Hematopoietic contribution to skeletal muscle regeneration in acid alpha-glucosidase knockout mice.

Recent studies have shown that cells from bone marrow (BM) can give rise to differentiated skeletal muscle fibers. However, the mechanisms and identities of the cell types involved remain unknown. We performed BM transplantation in acid alpha-glucosidase (GAA) knockout mice, a model of glycogen storage disease type II, and our observations suggested that the BM cells contribute to skeletal muscle fiber formation. Furthermore, we showed that most CD45+:Sca1+ cells have a donor character in regenerating muscle of recipient mice. Based on these findings, CD45+:Sca1+ cells were sorted from regenerating muscles. The cell number was increased with granulocyte colony-stimulating factor after cardiotoxin injury, and the cells were transplanted directly into the tibialis anterior (TA) muscles of GAA knockout mice. Sections of the TA muscles stained with anti-laminin-alpha2 antibody showed that the number of CD45+:Sca1+ cells contributing to muscle fiber formation and glycogen levels were decreased in transplanted muscles. Our results indicated that hematopoietic stem cells, such as CD45+:Sca1+ cells, are involved in skeletal muscle regeneration.     

Skeletal muscle injury induces hepatocyte growth factor expression in spleen

Hepatocyte growth factor (HGF) is present in skeletal muscle and facilitates skeletal muscle regeneration by activating quiescent satellite cells and stimulating their proliferation. However, possible involvement of HGF from non-muscle organs during muscle regeneration is still uncovered. Since liver injury induces HGF expression in distal HGF-producing organs such as lung, kidney and spleen, we examined if this is the case in muscle injury in analogy. In rat femoral muscle, HGF protein levels were elevated within 1 h after muscle injury, with a simultaneous proteolytic activation of HGF protein. Semiquantitative RT-PCR analysis revealed an elevation of HGF mRNA expression after muscle injury in the liver and spleen, and also an increase of HGF protein levels in the spleen, suggesting the presence of endocrine HGF-inducing factor(s) during muscle regeneration. Indeed, the sera from the rat with muscle regeneration were capable of inducing HGF mRNA expression when applied to primary cultured spleen cells from intact rats. These results indicated that skeletal muscle injury induces HGF expression in the non-muscle HGF-producing organs, especially in the spleen, and suggested the possible involvement of non-muscle organ-derived HGF in activation/proliferation of satellite cells during muscle regeneration.     

Muscle Regeneration with Intermuscular Adipose Tissue (IMAT) Accumulation Is Modulated by Mechanical Constraints

Sports trauma are able to induce muscle injury with fibrosis and accumulation of intermuscular adipose tissue (IMAT), which affect muscle function. This study was designed to investigate whether hypoactivity would influence IMAT accumulation in regenerating mouse skeletal muscle using the glycerol model of muscle regeneration. The animals were immediately hindlimb unloaded for 21 days after glycerol injection into the tibialis anterior (TA) muscle. Muscle fiber and adipocyte cross-sectional area (CSA) and IMAT accumulation were determined by histomorphometric analysis. Adipogenesis during regenerative processes was examined using RT-qPCR and Western blot quantification. Twenty-one days of hindlimb unloading resulted in decreases of 38% and 50.6% in the muscle weight/body weight ratio and CSA, respectively, in soleus muscle. Glycerol injection into TA induced IMAT accumulation, reaching 3% of control normal-loading muscle area. This IMAT accumulation was largely inhibited in unloading conditions (0.09%) and concomitant with a marked reduction in perilipin and FABP4 protein content, two key markers of mature adipocytes. Induction of PPARγ and C/EBPα mRNA, two markers of adipogenesis, was also decreased. Furthermore, the protein expression of PDGFRα, a cell surface marker of fibro/adipogenic progenitors, was much lower in regenerating TA from the unloaded group. Exposure of regenerating muscle to hypoactivity severely reduces IMAT development and accumulation. These results provide new insight into the mechanisms regulating IMAT development in skeletal muscle and highlight the importance of taking into account the level of mechanical constraint imposed on skeletal muscle during the regeneration processes.     

Early functional muscle regeneration after myotoxic injury in mice is unaffected by nNOS absence

Nitric oxide (NO) is an important signaling molecule produced in skeletal muscle primarily via the neuronal subtype of NO synthase (NOS1, or nNOS). While many studies have reported NO production to be important in muscle regeneration, none have examined the contribution of nNOS-derived NO to functional muscle regeneration (i.e., restoration of the muscle's ability to produce force) after acute myotoxic injury. In the present study, we tested the hypothesis that genetic deletion of nNOS would impair functional muscle regeneration after myotoxic injury in nNOS(-/-) mice. We found that nNOS(-/-) mice had lower body mass, lower muscle mass, and smaller myofiber cross-sectional area and that their tibialis anterior (TA) muscles produced lower absolute tetanic forces than those of wild-type littermate controls but that normalized or specific force was identical between the strains. In addition, muscles from nNOS(-/-) mice were more resistant to fatigue than those of wild-type littermates (P < 0.05). To determine whether deletion of nNOS affected muscle regeneration, TA muscles from nNOS(-/-) mice and wild-type littermates were injected with the myotoxin notexin to cause complete fiber degeneration, and muscle structure and function were assessed at 7 and 10 days postinjury. Myofiber cross-sectional area was lower in regenerating nNOS(-/-) mice than wild-type controls at 7 and 10 days postinjury; however, contrary to our original hypothesis, no difference in force-producing capacity of the TA muscle was evident between the two groups at either time point. Our findings reveal that nNOS is not essential for functional muscle regeneration after acute myotoxic damage.     

Mobilization of bone marrow stem cells with StemEnhance® improves muscle regeneration in cardiotoxin-induced muscle injury

Bone marrow-derived stem cells have the ability to migrate to sites of tissue damage and participate in tissue regeneration. The number of circulating stem cells has been shown to be a key parameter in this process. Therefore, stimulating the mobilization of bone marrow stem cells may accelerate tissue regeneration in various animal models of injury. In this study we investigated the effect of the bone marrow stem cells mobilizer StemEnhance (SE), a water-soluble extract of the cyanophyta Aphanizomenon flos-aquae (AFA), on hematopoietic recovery after myeloablation as well as recovery from cardiotoxin-induced injury of the anterior tibialis muscle in mice. Control and SE-treated female mice were irradiated, and then transplanted with GFP+ bone marrow stem cells and allowed to recover. Immediately after transplant, animals were gavaged daily with 300 mg/kg of SE in PBS or a PBS control. After hematopoietic recovery (23 days), mice were injected with cardiotoxin in the anterior tibialis muscle. Five weeks later, the anterior tibialis muscles were analyzed for incorporation of GFP+ bone marrow-derived cells using fluorescence imaging. SE significantly enhanced recovery from cardiotoxin-injury. However, StemEnhance did not affect the growth of the animal and did not affect hematopoietic recovery after myeloablation, when compared to control. This study suggests that inducing mobilization of stem cells from the bone marrow is a strategy for muscle regeneration.     

Impaired Macrophage and Satellite Cell Infiltration Occurs in a Muscle-Specific Fashion Following Injury in Diabetic Skeletal Muscle

Background

Systemic elevations in PAI-1 suppress the fibrinolytic pathway leading to poor collagen remodelling and delayed regeneration of tibialis anterior (TA) muscles in type-1 diabetic Akita mice. However, how impaired collagen remodelling was specifically attenuating regeneration in Akita mice remained unknown. Furthermore, given intrinsic differences between muscle groups, it was unclear if the reparative responses between muscle groups were different.

Principal Findings

Here we reveal that diabetic Akita muscles display differential regenerative responses with the TA and gastrocnemius muscles exhibiting reduced regenerating myofiber area compared to wild-type mice, while soleus muscles displayed no difference between animal groups following injury. Collagen levels in TA and gastrocnemius, but not soleus, were significantly increased post-injury versus controls. At 5 days post-injury, when degenerating/necrotic regions were present in both animal groups, Akita TA and gastrocnemius muscles displayed reduced macrophage and satellite cell infiltration and poor myofiber formation. By 10 days post-injury, necrotic regions were absent in wild-type TA but persisted in Akita TA. In contrast, Akita soleus exhibited no impairment in any of these measures compared to wild-type soleus. In an effort to define how impaired collagen turnover was attenuating regeneration in Akita TA, a PAI-1 inhibitor (PAI-039) was orally administered to Akita mice following cardiotoxin injury. PAI-039 administration promoted macrophage and satellite cell infiltration into necrotic areas of the TA and gastrocnemius. Importantly, soleus muscles exhibit the highest inducible expression of MMP-9 following injury, providing a mechanism for normative collagen degradation and injury recovery in this muscle despite systemically elevated PAI-1.

Conclusions

Our findings suggest the mechanism underlying how impaired collagen remodelling in type-1 diabetes results in delayed regeneration is an impairment in macrophage infiltration and satellite cell recruitment to degenerating areas; a phenomena that occurs differentially between muscle groups.     

Wnt signaling induces the myogenic specification of resident CD45+ adult stem cells during muscle regeneration

The observation that CD45(+) stem cells injected into the circulation participate in muscle regeneration raised the question of whether CD45(+) stem cells resident in muscle play a physiological role during regeneration. We found that CD45(+) cells cultured from uninjured muscle were uniformly nonmyogenic. However, CD45(+) cells purified from regenerating muscle readily gave rise to determined myoblasts. The number of CD45(+) cells in muscle rapidly expanded following injury, and a high proportion entered the cell cycle. Investigation of candidate pathways involved in embryonic myogenesis revealed that Wnt signaling was sufficient to induce the myogenic specification of muscle-derived CD45(+) stem cells. Moreover, injection of the Wnt antagonists sFRP2/3 into regenerating muscle markedly reduced CD45(+) stem cell proliferation and myogenic specification. Our data therefore suggest that mobilization of resident CD45(+) stem cells is an important factor in regeneration after injury and highlight the Wnt pathway as a potential therapeutic target for degenerative neuromuscular disease.     

Ultrastructural study of cardiotoxicity and skin alterations in the golden hamster after treatment with 8 different anthracyclines]

Golden hamsters were submitted to i.p. administration during 4 weeks of 8 anthracyclines, adriamycin (ADM), detorubicin (DTR), daunorubicin (DNR), 4'-epi-adriamycin (eADM), adriamycin hydrochloride (ADMh), rubidazon (RBZ), aclacinomycin (ACM) and AD32, at doses equivalent to 3/4 of those which are optimally oncostatic on murine L1210 leukemia. The comparative study of the mortality, the electron microscopic (EM) alterations of the myocardium, and the light microscopic (EM) alterations of the myocardium, and the light microscopic (LM) lesions of the skin, show that ACM and AD32 are the least toxic drugs. EM detected almost no early lesions of myocardium in ACM treated animals, but, after 4 week's treatment, severe cardiac alterations appeared which, like those after AD32 treatment, are non lethal and reversible. Similarly. LM revealed no histologic changes in the skin following ACM and AD32 administrations, but pathologic alterations, atrophy and alopecia, were observed in animals receiving all other drugs.     

Pulsing electromagnetic field therapy in nerve regeneration: an experimental study in the cat

A multidisciplinary approach to the study of peripheral nerve regeneration in the cat has been presented. The purpose of this work has been to determine if pulsing electromagnetic field (PEMF) therapy can enhance peripheral nerve regeneration after injury. In equal groups of animals, two types of pulsing electromagnetic field treatment were compared with untreated controls. All animals underwent quantitative electrophysiologic and morphologic assessment at the area of injury. In addition, muscle fiber sizing in the periphery and retrograde labeling of anterior horn motoneurons with horseradish peroxidase were studied. Results have shown no statistical differences between the groups in electrophysiologic or morphologic parameters. However, in animals treated with a pulse-burst electromagnetic field there was a statistically significant improvement in the labeling and localization of anterior horn cells in the central nervous system. These results indicate that pulse-burst electromagnetic radiation can increase the numbers of motor neurons that reestablish appropriate connections to the periphery after nerve injury. It remains to be seen if this improved spinal cord organization can translate to improved peripheral functional return.     

Electron microscope localization of acetylcholinesterase and butyrylcholinesterase in the superior cervical ganglion of the cat. I. Normal ganglion

The distributions of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in the superior cervical ganglion (SCG) of the cat were determined by electron microscopy (EM) with the bis- (thioacetoxy)aurate (I), or Au(TA)2, method. Before the infusion of fixative, one of the enzymes was selectively, irreversibly inactivated in vivo, as confirmed by light microscope (LM) examination of sections of the stellate ganglion stained by the more specific copper thiocholine method. Physostigmine-treated controls, for inhibition of AChE or BuChE, were stained concomitantly with tissue for enzyme localization by the Au(TA)2 method for EM examination in each experiment. It was concluded that most of the AChE of the cat SCG is present in the plasma membranes of the preganglionic axons and their terminals, and in the dendritic and perikaryonal plasma membranes of the postsynaptic ganglion cells. BuChE is confined largely to the postsynaptic neuronal plasma membranes. Reasons for the discrepancies between the localizations found by the present direct EM observations and those deduced earlier from LM comparisons of normal and denervated SCG are discussed. It is proposed that a trophic factor released by the preganglionic terminals is probably required for the synthesis of postsynaptic neuronal AChE, and that BuChE may serve as a precursor of AChE at that site.     

Effect of therapeutic pulsed ultrasound on parameters of oxidative stress in skeletal muscle after injury

Contusion injuries are a very common form of both athletic and non-athletic injury, that effect muscle function. Treatments to augment the normal repair and regeneration processes are important for a wide variety of patients. Therapeutic ultrasound has been claimed to promote tissue repair, especially by enhancing cell proliferation and protein synthesis. The present study aimed to investigate the effect of therapeutic pulsed ultrasound (TPU) on parameters of oxidative stress, namely thiobarbituric acid-reactive substances (TBARS), protein carbonyl content and the activities of antioxidant enzymes, catalase and superoxide dismutase (SOD), in skeletal muscle after injury. Wistar rats were submitted to an animal model of muscle (gastrocnemius) laceration. TPU was used once a day. One, three or five days after muscle laceration, the animals were killed by decapitation and oxidative stress parameters were evaluated. Serum CK levels were increased in muscle-injured animals, indicating that the laceration animal model was successful. TBARS were not altered after muscle injury, when compared to the sham group. Protein carbonyl content was increased after muscle laceration. Catalase and SOD activities were increased 1 day after muscle injury and not altered at days 3 and 5. TPU decreased TBARS levels after muscle laceration when compared to injured muscle animals without treatment. Protein carbonyl content evaluation presented similar results. It is tempting to speculate that TPU seems to protect the tissue from oxidative injury. TPU diminished catalase and SOD activities, especially on the first day following muscle laceration.     

Calcineurin-A alpha activation enhances the structure and function of regenerating muscles after myotoxic injury

Calcineurin signaling is essential for successful muscle regeneration. Although calcineurin inhibition compromises muscle repair, it is not known whether calcineurin activation can enhance muscle repair after injury. Tibialis anterior (TA) muscles from adult wild-type (WT) and transgenic mice overexpressing the constitutively active calcineurin-A alpha transgene under the control of the mitochondrial creatine kinase promoter (MCK-CnA alpha*) were injected with the myotoxic snake venom Notexin to destroy all muscle fibers. The TA muscle of the contralateral limb served as the uninjured control. Muscle structure was assessed at 5 and 9 days postinjury, and muscle function was tested in situ at 9 days postinjury. Calcineurin stimulation enhanced muscle regeneration and altered levels of myoregulatory factors (MRFs). Recovery of myofiber size and force-producing capacity was hastened in injured muscles of MCK-CnA alpha* mice compared with control. Myogenin levels were greater 5 days postinjury and myocyte enhancer factor 2a (MEF2a) expression was greater 9 days postinjury in muscles of MCK-CnA alpha* mice compared with WT mice. Higher MEF2a expression in regenerating muscles of MCK-CnA alpha* mice 9 days postinjury may be related to an increase of slow fiber genes. Calcineurin activation in uninjured and injured TA muscles slowed muscle contractile properties, reduced fatigability, and enhanced force recovery after 4 min of intermittent maximal stimulation. Therefore, calcineurin activation can confer structural and functional benefits to regenerating skeletal muscles, which may be mediated in part by differential expression of MRFs.     

An absolute requirement for Pax7-positive satellite cells in acute injury-induced skeletal muscle regeneration

Skeletal muscle tissue provides mechanical force for locomotion of all vertebrate animals. It is prone to damage from acute physical trauma and physiological stress. To cope with this, it possesses a tremendous capacity for rapid and effective repair that is widely held to be accomplished by the satellite cells lying between the muscle fiber plasmalemma and the basement membrane. Cell transplantation and lineage-tracing studies have demonstrated that Pax7-expressing (Pax7(+)) satellite cells can repair damaged muscle tissue repeatedly after several bouts of acute injury. These findings provided evidence that Pax7(+) cells are muscle stem cells. However, stem cells from a variety of other origins are also reported to contribute to myofibers upon engraftment into muscles, questioning whether satellite cells are the only stem cell source for muscle regeneration. Here, we have engineered genetic ablation of Pax7(+) cells to test whether there is any significant contribution to muscle regeneration after acute injury from cells other than this source. We find that such elimination of Pax7(+) cells completely blocks regenerative myogenesis either following injury to the tibialis anterior (TA) muscle or after transplantation of extensor digitorum longus (EDL) muscles into nude mice. As Pax7 is specifically expressed in satellite cells, we conclude that they are essential for acute injury-induced muscle regeneration. It remains to be established whether there is any significant role for stem cells of other origins. The implications of our results for muscle stem cell-based therapy are discussed.     

Neuromuscular rehabilitation by treadmill running or electrical stimulation after peripheral nerve injury and repair.

Numerous studies have been devoted to the regeneration of the motor pathway toward a denervated muscle after nerve injury. However, the regeneration of sensory muscle endings after repair by self-anastomosis are little studied. In previous electrophysiological studies, our laboratory showed that the functional characteristics of tibialis anterior muscle afferents are differentially affected after injury and repair of the peroneal nerve with and without chronic electrostimulation. The present study focuses on the axonal regeneration of mechano- (fibers I and II) and metabosensitive (fibers III and IV) muscle afferents by evaluating the recovery of their response to different test agents after nerve injury and repair by self-anastomosis during 10 wk of treadmill running (LSR). Data were compared with control animals (C), animals with nerve lesion and suture (LS), and animals with lesion, suture, and chronic muscle rehabilitation by electrostimulation (LSE) with a biphasic current modulated in pulse duration and frequency, eliciting a pattern mimicking the activity delivered by the nerve to the muscle. Compared with the C group, results indicated that 1) muscle weight was smaller in LS and LSR groups, 2) the fatigue index was greater in the LS group and smaller in the LSE group, 3) metabosensibility remained altered in the LS and LSE groups, and 4) mechanosensitivity presented a large increase of the activation pattern in the LS and LSE groups. Our data indicated that chronic muscle electrostimulation partially favors the recovery of muscle properties (i.e., muscle weight and twitch response were close to the C group) and that rehabilitation by treadmill running also efficiently induced a better functional muscle afferent recovery (i.e., the discharge pattern was similar to the C group). The effectiveness of the chronic electromyostimulation and the treadmill exercise on afferent recovery is discussed with regard to parameters listed above.     

Disruption of muscle renin-angiotensin system in AT1a-/- mice enhances muscle function despite reducing muscle mass but compromises repair after injury

The role of the renin-angiotensin system (RAS) in vasoregulation is well established, but a localized RAS exists in multiple tissues and exerts diverse functions including autonomic control and thermogenesis. The role of the RAS in the maintenance and function of skeletal muscle is not well understood, especially the role of angiotensin peptides, which appear to contribute to muscle atrophy. We tested the hypothesis that mice lacking the angiotensin type 1A receptor (AT(1A)(-/-)) would exhibit enhanced whole body and skeletal muscle function and improved regeneration after severe injury. Despite 18- to 20-wk-old AT(1A)(-/-) mice exhibiting reduced muscle mass compared with controls (P < 0.05), the tibialis anterior (TA) muscles produced a 25% higher maximum specific (normalized) force (P < 0.05). Average fiber cross-sectional area (CSA) and fiber oxidative capacity was not different between groups, but TA muscles from AT(1A)(-/-) mice had a reduced number of muscle fibers as well as a higher proportion of type IIx/b fibers and a lower proportion of type IIa fibers (P < 0.05). Measures of whole body function (grip strength, rotarod performance, locomotor activity) were all improved in AT(1A)(-/-) mice (P < 0.05). Surprisingly, the recovery of muscle mass and fiber CSA following myotoxic injury was impaired in AT(1A)(-/-) mice, in part by impaired myoblast fusion, prolonged collagen infiltration and inflammation, and delayed expression of myogenic regulatory factors. The findings support the therapeutic potential of RAS inhibition for enhancing whole body and skeletal muscle function, but they also reveal the importance of RAS signaling in the maintenance of muscle mass and for normal fiber repair after injury.     

Effects of L-NAME and L-arginine on ischemia-reperfusion injury in rat skeletal muscle

The involvement of nitric oxide in ischemia-reperfusion injury remains controversial and has been reported to be both beneficial and deleterious, depending on the tissue and model used. This study evaluated the effects of the nitric oxide synthase inhibitor N(G)-nitro-L-arginine-methyl ester (L-NAME) and the substrate for nitric oxide synthase, L-arginine on skeletal muscle necrosis in a rat model of ischemia-reperfusion injury. The rectus femoris muscle in male Wistar rats (250 to 500 g) was isolated on its vascular pedicle and subjected to 4 hours of complete arteriovenous occlusion. The animals were divided into five groups: (1) sham-raised control, no ischemia, no treatment (n = 6); (2) 4 hours of ischemia (n = 6); (3) vehicle control, 4 hours of ischemia + saline (n = 6); (4) 4 hours of ischemia + L-arginine infusion (n = 6); and (5) 4 hours of ischemia + L-NAME infusion (n = 6). The infusions (10 mg/kg) were administered into the contralateral femoral vein beginning 5 minutes before reperfusion and during the following 30 to 45 minutes. Upon reperfusion, the muscle was sutured in its anatomic position and all wounds were closed. The percentage of muscle necrosis was assessed after 24 hours of reperfusion by serial transections, nitroblue tetrazolium staining, digital photography, and computerized planimetry. Sham (group 1) animals sustained baseline necrosis of 11.9 +/- 3.0 (percentage necrosis +/- SEM). Four hours of ischemia (group 2) significantly increased necrosis to 79.2 +/- 1.4 (p < 0.01). Vehicle control (group 3) had no significant difference in necrosis (81.17 +/- 5.0) versus untreated animals subjected to 4 hours of ischemia (group 2). Animals treated with L-arginine (group 4) had significantly reduced necrosis to 34.6 +/- 7.5 versus untreated (group 2) animals (p < 0.01). Animals infused with L-NAME (group 5) had no significant difference in necrosis (68.2 +/- 6.7) versus untreated (group 2) animals. L-Arginine (nitric oxide donor) significantly decreased the severity of muscle necrosis in this rat model of ischemia-reperfusion injury. L-arginine is known to increase the amount of nitric oxide through the action of nitric oxide synthase, whereas L-NAME, known to inhibit nitric oxide synthase and decrease nitric oxide production, had comparable results to the untreated 4-hour ischemia group. These results suggest that L-arginine, presumably through nitric oxide mediation, appears beneficial to rat skeletal muscle subjected to ischemia-reperfusion injury.     

Combined Effect of AMPK/PPAR Agonists and Exercise Training in mdx Mice Functional Performance

The present investigation was undertaken to test whether exercise training (ET) associated with AMPK/PPAR agonists (EM) would improve skeletal muscle function in mdx mice. These drugs have the potential to improve oxidative metabolism. This is of particular interest because oxidative muscle fibers are less affected in the course of the disease than glycolitic counterparts. Therefore, a cohort of 34 male congenic C57Bl/10J mdx mice included in this study was randomly assigned into four groups: vehicle solution (V), EM [AICAR (AMPK agonist, 50 mg/Kg-1.day-1, ip) and GW 1516 (PPARδ agonist, 2.5 mg/Kg-1.day-1, gavage)], ET (voluntary running on activity wheel) and EM+ET. Functional performance (grip meter and rotarod), aerobic capacity (running test), muscle histopathology, serum creatine kinase (CK), levels of ubiquitined proteins, oxidative metabolism protein expression (AMPK, PPAR, myoglobin and SCD) and intracellular calcium handling (DHPR, SERCA and NCX) protein expression were analyzed. Treatments started when the animals were two months old and were maintained for one month. A significant functional improvement (p<0.05) was observed in animals submitted to the combination of ET and EM. CK levels were decreased and the expression of proteins related to oxidative metabolism was increased in this group. There were no differences among the groups in the intracellular calcium handling protein expression. To our knowledge, this is the first study that tested the association of ET with EM in an experimental model of muscular dystrophy. Our results suggest that the association of ET and EM should be further tested as a potential therapeutic approach in muscular dystrophies.     

Overexpression of inducible 70-kDa heat shock protein in mouse attenuates skeletal muscle damage induced by cryolesioning

Heat shock protein expression is elevated upon exposure to a variety of stresses and limits the extent of stress-induced damage. To investigate the putative role of inducible 70-kDa heat shock protein (HSP70) in skeletal muscle damage and regeneration, soleus and tibialis anterior (TA) muscles from HSP70-overexpressing transgenic mice were subjected to cryolesioning and analyzed after 1, 10, and 21 days. Histological analysis showed that the muscles from both HSP70 and wild-type mice treated with radicicol (a HSP inducer) had decreased necrosis after cryolesioning compared with controls. The decrease in muscle fiber cross-sectional area in both soleus and TA muscles in 10 days postlesioning was attenuated in HSP70 mice compared with wild-type mice. Glutathione peroxidase activity was increased 1 day after cryolesioning in both HSP70 and control mice and remained elevated for up to 21 days. Immunodetection of neuronal cell adhesion molecule (a satellite cell marker) and developmental/neonatal MHC were significantly lower in cryolesioned HSP70-overexpressing mice than in cryolesioned controls. These results suggest that HSP70 protects skeletal muscle against injury and radicicol might be useful as a skeletal muscle protective agent. regeneration; radicicol; transgenic mouse; myoprotection     

Abnormal Skeletal Muscle Regeneration plus Mild Alterations in Mature Fiber Type Specification in Fktn-Deficient Dystroglycanopathy Muscular Dystrophy Mice

Glycosylated α-dystroglycan provides an essential link between extracellular matrix proteins, like laminin, and the cellular cytoskeleton via the dystrophin-glycoprotein complex. In secondary dystroglycanopathy muscular dystrophy, glycosylation abnormalities disrupt a complex O-mannose glycan necessary for muscle structural integrity and signaling. Fktn-deficient dystroglycanopathy mice develop moderate to severe muscular dystrophy with skeletal muscle developmental and/or regeneration defects. To gain insight into the role of glycosylated α-dystroglycan in these processes, we performed muscle fiber typing in young (2, 4 and 8 week old) and regenerated muscle. In mice with Fktn disruption during skeletal muscle specification (Myf5/Fktn KO), newly regenerated fibers (embryonic myosin heavy chain positive) peaked at 4 weeks old, while total regenerated fibers (centrally nucleated) were highest at 8 weeks old in tibialis anterior (TA) and iliopsoas, indicating peak degeneration/regeneration activity around 4 weeks of age. In contrast, mature fiber type specification at 2, 4 and 8 weeks old was relatively unchanged. Fourteen days after necrotic toxin-induced injury, there was a divergence in muscle fiber types between Myf5/Fktn KO (skeletal-muscle specific) and whole animal knockout induced with tamoxifen post-development (Tam/Fktn KO) despite equivalent time after gene deletion. Notably, Tam/Fktn KO retained higher levels of embryonic myosin heavy chain expression after injury, suggesting a delay or abnormality in differentiation programs. In mature fiber type specification post-injury, there were significant interactions between genotype and toxin parameters for type 1, 2a, and 2x fibers, and a difference between Myf5/Fktn and Tam/Fktn study groups in type 2b fibers. These data suggest that functionally glycosylated α-dystroglycan has a unique role in muscle regeneration and may influence fiber type specification post-injury.     

Biochemical and organoleptic characteristics of muscle from early and late maturing bulls in different production systems

In grass-based beef production systems (PS), early maturing (EM) breed types may be preferable to late maturing (LM) breed types in achieving adequate carcass fat cover. Biochemical and organoleptic characteristics of muscle from suckler bulls were investigated in EM and LM (n=28/breed) assigned to one of two PS (ad libitum concentrates and grass silage to slaughter (C) or ad libitum silage plus 2 kg concentrate daily during winter followed by 99 days at pasture and then an indoor finishing period on C (GSPC)) in a 2 breed type×2 PS factorial arrangement of treatments. Bulls were managed to have a common target carcass weight of 380 kg. Intramuscular fat (IMF) content was higher (P<0.05) for EM than LM, and for C than GSPC bulls. Collagen solubility was higher (P<0.05) for C than GSPC bulls. Lactate dehydrogenase (LDH) and phosphofructokinase activities were higher (P<0.05) for LM than EM. Isocitrate dehydrogenase activity and the Type I myosin heavy chain (MyHC) proportion were higher (P<0.05) for EM than LM. The LDH activity and the Type IIX MyHC proportion were higher (P<0.05) for C than GSPC bulls. Sensory ratings for tenderness and juiciness were higher (P<0.01) for beef from EM than LM while sensory ratings for tenderness, flavour liking and overall liking were higher (P<0.001) for C than for GSPC bulls. Differences in sensory quality were largely eliminated when adjusted for IMF. Overall, carcass fat scores, IMF and sensory scores were higher in EM than LM and in C than GSPC bulls but most differences in sensory quality could be attributed to differences in IMF.