Effects of Combined Inorganic Nitrate and Nitrite Supplementation on Cardiorespiratory Fitness and Skeletal Muscle Oxidative Capacity in Type 2 Diabetes: A Pilot Randomized Controlled Trial.

Nitric oxide (NO) stimulates mitochondrial biogenesis in skeletal muscle. However, NO metabolism is disrupted in individuals with type 2 diabetes mellitus (T2DM) potentially contributing to their decreased cardiorespiratory fitness (i.e., VO2max) and skeletal muscle oxidative capacity. We used a randomized, double-blind, placebo-controlled, 8-week trial with beetroot juice containing nitrate (NO3−) and nitrite (NO2−) (250 mg and 20 mg/day) to test potential benefits on VO2max and skeletal muscle oxidative capacity in T2DM. T2DM (N = 36, Age = 59 ± 9 years; BMI = 31.9 ± 5.0 kg/m2) and age- and BMI-matched non-diabetic controls (N = 15, Age = 60 ± 9 years; BMI = 29.5 ± 4.6 kg/m2) were studied. Mitochondrial respiratory capacity was assessed in muscle biopsies from a subgroup of T2DM and controls (N = 19 and N = 10, respectively). At baseline, T2DM had higher plasma NO3− (100%; p < 0.001) and lower plasma NO2− levels (−46.8%; p < 0.0001) than controls. VO2max was lower in T2DM (−26.4%; p < 0.001), as was maximal carbohydrate- and fatty acid-supported oxygen consumption in permeabilized muscle fibers (−26.1% and −25.5%, respectively; p < 0.05). NO3−/NO2− supplementation increased VO2max (5.3%; p < 0.01). Further, circulating NO2−, but not NO3−, positively correlated with VO2max after supplementation (R2= 0.40; p < 0.05). Within the NO3−/NO2− group, 42% of subjects presented improvements in both carbohydrate- and fatty acid-supported oxygen consumption in skeletal muscle (vs. 0% in placebo; p < 0.05). VO2max improvements in these individuals tended to be larger than in the rest of the NO3−/NO2− group (1.21 ± 0.51 mL/(kg*min) vs. 0.31 ± 0.10 mL/(kg*min); p = 0.09). NO3−/NO2− supplementation increases VO2max in T2DM individuals and improvements in skeletal muscle oxidative capacity appear to occur in those with more pronounced increases in VO2max.

ULK2 is essential for degradation of ubiquitinated protein aggregates and homeostasis in skeletal muscle.

Basal protein turnover, which largely relies on the degradation of ubiquitinated substrates, is instrumental for maintenance of muscle mass and function. However, the regulation of ubiquitinated protein degradation in healthy, nonatrophying skeletal muscle is still evolving, and potential tissue-specific modulators remain unknown. Using an unbiased expression analysis of 34 putative autophagy genes across mouse tissues, we identified unc-51 like autophagy activating kinase (Ulk)2, a homolog of the yeast autophagy related protein 1, as particularly enriched in skeletal muscle. Subsequent experiments revealed accumulations of insoluble ubiquitinated protein aggregates associated with the adaptors sequestosome 1 (SQSTM1, also known as p62) and next to breast cancer type 1 susceptibility protein gene 1 protein (NBR1) in adult muscles with ULK2 deficiency. ULK2 deficiency also led to impaired muscle force and caused myofiber atrophy and degeneration. These features were not observed in muscles with deficiency of the ULK2 paralog, ULK1. Furthermore, short-term ULK2 deficiency did not impair autophagy initiation, autophagosome to lysosome fusion, or protease activities of the lysosome and proteasome. Altogether, our results indicate that skeletal muscle ULK2 has a unique role in basal selective protein degradation by stimulating the recognition and proteolytic sequestration of insoluble ubiquitinated protein aggregates associated with p62 and NBR1. These findings have potential implications for conditions of poor protein homeostasis in muscles as observed in several myopathies and aging.-Fuqua, J. D., Mere, C. P., Kronemberger, A., Blomme, J., Bae, D., Turner, K. D., Harris, M. P., Scudese, E., Edwards, M., Ebert, S. M., de Sousa, L. G. O., Bodine, S. C., Yang, L., Adams, C. M., Lira, V. A. ULK2 is essential for degradation of ubiquitinated protein aggregates and homeostasis in skeletal muscle.

Electrical pulse stimulation induces differential responses in insulin action in myotubes from severely obese individuals.

KEY POINTS: Exercise/exercise training can enhance insulin sensitivity through adaptations in skeletal muscle, the primary site of insulin-mediated glucose disposal; however, in humans the range of improvement can vary substantially. The purpose of this study was to determine if obesity influences the magnitude of the exercise response in relation to improving insulin sensitivity in human skeletal muscle. Electrical pulse stimulation (EPS; 24 h) of primary human skeletal muscle myotubes improved insulin action in tissue from both lean and severely obese individuals, but responses to EPS were blunted with obesity. EPS improved insulin signal transduction in myotubes from lean but not severely obese subjects and increased AMP accumulation and AMPK Thr172 phosphorylation, but to a lesser degree in myotubes from the severely obese. These data reveal that myotubes of severely obese individuals enhance insulin action and stimulate exercise-responsive molecules with contraction, but in a manner and magnitude that differs from lean subjects. ABSTRACT: Exercise/muscle contraction can enhance whole-body insulin sensitivity; however, in humans the range of improvements can vary substantially. In order, to determine if obesity influences the magnitude of the exercise response, this study compared the effects of electrical pulse stimulation (EPS)-induced contractile activity upon primary myotubes derived from lean and severely obese (BMI ≥ 40 kg/m2 ) women. Prior to muscle contraction, insulin action was compromised in myotubes from the severely obese as was evident from reduced insulin-stimulated glycogen synthesis, glucose oxidation, glucose uptake, insulin signal transduction (IRS1, Akt, TBC1D4), and insulin-stimulated GLUT4 translocation. EPS (24 h) increased AMP, IMP, AMPK Thr172 phosphorylation, PGC1α content, and insulin action in myotubes of both the lean and severely obese subjects. However, despite normalizing indices of insulin action to levels seen in the lean control (non-EPS) condition, responses to EPS were blunted with obesity. EPS improved insulin signal transduction in myotubes from lean but not severely obese subjects and EPS increased AMP accumulation and AMPK Thr172 phosphorylation, but to a lesser degree in myotubes from the severely obese. These data reveal that myotubes of severely obese individuals enhance insulin action and stimulate exercise-responsive molecules with contraction, but in a manner and magnitude that differs from lean subjects.