Fast-twitch Skeletal Muscles of Dystrophic Mouse Pups Are Resistant to Injury from Acute Mechanical Stress

American Journal of Physiology. Cell Physiology. Oct, 2002  |  Pubmed ID: 12225973

Loss of the dystrophin-glycoprotein complex from muscle sarcolemma in Duchenne's muscular dystrophy (DMD) renders the membrane susceptible to mechanical injury, leaky to Ca(2+), and disrupts signaling, but the precise mechanism(s) leading to the onset of DMD remain unclear. To assess the role of mechanical injury in the onset of DMD, extensor digitorum longus (EDL) muscles from C57 (control), mdx, and mdx-utrophin-deficient [mdx:utrn(-/-); dystrophic] pups aged 9-12 days were subjected to an acute stretch-injury or no-stretch protocol in vitro. Before the stretches, isometric stress was attenuated for mdx:utrn(-/-) compared with control muscles at all stimulation frequencies (P < 0.05). During the stretches, EDL muscles for each genotype demonstrated similar mean stiffness values. After the stretches, isometric stress during a tetanus was decreased significantly for both mdx and mdx:utrn(-/-) muscles compared with control muscles (P < 0.05). Membrane injury assessed by uptake of procion orange dye was greater for dystrophic compared with control EDL (P < 0.05), but, within each genotype, the percentage of total cells taking up dye was not different for the no-stretch vs. stretch condition. These data suggest that the sarcolemma of maturing dystrophic EDL muscles are resistant to acute mechanical injury.

Alterations in Slow-twitch Muscle Phenotype in Transgenic Mice Overexpressing the Ca2+ Buffering Protein Parvalbumin

The Journal of Physiology. Mar, 2003  |  Pubmed ID: 12562945

The purpose of this study was to determine whether induced expression of the Ca2+ buffering protein parvalbumin (PV) in slow-twitch fibres would lead to alterations in physiological, biochemical and molecular properties reflective of a fast fibre phenotype. Transgenic (TG) mice were generated that overexpressed PV in slow (type I) muscle fibres. In soleus muscle (SOL; 58 % type I fibres) total PV expression was 2- to 6-fold higher in TG compared to wild-type (WT) mice. Maximum twitch and tetanic tensions were similar in WT and TG but force at subtetanic frequencies (30 and 50 Hz) was reduced in TG SOL. Twitch time-to-peak tension and half-relaxation time were significantly decreased in TG SOL (time-to-peak tension: 39.3 +/- 2.6 vs. 55.1 +/- 4.7 ms; half-relaxation time: 42.1 +/- 3.5 vs. 68.1 +/- 9.6 ms, P < 0.05 for TG vs. WT, respectively; n = 8-10). There was a significant increase in expression of type IIa myosin heavy chain (MHC) and ryanodine receptor at the mRNA level in TG SOL but there were no differences in MHC expression at the protein level and thus no difference in fibre type. Whole muscle succinate dehydrogenase activity was reduced by 12 +/- 0.4 % in TG SOL and single fibre glycerol-3-phosphate dehydrogenase activity was decreased in a subset of type IIa fibres. These differences were associated with a 64 % reduction in calcineurin activity in TG SOL. These data show that overexpression of PV, resulting in decreased calcineurin activity, can alter the functional and metabolic profile of muscle and influence the expression of key marker genes in a predominantly slow-twitch muscle with minimal effects on the expression of muscle contractile proteins.

Genistein Activates the 3',5'-cyclic Adenosine Monophosphate Signaling Pathway in Vascular Endothelial Cells and Protects Endothelial Barrier Function

Endocrinology. Mar, 2005  |  Pubmed ID: 15591142

The soy phytoestrogen, genistein, has an array of biological actions, including weak estrogenic effects, inhibition of tyrosine kinase, and cellular antioxidant activity. Recent studies showed that genistein may improve vascular function, but the mechanism is unclear. We show that genistein stimulates intracellular cAMP accumulation in intact bovine aortic endothelial cells and human umbilical vein endothelial cells over an incubation period of 30 min. Increases in intracellular cAMP are evoked by as low as 10 nm genistein but not by estrogen. These increases in cAMP may result primarily from enhanced adenylate cyclase activity by a mechanism that does not involve genomic actions or estrogen receptors. The cAMP induced by genistein activates cAMP-dependent protein kinase (PKA) in bovine aortic endothelial cells. The activation of PKA phosphorylates and activates cAMP response element-binding protein, leading to up-regulation of cAMP response element-containing gene expression. In addition, activation of PKA protects thrombin-induced endothelial monolayer permeability, a novel cardioprotective effect of genistein mediated by the cAMP/PKA cascade. These findings demonstrate that a nongenomic action of genistein leads to activation of the cAMP/PKA signaling system to protect the vascular barrier function and alter the expression of cAMP-regulated genes, thereby providing a novel mechanism underlying some of the cardiovascular protective effects proposed for soy phytoestrogens.

Adeno-associated Virus-mediated Microdystrophin Expression Protects Young Mdx Muscle from Contraction-induced Injury

Molecular Therapy : the Journal of the American Society of Gene Therapy. Feb, 2005  |  Pubmed ID: 15668136

Duchenne muscular dystrophy (DMD) is the most common inherited lethal muscle degenerative disease. Currently there is no cure. Highly abbreviated microdystrophin cDNAs were developed recently for adeno-associated virus (AAV)-mediated DMD gene therapy. Among these, a C-terminal-truncated DeltaR4-R23/DeltaC microgene (DeltaR4/DeltaC) has been considered as a very promising therapeutic candidate gene. In this study, we packaged a CMV.DeltaR4/DeltaC cassette in AAV-5 and evaluated the transduction and muscle contractile profiles in the extensor digitorum longus muscles of young (7-week-old) and adult (9-month-old) mdx mice. At approximately 3 months post-gene transfer, 50-60% of the total myofibers were transduced in young mdx muscle and the percentage of centrally nucleated myofibers was reduced from approximately 70% in untreated mdx muscle to approximately 22% in microdystrophin-treated muscle. Importantly, this level of transduction protected mdx muscle from eccentric contraction-induced damage. In contrast, adult mdx muscle was more resistant to AAV-5 transduction, as only approximately 30% of the myofibers were transduced at 3 months postinfection. This transduction yielded marginal protection against eccentric contraction-induced injury. The extent of central nucleation was also more difficult to reverse in adult mdx muscle (from approximately 83% in untreated to approximately 58% in treated). Finally, we determined that the DeltaR4/DeltaC microdystrophin did not significantly alter the expression pattern of the endogenous full-length dystrophin in normal muscle. Neither did it have any adverse effects on normal muscle morphology or contractility. Taken together, our results suggest that AAV-mediated DeltaR4/DeltaC microdystrophin expression represents a promising approach to rescue muscular dystrophy in young mdx skeletal muscle.

Lumbar Extensor Fatigue and Circumferential Ankle Pressure Impair Ankle Joint Motion Sense

Neuroscience Letters. Dec, 2005  |  Pubmed ID: 16102900

Fatigue of the lumbar extensor muscles has been associated with a degradation of balance, but the mechanism is not well understood. The ankle plays a major role in upright standing, and loss of proprioceptive acuity at the ankle could contribute to a degradation of balance. Therefore, the first objective of this study was to investigate the effect of lumbar extensor fatigue on ankle proprioceptive acuity. The second objective was to investigate the effect of circumferential ankle pressure (CAP) on ankle proprioceptive acuity to evaluate CAP as a potential intervention to mitigate any loss of proprioceptive acuity at the ankle with lumbar extensor fatigue. To address these objectives, ankle joint motion sense was evaluated with and without CAP, both before and after the lumbar extensors were fatigued. Results showed an impairment in joint motion sense with both fatigue and CAP. These results indicate that lumbar extensor fatigue impairs ankle proprioceptive acuity, which may help explain observed increases in postural sway subsequent to lumbar extensor fatigue.

Molecular and Cellular Contractile Dysfunction of Dystrophic Muscle from Young Mice

Muscle & Nerve. Jul, 2006  |  Pubmed ID: 16634063

The purpose of this study was to determine whether contractile protein alterations are responsible for force deficits in young dystrophic muscle. Contractility of intact extensor digitorum longus muscles and permeabilized fibers from wild-type (wt), dystrophin-deficient (mdx), and dystrophin/utrophin-deficient (mdx:utrn-/-) mice aged 21 and 35 days was determined. Myosin structural dynamics were assessed by site-directed spin labeling and electron paramagnetic resonance spectroscopy. The principal finding was that force generation was depressed by approximately 20% in mdx muscles, but fiber Ca2+-activated force and myosin structure were not different from wt animals, suggesting that contractile proteins are not responsible for the force deficits in those muscles. For mdx:utrn-/- mice, muscle and fiber forces were approximately 40% lower than wt and the fraction of strong-binding myosin during contraction was reduced by 13%. These data indicate that contractile protein alterations, in addition to myosin dysfunction, cause force deficit in muscles from young mdx:utrn-/- mice. Elucidating the molecular mechanisms underlying muscle weakness at the onset of disease is important for designing treatment strategies.

Increased Extraocular Muscle Strength with Direct Injection of Insulin-like Growth Factor-I

Investigative Ophthalmology & Visual Science. Jun, 2006  |  Pubmed ID: 16723457

Previous work has demonstrated the effectiveness of insulin-like growth factor (IGF)-II in increasing force generation in extraocular muscle (EOM). Studies in the literature have suggested that IGF-I would be even more effective than IGF-II. This study was performed to assess the effects on muscle mass and force generation of IGF-I injection in adult rabbit superior rectus muscle.

Passive Mechanical Properties of Maturing Extensor Digitorum Longus Are Not Affected by Lack of Dystrophin

Muscle & Nerve. Sep, 2006  |  Pubmed ID: 16770793

Mechanical weakness of skeletal muscle is thought to contribute to onset and early progression of Duchenne muscular dystrophy, but this has not been systematically assessed. The purpose of this study was to determine in mice: (1) whether the passive mechanical properties of maturing dystrophic (mdx) muscles were different from control; and (2) if different, the time during maturation when these properties change. Prior to and following the overt onset of the dystrophic process (14-35 days), control and dystrophic extensor digitorum longus (EDL) muscles were subjected to two passive stretch protocols in vitro (5% strain at instantaneous and 1.5 L(0)/s strain rates). Force profiles were fit to a viscoelastic muscle model to determine stiffness and damping. The mdx and control EDL muscles exhibited similar passive mechanical properties at each age, suggesting a functional threshold for dystrophic muscle below which damage may be minimized. Determining this threshold may have important clinical implications for treatments of muscular dystrophy involving physical activity.

Recommendations to Define Exercise Prescription for Duchenne Muscular Dystrophy

Exercise and Sport Sciences Reviews. Jan, 2007  |  Pubmed ID: 17211188

Duchenne muscular dystrophy yields pervasive and progressive muscle weakness. This weakness may be attenuated by regular, low-intensity exercise. However, there is a critical lack of data to support appropriate exercise prescription. Because inappropriate activity may exacerbate the dystrophic process, a systematic analysis of muscle function to determine potential exercise load thresholds to avoid injury in dystrophic mice and dogs, and then in humans, is recommended.

Telomere Shortening in Diaphragm and Tibialis Anterior Muscles of Aged Mdx Mice

Muscle & Nerve. Sep, 2007  |  Pubmed ID: 17617801

The progression of Duchenne muscular dystrophy (DMD) is, in part, due to satellite cell senescence driven by high replicative pressure as these muscle stem cells repeatedly divide and fuse to damaged muscle fibers. We hypothesize that telomere shortening in satellite cells underlies their senescence. To test this hypothesis, we evaluated the diaphragm and a leg muscle from dystrophic mice of various ages for telomere dynamics. We found 30% telomere shortening in tibialis anterior muscles from 600-day-old mdx mice relative to age-matched wildtype mice. We also found a more severe shortening of telomere length in diaphragm muscles of old mdx mice. In those muscles, telomeres were shortened by approximately 15% and 40% in 100- and 600-day-old mdx mice, respectively. These findings indicate that satellite cells undergo telomere erosion, which may contribute to the inability of these cells to perpetually repair DMD muscle.

A Novel SNaPshot Assay to Detect the Mdx Mutation

Muscle & Nerve. Jun, 2008  |  Pubmed ID: 18506711

The mdx mouse is an animal model for Duchenne muscular dystrophy (DMD). In order to evaluate possible treatments and to carry out genetic studies, it is essential to distinguish between mice that carry the dystrophic (mutant) or wild-type (wt) allele(s). The current amplification-resistant mutation system (ARMS) assay is labor intensive and yields false negatives, which reduces its efficiency as a screening tool. An alternate assay based on single-nucleotide polymorphism (SNP) primer extension technology (i.e., SNaPshot) is described. The SNaPshot assay has been optimized to identify both wild-type and mutant alleles, providing a robust, potentially automatable assay for high-throughput analysis.

Endurance Capacity in Maturing Mdx Mice is Markedly Enhanced by Combined Voluntary Wheel Running and Green Tea Extract

Journal of Applied Physiology (Bethesda, Md. : 1985). Sep, 2008  |  Pubmed ID: 18583385

Duchenne muscular dystrophy is characterized by the absence of dystrophin from muscle cells. Dystrophic muscle cells are susceptible to oxidative stress. We tested the hypothesis that 3 wk of endurance exercise starting at age 21 days in young male mdx mice would blunt oxidative stress and improve dystrophic skeletal muscle function, and these effects would be enhanced by the antioxidant green tea extract (GTE). In mice fed normal diet, average daily running distance increased 300% from week 1 to week 3, and total distance over 3 wk was improved by 128% in mice fed GTE. Running, independent of diet, increased serum antioxidant capacity, extensor digitorum longus tetanic stress, and total contractile protein content, heart citrate synthase, and heart and quadriceps beta-hydroxyacyl-CoA dehydrogenase activities. GTE, independent of running, decreased serum creatine kinase and heart and gastrocnemius lipid peroxidation and increased gastrocnemius citrate synthase activity. These data suggest that both endurance exercise and GTE may be beneficial as therapeutic strategies to improve muscle function in mdx mice.

Dysregulated Intracellular Signaling and Inflammatory Gene Expression During Initial Disease Onset in Duchenne Muscular Dystrophy

American Journal of Physical Medicine & Rehabilitation / Association of Academic Physiatrists. Jun, 2009  |  Pubmed ID: 19454857

Duchenne muscular dystrophy is a debilitating genetic disorder characterized by severe muscle wasting and early death in affected boys. The primary cause of this disease is mutations in the dystrophin gene that result in the absence of the protein dystrophin and the associated dystrophin-glycoprotein complex in the plasma membrane of muscle fibers. In normal muscle, this complex forms a link between the extracellular matrix and the cytoskeleton that is thought to protect muscle fibers from contraction-induced membrane lesions and to regulate cell signaling cascades. Although the primary defect is known, the mechanisms that initiate disease onset have not been characterized. Data collected during early maturation suggest that inflammatory and immune responses are key contributors to disease pathogenesis and may be initiated by aberrant signaling in dystrophic muscle. However, detailed time course studies of the inflammatory and immune processes are incomplete and need to be characterized further to understand the disease progression. The purposes of this review are to examine the possibility that initial disease onset in dystrophin-deficient muscle results from aberrant inflammatory signaling pathways and to highlight the potential clinical relevance of targeting these pathways to treat Duchenne muscular dystrophy.

The F-BAR Protein CIP4 Promotes GLUT4 Endocytosis Through Bidirectional Interactions with N-WASp and Dynamin-2

Journal of Cell Science. Jul, 2009  |  Pubmed ID: 19509061

F-BAR proteins are a newly described family of proteins with unknown physiological significance. Because F-BAR proteins, including Cdc42 interacting protein-4 (CIP4), drive membrane deformation and affect endocytosis, we investigated the role of CIP4 in GLUT4 traffic by flow cytometry in GLUT4myc-expressing L6 myoblasts (L6 GLUT4myc). L6 GLUT4myc cells express CIP4a as the predominant F-BAR protein. siRNA knockdown of CIP4 increased insulin-stimulated (14)C-deoxyglucose uptake by elevating cell-surface GLUT4. Enhanced surface GLUT4 was due to decreased endocytosis, which correlated with lower transferrin internalization. Immunoprecipitation of endogenous CIP4 revealed that CIP4 interacted with N-WASp and Dynamin-2 in an insulin-dependent manner. FRET confirmed the insulin-dependent, subcellular properties of these interactions. Insulin exposure stimulated specific interactions in plasma membrane and cytosolic compartments, followed by a steady-state response that underlies the coordination of proteins needed for GLUT4 traffic. Our findings reveal a physiological function for F-BAR proteins, supporting a previously unrecognized role for the F-BAR protein CIP4 in GLUT4 endocytosis, and show that interactions between CIP4 and Dynamin-2 and between CIP4 and NWASp are spatially coordinated to promote function.

Immune-mediated Mechanisms Potentially Regulate the Disease Time-course of Duchenne Muscular Dystrophy and Provide Targets for Therapeutic Intervention

PM & R : the Journal of Injury, Function, and Rehabilitation. Aug, 2009  |  Pubmed ID: 19695529

Duchenne muscular dystrophy is a lethal muscle-wasting disease that affects boys. Mutations in the dystrophin gene result in the absence of the dystrophin glycoprotein complex (DGC) from muscle plasma membranes. In healthy muscle fibers, the DGC forms a link between the extracellular matrix and the cytoskeleton to protect against contraction-induced membrane lesions and to regulate cell signaling. The absence of the DGC results in aberrant regulation of inflammatory signaling cascades. Inflammation is a key pathological characteristic of dystrophic muscle lesion formation. However, the role and regulation of this process in the disease time-course has not been sufficiently examined. The transcription factor nuclear factor-kappaB has been shown to contribute to the disease process and is likely involved with increased inflammatory gene expression, including cytokines and chemokines, found in dystrophic muscle. These aberrant signaling processes may regulate the early time-course of inflammatory events that contribute to the onset of disease. This review critically evaluates the possibility that dystrophic muscle lesions in both patients with Duchenne muscular dystrophy and mdx mice are the result of immune-mediated mechanisms that are regulated by inflammatory signaling and also highlights new therapeutic directions.

Green Tea Extract Decreases Muscle Pathology and NF-kappaB Immunostaining in Regenerating Muscle Fibers of Mdx Mice

Clinical Nutrition (Edinburgh, Scotland). Jun, 2010  |  Pubmed ID: 19897286

Duchenne muscular dystrophy is a debilitating genetic disorder characterized by severe muscle wasting and early death in afflicted boys. The primary cause of this disease is mutations in the dystrophin gene resulting in massive muscle degeneration and inflammation. The purpose of this study was to determine if dystrophic muscle pathology and inflammation were decreased by pre-natal and early dietary intervention with green tea extract.

Foxj3 Transcriptionally Activates Mef2c and Regulates Adult Skeletal Muscle Fiber Type Identity

Developmental Biology. Jan, 2010  |  Pubmed ID: 19914232

The mechanisms that regulate skeletal muscle differentiation, fiber type diversity and muscle regeneration are incompletely defined. Forkhead transcription factors are critical regulators of cellular fate determination, proliferation, and differentiation. We identified a forkhead/winged helix transcription factor, Foxj3, which was expressed in embryonic and adult skeletal muscle. To define the functional role of Foxj3, we examined Foxj3 mutant mice. Foxj3 mutant mice are viable but have significantly fewer Type I slow-twitch myofibers and have impaired skeletal muscle contractile function compared to their wild type controls. In response to a severe injury, Foxj3 mutant mice have impaired muscle regeneration. Foxj3 mutant myogenic progenitor cells have perturbed cell cycle kinetics and decreased expression of Mef2c. Examination of the skeletal muscle 5' upstream enhancer of the Mef2c gene revealed an evolutionary conserved forkhead binding site (FBS). Transcriptional assays in C2C12 myoblasts revealed that Foxj3 transcriptionally activates the Mef2c gene in a dose dependent fashion and binds to the conserved FBS. Together, these studies support the hypothesis that Foxj3 is an important regulator of myofiber identity and muscle regeneration through the transcriptional activation of the Mef2c gene.

Eccentric Contractions Induce Rapid Isometric Torque Drop in Dystrophin-deficient Dogs

Muscle & Nerve. Jul, 2010  |  Pubmed ID: 20544944

We tested the hypothesis that eccentric contractions (ECCs) rapidly induce greater-than-normal isometric torque drop in dystrophin-deficient golden retriever muscular dystrophy (GRMD) muscles. ECCs were imposed by forcibly stretching activated muscles. The results indicate that isometric torque drop was greater in GRMD versus controls (P < 0.0001). Our findings support the hypothesis that ECCs induce greater-than-normal isometric torque drop in GRMD muscles. The magnitude of ECC-induced isometric torque loss may be an ideal clinical endpoint in the GRMD model.

Golden Retriever Muscular Dystrophy (GRMD): Developing and Maintaining a Colony and Physiological Functional Measurements

Methods in Molecular Biology (Clifton, N.J.). 2011  |  Pubmed ID: 21194024

Studies of canine models of Duchenne muscular dystrophy (DMD) provide insight regarding disease pathogenesis and treatment efficacy. To take maximal advantage, colonies of affected dogs must be maintained and outcome parameters developed. In this chapter, we review our 25 years of experience with the golden retriever muscular dystrophy (GRMD) model. Key challenges in colony development (breeding, neonatal death, and the risk of inbreeding) and representative functional measurements (tibiotarsal joint angle and torque force; and eccentric contraction decrement) are discussed.

Exercise and Duchenne Muscular Dystrophy: Toward Evidence-based Exercise Prescription

Muscle & Nerve. Apr, 2011  |  Pubmed ID: 21404285

To develop a rational framework for answering questions about the role of exercise in Duchenne muscular dystrophy (DMD), we focused on five pathophysiological mechanisms and offer brief hypotheses regarding how exercise may beneficially modulate pertinent cellular and molecular pathways. We aimed to provide an integrative overview of mechanisms of DMD pathology that may improve or worsen as a result of exercise. We also sought to stimulate discussion of what outcomes/dependent variables most appropriately measure these mechanisms, with the purpose of defining criteria for well-designed, controlled studies of exercise in DMD. The five mechanisms include pathways that are both intrinsic and extrinsic to the diseased muscle cells.

The Passive Mechanical Properties of the Extensor Digitorum Longus Muscle Are Compromised in 2- to 20-mo-old Mdx Mice

Journal of Applied Physiology (Bethesda, Md. : 1985). Jun, 2011  |  Pubmed ID: 21415170

Muscle rigidity and myotendinous junction (MTJ) deficiency contribute to immobilization in Duchenne muscular dystrophy (DMD), a lethal disease caused by the absence of dystrophin. However, little is known about the muscle passive properties and MTJ strength in a diseased muscle. Here, we hypothesize that dystrophin-deficient muscle pathology renders skeletal muscle stiffer and MTJ weaker. To test our hypothesis, we examined the passive properties of an intact noncontracting muscle-tendon unit in mdx mice, a mouse model for DMD. The extensor digitorum longus (EDL) muscle-tendon preparations of 2-, 6-, 14-, and 20-mo-old mdx and normal control mice were strained stepwisely from 110% to 160% of the muscle optimal length. The stress-strain response and failure position were analyzed. In support of our hypothesis, the mdx EDL preparation consistently developed higher stress before muscle failure. Postfailure stresses decreased dramatically in mdx but not normal preparations. Further, mdx showed a significantly faster stress relaxation rate. Consistent with stress-strain assay results, we observed significantly higher fibrosis in mdx muscle. In 2- and 6-mo-old mdx and 20-mo-old BL10 mice failure occurred within the muscle (2- to 14-mo-old BL10 preparations did not fail). Interestingly, in ≥14-mo-old mdx mice the failure site shifted toward the MTJ. Electron microscopy revealed substantial MTJ degeneration in aged but not young mdx mice. In summary, our results suggest that the passive properties of the EDL muscle and the strength of MTJ are compromised in mdx in an age-dependent manner. These findings offer new insights in studying DMD pathogenesis and developing novel therapies.

Myosin Light-chain Phosphorylation and Potentiation of Dynamic Function in Mouse Fast Muscle

Pflügers Archiv : European Journal of Physiology. Aug, 2011  |  Pubmed ID: 21499697

The intent of this study was to determine if the stimulation-induced increase or "potentiation" of dynamic function of mouse extensor digitorum longus muscle (in vitro 25°C) during work cycles is graded to myosin regulatory light-chain (RLC) phosphorylation. To do this, concentric force and muscle work output during sinusoidal length changes were determined before (unpotentiated) and after (potentiated) the application of conditioning stimuli (CS) producing incremental elevations in RLC phosphorylation from rest. Sine wave excursion was from 1.09 to 0.91 of L (o) with a period of 142 ms; stimulating muscles to twitch and generate force during these cycles produced plots of force × displacement termed work loops. Stimulation at 2.5-, 5.0-, and 100-Hz elevated RLC phosphorylation from 0.16±0.02 (rest) to 0.29±0.03, 0.45±0.02 and 0.56±0.02 mol phos per mole RLC, respectively (n= 6-7, P<0.05). These CS potentiated mean concentric force (at all lengths) to 1.14±0.02, 1.26±0.04 and 1.41±0.06 of pre-stimulus, control levels (all n= 5-7, P<0.05) while work was increased to 1.07±0.02, 1.17±0.02 and 1.34±0.03 of controls, respectively. In a No CS condition that did not elevate RLC phosphorylation, neither mean concentric force nor work was altered. Thus, strong correlations between RLC phosphorylation and mean concentric force and work support the hypothesis that this molecular mechanism modulates muscle power output. No length-dependence for concentric force potentiation was observed in any condition, an outcome suggesting that interactions between instantaneous variations in muscle length and shortening velocity during work cycles modulates the potentiation response.

Dietary Epicatechin Promotes Survival of Obese Diabetic Mice and Drosophila Melanogaster

The Journal of Nutrition. Jun, 2011  |  Pubmed ID: 21525262

The lifespan of diabetic patients is 7-8 y shorter than that of the general population because of hyperglycemia-induced vascular complications and damage to other organs such as the liver and skeletal muscle. Here, we investigated the effects of epicatechin, one of the major flavonoids in cocoa, on health-promoting effects in obese diabetic (db/db) mice (0.25% in drinking water for 15 wk) and Drosophila melanogaster (0.01-8 mmol/L in diet). Dietary intake of epicatechin promoted survival in the diabetic mice (50% mortality in diabetic control group vs. 8.4% in epicatechin group after 15 wk of treatment), whereas blood pressure, blood glucose, food intake, and body weight gain were not significantly altered. Pathological analysis showed that epicatechin administration reduced the degeneration of aortic vessels and blunted fat deposition and hydropic degeneration in the liver caused by diabetes. Epicatechin treatment caused changes in diabetic mice that are associated with a healthier and longer lifespan, including improved skeletal muscle stress output, reduced systematic inflammation markers and serum LDL cholesterol, increased hepatic antioxidant glutathione concentration and total superoxide dismutase activity, decreased circulating insulin-like growth factor-1 (from 303 ± 21 mg/L in the diabetic control group to 189 ± 21 mg/L in the epicatechin-treated group), and improved AMP-activated protein kinase-α activity in the liver and skeletal muscle. Consistently, epicatechin (0.1-8 mmol/L) also promoted survival and increased mean lifespan of Drosophila. Therefore, epicatechin may be a novel food-derived, antiaging compound.

Concerted Regulation of Myofiber-specific Gene Expression and Muscle Performance by the Transcriptional Repressor Sox6

Proceedings of the National Academy of Sciences of the United States of America. Jun, 2011  |  Pubmed ID: 21633012

In response to physiological stimuli, skeletal muscle alters its myofiber composition to significantly affect muscle performance and metabolism. This process requires concerted regulation of myofiber-specific isoforms of sarcomeric and calcium regulatory proteins that couple action potentials to the generation of contractile force. Here, we identify Sox6 as a fast myofiber-enriched repressor of slow muscle gene expression in vivo. Mice lacking Sox6 specifically in skeletal muscle have an increased number of slow myofibers, elevated mitochondrial activity, and exhibit down-regulation of the fast myofiber gene program, resulting in enhanced muscular endurance. In addition, microarray profiling of Sox6 knockout muscle revealed extensive muscle fiber-type remodeling, and identified numerous genes that display distinctive fiber-type enrichment. Sox6 directly represses the transcription of slow myofiber-enriched genes by binding to conserved cis-regulatory elements. These results identify Sox6 as a robust regulator of muscle contractile phenotype and metabolism, and elucidate a mechanism by which functionally related muscle fiber-type specific gene isoforms are collectively controlled.

Mice Lacking MicroRNA 133a Develop Dynamin 2–dependent Centronuclear Myopathy

The Journal of Clinical Investigation. Aug, 2011  |  Pubmed ID: 21737882

MicroRNAs modulate cellular phenotypes by inhibiting expression of mRNA targets. In this study, we have shown that the muscle-specific microRNAs miR-133a-1 and miR-133a-2 are essential for multiple facets of skeletal muscle function and homeostasis in mice. Mice with genetic deletions of miR-133a-1 and miR-133a-2 developed adult-onset centronuclear myopathy in type II (fast-twitch) myofibers, accompanied by impaired mitochondrial function, fast-to-slow myofiber conversion, and disarray of muscle triads (sites of excitation- contraction coupling). These abnormalities mimicked human centronuclear myopathies and could be ascribed, at least in part, to dysregulation of the miR-133a target mRNA that encodes dynamin 2, a GTPase implicated in human centronuclear myopathy. Our findings reveal an essential role for miR-133a in the maintenance of adult skeletal muscle structure, function, bioenergetics, and myofiber identity; they also identify a potential modulator of centronuclear myopathies.

Aging and Physical Mobility in Group-housed Old World Monkeys

Age (Dordrecht, Netherlands). Dec, 2011  |  Pubmed ID: 22203457

While indices of physical mobility such as gait speed are significant predictors of future morbidity/mortality in the elderly, mechanisms of these relationships are not understood. Relevant animal models of aging and physical mobility are needed to study these relationships. The goal of this study was to develop measures of physical mobility including activity levels and gait speed in Old World monkeys which vary with age in adults. Locomotor behaviors of 21 old ([Formula: see text] = 20 yoa) and 24 young ([Formula: see text] = 9 yoa) socially housed adult females of three species were recorded using focal sample and ad libitum behavior observation methods. Self-motivated walking speed was 17% slower in older than younger adults. Likewise, young adults climbed more frequently than older adults. Leaping and jumping were more common, on average, in young adults, but this difference did not reach significance. Overall activity levels did not vary significantly by age, and there were no significant age by species interactions in any of these behaviors. Of all the behaviors evaluated, walking speed measured in a simple and inexpensive manner appeared most sensitive to age and has the added feature of being least affected by differences in housing characteristics. Thus, walking speed may be a useful indicator of decline in physical mobility in nonhuman primate models of aging.

Chronic Administration of a Leupeptin-derived Calpain Inhibitor Fails to Ameliorate Severe Muscle Pathology in a Canine Model of Duchenne Muscular Dystrophy

Frontiers in Pharmacology. 2011  |  Pubmed ID: 22291646

Calpains likely play a role in the pathogenesis of Duchenne muscular dystrophy (DMD). Accordingly, calpain inhibition may provide therapeutic benefit to DMD patients. In the present study, we sought to measure benefit from administration of a novel calpain inhibitor, C101, in a canine muscular dystrophy model. Specifically, we tested the hypothesis that treatment with C101 mitigates progressive weakness and severe muscle pathology observed in young dogs with golden retriever muscular dystrophy (GRMD). Young (6-week-old) GRMD dogs were treated daily with either C101 (17 mg/kg twice daily oral dose, n = 9) or placebo (vehicle only, n = 7) for 8 weeks. A battery of functional tests, including tibiotarsal joint angle, muscle/fat composition, and pelvic limb muscle strength were performed at baseline and every 2 weeks during the 8-week study. Results indicate that C101-treated GRMD dogs maintained strength in their cranial pelvic limb muscles (tibiotarsal flexors) while placebo-treated dogs progressively lost strength. However, concomitant improvement was not observed in posterior pelvic limb muscles (tibiotarsal extensors). C101 treatment did not mitigate force drop following repeated eccentric contractions and no improvement was seen in the development of joint contractures, lean muscle mass, or muscle histopathology. Taken together, these data do not support the hypothesis that treatment with C101 mitigates progressive weakness or ameliorates severe muscle pathology observed in young dogs with GRMD.

Canine Models of Duchenne Muscular Dystrophy and Their Use in Therapeutic Strategies

Mammalian Genome : Official Journal of the International Mammalian Genome Society. Feb, 2012  |  Pubmed ID: 22218699

Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder in which the loss of dystrophin causes progressive degeneration of skeletal and cardiac muscle. Potential therapies that carry substantial risk, such as gene- and cell-based approaches, must first be tested in animal models, notably the mdx mouse and several dystrophin-deficient breeds of dogs, including golden retriever muscular dystrophy (GRMD). Affected dogs have a more severe phenotype, in keeping with that of DMD, so may better predict disease pathogenesis and treatment efficacy. Various phenotypic tests have been developed to characterize disease progression in the GRMD model. These biomarkers range from measures of strength and joint contractures to magnetic resonance imaging. Some of these tests are routinely used in clinical veterinary practice, while others require specialized equipment and expertise. By comparing serial measurements from treated and untreated groups, one can document improvement or delayed progression of disease. Potential treatments for DMD may be broadly categorized as molecular, cellular, or pharmacologic. The GRMD model has increasingly been used to assess efficacy of a range of these therapies. A number of these studies have provided largely general proof-of-concept for the treatment under study. Others have demonstrated efficacy using the biomarkers discussed. Importantly, just as symptoms in DMD vary among patients, GRMD dogs display remarkable phenotypic variation. Though confounding statistical analysis in preclinical trials, this variation offers insight regarding the role that modifier genes play in disease pathogenesis. By correlating functional and mRNA profiling results, gene targets for therapy development can be identified.

The Paradox of Muscle Hypertrophy in Muscular Dystrophy

Physical Medicine and Rehabilitation Clinics of North America. Feb, 2012  |  Pubmed ID: 22239881

Mutations in the dystrophin gene cause Duchenne and Becker muscular dystrophy in humans and syndromes in mice, dogs, and cats. Affected humans and dogs have progressive disease that leads primarily to muscle atrophy. Mdx mice progress through an initial phase of muscle hypertrophy followed by atrophy. Cats have persistent muscle hypertrophy. Hypertrophy in humans has been attributed to deposition of fat and connective tissue (pseudohypertrophy). Increased muscle mass (true hypertrophy) has been documented in animal models. Muscle hypertrophy can exaggerate postural instability and joint contractures. Deleterious consequences of muscle hypertrophy should be considered when developing treatments for muscular dystrophy.

Nutrition Strategies to Improve Physical Capabilities in Duchenne Muscular Dystrophy

Physical Medicine and Rehabilitation Clinics of North America. Feb, 2012  |  Pubmed ID: 22239883

There is no current cure for Duchenne muscular dystrophy (DMD), and palliative and prophylactic interventions to improve the quality of life of patients remain limited, with the exception of corticosteroids. This article describes 2 potential nutritional interventions for the treatment of DMD, green tea extract (GTE) and the branched-chain amino acid leucine, and their positive effects on physical activity. Both GTE and leucine are suitable for human consumption, are easily tolerated with no side effects, and, with appropriate preclinical data, could be brought forward to clinical trials rapidly.

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Journal of Applied Physiology (Bethesda, Md. : 1985). Jan, 2012  |  Pubmed ID: 22253404