tetramethyl-p-phenylendiamine (TMPD) and reduced cytochrome c are used as electron donors specific for cytochrome c oxidase. Resting cells of strain MON-1 had TMPD oxidase activity and volatilized metal mercury with TMPD as an electron donor. Cytochrome c oxidase purified from strain MON-1 reduced mercuric ions to metalic mercury with reduced mammalian cytochrome c as well as TMPD. These mercury volatilization activities with reduced cytochrome c and TMPD were completely inhibited by 1 mM NaCN. These results indicate that cytochrome c oxidase is involved in mercury reduction in A. ferrooxidans cells. The cytochrome c oxidase activities of strains AP19-3 and ATCC 23270 were completely inhibited by 1 μM and 5 μM of mercuric chloride respectively. In contrast, the activity of strain MON-1 was inhibited 33% by 5 μM, and 70% by 10 μM of mercuric chloride, suggesting that the levels of mercury resistance in A. ferrooxidans strains correspond well with the levels of mercury resistance of cytochrome c oxidase.     

Mitochondrial function in glucocorticoid triggered T-ALL cells with transgenic bcl-2 expression

Independent of apoptosis, dexamethasone induced and a decrease of respiration and citrate synthase activity per cell in cells with and without transgenic Bcl-2 expression. The reduction of respiration, however, was slightly, but statistically more pronounced in apoptotic cells compared to non-apoptotic Bcl-2 over-expressing cells. A slight cytochrome c release was detected in apoptotic cells only. Importantly, the stimulatory effect of FCCP was maintained, indicating that oxidative phosphorylation remained coupled in active mitochondria. Coupled and uncoupled respiration were reduced to almost identical degrees as the activities of the marker enzymes citrate synthase (matrix) and cytochrome c oxidase (respiratory chain). Therefore, the reduction of cellular respiration was mainly caused by a decrease in mitochondrial content per cell. The functional integrity of mitochondria was preserved, apart from the slight degree of cytochrome c release, either through a pore formed by the oligomerisation of BAK in coupled mitochondria or by permeability transition of a small fraction of injured mitochondria.