scholarly journals Methylene Blue Bridges the Inhibition and Produces Unusual Respiratory Changes in Complex III-Inhibited Mitochondria. Studies on Rats, Mice and Guinea Pigs

Antioxidants ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 305
Author(s):  
Gergely Sváb ◽  
Márton Kokas ◽  
Ildikó Sipos ◽  
Attila Ambrus ◽  
László Tretter
Keyword(s):  
Methylene Blue ◽  
Oxygen Consumption ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Complex I ◽  
Complex Iii ◽  
Present Contribution ◽  
Cellular Targets ◽  
Beneficial Effects ◽  
Human Therapy

Methylene blue (MB) is used in human therapy in various pathological conditions. Its effects in neurodegenerative disease models are promising. MB acts on multiple cellular targets and mechanisms, but many of its potential beneficial effects are ascribed to be mitochondrial. According to the “alternative electron transport” hypothesis, MB is capable of donating electrons to cytochrome c bypassing complex I and III. As a consequence of this, the deleterious effects of the inhibitors of complex I and III can be ameliorated by MB. Recently, the beneficial effects of MB exerted on complex III-inhibited mitochondria were debated. In the present contribution, several pieces of evidence are provided towards that MB is able to reduce cytochrome c and improve bioenergetic parameters, like respiration and membrane potential, in mitochondria treated with complex III inhibitors, either antimycin or myxothiazol. These conclusions were drawn from measurements for mitochondrial oxygen consumption, membrane potential, NAD(P)H steady state, MB uptake and MB-cytochrome c oxidoreduction. In the presence of MB and complex III inhibitors, unusual respiratory reactions, like decreased oxygen consumption as a response to ADP addition as well as stimulation of respiration upon administration of inhibitors of ATP synthase or ANT, were observed. Qualitatively identical results were obtained in three rodent species. The actual metabolic status of mitochondria is well reflected in the distribution of MB amongst various compartments of this organelle.

scholarly journals Impact of diabetes-associated lipoproteins on oxygen consumption and mitochondrial enzymes in porcine aortic endothelial cells.

2010 ◽  
Vol 57 (4) ◽  
Author(s):  
Xueping Xie ◽  
Subir Roy Chowdhury ◽  
Ganesh Sangle ◽  
Garry X Shen
Keyword(s):  
Endothelial Cells ◽  
Oxygen Consumption ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Respiration ◽  
Complex I ◽  
Complex Iii ◽  
Mitochondrial Enzymes ◽  
Key Enzymes ◽  
Cytochrome C Reductase

Impairments in mitochondrial function have been proposed to play an important role in the pathogenesis of diabetes. Atherosclerotic coronary artery disease (CAD) is the leading cause of mortality in diabetic patients. Mitochondrial dysfunction and increased production of reactive oxygen species (ROS) are associated with diabetes and CAD. Elevated levels of glycated low density lipoproteins (glyLDL) and oxidized LDL (oxLDL) were detected in patients with diabetes. Our previous studies demonstrated that oxLDL and glyLDL increased the generation of ROS and altered the activities of antioxidant enzymes in vascular endothelial cells (EC). The present study examined the effects of glyLDL and oxLDL on mitochondrial respiration, membrane potential and the activities and proteins of key enzymes in mitochondrial electron transport chain (mETC) in cultured porcine aortic EC (PAEC). The results demonstrated that glyLDL or oxLDL significantly reduced oxygen consumption in Complex I, II/III and IV of mETC in PAEC compared to LDL or vehicle control using oxygraphy. Incubation with glyLDL or oxLDL significantly reduced mitochondrial membrane potential, the activities of mitochondrial ETC enzymes - NADH dehydrogenase (Complex I), succinate cytochrome c reductase (Complex II + III), ubiquinol cytochrome c reductase (Complex III), and cytochrome c oxidase (Complex IV) in PAEC compared to LDL or control. Treatment with oxLDL or glyLDL reduced the abundance of subunits of Complex I, ND1 and ND6 in PAEC. However, the effects of oxLDL on mitochondrial activity and proteins were not significantly different from glyLDL. The findings suggest that the glyLDL or oxLDL impairs mitochondrial respiration, as a result from the reduction of the abundance of several key enzymes in mitochondria of vascular EC, which potentially may lead to oxidative stress in vascular EC, and the development of diabetic vascular complications.

scholarly journals Caspase-mediated loss of mitochondrial function and generation of reactive oxygen species during apoptosis

2003 ◽  
Vol 160 (1) ◽  
pp. 65-75 ◽  
Author(s):  
Jean-Ehrland Ricci ◽  
Roberta A. Gottlieb ◽  
Douglas R. Green
Keyword(s):  
Reactive Oxygen Species ◽  
Oxygen Consumption ◽  
Electron Transport ◽  
Cytochrome C ◽  
Mitochondrial Function ◽  
Complex I ◽  
Caspase 3 ◽  
Reactive Oxygen ◽  
Complex Iii ◽  
Oxygen Species

During apoptosis, the permeabilization of the mitochondrial outer membrane allows the release of cytochrome c, which induces caspase activation to orchestrate the death of the cell. Mitochondria rapidly lose their transmembrane potential (ΔΨm) and generate reactive oxygen species (ROS), both of which are likely to contribute to the dismantling of the cell. Here we show that both the rapid loss of ΔΨm and the generation of ROS are due to the effects of activated caspases on mitochondrial electron transport complexes I and II. Caspase-3 disrupts oxygen consumption induced by complex I and II substrates but not that induced by electron transfer to complex IV. Similarly, ΔΨm generated in the presence of complex I or II substrates is disrupted by caspase-3, and ROS are produced. Complex III activity measured by cytochrome c reduction remains intact after caspase-3 treatment. In apoptotic cells, electron transport and oxygen consumption that depends on complex I or II was disrupted in a caspase-dependent manner. Our results indicate that after cytochrome c release the activation of caspases feeds back on the permeabilized mitochondria to damage mitochondrial function (loss of ΔΨm) and generate ROS through effects of caspases on complex I and II in the electron transport chain.

scholarly journals Response of Acanthamoeba castellanii mitochondria to oxidative stress.

2007 ◽  
Vol 54 (4) ◽  
pp. 797-803 ◽  
Author(s):  
Lidia K Trocha ◽  
Olgierd Stobienia
Keyword(s):  
Oxidative Stress ◽  
Oxygen Consumption ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Accumulation Rate ◽  
Acanthamoeba Castellanii ◽  
Stress Conditions ◽  
Cytochrome C Release ◽  
Stress Treatment ◽  
Four Levels

The purpose of this study was to examine the effects of oxidative stress caused by hydroperoxide (H(2)O(2)) in the presence of iron ions (Fe(2+)) on mitochondria of the amoeba Acanthamoeba castellanii. We used isolated mitochondria of A. castellanii and exposed them to four levels of H(2)O(2) concentration: 0.5, 5, 15, and 25 mM. We measured basic energetics of mitochondria: oxygen consumption in phosphorylation state (state 3) and resting state (state 4), respiratory coefficient rates (RC), ADP/O ratios, membrane potential (DeltaPsi(m)), ability to accumulate Ca(2+) , and cytochrome c release. Our results show that the increasing concentrations of H(2)O(2) stimulates respiration in states 3 and 4. The highest concentration of H(2)O(2) caused a 3-fold increase in respiration in state 3 compared to the control. Respiratory coefficients and ADP/O ratios decreased with increasing stress conditions. Membrane potential significantly collapsed with increasing hydroperoxide concentration. The ability to accumulate Ca(2+) also decreased with the increasing stress treatment. The lowest stress treatment (0.5 mM H(2)O(2)) significantly decreased oxygen consumption in state 3 and 4, RC, and membrane potential. The ADP/O ratio decreased significantly under 5 mM H(2)O(2) treatment, while Ca(2+) accumulation rate decreased significantly at 15 mM H(2)O(2). We also observed cytochrome c release under increasing stress conditions. However, this release was not linear. These results indicate that as low as 0.5 mM H(2)O(2) with Fe(2+) damage the basic energetics of mitochondria of the unicellular eukaryotic organism Acanthamoeba castellanii.

scholarly journals The effects of lipid phase transitions on the interaction of mitochondrial NADH–ubiquinone oxidoreductase with ubiquinol–cytochrome c oxidoreductase

1979 ◽  
Vol 178 (2) ◽  
pp. 415-426 ◽  
Author(s):  
C Heron ◽  
M G Gore ◽  
C I Ragan
Keyword(s):  
Cytochrome C ◽  
Cytochrome B ◽  
Complex I ◽  
Complex Iii ◽  
Molar Ratio ◽  
Lipid Phase ◽  
Oxidoreductase Activity ◽  
Arrhenius Plots ◽  
Nadh Cytochrome ◽  
Ubiquinone 10

1. The endogenous phosphatidylcholine and phosphatidylethanolamine of Complexes I and III from bovine heart mitochondria may be completely replaced with 1,2-ditetradecanoyl-sn-glycero-3-phosphocholine with at least partial retention of activity. 2. The lipid-replaced enzymes associate in 1:1 molar ratio to give a Complex I–III unit catalysing NADH-cytochrome c oxidoreductase activity. 3. On increasing the concentration of ubiquinone-10 and the synthetic phospholipid, the lipid-replaced Complexes appear to operate independently of each other as in the natural membrane. Thus the lipid-replaced enzymes associate in exactly the same ways as the enzymes containing natural phospholipids. 4. Arrhenius plots of NADH–cytochrome c oxidoreductase activity reconstituted from lipid-replaced Complexes I and III exhibit changes in slope at 24 degrees C. When the concentrations of phospholipid and ubiquinone-10 are increased, the Arrhenius plots show discontinuities at 24 degrees C as well as changes in slope. 5. The kinetics of cytochrome b reduction by NADH were measured in mixtures containing 2 mol of Complex III/mol of Complex I. When the enzymes contained natural phospholipids. the reduction kinetics were biphasic. When the enzymes had been supplemented with further phospholipid and ubiquinone-10 the kinetics were monophasic. When lipid-replaced enzymes were supplemented with 1,2-ditetradecanoyl-sn-glycero-3-phosphocholine and ubiquinone-10, reduction of cytochrome b was monophasic above the phase-transition temperature of the lipid but biphasic below it. 6. These findings are interpreted in terms of the model for the interaction of Complexes in the natural membrane proposed by Heron, Ragan & Trum-power [(1978) Biochem. J. 174, 791–800].

scholarly journals Potent inhibition of tumour cell proliferation and immunoregulatory function by mitochondria-targeted atovaquone

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Gang Cheng ◽  
Micael Hardy ◽  
Paytsar Topchyan ◽  
Ryan Zander ◽  
Peter Volberding ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Cancer Cells ◽  
Complex I ◽  
Complex Iii ◽  
Antiproliferative Effect ◽  
Mitochondrial Complex ◽  
Mitochondrial Target ◽  
Potent Inhibition ◽  
Alkyl Side Chains ◽  
First Time

Abstract The FDA-approved prophylactic antimalarial drug atovaquone (ATO) recently was repurposed as an antitumor drug. Studies show that ATO exerts a profound antiproliferative effect in several cancer cells, including breast, ovarian, and glioma. Analogous to the mechanism of action proposed in parasites, ATO inhibits mitochondrial complex III and cell respiration. To enhance the chemotherapeutic efficacy and oxidative phosphorylation inhibition, we developed a mitochondria-targeted triphenylphosphonium-conjugated ATO with varying alkyl side chains (Mito4-ATO, Mito10-ATO, Mito12-ATO, and Mito16-ATO). Results show, for the first time, that triphenylphosphonium-conjugated ATO potently enhanced the antiproliferative effect of ATO in cancer cells and, depending upon the alkyl chain length, the molecular target of inhibition changes from mitochondrial complex III to complex I. Mito4-ATO and Mito10-ATO inhibit both pyruvate/malate-dependent complex I and duroquinol-dependent complex III-induced oxygen consumption whereas Mito12-ATO and Mito16-ATO inhibit only complex I-induced oxygen consumption. Mitochondrial target shifting may have immunoregulatory implications.

scholarly journals The interaction between mitochondrial NADH-ubiquinone oxidoreductase and ubiquinol-cytochrome c oxidoreductase. Evidence for stoicheiometric association

1978 ◽  
Vol 174 (3) ◽  
pp. 783-790 ◽  
Author(s):  
C I Ragan ◽  
C Heron
Keyword(s):  
Cytochrome C ◽  
Complex I ◽  
Structural Features ◽  
Complex Iii ◽  
Molar Ratio ◽  
Oxidoreductase Activity ◽  
Ubiquinone Oxidoreductase ◽  
Nadh Ubiquinone Oxidoreductase ◽  
Cytochrome C Reduction ◽  
Nadh Cytochrome

1. The NADH-ubiquinone oxidoreductase complex (Complex I) and the ubiquinol-cytochrome c oxidoreductase complex (Complex III) combine in a 1:1 molar ratio to give NADH-cytochrome c oxidoreductase (Complex I-Complex III). 2. Experiments on the inhibition of the NADH-cytochrome c oxidoreductase activity of mixtures of Complexes I and III by rotenone and antimycin indicate that electron transfer between a unit of Complex I-Complex III and extra molecules of Complexes I or III does not contribute to the overall rate of cytochrome c reduction. 3. The reduction by NADH of the cytochrome b of mixtures of Complexes I and III is biphasic. The extents of the fast and slow phases of reduction are determined by the proportion of the total Complex III specifically associated with Complex I. 4. Activation-energy measurements suggest that the structural features of the Complex I-Complex III unit promote oxidoreduction of endogenous ubiquinone-10.

scholarly journals Complex I-mediated reactive oxygen species generation: modulation by cytochrome c and NAD(P)+ oxidation–reduction state

2002 ◽  
Vol 368 (2) ◽  
pp. 545-553 ◽  
Author(s):  
Yulia KUSHNAREVA ◽  
Anne N. MURPHY ◽  
Alexander ANDREYEV
Keyword(s):  
Reactive Oxygen Species ◽  
Cytochrome C ◽  
Complex I ◽  
Electron Flow ◽  
Reactive Oxygen ◽  
Complex Iii ◽  
Ros Generation ◽  
Ros Production ◽  
Oxygen Species ◽  
Reduced State

Several lines of evidence indicate that mitochondrial reactive oxygen species (ROS) generation is the major source of oxidative stress in the cell. It has been shown that ROS production accompanies cytochrome c release in different apoptotic paradigms, but the site(s) of ROS production remain obscure. In the current study, we demonstrate that loss of cytochrome c by mitochondria oxidizing NAD+-linked substrates results in a dramatic increase of ROS production and respiratory inhibition. This increased ROS production can be mimicked by rotenone, a complex I inhibitor, as well as other chemical inhibitors of electron flow that act further downstream in the electron transport chain. The effects of cytochrome c depletion from mitoplasts on ROS production and respiration are reversible upon addition of exogenous cytochrome c. Thus in these models of mitochondrial injury, a primary site of ROS generation in both brain and heart mitochondria is proximal to the rotenone inhibitory site, rather than in complex III. ROS production at complex I is critically dependent upon a highly reduced state of the mitochondrial NAD(P)+ pool and is achieved upon nearly complete inhibition of the respiratory chain. Redox clamp experiments using the acetoacetate/d-β-hydroxybutyrate couple in the presence of a maximally inhibitory rotenone concentration suggest that the site is approx. 50mV more electronegative than the NADH/NAD+ couple. In the absence of inhibitors, this highly reduced state of mitochondria can be induced by reverse electron flow from succinate to NAD+, accounting for profound ROS production in the presence of succinate. These results lead us to propose a model of thermodynamic control of mitochondrial ROS production which suggests that the ROS-generating site of complex I is the Fe—S centre N-1a.

scholarly journals Dynamic Modeling of Mitochondrial Membrane Potential Upon Exposure to Mitochondrial Inhibitors

2021 ◽  
Vol 12 ◽  
Author(s):  
Huan Yang ◽  
Wanda van der Stel ◽  
Randy Lee ◽  
Caroline Bauch ◽  
Sam Bevan ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Complex I ◽  
Mitochondrial Membrane ◽  
Energy Production ◽  
Cellular Response ◽  
Chemical Exposure ◽  
Confocal Imaging ◽  
Complex Iii ◽  
Ion Leakage

Mitochondria are the main bioenergetic organelles of cells. Exposure to chemicals targeting mitochondria therefore generally results in the development of toxicity. The cellular response to perturbations in cellular energy production is a balance between adaptation, by reorganisation and organelle biogenesis, and sacrifice, in the form of cell death. In homeostatic conditions, aerobic mitochondrial energy production requires the maintenance of a mitochondrial membrane potential (MMP). Chemicals can perturb this MMP, and the extent of this perturbation depends both on the pharmacokinetics of the chemicals and on downstream MMP dynamics. Here we obtain a quantitative understanding of mitochondrial adaptation upon exposure to various mitochondrial respiration inhibitors by applying mathematical modeling to partially published high-content imaging time-lapse confocal imaging data, focusing on MMP dynamics in HepG2 cells over a period of 24 h. The MMP was perturbed using a set of 24 compounds, either acting as uncoupler or as mitochondrial complex inhibitor targeting complex I, II, III or V. To characterize the effect of chemical exposure on MMP dynamics, we adapted an existing differential equation model and fitted this model to the observed MMP dynamics. Complex III inhibitor data were better described by the model than complex I data. Incorporation of pharmacokinetic decay into the model was required to obtain a proper fit for the uncoupler FCCP. Furthermore, oligomycin (complex V inhibitor) model fits were improved by either combining pharmacokinetic (PK) decay and ion leakage or a concentration-dependent decay. Subsequent mass spectrometry measurements showed that FCCP had a significant decay in its PK profile as predicted by the model. Moreover, the measured oligomycin PK profile exhibited only a limited decay at high concentration, whereas at low concentrations the compound remained below the detection limit within cells. This is consistent with the hypothesis that oligomycin exhibits a concentration-dependent decay, yet awaits further experimental verification with more sensitive detection methods. Overall, we show that there is a complex interplay between PK and MMP dynamics within mitochondria and that data-driven modeling is a powerful combination to unravel such complexity.

scholarly journals Propofol-Induced Mitochondrial and Contractile Dysfunction of the Rat Ventricular Myocardium

2016 ◽  
pp. S601-S609 ◽  
Author(s):  
M. GRUNDMANOVÁ ◽  
D. JARKOVSKÁ ◽  
A. SÜß ◽  
Z. TŮMA ◽  
M. MARKOVÁ ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Complex I ◽  
Ventricular Myocardium ◽  
Myocardial Contraction ◽  
Complex Iii ◽  
Adult Rats ◽  
Arterial Blood ◽  
Cardiovascular Side Effects ◽  
Hypnotic Agent ◽  
The Right

Propofol is a short-acting hypnotic agent used in human medicine for sedation and general anesthesia. Its administration can be associated with serious cardiovascular side-effects that include decrease in arterial blood pressure and cardiac output. The aim of the present study was to evaluate propofol effects on mitochondrial respiration, myocardial contractility and electrophysiology in the same samples isolated from the heart ventricles of adult rats. Mitochondrial oxygen consumption was measured in permeabilized samples dissected from free walls of both ventricles using high-resolution respirometry. State LEAK was determined with malate and glutamate. Active respiration was induced by ADP (state PI) and further by succinate, a Complex II substrate (PI+II). Rotenone was injected to measure state PII. Antimycin A, a Complex III inhibitor was used to determine residual oxygen consumption (ROX). N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride and ascorbate were injected simultaneously for respirometric assay of cytochrome c oxidase activity (CIV). Isometric contractions and membrane potentials were determined on multicellular preparations isolated from right and left ventricles. Propofol concentrations used ranged from 0.005 to 0.5 mmol/l. All respiratory parameters were significantly higher in the left control ventricles compared to the right ones. Propofol significantly decreased Complex I activity at concentration 0.025 mmol/l and papillary muscle contraction force at 0.1 mmol/l. Propofol did not affect action potential duration at any concentration studied. Our study suggests that mechanisms contributing to the impaired myocardial contraction during propofol anesthesia might include also mitochondrial dysfunction manifested by compromised activity of the respiratory Complex I.

Sign in / Sign up

Export Citation Format

Share Document