SIGNIFICANCE: Mitochondria are the energetic, metabolic, redox, and information signaling centers of the cell. Substrate pressure, mitochondrial network dynamics, and cristae morphology state are integrated by the protonmotive force Δp or its potential component, ΔΨ, which are attenuated by proton backflux into the matrix, termed uncoupling. The mitochondrial uncoupling proteins (UCP1-5) play an eminent role in the regulation of each of the mentioned aspects, being involved in numerous physiological events including redox signaling. Recent Advances: UCP2 structure, including purine nucleotide and fatty acid (FA) binding sites, strongly support the FA cycling mechanism: UCP2 expels FA anions, whereas uncoupling is achieved by the membrane backflux of protonated FA. Nascent FAs, cleaved by phospholipases, are preferential. The resulting Δp dissipation decreases superoxide formation dependent on Δp. UCP-mediated antioxidant protection and its impairment are expected to play a major role in cell physiology and pathology. Moreover, UCP2-mediated aspartate, oxaloacetate, and malate antiport with phosphate is expected to alter metabolism of cancer cells. CRITICAL ISSUES: A wide range of UCP antioxidant effects and participations in redox signaling have been reported; however, mechanisms of UCP activation are still debated. Switching off/on the UCP2 protonophoretic function might serve as redox signaling either by employing/releasing the extra capacity of cell antioxidant systems or by directly increasing/decreasing mitochondrial superoxide sources. Rapid UCP2 degradation, FA levels, elevation of purine nucleotides, decreased Mg2+, or increased pyruvate accumulation may initiate UCP-mediated redox signaling. FUTURE DIRECTIONS: Issues such as UCP2 participation in glucose sensing, neuronal (synaptic) function, and immune cell activation should be elucidated. Antioxid. Redox Signal. 29, 667-714.

• Keywords
• UCP2, anion transport, attenuation of superoxide formation, fatty acid cycling, mitochondrial uncoupling proteins, redox signaling,
• MeSH
• Antioxidants metabolism   MeSH
• Humans MeSH
• Mitochondrial Uncoupling Proteins metabolism   MeSH
• Oxidation-Reduction MeSH
• Signal Transduction * MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Antioxidants MeSH
• Mitochondrial Uncoupling Proteins MeSH

Quercetin and dehydrosilybin are polyphenols which are known to behave like uncouplers of respiration in isolated mitochondria. Here we investigated whether the effect is conserved in whole cells. Following short term incubation, neither compound uncouples mitochondrial respiration in whole H9c2 cells below 50μM. However, following hypoxia, or long term incubation, leak (state IV with oligomycin) oxygen consumption is increased by quercetin. Both compounds partially protected complex I respiration, but not complex II in H9c2 cells following hypoxia. In a permeabilised H9c2 cell model, the increase in leak respiration caused by quercetin is lowered by increased [ADP] and is increased by adenine nucleotide transporter inhibitor, atractyloside, but not bongkrekic acid. Both quercetin and dehydrosilybin dissipate mitochondrial membrane potential in whole cells. In the case of quercetin, the effect is potentiated post hypoxia. Genetically encoded Ca++ sensors, targeted to the mitochondria, enabled the use of fluorescence microscopy to show that quercetin decreased mitochondrial [Ca++] while dehydrosilybin did not. Likewise, quercetin decreases accumulation of [Ca++] in mitochondria following hypoxia. Fluorescent probes were used to show that both compounds decrease plasma membrane potential and increase cytosolic [Ca++]. We conclude that the uncoupler-like effects of these polyphenols are attenuated in whole cells compared to isolated mitochondria, but downstream effects are nevertheless apparent. Results suggest that the effect of quercetin observed in whole and permeabilised cells may originate in the mitochondria, while the mechanism of action of cardioprotection by dehydrosilybin may be less dependent on mitochondrial uncoupling than originally thought. Rather, protective effects may originate due to interactions at the plasma membrane.

• MeSH
• Cell Line MeSH
• Digitonin pharmacology   MeSH
• Microscopy, Fluorescence MeSH
• Microscopy, Confocal MeSH
• Membrane Potential, Mitochondrial drug effects   MeSH
• Mitochondrial ADP, ATP Translocases metabolism   MeSH
• Quercetin pharmacology   MeSH
• Silymarin pharmacology   MeSH
• Calcium metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• dehydrosilybin MeSH Browser
• Digitonin MeSH
• Mitochondrial ADP, ATP Translocases MeSH
• Quercetin MeSH
• Silymarin MeSH
• Calcium MeSH

Flavonol (-)-epicatechin and its derived dimer procyanidin B2, present in high amounts in cocoa products, have been shown to exert beneficial effects on the heart and cardiovascular system; however, their mechanism of action has not been fully elucidated. We studied effects of (-)-epicatechin and procyanidin B2 on the oxidative phosphorylation of isolated rat heart mitochondria. (-)-Epicatechin and procyanidin B2 had stimulating effect (up to 30% compared to control) on substrate-driven (State 2) mitochondrial respiration. Their effect was dependent on the respiratory substrates used. (-)-Epicatechin at higher concentrations (from 0.27 µg/mL) significantly decreased (up to 15%) substrate- and ADP-driven (State 3) mitochondrial respiration in case of pyruvate and malate oxidation only. Procyanidin B2 (0.7-17.9 ng/mL) inhibited State 3 respiration rate up to 19%, the most profound effect being expressed with succinate as the substrate. (-)-Epicatechin at concentrations of 0.23 µg/mL and 0.46 µg/mL prevented loss of the cytochrome c from mitochondria when substrate was succinate, supporting the evidence of membrane stabilizing properties of this flavonol. Thus, both (-)-epicatechin and procyanidin B2 directly influenced mitochondrial functions and the observed effects could help to explain cardiometabolic risk reduction ascribed to the consumption of modest amounts of cocoa products.

• MeSH
• Adenosine Diphosphate pharmacology   MeSH
• Biflavonoids chemistry   pharmacology   MeSH
• Cell Respiration drug effects   MeSH
• Cytochromes c metabolism   MeSH
• Catechin chemistry   pharmacology   MeSH
• Rats MeSH
• Oxidation-Reduction MeSH
• Proanthocyanidins chemistry   pharmacology   MeSH
• Mitochondria, Heart drug effects   metabolism   MeSH
• Substrate Specificity drug effects   MeSH
• Succinates metabolism   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Adenosine Diphosphate MeSH
• Biflavonoids MeSH
• Cytochromes c MeSH
• Catechin MeSH
• Proanthocyanidins MeSH
• procyanidin B2 MeSH Browser
• Succinates MeSH

Reactive oxygen species (ROS) originating from mitochondria are perceived as a factor contributing to cell aging and means have been sought to attenuate ROS formation with the aim of extending the cell lifespan. Silybin and dehydrosilybin, two polyphenolic compounds, display a plethora of biological effects generally ascribed to their known antioxidant capacity. When investigating the cytoprotective effects of these two compounds in the primary cell cultures of neonatal rat cardiomyocytes, we noted the ability of dehydrosilybin to de-energize the cells by monitoring JC-1 fluorescence. Experiments evaluating oxygen consumption and membrane potential revealed that dehydrosilybin uncouples the respiration of isolated rat heart mitochondria albeit with a much lower potency than synthetic uncouplers. Furthermore, dehydrosilybin revealed a very high potency in suppressing ROS formation in isolated rat heart mitochondria with IC(50) = 0.15 μM. It is far more effective than its effect in a purely chemical system generating superoxide or in cells capable of oxidative burst, where the IC(50) for dehydrosilybin exceeds 50 μM. Dehydrosilybin also attenuated ROS formation caused by rotenone in the primary cultures of neonatal rat cardiomyocytes. We infer that the apparent uncoupler-like activity of dehydrosilybin is the basis of its ROS modulation effect in neonatal rat cardiomyocytes and leads us to propose a hypothesis on natural ischemia preconditioning by dietary polyphenols.

• MeSH
• Analysis of Variance MeSH
• Benzimidazoles MeSH
• Fluorescent Dyes MeSH
• Inhibitory Concentration 50 MeSH
• Carbocyanines MeSH
• Myocytes, Cardiac metabolism   MeSH
• Rats MeSH
• Mitochondria metabolism   MeSH
• Molecular Structure MeSH
• Rats, Wistar MeSH
• Reactive Oxygen Species metabolism   MeSH
• Rotenone toxicity   MeSH
• Silymarin chemistry   pharmacology   MeSH
• Oxygen Consumption drug effects   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine MeSH Browser
• Benzimidazoles MeSH
• dehydrosilybin MeSH Browser
• Fluorescent Dyes MeSH
• Carbocyanines MeSH
• Reactive Oxygen Species MeSH
• Rotenone MeSH
• Silymarin MeSH