Oxidative Phosphorylation Capacity Research Articles

The effect of trimetazidine on cardiac haemodynamics and mitochondrial function in wild-type transthyretin amyloidosis.

Wild-type transthyretin cardiac amyloidosis (ATTRwt) is a cardiomyopathy causing myocardial hypoperfusion and impaired cardiac mitochondrial function. Trimetazidine is an antianginal agent used in patients with stable angina pectoris, which improves cardiac contractility and mitochondrial function. The aim of the study was to investigate the effect of trimetazidine on invasive haemodynamics and cardiac mitochondrial function in ATTRwt. In a randomized, double-blind, placebo-controlled, crossover trial, 22 patients with ATTRwt received 4weeks of trimetazidine and placebo in randomized order. After each treatment period followed examinations with endomyocardial biopsies taken for high-resolution respirometry and right heart catheterization at rest and during a cardiopulmonary exercise test. The primary endpoint was mean pulmonary artery wedge pressure (mPAWP) during peak exercise. The secondary endpoint was cardiac mitochondrial oxidative phosphorylation capacity. Exploratory endpoints were echocardiographic parameters, cardiac biomarker levels and quality of life. Trimetazidine did not significantly reduce mPAWP during peak exercise (31±12 vs. 31±13mmHg, P=0.61) or improve the cardiac mitochondrial oxidative phosphorylation capacity (73.4±7.7 vs. 75.3±7.7pmol O2/(mg*s), P=0.81) compared with placebo, nor did treatment with trimetazidine improve ejection fraction (P=0.93), global longitudinal strain (P=0.23), N-terminal pro-brain natriuretic peptide (NT-proBNP) levels (P=0.92) or the patients' quality of life (P=0.98). In ATTRwt, treatment with trimetazidine did not improve mPAWP or cardiac mitochondrial oxidative phosphorylation capacity compared with placebo.

Effects of Hyperthermia and Acidosis on Mitochondrial Oxidative Phosphorylation.

The intracellular environment of skeletal muscle can develop pronounced hyperthermia and acidosis during strenuous exercise, and these alterations in the typical intracellular conditions have been shown to alter mitochondrial respiration. However, the impact of these conditions on ATP synthesis is poorly understood. We used Thoroughbred racehorses to test the hypothesis that both hyperthermia and acidosis decrease the rate of ATP synthesis, but that athletic conditioning mitigates this loss of phosphorylation capacity. Isolated mitochondria were harvested from skeletal muscle before and after a 9-week racetrack conditioning program that increased whole-body aerobic capacity by 19%, and oxidative phosphorylation capacity was tested ex vivo under normothermic and hyperthermic conditions, as well as normal pH and acidic pH created by the addition of lactic acid. In unfit horses, hyperthermia caused a 30-55% decrease in the rate of ATP synthesis and loss of phosphorylation efficiency (P/O ratio decreased from 4.2 to 1.7 during maximal oxidative phosphorylation). Aerobic conditioning resulted in increased phosphorylation efficiency under hyperthermic conditions. Lactic acidosis had a small negative effect on ATP synthesis in unfit horses, but aerobic conditioning increased the sensitivity of isolated mitochondria to the deleterious effects of lactic acidosis. These data support a prominent role of hyperthermia in skeletal muscle fatigue during exercise, particularly in unfit subjects. However, acidosis may be a more important cause of failure of ATP synthesis in fit subjects.

Effects of concentric and eccentric exercise regimens on bioenergetic efficiency of lymphocytes in sedentary males.

Eccentric exercise training (EET) increases physical performance while having lower metabolic demand than concentric exercise training (CET). Whether EET influences bioenergetic efficiency in peripheral blood mononuclear cells (PBMCs) remains unclear. This study investigates the effects of EET and CET on PBMC phenotypes and mitochondrial functions in blood. Thirty three sedentary healthy males were randomly assigned to either EET (n=11) or CET (n=11) that performed at progressively increased from 60% to 80% of maximal absolute workload for 30min/day, 5days/week for 6weeks, or a control group (n=11) that did not receive any exercise intervention. A graded exercise stress test (GXT) was performed before and after the intervention. PBMC phenotypes and mitochondrial respiratory capacity were analyzed using flowcytometry and high-resolution respirometry, respectively. In the same absolute workload, EET elicited lower heart rate and rating of perceived exertion than CET. However, EET as CET increased the VO2 level at the ventilatory threshold. Notably, both EET and CET increased central memory (CD45RO+/CD62+/CD3+) T cells and decreased effector memory T cells reexpressing CD45RA (CD45RA+/CD62-/CD3+). Moreover, the two exercise regimens diminished the loss of mitochondrial membrane potential (ΔΨm) caused by GXT, increased maximal/reserve O2 consumption rates (OCR), and bioenergetic health index in intact PBMCs and enhanced complex I-/II-related OCR in PBMCs with a substrate-rich environment. EET improves aerobic fitness with a lower cardiovascular response to exercise than CET. Moreover, EET as CET reduces senescent T-cell distribution in blood and improves PBMC bioenergetic efficiency by stabilizing ΔΨm and increasing capacity of oxidative phosphorylation.

Cellular ATP demand creates metabolically distinct subpopulations of mitochondria.

Mitochondria serve a crucial role in cell growth and proliferation by supporting both ATP synthesis and the production of macromolecular precursors. Whereas oxidative phosphorylation (OXPHOS) depends mainly on the oxidation of intermediates from the tricarboxylic acid cycle, the mitochondrial production of proline and ornithine relies on reductive synthesis1. How these competing metabolic pathways take place in the same organelle is not clear. Here we show that when cellular dependence on OXPHOS increases, pyrroline-5-carboxylate synthase (P5CS)-the rate-limiting enzyme in the reductive synthesis of proline and ornithine-becomes sequestered in a subset of mitochondria that lack cristae and ATP synthase. This sequestration is driven by both the intrinsic ability of P5CS to form filaments and the mitochondrial fusion and fission cycle. Disruption of mitochondrial dynamics, by impeding mitofusin-mediated fusion or dynamin-like-protein-1-mediated fission, impairs the separation of P5CS-containing mitochondria from mitochondria that are enriched in cristae and ATP synthase. Failure to segregate these metabolic pathways through mitochondrial fusion and fission results in cells either sacrificing the capacity for OXPHOS while sustaining the reductive synthesis of proline, or foregoing proline synthesis while preserving adaptive OXPHOS. These findings provide evidence of the key role of mitochondrial fission and fusion in maintaining both oxidative and reductive biosyntheses in response to changing nutrient availability and bioenergetic demand.

The mitochondrial physiology of torpor in ruby-throated hummingbirds, Archilochus colubris.

Hummingbirds save energy by facultatively entering torpor, but the physiological mechanisms underlying this metabolic suppression are largely unknown. We compared whole-animal and pectoralis mitochondrial metabolism between torpid and normothermic ruby-throated hummingbirds (Archilochus colubris). When fasting, hummingbirds were exposed to 10°C ambient temperature at night and they entered torpor; average body temperature decreased by nearly 25°C (from ∼37 to ∼13°C) and whole-animal metabolic rate (V̇O2) decreased by 95% compared with normothermia, a much greater metabolic suppression compared with that of mammalian daily heterotherms. We then measured pectoralis mitochondrial oxidative phosphorylation (OXPHOS) fueled by either carbohydrate or fatty acid substrates at both 39°C and 10°C in torpid and normothermic hummingbirds. Aside from a 20% decrease in electron transport system complex I-supported respiration with pyruvate, the capacity for OXPHOS at a common in vivo temperature did not differ in isolated mitochondria between torpor and normothermia. Similarly, the activities of pectoralis pyruvate dehydrogenase and 3-hydroxyacyl-CoA dehydrogenase did not differ between the states. Unlike heterothermic mammals, hummingbirds do not suppress muscle mitochondrial metabolism in torpor by active, temperature-independent mechanisms. Other mechanisms that may underly this impressive whole-animal metabolic suppression include decreasing ATP demand or relying on rapid passive cooling facilitated by the very small body size of A. colubris.

Higher myocardial mitochondrial ketone body oxidation capacity in human heart failure

Abstract Introduction/Purpose Recent studies suggest that the ketone bodies (KB) beta-hydroxybutyrate (HBA) and acetoacetate (ACA) are relevant substrates for myocardial energy metabolism in heart failure (HF). However, the amount of KB-dependent mitochondrial respiration in HF has not yet been quantified. High-resolution respirometry (HRR) is the gold-standard method for quantifying substrate-dependent mitochondrial oxidative capacity. Therefore, we aimed to evaluate the association between myocardial KB-dependent oxidative capacity and human HF using HRR. Methods The primary endpoint of this single-center prospective cohort study was KB-dependent and overall mitochondrial oxidative capacity in permeabilized myocardium. We used two different protocols for our measurements. In the first HRR protocol, conventional respirometry substrates including fatty acids, complex I- and complex II substrates were used to quantify the maximum coupled oxidative phosphorylation capacity (OXPHOS). Leak respiration was quantified using oligomycin to calculate the respiratory control ratio (RCR, state 3/state 4o), and leak control ratio (LCR, state 4o/state u). In the second protocol which was recently established by our working group, HBA and ACA were used as respirometry substrates to quantify KB-dependent respiration down to the enzyme level. The relative contribution of the KBs was obtained by comparing HBA- and ACA-derived respiration to OXPHOS. We applied both protocols to myocardial samples from individuals with advanced HF (HF group, catheter-acquired endomyocardial biopsies from non-ischemic HF patients or samples of explanted hearts collected during orthotopic heart transplantation surgery in end-stage heart failure patients) as well as previously heart transplanted individuals without current heart failure (Control group). Results Controls and HF had similar demographic characteristics, with comparable age distribution (mean±SD, 56.3±10.3 vs. 54.2±10.7 years, p=0.36), sex (68.75% vs. 66.67% male, p=0.87), or body mass index (25.0±5.8 vs. 29.0±10.6 kg/m2, p=0.08) and differed significantly in cardiac index (3.06±0.67 vs. 1.93±0.44) and ejection fraction (55.8±10.2 vs. 26.6±9.1) (both p<0.0001). The HF group exhibited lower OXPHOS and respiration for all substrate combinations tested (Fig. 1A). In contrast, KB-dependent respiration did not differ between HF and controls (Fig. 1B). The relative HBA-dependent respiration (Fig. 1C) and the relative and absolute ACA-dependent respiration (Fig. 1C and D) were higher in the HF group compared to controls. Mitochondrial uncoupling (LCR) (0.47±0.29 vs. 0.44±0.09 [AU], p=0.79) and coupling efficiency (RCR) (2.75±1.38 vs. 2.11±0.39 [AU], p=0.15) were not different between the groups. Conclusion These findings suggest that mitochondria in the failing heart can utilize KBs for OXPHOS to a greater extent and further hint towards KB metabolism as a potential therapeutic target for HF.

Mitochondrial oxidative phosphorylation capacity in skeletal muscle measured by ultrafast Z-spectroscopy (UFZ) MRI at 3T.

To investigate the feasibility of rapid CEST MRI acquisition for evaluating oxidative phosphorylation (OXPHOS) in human skeletal muscle at 3T, utilizing ultrafast Z-spectroscopy (UFZ) combined with MRI and the Polynomial and Lorentzian line-shape Fitting (PLOF) technique. UFZ MRI on muscle was evaluated with turbo spin echo (TSE) and 3D EPI readouts. Five healthy subjects performed in-magnet plantar flexion exercise (PFE) and subsequent changes of amide, PCr, and partial PCr mixed Cr (Cr+) CEST dynamic signals post-exercise were enabled by PLOF fitting. PCr/Cr CEST signal was further refined through pH correction by using the ratios between PCr/Cr and amide signals, named PCAR/CAR, respectively. UFZ MRI with TSE readout significantly reduces acquisition time, achieving a temporal resolution of <50 s for collecting high-resolution Z-spectra. Following PFE, the recovery/decay times (τ) for both PCr and Cr in the gastrocnemius muscle of the calf were notably longer when determined using PCr/Cr CEST compared to those after pH correction with amideCEST, namely = 87.1 ± 15.8 s and = 98.1 ± 20.4 s versus = 32.9 ± 19.7 s and = 43.0 ± 13.0 s, respectively. obtained via 31P MRS ( = 50.3 ± 6.2 s) closely resemble those obtained from pH-corrected PCr/Cr CEST signals. The outcomes suggest potential of UFZ MRI as a robust tool for non-invasive assessment of mitochondrial function in skeletal muscles. pH correction is critical for the reliable OXPHOS measurement by CEST.

Mitochondrial Abundance and Function Differ Across Muscle Within Species.

Background: Mitochondria are considered the powerhouse of cells, and skeletal muscle cells are no exception. However, information regarding muscle mitochondria from different species is limited. Methods: Different muscles from cattle, pigs and chickens were analyzed for mitochondrial DNA (mtDNA), protein and oxygen consumption. Results: Bovine oxidative muscle mitochondria contain greater mtDNA (p < 0.05), protein (succinate dehydrogenase, SDHA, p < 0.01; citrate synthase, CS, p < 0.01; complex I, CI, p < 0.05), and oxygen consumption (p < 0.01) than their glycolytic counterpart. Likewise, porcine oxidative muscle contains greater mtDNA (p < 0.01), mitochondrial proteins (SDHA, p < 0.05; CS, p < 0.001; CI, p < 0.01) and oxidative phosphorylation capacity (OXPHOS, p < 0.05) in comparison to glycolytic muscle. However, avian oxidative skeletal muscle showed no differences in absolute mtDNA, SDHA, CI, complex II, lactate dehydrogenase, or glyceraldehyde 3 phosphate dehydrogenase compared to their glycolytic counterpart. Even so, avian mitochondria isolated from oxidative muscles had greater OXPHOS capacity (p < 0.05) than glycolytic muscle. Conclusions: These data show avian mitochondria function is independent of absolute mtDNA content and protein abundance, and argue that multiple levels of inquiry are warranted to determine the wholistic role of mitochondria in skeletal muscle.

EMC1 Is Required for the Sarcoplasmic Reticulum and Mitochondrial Functions in the Drosophila Muscle

EMC1 is part of the endoplasmic reticulum (ER) membrane protein complex, whose functions include the insertion of transmembrane proteins into the ER membrane, ER–mitochondria contact, and lipid exchange. Here, we show that the Drosophila melanogaster EMC1 gene is expressed in the somatic musculature and the protein localizes to the sarcoplasmic reticulum (SR) network. Muscle-specific EMC1 RNAi led to severe motility defects and partial late pupae/early adulthood lethality, phenotypes that are rescued by co-expression with an EMC1 transgene. Motility impairment in EMC1-depleted flies was associated with aberrations in muscle morphology in embryos, larvae, and adults, including tortuous and misaligned fibers with reduced size and weakness. They were also associated with an altered SR network, cytosolic calcium overload, and mitochondrial dysfunction and dysmorphology that impaired membrane potential and oxidative phosphorylation capacity. Genes coding for ER stress sensors, mitochondrial biogenesis/dynamics, and other EMC components showed altered expression and were mostly rescued by the EMC1 transgene expression. In conclusion, EMC1 is required for the SR network’s mitochondrial integrity and influences underlying programs involved in the regulation of muscle mass and shape. We believe our data can contribute to the biology of human diseases caused by EMC1 mutations.

7086 Disrupted Mitochondrial Function in Alström Syndrome- A Monogenic Model of Insulin Resistance and Obesity

Abstract Disclosure: S. Ali: None. S. Heising: None. V. Veeranna: None. A. Vincent: None. G.S. Xavier: None. T. Geberhiwot: None. Alström syndrome (ALMS) is a rare monogenic disease typified by severe insulin resistance (IR) and obesity. IR and obesity are cardinal features of metabolic syndrome. The metabolic effects of insulin at key target tissues leads to metabolic haemostasis by promoting glucose and lipid uptake and metabolism to generate energy in cells. Although insulin is a major regulator of fuel metabolism, its effects on mitochondrial ATP production in severe insulin resistance are unclear. RNASeq data from adipose tissue (AT) from patients with ALMS show downregulation of the expression of genes associated with mitochondrial respiratory chain function, pointing towards possible altered mitochondrial function in ALMS. We therefore undertook a detailed assessment of mitochondrial oxidative phosphorylation capacity (OXPHOS) in skeletal muscle (SM) in ALMS. We performed SM biopsy in 10 ALMS and 10 healthy control volunteers that were age, gender and Body Mass Index matched. The SM was subjected to high resolution respirometry, the gold standard method to quantify mitochondrial function ex vivo, using OROBOROS Oxygraph -2k (O2k) system. Overall OXPHOS was higher in control SM compared to ALMS, with maximal respiration of 62.96±19.2 and 44.24 ± 12.53 pmol/(s*mg), respectively (p&amp;lt;0.05).Reduced oxygen influx was observed in ALMS SM in comparison to controls for fatty acid ß oxidation (6.7±5.96 vs 7.87±2.7 pmol/(s*mg);p&amp;lt;0.05, at complex I (19.4 ± 8.98 vs 29.7 ±5.3 pmol/(s*mg);p&amp;lt;0.05, and at complex II (34.98 ±11.75 vs 53.52± 13.2 pmol/(s*mg) ;p&amp;lt;0.05. Mitochondrial membrane integrity was unaffected during these experiments. Therefore, our analysis unveils a consistent reduction in mitochondrial respiration across all the electron transport pathway. This noteworthy finding suggests impairment in respiratory function and the electron transport chain in mitochondria, a crucial aspect of cellular energy metabolism. Several studies have previously linked IR with mitochondrial dysfunction, where insulin-resistant rodents and humans have shown blunted mitochondrial response to ATP generation. Our results align with this body of evidence emphasizing the significance of investigating the underlying mechanism contributing to this phenomenon. These findings could pave the way to exploring potential mitochondrial-targeted therapies to restore mitochondrial function and therefore, improve insulin sensitivity to tackle IR in ALMS. Presentation: 6/2/2024

Brain-body mitochondrial distribution patterns lack coherence and point to tissue-specific and individualized regulatory mechanisms.

Energy transformation capacity is generally assumed to be a coherent individual trait driven by genetic and environmental factors. This predicts that some individuals should have high and others low mitochondrial oxidative phosphorylation (OxPhos) capacity across organ systems. Here, we test this assumption using multi-tissue molecular and enzymatic activities in mice and humans. Across up to 22 mouse tissues, neither mitochondrial OxPhos capacity nor mtDNA density were correlated between tissues (median r = -0.01-0.16), indicating that animals with high mitochondrial capacity in one tissue can have low capacity in other tissues. Similarly, the multi-tissue correlation structure of RNAseq-based indices of mitochondrial gene expression across 45 tissues from 948 women and men (GTEx) showed small to moderate coherence between only some tissues (regions of the same brain), but not between brain-body tissue pairs in the same person (median r = 0.01). Mitochondrial DNA copy number (mtDNAcn) also lacked coherence across organs and tissues. Mechanistically, tissue-specific differences in mitochondrial gene expression were attributable in part to i) tissue-specific activation of canonical energy sensing pathways including the transcriptional coactivator PGC-1 and the integrated stress response (ISR), and ii) proliferative activity across tissues. Finally, we identify subgroups of individuals with high mitochondrial gene expression in some tissues (e.g., heart) but low expression in others (e.g., skeletal muscle) who display different clinical phenotypic patterns. Taken together, these data raise the possibility that tissue-specific energy sensing pathways may contribute to the idiosyncratic mitochondrial distribution patterns associated with the inter-organ heterogeneity and phenotypic diversity among individuals.

Exploring the transcriptomic and metabolomic profiles of adipose tissues: Insights and implications for fat grafting

BackgroundExpanding on our prior research on murine fat grafts' metabolic shift, this study delves deeper into the metabolic profiles of human adipose tissues, specifically the superficial subcutaneous adipose tissue (SSAT) and deep subcutaneous adipose tissue (DSAT). MethodsUtilizing RNA-sequencing, metabolome, and metabolic flux analyses, we examined SSAT and DSAT samples obtained during deep inferior epigastric perforator flap breast reconstructions. Transcript data underwent unsupervised hierarchical clustering and Gene set enrichment analysis. Metabolomics involved analyzing samples for cationic and anionic metabolites via Capillary Electrophoresis Time-of-Flight Mass Spectrometry (CE-TOFMS), followed by principal component analysis (PCA) and heat map generation. Additionally, primary adipocytes from SSAT and DSAT were assessed using the Seahorse® extracellular flux analyzer. ResultsPCA and heat map analyses highlighted distinct transcriptome and metabolome differences between SSAT and DSAT. SSAT predominantly upregulated genes linked to adipogenesis (false discovery rate [FDR] q < 0.0001), oxidative phosphorylation (FDR q < 0.0001), fatty acid metabolism (FDR q <0.0001), and glycolysis (FDR q = 0.001). In contrast, DSAT showed a significant upregulation in inflammatory response genes (FDR q <0.05). Metabolite analysis revealed an abundance of glycolytic metabolites in SSAT, whereas DSAT was rich in metabolites associated with fatty acid metabolism and oxidative phosphorylation. Cellular flux analysis further confirmed SSAT's elevated glycolysis and spare oxidative phosphorylation capacities. ConclusionOur results highlight the metabolic uniqueness of SSAT and DSAT in humans, with SSAT exhibiting superior metabolic flexibility. The implications of these metabolic differences, especially in fat grafting procedures, necessitate further research and exploration in future studies. Data availabilityThe ribonucleic acid sequencing data were deposited in the Gene Expression Omnibus database (GSE216803). https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE216803 (GEO token: qnsngiuqplqdpaf).

Panaxadiol Saponin alleviates LPS-induced cardiomyopathy similar to dexamethasone via improving mitochondrial quality control.

Septic cardiomyopathy is linked to a dysregulation in mitochondrial integrity and elevated mortality rates, for which an efficacious treatment remains elusive. PDS, a panaxadiol saponin extracted from ginseng stem and leaf. This study identified the protective effects of PDS and DEX in LPS-induced cardiomyopathy and explored the mechanism of them treating LPS-induced cardiomyopathy from the perspectives of mitochondrial quality control. DEX and PDS enhance antioxidant defense by degrading Keap1 to activate Nrf2, activate mitochondrial occurrence protein PGC-1α and fusion protein OPA1, Mfn1, Mfn2 expression, inhibit phosphorylation of mitochondrial fission protein Drp1, aiming to maintain normal structure and function of mitochondrial, thereby preserving oxidative phosphorylation capacity. In summary, our findings highlighted that the protective efficacy of PDS and DEX in maintaining mitochondrial in LPS-induced cardiomyopathy, and mechanism improving mitochondrial quality control at least in part by promoting Nrf2 activation.

Insulin sensitivity and insulin secretion in adults with Friedreich's Ataxia: the role of skeletal muscle.

Friedreich's Ataxia (FRDA) is a multi-system disorder caused by frataxin deficiency. FRDA-related diabetes mellitus (DM) is common. Frataxin supports skeletal muscle mitochondrial oxidative phosphorylation (OXPHOS) capacity, a mediator of insulin sensitivity. Our objective was to test the association between skeletal muscle health and insulin sensitivity and secretion in adults with FRDA without DM. Case-control study (NCT02920671). Glucose and insulin metabolism (stable-isotope oral glucose tolerance tests), body composition (dual-energy x-ray absorptiometry), physical activity (self-report), and skeletal muscle OXPHOS capacity (creatine chemical exchange saturation transfer MRI) were assessed. Participants included 11 individuals with FRDA (4 female), median age 27y (IQR 23, 39), BMI 26.9kg/m2 (24.1, 29.4), and 24 controls (11 female), 29y (26, 39), 24.4kg/m2 (21.8, 27.0). Fasting glucose was higher in FRDA (91 vs. 83mg/dL (5.0 vs. 4.6mmol/L), p<0.05). Individuals with FRDA had lower insulin sensitivity (WBISI 2.8 vs. 5.3, p<0.01), higher post-prandial insulin secretion (insulin secretory rate iAUC 30-180 minutes, 24,652 vs. 17,858, p<0.05), and more suppressed post-prandial endogenous glucose production (-0.9% vs. 26.9% of fasting EGP, p<0.05). In regression analyses, lower OXPHOS and inactivity explained some of the difference in insulin sensitivity. More visceral fat contributed to lower insulin sensitivity independent of FRDA. Insulin secretion accounting for sensitivity (disposition index) was not different. Lower mitochondrial OXPHOS capacity, inactivity, and visceral adiposity contribute to lower insulin sensitivity in FRDA. Higher insulin secretion appears compensatory, and when inadequate, could herald DM. Further studies are needed to determine if muscle- or adipose-focused interventions could delay FRDA-related DM.

NRF2 is a spatiotemporal metabolic hub essential for the polyfunctionality of Th2 cells

Upon encountering allergens, CD4+ T cells differentiate into IL-4-producing Th2 cells in lymph nodes, which later transform into polyfunctional Th2 cells producing IL-5 and IL-13 in inflamed tissues. However, the precise mechanism underlying their polyfunctionality remains elusive. In this study, we elucidate the pivotal role of NRF2 in polyfunctional Th2 cells in murine models of allergic asthma and in human Th2 cells. We found that an increase in reactive oxygen species (ROS) in immune cells infiltrating the lungs is necessary for the development of eosinophilic asthma and polyfunctional Th2 cells in vivo. Deletion of the ROS sensor NRF2 specifically in T cells, but not in dendritic cells, significantly abolished eosinophilia and polyfunctional Th2 cells in the airway. Mechanistically, NRF2 intrinsic to T cells is essential for inducing optimal oxidative phosphorylation and glycolysis capacity, thereby driving Th2 cell polyfunctionality independently of IL-33, partially by inducing PPARγ. Treatment with an NRF2 inhibitor leads to a substantial decrease in polyfunctional Th2 cells and subsequent eosinophilia in mice and a reduction in the production of Th2 cytokines from peripheral blood mononuclear cells in asthmatic patients. These findings highlight the critical role of Nrf2 as a spatial and temporal metabolic hub that is essential for polyfunctional Th2 cells, suggesting potential therapeutic implications for allergic diseases.

Exercise Hemodynamics and Mitochondrial Oxidative Capacity in Disease Stages of Wild-Type Transthyretin Amyloid Cardiomyopathy.

Wild-type transthyretin amyloid (ATTRwt) cardiomyopathy is increasingly recognized in the development of heart failure. The link between cardiac performance, hemodynamics, and mitochondrial function in disease stages of ATTRwt has not previously been studied but may provide new insights into the pathophysiology and clinical performance of the patients. The study investigated 47 patients diagnosed with ATTRwt at Aarhus University Hospital, Denmark. Patients were stratified according to the disease stages of the National Amyloidosis Centre (NAC) as NAC I with low levels of NT-proBNP (N-terminal pro-B-type natriuretic peptide) (NAC I-L, n=14), NAC I with high levels NT-proBNP (NAC I-H, n=20), and NAC II-III (n=13). Exercise testing with simultaneous right heart catheterization was performed in all patients. Endomyocardial biopsies were collected from the patients and the mitochondrial oxidative phosphorylation capacity was assessed. All NAC disease groups, even in the NAC I-L group, a significant abnormal increase in biventricular filling pressures were noted during exercise while the filling pressures was normal or near normal at rest. The inotropic response to exercise was reduced with diminished increase in cardiac output which was significantly more pronounced in the NAC I-H (Diff. -2.4, 95% CI (-4.2: -0.7), P=0.00) and the NAC II-III group (Diff: -3.1 L/min, 95% CI (-5.2: -1.1), P=0.00) compared with the NAC I-L group. The pulmonary artery wedge pressure to cardiac output ratio at peak exercise was significantly different between NAC I-L and NAC II-III (Diff: 1.6 mm Hg*min/L, 95% CI (0.01:3.3, P=0.04)). Patients with ATTRwt had a reduced oxidative phosphorylation capacity which correlated to left ventricular mass but not to cardiac output capacity. An abnormal restrictive left ventricle and right ventricle response to exercise was demonstrated, even present in patients with early-stage ATTRwt. In more advanced disease stages a progressive impairment of the pressure-flow relationship was noted. The myocyte energetics is deranged but not associated to the contractile reserve or restrictive filling characteristics in ATTRwt.

Importance of Michaelis Constants for Cancer Cell Redox Balance and Lactate Secretion-Revisiting the Warburg Effect.

Cancer cells metabolize a large fraction of glucose to lactate, even under a sufficient oxygen supply. This phenomenon-the "Warburg Effect"-is often regarded as not yet understood. Cancer cells change gene expression to increase the uptake and utilization of glucose for biosynthesis pathways and glycolysis, but they do not adequately up-regulate the tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS). Thereby, an increased glycolytic flux causes an increased production of cytosolic NADH. However, since the corresponding gene expression changes are not neatly fine-tuned in the cancer cells, cytosolic NAD+ must often be regenerated by loading excess electrons onto pyruvate and secreting the resulting lactate, even under sufficient oxygen supply. Interestingly, the Michaelis constants (KM values) of the enzymes at the pyruvate junction are sufficient to explain the priorities for pyruvate utilization in cancer cells: 1. mitochondrial OXPHOS for efficient ATP production, 2. electrons that exceed OXPHOS capacity need to be disposed of and secreted as lactate, and 3. biosynthesis reactions for cancer cell growth. In other words, a number of cytosolic electrons need to take the "emergency exit" from the cell by lactate secretion to maintain the cytosolic redox balance.

Mechanisms that Alter Capacity for Adenosine Triphosphate Production and Oxidative Phosphorylation: Insights from Avian Migration

Synopsis Avian migration is among the most energetically demanding feats observed in animals. Studies evaluating the physiological underpinnings of migration have repeatedly shown that migratory birds display numerous adaptations that ultimately supply the flight muscle mitochondria with abundant fuel and oxygen during long-distance flights. To make use of this high input, the organs and mitochondria of migrants are predicted to display several traits that maximize their capacity to produce adenosine triphosphate (ATP). This review aims to introduce readers to several mechanisms by which organs and mitochondria can alter their capacity for oxidative phosphorylation and ATP production. The role of organ size, mitochondrial volume, substrate, and oxygen delivery to the electron transport system are discussed. A central theme of this review is the role of changes in electron chain complex activity, mitochondrial morphology and dynamics, and supercomplexes in allowing avian migrants and other taxa to alter the performance of the electron transport system with predictable shifts in demand. It is my hope that this review will serve as a springboard for future studies exploring the mechanisms that alter bioenergetic capacity across animal species.

In Vitro Hypoxia/Reoxygenation Induces Mitochondrial Cardiolipin Remodeling in Human Kidney Cells.

Renal ischemia/reperfusion is a serious condition that not only causes acute kidney injury, a severe clinical syndrome with high mortality, but is also an inevitable part of kidney transplantation or other kidney surgeries. Alterations of oxygen levels during ischemia/reperfusion, namely hypoxia/reoxygenation, disrupt mitochondrial metabolism and induce structural changes that lead to cell death. A signature mitochondrial phospholipid, cardiolipin, with many vital roles in mitochondrial homeostasis, is one of the key players in hypoxia/reoxygenation-induced mitochondrial damage. In this study, we analyze the effect of hypoxia/reoxygenation on human renal proximal tubule epithelial cell (RPTEC) cardiolipins, as well as their metabolism and mitochondrial functions. RPTEC cells were placed in a hypoxic chamber with a 2% oxygen atmosphere for 24 h to induce hypoxia; then, they were replaced back into regular growth conditions for 24 h of reoxygenation. Surprisingly, after 24 h, hypoxia cardiolipin levels substantially increased and remained higher than control levels after 24 h of reoxygenation. This was explained by significantly elevated levels of cardiolipin synthase and lysocardiolipin acyltransferase 1 (LCLAT1) gene expression and protein levels. Meanwhile, hypoxia/reoxygenation decreased ADP-dependent mitochondrial respiration rates and oxidative phosphorylation capacity and increased reactive oxygen species generation. Our findings suggest that hypoxia/reoxygenation induces cardiolipin remodeling in response to reduced mitochondrial oxidative phosphorylation in a way that protects mitochondrial function.

Mitochondrial respiration is lower in the intrauterine growth-restricted fetal sheep heart.

Fetuses affected by intrauterine growth restriction have an increased risk of developing heart disease and failure in adulthood. Compared with controls, late gestation intrauterine growth-restricted (IUGR) fetal sheep have fewer binucleated cardiomyocytes, reflecting a more immature heart, which may reduce mitochondrial capacity to oxidize substrates. We hypothesized that the late gestation IUGR fetal heart has a lower capacity for mitochondrial oxidative phosphorylation. Left (LV) and right (RV) ventricles from IUGR and control (CON) fetal sheep at 90% gestation were harvested. Mitochondrial respiration (states 1-3, LeakOmy, and maximal respiration) in response to carbohydrates and lipids, citrate synthase (CS) activity, protein expression levels of mitochondrial oxidative phosphorylation complexes (CI-CV), and mRNA expression levels of mitochondrial biosynthesis regulators were measured. The carbohydrate and lipid state 3 respiration rates were lower in IUGR than CON, and CS activity was lower in IUGR LV than CON LV. However, relative CII and CV protein levels were higher in IUGR than CON; CV expression level was higher in IUGR than CON. Genes involved in lipid metabolism had lower expression in IUGR than CON. In addition, the LV and RV demonstrated distinct differences in oxygen flux and gene expression levels, which were independent from CON and IUGR status. Low mitochondrial respiration and CS activity in the IUGR heart compared with CON are consistent with delayed cardiomyocyte maturation, and CII and CV protein expression levels may be upregulated to support ATP production. These insights will provide a better understanding of fetal heart development in an adverse in utero environment. KEY POINTS: Growth-restricted fetuses have a higher risk of developing and dying from cardiovascular diseases in adulthood. Mitochondria are the main supplier of energy for the heart. As the heart matures, the substrate preference of the mitochondria switches from carbohydrates to lipids. We used a sheep model of intrauterine growth restriction to study the capacity of the mitochondria in the heart to produce energy using either carbohydrate or lipid substrates by measuring how much oxygen was consumed. Our data show that the mitochondria respiration levels in the growth-restricted fetal heart were lower than in the normally growing fetuses, and the expression levels of genes involved in lipid metabolism were also lower. Differences between the right and left ventricles that are independent of the fetal growth restriction condition were identified. These results indicate an impaired metabolic maturation of the growth-restricted fetal heart associated with a decreased capacity to oxidize lipids postnatally.