Chronic kidney disease (CKD) impacts more than 25 million Americans and is associated with higher risk of all‐cause and cardiovascular mortality. Impaired kidney function leads to retention of metabolic waste products, termed uremic toxins, that negatively impact skeletal muscle resulting in increased fatigue, weakness, and muscle atrophy. Previous evidence has implicated mitochondria within the skeletal muscle as a primary mediator of muscle dysfunction in CKD, yet the underlying mechanisms are unknown. Therefore, the purpose of this study was to define the impact of uremic toxins on mitochondrial energetics. Skeletal muscle mitochondria were isolated from healthy C57BL/6N mice and exposed to vehicle (DMSO) or varying doses of the following uremic toxins: indoxyl sulfate, indole‐3‐acetic‐acid, L‐kynurenine, kynurenic acid, and methylguanidine. We employed a comprehensive mitochondrial phenotyping platform that included assessments of mitochondrial OXPHOS conductance across several levels of energy demand, hydrogen peroxide production (JH2O2), and dehydrogenase flux (using NADH autofluorescence). Exposure to uremic toxins resulted in a dose‐dependent decrease in OXPHOS conductance for all toxins, with 100nM exposure resulting in an average decrease of ~22% supported by pyruvate/malate (all P<0.05, n= 5–6/group). Uremic toxins did not decrease pyruvate dehydrogenase activity even at millimolar concentrations (all P>0.64), suggesting the decreased OXPHOS conductance occurs downstream of matrix dehydrogenases. Consistent with decreased OXPHOS conductance, uremic toxins dose‐dependently increased JH2O2 by 2–5‐fold (all P<0.01, n=4/group). These findings provide direct evidence that uremic toxins negatively impact skeletal muscle mitochondrial energetics, resulting in decreased energy transfer. Impaired mitochondrial energetics appears to be mediated downstream of matrix dehydrogenases, through either direct interaction within the electron transport system or ATP synthase. Support or Funding Information Partially supported by AHA Grant 18CDA34110044 to TER This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The decline in stroke volume (SV) during exercise in the heat has been attributed to either an increase in cutaneous blood flow (CBF) that reduces venous return or an increase in heart rate (HR) that reduces cardiac filling time. However, the evidence supporting each mechanism arises under experimental conditions with different skin temperatures (T; e.g., ≥38°C vs. ≤36°C, respectively). We systematically studied cardiovascular responses to progressively increased T (32°C-39°C) with narrowing of the core-to-skin gradient during moderate intensity exercise. Eight men cycled at 63 ± 1% peak oxygen consumption for 20-30 min. T was manipulated by having subjects wear a water-perfused suit that covered most of the body and maintained T that was significantly different between trials and averaged 32.4 ± 0.2, 35.5 ± 0.1, 37.5 ± 0.1, and 39.5 ± 0.1°C, respectively. The graded heating of T ultimately produced a graded elevation of esophageal temperature (T) at the end of exercise. Incrementally increasing T resulted in a graded increase in HR and a graded decrease in SV. CBF reached a similar average plateau value in all trials when T was above ~38°C, independent of T. T had no apparent effect on forearm venous volume (FVV). In conclusion, the CBF and FVV responses suggest no further pooling of blood in the skin when T is increased from 32.4°C to 39.5°C. The decrease in SV during moderate intensity exercise when heating the skin to high levels appears related to an increase in HR and not an increase in CBF. NEW & NOTEWORTHY This study systematically investigated the effect of increasing skin temperature (T) to high levels on cardiovascular responses during moderate intensity exercise. We conclude that the declines in stroke volume were related to the increases in heart rate but not the changes in cutaneous blood flow (CBF) and forearm venous volume (FVV) during moderate intensity exercise when T increased from ~32°C to ~39°C. High T (≥38°C) did not further elevate CBF and FVV compared with lower T during moderate intensity exercise.

Besides the known beneficial effect on the LV, ERT improves RV mass and EDV.

Duchenne muscular dystrophy (DMD) is a severe, progressive, and ultimately fatal disease of skeletal muscle wasting, respiratory insufficiency, and cardiomyopathy. The identification of the dystrophin gene as central to DMD pathogenesis has led to the understanding of the muscle membrane and the proteins involved in membrane stability as the focal point of the disease. The lessons learned from decades of research in human genetics, biochemistry, and physiology have culminated in establishing the myriad functionalities of dystrophin in striated muscle biology. Here, we review the pathophysiological basis of DMD and discuss recent progress toward the development of therapeutic strategies for DMD that are currently close to or are in human clinical trials. The first section of the review focuses on DMD and the mechanisms contributing to membrane instability, inflammation, and fibrosis. The second section discusses therapeutic strategies currently used to treat DMD. This includes a focus on outlining the strengths and limitations of approaches directed at correcting the genetic defect through dystrophin gene replacement, modification, repair, and/or a range of dystrophin-independent approaches. The final section highlights the different therapeutic strategies for DMD currently in clinical trials.

Chronic diseases such as cancer, COPD, and heart failure impair diaphragm metabolism and mitochondrial function. Mice are used extensively to replicate disease conditions and offer the advantage of studying genetically modified animals. Mouse diaphragm contains a high abundance of connective tissue, with relatively thin (15–25 μm diameter) and fragile fibers. In preliminary studies, we discovered that standard fiber separation approaches utilized for limb muscles are unsuitable for the diaphragm. Thus, the purpose of this study was to optimize a protocol for diaphragm (Dia) fiber bundle preparation to evaluate the mitochondrial respiration and reactive oxygen species emission. We used the red gastrocnemius (Gast) muscle as a ‘standard’ for comparison. Michaelis‐Menten kinetics of ADP‐stimulated O2 consumption with complex I substrates (JO2 in pmol/s/mg wet weight) showed 3‐fold higher Vmax in Dia than Gast (Gast 69 ± 18, Dia 211 ± 20; p < 0.05, n = 9 mice), whereas Km was not significantly different. Maximal JO2 in Dia was 3‐fold higher with complex I + II substrates (Gast 148 ± 25, Dia 387 ± 45; p < 0.05), and 2‐fold higher with palmitoyl‐CoA + carnitine (Gast 7 ± 0, Dia 15 ± 2, p < 0.05) compared to Gast. Baseline JH2O2 (pmol/min/mg wet weight) was higher in Dia (Gast 0.8 ± 0.2, Dia 1.7 ± 0.3; p < 0.05), but succinate‐induced JH2O2 was not different between muscles. Citrate synthase activity was 3‐fold higher in Dia (μmol/min/mg protein: Gast 43 ± 3, Dia 118 ± 4; p < 0.05). The protein abundance of electron transport chain complexes I‐V were 2–4 fold higher in the Dia than Gast (p < 0.05). When we normalized maximal JO2 by citrate synthase activity, there were no differences between Dia and Gast for complex I and complex I + II substrates. JH2O2 normalized to citrate synthase activity was higher for Gast at baseline (pmol/min/mg wet weight/U CS: Gast 2.1 ± 0.3, Dia 0.3 ± 0.1; p < 0.05), but there was no difference in succinate‐induced JH2O2 normalized to citrate synthase activity. The values for maximal diaphragm mitochondria JO2 in our study are 2–3 fold higher than reported in the literature using the standard ‘limb muscle’ approach for the diaphragm fiber preparation. The similar maximal JO2 normalized to citrate synthase activity between diaphragm and red gastrocnemius muscles suggest that our new approach is valid for the assessment of intact diaphragm mitochondrial function in permeabilized fiber bundles. Support or Funding Information Funding source: NIH 1R01HL130318‐01 This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

A group of 139 patients with symptoms of bladder outflow obstruction due to benign prostatic hypertrophy were entered into a double-blind, parallel group, multicentre study of 2 doses of indoramin versus placebo. There were 18 withdrawals or exclusions, leaving 121 patients for analysis. After 8 weeks, mean peak flow rates increased more in patients treated with indoramin 20 mg bd than in those with placebo. The difference between indoramin 20 mg nocte and placebo was not significant. The change in mean peak flow rate for the higher dose of indoramin represented an increase of 50%. Both patients and investigators reported that the patients' symptoms had improved significantly on both indoramin 20 mg bd and 20 mg nocte; 9 patients were withdrawn because of adverse events, 5 taking placebo and 2 in each of the treatment groups. It was concluded that indoramin 20 mg bd was both effective and well tolerated in the management of symptomatic benign prostatic hypertrophy.

Background/Aims: Diaphragm dysfunction with increased reactive oxygen species (ROS) occurs within 72 hrs post-myocardial infarction (MI) in mice and may contribute to loss of inspiratory maximal pressure and endurance in patients. Methods: We used wild-type (WT) and whole-body Nox4 knockout (Nox4KO) mice to measure diaphragm bundle force in vitro with a force transducer, mitochondrial respiration in isolated fiber bundles with an O2 sensor, mitochondrial ROS by fluorescence, mRNA (RT-PCR) and protein (immunoblot), and fiber size by histology 72 hrs post-MI. Results: MI decreased diaphragm fiber cross-sectional area (CSA) (~15%, p = 0.015) and maximal specific force (10%, p = 0.005), and increased actin carbonylation (5-10%, p = 0.007) in both WT and Nox4KO. Interestingly, MI did not affect diaphragm mRNA abundance of MAFbx/atrogin-1 and MuRF-1 but Nox4KO decreased it by 20-50% (p < 0.01). Regarding the mitochondria, MI and Nox4KO decreased the protein abundance of citrate synthase and subunits of electron transport system (ETS) complexes and increased mitochondrial O2 flux (JO2) and H2O2 emission (JH2O2) normalized to citrate synthase. Mitochondrial electron leak (JH2O2/JO2) in the presence of ADP was lower in Nox4KO and not changed by MI. Conclusion: Our study shows that the early phase post-MI causes diaphragm atrophy, contractile dysfunction, sarcomeric actin oxidation, and decreases citrate synthase and subunits of mitochondrial ETS complexes. These factors are potential causes of loss of inspiratory muscle strength and endurance in patients, which likely contribute to the pathophysiology in the early phase post-MI. Whole-body Nox4KO did not prevent the diaphragm abnormalities induced 72 hrs post-MI, suggesting that systemic pharmacological inhibition of Nox4 will not benefit patients in the early phase post-MI.

Here we report greater in vitro respiratory muscle contractile function in old mice receiving supplemental NaNO 3 for 14 days compared with age-matched controls. r Myofibrillar protein phosphorylation, which enhances contractile function, did not change in our study-a finding inconsistent with the hypothesis that this post-translational modification is a mechanism for dietary nitrate to improve muscle contractile function. r Nitrate supplementation did not change the abundance of calcium-handling proteins in the diaphragm of old mice, in contrast with findings from the limb muscles of young mice in previous studies. r Our findings suggest that nitrate supplementation enhances myofibrillar protein function without affecting the phosphorylation status of key myofibrillar proteins.