A single bout of resistance exercise triggers mitophagy, potentially involving the ejection of mitochondria in human skeletal muscle.

AIM: The present study aimed to investigate the effects of a single bout of resistance exercise on mitophagy in human skeletal muscle (SkM). METHODS: Eight healthy men were recruited to complete an acute bout of one-leg resistance exercise. SkM biopsies were obtained one hour after exercise in the resting leg (Rest-leg) and the contracting leg (Ex-leg). Mitophagy was assessed using protein-related abundance, transmission electron microscopy (TEM), and fluorescence microscopy. RESULTS: Our results show that acute resistance exercise increased pro-fission protein phosphorylation (DRP1Ser616) and decreased mitophagy markers such as PARKIN and BNIP3L/NIX protein abundance in the Ex-leg. Additionally, mitochondrial complex IV decreased in the Ex-leg when compared to the Rest-leg. In the Ex-leg, TEM and immunofluorescence images showed mitochondrial cristae abnormalities, a mitochondrial fission phenotype, and increased mitophagosome-like structures in both subsarcolemmal and intermyofibrillar mitochondria. We also observed increased mitophagosome-like structures on the subsarcolemmal cleft and mitochondria in the extracellular space of SkM in the Ex-leg. We stimulated human primary myotubes with CCCP, which mimics mitophagy induction in the Ex-leg, and found that BNIP3L/NIX protein abundance decreased independently of lysosomal degradation. Finally, in another human cohort, we found a negative association between BNIP3L/NIX protein abundance with both mitophagosome-like structures and mitochondrial cristae density in the SkM. CONCLUSION: The findings suggest that a single bout of resistance exercise can initiate mitophagy, potentially involving mitochondrial ejection, in human skeletal muscle. BNIP3L/NIX is proposed as a sensitive marker for assessing mitophagy flux in SkM.

Effect of epicatechin consumption on the inflammatory pathway and mitochondria morphology in PBMC from a R350P desminopathy patient: A case report.

Desminopathy R350P is a human myopathy that is characterized by the progressive loss of muscle fiber organization. This results in the loss of muscle size, mobility, and strength. In desminopathy, inflammation affects muscle homeostasis and repair, and contributes to progressive muscle deterioration. Mitochondria morphology was also suggested to affect desminopathy progression. Epicatechin (Epi)-a natural compound found in cacao-has been proposed to regulate inflammatory signaling and mitochondria morphology in human and animal models. Hence, we hypothesize chronic Epi consumption to improve inflammatory pathway and mitochondria morphology in the peripheral blood mononuclear cells (PBMCs) of a desminopathy R350P patient. We found that 12 weeks of Epi consumption partially restored TRL4 signaling, indicative of inflammatory signaling and mitochondria morphology in the desminopathy patient. Moreover, Epi consumption improved blood health parameters, including reduced HOMA-IR and IL-6 levels in the desminopathy patient. This indicates that Epi consumption could be a useful tool to slow disease progression in desminopathy patients.

Effects of an antioxidants cocktail on glucose metabolism at rest, during exercise, and during a glucose load in healthy young subjects

Background: Reactive oxygen species (ROS) regulate glucose metabolism (GM) in skeletal muscle by improving the translocation of GLUT4. Antioxidant supplementation could block this physiological effect, altering glucose signaling during exercise. However, there is limited evidence in humans on whether antioxidant intake affects GM. Therefore, we aimed to determine the effect of an antioxidant cocktail (AOC) on GM at rest and during metabolic challenges. Methods: Ten healthy male subjects received AOC supplementation (1000 mg of Vitamin C, 600 IU of Vitamin E, and 600 mg of α-lipoic acid) or placebo (2.000 mg of talc) before two trials conducted 7 days apart. Trial 1: AOC 120 and 90 minutes before an endurance exercise (EEX) bout at 60 % of maximal oxygen uptake (VO2max); Trial 2: AOC 120 and 90 minutes before an oral glucose tolerance test (OGTT; 75 g glucose). Measurements of gas exchange and capillary blood samples were collected every 15 minutes during both trials. Results: AOC supplementation increased resting glucose levels (p<0.05). During Trial 1 (EEX), the AOC increased carbohydrate oxidation (CHOox) (p= 0.03), without effect in glucose blood levels. During Trial 2 (OGTT), the AOC supplementation had no significant effect on GM parameters. Conclusion: Acute supplementation with AOC increased resting glucose levels and CHOox during EEX in healthy subjects, with no effect on GM during the OGTT.

Antecedentes: Las especies reactivas de oxígeno (ROS) regulan el metabolismo de la glucosa (GM) en el músculo esquelético al mejorar la translocación de GLUT4. La suplementación con antioxidantes podría bloquear este efecto fisiológico, alterando la señalización de la glucosa durante el ejercicio. Sin embargo, existe evidencia limitada en humanos sobre si la ingesta de antioxidantes afecta el GM. Por lo tanto, nuestro objetivo fue determinar el efecto de un cóctel de antioxidantes (AOC) en el GM en reposo y durante desafíos metabólicos. Métodos: Sujetos sanos (sexo masculino; n= 10) recibieron suplementos de AOC (1.000 mg de vitamina C, 600 UI de vitamina E y 600 mg de ácido α-lipoico) o placebo (2.000 mg de talco) previo a dos pruebas realizadas con 7 días de diferencia. Prueba 1: AOC 120 y 90 minutos antes de una serie de ejercicio de resistencia (EEX) al 60% del consumo máximo de oxígeno (VO2max); prueba 2: AOC 120 y 90 minutos antes de una prueba de tolerancia oral a la glucosa (OGTT; 75 g de glucosa). Se obtuvieron datos de intercambio de gaseoso y muestras de sangre capilar cada 15 minutos durante ambas pruebas. Resultados: la suplementación con AOC aumentó los niveles de glucosa en reposo (p<0,05). Durante la prueba 1 (EEX), el AOC aumentó la oxidación de carbohidratos (CHOox) (p= 0,03), sin efecto en los niveles de glucosa en sangre. Durante la prueba 2 (OGTT), la suplementación con AOC no tuvo un efecto significativo en los parámetros de GM. Conclusión: Una suplementación aguda con AOC aumentó los niveles de glucosa en reposo y la CHOox durante EEX en sujetos sanos, sin efecto sobre el GM durante la OGTT.

Regulation of mitochondrial morphology and cristae architecture by the TLR4 pathway in human skeletal muscle.

In skeletal muscle (SkM), a reduced mitochondrial elongate phenotype is associated with several metabolic disorders like type 2 diabetes mellitus (T2DM). However, the mechanisms contributing to this reduction in mitochondrial elongate phenotype in SkM have not been fully elucidated. It has recently been shown in a SkM cell line that toll-like receptor 4 (TLR4) contributes to the regulation of mitochondrial morphology. However, this has not been investigated in human SkM. Here we found that in human SkM biopsies, TLR4 protein correlated negatively with Opa1 (pro-mitochondrial fusion protein). Moreover, the incubation of human myotubes with LPS reduced mitochondrial size and elongation and induced abnormal mitochondrial cristae, which was prevented with the co-incubation of LPS with TAK242. Finally, T2DM myotubes were found to have reduced mitochondrial elongation and mitochondrial cristae density. Mitochondrial morphology, membrane structure, and insulin-stimulated glucose uptake were restored to healthy levels in T2DM myotubes treated with TAK242. In conclusion, mitochondrial morphology and mitochondrial cristae seem to be regulated by the TLR4 pathway in human SkM. Those mitochondrial alterations might potentially contribute to insulin resistance in the SkM of patients with T2DM.

Plant-Based Nutrition: Exploring Health Benefits for Atherosclerosis, Chronic Diseases, and Metabolic Syndrome-A Comprehensive Review.

Atherosclerosis, chronic non-communicable diseases, and metabolic syndrome are highly interconnected and collectively contribute to global health concerns that reduce life expectancy and quality of life. These conditions arise from multiple risk factors, including inflammation, insulin resistance, impaired blood lipid profile, endothelial dysfunction, and increased cardiovascular risk. Adopting a plant-based diet has gained popularity as a viable alternative to promote health and mitigate the incidence of, and risk factors associated with, these three health conditions. Understanding the potential benefits of a plant-based diet for human health is crucial, particularly in the face of the rising prevalence of chronic diseases like diabetes, hypertension, dyslipidemia, atherosclerosis, and cancer. Thus, this review focused on the plausible advantages of consuming a type of food pattern for the prevention and/or treatment of chronic diseases, emphasizing the dietary aspects that contribute to these conditions and the evidence supporting the benefits of a plant-based diet for human health. To facilitate a more in-depth analysis, we present separate evidence for each of these three concepts, acknowledging their intrinsic connection while providing a specific focus on each one. This review underscores the potential of a plant-based diet to target the underlying causes of these chronic diseases and enhance health outcomes for individuals and populations.

Mitochondria-SR interaction and mitochondrial fusion/fission in the regulation of skeletal muscle metabolism.

Mitochondria-endoplasmic/sarcoplasmic reticulum (ER/SR) interaction and mitochondrial fusion/fission are critical processes that influence substrate oxidation. This narrative review summarizes the evidence on the effects of substrate availability on mitochondrial-SR interaction and mitochondria fusion/fission dynamics to modulate substrate oxidation in human skeletal muscle. Evidence shows that an increase in mitochondria-SR interaction and mitochondrial fusion are associated with elevated fatty acid oxidation. In contrast, a decrease in mitochondria-SR interaction and an increase in mitochondrial fission are associated with an elevated glycolytic activity. Based on the evidence reviewed, we postulate two hypotheses for the link between mitochondrial dynamics and insulin resistance in human skeletal muscle. First, glucose and fatty acid availability modifies mitochondria-SR interaction and mitochondrial fusion/fission to help the cell to adapt substrate oxidation appropriately. Individuals with an impaired response to these substrate challenges will accumulate lipid species and develop insulin resistance in skeletal muscle. Second, a chronically elevated substrate availability (e.g. overfeeding) increases mitochondrial production of reactive oxygen species and induced mitochondrial fission. This decreases fatty acid oxidation, thus leading to the accumulation of lipid species and insulin resistance in skeletal muscle. Altogether, we propose mitochondrial dynamics as a potential target for disturbances associated with low fatty acid oxidation.

Time reallocation of physical behaviours induced by endurance exercise in physically active individuals.

Increasing moderate-vigorous physical activity (MVPA) through exercise requires reallocating time from other physical behaviour(s). We aimed to determine the reallocations induced by endurance exercise in physically active individuals. We also searched for behavioural compensatory responses, and explored the effect of exercise on daily energy expenditure. Fourteen participants (8 women; median age 37.8 [IQR 29.9-48.5] yr) exercised on Monday, Wednesday, and Friday mornings (cycling MVPA, 65 min/session; "exercise days"), and avoided exercising on Tuesday and Thursday ("rest days"). Time spent on sleep, sedentary behaviour, light-intensity physical activity, and MVPA was determined each day by accelerometers and logs. An energy expenditure index was computed considering minutes spent on each behaviour and fixed metabolic equivalents. We found that all participants had lower sleep and higher total (including exercise) MVPA on exercise days compared to rest days. Thus, on exercise vs. rest days, sleep was lower (490 [453-553] vs. 553 [497-599] min/day, respectively, P < 0.001), and total MVPA was higher (86 [80-101] vs. 23 [15-45] min/day, respectively; P < 0.001). No differences in other physical behaviours were detected. Notably, exercise not only induced reallocations (i.e. less time in other behaviours) but also behavioural compensatory responses in some participants (e.g. increased sedentary behaviour). This rearrangement of physical behaviours manifested in exercise-induced increases in energy expenditure from 96 to 232 MET × min/day. In conclusion, active individuals reallocated time from sleep to accommodate morning exercise. Yet exercise induced variable rearrangements of behaviours, with some individuals manifesting compensatory responses. Understanding individual rearrangements may help improve exercise interventions.

Adults are recommended to engage in moderate-vigorous physical activity (MVPA) to maintain health. But including exercise sessions within a day inevitably requires reallocating time from other physical behaviour(s): sleep, sedentary behaviour, or physical activity.We studied the time reallocations induced by 65 min/day of morning exercise (cycling MVPA) in physically active participants.Participants spent less time sleeping and higher time on total (including exercise) MVPA on days that included exercise compared to days without exercise. Thus, participants reallocated sleep time to accommodate morning exercise sessions.Some participants also spent higher time on sedentary behaviour during days that included exercise compared to days without exercise. This probably represents a behavioural compensatory response to exercise-induced fatigue.Together, time reallocations and behavioural compensatory responses led to a rearrangement of daily time spent on physical behaviours. This rearrangement was estimated to produce large interindividual variability in the increase in energy expenditure induced by exercise.

Neither Chia Flour nor Whey Protein Supplementation Further Improves Body Composition or Strength Gains after a Resistance Training Program in Young Subjects with a Habitual High Daily Protein Intake.

The aim of this study was to compare the potential additional effect of chia flour, whey protein, and a placebo juice to resistance training on fat-free mass (FFM) and strength gains in untrained young men. Eighteen healthy, untrained young men underwent an 8-week whole-body resistance training program, comprising three sessions per week. Subjects were randomized into three groups that after each training session consumed: (1) 30 g whey protein concentrate containing 23 g protein (WG), (2) 50 g chia flour containing 20 g protein (CG), or (3) a placebo not containing protein (PG). Strength tests (lower- and upper-limb one repetition maximum (1 RM) tests) and body composition analyses (dual-energy X-ray absorptiometry; DXA) were performed before (PRE) and after (POST) the intervention. Resistance training increased FFM and the 1 RM for each of the strength tests similarly in the three groups. FFM increased by 2.3% in WG (p = 0.04), by 3.6% in CG (p = 0.004), and by 3.0% in PG (p = 0.002)., and 1 RM increased in the different strength tests in the three groups (p < 0.05) with no difference between PG, CG, and WG. In conclusion, neither chia flour nor whey protein supplementation elicited an enhanced effect on FFM and strength gains after an 8-week resistance training program in healthy, untrained young men consuming a habitual high protein mixed diet (>1.2 g/kg/day).

Plasma α-Actin as an Early Marker of Muscle Damage After Repeated Bouts of Eccentric Cycling.

Purpose: This study aimed to examine the changes in skeletal muscle (SM) α-actin, myoglobin (Mb) and hydroxyproline (HP) in plasma and other indirect markers of muscle damage after repeated bouts of eccentric cycling. Methods: Ten healthy men (23.3 ± 2.8 years) performed two 30-min eccentric cycling bouts at 100% of maximal concentric power output (230.7 ± 36.9 W) separated by 2 weeks (ECC1 and ECC2). Maximal voluntary isometric contraction (MVIC) peak force of the knee extensor muscles, muscle soreness (SOR), pain pressure threshold (PPT) and plasma levels of SM α-actin, Mb, and HP were measured before, 0.5, 3, 24-168 h after each cycling bout. Results: MVIC peak force decreased on average 10.7 ± 13.1% more after ECC1 than ECC2. SOR was 80% greater and PPT was 12-14% lower after ECC1 than ECC2. Plasma SM α-actin levels increased at 0.5, 3, and 24-72 h after ECC1 (26.1-47.9%), and SM α-actin levels at 24 h after ECC1 were associated with muscle strength loss (r = -0.56, P = .04) and SOR (r = 0.88, P = .001). Mb levels increased at 0.5, 3, and 24 h after ECC1 (200-502%). However, Mb levels at 24 h after ECC1were not associated with muscle strength loss and SOR. HP levels remained unchanged after ECC1. ECC2 did not increase SM α-actin, Mb and HP levels. Conclusion: Our results indicate that α-actin could be used as a potential marker for the early identification of SM damage due to its early appearance in plasma and its association with other indirect markers of muscle damage.

Severe COVID-19 correlates with lower mitochondrial cristae density in PBMCs and greater sitting time in humans.

An interaction between mitochondrial dynamics, physical activity levels, and COVID-19 severity has been previously hypothesized. However, this has not been tested. We aimed to compare mitochondrial morphology and cristae density of PBMCs between subjects with non-severe COVID-19, subjects with severe COVID-19, and healthy controls. Additionally, we compared the level of moderate-vigorous physical activity (MVPA) and sitting time between groups. Blood samples were taken to obtain PBMCs. Mitochondrial dynamics were assessed by electron microscopy images and western blot of protein that regulate mitochondrial dynamics. The International Physical Activity Questionnaire (IPAQ; short version) was used to estimate the level of MVPA and the sitting time The patients who develop severe COVID-19 (COVID-19++) not present alterations of mitochondrial size neither mitochondrial density in comparison to non-severe patients COVID-19 (COVID-19) and control subjects (CTRL). However, compared to CTRL, COVID-19 and COVID-19++ groups have lower mitochondrial cristae length, a higher proportion of abnormal mitochondrial cristae. The COVID-19++ group has lower number (trend) and length of mitochondrial cristae in comparison to COVID-19 group. COVID-19, but not COVID-19++ group had lower Opa 1, Mfn 2 and SDHB (Complex II) proteins than CTRL group. Besides, COVID-19++ group has a higher time sitting. Our results show that low mitochondrial cristae density, potentially due to physical inactivity, is associated with COVID-19 severity.

Effect of Epicatechin on Skeletal Muscle.

Loss of skeletal muscle (SkM) quality is associated with different clinical conditions such as aging, diabetes, obesity, cancer, and heart failure. Nutritional research has focused on identifying naturally occurring molecules that mitigate the loss of SkM quality induced by pathology or syndrome. In this context, although few human studies have been conducted, epicatechin (Epi) is a prime candidate that may positively affect SkM quality by its potential ability to mitigate muscle mass loss. This seems to be a consequence of its antioxidant and anti-inflammatory properties and its stimulation of mitochondrial biogenesis to increase myogenic differentiation, as well as its modulation of key proteins involved in SkM structure, function, metabolism, and growth. In conclusion, the Epi could prevent, mitigate, delay, and even treat muscle-related disorders caused by aging and diseases. However, studies in humans are needed.

Protection against indomethacin-induced loss of intestinal epithelial barrier function by a quercetin oxidation metabolite present in onion peel: In vitro and in vivo studies.

Oxidative stress is directly implicated in the loss of intestinal epithelial barrier function (IEBF) induced by non-steroidal anti-inflammatory drugs (NSAIDs). Previous studies by our research team demonstrated that 2-(3,4-dihydroxybenzoyl)-2,4,6-trihydroxy-3(2H)-benzofuranone (BZF), a quercetin oxidation metabolite that naturally occurs in onion peels, exhibits an antioxidant potency notably higher than quercetin. Thus, we assessed the potential of BZF and a BZF-rich onion peel aqueous extract (OAE) to protect against the loss of IEBF in Caco-2 cell monolayers and in rats exposed to indomethacin. In vitro, pure BZF and OAE standardized in BZF (100 nM), protected against the drop in transepithelial electrical resistance by 70 - 73%. Likewise, it prevented the increase in fluorescein-isothiocyanate labelled dextran (FITC-dextran) paracellular transport by 74% and oxidative stress by 84 - 86%. In vivo, BZF, given orally at a dose 80 µg/Kg bw as OAE, totally abolished a 30-fold increase in FITC-dextran serum concentration induced by indomethacin. This effect was dose-dependent and largely conserved (85%) when OAE was given 180-min prior to indomethacin. The IEBF-protective effect of OAE was accompanied by a full prevention of the NF-ĸB activation, and the increases in interleukine-8 secretion and myeloperoxidase activity induced by indomethacin. The protection was also associated with a 21-fold increase in Nrf2, and a 7-fold and 9-fold increase in heme oxygenase-1 and NAD(P)H-quinone oxidoreductase 1, respectively. The IEBF-protecting effect of OAE involves, most likely, its dual capacity to activate Nrf2 while inhibiting NF-ĸB activation. The extremely low doses of BZF needed to promote such actions warrants extending its IEBF-protective effects to other NSAIDs.

The fasting-feeding metabolic transition regulates mitochondrial dynamics.

In humans, insulin resistance has been linked to an impaired metabolic transition from fasting to feeding (metabolic flexibility; MetFlex). Previous studies suggest that mitochondrial dynamics response is a putative determinant of MetFlex; however, this has not been studied in humans. Thus, the aim of this study was to investigate the mitochondrial dynamics response in the metabolic transition from fasting to feeding in human peripheral blood mononuclear cells (PBMCs). Six male subjects fasted for 16 h (fasting), immediately after which they consumed a 75-g oral glucose load (glucose). In both fasting and glucose conditions, blood samples were taken to obtain PBMCs. Mitochondrial dynamics were assessed by electron microscopy images. We exposed in vitro acetoacetate-treated PBMCs to the specific IP3R inhibitor Xestospongin B (XeB) to reduce IP3R-mediated mitochondrial Ca2+ accumulation. This allowed us to evaluate the role of ER-mitochondria Ca2+ exchange in the mitochondrial dynamic response to substrate availability. To determine whether PBMCs could be used in obesity context (low MetFlex), we measured mitochondrial dynamics in mouse spleen-derived lymphocytes from WT and ob/ob mice. We demonstrated that the transition from fasting to feeding reduces mitochondria-ER interactions, induces mitochondrial fission and reduces mitochondrial cristae density in human PBMCs. In addition, we demonstrated that IP3R activity is key in the mitochondrial dynamics response when PBMCs are treated with a fasting-substrate in vitro. In murine mononuclear-cells, we confirmed that mitochondria-ER interactions are regulated in the fasted-fed transition and we further highlight mitochondria-ER miscommunication in PBMCs of diabetic mice. In conclusion, our results demonstrate that the fasting/feeding transition reduces mitochondria-ER interactions, induces mitochondrial fission and reduces mitochondrial cristae density in human PBMCs, and that IP3R activity may potentially play a central role.

Low abundance of Mfn2 protein correlates with reduced mitochondria-SR juxtaposition and mitochondrial cristae density in human men skeletal muscle: Examining organelle measurements from TEM images.

The role of mitofusin 2 (Mfn2) in the regulation of skeletal muscle (SM) mitochondria-sarcoplasmic (SR) juxtaposition, mitochondrial morphology, mitochondrial cristae density (MCD), and SM quality has not been studied in humans. In in vitro studies, whether Mfn2 increases or decreases mitochondria-SR juxtaposition remains controversial. Transmission electron microscopy (TEM) images are commonly used to measure the organelle juxtaposition, but the measurements are performed "by-hand," thus potentially leading to between-rater differences. The purposes of this study were to: (1) examine the repeatability and reproducibility of mitochondrial-SR juxtaposition measurement from TEM images of human SM between three raters with different experience and (2) compare the mitochondrial-SR juxtaposition, mitochondrial morphology, MCD (stereological-method), and SM quality (cross-sectional area [CSA] and the maximum voluntary contraction [MVC]) between subjects with high abundance (Mfn2-HA; n = 6) and low abundance (Mfn2-LA; n = 6) of Mfn2 protein. The mitochondria-SR juxtaposition had moderate repeatability and reproducibility, with the most experienced raters showing the best values. There were no differences between Mfn2-HA and Mfn2-LA groups in mitochondrial size, distance from mitochondria to SR, CSA, or MVC. Nevertheless, the Mfn2-LA group showed lower mitochondria-SR interaction, MCD, and VO2max . In conclusion, mitochondrial-SR juxtaposition measurement depends on the experience of the rater, and Mfn2 protein seems to play a role in the metabolic control of human men SM, by regulating the mitochondria-SR interaction.

Acute and Chronic Effects of High Frequency Electric Pulse Stimulation on the Akt/mTOR Pathway in Human Primary Myotubes.

Electrical pulse stimulation (EPS) has been suggested to be a useful method to investigate the mechanisms underlying the adaptations of human skeletal muscle to both endurance and resistance exercise. Although different myotube stimulation protocols mimicking acute and chronic endurance exercise have been developed, no convincing protocol mimicking resistance exercise exists. Adaptations to resistance exercise mainly ensue via the Akt/mTOR pathway. Therefore, the aim of this study was to develop a high frequency EPS protocol mimicking resistance exercise both acutely (100 Hz, 15 V, 0.4 ms with 4 s rest between each contraction for 30 min) and chronically (acute EPS protocol repeated on three consecutive days) on human myotubes. Compared to control conditions, the acute EPS protocol increased the phosphorylation of AktSer473 at 0 h (+91%, p = 0.02) and 3 h (+95%, p = 0.01), and mTORSer2448 at 0 h (+93%, p = 0.03), 1 h (+129%, p = 0.01), and 3 h (+104%, p = 0.0250) post-stimulation. The phosphorylation of ERK1/2Thr202/Tyr204 was increased at 0 h (+69%, p = 0.02) and 3 h (+117%, p = 0.003) post-stimulation compared to control conditions. In addition, both S6K1Thr389 (+157%, p = 0.009) and S6Ser240/244 (+153%, p = 0.003) phosphorylation increased 1 h after EPS compared to control conditions. Chronic EPS protocol increased the phosphorylation of S6K1Thr389 1 h (+105%, p = 0.03) and 3 h (+126%, p = 0.02) and the phosphorylation of S6Ser240/244 1 h (+32%, p = 0.02) after the end of the last stimulation. In conclusion, the present work shows that human muscle cells subjected to EPS can be used as an in vitro model of acute and chronic resistance exercise.

Generation of desminopathy in rats using CRISPR-Cas9.

BACKGROUND: Desminopathy is a clinically heterogeneous muscle disease caused by over 60 different mutations in desmin. The most common mutation with a clinical phenotype in humans is an exchange of arginine to proline at position 350 of desmin leading to p.R350P. We created the first CRISPR-Cas9 engineered rat model for a muscle disease by mirroring the R350P mutation in humans. METHODS: Using CRISPR-Cas9 technology, Des c.1045-1046 (AGG > CCG) was introduced into exon 6 of the rat genome causing p.R349P. The genotype of each animal was confirmed via quantitative PCR. Six male rats with a mutation in desmin (n = 6) between the age of 120-150 days and an equal number of wild type littermates (n = 6) were used for experiments. Maximal plantar flexion force was measured in vivo and combined with the collection of muscle weights, immunoblotting, and histological analysis. In addition to the baseline phenotyping, we performed a synergist ablation study in the same animals. RESULTS: We found a difference in the number of central nuclei between desmin mutants (1 ± 0.4%) and wild type littermates (0.2 ± 0.1%; P < 0.05). While muscle weights did not differ, we found the levels of many structural proteins to be altered in mutant animals. Dystrophin and syntrophin were increased 54% and 45% in desmin mutants, respectively (P < 0.05). Dysferlin and Annexin A2, proteins associated with membrane repair, were increased two-fold and 32%, respectively, in mutants (P < 0.05). Synergist ablation caused similar increases in muscle weight between mutant and wild type animals, but changes in fibre diameter revealed that fibre hypertrophy in desmin mutants was hampered compared with wild type animals (P < 0.05). CONCLUSIONS: We created a novel animal model for desminopathy that will be a useful tool in furthering our understanding of the disease. While mutant animals at an age corresponding to a preclinical age in humans show no macroscopic differences, microscopic and molecular changes are already present. Future studies should aim to further decipher those biological changes that precede the clinical progression of disease and test therapeutic approaches to delay disease progression.

Effects of A High Intensity Interval Session on Mucosal Immune Function and Salivary Hormones in Male and Female Endurance Athletes.

Although the effects of high intensity interval training (HIIT) on health and sports performance are well documented, the effects of this training type on mucosal immune function remain unclear. The aim of this study was to assess the impact of an acute HIIT session on salivary immune and endocrine marker levels (immunoglobulin A (sIgA), alpha amylase (sAA), cortisol (C), and testosterone (T)) in male and female endurance athletes. Twenty subjects (ten males and ten females) underwent ten bouts of treadmill running using a 4 min:2 min work:rest ratio at ~90% of peak oxygen uptake (VO2peak). Saliva samples were collected 5 min before and 20 min post-exercise. During work intervals, female participants had a higher HR than male participants (+4.0 ± 5%; p = 0.008). Rating of perceived exertion (RPE) increased throughout the duration of the HIIT session in both males and females (main time effect: p < 0.001), but was higher in males than females (+17 ± 4%; time x gender main effect: p < 0.001). Lactate concentrations were similar in both males and females. Exercise increased the concentration of salivary IgA (males: +24 ± 6%, p = 0.004; females: +27 ± 3%, p = 0.03), salivary alpha-amylase (males: +44 ± 22%, p = 0.036; females: +71 ± 26%, p = 0.026) and salivary cortisol (males: +41 ± 24%, p = 0.015; females: +55 ± 24%, p = 0.005). Testosterone levels and the Testosterone/Cortisol ratio remained stable in both males and females. These findings suggest that the physiological stress produced by a HIIT session does not affect immune function and does not disturb the anabolic/catabolic balance.

Effect of a 12-week endurance training program on force transfer and membrane integrity proteins in lean, obese, and type 2 diabetic subjects.

The mechanisms accounting for the loss of muscle function with obesity and type 2 diabetes are likely the result of a combination of neural and muscular factors. One muscular factor that is important, yet has received little attention, is the protein machinery involved in longitudinal and lateral force transmission. The purpose of this study was to compare the levels of force transfer and membrane integrity proteins before and after a 12-week endurance training program in lean, obese, and obese type 2 diabetic adults. Nineteen sedentary subjects (male = 8 and female = 11) were divided into three groups: Lean (n = 7; 50.3 ± 4.1 y; 69.1 ± 7.2 kg); Obese (n = 6; 49.8 ± 4.1 y; 92.9 ± 19.5 kg); and Obese with type 2 diabetes (n = 6; 51.5 ± 7.9 years; 88.9 ± 15.1 kg). Participants trained 150 min/week between 55% and 75% of VO2max for 12 weeks. Skeletal muscle biopsies were taken before and after the training intervention. Baseline dystrophin and muscle LIM protein levels were higher (~50% p < .01) in lean compared to obese and type 2 diabetic adults, while the protein levels of the remaining force transfer and membrane integrity proteins were similar between groups. After training, obese individuals decreased (-53%; p < .01) the levels of the muscle ankyrin repeat protein and lean individuals decreased dystrophin levels (-45%; p = .01), while the levels of the remaining force transfer and membrane integrity proteins were not affected by training. These results suggest that there are modest changes to force transfer and membrane integrity proteins in middle-aged individuals as a result of 12 weeks of lifestyle and training interventions.

Relative lipid oxidation associates directly with mitochondrial fusion phenotype and mitochondria-sarcoplasmic reticulum interactions in human skeletal muscle.

Disturbances in skeletal muscle lipid oxidation might induce ectopic fat deposition and lipotoxicity. Nevertheless, the cellular mechanisms that regulate skeletal muscle lipid oxidation have not been fully determined. We aimed to determine whether there was an association between relative whole body lipid oxidation and mitochondrial size or mitochondria-sarcoplasmic reticulum interactions in the skeletal muscle. Twelve healthy men were included [mean (standard deviation), 24.7 (1.5) yr old, 24.4 (2.6) kg/m2]. The respiratory quotient (RQ) was used to estimate relative lipid oxidation at rest and during exercise (50% maximal oxygen consumption, 600 kcal expended). A skeletal muscle biopsy was obtained from the vastus lateralis at rest. Transmission electron microscopy was used to determine mitochondrial size and mitochondria-sarcoplasmic reticulum interactions (≤50 nm of distance between organelles). Protein levels of fusion/fission regulators were measured in skeletal muscle by Western blot. Resting RQ and exercise RQ associated inversely with intermyofibrillar mitochondrial size (r = -0.66 and r = -0.60, respectively, P < 0.05). Resting RQ also associated inversely with the percentage of intermyofibrillar mitochondria-sarcoplasmic reticulum interactions (r = -0.62, P = 0.03). Finally, intermyofibrillar mitochondrial size associated inversely with lipid droplet density (r = -0.66, P = 0.01) but directly with mitochondria fusion-to-fission ratio (r = 0.61, P = 0.03). Our results show that whole body lipid oxidation is associated with skeletal muscle intermyofibrillar mitochondrial size, fusion phenotype, and mitochondria-sarcoplasmic-reticulum interactions in nondiabetic humans.

Role of Basal Hormones on Sweat Rate and Sweat Na+ Loss in Elite Women Soccer Players.

We aimed to determine whether basal concentrations of testosterone, cortisol or the ratio testosterone/cortisol were related to sweat Na+ loss, sweat Na+ concentration ([Na+]) and sweat rate during exercise. Twenty-two female elite soccer players participated in the study. Testosterone and cortisol were measured in blood samples before exercise. Sweat samples were collected during a training session (~20°C, ~30% RH, and ~0.55 m/s of wind speed) to measure sweat [Na+]. Sweat rate was determined by considering the difference between post-and pre-body weight, along with the amount of liquid consumed. During exercise, sweat Na+ loss (0.33[0.19] g/h) and sweat rate (0.49[0.20] L/h) were related to basal testosterone concentration (1.4[0.4] pg/mL) (r=0.54; r=0.55, respectively; p<0.05), but not with basal cortisol concentration (119.2[24.2] ng/mL) nor testosterone/cortisol ratio (0.012[0.003]) (p>0.05). However, when Na+ loss was adjusted to sweat rate, no association was found between Na+ loss and testosterone (p>0.05). In addition, no differences were found between players with high vs. low Na+ loss adjusted to sweat loss in menstrual phase or intensity during exercise (p>0.05). In conclusion, these results suggest that in these specific environmental conditions, basal levels of testosterone might increase sweat rate and therefore, the amount of Na+ lost during exercise in elite women soccer players.