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Background Skeletal muscle stem cells (satellite cells) are well known to participate in regeneration and maintenance of the tissue over time. Studies have shown increases in the number of satellite cells after exercise, but their functional role in endurance training remains unexplored. Methods Young adult mice were submitted to endurance exercise training and the function, differentiation, and metabolic characteristics of satellite cells were investigated in vivo and in vitro. Results We found that injured muscles from endurance-exercised mice display improved regenerative capacity, demonstrated through higher densities of newly formed myofibres compared with controls (evidenced by an increase in embryonic myosin heavy chain expression), as well as lower inflammation (evidenced by quantifying CD68-marked macrophages), and reduced fibrosis. Enhanced myogenic function was accompanied by an increased fraction of satellite cells expressing self-renewal markers, while control satellite cells had morphologies suggestive of early differentiation. The beneficial effects of endurance exercise were associated with satellite cell metabolic reprogramming, including reduced mitochondrial respiration (O 2 consumption) under resting conditions (absence of muscle injury) and increased stemness. During proliferation or activated states (3 days after injury), O 2 consumption was equal in control and exercised cells, while exercise enhanced myogenic colony formation. Surprisingly, inhibition of mitochondrial O 2 consumption was sufficient to enhance muscle stem cell self-renewal characteristics in vitro. Moreover, transplanted muscle satellite cells from exercised mice or cells with reduced mitochondrial respiration promoted a significant reduction in inflammation compared with controls. Conclusions Our results indicate that endurance exercise promotes self-renewal and inhibits differentiation in satellite cells, an effect promoted by metabolic reprogramming and respiratory inhibition, which is associated with a more favourable muscular response to injury.

UCH-L1 is a deubiquitinating enzyme (DUB), highly abundant in neurons, with a sub-cellular localization dependent on its farnesylation state. Despite UCH-L1′s association with familial Parkinson's Disease (PD), the effects on mitochondrial bioenergetics and quality control remain unexplored. Here we investigated the role of UCHL-1 in mitochondrial dynamics and bioenergetics. We demonstrate that knock-down (KD) of UCH-L1 in different cell lines reduces the levels of the mitochondrial fusion protein Mitofusin-2, but not Mitofusin-1, resulting in mitochondrial enlargement and disruption of the tubular network. This was associated with lower tethering between mitochondria and the endoplasmic reticulum, consequently altering mitochondrial calcium uptake. Respiratory function was also altered, as UCH-L1 KD cells displayed higher proton leak and maximum respiratory capacity. Conversely, overexpression of UCH-L1 increased Mfn2 levels, an effect dramatically enhanced by the mutation of the farnesylation site (C220S), which drives UCH-L1 binding to membranes. These data indicate that the soluble cytosolic form of UCH-L1 regulates Mitofusin-2 levels and mitochondrial function. These effects are biologically conserved, since knock-down of the corresponding UCH-L1 ortholog in D. melanogaster reduces levels of the mitofusin ortholog Marf and also increases mitochondrial respiratory capacity. We thus show that Mfn-2 levels are directly affected by UCH-L1, demonstrating that the mitochondrial roles of DUBs go beyond controlling mitophagy rates.

The existence of a K+ cycle in mitochondria has been predicted since the development of the chemiosmotic theory and has been shown to be crucial for several cellular phenomena, including regulation of mitochondrial volume and redox state. One of the pathways known to participate in K+ cycling is the ATP-sensitive K+ channel, MitoKATP. This channel was vastly studied for promoting protection against ischemia reperfusion when pharmacologically activated, although its molecular identity remained unknown for decades. The recent molecular characterization of MitoKATP has opened new possibilities for modulation of this channel as a mechanism to control cellular processes. Here, we discuss different strategies to control MitoKATP activity and consider how these could be used as tools to regulate metabolism and cellular events.

Exposure to toxic levels of fatty acids (lipotoxicity) leads to cell damage and death and is involved in the pathogenesis of the metabolic syndrome. Since the metabolic consequences of lipotoxicity are still poorly understood, we studied the bioenergetic effects of the saturated fatty acid palmitate, quantifying changes in mitochondrial morphology, real-time oxygen consumption, ATP production sources, and extracellular acidification in hepatoma cells. Surprisingly, glycolysis was enhanced by the presence of palmitate as soon as 1 h after stimulus, while oxygen consumption and oxidative phosphorylation were unchanged, despite overt mitochondrial fragmentation. Palmitate only induced mitochondrial fragmentation if glucose and glutamine were available, while glycolytic enhancement did not require glutamine, showing it is independent of mitochondrial morphological changes. Redox state was altered by palmitate, as indicated by NAD(P)H quantification. Furthermore, the mitochondrial antioxidant mitoquinone, or a selective inhibitor of complex I electron leakage (S1QEL) further enhanced palmitate-induced glycolysis. Our results demonstrate that palmitate overload and lipotoxicity involves an unexpected and early increase in glycolytic flux, while, surprisingly, no changes in oxidative phosphorylation are observed. Interestingly, enhanced glycolysis involves signaling by mitochondrially-generated oxidants, uncovering a novel regulatory mechanism for this pathway.

Background: Cardiac hypertrophy involves marked wall thickening or chamber enlargement. If sustained, this condition will lead to dysfunctional mitochondria and oxidative stress. Mitochondria have ATP-sensitive K+ channels (mitoKATP) in the inner membrane that modulate the redox status of the cell. Objective: We investigated the in vivo effects of mitoKATP opening on oxidative stress in isoproterenol- induced cardiac hypertrophy. Methods: Cardiac hypertrophy was induced in Swiss mice treated intraperitoneally with isoproterenol (ISO - 30 mg/kg/day) for 8 days. From day 4, diazoxide (DZX - 5 mg/kg/day) was used in order to open mitoKATP (a clinically relevant therapy scheme) and 5-hydroxydecanoate (5HD - 5 mg/kg/day) or glibenclamide (GLI - 3 mg/kg/day) were used as mitoKATP blockers. Results: Isoproterenol-treated mice had elevated heart weight/tibia length ratios (HW/TL). Additionally, hypertrophic hearts had elevated levels of carbonylated proteins and Thiobarbituric Acid Reactive Substances (TBARS), markers of protein and lipid oxidation. In contrast, mitoKATP opening with DZX avoided ISO effects on gross hypertrophic markers (HW/TL), carbonylated proteins and TBARS, in a manner reversed by 5HD and GLI. Moreover, DZX improved mitochondrial superoxide dismutase activity. This effect was also blocked by 5HD and GLI. Additionally, ex vivo treatment of isoproterenol- induced hypertrophic cardiac tissue with DZX decreased H2O2 production in a manner sensitive to 5HD, indicating that this drug also acutely avoids oxidative stress. Conclusion: Our results suggest that diazoxide blocks oxidative stress and reverses cardiac hypertrophy. This pharmacological intervention could be a potential therapeutic strategy to prevent oxidative stress associated with cardiac hypertrophy.

Metabolic analysis in animals is usually either evaluated as whole-body measurements or in isolated tissue samples. To reveal tissue specificities in vivo , this study uses scanning electrochemical microscopy (SECM) to provide localized oxygen consumption rates (OCRs) in different regions of single adult Caenorhabditis elegans individuals. This is achieved by measuring the oxygen reduction current at the SECM tip electrode and using a finite element method model of the experiment that defines oxygen concentration and flux at the surface of the organism. SECM mapping measurements uncover a marked heterogeneity of OCR along the worm, with high respiration rates at the reproductive system region. To enable sensitive and quantitative measurements, a self-referencing approach is adopted, whereby the oxygen reduction current at the SECM tip is measured at a selected point on the worm and in bulk solution (calibration). Using genetic and pharmacological approaches, our SECM measurements indicate that viable eggs in the reproductive system are the main contributors in the total oxygen consumption of adult Caenorhabditis elegans . The finding that large regional differences in OCR exist within the animal provides a new understanding of oxygen consumption and metabolic measurements, paving the way for tissue-specific metabolic analyses and toxicity evaluation within single organisms.

Caloric restriction (CR) prevents obesity and increases resilience against pathological stimuli in laboratory rodents. At the mitochondrial level, protection promoted by CR in the brain and liver is related to higher calcium uptake rates and capacities, avoiding Ca2+-induced mitochondrial permeability transition. Dietary restriction has also been shown to increase kidney resistance against damaging stimuli, but if these effects are related to similar mitochondrial adaptations has not been uncovered. Here, we characterized changes in mitochondrial function in response to six months CR in rats, measuring bioenergetic parameters, redox balance and calcium homeostasis. CR promoted an increase in succinate-supported mitochondrial oxygen consumption rates. While CR prevents mitochondrial reactive oxygen species production in many tissues, in kidney we found that mitochondrial H2O2 release was enhanced in a succinate-dependent manner. Surprisingly, and opposite to the effects observed in brain and liver, mitochondria from CR animals are more prone to Ca2+-induced mitochondrial permeability transition, in a manner reversed by antioxidant dithiothreitol. CR mitochondria also displayed higher calcium uptake rates, which were not accompanied by changes in calcium efflux rates, nor related to altered inner mitochondrial membrane potentials or amounts of the mitochondrial calcium uniporter (MCU). Instead, increased mitochondrial calcium uptake rates in CR kidneys correlate with a loss of MICU2, an MCU modulator. Interestingly, MICU2 is also modulated by CR in liver, suggesting it has a broader diet-sensitive regulatory role controlling mitochondrial calcium homeostasis. Together, our results highlight the organ-specific bioenergetic, redox, and ionic transport effects of CR, with some unexpected deleterious effects in kidney.