Supplemental Digital Content is available in the text.

Purpose: It is well established that high-load resistance exercise (HLRE) can stimulate myofibrillar accretion. Additionally, recent studies suggest that HLRE can also stimulate mitochondrial biogenesis and respiratory function. However, in several clinical situations, the use of resistance exercise with high loading may not constitute a viable approach. Low-load blood flow restricted resistance exercise (BFRRE) has emerged as a time-effective low-load alternative to stimulate myofibrillar accretion. It is unknown if BFRRE can also stimulate mitochondrial biogenesis and respiratory function. If so, BFRRE could provide a feasible strategy to stimulate muscle metabolic health.Methods: To study this, 34 healthy previously untrained individuals (24 ± 3 years) participated in BFRRE, HLRE, or non-exercise control intervention (CON) 3 times per week for 6 weeks. Skeletal muscle biopsies were collected; (1) before and after the 6-week intervention period to assess mitochondrial biogenesis and respiratory function and; (2) during recovery from single-bout exercise to assess myocellular signaling events involved in transcriptional regulation of mitochondrial biogenesis. During the 6-week intervention period, deuterium oxide (D2O) was continuously administered to the participants to label newly synthesized skeletal muscle mitochondrial proteins. Mitochondrial respiratory function was assessed in permeabilized muscle fibers with high-resolution respirometry. Mitochondrial content was assessed with a citrate synthase activity assay. Myocellular signaling was assessed with immunoblotting.Results: Mitochondrial protein synthesis rate was higher with BFRRE (1.19%/day) and HLRE (1.15%/day) compared to CON (0.92%/day) (P < 0.05) but similar between exercise groups. Mitochondrial respiratory function increased to similar degree with both exercise regimens and did not change with CON. For instance, coupled respiration supported by convergent electron flow from complex I and II increased 38% with BFRRE and 24% with HLRE (P < 0.01). Training did not alter citrate synthase activity compared to CON. BFRRE and HLRE elicited similar myocellular signaling responses.Conclusion: These results support recent findings that resistance exercise can stimulate mitochondrial biogenesis and respiratory function to support healthy skeletal muscle and whole-body metabolism. Intriquingly, BFRRE produces similar mitochondrial adaptations at a markedly lower load, which entail great clinical perspective for populations in whom exercise with high loading is untenable.

Reducing infarct size during a cardiac ischaemic‐reperfusion episode is still of paramount importance, because the extension of myocardial necrosis is an important risk factor for developing heart failure. Cardiac ischaemia‐reperfusion injury (IRI) is in principle a metabolic pathology as it is caused by abruptly halted metabolism during the ischaemic episode and exacerbated by sudden restart of specific metabolic pathways at reperfusion. It should therefore not come as a surprise that therapy directed at metabolic pathways can modulate IRI. Here, we summarize the current knowledge of important metabolic pathways as therapeutic targets to combat cardiac IRI. Activating metabolic pathways such as glycolysis (eg AMPK activators), glucose oxidation (activating pyruvate dehydrogenase complex), ketone oxidation (increasing ketone plasma levels), hexosamine biosynthesis pathway (O‐GlcNAcylation; administration of glucosamine/glutamine) and deacetylation (activating sirtuins 1 or 3; administration of NAD+‐boosting compounds) all seem to hold promise to reduce acute IRI. In contrast, some metabolic pathways may offer protection through diminished activity. These pathways comprise the malate‐aspartate shuttle (in need of novel specific reversible inhibitors), mitochondrial oxygen consumption, fatty acid oxidation (CD36 inhibitors, malonyl‐CoA decarboxylase inhibitors) and mitochondrial succinate metabolism (malonate). Additionally, protecting the cristae structure of the mitochondria during IR, by maintaining the association of hexokinase II or creatine kinase with mitochondria, or inhibiting destabilization of FOF1‐ATPase dimers, prevents mitochondrial damage and thereby reduces cardiac IRI. Currently, the most promising and druggable metabolic therapy against cardiac IRI seems to be the singular or combined targeting of glycolysis, O‐GlcNAcylation and metabolism of ketones, fatty acids and succinate.

ObjectivesThe aim of this study was to determine if hyperpolarized [1,4–13C2]malate imaging could measure cardiomyocyte necrosis after myocardial infarction (MI).BackgroundMI is defined by an acute burst of cellular necrosis and the subsequent cascade of structural and functional adaptations. Quantifying necrosis in the clinic after MI remains challenging. Magnetic resonance-based detection of the conversion of hyperpolarized [1,4–13C2]fumarate to [1,4–13C2]malate, enabled by disrupted cell membrane integrity, has measured cellular necrosis in vivo in other tissue types. Our aim was to determine whether hyperpolarized [1,4–13C2]malate imaging could measure necrosis after MI.MethodsIsolated perfused hearts were given hyperpolarized [1,4–13C2]fumarate at baseline, immediately after 20 min of ischemia, and after 45 min of reperfusion. Magnetic resonance spectroscopy measured conversion into [1,4–13C2]malate. Left ventricular function and energetics were monitored throughout the protocol, buffer samples were collected and hearts were preserved for further analyses. For in vivo studies, magnetic resonance spectroscopy and a novel spatial-spectral magnetic resonance imaging sequence were implemented to assess cardiomyocyte necrosis in rats, 1 day and 1 week after cryo-induced MI.ResultsIn isolated hearts, [1,4–13C2]malate production became apparent after 45 min of reperfusion, and increased 2.7-fold compared with baseline. Expression of dicarboxylic acid transporter genes were negligible in healthy and reperfused hearts, and lactate dehydrogenase release and infarct size were significantly increased in reperfused hearts. Nonlinear regression revealed that [1,4–13C2]malate production was induced when adenosine triphosphate was depleted by >50%, below 5.3 mmol/l (R2 = 0.904). In vivo, the quantity of [1,4–13C2]malate visible increased 82-fold over controls 1 day after infarction, maintaining a 31-fold increase 7 days post-infarct. [1,4–13C2]Malate could be resolved using hyperpolarized magnetic resonance imaging in the infarct region one day after MI; [1,4–13C2]malate was not visible in control hearts.ConclusionsMalate production in the infarcted heart appears to provide a specific probe of necrosis acutely after MI, and for at least 1 week afterward. This technique could offer an alternative noninvasive method to measure cellular necrosis in heart disease, and warrants further investigation in patients.

Mitochondrial dysfunction plays a central role in ischaemia-reperfusion (IR) injury. Pre-ischaemic administration of aminooxyacetate (AOA), an inhibitor of the malate-aspartate shuttle (MAS), provides cardioprotection against IR injury, although the underlying mechanism remains unknown. We hypothesized that a transient inhibition of the MAS during ischaemia and early reperfusion could preserve mitochondrial function at later phase of reperfusion in the IR-injured heart to the same extent as ischaemic preconditioning (IPC), which is a well-validated cardioprotective strategy against IR injury. In the present study, we show that pre-ischaemic administration of AOA preserved mitochondrial complex I-linked state 3 respiration and fatty acid oxidation during late reperfusion in IR-injured isolated rat hearts. AOA treatment also attenuated the excessive emission of mitochondrial reactive oxygen species during state 3 with complex I-linked substrates during late reperfusion, which was consistent with reduced oxidative damage in the IR-injured heart. As a result, AOA treatment reduced infarct size after reperfusion. These protective effects of MAS inhibition on the mitochondria were similar to those of IPC. Intriguingly, the protection of mitochondrial function by AOA treatment appears to be different from that of IPC because AOA treatment, but not IPC, downregulated myocardial tricarboxilic acid (TCA)-cycle intermediates at the onset of reperfusion. MAS inhibition thus preserved mitochondrial respiratory capacity and decreased mitochondrial oxidative stress during late reperfusion in the IR-injured heart, at least in part, via metabolic regulation of TCA cycle intermediates in the mitochondria at the onset of reperfusion.

Hepatic and renal energy status prior to transplantation correlates with graft survival. However, effects of brain death (BD) on organ-specific energy status are largely unknown. We studied metabolism, perfusion, oxygen consumption, and mitochondrial function in the liver and kidneys following BD. BD was induced in mechanically-ventilated rats, inflating an epidurally-placed Fogarty-catheter, with sham-operated rats as controls. A 9.4T-preclinical MRI system measured hourly oxygen availability (BOLD-related R2*) and perfusion (T1-weighted). After 4 hrs, tissue was collected, mitochondria isolated and assessed with high-resolution respirometry. Quantitative proteomics, qPCR, and biochemistry was performed on stored tissue/plasma. Following BD, the liver increased glycolytic gene expression (Pfk-1) with decreased glycogen stores, while the kidneys increased anaerobic- (Ldha) and decreased gluconeogenic-related gene expression (Pck-1). Hepatic oxygen consumption increased, while renal perfusion decreased. ATP levels dropped in both organs while mitochondrial respiration and complex I/ATP synthase activity were unaffected. In conclusion, the liver responds to increased metabolic demands during BD, enhancing aerobic metabolism with functional mitochondria. The kidneys shift towards anaerobic energy production while renal perfusion decreases. Our findings highlight the need for an organ-specific approach to assess and optimise graft quality prior to transplantation, to optimise hepatic metabolic conditions and improve renal perfusion while supporting cellular detoxification.

Augmented mortality and morbidity following an acute myocardial infarction in patients with diabetes mellitus Type 2 (T2DM) may be caused by increased sensitivity to ischemia reperfusion (IR) injury or altered activation of endogenous cardioprotective pathways modified by T2DM per se or ischemic preconditioning (IPC). We aimed to investigate, whether the duration of T2DM influences sensitivity against IR injury and the efficacy of IPC, and how myocardial glucose oxidation rate was involved. Male Zucker diabetic fatty rats (homozygote (fa/fa)) at ages 6-(prediabetic), 12- (onset diabetes) and 24-weeks of age (late diabetes) and their age-matched non-diabetic controls (heterozygote (fa/+) were subjected to IR injury in the Langendorff model and randomised to IPC stimulus or control. T2DM rats were endogenously protected at onset of diabetes, as infarct size was lower in 12-weeks T2DM animals than in 6- (35±2% vs 53±4%; P = 0.006) and 24-weeks animals (35±2% vs 72±4%; P<0.0001). IPC reduced infarct size in all groups irrespective of the presence of T2DM and its duration (32±3%; 20±2%; 36±4% respectively; (ANOVA P<0.0001). Compared to prediabetic rats, myocardial glucose oxidation rates were reduced during stabilisation and early reperfusion at onset of T2DM, but these animals retained the ability to increase oxidation rate in late reperfusion. Late diabetic rats had low glucose oxidation rates throughout stabilisation and reperfusion. Despite inherent differences in sensitivity to IR injury, the cardioprotective effect of IPC was preserved in our animal model of pre-, early and late stage T2DM and associated with adaptations to myocardial glucose oxidation capacity.

Background: Patients with congestive heart failure (CHF) have impaired functional capacity and inferior quality of life. The clinical manifestations are associated with structural and functional impairments in skeletal muscle, emphasizing a need for feasible rehabilitation strategies beyond optimal anticongestive medical treatment. We investigated whether low-load blood flow restricted resistance exercise (BFRRE) or remote ischemic conditioning (RIC) could improve functional capacity and quality of life in patients with CHF and stimulate skeletal muscle myofibrillar and mitochondrial adaptations. Methods: We randomized 36 patients with CHF to BFRRE, RIC, or nontreatment control. BFRRE and RIC were performed 3× per week for 6 weeks. Before and after intervention, muscle biopsies, tests of functional capacity, and quality of life assessments were performed. Deuterium oxide was administered throughout the intervention to measure cumulative RNA and subfraction protein synthesis. Changes in muscle fiber morphology and mitochondrial respiratory function were also assessed. Results: BFRRE improved 6-minute walk test by 39.0 m (CI, 7.0–71.1, P =0.019) compared with control. BFRRE increased maximum isometric strength by 29.7 Nm (CI, 10.8–48.6, P =0.003) compared with control. BFRRE improved quality of life by 5.4 points (CI, −0.04 to 10.9; P =0.052) compared with control. BFRRE increased mitochondrial function by 19.1 pmol/s per milligram (CI, 7.3–30.8; P =0.002) compared with control. RIC did not produce similar changes. Conclusions: Our results demonstrate that BFRRE, but not RIC, improves functional capacity, quality of life, and muscle mitochondrial function. Our findings have clinical implications for rehabilitation of patients with CHF and provide new insights on the myopathy accompanying CHF. Clinical Trial Registration: URL: https://www.clinicaltrials.gov . Unique identifier: NCT03380663.