Respiration In Skeletal Muscle Research Articles

Localization Of Myoglobin In Mitochondria

PURPOSE: Mitochondria play a principal role for metabolism and have a primary role in regulating respiration and energy expenditure. Recently, we showed that the muscle- specific protein myoglobin (Mb) interacted with complex IV to augment mitochondrial respiratory capacity in skeletal muscles. However, the precise mechanism for the Mb- mediated upregulation remains under debate. The present study has focused on localizing Mb within the mitochondria. METHODS: Muscle specimen from deep portion of m. Gastrocnemius in Wistar rat was homogenized. Crude mitochondria were isolated by differential centrifugations and washed with the mitochondrial isolation buffer. The isolated mitochondria were treated with proteinase K (PK), osmotic shock (OS), and SDS (or TriX) in order to digest proteins on the outer membrane and in the intramembrane. The final samples were subjected to SDS-PAGE and immunoblotting using antibodies to localize the proteins in the mitochondria. RESULTS: Western blotting analysis revealed that the PK digests Tom20, which localized on the outer membrane of mitochondria. The Tom20 band intensity decreased with the amount of PK used. Other mitochondrial proteins such as cytochrome c (intermembrane space), COX-IV (inner membrane), and PDH (matrix), were not affected by PK treatment. PK treatment did not affect Mb. The results suggested that Mb did not localize on the outer membrane of mitochondria. The combined treatment of PK, OS and SDS (or TrX) allowed immunoblotting detection of the mitochondrial proteins in specific regions of the mitochondria. For example, cytochrome c disappeared with OS treatment. Timm22 disappeared with PK+OS treatment. However, Mb was detected with either PK or OS treatment. But it cannot be detected with a combined PK+OS treatment. The results suggest that Mb associated with the inner membrane (intramembrane side, not matrix side) of the mitochondria. CONCLUSIONS: We conclude that Mb in muscle cells localizes in the cytosol and in the mitochondrial intermembrane space. Since exercise training increases Mb expressions of skeletal muscle, the increased Mb concentration may play a direct role in modulation skeletal muscle respiration and oxidative phosphorylation capacity.

Chemotherapeutic agents induce mitochondrial superoxide production and toxicity but do not alter respiration in skeletal muscle in vitro

Chemotherapeutic agents (CAs) can independently promote skeletal muscle dysfunction, fatigue and wasting with mitochondrial toxicity implicated as a possible mechanism. Thus, we aimed to characterise the effects of various CAs on mitochondrial function, viability and oxidant production in C2C12 myoblasts and myotubes. All CAs significantly reduced the viable mitochondrial pool but did not affect mitochondrial functional parameters. Doxorubicin and oxaliplatin increased oxidant production in myotubes while all CAs, except for irinotecan, increased oxidant production in myoblasts and reduced myotube diameter. Our data demonstrate CAs mito-toxic effects, highlighting the potential for mitochondria-protective therapeutics to address chemotherapy-induced skeletal muscle damage.

Long-term rates of mitochondrial protein synthesis are increased in mouse skeletal muscle with high-fat feeding regardless of insulin-sensitizing treatment.

Skeletal muscle mitochondrial protein synthesis is regulated in part by insulin. The development of insulin resistance with diet-induced obesity may therefore contribute to impairments to protein synthesis and decreased mitochondrial respiration. Yet the impact of diet-induced obesity and insulin resistance on mitochondrial energetics is controversial, with reports varying from decreases to increases in mitochondrial respiration. We investigated the impact of changes in insulin sensitivity on long-term rates of mitochondrial protein synthesis as a mechanism for changes to mitochondrial respiration in skeletal muscle. Insulin resistance was induced in C57BL/6J mice using 4 wk of a high-fat compared with a low-fat diet. For 8 additional weeks, diets were enriched with pioglitazone to restore insulin sensitivity compared with nonenriched control low-fat or high-fat diets. Skeletal muscle mitochondrial protein synthesis was measured using deuterium oxide labeling during weeks 10-12 High-resolution respirometry was performed using palmitoyl-l-carnitine, glutamate+malate, and glutamate+malate+succinate as substrates for mitochondria isolated from quadriceps. Mitochondrial protein synthesis and palmitoyl- l-carnitine oxidation were increased in mice consuming a high-fat diet, regardless of differences in insulin sensitivity with pioglitazone treatment. There was no effect of diet or pioglitazone treatment on ADP-stimulated respiration or H2O2 emission using glutamate+malate or glutamate+malate+succinate. The results demonstrate no impairments to mitochondrial protein synthesis or respiration following induction of insulin resistance. Instead, mitochondrial protein synthesis was increased with a high-fat diet and may contribute to remodeling of the mitochondria to increase lipid oxidation capacity. Mitochondrial adaptations with a high-fat diet appear driven by nutrient availability, not intrinsic defects that contribute to insulin resistance.

Impaired Insulin Signaling is Associated with Hepatic Mitochondrial Dysfunction in IR+/--IRS-1+/- Double Heterozygous (IR-IRS1dh) Mice.

Mitochondria play a pivotal role in energy metabolism, but whether insulin signaling per se could regulate mitochondrial function has not been identified yet. To investigate whether mitochondrial function is regulated by insulin signaling, we analyzed muscle and liver of insulin receptor (IR)+/−-insulin receptor substrate-1 (IRS-1)+/− double heterozygous (IR-IRS1dh) mice, a well described model for insulin resistance. IR-IRS1dh mice were studied at the age of 6 and 12 months and glucose metabolism was determined by glucose and insulin tolerance tests. Mitochondrial enzyme activities, oxygen consumption, and membrane potential were assessed using spectrophotometric, respirometric, and proton motive force analysis, respectively. IR-IRS1dh mice showed elevated serum insulin levels. Hepatic mitochondrial oxygen consumption was reduced in IR-IRS1dh animals at 12 months of age. Furthermore, 6-month-old IR-IRS1dh mice demonstrated enhanced mitochondrial respiration in skeletal muscle, but a tendency of impaired glucose tolerance. On the other hand, 12-month-old IR-IRS1dh mice showed improved glucose tolerance, but normal muscle mitochondrial function. Our data revealed that deficiency in IR/IRS-1 resulted in normal or even elevated skeletal muscle, but impaired hepatic mitochondrial function, suggesting a direct cross-talk between insulin signaling and mitochondria in the liver.

Neuregulin 1 improves complex 2-mediated mitochondrial respiration in skeletal muscle of healthy and diabetic mice

It has been reported that neuregulin1 (NRG1) improves glucose tolerance in healthy and diabetic rodents. In vitro studies also suggest that NRG1 regulates myocyte oxidative capacity. To confirm this observation in vivo, we evaluated the effect on mitochondrial function of an 8-week treatment with NRG1 in db/db diabetic mice and C57BL/6JRJ healthy controls. NRG1 treatment improved complex 2-mediated mitochondrial respiration in the gastrocnemius of both control and diabetic mice and increased mitochondrial complex 2 subunit content by 2-fold. This effect was not associated with an increase in mitochondrial biogenesis markers. Enhanced ERBB4 phosphorylation could mediate NRG1 effects on mitochondrial function through signalling pathways, independently of ERK1/2, AKT or AMPK.

Effects of WIN 55,212‐2 and AM281 in mitochondrial respiration of skeletal muscle

The endocannabinoid system is emerging as a critical agent of cellular metabolism regulation, through its effects on the cellular respiration and energy production. Most of the cannabinoid and endocannabinoid effects have been mediated predominantly via the activation of the cannabinoid type 1 (CB1) receptor, which is widely distributed in the brain and peripheral organs as the skeletal muscle and recently were confirmed to be localized on the outer mitochondrial membrane of these tissues. It is known that their activation by synthetic cannabinoids is able to cause changes in mitochondrial respiration of brain mitochondria, however, little is known about its effect in skeletal muscle. Thus, the aim of this work was to explore the effects of the cannabinoid agonist WIN 55,212‐2 and the CB1 antagonist receptor AM281 on oxygen consumption in the state 3 from mitochondrial respiration in isolated mitochondria derived from rat skeletal muscle.MethodsMitochondria from hindlimb muscles of male Wistar rats weighing between 300 and 350 were isolated by differential centrifugation. The rate of oxygen consumption was determined in a sealed glass chamber with constant stirring at room temperature, using a Clark‐type oxygen electrode coupled to an oxygen monitor YSI 5300 and a computer for data acquisition. The determinations started after adding 10 mM glutamate/malate as respiratory substrate for complex I (state 4) and 1 mM ADP was added to determine oxygen consumption in the phosphorylating state (state 3). Finally, the inhibitor of complex IV (1mM KCN) were added to inhibit mitochondrial respiration. To explore the effects of the experimental drugs on mitochondrial respiration, a concentration‐response curve was performed for each of them: WIN 55,212‐2 (0.1, 1, 5 and 10 μM) and AM281(1, 5, 10 and 20 μM).Our results show that both drugs have similar effects by decrease mitochondrial respiration, this inhibitory effects on oxygen consumption is dose‐dependent by WIN 55,212‐2 being more prominent with 10 μM (40%) and to a lesser extent with 0.1 μM (≈ 20%), while with AM281 decreased 50% with 1 μM. These results suggest that the effect by this drugs on mitochondrial respiration exert a similar effect as has been previously reported in brain mitochondria and that could be acting on other respiratory chain components.Support or Funding InformationWe thank to Universidad de Colima, Universidad Michoacana de San Nicolás de Hidalgo (UMSNH) and Consejo Nacional de de Ciencia y Tecnología (CONACyT) the partial financial of this research. ESD was a receipient of a CONACyT PhD fellow. We acknowledge the partial financing to Coordinación de la Investigación Científica of UMSNH: ASM‐CIC‐2.16; RMP‐CIC 182171.

Relationship of pectoralis muscle area and skeletal muscle strength with exercise tolerance and dyspnea in interstitial lung disease.

Background: Pectoralis muscle area (PMA) is an easily derived computed tomography-based assessment that can provide insight into clinical features of other skeletal muscles. Respiratory and locomotor muscle dysfunction has been increasingly recognized in patients with interstitial lung disease (ILD). Its contribution to exercise performance has been controversial. Objective: We aimed to investigate if PMA is related with respiratory and locomotor skeletal muscle strength in ILD patients, and if skeletal muscle function is compromised and independently related with exercise capacity and dyspnea. Methods: Cross-sectional study where subjects performed incremental cycling cardiopulmonary exercise testing with maximal inspiratory (MIP) and expiratory (MEP) pressure measurements, and quadriceps maximal voluntary contraction (MVC) before and after exercise. Results: Thirty ILD patients (forced vital capacity [FVC] and lung diffusing capacity [DLCO] of 60±15% and 38±10% of predicted, respectively) and 15 healthy control subjects were studied. Patients presented significantly lower MIP and qMVC compared to controls. PMA was significantly associated with qMVC only (r=0.506; p<0.01). Only expiratory muscles showed a significant strength decline after exercise, both in patients and controls. In multivariate regression analysis, only FVC remained as independent predictor of peak aerobic capacity and MEP post exercise remained as independent predictor of peak exercise dyspnea even adjusting for FVC. Conclusion: ILD patients exhibited reduced inspiratory and quadriceps strength, but PMA was associated with the later only. Muscle strength was not associated with exercise capacity while expiratory muscle fatigue might underlie exertional dyspnea. (Sarcoidosis Vasc Diffuse Lung Dis 2017; 34: 200-208)

Similar efficacy from specific and non-specific mineralocorticoid receptor antagonist treatment of muscular dystrophy mice.

Combined treatment with an angiotensin-converting enzyme inhibitor and a mineralocorticoid receptor (MR) antagonist improved cardiac and skeletal muscle function and pathology in a mouse model of Duchenne muscular dystrophy. MR is present in limb and respiratory skeletal muscles and functions as a steroid hormone receptor. The goals of the current study were to compare the efficacy of the specific MR antagonist eplerenone with the non-specific MR antagonist spironolactone, both in combination with the angiotensin-converting enzyme inhibitor lisinopril. Three groups of n=18 dystrophin-deficient, utrophin-haploinsufficient male mice were given chow containing: lisinopril plus spironolactone, lisinopril plus eplerenone, or no drug, from four to 20 weeks-of-age. Eighteen C57BL/10 male mice were used as wild-type controls. In vivo measurements included cardiac magnetic resonance imaging, conscious electrocardiography, and grip strength. From each mouse in the study, diaphragm, extensor digitorum longus, and cardiac papillary muscle force was measured ex vivo, followed by histological quantification of muscle damage in heart, diaphragm, quadriceps, and abdominal muscles. MR protein levels were also verified in treated muscles. Treatment with specific and non-specific MR antagonists did not result in any adverse effects to dystrophic skeletal muscles or heart. Both treatments resulted in similar functional and pathological improvements across a wide array of parameters. MR protein levels were not reduced by treatment. These data suggest that spironolactone and eplerenone show similar effects in dystrophic mice and support the clinical development of MR antagonists for treating skeletal muscles in Duchenne muscular dystrophy.

Hypoxic Preconditioning Strengthens Respiratory Skeletal Muscle Function during Reoxygenation via Mitochondrial Regulation

Hypoxic Preconditioning Strengthens Respiratory Skeletal Muscle Function during Reoxygenation via Mitochondrial Regulation

The Angiotensin Converting Enzyme Inhibitor Lisinopril Improves Muscle Histopathology but not Contractile Function in a Mouse Model of Duchenne Muscular Dystrophy.

Background: Angiotensin converting enzyme inhibitors (ACEi) are the current standard of care treatment for cardiac dysfunction in Duchenne muscular dystrophy patients. We previously showed treatment with an ACEi plus mineralocorticoid receptor (MR) antagonist improves limb and respiratory skeletal muscles, in addition to cardiac muscles, in a dystrophic mouse model at 20 weeks-of-age. Objective: To determine whether previously observed preclinical benefits of an ACEi plus MR antagonist on dystrophic skeletal muscles can be reproduced by increasing ACEi dosage alone. We also compared functional and histological outcome measures at 10 and 20 weeks-of-age. Methods: Dystrophin deficient utrophin haplo-insufficient (utrn +/- ; mdx) “het” mice were treated with 10, 20, or 50 mg/kg × day of the ACEi lisinopril from 4 to 10 weeks-of-age via water bottles and compared with C57BL/10 wild-type control mice and untreated hets. Data from 10 week-old het mice were also compared to data collected from an untreated het group at 20 weeks-old. In vivo cardiac and grip strength measurements, in vitro diaphragm and extensor digitorum longus muscle force measurements, and histopathological analyses were performed. One-way ANOVA followed by Dunnett post hoc comparison was used to determine significance. Results: ACEi treatment reduced skeletal muscle damage but had no significant effect on muscle force. Body weight, heart rate, grip strength and blood pressure were unaffected by treatment. Limb muscle histopathology was more informative at 10 than 20 weeks-of-age.Conclusions: These results suggest increased ACEi dosage alone cannot improve all dystrophic parameters. Further optimization of MR antagonists in 20 week-old mice is warranted.

The Effect of Succinate on Skeletal Muscle Respiration in Diet Induced Obesity and Insulin Resistance

Obesity and insulin resistance are associated with mitochondrial dysfunction. We sought to determine the effect of succinate, an electron donor at complex II of the mitochondria, on insulin action, energy homeostasis, and muscle mitochondrial function. C57B6 male mice were randomly assigned to a low fat diet (LFD) or a high fat diet (HFD) for 20 weeks. After 14 weeks of the dietary intervention, mice from within a diet group were assigned to a control or succinate (0.75 mg/ml in drinking water) group. Caloric intake, water consumption, and body mass were assessed weekly, while glucose tolerance (GT), insulin tolerance (IT), energy expenditure (EE), and spontaneous physical activity (PA) were assessed during the last 2 weeks of succinate treatment. In vitro measurements of mitochondrial function in soleus muscle were performed using permeabilized fiber respirometry. As expected, the HFD increased body mass, epididymal white adipose tissue (EWAT) mass, and reduced GT and IT. However, succinate treatment had no effect on any in vivo measurements (body mass, EWAT mass, GT, IT, EE, or PA). Despite impaired insulin action, there was no effect of diet on soleus muscle mitochondrial respiration, however, a trend to increase complex I+II driven state 3 respiration, and complex IV activity was evident in mice that received succinate. In conclusion, we observed no effect of succinate treatment on insulin action, energy homeostasis, or physical activity. In oxidative skeletal muscle, there was a tendency for increased state 3 respiration, owed to greater mitochondrial content, suggestive of a succinate‐induced mitochondrial biogenesis.

The clinical heterogeneity of coenzyme Q10 deficiency results from genotypic differences in the Coq9 gene.

Primary coenzyme Q10 (CoQ10) deficiency is due to mutations in genes involved in CoQ biosynthesis. The disease has been associated with five major phenotypes, but a genotype–phenotype correlation is unclear. Here, we compare two mouse models with a genetic modification in Coq9 gene (Coq9Q95X and Coq9R239X), and their responses to 2,4-dihydroxybenzoic acid (2,4-diHB). Coq9R239X mice manifest severe widespread CoQ deficiency associated with fatal encephalomyopathy and respond to 2,4-diHB increasing CoQ levels. In contrast, Coq9Q95X mice exhibit mild CoQ deficiency manifesting with reduction in CI+III activity and mitochondrial respiration in skeletal muscle, and late-onset mild mitochondrial myopathy, which does not respond to 2,4-diHB. We show that these differences are due to the levels of COQ biosynthetic proteins, suggesting that the presence of a truncated version of COQ9 protein in Coq9R239X mice destabilizes the CoQ multiprotein complex. Our study points out the importance of the multiprotein complex for CoQ biosynthesis in mammals, which may provide new insights to understand the genotype–phenotype heterogeneity associated with human CoQ deficiency and may have a potential impact on the treatment of this mitochondrial disorder.

Improvement Characteristics of Bio-active Materials Coated Fabric on Rat Muscular Mitochondria.

This study surveys the improvement characteristics in old-aged muscular mitochondria by bio-active materials coated fabric (BMCF). To observe the effects, the fabric (10 and 30%) was worn to old-aged rat then the oxygen consumption efficiency and copy numbers of mitochondria, and mRNA expression of apoptosis- and mitophagy-related genes were verified. By wearing the BMCF, the oxidative respiration significantly increased when using the 30% materials coated fabric. The mitochondrial DNA copy number significantly decreased and subsequently recovered in a dose-dependent manner. The respiratory control ratio to mitochondrial DNA copy number showed a dose-dependent increment. As times passed, Bax, caspase 9, PGC-1α and β-actin increased, and Bcl-2 decreased in a dose-dependent manner. However, the BMCF can be seen to have had no effect on Fas receptor. PINK1 expression did not change considerably and was inclined to decrease in control group, but the expression was down-regulated then subsequently increased with the use of the BMCF in a dose-dependent manner. Caspase 3 increased and subsequently decreased in a dose-dependent manner. These results suggest that the BMCF invigorates mitophagy and improves mitochondrial oxidative respiration in skeletal muscle, and in early stage of apoptosis induced by the BMCF is not related to extrinsic death-receptor mediated but mitochondria-mediated signaling pathway.

Evidence for a direct effect of the NAD+ precursor acipimox on muscle mitochondrial function in humans.

Recent preclinical studies showed the potential of nicotinamide adenine dinucleotide (NAD+) precursors to increase oxidative phosphorylation and improve metabolic health, but human data are lacking. We hypothesize that the nicotinic acid derivative acipimox, an NAD+ precursor, would directly affect mitochondrial function independent of reductions in nonesterified fatty acid (NEFA) concentrations. In a multicenter randomized crossover trial, 21 patients with type 2 diabetes (age 57.7 ± 1.1 years, BMI 33.4 ± 0.8 kg/m2) received either placebo or acipimox 250 mg three times daily dosage for 2 weeks. Acipimox treatment increased plasma NEFA levels (759 ± 44 vs. 1,135 ± 97 μmol/L for placebo vs. acipimox, P < 0.01) owing to a previously described rebound effect. As a result, skeletal muscle lipid content increased and insulin sensitivity decreased. Despite the elevated plasma NEFA levels, ex vivo mitochondrial respiration in skeletal muscle increased. Subsequently, we showed that acipimox treatment resulted in a robust elevation in expression of nuclear-encoded mitochondrial gene sets and a mitonuclear protein imbalance, which may indicate activation of the mitochondrial unfolded protein response. Further studies in C2C12 myotubes confirmed a direct effect of acipimox on NAD+ levels, mitonuclear protein imbalance, and mitochondrial oxidative capacity. To the best of our knowledge, this study is the first to demonstrate that NAD+ boosters can also directly affect skeletal muscle mitochondrial function in humans.

Hypoxic Preconditioning Alleviates Reoxygenation Injury via AKT in Respiratory Skeletal Muscle

Hypoxic Preconditioning Alleviates Reoxygenation Injury via AKT in Respiratory Skeletal Muscle

Diagnostic Ultrasound Imaging of Mouse Diaphragm Function

Function analysis of rodent respiratory skeletal muscles, particularly the diaphragm, is commonly performed by isolating muscle strips using invasive surgical procedures. Although this is an effective method of assessing in vitro diaphragm activity, it involves non-survival surgery. The application of non-invasive ultrasound imaging as an in vivo procedure is beneficial since it not only reduces the number of animals sacrificed, but is also suitable for monitoring disease progression in live mice. Thus, our ultrasound imaging method may likely assist in the development of novel therapies that alleviate muscle injury induced by various respiratory diseases. Particularly, in clinical diagnoses of obstructive lung diseases, ultrasound imaging has the potential to be used in conjunction with other standard tests to detect the early onset of diaphragm muscle fatigue. In the current protocol, we describe how to accurately evaluate diaphragm contractility in a mouse model using a diagnostic ultrasound imaging technique.

Microbiopsies versus Bergström needle for skeletal muscle sampling: impact on maximal mitochondrial respiration rate

Microbiopsies are increasingly used as an alternative to the standard Bergström technique for skeletal muscle sampling. The potential impact of these two different procedures on mitochondrial respiration rate is unknown. The objective of this work was to compare microbiopsies versus Bergström procedure on mitochondrial respiration in skeletal muscle. 52 vastus lateralis muscle samples were obtained from 13 anesthetized pigs, either with a Bergström [6 gauges (G)] needle or with microbiopsy needles (12, 14, 18G). Maximal mitochondrial respiration (V GM-ADP) was assessed using an oxygraphic method on permeabilized fibers. The weight of the muscle samples and V GM-ADP decreased with the increasing gauge of the needles. A positive nonlinear relationship was observed between the weight of the muscle sample and the level of maximal mitochondrial respiration (r = 0.99, p < 0.05) and between needle size and maximal mitochondrial respiration (r = 0.99, p < 0.05). Microbiopsies give lower muscle sample weight and maximal rate of mitochondrial respiration compared to the standard Bergström needle.Therefore, the higher the gauge (i.e. the smaller the size) of the microbiopsy needle, the lower is the maximal rate of respiration. Microbiopsies of skeletal muscle underestimate the maximal mitochondrial respiration rate, and this finding needs to be highlighted for adequate interpretation and comparison with literature data.

The Effect of Hypoxic Preconditioning on Intracellular Reactive Oxygen Species Formation in Hypoxic Skeletal Muscle

Exposure to acute hypoxia may stimulate an increase in intracellular reactive oxygen species (ROS) formation leading to subsequent muscular injuries. Hypoxic preconditioning (HPC), which involves cycles of brief exposure to low and normal oxygen, may alleviate this ROS induced damage. However, the mechanism of HPC has not been fully elucidated. In this study, we hypothesized that HPC can mitigate intramuscular ROS in mouse skeletal muscle during hypoxia. Mouse diaphragm was isolated and loaded with dihydrofluorescein (a fluorescent probe) in order to detect ROS production in real time via confocal microscopy during a hypoxic period. Diaphragm strips were either non-treated (n = 5), treated with HPC (5 cycles; n = 5), or subject to antioxidant treatment (ebselen; n = 4), followed by a 30 min hypoxic exposure (PO2 = 5 Torr). Data were analyzed using a multi-way ANOVA, and expressed as means ± SE. In the control, intracellular ROS levels increased after 20 min of hypoxia, and this increase continued until the end of the 30 min hypoxic exposure. However, there was no substantial ROS formation in muscles treated with either HPC or the antioxidant, ebselen. Our data suggest that HPC suppresses intracellular ROS generation in respiratory skeletal muscle subjected to a hypoxic condition. Thus, we speculate that HPC can be used as a potential drug-free treatment to prevent oxidative injuries in respiratory muscle during hypoxia. Supported by OU General Fund G110, and Research Excellence Fund of Biomedical Research, OSU-HRS Fund 013000.

Ceramide alters mitochondrial dynamics and reduces mitochondrial respiration in skeletal muscle

Fat gain is associated with ceramide accrual in tissues of high‐metabolic rate, such as liver, brain and skeletal muscle. We predict that ceramide accumulation in skeletal muscle affects mitochondrial morphology and physiology. We found that ceramide treatment on murine myoblasts decreased viable mitochondrial density and induced mitochondrial fission, possibly via dynamin‐related protein 1 (Drp1), which was increased with ceramide accrual. Inhibition of endogenous ceramide synthesis via myriocin and serine palmitoyltransferase 2 (SPT2) siRNA prevented fatty acid‐induced mitochondrial fission in myoblasts. Moreover, inhibition of ceramide biosynthesis prevented the reduction in mitochondrial O2 consumption in myoblasts treated with fatty acids. Altogether, these findings suggest a critical role for ceramide in altering mitochondrial dynamics and function. Ongoing work will determine the extent to which mitochondrial fission mediates the adverse alterations in mitochondrial function. This work was supported by a BYU Graduate Research Fellowship (MES) and a BYU Mentoring Environment Grant (BTB).

Altered skeletal muscle respiration in patients with COPD: mitochondrial density or dysfunction?

INTRODUCTIONWe sought to determine if the blunted mitochondrial respiration typically observed in the skeletal muscle of patients with chronic obstructive pulmonary disease (COPD) is solely related to changes in mitochondrial density or if qualitative changes in respiration occur.METHODSCombinations of substrates (malate, glutamate, succinate, and ADP) and inhibitors (oligomycin) were used to measure complex I (CI) and complex II (CII) driven state 3 respiration separately and in combination in permeabilized muscle fibers from the vastus lateralis of 9 healthy controls (67±2 years) and 11 COPD patients (FEV1= 56±6%; 63±3 years). State 4 respiration and citrate synthase (CS) activity were measured using these same fibers.RESULTSCompared to controls COPD patients had blunted CI driven state 3 respiration (37.2±4.1, 23.7±2.7 nM/min/mg, respectively) and blunted state 3 respiration with CI and CII (47.2±5.1, 33.7±3.8 nM/min/mg, respectively) (P&lt;0.05) while state 3 driven by CII alone was similar between groups. CS activity and state 4 respiration were not different between groups.CONCLUSIONSGiven that CS activity, an indicator of density, was similar between groups, the blunted respiratory capacity in these COPD patients appears to be related to an attenuation of CI driven respiration. These data indicate that COPD results in qualitative changes in mitochondrial respiration of skeletal muscle.