Mitochondrial respiration in peripheral arterial disease depends on stage severity.

Peripheral arterial disease (PAD) is an increasing cause of morbidity and its severity is graded based on clinical manifestation. To investigate the influence of the different stages on myopathy of ischemic muscle we analysed severity-dependent effects of mitochondrial respiration in PAD. Eighteen patients with severe PAD, defined as chronic limb-threatening ischemia, 47 patients with intermittent claudication (IC) and 22 non-ischemic controls were analysed. High-resolution respirometry (HRR) was performed on muscle biopsies of gastrocnemius and vastus lateralis muscle of patients in different PAD stages to investigate different respiratory states. Results from HRR are given as median and interquartile range and were normalized to citrate synthase activity (CSA), a marker for mitochondrial content. In order to account for inter-individual differences between patients and controls, we calculated the ratio of O2-flux in gastrocnemius muscle over vastus muscle ('GV ratio'). CSA of the gastrocnemius muscle as a proxy for mitochondrial content was significantly lower in critical ischemia compared to controls. Mitochondrial respiration normalized to CSA was higher in IC compared to controls. Likewise, the GV ratio was significantly higher in IC compared to control. Mitochondrial respiration and CSA of PAD patients showed stage-dependent modifications with greater changes in the mild PAD stage group (IC).

Impact of physical fitness and exercise training on subcutaneous adipose tissue beiging markers in humans with and without diabetes and a high-fat diet-fed mouse model.

AIMS: Exercise training induces white adipose tissue (WAT) beiging and improves glucose homeostasis and mitochondrial function in rodents. This could be relevant for type 2 diabetes in humans, but the effect of physical fitness on beiging of subcutaneous WAT (scWAT) remains unclear. This translational study investigates if beiging of scWAT associates with physical fitness in healthy humans and recent-onset type 2 diabetes and if a voluntary running wheel intervention is sufficient to induce beiging in mice. MATERIALS AND METHODS: Gene expression levels of established beiging markers were measured in scWAT biopsies of humans with (n = 28) or without type 2 diabetes (n = 28), stratified by spiroergometry into low (L-FIT; n = 14 each) and high physical fitness (H-FIT; n = 14 each). High-fat diet-fed FVB/N mice underwent voluntary wheel running, treadmill training or no training (n = 8 each group). Following the training intervention, mitochondrial respiration and content of scWAT were assessed by high-resolution respirometry and citrate synthase activity, respectively. RESULTS: Secreted CD137 antigen (Tnfrsf9/Cd137) expression was three-fold higher in glucose-tolerant H-FIT than in L-FIT, but not different between H-FIT and L-FIT with type 2 diabetes. In mice, both training modalities increased Cd137 expression and enhanced mitochondrial content without changing respiration in scWAT. Treadmill but not voluntary wheel running led to improved whole-body insulin sensitivity. CONCLUSIONS: Higher physical fitness and different exercise interventions associated with higher gene expression levels of the beiging marker CD137 in healthy humans and mice on a high-fat diet. Humans with recent-onset type 2 diabetes show an impaired adipose tissue-specific response to physical activity.

Adjuvant pharmacological strategies for the musculoskeletal system during long-term space missions.

Despite 2 h of daily exercise training, muscle wasting and bone loss are still present after 6-month missions to the international space station. Some crew members lose bone much faster than others. In preparation for missions to the Moon and Mars, space agencies are therefore reviewing their countermeasure portfolios. Here, we discuss the potential of current pharmacological strategies. Bone loss in space is fuelled by bone resorption. Alendronate, an oral bisphosphonate, reduced bone losses in experimental bed rest and space. However, gastrointestinal side effects precluded its further utilization in space. Zoledronate (a potent bisphosphonate), denosumab (RANKL antagonist) and romosozumab (sclerostin antagonist) are all administered via injection. They effectively suppress bone resorption and are routinely prescribed against osteoporosis. Their serious adverse effects, namely, osteonecrosis of the jaw and atypical femur fractures occur very rarely when the usage is limited to 1 or 2 years. Hence, utilization of one of these compounds may outweigh the bone risks of space travelling, in particular in those with high bone resorption rates. Muscle wasting in space is likely due to hampered muscle protein synthesis. Even though this might theoretically be countered by the synthesis-boosting effects of anabolic steroids, the practical grounds for such recommendation are currently weak. Moreover, they reveal their full potential only when combined with an anabolic exercise stimulus, for example, via strength training. It therefore seems that a combination of exercise and pharmacological countermeasures should be considered for musculoskeletal health on the way to the Moon and Mars and back.

High-intensity interval training for 12 weeks improves cardiovascular autonomic function but not somatosensory nerve function and structure in overweight men with type 2 diabetes.

AIMS/HYPOTHESIS: It remains unclear whether and which modality of exercise training as a component of lifestyle intervention may exert favourable effects on somatosensory and autonomic nerve tests in people with type 2 diabetes. METHODS: Cardiovascular autonomic and somatosensory nerve function as well as intraepidermal nerve fibre density (IENFD) were assessed in overweight men with type 2 diabetes (type 2 diabetes, n = 20) and male glucose-tolerant individuals (normal glucose tolerance [NGT], n = 23), comparable in age and BMI and serving as a control group, before and after a supervised high-intensity interval training (HIIT) intervention programme over 12 weeks. Study endpoints included clinical scores, nerve conduction studies, quantitative sensory testing, IENFD, heart rate variability, postural change in systolic blood pressure and spontaneous baroreflex sensitivity (BRS). RESULTS: After 12 weeks of HIIT, resting heart rate decreased in both groups ([mean ± SD] baseline/12 weeks: NGT: 65.1 ± 8.2/60.2 ± 9.0 beats per min; type 2 diabetes: 68.8 ± 10.1/63.4 ± 7.8 beats per min), while three BRS indices increased (sequence analysis BRS: 8.82 ± 4.89/14.6 ± 11.7 ms2/mmHg; positive sequences BRS: 7.19 ± 5.43/15.4 ± 15.9 ms2/mmHg; negative sequences BRS: 12.8 ± 5.4/14.6 ± 8.7 ms2/mmHg) and postural change in systolic blood pressure decreased (-13.9 ± 11.6/-9.35 ± 9.76 mmHg) in participants with type 2 diabetes, and two heart rate variability indices increased in the NGT group (standard deviation of R-R intervals: 36.1 ± 11.8/55.3 ± 41.3 ms; coefficient of R-R interval variation: 3.84 ± 1.21/5.17 ± 3.28) (all p<0.05). In contrast, BMI, clinical scores, nerve conduction studies, quantitative sensory testing, IENFD and the prevalence rates of diabetic sensorimotor polyneuropathy and cardiovascular autonomic neuropathy remained unchanged in both groups. In the entire cohort, correlations between the changes in two BRS indices and changes in [Formula: see text] over 12 weeks of HIIT (e.g. sequence analysis BRS: r = 0.528, p=0.017) were observed. CONCLUSIONS/INTERPRETATION: In male overweight individuals with type 2 diabetes, BRS, resting heart rate and orthostatic blood pressure regulation improved in the absence of weight loss after 12 weeks of supervised HIIT. Since no favourable effects on somatic nerve function and structure were observed, cardiovascular autonomic function appears to be more amenable to this short-term intervention, possibly due to improved cardiorespiratory fitness.

Dietary palmitate and oleate differently modulate insulin sensitivity in human skeletal muscle.

AIMS/HYPOTHESIS: Energy-dense nutrition generally induces insulin resistance, but dietary composition may differently affect glucose metabolism. This study investigated initial effects of monounsaturated vs saturated lipid meals on basal and insulin-stimulated myocellular glucose metabolism and insulin signalling. METHODS: In a randomised crossover study, 16 lean metabolically healthy volunteers received single meals containing safflower oil (SAF), palm oil (PAL) or vehicle (VCL). Whole-body glucose metabolism was assessed from glucose disposal (Rd) before and during hyperinsulinaemic-euglycaemic clamps with D-[6,6-2H2]glucose. In serial skeletal muscle biopsies, subcellular lipid metabolites and insulin signalling were measured before and after meals. RESULTS: SAF and PAL raised plasma oleate, but only PAL significantly increased plasma palmitate concentrations. SAF and PAL increased myocellular diacylglycerol and activated protein kinase C (PKC) isoform θ (p < 0.05) but only PAL activated PKCɛ. Moreover, PAL led to increased myocellular ceramides along with stimulated PKCζ translocation (p < 0.05 vs SAF). During clamp, SAF and PAL both decreased insulin-stimulated Rd (p < 0.05 vs VCL), but non-oxidative glucose disposal was lower after PAL compared with SAF (p < 0.05). Muscle serine1101-phosphorylation of IRS-1 was increased upon SAF and PAL consumption (p < 0.05), whereas PAL decreased serine473-phosphorylation of Akt more than SAF (p < 0.05). CONCLUSIONS/INTERPRETATION: Lipid-induced myocellular insulin resistance is likely more pronounced with palmitate than with oleate and is associated with PKC isoforms activation and inhibitory insulin signalling. TRIAL REGISTRATION: ClinicalTrials.gov .NCT01736202. FUNDING: German Federal Ministry of Health, Ministry of Culture and Science of the State North Rhine-Westphalia, German Federal Ministry of Education and Research, European Regional Development Fund, German Research Foundation, German Center for Diabetes Research.

Mitochondrial respiration is decreased in visceral but not subcutaneous adipose tissue in obese individuals with fatty liver disease.

BACKGROUND & AIMS: Adipose tissue dysfunction is involved in the development of insulin resistance and is responsible for excessive lipid delivery to other organs such as the liver. We tested the hypothesis that impaired mitochondrial function is a common feature of subcutaneous (SAT) and visceral adipose tissue (VAT), but may differently contribute to adipose tissue insulin resistance (IR) in obesity, non-alcoholic fatty liver (NAFL) and steatohepatitis (NASH). METHODS: In this cross-sectional study, we analyzed tissue-specific insulin sensitivity using stable isotope dilution and hyperinsulinemic-normoglycemic clamp tests. We also assessed mitochondrial respiration, mRNA and protein expression, and tissue morphology in biopsies of SAT and VAT from obese humans without NAFL, with NAFL or with NASH (n = 22/group). RESULTS: Compared to individuals without liver disease, persons with NAFL and NASH had about 30% (p = 0.010) and 33% (p = 0.002) lower maximal mitochondrial respiration, respectively, in VAT, but not in SAT. The lower maximal mitochondrial respiration of VAT was associated with lower adipose tissue insulin sensitivity (ß = 0.985, p = 0.041) and with increased VAT protein expression of tumor necrosis factor A across all groups (ß = -0.085, p = 0.040). VAT from individuals with NASH was characterized by lower expression of oxidative phosphorylation complex IV (p = 0.042) and higher mRNA expression of the macrophage marker CD68 (p = 0.002) than VAT from participants without NAFL. CONCLUSIONS: Humans with non-alcoholic fatty liver disease have distinct abnormalities of VAT energy metabolism, which correlate with adipose tissue dysfunction and may favor progression of NAFL to NASH. LAY SUMMARY: Adipose tissue (commonly called body fat) can be found under the skin (subcutaneous) or around internal organs (visceral). Dysfunction of adipose tissue can cause insulin resistance and lead to excess delivery of fat to other organs such as the liver. Herein, we show that dysfunction specifically in visceral adipose tissue was associated with fatty liver disease. CLINICAL TRIAL NUMBER: NCT01477957.

Dietary lipid droplet structure in postnatal life improves hepatic energy and lipid metabolism in a mouse model for postnatal programming.

Early-life diets may have a long-lasting impact on metabolic health. This study tested the hypothesis that an early-life diet with large, phospholipid-coated lipid droplets (Concept) induces sustained improvements of hepatic mitochondrial function and metabolism. Young C57BL/6j mice were fed Concept or control (CTRL) diet from postnatal day 15 (PN15) to PN42, followed by western style (WSD) or standard rodent diet (AIN) until PN98. Measurements comprised body composition, insulin resistance (HOMA-IR), tricarboxylic acid (TCA) cycle- and ß-oxidation-related hepatic oxidative capacity using high-resolution respirometry, mitochondrial dynamics, mediators of insulin resistance (diacylglycerols, DAG) or ceramides) in subcellular compartments as well as systemic oxidative stress. Concept feeding increased TCA cycle-related respiration by 33% and mitochondrial fusion protein-1 by 65% at PN42 (both p 0.05). At PN98, CTRL, but not Concept, mice developed hyperinsulinemia (CTRL/AIN 0.22 ± 0.44 vs. CTRL/WSD 1.49 ± 0.53 nmol/l, p 0.05 and Concept/AIN 0.20 ± 0.38 vs. Concept/WSD 1.00 ± 0.29 nmol/l, n.s.) and insulin resistance after WSD (CTRL/AIN 107 ± 23 vs. CTRL/WSD 738 ± 284, p 0.05 and Concept/AIN 109 ± 24 vs. Concept/WSD 524 ± 157, n.s.). WSD-induced liver weight was 18% lower in adult Concept-fed mice and ß-oxidation-related respiration was 69% higher (p 0.05; Concept/WSD vs. Concept/AIN) along with lower plasma lipid peroxides (CTRL/AIN 4.85 ± 0.28 vs. CTRL/WSD 5.73 ± 0.47 µmol/l, p 0.05 and Concept/AIN 4.49 ± 0.31 vs. Concept/WSD 4.42 ± 0.33 µmol/l, n.s.) and were in part protected from WSD-induced increase in hepatic cytosolic DAG C16:0/C18:1. Early-life feeding of Concept partly protected from WSD-induced insulin resistance and systemic oxidative stress, potentially via changes in specific DAG and mitochondrial function, highlighting the role of early life diets on metabolic health later in life.

The effect of revascularization on recovery of mitochondrial respiration in peripheral artery disease: a case control study.

BACKGROUND: Peripheral arterial disease (PAD) is accompanied by myopathy characterized by mitochondrial dysfunction. The aim of this experimental study was to investigate the effect of revascularization procedures on mitochondrial function in ischemic and non-ischemic muscle. METHODS: Muscle biopsies from patients with symptomatic stage IIB/III PAD caused by isolated pathologies of the superficial femoral artery were obtained from muscle regions within the chronic ischemic muscle (gastrocnemius) and from non-ischemic muscle (vastus lateralis) before and 6 weeks after invasive revascularization. High-resolution respirometry was used to investigate mitochondrial function and results were normalized to citrate synthase activity (CSA). Results are given in absolute values and fold over basal (FOB). RESULTS: Respiratory states (OXPHOS (P) and electron transfer (E) capacity) normalized to CSA decreased while CSA was increased in chronic ischemic muscle after revascularization. There were no changes in in non-ischemic muscle. The FOB of chronic ischemic muscle was significantly higher for CSA (chronic ischemic 1.37 (IQR 1.10-1.64) vs. non-ischemic 0.93 (IQR 0.69-1.16) p = 0.020) and significantly lower for respiratory states normalized to CSA when compared to the non-ischemic muscle (P per CSA chronic ischemic 0.64 (IQR 0.46-0.82) vs non-ischemic 1.16 (IQR 0.77-1.54) p = 0.011; E per CSA chronic ischemic 0.61 (IQR 0.47-0.76) vs. non-ischemic 1.02 (IQR 0.64-1.40) p = 0.010). CONCLUSIONS: Regeneration of mitochondrial content and function following revascularization procedures only occur in muscle regions affected by malperfusion. This indicates that the restoration of blood and oxygen supply are important mediators aiding mitochondrial recovery.

Improving insulin sensitivity, liver steatosis and fibrosis in type 2 diabetes by a food-based digital education-assisted lifestyle intervention program: a feasibility study.

PURPOSE: Recent trials demonstrated remission of type 2 diabetes and non-alcoholic fatty liver disease (NAFLD) following formula diet-induced weight loss. To improve the outreach for populations in need, many mobile health apps targeting weight loss have been developed with limited scientific evaluation of these apps. The present feasibility study investigated the effects of a novel approach incorporating a regular 'whole food-based' low-calorie diet combined with app-based digital education and behavioral change program on glucose metabolism and disease management. METHODS: Twenty-four individuals with type 2 diabetes followed this approach supported by weekly coaching calls for 12 weeks. Phenotyping included bioimpedance analysis, mixed-meal tolerance test, magnetic resonance spectroscopy and transient elastography for assessing liver fat content and liver stiffness. RESULTS: Over 12 weeks, participants reduced their body weight by 9% (97 ± 13 to 88 ± 12 kg), body mass index (BMI; 33 ± 5 to 29 ± 4 kg/m2), total fat mass (31 ± 10 to 27 ± 10%) (all p < 0.01) and liver fat by 50% alongside with decreased liver stiffness. Target HbA1c (< 6.5%) was achieved by 38% and resolution of NAFLD (liver fat content < 5.6%) was observed in 30% of the participants. CONCLUSION: This novel approach combining digital education with a low-calorie diet results in effective improvements of body weight, glycemic control and NAFLD and could complement existing care for patients with type 2 diabetes. TRIAL REGISTRATION: NCT04509245.

The Role of Mitochondrial Function in Peripheral Arterial Disease: Insights from Translational Studies.

Recent evidence demonstrates an involvement of impaired mitochondrial function in peripheral arterial disease (PAD) development. Specific impairments have been assessed by different methodological in-vivo (near-infrared spectroscopy, 31P magnetic resonance spectroscopy), as well as in-vitro approaches (Western blotting of mitochondrial proteins and enzymes, assays of mitochondrial function and content). While effects differ with regard to disease severity, chronic malperfusion impacts subcellular energy homeostasis, and repeating cycles of ischemia and reperfusion contribute to PAD disease progression by increasing mitochondrial reactive oxygen species production and impairing mitochondrial function. With the leading clinical symptom of decreased walking capacity due to intermittent claudication, PAD patients suffer from a subsequent reduction of quality of life. Different treatment modalities, such as physical activity and revascularization procedures, can aid mitochondrial recovery. While the relevance of these modalities for mitochondrial functional recovery is still a matter of debate, recent research indicates the importance of revascularization procedures, with increased physical activity levels being a subordinate contributor, at least during mild stages of PAD. With an additional focus on the role of revascularization procedures on mitochondria and the identification of suitable mitochondrial markers in PAD, this review aims to critically evaluate the relevance of mitochondrial function in PAD development and progression.

Regeneration of Mitochondrial Function in Gastrocnemius Muscle in Peripheral Arterial Disease After Successful Revascularisation.

OBJECTIVE: Myopathy, characterised by altered mitochondrial function, is a central part of the pathophysiology of peripheral arterial disease and the aim of this study was to investigate the effect of revascularisation on mitochondrial function. METHODS: High resolution respirometry was used to investigate mitochondrial respiration and the results were normalised to citrate synthase activity (CSA), a marker of mitochondrial content. Ten patients with symptomatic peripheral arterial disease (study group) and 10 subjects without ischaemia (control group) were included. Ankle brachial index and ultrasound imaging were performed before and after vascular intervention to confirm technically successful revascularisation. Within the study group, muscle biopsies from the gastrocnemius muscle were taken before vascular intervention and six weeks after revascularisation. Within the control group, tissue was harvested once. RESULTS: There were no significant group differences regarding anthropometric data. CSA showed a significant increase after successful revascularisation (CSA pre-operative 281.4 (252.4-391.8) nmol/min/mg protein vs. CSA post-operative 438.5 (361.4-471.3) nmol/min/mg protein; p = .01) with post-operative return of values to the range of control subjects (CSA control 396.6 (308.2-435.9)). Mitochondrial respiration normalised to CSA in oxidative phosphorylation (P) as well as in electron transfer (E) capacity were significantly reduced post-operatively when compared with pre-operative values (P pre-operative 0.218 (0.196-0.266) pmol/(sec×mg) per CSA vs. post-operative 0.132 (0.116-0.150) pmol/(sec×mg) per CSA, p = .007; E pre-operative 0.230 (0.195-0.279) pmol/(sec×mg) per CSA vs. post-operative 0.129 (0.120-0.154) pmol/(sec×mg) per CSA, p = .005) meaning a post-operative return of values to within the range of control subjects (P control 0.124 (0.080-0.155) pmol/(sec×mg) per CSA; E control 0.121 (0.079-0.125) pmol/(sec×mg) per CSA). CONCLUSION: With these results, it has been shown that the initially impaired mitochondrial function and content can normalise after revascularisation.

Breathlessness and Restrictive Lung Disease: An Important Diabetes-Related Feature in Patients with Type 2 Diabetes.

BACKGROUND: Diabetes mellitus is a significant comorbidity of interstitial lung disease (ILD). OBJECTIVES: The aim of this study was to investigate the incidence of restrictive lung disease (RLD) and ILD in patients with prediabetes and type 2 diabetes (T2D). METHODS: Forty-eight nondiabetics, 68 patients with prediabetes, 29 newly diagnosed T2D, and 110 patients with long-term T2D were examined for metabolic control, diabetes-related complications, breathlessness, and lung function. Five participants with T2D, breathlessness, and RLD underwent multidetector computed tomography (MDCT) and a Six-Minute Walk Test (6MWT). Lung tissue from 4 patients without diabetes and from 3 patients with T2D was histologically examined for presence of pulmonary fibrosis. RESULTS: Breathlessness in combination with RLD was significantly increased in patients with prediabetes and T2D (p < 0.01). RLD was found in 9% of patients with prediabetes, in 20% of patients with newly diagnosed T2D, and in 27% of patients with long-term T2D. Thus, patients with long-term T2D had an increased risk of RLD (OR 5.82 [95% CI 1.71-20.5], p < 0.01). RLD was significantly associated with glucose metabolism and albuminuria (p < 0.01); furthermore, presence of nephropathy increased the risk of RLD (OR 8.57 [95% CI 3.4-21.9], p < 0.01) compared to nondiabetics. MDCT revealed ILD in 4 patients, the 6MWT correlated with the extent of ILD, and histological analysis showed fibrosing ILD in patients with T2D. CONCLUSIONS: This study demonstrates increased breathlessness and a high prevalence of RLD in patients with T2D, indicating an association between diabetes and fibrosing ILD.

Evaluation of mitochondrial function in chronic myofascial trigger points - a prospective cohort pilot study using high-resolution respirometry.

BACKGROUND: Myofascial trigger points (MTrPs) are hyperirritable areas in the fascia of the affected muscle, possibly related to mitochondrial impairment. They can result in pain and hypoxic areas within the muscle. This pilot study established a minimally invasive biopsy technique to obtain high-quality MTrP tissue samples to evaluate mitochondrial function via high-resolution respirometry. Secondary objectives included the feasibility and safety of the biopsy procedure. METHODS: Twenty healthy males participated in this study, 10 with a diagnosis of myofascial pain in the musculus (m.) trapezius MTrP (TTP group) and 10 with a diagnosis of myofascial pain in the m. gluteus medius (GTP group). Each participant had 2 muscle biopsies taken in one session. The affected muscle was biopsied followed by a biopsy from the m. vastus lateralis to be used as a control. Measurements of oxygen consumption were carried out using high-resolution respirometry. RESULTS: Mitochondrial respiration was highest in the GTP group compared to the TTP group and the control muscle whereas no differences were observed between the GTP and the control muscle. When normalizing respiration to an internal reference state, there were no differences between muscle groups. None of the participants had hematomas or reported surgical complications. Patient-reported pain was minimal for all 3 groups. All participants reported a low procedural burden. CONCLUSIONS: This pilot study used a safe and minimally invasive technique for obtaining biopsies from MTrPs suitable for high-resolution respirometry analysis of mitochondrial function. The results suggest that there are no qualitative differences in mitochondrial function of MTrPs of the trapezius and gluteus medius muscles compared to the vastus lateralis control muscle, implying that alterations of mitochondrial function do not appear to have a role in the development of MTrPs. TRIAL REGISTRATION: Registered as No. 20131128-850 at the Coordinating Center for Clinical Studies of the Medical University of Innsbruck, trial registration date: 28th November 2013 and retrospectively registered on 11th of October 2018 at ClinicalTrials.gov with the ID NCT03704311 .

Neuronal UCP1 expression suggests a mechanism for local thermogenesis during hibernation.

Hibernating mammals possess a unique ability to reduce their body temperature to ambient levels, which can be as low as -2.9 °C, by active down-regulation of metabolism. Despite such a depressed physiologic phenotype, hibernators still maintain activity in their nervous systems, as evidenced by their continued sensitivity to auditory, tactile, and thermal stimulation. The molecular mechanisms that underlie this adaptation remain unknown. We report, using differential transcriptomics alongside immunohistologic and biochemical analyses, that neurons from thirteen-lined ground squirrels (Ictidomys tridecemlineatus) express mitochondrial uncoupling protein 1 (UCP1). The expression changes seasonally, with higher expression during hibernation compared with the summer active state. Functional and pharmacologic analyses show that squirrel UCP1 acts as the typical thermogenic protein in vitro. Accordingly, we found that mitochondria isolated from torpid squirrel brain show a high level of palmitate-induced uncoupling. Furthermore, torpid squirrels during the hibernation season keep their brain temperature significantly elevated above ambient temperature and that of the rest of the body, including brown adipose tissue. Together, our findings suggest that UCP1 contributes to local thermogenesis in the squirrel brain, and thus supports nervous tissue function at low body temperature during hibernation.

Hypophosphatemia promotes lower rates of muscle ATP synthesis.

Hypophosphatemia can lead to muscle weakness and respiratory and heart failure, but the mechanism is unknown. To address this question, we noninvasively assessed rates of muscle ATP synthesis in hypophosphatemic mice by using in vivo saturation transfer [31P]-magnetic resonance spectroscopy. By using this approach, we found that basal and insulin-stimulated rates of muscle ATP synthetic flux (VATP) and plasma inorganic phosphate (Pi) were reduced by 50% in mice with diet-induced hypophosphatemia as well as in sodium-dependent Pi transporter solute carrier family 34, member 1 (NaPi2a)-knockout (NaPi2a-/-) mice compared with their wild-type littermate controls. Rates of VATP normalized in both hypophosphatemic groups after restoring plasma Pi concentrations. Furthermore, VATP was directly related to cellular and mitochondrial Pi uptake in L6 and RC13 rodent myocytes and isolated muscle mitochondria. Similar findings were observed in a patient with chronic hypophosphatemia as a result of a mutation in SLC34A3 who had a 50% reduction in both serum Pi content and muscle VATP After oral Pi repletion and normalization of serum Pi levels, muscle VATP completely normalized in the patient. Taken together, these data support the hypothesis that decreased muscle ATP synthesis, in part, may be caused by low blood Pi concentrations, which may explain some aspects of muscle weakness observed in patients with hypophosphatemia.-Pesta, D. H., Tsirigotis, D. N., Befroy, D. E., Caballero, D., Jurczak, M. J., Rahimi, Y., Cline, G. W., Dufour, S., Birkenfeld, A. L., Rothman, D. L., Carpenter, T. O., Insogna, K., Petersen, K. F., Bergwitz, C., Shulman, G. I. Hypophosphatemia promotes lower rates of muscle ATP synthesis.

The Janus Head of Oxidative Stress in Metabolic Diseases and During Physical Exercise.

PURPOSE OF REVIEW: Oxidative stress describes an imbalance between production and degradation of reactive oxygen species (ROS), which can damage macromolecules. However, ROS may also serve as signaling molecules activating cellular pathways involved in cell proliferation and adaptation. This review describes alterations in metabolic diseases including obesity, insulin resistance, and/or diabetes mellitus as well as responses to acute and chronic physical exercise. RECENT FINDINGS: Chronic upregulation of oxidative stress associates with the development of insulin resistance and type 2 diabetes (T2D). While single bouts of exercise can transiently induce oxidative stress, chronic exercise promotes favorable oxidative adaptations with improvements in muscle mitochondrial biogenesis and glucose uptake. Although impaired antioxidant defense fails to scavenge ROS in metabolic diseases, chronic exercising can restore this abnormality. The different metabolic effects are likely due to variability of reactive species and discrepancies in temporal (acute vs. chronic) and local (subcellular distribution) patterns of production.

Genetic activation of pyruvate dehydrogenase alters oxidative substrate selection to induce skeletal muscle insulin resistance.

The pyruvate dehydrogenase complex (PDH) has been hypothesized to link lipid exposure to skeletal muscle insulin resistance through a glucose-fatty acid cycle in which increased fatty acid oxidation increases acetyl-CoA concentrations, thereby inactivating PDH and decreasing glucose oxidation. However, whether fatty acids induce insulin resistance by decreasing PDH flux remains unknown. To genetically examine this hypothesis we assessed relative rates of pyruvate dehydrogenase flux/mitochondrial oxidative flux and insulin-stimulated rates of muscle glucose metabolism in awake mice lacking pyruvate dehydrogenase kinase 2 and 4 [double knockout (DKO)], which results in constitutively activated PDH. Surprisingly, increased glucose oxidation in DKO muscle was accompanied by reduced insulin-stimulated muscle glucose uptake. Preferential myocellular glucose utilization in DKO mice decreased fatty acid oxidation, resulting in increased reesterification of acyl-CoAs into diacylglycerol and triacylglycerol, with subsequent activation of PKC-θ and inhibition of insulin signaling in muscle. In contrast, other putative mediators of muscle insulin resistance, including muscle acylcarnitines, ceramides, reactive oxygen species production, and oxidative stress markers, were not increased. These findings demonstrate that modulation of oxidative substrate selection to increase muscle glucose utilization surprisingly results in muscle insulin resistance, offering genetic evidence against the glucose-fatty acid cycle hypothesis of muscle insulin resistance.

Immortalised murine R349P desmin knock-in myotubes exhibit a reduced proton leak and decreased ADP/ATP translocase levels in purified mitochondria.

Desmin gene mutations cause myopathies and cardiomyopathies. Our previously characterised R349P desminopathy mice, which carry the ortholog of the common human desmin mutation R350P, showed marked alterations in mitochondrial morphology and function in muscle tissue. By isolating skeletal muscle myoblasts from offspring of R349P desminopathy and p53 knock-out mice, we established an immortalised cellular disease model. Heterozygous and homozygous R349P desmin knock-in and wild-type myoblasts could be well differentiated into multinucleated spontaneously contracting myotubes. The desminopathy myoblasts showed the characteristic disruption of the desmin cytoskeleton and desmin protein aggregation, and the desminopathy myotubes showed the characteristic myofibrillar irregularities. Long-term electrical pulse stimulation promoted myotube differentiation and markedly increased their spontaneous contraction rate. In both heterozygous and homozygous R349P desminopathy myotubes, this treatment restored a regular myofibrillar cross-striation pattern as seen in wild-type myotubes. High-resolution respirometry of mitochondria purified from myotubes by density gradient ultracentrifugation revealed normal oxidative phosphorylation capacity, but a significantly reduced proton leak in mitochondria from the homozygous R349P desmin knock-in cells. Consistent with a reduced proton flux across the inner mitochondrial membrane, our quantitative proteomic analysis of the purified mitochondria revealed significantly reduced levels of ADP/ATP translocases in the homozygous R349P desmin knock-in genotype. As this alteration was also detected in the soleus muscle of R349P desminopathy mice, which, in contrast to the mitochondria purified from cultured cells, showed a variety of other dysregulated mitochondrial proteins, we consider this finding to be an early step in the pathogenesis of secondary mitochondriopathy in desminopathy.

Dietary Intake of Athletes at the World Masters Athletics Championships as Assessed by Single 24 h Recall.

Proper dietary intake is important for masters athletes because of the physiological changes that occur with aging and the unique nutritional needs when competing at high levels. We evaluated the dietary intake of masters athletes competing at the World Masters Athletics Championships (outdoor games, Tampere, Finland, 2022, and indoor games, Torun, Poland, 2023). A total of 43 athletes (16 females and 27 males, mean age 59.2 ± 10.3 y, height 168 ± 8 cm, and body mass 62.3 ± 10.8 kg) participating in endurance (n = 21), sprint (n = 16), jumping (2), multi-component (e.g., decathlon; n = 3), and throwing (n = 1) events provided 24 h dietary recalls while participating in the games. Carbohydrate intake was below the recommended levels for endurance athletes. Protein intake was below the recommended levels for masters athletes, except for female athletes involved in power events (i.e., sprinters and jumpers). Other nutrient intakes that were below the recommended levels included vitamins D and E, calcium, potassium, vitamin A (except for female endurance athletes), folate (except for female power athletes), vitamin C for female endurance athletes, vitamin K and fiber for males, and zinc for endurance athletes. We conclude that while competing at world championships, many athletes are not consuming the recommended levels of carbohydrates, protein, and micronutrients. Athletes attending these games would benefit from increased nutritional support.

Loss of mitochondrial adaptation associates with deterioration of mitochondrial turnover and structure in metabolic dysfunction-associated steatotic liver disease.

BACKGROUND: Obesity and type 2 diabetes frequently have metabolic dysfunction-associated steatotic liver disease (MASLD) including steatohepatitis (MASH). In obesity, the liver may adapt its oxidative capacity, but the role of mitochondrial turnover in MASLD remains uncertain. METHODS: This cross-sectional study compared individuals with class III obesity (n = 8/group) without (control, OBE CON; NAFLD activity score: 0.4 ± 0.1) or with steatosis (OBE MASL, 2.3 ± 0.4), or MASH (OBE MASH, 5.3 ± 0.3, p < 0.05 vs. other groups). Hepatic mitochondrial ultrastructure was assessed by transmission electron microscopy, mitochondrial respiration by high-resolution respirometry, biomarkers of mitochondrial quality control and endoplasmic reticulum (ER) stress by Western Blot. RESULTS: Mitochondrial oxidative capacity was 31 % higher in OBE MASL, but 25 % lower in OBE MASH (p < 0.05 vs. OBE CON). OBE MASH showed ~1.5fold lower mitochondrial number, but ~1.2-1.5fold higher diameter and area (p < 0.001 vs. other groups). Biomarkers of autophagy (p62), mitophagy (PINK1, PARKIN), fission (DRP-1, FIS1) and fusion (MFN1/2, OPA1) were reduced in OBE MASH (p < 0.05 vs. OBE CON). OBE MASL showed lower p62, p-PARKIN/PARKIN, and p-DRP-1 (p < 0.05 vs. OBE CON). OBE MASL and MASH showed higher ER stress markers (PERK, ATF4, p-eIF2α-S51/eIF2α; p < 0.05 vs. OBE CON). Mitochondrial diameter associated inversely with fusion/fission biomarkers and with oxidative capacity, but positively with H2O2. CONCLUSION: Humans with hepatic steatosis already exhibit impaired mitochondrial turnover, despite upregulated oxidative capacity, and evidence for ER stress. In MASH, oxidative stress likely mediates progressive decline of mitochondrial turnover, ultrastructure and respiration indicating that mitochondrial quality control is key for energy metabolism and may have potential for targeting MASH. ClinGovTrial:NCT01477957.