Tibialis Anterior Muscle Fibers Research Articles

NNOS Increases Fiber Type-Specific Angiogenesis in Skeletal Muscle of Mice in Response to Endurance Exercise.

We studied the relationship between neuronal NO synthase (nNOS) expression and capillarity in the tibialis anterior (TA) muscle of mice subjected to treadmill training. The mRNA (+131%) and protein (+63%) levels of nNOS were higher (p ≤ 0.05) in the TA muscle of C57BL/6 mice undergoing treadmill training for 28 days than in those of littermates remaining sedentary, indicating an up-regulation of nNOS by endurance exercise. Both TA muscles of 16 C57BL/6 mice were subjected to gene electroporation with either the pIRES2-ZsGreen1 plasmid (control plasmid) or the pIRES2-ZsGreen1-nNOS gene-inserted plasmid (nNOS plasmid). Subsequently, one group of mice (n = 8) underwent treadmill training for seven days, while the second group of mice (n = 8) remained sedentary. At study end, 12-18% of TA muscle fibers expressed the fluorescent reporter gene ZsGreen1. Immunofluorescence for nNOS was 23% higher (p ≤ 0.05) in ZsGreen1-positive fibers than ZsGreen1-negative fibers from the nNOS-transfected TA muscle of mice subjected to treadmill training. Capillary contacts around myosin heavy-chain (MHC)-IIb immunoreactive fibers (14.2%; p ≤ 0.05) were only higher in ZsGreen1-positive fibers than ZsGreen1-negative fibers in the nNOS-plasmid-transfected TA muscles of trained mice. Our observations are in line with an angiogenic effect of quantitative increases in nNOS expression, specifically in type-IIb muscle fibers after treadmill training.

Depleting Ly6G Positive Myeloid Cells Reduces Pancreatic Cancer-Induced Skeletal Muscle Atrophy.

Immune cells can mount desirable anti-cancer immunity. However, some immune cells can support cancer disease progression. The presence of cancer can lead to production of immature myeloid cells from the bone marrow known as myeloid-derived suppressor cells (MDSCs). The immunosuppressive and pro-tumorigenic effects of MDSCs are well understood. Whether MDSCs are involved in promoting cancer cachexia is not well understood. We orthotopically injected the pancreas of mice with KPC cells or PBS. One group of tumor-bearing mice was treated with an anti-Ly6G antibody that depletes granulocytic MDSCs and neutrophils; the other received a control antibody. Anti-Ly6G treatment delayed body mass loss, reduced tibialis anterior (TA) muscle wasting, abolished TA muscle fiber atrophy, reduced diaphragm muscle fiber atrophy of type IIb and IIx fibers, and reduced atrophic gene expression in the TA muscles. Anti-ly6G treatment resulted in greater than 50% Ly6G+ cell depletion efficiency in the tumors and TA muscles. These data show that, in the orthotopic KPC model, anti-Ly6G treatment reduces the number of Ly6G+ cells in the tumor and skeletal muscle and reduces skeletal muscle atrophy. These data implicate Ly6G+ cells, including granulocytic MDSCs and neutrophils, as possible contributors to the development of pancreatic cancer-induced skeletal muscle wasting.

Effect of Photobiomodulation on Denervation-Induced Skeletal Muscle Atrophy and Autophagy: A Study in Mice

ObjectiveThe purpose of this study was to investigate whether photobiomodulation (PBM) can protect against and attenuate muscle atrophy owing to complete peripheral nerve lesion in mice by acting on autophagy. MethodsC57BL/10 mice underwent right sciatic nerve transection to induce tibialis anterior muscle atrophy. After 6 hours of denervation, the mice received PBM (wavelength, 830 nm) daily, transcutaneously over the tibialis anterior muscle region for 5 or 14 days. Some mice with sciatic nerve lesion did not receive PBM. Mice that did not have sciatic nerve lesion and PBM were used as controls. After 5 and 14 days, the right tibialis anterior muscle was examined using histomorphometric (cross-sectional area of muscle fibers), Western blot (levels of the autophagy marker LC3), and immunofluorescence analyses (number of LC3 puncta in the muscle fibers). ResultsThe cross-sectional area of the tibialis anterior muscle fibers decreased after 5 and 14 days of denervation. PBM protected against muscle fiber atrophy after 5 days of denervation and attenuated muscle fiber atrophy after 14 days of denervation. After 5 days of muscle denervation, autophagy did not change, as demonstrated by the comparable levels of LC3-I/II ratio and LC3 puncta between the controls and the mice with atrophic muscle; PBM did not change this profile. After 14 days of denervation, an increased LC3-I/II ratio suggested an ongoing autophagy, which was not affected by PBM. ConclusionPBM attenuated the tibialis anterior muscle atrophy induced by sciatic nerve transection in the mice after at least 5 and 14 days of muscle denervation, without affecting autophagy. The transient protective effect of PBM was observed as early as 5 days after the of complete nerve lesion.

Impaired neuromuscular transmission of the tibialis anterior in a rodent model of hypertonia.

Early-onset hypertonia is characteristic of developmental neuromotor disorders, including cerebral palsy (CP). The spa transgenic mouse displays early-onset spasticity, abnormal gait, and motor impairments that are remarkably similar to symptoms of human CP. Previously, we showed that spa mice have fewer motor neurons innervating the tibialis anterior (TA). An expanded innervation ratio may result in increased susceptibility to neuromuscular transmission failure (NMTF). We assessed NMTF in an ex vivo TA muscle nerve preparation from spa and wild-type (WT) mice by comparing forces elicited by nerve versus muscle stimulation. TA muscle innervation ratio was assessed by counting the number of muscle fibers and dividing by the number of TA motor neurons. Muscle fiber cross-sectional areas were also assessed in the TA muscle. We observed that NMTF was immediately present in spa mice, increased with repetitive stimulation, and associated with increased innervation ratio. These changes were concomitant with reduced TA muscle fiber cross-sectional area in spa mice compared with WT. Early-onset hypertonia is associated with increased innervation ratio and impaired neuromuscular transmission. These disturbances may exacerbate the underlying gait abnormalities present in individuals with hypertonia.NEW & NOTEWORTHY Nerve-muscle interaction is poorly understood in the context of early-onset spasticity and hypertonia. In an animal model of early-onset spasticity, spa mice, we found a marked impairment of tibialis anterior neuromuscular transmission. This impairment is associated with an increased innervation ratio (mean number of muscle fibers innervated by a single motor neuron). These disturbances may underlie weakness and gait disturbances observed in individual with developmental hypertonia and spasticity.

Intraoperative single administration of neutrophil peptide 1 accelerates the early functional recovery of peripheral nerves after crush injury.

Neutrophil peptide 1 belongs to a family of peptides involved in innate immunity. Continuous intramuscular injection of neutrophil peptide 1 can promote the regeneration of peripheral nerves, but clinical application in this manner is not convenient. To this end, the effects of a single intraoperative administration of neutrophil peptide 1 on peripheral nerve regeneration were experimentally observed. A rat model of sciatic nerve crush injury was established using the clamp method. After model establishment, a normal saline group and a neutrophil peptide 1 group were injected with a single dose of normal saline or 10 μg/mL neutrophil peptide 1, respectively. A sham group, without sciatic nerve crush was also prepared as a control. Sciatic nerve function tests, neuroelectrophysiological tests, and hematoxylin-eosin staining showed that the nerve conduction velocity, sciatic functional index, and tibialis anterior muscle fiber cross-sectional area were better in the neutrophil peptide 1 group than in the normal saline group at 4 weeks after surgery. At 4 and 8 weeks after surgery, there were no differences in the wet weight of the tibialis anterior muscle between the neutrophil peptide 1 and saline groups. Histological staining of the sciatic nerve showed no significant differences in the number of myelinated nerve fibers or the axon cross-sectional area between the neutrophil peptide 1 and normal saline groups. The above data confirmed that a single dose of neutrophil peptide 1 during surgery can promote the recovery of neurological function 4 weeks after sciatic nerve injury. All the experiments were approved by the Medical Ethics Committee of Peking University People's Hospital, China (approval No. 2015-50) on December 9, 2015.

Lipid accumulation and mitochondrial abnormalities are associated with fiber atrophy in the skeletal muscle of rats with collagen-induced arthritis

Rheumatoid arthritis (RA) has a negative impact on muscle mass, and reduces patient's mobility and autonomy. Furthermore, RA is associated with metabolic comorbidities, notably in lipid homeostasis by unknown mechanisms. To understand the links between the loss in muscle mass and the metabolic abnormalities, arthritis was induced in male Sprague Dawley rats (n = 11) using the collagen-induced arthritis model. Rats immunized with bovine type II collagen were compared to a control group of animals (n = 11) injected with acetic acid and complete Freund's adjuvant. The clinical severity of the ensuing arthritis was evaluated weekly by a semi-quantitative score. Skeletal muscles from the hind limb were used for the histological analysis and exploration of mitochondrial activity, lipid accumulation, metabolism and regenerative capacities. A significant atrophy in tibialis anterior muscle fibers was observed in the arthritic rats despite a non-significant decrease in the weight of the muscles. Despite moderate inflammation, accumulation of triglycerides (P < 0.05), reduced mitochondrial DNA copy number (P < 0.05) and non-significant dysfunction in mitochondrial cytochrome c oxidase activity were found in the gastrocnemius muscle. Concomitantly, our results suggested an activation of the muscle specific E3 ubiquitin ligases MuRF-1 and MAFbx. Finally, the adipose tissue from the arthritic rats exhibited decreased PPARγ mRNA suggesting reduced adipogenic capacities. In conclusion, the reduced adipose tissue adipogenic capacity and skeletal muscle mitochondrial capacity are probably involved in the activation of protein catabolism, inhibition of myogenesis, accumulation of lipids and fiber atrophy in the skeletal muscle during RA.

Redox responses in skeletal muscle following denervation

Previous studies have shown a significant increase in the mitochondrial generation of hydrogen peroxide (H2O2) and other peroxides in recently denervated muscle fibers. The mechanisms for generation of these peroxides and how the muscle responds to these peroxides are not fully established. The aim of this work was to determine the effect of denervation on the muscle content of proteins that may contribute to mitochondrial peroxide release and the muscle responses to this generation. Denervation of the tibialis anterior (TA) and extensor digitorum longus (EDL) muscles in mice was achieved by surgical removal of a small section of the peroneal nerve prior to its entry into the muscle. An increase in mitochondrial peroxide generation has been observed from 7 days and sustained up to 21 days following denervation in the TA muscle fibers. This increased peroxide generation was reduced by incubation of skinned fibers with inhibitors of monoamine oxidases, NADPH oxidases or phospholipase A2 enzymes and the muscle content of these enzymes together with peroxiredoxin 6 were increased following denervation. Denervated muscle also showed significant adaptations in the content of several enzymes involved in the protection of cells against oxidative damage. Morphological analyses indicated a progressive significant loss of muscle mass in the TA muscle from 7 days up to 21 days following denervation due to fiber atrophy but without fiber loss. These results support the possibility that, at least initially, the increase in peroxide production may stimulate adaptations in an attempt to protect the muscle fibers, but that these processes are insufficient and the increased peroxide generation over the longer term may activate degenerative and atrophic processes in the denervated muscle fibers.

Effects of activated vitamin D, alfacalcidol, and low-intensity aerobic exercise on osteopenia and muscle atrophy in type 2 diabetes mellitus model rats.

Diabetes mellitus causes secondary osteoporosis and muscle atrophy. The ability of alfacalcidol (ALF) and exercise (Exe) to inhibit osteoporosis and muscle atrophy in type 2 diabetes mellitus (T2DM) model rats was examined. Twenty-week-old Otsuka Long-Evans Tokushima Fatty rats were randomized to ALF (orally 0.1 μg/kg/day), Exe (treadmill exercise at 10 m/min, 60 min/day, 5 days/week), Comb (ALF and Exe), and Cont (T2DM control treated with vehicle and no exercise) groups (n = 8–10 per group). Sedentary Long-Evans Tokushima Otsuka rats were used as a non-hyperphagic control. After treatment for 2 or 6 weeks, blood glucose (BG) levels, cross-sectional area (CSA) of tibialis anterior muscle fibers, femoral bone mineral density (BMD), and relative quantities of muscle anabolic markers (Pax7, MyoD, and myogenin) and catabolic markers (Atrogin-1, MuRF1, and REDD1) of the soleus muscle assessed by real-time polymerase chain reaction assays were measured. Exe and Comb treatments for 6 weeks decreased BG levels compared with those of the Cont group. ALF, Exe, and Comb treatments for 2 and 6 weeks recovered the CSA compared with that of the Cont group. ALF and Comb treatments for 6 weeks increased femoral BMDs compared with those of the Cont group. After 2 weeks of treatment, Comb treatment increased MyoD expression and decreased MuRF1 expression. ALF or Exe monotherapy significantly decreased Atrogin-1 or MuRF1 expression after 2 weeks of treatment, respectively. After 6 weeks of treatment, ALF and Comb treatments decreased Atrogin-1 and REDD1. These results demonstrate that a combination of ALF and Exe improved CSA from the early phase of treatment by stimulating skeletal muscle differentiation and suppressing muscle catabolic genes. Improvements in BG, BMD, and CSA were observed as long-term effects of the combination therapy. Continued suppression of muscle catabolic genes was observed as a background to these effects.

Changes In Muscle Biomarkers Following Swimming Exercise in Streptozotoxin-induced Rats

[PURPOSE] The purpose of this study was to assess the effect of swimming exercise in diabetic atrophy by monitoring changes in biomarkers associated with musculoskeletal growth and abnormalities. [METHODS] Ninety-two male Sprague-Dawley rats were randomly divided into four groups: control plus exercise group (CO+Ex), control and no exercise group (CO+No Ex), diabetes plus exercise group (DM+Ex) and diabetes and no exercise group (DM+No Ex). Swimming exercise was implemented for 60 min/day, 5 day/week for a total of eight weeks. Muscle strength and atrophy biomarkers as well as muscle morphology were compared at the first, second, fourth and eighth week of experimentation. [RESULTS] ACE levels were significantly higher in the DM groups (p<.05), however swimming exercise did not significantly affect ACE levels. Irisin levels were significantly higher in the DM and DM+Ex groups compared to the CON, in week one (p<.05), and greater in the DM+Ex group at week two compared to CON+Ex (p<.05), but swimming did not significantly affect irisin levels over the eight-week period in DM rats. In terms of muscle damage, Myl3 expression was unchanged in the CON groups, but significantly higher in the DM+Ex group compared to the CON+Ex group (p<.05). AST was found to be in higher levels in the DM+No Ex group, compared to CON and CON+Ex (p<.05); and DM+Ex group, compared to the CON+Ex group at weeks one, four and eight (p<.05). Tibialis anterior muscle fibers were thicker in the DM+Ex group compared to DM+No Ex, but only at week four (p<.05). Otherwise, no other notable differences were recorded in muscle morphology. [CONCLUSIONS] These results suggest that swimming exercise alone may not be sufficient to induce muscle strength and/or protective effects against muscle atrophy associated with diabetes mellitus. However, this mode of exercise may be considered safe by not compounding the deteriorative effects of this disease.

Denervated muscle fibers induce mitochondrial peroxide generation in neighboring innervated fibers: Role in muscle aging

Disruption of neuromuscular junctions and denervation of some muscle fibers occurs in ageing skeletal muscle and contribute to loss of muscle mass and function. Aging is associated with mitochondrial dysfunction and loss of redox homeostasis potentially occurs through increased mitochondrial generation of reactive oxygen species (ROS). No specific link between increased mitochondrial ROS generation and denervation has been defined in muscle ageing. To address this, we have examined the effect of experimental denervation of all fibers, or only a proportion of the fibers, in the mouse tibialis anterior (TA) muscle on muscle mitochondrial peroxide generation. Transection of the peroneal nerve of mice caused loss of pre-synaptic axons within 1–3 days with no significant morphological changes in post-synaptic structures up to 10 days post-surgery when decreased TA mass and fiber size were apparent. Mitochondria in the denervated muscle showed increased peroxide generation by 3 days post-transection. Use of electron transport chain (ETC) substrates and inhibitors of specific pathways indicated that the ETC was unlikely to contribute to increased ROS generation, but monoamine oxidase B, NADPH oxidase and phospholipase enzymes were implicated. Transection of one of the 3 branches of the peroneal nerve caused denervation of some TA muscle fibers while others retained innervation, but increased mitochondrial peroxide generation occurred in both denervated and innervated fibers. Thus the presence of recently denervated fibers leads to increased ROS generation by mitochondria in neighboring innervated fibers providing a novel explanation for the increased mitochondrial oxidative stress and damage seen with aging in skeletal muscles.

The effect of chlorpyrifos upon ATPase activity of sarcoplasmic reticulum and biomechanics of skeleta l muscle contraction.

We investigated the effect of chlorpyrifos, an organophosphate insecticide, on Ca2+,Mg2+-ATPase activity of sarcoplasmic reticulum and on contraction dynamics (force and length changes) of Rana temporaria m. tibialis anterior muscle fiber bundles. All of the used concentrations of chlorpyrifos (10-6 to 10-5 M) caused decrease of Ca2+,Mg2+-ATPase activity. The inhibition of Ca2+,Mg2+-ATPase activity by chlorpyriphos in concentrations of 10-6 M to 7.5·10-6 M is due to permeation of sarcoplasmic reticulum rather than due to direct enzyme inhibition by organophosphate insecticides. The inhibitory properties of the compound were higher at increased concentration and exposure timeframes. Chlorpyrifos in concentration range of 10-6 to 7.5·10-6 M causes changes in muscle fiber response force that were more pronounced than changes in contractile length. We demonstrated inhibition of Ca2+,Mg2+-ATPase activity caused by noncholinergic effects of chlorpyriphos. It is possible to conclude that influence of organophosphate insecticides happens not only in the neuromuscular transmission but also on the level of subcellular structures.

Disruption of nuclear factor (erythroid-derived-2)-like 2 antioxidant signaling: a mechanism for impaired activation of stem cells and delayed regeneration of skeletal muscle.

Recently we have reported that age-dependent decline in antioxidant levels accelerated apoptosis and skeletal muscle degeneration. Here, we demonstrate genetic ablation of the master cytoprotective transcription factor, nuclear factor (erythroid-derived-2)-like 2 (Nrf2), aggravates cardiotoxin (CTX)-induced tibialis anterior (TA) muscle damage. Disruption of Nrf2 signaling sustained the CTX-induced burden of reactive oxygen species together with compromised expression of antioxidant genes and proteins. Transcript/protein expression of phenotypic markers of muscle differentiation, namely paired box 7 (satellite cell) and early myogenic differentiation and terminal differentiation (myogenin and myosin heavy chain 2) were increased on d 2 and 4 postinjury but later returned to baseline levels on d 8 and 15 in wild-type (WT) mice. In contrast, these responses were persistently augmented in Nrf2-null mice suggesting that regulation of the regeneration-related signaling mechanisms require Nrf2 for normal functioning. Furthermore, Nrf2-null mice displayed slower regeneration marked by dysregulation of embryonic myosin heavy chain temporal expression. Histologic observations illustrated that Nrf2-null mice displayed smaller, immature TA muscle fibers compared with WT counterparts on d 15 after CTX injury. Improvement in TA muscle morphology and gain in muscle mass evident in the WT mice was not noticeable in the Nrf2-null animals. Taken together these data show that the satellite cell activation, proliferation, and differentiation requires a functional Nrf2 system for effective healing following injury.-Shelar, S. B., Narasimhan, M., Shanmugam, G., Litovsky, S. H., Gounder, S. S., Karan, G., Arulvasu, C., Kensler, T. W., Hoidal, J. R., Darley-Usmar, V. M., Rajasekaran, N. S. Disruption of nuclear factor (erythroid-derived-2)-like 2 antioxidant signaling: a mechanism for impaired activation of stem cells and delayed regeneration of skeletal muscle.

Combinatorial therapeutic activation with heparin and AICAR stimulates additive effects on utrophin A expression in dystrophic muscles.

Upregulation of utrophin A is an attractive therapeutic strategy for treating Duchenne muscular dystrophy (DMD). Over the years, several studies revealed that utrophin A is regulated by multiple transcriptional and post-transcriptional mechanisms, and that pharmacological modulation of these pathways stimulates utrophin A expression in dystrophic muscle. In particular, we recently showed that activation of p38 signaling causes an increase in the levels of utrophin A mRNAs and protein by decreasing the functional availability of the destabilizing RNA-binding protein called K-homology splicing regulatory protein, thereby resulting in increases in the stability of existing mRNAs. Here, we treated 6-week-old mdx mice for 4 weeks with the clinically used anticoagulant drug heparin known to activate p38 mitogen-activated protein kinase, and determined the impact of this pharmacological intervention on the dystrophic phenotype. Our results show that heparin treatment of mdx mice caused a significant ∼1.5- to 3-fold increase in utrophin A expression in diaphragm, extensor digitorum longus and tibialis anterior (TA) muscles. In agreement with these findings, heparin-treated diaphragm and TA muscle fibers showed an accumulation of utrophin A and β-dystroglycan along their sarcolemma and displayed improved morphology and structural integrity. Moreover, combinatorial drug treatment using both heparin and 5-amino-4-imidazolecarboxamide riboside (AICAR), the latter targeting 5' adenosine monophosphate-activated protein kinase and the transcriptional activation of utrophin A, caused an additive effect on utrophin A expression in dystrophic muscle. These findings establish that heparin is a relevant therapeutic agent for treating DMD, and illustrate that combinatorial treatment of heparin with AICAR may serve as an effective strategy to further increase utrophin A expression in dystrophic muscle via activation of distinct signaling pathways.

Effects of Hypoxia on the Contractile and Electrical Responses of Chick Embryo Skeletal Muscles in the Last Third of Embryogenesis

Developmental changes in the contractile and electrical responses of isolated fast (tibialis anterior) and slow (soleus) muscles from chick embryo (16–20 days of embryonic development) were studied, as were the effects of hypoxia on them. Normalized contractile response forces for tibialis anterior were significantly greater than those for soleus. On days 16–17 of embryogenesis, soleus and tibialis anterior muscle fibers showed slow decaying oscillation waves of excitation, while extracellular recording identified propagating action potentials in 20% of the tibialis anterior muscle fibers studied. By day 20 of embryonic development, the number of muscle fibers able to generate action potentials in fast muscles approached 100%. In slow muscles, this value was close to 50%. Hypoxia induced a reduction in the strength of the contractile responses of muscles at all study time points during the incubatory development of embryos, but was not found to have any effect on the magnitude of the contractile response evoked by caffeine. These results lead to the conclusion that in these conditions, hypoxia has no influence on the functional state of ryanodine receptors. Muscles treated with insulin and ouabain showed significant reductions in the sensitivity of contractile responses to hypoxia. It is suggested that the effects of hypoxia involve an active role for membrane Na+,K+-ATPase.

Effects of eldecalcitol on bone and skeletal muscles in glucocorticoid-treated rats.

Glucocorticoids cause secondary osteoporosis and myopathy, characterized by type II muscle fiber atrophy. We examined whether a new vitamin D3 analogue, eldecalcitol, could inhibit glucocorticoid-induced osteopenia or myopathy in rats, and also determined the effects of prednisolone (PSL) and/or eldecalcitol on muscle-related gene expression. Six-month-old female Wistar rats were randomized into four groups: PSL group (10mg/kg PSL); E group (0.05µg/kg eldecalcitol); PSL+E group; and control group. PSL, eldecalcitol, and vehicles were administered daily for 2 or 4weeks. Right calf muscle strength, muscle fatigue, cross-sectional areas (CSAs) of left tibialis anterior muscle fibers, and bone mineral density (BMD) were measured following administration. Pax7, MyoD, and myogenin mRNA levels in gastrocnemius muscles were also determined. Muscle strength was significantly higher in the PSL+E group than in the PSL group (p<0.05) after 4weeks, but not after 2weeks. No significant difference in muscle fatigue was seen between groups at 2 or 4weeks. CSAs of type II muscle fibers were significantly larger in the E group and the PSL+E group than in the PSL group at 4weeks (p=0.0093, p=0.0443, respectively). Eldecalcitol treatment for 4weeks maintained the same BMD as the PSL+E group. After 2weeks, but not 4weeks, eldecalcitol treatment significantly increased Pax7 and myogenin mRNA expression in gastrocnemius muscle, and PSL also stimulated myogenin expression. Eldecalcitol appears to increase muscle volume and to protect against femur BMD loss in PSL-administered rats, and it may also stimulate myoblast differentiation into early myotubes.

Structure of the cortical cytoskeleton in fibers of postural muscles and cardiomyocytes of mice after 30-day 2-g centrifugation.

Altered external mechanical loading during spaceflights causes negative effects on muscular and cardiovascular systems. The aim of the study was estimation of the cortical cytoskeleton statement of the skeletal muscle cells and cardiomyocytes. The state of the cortical cytoskeleton in C57BL6J mice soleus, tibialis anterior muscle fibers, and left ventricle cardiomyocytes was investigated after 30-day 2-g centrifugation ("2-g" group) and within 12 h after its completion ("2-g + 12-h" group). We used atomic force microscopy for estimating cell's transverse stiffness, Western blotting for measuring protein content, and RT-PCR for estimating their expression level. The transverse stiffness significantly decreased in cardiomyocytes (by 16%) and increased in skeletal muscles fibers (by 35% for soleus and by 29% for tibialis anterior muscle fibers) in animals of the 2-g group (compared with the control group). For cardiomyocytes, we found that, in the 2-g + 12-h group, α-actinin-1 content decreased in the membranous fraction (by 27%) and increased in cytoplasmic fraction (by 28%) of proteins (compared with the levels in the 2-g group). But for skeletal muscle fibers, similar changes were noted for α-actinin-4, but not for α-actinin-1. In conclusion, we showed that the different isoforms of α-actinins dissociate from cortical cytoskeleton under increased/decreased of mechanical load.

Effect of Low-Level Laser Therapy (808 nm) in Skeletal Muscle After Resistance Exercise Training in Rats

The aim of this study was to evaluate the effects of 808 nm laser applied after a resistance training protocol, on biochemical markers and the morphology of skeletal muscle in rats. Strenuous physical activity results in fatigue and decreased muscle strength, impaired motor control, and muscle pain. Many biochemical and biophysical interventions have been studied in an attempt to accelerate the recovery process of muscle fatigue. Among these, low-level laser therapy (LLLT) has been demonstrated to be effective in increasing skeletal muscle performance in in vivo studies and in clinical trials. However, little is known about the effects of LLLT on muscle performance after resistance training. Thirty Wistar rats were randomly divided into three groups: control group (CG), trained group (TG), and trained and laser-irradiated group (TGL). The resistance training program was performed three times per week for 5 weeks, and consisted of a climbing exercise, with weights attached to the tail of the animal. Furthermore, laser irradiation was performed in the middle region of tibialis anterior (TA) muscle of both legs, after the exercise protocol. Analysis demonstrated that TGL demonstrated significantly reduced resting lactate level and decreased muscle glycogen depletion than the animals that were exercised only, and significantly increased the cross-section area of TA muscle fibers compared with thoseo in the other groups. These results suggest that LLLT could be an effective therapeutic approach in increasing muscle performance during a resistance exercise protocol.

Effects of repeated crush injuries on motor functional recovery of the sciatic nerve

Objectives: The present study was conducted to examine whether repeated crush injuries have significant effects on motor functional recovery of peripheral nerves.Methods: Repeated crush injuries of the sciatic nerve were inflicted on adult rats at 1-week intervals, and functionality of the sciatic nerve was assessed by the static sciatic index each week for 8 weeks after the final injury. To determine the effects of repeated crush injuries on motor functional recovery of the sciatic nerve, tibialis anterior muscle fibers from single and triple crush injuries were examined, and fiber size and fiber reinnervation during the 2- to 4-week period after the final injury were measured.Results: Compared to single crush injuries, which completely recovered by post-injury week 4, double crush injuries resulted in retarded, but complete recovery by post-injury week 6, whereas triple crush injuries resulted in marked retardation in the regenerative process with incomplete recovery during week 8 of the experimental period. Muscle fiber size for rats with triple crush did not recover to normal range at post-injury week 4, despite its normal size for rats with single crush. The rate of reinnervation increased prominently between post-injury weeks 2 and 3 in both injuries, but the rate with triple crush was lower than that with single crush at post-injury week 3.Discussion: These results, which contradict those of a previous study that reported early functional recovery, indicate that repeated crush injuries inhibit motor functional recovery of the damaged sciatic nerve, as evidenced by delayed and incomplete regeneration, atrophied muscle fibers, and delayed reinnervation.

Motor Unit Abnormalities in Dystonia musculorum Mice

Dystonia musculorum (dt) is a mouse inherited sensory neuropathy caused by mutations in the dystonin gene. While the primary pathology lies in the sensory neurons of dt mice, the overt movement disorder suggests motor neurons may also be affected. Here, we report on the contribution of motor neurons to the pathology in dt27J mice. Phenotypic dt27J mice display reduced alpha motor neuron cell number and eccentric alpha motor nuclei in the ventral horn of the lumbar L1 spinal cord region. A dramatic reduction in the total number of motor axons in the ventral root of postnatal day 15 dt27J mice was also evident. Moreover, analysis of the trigeminal nerve of the brainstem showed a 2.4 fold increase in number of degenerating neurons coupled with a decrease in motor neuron number relative to wild type. Aberrant phosphorylation of neurofilaments in the perikaryon region and axonal swellings within the pre-synaptic terminal region of motor neurons were observed. Furthermore, neuromuscular junction staining of dt27J mouse extensor digitorum longus and tibialis anterior muscle fibers showed immature endplates and a significant decrease in axon branching compared to wild type littermates. Muscle atrophy was also observed in dt27J muscle. Ultrastructure analysis revealed amyelinated motor axons in the ventral root of the spinal nerve, suggesting a possible defect in Schwann cells. Finally, behavioral analysis identified defective motor function in dt27J mice. This study reveals neuromuscular defects that likely contribute to the dt27J pathology and identifies a critical role for dystonin outside of sensory neurons.

Increased Expression of Wild-Type or a Centronuclear Myopathy Mutant of Dynamin 2 in Skeletal Muscle of Adult Mice Leads to Structural Defects and Muscle Weakness

Dynamin 2 (DNM2) is a large GTPase implicated in many cellular functions, including cytoskeleton regulation and endocytosis. Although ubiquitously expressed, DNM2 was found mutated in two genetic disorders affecting different tissues: autosomal dominant centronuclear myopathy (ADCNM; skeletal muscle) and peripheral Charcot-Marie-Tooth neuropathy (peripheral nerve). To gain insight into the function of DNM2 in skeletal muscle and the pathological mechanisms leading to ADCNM, we introduced wild-type DNM2 (WT-DNM2) or R465W DNM2 (RW-DNM2), the most common ADCNM mutation, into adult wild-type mouse skeletal muscle by intramuscular adeno-associated virus injections. We detected altered localization of RW-DNM2 in mouse muscle. Several ADCNM features were present in RW-DNM2 mice: fiber atrophy, nuclear mislocalization, and altered mitochondrial staining, with a corresponding reduction in specific maximal muscle force. The sarcomere and triad structures were also altered. We report similar findings in muscle biopsy specimens from an ADCNM patient with the R465W mutation. In addition, expression of wild-type DNM2 induced some muscle defects, albeit to a lesser extent than RW-DNM2, suggesting that the R465W mutation has enhanced activity in vivo. In conclusion, we show the RW-DNM2 mutation acts in a dominant manner to cause ADCNM in adult muscle, and the disease arises from a primary defect in skeletal muscle rather than secondary to peripheral nerve involvement. Therefore, DNM2 plays important roles in the maintenance of adult muscle fibers.