Magnetic resonance imaging of graded skeletal muscle injury in live rats.

INTRODUCTION: Increasing number of stretch-shortening contractions (SSCs) results in increased muscle injury. METHODS: Fischer Hybrid rats were acutely exposed to an increasing number of SSCs in vivo using a custom-designed dynamometer. Magnetic resonance imaging (MRI) imaging was conducted 72 hours after exposure when rats were infused with Prohance and imaged using a 7T rodent MRI system (GE Epic 12.0). Images were acquired in the transverse plane with typically 60 total slices acquired covering the entire length of the hind legs. Rats were euthanized after MRI, the lower limbs removed, and tibialis anterior muscles were prepared for histology and quantified stereology. RESULTS: Stereological analyses showed myofiber degeneration, and cellular infiltrates significantly increased following 70 and 150 SSC exposure compared to controls. MRI images revealed that the percent affected area significantly increased with exposure in all SSC groups in a graded fashion. Signal intensity also significantly increased with increasing SSC repetitions. DISCUSSION: These results suggest that contrast-enhanced MRI has the sensitivity to differentiate specific degrees of skeletal muscle strain injury, and imaging data are specifically representative of cellular histopathology quantified via stereological analyses.

Effects of glutathione depletion and age on skeletal muscle performance and morphology following chronic stretch-shortening contraction exposure.

The involvement of glutathione in the response of skeletal muscle following repetitive, high-intensity mechanical loading is not known. We examined the influence of a glutathione antagonist [L: -Buthionine Sulfoximine (BSO)] had on the adaptability of skeletal muscle during chronic mechanical loading via stretch-shortening contractions (SSCs) in young and old rats. Left dorsiflexor muscles of young (12 weeks, N = 16) and old (30 months, N = 16), vehicle- and BSO-treated rats were exposed three times per week for 4.5-weeks to a protocol of 80 maximal SSCs per exposure in vivo. Skeletal muscle response to the SSC exposure was characterized by muscle performance, as well as muscle wet-weight and quantitative morphological analyses following the exposure period. Results reveal that generally, muscle performance increased in the young rats only following chronic SSC exposure. BSO treatment had no effect on muscle performance or morphology following the chronic SSC exposure in old rats. Muscle wet-weight was increased following exposure compared with the contra-lateral control limb, irrespective of age (p < 0.05). Muscle cross-sectional area increased approximately 20% with SSC loading in the young, vehicle rats, while increasing approximately 10% with SSC loading in old, vehicle rats compared with control rat muscle. No degenerative myofibers were noted in either age group, but edema were increased as a result of aging (p < 0.05). We conclude that our results indicate that glutathione depletion does not adversely affect muscle performance or morphology in old rats. Nevertheless, we continue to show that aging negatively influences performance and morphology following chronic SSC exposure.

Effects of age and glutathione levels on oxidative stress in rats after chronic exposure to stretch-shortening contractions.

We investigated effects of age and glutathione synthesis inhibition on the oxidative stress status of tibialis anterior muscles from young and old Fisher 344 x Brown Norway male rats after chronic administration of stretch-shortening contractions. Oral supplementation of L: -buthionine-(S,R)-sulfoximine (BSO) inhibited glutathione synthesis. Dorsiflexor muscles in the hindlimb were exposed to 80 maximal stretch-shortening contractions (SSCs) three times per week for 4.5 weeks. We measured malondialdehyde, hydrogen peroxide (H(2)O(2)), and free isoprostanes to determine oxidative stress. Glutathione peroxidase activity was measured as an indicator of H(2)O(2) scavenging. Glutathione measurements confirmed the effectiveness of BSO treatment. In young rats, the SSC exposure protocol prevented oxidative stress and enhanced H(2)O(2) scavenging. In old rats, malondialdehyde was increased in the exposed muscle and a BSO-induced increase in H(2)O(2) was not alleviated with SSC exposure as seen in young rats. In addition, glutathione peroxidase activity and total glutathione were increased in old rats relative to their young counterparts. All comparisons were significant at the 0.05 level. Overall, BSO administration was effective in decreasing total glutathione levels and increasing H(2)O(2) levels in old and young rats exposed to SSCs. In addition, effects of chronic exposure to high-force resistive loading SSCs in active muscle from old animals are: (1) antioxidant capacity is enhanced similar to what is seen with endurance training and (2) oxidative stress is increased, probably as a consequence of the enhanced vulnerability due to aging.

Response of tibialis anterior tendon to a chronic exposure of stretch-shortening cycles: age effects.

BACKGROUND: The purpose of the current study was to investigate the effects of aging on tendon response to repetitive exposures of stretch-shortening cycles (SSC's). METHODS: The left hind limb from young (3 mo, N = 4) and old (30 mo, N = 9) male Fisher 344 x Brown Norway rats were exposed to 80 maximal SSCs (60 deg/s, 50 deg range of motion) 3 x/week for 4.5 weeks in vivo. After the last exposure, tendons from the tibialis anterior muscle were isolated, stored at -80 degrees C, and then tested using a micro-mechanical testing machine. Deformation of each tendon was evaluated using both relative grip-to-grip displacements and reference marks via a video system. RESULTS: At failure, the young control tendons had higher strain magnitude than the young exposed (p < 0.01) and the old control tendons (p < .0001). Total load at inflection was affected by age only (p < 0.01). Old exposed and control tendons exhibited significantly higher loads at the inflection point than their young counterparts (p < 0.05 for both comparisons). At failure, the old exposed tendons carried higher loads than the young exposed tendons (p < 0.05). Stiffness was affected by age only at failure where the old tendons exhibited higher stiffness in both exposed and control tendons than their young counterparts (p < 0.05 and p < 0.01, respectively). CONCLUSION: The chronic protocol enhanced the elastic stiffness of young tendon and the loads in both the young and old tendons. The old exposed tendons were found to exhibit higher load capacity than their younger counterparts, which differed from our initial hypothesis.

Injury and adaptive mechanisms in skeletal muscle.

Work-related musculoskeletal disorders (MSD) are a major concern in the United States. Overexertion and repetitive motion injuries dominate reporting of lost-time MSD incidents. Over the past three decades, there has been much study on contraction-induced skeletal muscle injury. The effect of the biomechanical loading signature that includes velocity, range of motion, the number of repetitions, force, work-rest cycle, and exposure duration has been studied. More recently, the effect of aging on muscle injury susceptibility and regeneration has been studied. This review will focus on contraction-induced skeletal muscle injury, the effects of repetitions, range of motion, work-rest cycles, and aging on injury susceptibility and regenerative and adaptive pathways. The different physiological phenomena responsive to overt muscle injury versus adaptation will be distinguished. The inherent capability of skeletal muscle to adapt to mechanical loading, given the appropriate exposure signature will also be discussed. Finally, we will submit that repeated high-intensity mechanical loading is a desirable means to attenuate the effects of sarcopenia, and may be the most effective and appealing mode of physical activity to counteract the effects often observed with musculo-skeletal dysfunction in the workplace.

Adaptive stretch-shortening contractions: diminished regenerative capacity with aging.

This study determined the age-related changes in acute events responsible for initiating skeletal muscle remodeling and (or) regeneration in the tibialis anterior muscle following a bout of stretch-shortening contractions (SSCs). Changes in muscle performance and morphology were quantified in young and old rats, following an acute exposure to adaptive SSCs at 6, 24, 48, 72, and 120 h postexposure (n = 6 for each age at each recovery period). Following SSC exposure, all performance measures were decreased in old rats throughout the 120 h acute phase. Estimates of edema were increased in the old vs. young exposed muscle at 120 h recovery. Both young and old rats displayed an increase in developmental myosin heavy chain (MHCdev+) labeling in the exposed muscle, indicating muscle regeneration. However, old rats displayed diminished MHCdev+ labeling, compared with young rats, suggesting limited remodeling and (or) regenerative capacity. Based on these data, diminished local muscle remodeling and (or) regeneration with aging may limit skeletal muscle adaptation following mechanical loading.

Embryonic atrial function is essential for mouse embryogenesis, cardiac morphogenesis and angiogenesis.

The requirement for atrial function in developing heart is unknown. To address this question, we have generated mice deficient in atrial myosin light chain 2 (MLC2a), a major structural component of the atrial myofibrillar apparatus. Inactivation of the Mlc2a gene resulted in severely diminished atrial contraction and consequent embryonic lethality at ED10.5-11.5, demonstrating that atrial function is essential for embryogenesis. Our data also address two longstanding questions in cardiovascular development: the connection between function and form during cardiac morphogenesis, and the requirement for cardiac function during vascular development. Diminished atrial function in MLC2a-null embryos resulted in a number of consistent secondary abnormalities in both cardiac morphogenesis and angiogenesis. Our results unequivocally demonstrate that normal cardiac function is directly linked to normal morphogenic development of heart and vasculature. These data have important implications for the etiology of congenital heart disease.

Characterization and in vivo functional analysis of splice variants of cypher.

Previously, we reported two splice variants of Cypher, a striated muscle-specific PDZLIM domain protein, Cypher1 and Cypher2. We have now characterized four additional splice isoforms, two of which are novel. The six isoforms can be divided into skeletal or cardiac specific classes, based on the inclusion of skeletal or cardiac specific domains. Short and long isoforms share an N-terminal PDZ domain, but the three C-terminal LIM domains are unique to long isoforms. By RNA and protein analysis, we have demonstrated that Cypher isoforms are developmentally regulated in both skeletal and cardiac muscle. We have previously shown that knockout of Cypher is neonatal lethal. To investigate the function of splice variants in vivo, we have performed a rescue experiment of the Cypher null mutant by replacing the endogenous Cypher gene with cDNAs encoding either a short or long skeletal muscle isoform. In contrast to Cypher null mice, a percentage of mice that express only a short or a long skeletal muscle-specific isoform can survive to at least 1 year of age. Although surviving mice exhibit muscle pathology, these results suggest that either isoform is sufficient to rescue the lethality associated with the absence of Cypher.