Repeated sprint training under hypoxia improves aerobic performance and repeated sprint ability by enhancing muscle deoxygenation and markers of angiogenesis in rugby sevens.

OBJECTIVE: To evaluate the effects of repeated sprint (RS) training in hypoxia on aerobic performance, repeated sprint ability (RSA), and muscle oxygenation in Rugby Sevens. METHODS: Fourteen Rugby Sevens players were randomly allocated into hypoxic (RSH, FIO2 = 14.5%, n = 7) or normoxic (RSN, FIO2 = 20.9%, n = 7) groups. Both groups underwent RS training consisting of 3 sets of 6-s × 10 sprints at 140% of velocity at peak oxygen uptake ([Formula: see text]) on a motorized treadmill, 3 days/week for 6 weeks in addition to usual training. Hematological variables, hypoxia-inducible factor-1 alpha (HIF-1α), and vascular endothelial growth factor (VEGF) concentrations were measured. Aerobic performance, RSA, and muscle oxygenation during the running-based anaerobic sprint (RAS) test were analyzed. RESULTS: RSH caused no changes in hemoglobin concentration and hematocrit but significant improvements in [Formula: see text] (7.5%, p = 0.03, ES = 1.07), time to exhaustion (17.6%, p = 0.05, ES = 0.92), and fatigue index (FI, - 12.3%, p = 0.01, ES = 1.39) during the RSA test compared to baseline but not RSN. While ∆deoxygenated hemoglobin was significantly increased both after RSH and RSN (p < 0.05), ∆tissue saturation index (- 56.1%, p = 0.01, ES = 1.35) and ∆oxygenated hemoglobin (- 54.7%, p = 0.04, ES = 0.97) were significantly decreased after RSH. These changes were concomitant with increased levels of HIF-1α and VEGF in serum after RSH with a strong negative correlation between ∆FI and ∆deoxygenated hemoglobin after RSH (r = - 0.81, p = 0.03). CONCLUSION: There was minimal benefit from adding RSH to standard Rugby Sevens training, in eliciting improvements in aerobic performance and resistance to fatigue, possibly by enhanced muscle deoxygenation and increased serum HIF-1α and VEGF concentrations.

Heat stress promotes extracellular matrix remodelling via TGF-ß1 and MMP-2/TIMP-2 modulation in tenotomised soleus and plantaris muscles.

PURPOSE: Heat stress has been shown to reduce muscle atrophy and enhance muscle regeneration. However, the role of heat stress on extracellular matrix (ECM) remodelling remains poorly understood. Here, we examined the effect of heat exposure on intramuscular fibrosis and its associated signalling in soleus and plantaris muscles after tenotomy. MATERIALS AND METHODS: Male Wistar rats were randomly divided into four groups: sedentary control (CON), control plus heat stress (CON+HEAT), tenotomy (TEN) for 8 days, and tenotomy for 8 days plus heat stress (TEN+HEAT). Whole body heat stress was maintained at 40.5-41.5 °C for 30 min, 24 h before and 1-6 days after tenotomy. RESULTS: Tenotomy resulted in muscle atrophy and a substantial increase in intramuscular collagen content, which was more pronounced in soleus than in plantaris muscles, whereas laminin content remained unaffected. These effects were associated with increases in MMP-2 activity, TIMP-2, and TGF-ß1 protein expressions. Heat stress, however, attenuated tenotomy-induced intramuscular collagen accumulation in soleus muscle and reduced TIMP-2 and TGF-ß1 protein expressions, but had no effect on MMP-2 activity in both muscles. These alterations were concomitant with the induction of heat shock protein 72 (Hsp72). CONCLUSION: These data demonstrated that heat stress could reduce intramuscular fibrosis, at least in part, through decreasing TGF-ß1 and TIMP-2 protein expressions of tenotomised soleus muscle. The results from this study shed light on the mechanism and suggest the potential therapeutic effect of heat stress in alleviating intramuscular fibrosis after tenotomy.

Effect of supervised rehabilitation combined with blood flow restriction training in athletes with chronic ankle instability: a randomized placebo-controlled trial.

Blood flow restriction (BFR) resistance exercise has been advocated as an alternative approach for improving muscle strength in patients undergoing musculoskeletal rehabilitation. The present study aimed to evaluate the effectiveness of a 4-week supervised rehabilitation (R) with and without BFR on muscle strength, cross-sectional area (CSA), dynamic balance, and functional performance in athletes with chronic ankle instability (CAI). A total of 16 collegiate athletes with CAI participated in this study. They were randomly assigned to the BFR+R group (n=8) or the R group (n=8). Both groups underwent supervised rehabilitation 3 times weekly for 4 consecutive weeks. Additionally, the BFR+R group was applied with a cuff around the proximal thigh at 80% arterial occlusion pressure in addition to the traditional rehabilitation program, whereas the R group received the sham BFR only. Before and after 4 weeks of intervention, isokinetic muscle strength, CSA, Y-balance test, and side hop test (SHT) were measured. Following a 4-week intervention, the BFR+R group exhibited significant improvements in muscle strength of ankle plantarflexor and evertor, CSA of fibularis longus, and SHT timed performance compared with prior training and the R group (all, P<0.05). However, no significant difference was observed on dynamic balance among the groups. The present finding indicated that a 4-week supervised rehabilitation combined with BFR is more effective in improving muscle strength and size and functional performance compared with the traditional rehabilitation alone. This information could have implications for physical therapists and clinician in developing and designing a rehabilitation program for athletes with CAI.

Satellite cell activity in muscle regeneration after contusion in rats.

1. The role of satellite cells in muscle growth during development is well documented, but the involvement of these cells in muscle repair after contusion is less well known. In the present study, we investigated the time-course of satellite cell activity (from 3h to 7days) after contusion of rat gastrocnemius muscle using specific molecular markers for immunofluorescence and real-time polymerase chain reaction (PCR). 2. Inflammation of the injured muscle occurred within 6h, followed by disintegration of the damaged myofibres within 12h. Newly formed myofibres appeared by Day 7. 3. The number of MyoD-positive nuclei (activated satellite cells) in the injured muscle was significantly increased by 6h, reaching a maximum by 12h after contusion. However, the number of MyoD-positive nuclei decreased towards control levels by Day 7. Changes in the number of bromodeoxyuridine-labelled nuclei (proliferating satellite cells) paralleled the changes seen in the number of MyoD-positive nuclei. Conversely, expression of myogenin protein was not apparent until Day 3 and increased further by Day 7. Colabelling of MyoD and myogenin was seen in only a few cells. 4. The time-course of MyoD mRNA expression corresponded with MyoD protein expression. However, there were two peaks in myogenin mRNA expression: 6h and Day 7 after contusion. The second peak coincided with upregulation of myostatin mRNA levels. 5. The results of the present study suggest that contusion activates a homogeneous population of satellite cells to proliferate within 3days, followed by differentiation to form new myofibres. The latter may be regulated, in part, by myostatin.

Effects of Active and Passive Recovery on Muscle Oxygenation and Swimming Performance.

PURPOSE: To compare the effectiveness of 3 recovery protocols on muscle oxygenation, blood lactate, and subsequent performance during a 200-m repeated swim session. METHODS: Twelve collegte swimmers completed 3 sessions of 2 consecutive 200-m front-crawl trials separated by 1 of 3 recovery protocols: a 15-minute active recovery (AR), a 15-minute passive recovery (PR), and a combination of 5-minute AR and 10-minute PR (CR) in a counterbalanced design. Tissue saturation index at biceps femoris, blood lactate concentration, arterial oxygen saturation, and heart rate were measured at rest, immediately after the trial, and at 5, 10, and 15 minutes of recovery. Two-way analysis of variance (recovery × time) with repeated measures was used to determine measurement variables. A level of significance was set at P < .05. RESULTS: No significant changes in swimming time were observed between trials (AR: 156.79 [4.09] vs 157.79 [4.23] s, CR: 156.50 [4.89] vs 155.55 [4.86] s, PR: 156.54 [4.70] vs 156.30 [4.52] s) across recovery conditions. Interestingly, tissue saturation index rapidly declined immediately after a 200-m swim and then gradually returned to baseline, with a greater value observed during CR compared with AR and PR after 15-minute recovery (P = .04). These changes were concomitant with significant reductions in blood lactate and heart rate during the recovery period (P = .00). CONCLUSION: The CR in the present study was more effective in enhancing muscle reoxygenation after a 200-m swim compared with AR and PR, albeit its beneficial effect on subsequent performance warrants further investigation.

Therapeutic Pulsed Ultrasound Promotes Revascularization and Functional Recovery of Rat Skeletal Muscle after Contusion Injury.

The mechanism by which therapeutic pulsed ultrasound (TPU) promotes the repair of damaged gastrocnemius muscle was investigated. Male Wistar rats were divided into uninjured, sham-treated injured and TPU-treated injured (TPU) groups. Injury was induced by mass-drop technique. TPU was applied to the injured muscle for 5 min, daily, started at day 1 post-injury and continuing for 3, 7 and 14 d. For 3 d post-injury, a significant reduction in muscle force was observed in both the sham-treated injured and TPU groups. TPU treatment significantly increased recovery force of the injured muscle after day 7 post-injury. This effect of TPU is associated with increased centronucleated fibers and cross-sectional area, mRNA expression of the vascular endothelial growth factor and capillary density of the regenerated fibers, but not with mRNA expression of nitric oxide synthase. We conclude that TPU hastens muscle recovery, at least in part, by upregulating angiogenesis.

Effect of 20-Hydroxyecdysone on Proteolytic Regulation in Skeletal Muscle Atrophy.

BACKGROUND/AIM: 20-Hydroxyecdystone (20E) is an ecdysteroid hormone which controls molting and reproduction in arthropods. 20E also produces a variety of effects in vertebrates, including enhancing protein synthesis and skeletal muscle regeneration. The effect of 20E on disuse muscle atrophy has not been reported to date. This study examined the proteolytic regulation of 20E in tenotomized rat slow soleus and fast plantaris muscles. MATERIALS AND METHODS: Male Wistar rats were randomly divided into three groups: sedentary control (CON), tenotomy without 20E treatment (TEN), and tenotomy with treatment of 5 mg/kg BW of 20E (TEN+20E). The TEN+20E group was administered 20E via subcutaneous injection to the right thigh for 7 days after tenotomy. RESULTS: 20E treatment tended to attenuate disuse muscle atrophy and reduced ubiquitination only in soleus muscle. CONCLUSION: 20E treatment alleviates skeletal muscle atrophy partially mediated by ubiquitinate pathway, dependent on the muscle phenotype.

Life long calorie restriction increases heat shock proteins and proteasome activity in soleus muscles of Fisher 344 rats.

Heat shock proteins (HSP's) closely interact with 20S proteasome and have been shown to maintain catalytic activity, responsible for the prevention of protein aggregation. A decrease in both proteasome activity and heat shock proteins (HSP's) has been observed with age. We investigated whether life-long calorie restriction (CR), a natural intervention, which prolongs life span, could prevent the age-associated decline in HSP's and restore the proteolytic activity of the 20S proteasome in skeletal muscle. Hence, we investigated HSP's and proteasome activity in the soleus muscle from 12-mo-old (Adult) and 26-28 mo old ad libitum fed (Old), and 26-28 mo old CR (Old-CR; fed 40% of ad libitum for their lifespan) male Fisher 344 rats. Trypsin-like proteasome activity in Old rats was significantly less than both Adult and Old-CR rats. Furthermore, no significant changes where found in chymotrypsin-like proteasome activity due to age or diet. Levels of HSP 72 and 25 were significantly less in Old animals when compared to both Adult and Old-CR rats. In contrast, HSP 90 was elevated in Old rats by 220% compared to adult animals and life-long calorie restriction caused a significant induction (150%) compared to age-matched ad libitum fed animals. Protein carbonyls were significantly elevated in Old when compared to Adult rats, but showed no significant decline due to life long CR. This study shows that HSP's may be largely responsible for the restoration of the trypsin-like activity of the 20S proteasome with age. The large increase in HSP 90 is intriguing and further studies are required to elucidate its role in maintaining 20S proteasome function.

Clenbuterol induces muscle-specific attenuation of atrophy through effects on the ubiquitin-proteasome pathway.

The ubiquitin-proteasome pathway is primarily responsible for myofibrillar protein degradation during hindlimb unweighting (HU). Beta-adrenergic agonists such as clenbuterol (CB) induce muscle hypertrophy and attenuate muscle atrophy due to disuse or inactivity. However, the molecular mechanism by which CB exerts these effects remains poorly understood. The aims of this study were to investigate whether CB attenuates HU-induced muscle atrophy through an inhibition of the ubiquitin-proteasome pathway and whether insulin-like growth factor I (IGF-I) mediates this inhibition. Rats were randomized to the following groups: weight-bearing control, 14-day CB-treated, 14-day HU, and CB + HU. HU-induced atrophy was associated with increased proteolysis and upregulation of components of the ubiquitin-proteasome pathway (ubiquitin conjugates, ubiquitin conjugating enzyme E2-14 kDa, and 20S proteasome activity). Upregulation of the ubiquitin proteasome occurred in all muscles tested but was more pronounced in muscles composed primarily of slow-twitch fibers (soleus) than in fast-twitch muscles (plantaris and tibialis anterior). Although CB induced hypertrophy in all muscles, CB attenuated the HU-induced atrophy and reduced ubiquitin conjugates only in the fast plantaris and tibialis anterior and not in the slow soleus muscle. CB did not elevate IGF-I protein content in either of the muscles examined. These results suggest that CB induces hypertrophy and alleviates HU-induced atrophy, particularly in the fast muscles, at least in part through a muscle-specific inhibition of the ubiquitin-proteasome pathway and that these effects are not mediated by the local production of IGF-I in skeletal muscle.

Mechanical ventilation induces alterations of the ubiquitin-proteasome pathway in the diaphragm.

Prolonged mechanical ventilation (MV) results in diaphragmatic atrophy due, in part, to an increase in proteolysis. These experiments tested the hypothesis that MV-induced diaphragmatic proteolysis is accompanied by increased expression of key components of the ubiquitin-proteasome pathway (UPP). To test this postulate, we investigated the effect of prolonged MV on UPP components and determined the trypsin-like and peptidylglutamyl peptide hydrolyzing activities of the 20S proteasome. Adult Sprague-Dawley rats were assigned to either control or 12-h MV groups (n=7/group). MV animals were anesthetized, tracheostomized, and ventilated with room air for 12 h. Animals in the control group were acutely anesthetized but not exposed to MV. Compared with controls, MV animals demonstrated increased diaphragmatic mRNA levels of two ubiquitin ligases, muscle atrophy F-box (+8.3-fold) and muscle ring finger 1 (+19.0-fold). However, MV did not alter mRNA levels of 14-kDa ubiquitin-conjugating enzyme, polyubiquitin, proteasome-activating complex PA28, or 20S alpha-subunit 7. Protein levels of 14-kDa ubiquitin-conjugating enzyme and proteasome-activating complex PA28 were not altered following MV, but 20S alpha-subunit 7 levels declined (-17.7%). MV increased diaphragmatic trypsin-like activity (+31%) but did not alter peptidylglutamyl peptide hydrolyzing activity. Finally, compared with controls, MV increased ubiquitin-protein conjugates in both the myofibrillar (+24.9%) and cytosolic (+54.7%) fractions of the diaphragm. These results are consistent with the hypothesis that prolonged MV increases diaphragmatic levels of key components within the UPP and that increases in 20S proteasome activity contribute to MV-induced diaphragmatic proteolysis and atrophy.

Stevioside enhances satellite cell activation by inhibiting of NF-κB signaling pathway in regenerating muscle after cardiotoxin-induced injury.

Stevioside, a noncaloric sweetener isolated from Stevia rebaudiana, exhibits anti-inflammatory and immunomodulatory effects through interference of nuclear factor (NF)-kappa B pathway. We investigated whether this anti-inflammatory property of stevioside could improve muscle regeneration following cardiotoxin-induced muscle injury. Adult male Wistar rats received stevioside orally at an accepted daily dosage of 10 mg kg⁻¹ for 7 days before cardiotoxin injection at the tibialis anterior (TA) muscle of the right hindlimb (the left hindlimb served as control), and stevioside administration was continued for 3 and 7 days. TA muscle was examined at days 3 and 7 postinjury. Although stevioside treatment had no significant effect in enhancing muscle regeneration as indicated by the absence of decreased muscle inflammation or improved myofibrillar protein content compared with vehicle treated injured group at day 7 postinjury, the number of MyoD-positive nuclei were increased (P < 0.05), with a corresponding decrease in NF-κB nuclear translocation (P < 0.05). This is the first study to demonstrate that stevioside could enhance satellite cell activation by modulation of the NF-κB signaling pathway in regenerating muscle following injury. Thus, stevioside may be beneficial as a dietary supplementation for promoting muscle recovery from injury. However, its pharmacological effect on muscle function recovery warrants further investigation.

Trolox attenuates mechanical ventilation-induced diaphragmatic dysfunction and proteolysis.

Prolonged mechanical ventilation results in diaphragmatic oxidative injury, elevated proteolysis, fiber atrophy, and reduced force-generating capacity. We tested the hypothesis that antioxidant infusion during mechanical ventilation would function as an antioxidant to maintain redox balance within diaphragm muscle fibers and therefore prevent oxidative stress and subsequent proteolysis and contractile dysfunction. Sprague-Dawley rats were anesthetized, tracheostomized, and mechanically ventilated with 21% O(2) for 12 hours. The antioxidant Trolox was intravenously infused in a subset of ventilated animals. Compared with acutely anesthetized, nonventilated control animals, mechanical ventilation resulted in a significant reduction (-17%) in diaphragmatic maximal tetanic force. Importantly, Trolox completely attenuated this mechanical ventilation-induced diaphragmatic contractile deficit. Total diaphragmatic proteolysis was increased 105% in mechanical ventilation animals compared with controls. In contrast, diaphragmatic proteolysis did not differ between controls and mechanical ventilation-Trolox animals. Moreover, 20S proteasome activity in the diaphragm was elevated in the mechanical ventilation animals (+76%); Trolox treatment attenuated this mechanical ventilation-induced rise in protease activity. These results are consistent with the hypothesis that mechanical ventilation-induced oxidative stress is an important factor regulating mechanical ventilation-induced diaphragmatic proteolysis and contractile dysfunction. Our findings suggest that antioxidant therapy could be beneficial during prolonged mechanical ventilation.

Mechanical ventilation-induced diaphragmatic atrophy is associated with oxidative injury and increased proteolytic activity.

Prolonged mechanical ventilation (MV) results in reduced diaphragmatic maximal force production and diaphragmatic atrophy. To investigate the mechanisms responsible for MV-induced diaphragmatic atrophy, we tested the hypothesis that controlled MV results in oxidation of diaphragmatic proteins and increased diaphragmatic proteolysis due to elevated protease activity. Further, we postulated that MV would result in atrophy of all diaphragmatic muscle fiber types. Mechanically ventilated animals were anesthetized, tracheostomized, and ventilated with 21% O2 for 18 hours. MV resulted in a decrease (p < 0.05) in diaphragmatic myofibrillar protein and the cross-sectional area of all muscle fiber types (i.e., I, IIa, IId/x, and IIb). Further, MV promoted an increase (p < 0.05) in diaphragmatic protein degradation along with elevated (p < 0.05) calpain and 20S proteasome activity. Finally, MV was also associated with a rise (p < 0.05) in both protein oxidation and lipid peroxidation. These data support the hypothesis that MV is associated with atrophy of all diaphragmatic fiber types, increased diaphragmatic protease activity, and augmented diaphragmatic oxidative stress.