Kevin A. Zwetsloot

Background: Quercetin (QCT) and citrulline (CIT) have been independently associated with improved antioxidant capacity and nitric oxide (NO) production, potentially enhancing cardiovascular function and exercise performance. This study aimed to evaluate the combined and independent effects of QCT and CIT supplementation on NO metabolites and antioxidant biomarkers in 50 trained cyclists undergoing a 20 km cycling time trial (TT). Methods: In a randomized, double-blind, placebo-controlled design, forty-two male and eight female trained cyclists were assigned to QCT + CIT, QCT, CIT, or placebo (PL) groups. Supplements were consumed twice daily for 28 days. Biochemical assessments included NO metabolites (nitrate/nitrite), ferric reducing antioxidant power (FRAP), superoxide dismutase (SOD) activity, and antioxidant capacity, measured pre- and post-TT. Results: NO metabolites were significantly elevated post-supplementation (p = 0.03); however, no significant interaction effects were observed for NO metabolites, FRAP, SOD, or antioxidant capacity across the groups (p > 0.05). Post-hoc analyses revealed that QCT significantly reduced FRAP concentrations compared to PL (p = 0.01), while no significant changes in SOD or antioxidant capacity were found across any groups. Conclusions: These findings suggest that combined and independent QCT and CIT supplementation did not significantly improve these biomarkers, suggesting that baseline training adaptations, supplementation timing, and individual variability may influence the efficacy of these compounds in enhancing exercise performance and oxidative stress markers. The ergogenic efficacy of QCT + CIT on antioxidant-related markers remains inconclusive.

Murine exercise models are developed to study the molecular and cellular mechanisms regulating muscle mass. A progressive weighted wheel running model, named ‘PoWeR’, was previously developed to serve as a more translatable alternative to involuntary resistance‐type exercise models in rodents, such as synergist ablation. However, mice still run great distances despite the added resistance as evidenced by a large glycolytic‐to‐oxidative shift in muscle fibre type. Thus, PoWeR reflects a blended resistance/endurance model. In an attempt to bias PoWeR further towards resistance‐type exercise, we developed a novel heavy PoWeR model (hPoWeR) utilizing higher wheel loads (max of 12.5 g vs 6 g). Adult male C57BL/6 mice voluntarily performed an 8‐week progressive loading protocol (PoWeR or hPoWeR). Running distance peaked at ∼5–6 km day⁻¹ in both treatments and was maintained by PoWeR mice, but declined in the hPoWeR mice as load increased beyond 7.5 g. Peak isometric force of the gastrocnemius–soleus–plantaris complex tended to increase in wheel running treatments. Soleus mass increased by 19% and 24% in PoWeR and hPoWeR treatments, respectively, and plantaris fibre cross‐sectional area was greater in hPoWeR, compared to PoWeR. There were fewer glycolytic and more oxidative fibres in the soleus and plantaris muscles in the PoWeR treatment, but not hPoWeR. Collectively, these data suggest hPoWeR may modestly alter skeletal muscle supporting the aim of better reflecting typical resistance training adaptations, in line with decreased running volume and exposure to higher resistance. Regardless, PoWeR remains an effective hypertrophic concurrent training model in mice.

The immune system is critical in defending against infection from pathogenic microorganisms. Individuals with weakened immune systems, such as the elderly, are more susceptible to infections and developing autoimmune and inflammatory diseases. The gut microbiome contains a plethora of bacteria and other microorganisms, which collectively plays a significant role in immune function and homeostasis. Gut microbiota are considered to be highly influential on host health and immune function. Therefore, dysbiosis of the microbiota could be a major contributor to the elevated incidence of multiple age-related pathologies. While there seems to be a general consensus that the composition of gut microbiota changes with age, very little is known about how diet and exercise might influence the aging microbiome. Here, we examine the current state of the literature regarding alterations to the gut microbiome as hosts age, drawing particular attention to the knowledge gaps in addressing how diet and exercise influence the aging microbiome. Further, we will demonstrate the need for more controlled studies to investigate the roles that diet and exercise play driving the composition, diversity, and function of the microbiome in an aging population.

Muscle fibers are multinucleated, and muscle fiber nuclei (myonuclei) are believed to be post-mitotic and are typically situated near the periphery of the myofiber. Due to the unique organization of muscle fibers and their nuclei, the cellular and molecular mechanisms regulating myofiber homeostasis in unstressed and stressed conditions (e.g., exercise) are unique. A key role myonuclei play in regulating muscle during exercise is gene transcription. Only recently have investigators had the capability to identify molecular changes at high resolution exclusively in myonuclei in response to perturbations in vivo. The purpose of this review is to describe how myonuclei modulate their transcriptome, epigenetic status, mobility and shape, and microRNA expression in response to exercise in vivo. Given the relative paucity of high-fidelity information on myonucleus-specific contributions to exercise adaptation, we identify specific gaps in knowledge and provide perspectives on future directions of research.

A significant challenge that non-elite collegiate triathletes encounter during competition is the decline in running performance immediately after cycling. Therefore, the purpose of this study was to determine if performing a 40-km bout of cycling immediately before running would negatively influence running economy and mechanical efficiency of running during simulated race conditions in collegiate triathletes. Eight competitive club-level collegiate triathletes randomly performed two trials: cycling for 40 km (Cycle-Run) or running for 5 km (Run–Run), immediately followed by a four-minute running economy and mechanical efficiency of running test at race pace on an instrumented treadmill. Blood lactate, respiratory exchange ratio, mechanical work, energy expenditure, and muscle glycogen were also measured during the four-minute running test. Mechanical efficiency of running, but not running economy, was significantly lower in Cycle-Run, compared to Run–Run (42.1 ± 2.5% vs. 48.1 ± 2.5%, respectively; p = 0.027). Anaerobic energy expenditure was significantly higher in the Cycle-Run trial, compared to the Run–Run trial (16.3 ± 2.4 vs. 7.6 ± 1.1 kJ; p = 0.004); while net (151.0 ± 12.3 vs. 136.6 ± 9.6 kJ; p = 0.204) and aerobic energy expenditure (134.7 ± 12.3 vs. 129.1 ± 10.5 kJ; p = 0.549) were not statistically different between trials. Analysis of blood lactate, respiratory exchange ratio, mechanical work, and changes in muscle glycogen revealed no statistically significant differences between trials. These results suggest that mechanical efficiency of running, but not running economy, is decreased and anaerobic energy expenditure is increased when a 40-km bout of cycling is performed immediately before running in collegiate triathletes.

Previously developed rodent resistance-based exercise models, including synergistic ablation, electrical stimulation, weighted-ladder climbing, and most recently, weighted-sled pulling, are highly effective at providing a hypertrophic stimulus to induce skeletal muscle adaptations. While these models have proven invaluable for skeletal muscle research, they are either invasive or involuntary and labor-intensive. Fortunately, many rodent strains voluntarily run long distances when given access to a running wheel. Loaded wheel running (LWR) models in rodents are capable of inducing adaptations commonly observed with resistance training in humans, such as increased muscle mass and fiber hypertrophy, as well as stimulation of muscle protein synthesis. However, the addition of moderate wheel load either fails to deter mice from running great distances, which is more reflective of an endurance/resistance training model, or the mice discontinue running nearly entirely due to the method of load application. Therefore, a novel high-load wheel running model (HLWR) has been developed for mice where external resistance is applied and progressively increased, enabling mice to continue running with much higher loads than previously utilized. Preliminary results from this novel HLWR model suggest it provides sufficient stimulus to induce hypertrophic adaptations over the 9 week training protocol. Herein, the specific procedures to execute this simple yet inexpensive progressive resistance-based exercise training model in mice are described.

The purpose of this investigation was to demonstrate that muscle fiber mechanics can be assessed on micro-biopsies obtained from human medial gastrocnemii. Three micro-biopsy samples were collected from female dancers (n=15). Single fibers and fiber bundles were isolated and passively stretched from 2.4 to 3.0 µm at 0.015 and 0.04 µm s−1 (n=50 fibers total) and in five increments at 0.12 µm s−1 (n=42 fibers total). Muscle fibers were then activated isometrically at 2.4 µm (n=4 fibers total) and 3.0 µm (n=3 fibers total). Peak stress and steady-state stress were significantly greater (P<0.0001) after stretching at 0.04 µm s−1 than at 0.015 µm s−1. Furthermore, peak stresses and steady-state stresses increased non-linearly with fiber length (P<0.0001). We conclude that active and passive muscle fiber mechanics can be investigated using tissue from micro-biopsies.

Background: Eccentric muscle contractions are commonly used in exercise regimens, as well as in rehabilitation as a treatment against muscle atrophy and weakness. If repeated multiple times, eccentric contractions may result in skeletal muscle injury and loss of function. Skeletal muscle possesses the remarkable ability to repair and regenerate after an injury or damage; however, this ability is impaired with aging. Phytoecdysteroids are natural plant steroids that possess medicinal, pharmacological, and biological properties, with no adverse side effects in mammals. Previous research has demonstrated that administration of phytoecdysteroids, such as 20-hydroxyecdysone (20E), leads to an increase in protein synthesis signaling and skeletal muscle strength.Methods: To investigate whether 20E enhances skeletal muscle recovery from eccentric contraction-induced damage, adult (7–8 mo) and old (26–27 mo) mice were subjected to injurious eccentric contractions (EC), followed by 20E or placebo (PLA) supplementation for 7 days. Contractile function via torque-frequency relationships (TF) was measured three times in each mouse: pre- and post-EC, as well as after the 7-day recovery period. Mice were anesthetized with isoflurane and then electrically-stimulated isometric contractions were performed to obtain in vivo muscle function of the anterior crural muscle group before injury (pre), followed by 150 EC, and then again post-injury (post). Following recovery from anesthesia, mice received either 20E (50 mg•kg⁻¹ BW) or PLA by oral gavage. Mice were gavaged daily for 6 days and on day 7, the TF relationship was reassessed (7-day).Results: EC resulted in significant reductions of muscle function post-injury, regardless of age or treatment condition (p < 0.001). 20E supplementation completely recovered muscle function after 7 days in both adult and old mice (pre vs. 7-day; p > 0.05), while PLA muscle function remained reduced (pre vs. 7-day; p < 0.01). In addition, histological markers of muscle damage appear lower in damaged muscle from 20E-treated mice after the 7-day recovery period, compared to PLA.Conclusions: Taken together, these findings demonstrate that 20E fully recovers skeletal muscle function in both adult and old mice just 7 days after eccentric contraction-induced damage. However, the underlying mechanics by which 20E contributes to the accelerated recovery from muscle damage warrant further investigation.

Skeletal muscle mass and strength are lost with aging. Phytoecdysteroids, in particular 20-hydroxyecdysone (20E), increase protein synthesis in C2C12 skeletal muscle cells and muscle strength in young rats. The objective of this study was to determine whether an extract from Ajuga turkestanica (ATE), enriched in phytoecdysteroids, and 20E affect skeletal muscle mass and fiber size, fiber type, activation of the PI3K–Akt signaling pathway, and the mRNA levels of MAFbx, MuRF-1, and myostatin in sedentary aging mice. Aging male C57BL/6 mice (20 months old) received ATE, 20E, or vehicle (CT) once per day for 28 days or a single acute dose. Treatment did not alter body, muscle, or organ mass; fiber cross-sectional area; or fiber type in the triceps brachii or plantaris muscles. Likewise, protein synthesis signaling markers (i.e., phosphorylation of Akt^Ser473 and p70S6k^Thr389) measured after either 28 days or acutely were unchanged. Neither ATE nor 20E treatment for 28 days affected the mRNA levels of MAFbx, MuRF-1, and myostatin. In conclusion, these data indicate that phytoecdysteroid treatment does not alter muscle mass or fiber type, nor does it activate protein synthesis signaling in the skeletal muscle of sedentary aging mice.

This randomized trial compared pea protein, whey protein, and water-only supplementation on muscle damage, inflammation, delayed onset of muscle soreness (DOMS), and physical fitness test performance during a 5-day period after a 90-min eccentric exercise bout in non-athletic non-obese males (n = 92, ages 18–55 years). The two protein sources (0.9 g protein/kg divided into three doses/day) were administered under double blind procedures. The eccentric exercise protocol induced significant muscle damage and soreness, and reduced bench press and 30-s Wingate performance. Whey protein supplementation significantly attenuated post-exercise blood levels for biomarkers of muscle damage compared to water-only, with large effect sizes for creatine kinase and myoglobin during the fourth and fifth days of recovery (Cohen’s d > 0.80); pea protein versus water supplementation had an intermediate non-significant effect (Cohen’s d < 0.50); and no significant differences between whey and pea protein were found. Whey and pea protein compared to water supplementation had no significant effects on post-exercise DOMS and the fitness tests. In conclusion, high intake of whey protein for 5 days after intensive eccentric exercise mitigated the efflux of muscle damage biomarkers, with the intake of pea protein having an intermediate effect.