MBNL1-Associated Mitochondrial Dysfunction and Apoptosis in C2C12 Myotubes and Mouse Skeletal Muscle.

We explored the interrelationship between a tissue-specific alternative splicing factor muscleblind-like 1 (MBNL1) and peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC-1α), B-cell lymphoma 2 (Bcl-2) or Bcl-2-associated X protein (Bax) in C2C12 myotubes and mouse skeletal muscle to investigate a possible physiological role of MBNL1 in mitochondrial-associated apoptosis of skeletal muscle. Expression level of PGC-1α and mitochondrial membrane potential evaluated by the fluorescence ratio of JC-1 aggregate to monomer in C2C12 myotubes were suppressed by knockdown of MBNL1. Conversely, the ratio of Bax to Bcl-2 as well as the apoptotic index in C2C12 myotubes was increased by MBNL1 knockdown. In plantaris muscle, on the other hand, not only the minimum muscle fiber diameter but also the expression level of MBNL1 and PGC-1α in of 100-week-old mice were significantly lower than that of 10-week-old mice. Furthermore, the ratio of Bax to Bcl-2 in mouse plantaris muscle was increased by aging. These results suggest that MBNL1 may play a key role in aging-associated muscle atrophy accompanied with mitochondrial dysfunction and apoptosis via mediating PGC-1α expression in skeletal muscle.

AMPK Mediates Muscle Mass Change But Not the Transition of Myosin Heavy Chain Isoforms during Unloading and Reloading of Skeletal Muscles in Mice.

5'AMP-activated protein kinase (AMPK) plays an important role in the regulation of skeletal muscle mass and fiber-type distribution. However, it is unclear whether AMPK is involved in muscle mass change or transition of myosin heavy chain (MyHC) isoforms in response to unloading or increased loading. Here, we checked whether AMPK controls muscle mass change and transition of MyHC isoforms during unloading and reloading using mice expressing a skeletal-muscle-specific dominant-negative AMPKα1 (AMPK-DN). Fourteen days of hindlimb unloading reduced the soleus muscle weight in wild-type and AMPK-DN mice, but reduction in the muscle mass was partly attenuated in AMPK-DN mice. There was no difference in the regrown muscle weight between the mice after 7 days of reloading, and there was concomitantly reduced AMPKα2 activity, however it was higher in AMPK-DN mice after 14 days reloading. No difference was observed between the mice in relation to the levels of slow-type MyHC I, fast-type MyHC IIa/x, and MyHC IIb isoforms following unloading and reloading. The levels of 72-kDa heat-shock protein, which preserves muscle mass, increased in AMPK-DN-mice. Our results indicate that AMPK mediates the progress of atrophy during unloading and regrowth of atrophied muscles following reloading, but it does not influence the transition of MyHC isoforms.

Potential involvement of dietary advanced glycation end products in impairment of skeletal muscle growth and muscle contractile function in mice.

Diets enriched with advanced glycation end products (AGE) have recently been related to muscle dysfunction processes. However, it remains unclear whether long-term exposure to an AGE-enriched diet impacts physiological characteristics of skeletal muscles. Therefore, we explored the differences in skeletal muscle mass, contractile function and molecular responses between mice receiving a diet high in AGE (H-AGE) and low in AGE (L-AGE) for 16 weeks. There were no significant differences between L-AGE and H-AGE mice with regard to body weight, food intake or epididymal fat pad weight. However, extensor digitorum longus (EDL) and plantaris (PLA) muscle weights in H-AGE mice were lower compared with L-AGE mice. Higher levels of N ε -(carboxymethyl)-l-lysine, a marker for AGE, in EDL muscles of H-AGE mice were observed compared with L-AGE mice. H-AGE mice showed lower muscle strength and endurance in vivo and lower muscle force production of PLA muscle in vitro. mRNA expression levels of myogenic factors including myogenic factor 5 and myogenic differentiation in EDL muscle were lower in H-AGE mice compared with L-AGE mice. The phosphorylation status of 70-kDa ribosomal protein S6 kinase Thr389, an indicator of protein synthesis signalling, was lower in EDL muscle of H-AGE mice than that of L-AGE mice. These findings suggest that long-term exposure to an AGE-enriched diet impairs skeletal muscle growth and muscle contractile function, and that these muscle dysfunctions may be attributed to the inhibition of myogenic potential and protein synthesis.

Long Term Changes in Muscles around the Knee Joint after ACL Resection in Rats: Comparisons of ACL-Resected, Contralateral and Normal Limb.

The purpose of this study was to investigate the long-term effects of anterior cruciate ligament (ACL) resection on the morphological and contractile characteristics of rectus femoris (RF) and semimembranosus (SM) muscles in both injured and contralateral hindlimbs in rats. Wistar male rats (8-week old) were used. Rats were divided into two groups; ACL-resected and (sham-operated) control groups. Furthermore, right and left limbs of rats in the ACL-resected group were assigned as ACL-resected and contralateral groups, respectively, at 1 day, 1, 4, and 48 weeks after ACL resection. No ACL-resection-associated changes in the mass of both muscles were observed 1 week after ACL resection. On the other hand, ACL-resection-associated reduction on mean fiber cross-sectional area (fiber CSA) in RF muscle lasted 48 weeks after ACL resection. Furthermore, ACL-resection associated increase in fiber composition of type I fiber in RF muscle in contralateral limbs. In addition, long-term effects of ACL resection were observed in both ACL-resected and contralateral limbs. Evidences from this study suggested that ACL resection may cause to change in the morphological (fiber CSA) and contractile (distribution of fiber types) properties of skeletal muscles around the knee joint in not only injured but also contralateral limb. Rehabilitation for quantitative and qualitative muscle changes by ACL resection may be required a special care for a long-term period.

Suppression of Myostatin Stimulates Regenerative Potential of Injured Antigravitational Soleus Muscle in Mice under Unloading Condition.

Effects of myostatin (MSTN)-suppression on the regeneration of injured skeletal muscle under unloading condition were investigated by using transgenic mice expressing a dominant-negative form of MSTN (MSTN-DN). Both MSTN-DN and wild-type (WT) mice were subjected to continuous hindlimb suspension (HS) for 6 weeks. Cardiotoxin (CTX) was injected into left soleus muscle under anesthesia 2 weeks after the initiation of HS. Then, the soleus muscles were excised following 6-week HS (4 weeks after CTX-injection). CTX-injection caused to reduce the soleus fiber cross-sectional area (CSA) in WT mice under both unloading and weight-bearing conditions, but not in MSTN-DN mice. Under unloading condition, CTX-injected muscle weight and fiber CSA in MSTN-DN mice were significantly higher than those in WT mice. CTX-injected muscle had many damaged and regenerating fibers having central nuclei in both WT and MSTN-DN mice. Significant increase in the population of Pax7-positive nuclei in CTX-injected muscle was observed in MSTN-DN mice, but not in WT mice. Evidences indicate that the suppression of MSTN cause to increase the regenerative potential of injured soleus muscle via the increase in the population of muscle satellite cells regardless of unloading conditions.

Involvement of AMPK in regulating slow-twitch muscle atrophy during hindlimb unloading in mice.

AMPK is considered to have a role in regulating skeletal muscle mass. However, there are no studies investigating the function of AMPK in modulating skeletal muscle mass during atrophic conditions. In the present study, we investigated the difference in unloading-associated muscle atrophy and molecular functions in response to 2-wk hindlimb suspension between transgenic mice overexpressing the dominant-negative mutant of AMPK (AMPK-DN) and their wild-type (WT) littermates. Male WT (n = 24) and AMPK-DN (n = 24) mice were randomly divided into two groups: an untreated preexperimental control group (n = 12 in each group) and an unloading (n = 12 in each group) group. The relative soleus muscle weight and fiber cross-sectional area to body weight were decreased by ∼30% in WT mice by hindlimb unloading and by ∼20% in AMPK-DN mice. There were no changes in puromycin-labeled protein or Akt/70-kDa ribosomal S6 kinase signaling, the indicators of protein synthesis. The expressions of ubiquitinated proteins and muscle RING finger 1 mRNA and protein, markers of the ubiquitin-proteasome system, were increased by hindlimb unloading in WT mice but not in AMPK-DN mice. The expressions of molecules related to the protein degradation system, phosphorylated forkhead box class O3a, inhibitor of κBα, microRNA (miR)-1, and miR-23a, were decreased only in WT mice in response to hindlimb unloading, and 72-kDa heat shock protein expression was higher in AMPK-DN mice than in WT mice. These results imply that AMPK partially regulates unloading-induced atrophy of slow-twitch muscle possibly through modulation of the protein degradation system, especially the ubiquitin-proteasome system.

Microcurrent electrical neuromuscular stimulation facilitates regeneration of injured skeletal muscle in mice.

Conservative therapies, mainly resting care for the damaged muscle, are generally used as a treatment for skeletal muscle injuries (such as muscle fragmentation). Several past studies reported that microcurrent electrical neuromuscular stimulation (MENS) facilitates a repair of injured soft tissues and shortens the recovery period. However, the effects of MENS on the regeneration in injured skeletal muscle are still unclear. The purpose of this study was to investigate the effect of MENS on the regenerative process of injured skeletal muscle and to elucidate whether satellite cells in injured skeletal muscle are activated by MENS by using animal models. Male C57BL/6J mice, aged 7 weeks old, were used (n = 30). Mice were randomly divided into two groups: (1) cardiotoxin (CTX)-injected (CX, n = 15) and (2) CTX-injected with MENS treatment (MX, n=15) groups. CTX was injected into tibialis anterior muscle (TA) of mice in CX and MX groups to initiate the necrosis-regeneration cycle of the muscle. TA was dissected 1, 2, and 3 weeks after the injection. Muscle weight, muscle protein content, the mean cross-sectional areas of muscle fibers, the relative percentage of fibers having central nuclei, and the number of muscle satellite cells were evaluated. MENS facilitated the recovery of the muscle dry weight and protein content relative to body weight, and the mean cross-sectional areas of muscle fibers in CTX-induced injured TA muscle. The number of Pax7-positive muscle satellite cells was increased by MENS during the regenerating period. Decrease in the percentages of fibers with central nuclei after CTX-injection was facilitated by MENS. MENS may facilitate the regeneration of injured skeletal muscles by activating the regenerative potential of skeletal muscles. Key pointsMicrocurrent electrical neuromuscular stimulation (MENS) facilitated the recovery of the relative muscle dry weight, the relative muscle protein content, and the mean cross-sectional areas of muscle fibers of injured TA muscle in mice.The number of satellite cells was increased by MENS during the regenerating phase of injured skeletal muscle.Decrease in the percentages of fibers with central nuclei was facilitated by MENS.MENS may facilitate the regeneration of injured skeletal muscles.

AICAR-induced activation of AMPK negatively regulates myotube hypertrophy through the HSP72-mediated pathway in C2C12 skeletal muscle cells.

5'-AMP-activated protein kinase (AMPK) plays an important role as a negative regulator of skeletal muscle mass. However, the precise mechanism of AMPK-mediated regulation of muscle mass is not fully clarified. Heat shock proteins (HSPs), stress-induced molecular chaperones, are related with skeletal muscle adaptation, but the association between AMPK and HSPs in skeletal muscle hypertrophy is unknown. Thus, we investigated whether AMPK regulates hypertrophy by mediating HSPs in C2C12 cells. The treatment with AICAR, a potent stimulator of AMPK, decreased 72-kDa HSP (HSP72) expression, whereas there were no changes in the expressions of 25-kDa HSP, 70-kDa heat shock cognate, and heat shock transcription factor 1 in myotubes. Protein content and diameter were less in the AICAR-treated myotubes in those without treatment. AICAR-induced suppression of myotube hypertrophy and HSP72 expression was attenuated in the siRNA-mediated AMPKα knockdown myotubes. AICAR increased microRNA (miR)-1, a modulator of HSP72, and the increase of miR-1 was not induced in AMPKα knockdown condition. Furthermore, siRNA-mediated HSP72 knockdown blocked AICAR-induced inhibition of myotube hypertrophy. AICAR upregulated the gene expression of muscle Ring-finger 1, and this alteration was suppressed in either AMPKα or HSP72 knockdown myotubes. The phosphorylation of p70 S6 kinase Thr(389) was downregulated by AICAR, whereas this was attenuated in AMPKα, but not in HSP72, knockdown myotubes. These results suggest that AMPK inhibits hypertrophy through, in part, an HSP72-associated mechanism via miR-1 and protein degradation pathways in skeletal muscle cells.

Glycative stress inhibits hypertrophy and impairs cell membrane integrity in overloaded mouse skeletal muscle.

BACKGROUND: Glycative stress, characterized by the formation and accumulation of advanced glycation end products (AGEs) associated with protein glycation reactions, has been implicated in inducing a decline of muscle function. Although the inverse correlation between glycative stress and muscle mass and strength has been demonstrated, the underlying molecular mechanisms are not fully understood. This study aimed to elucidate how glycative stress affects the skeletal muscle, particularly the adaptive muscle response to hypertrophic stimuli and its molecular mechanism. METHODS: Male C57BL/6NCr mice were randomly divided into the following two groups: the bovine serum albumin (BSA)-treated and AGE-treated groups. Mice in the AGE-treated group were intraperitoneally administered AGEs (0.5 mg/g) once daily, whereas those in the BSA-treated group received an equal amount of BSA (0.5 mg/g) as the vehicle control. After 7 days of continuous administration, the right leg plantaris muscle of mice in each group underwent functional overload treatment by synergist ablation for 7 days to induce muscle hypertrophy. In in vitro studies, cultured C2C12 myocytes were treated with AGEs (1 mg/mL) to examine cell adhesion and cell membrane permeability. RESULTS: Continuous AGE administration increased the levels of fluorescent AGEs, Nε-(carboxymethyl) lysine, and methylglyoxal-derived hydroimidazolone-1 in both plasma and skeletal muscle. Plantaris muscle weight, muscle fibre cross-sectional area, protein synthesis rate, and the number of myonuclei increased with functional overload in both groups; however, the increase was significantly reduced by AGE treatment. Some muscles of AGE-treated mice were destroyed by functional overload. Proteomic analysis was performed to explore the mechanisms of muscle hypertrophy suppression and myofibre destruction by AGEs. When principal component analysis was performed on 4659 data obtained by proteomic analysis, AGE treatment was observed to affect protein expression only in functionally overloaded muscles. Enrichment analysis of the 436 proteins extracted using the K-means method further identified a group of proteins involved in cell adhesion. Consistent with this finding, dystrophin-glycoprotein complex proteins and cell adhesion-related proteins were confirmed to increase with functional overload; however, this was attenuated by AGE treatment. Additionally, the treatment of C2C12 muscle cells with AGEs inhibited their ability to adhere and increased cell membrane permeability. CONCLUSIONS: This study indicates that glycative stress may be a novel pathogenic factor in skeletal muscle dysfunctions by causing loss of membrane integrity and preventing muscle mass gain.

Microcurrent electrical nerve stimulation facilitates regrowth of mouse soleus muscle.

Microcurrent electrical nerve stimulation (MENS) has been used to facilitate recovery from skeletal muscle injury. However, the effects of MENS on unloading-associated atrophied skeletal muscle remain unclear. Effects of MENS on the regrowing process of unloading-associated atrophied skeletal muscle were investigated. Male C57BL/6J mice (10-week old) were randomly assigned to untreated normal recovery (C) and MENS-treated (M) groups. Mice of both groups are subjected to continuous hindlimb suspension (HS) for 2 weeks followed by 7 days of ambulation recovery. Mice in M group were treated with MENS for 60 min 1, 3, and 5 days following HS, respectively, under anesthesia. The intensity, the frequency, and the pulse width of MENS were set at 10 µA, 0.3 Hz, and 250 msec, respectively. Soleus muscles were dissected before and immediately after, 1, 3 and 7 days after HS. Soleus muscle wet weight and protein content were decreased by HS. The regrowth of atrophied soleus muscle in M group was faster than that in C group. Decrease in the reloading-induced necrosis of atrophied soleus was facilitated by MENS. Significant increases in phosphorylated levels of p70 S6 kinase and protein kinase B (Akt) in M group were observed, compared with C group. These observations are consistent with that MENS facilitated regrowth of atrophied soleus muscle. MENS may be a potential extracellular stimulus to activate the intracellular signals involved in protein synthesis.

Skeletal Muscle Denervation: Sciatic and Tibial Nerve Transection Technique.

The nerve transection model is an established and validated experimental model of skeletal muscle atrophy prepared by denervating the skeletal muscle in rodents. While a number of denervation techniques are available in rats, the development of various transgenic and knockout mice has also led to the wide use of mouse models of nerve transection. Skeletal muscle denervation experiments expand our knowledge of the physiological role of nerval activity and/or neurotrophic factors in the plasticity of skeletal muscle. The denervation of the sciatic or tibial nerve is a common experimental procedure in mice and rats, as these nerves can be resected without great difficulty. An increasing number of reports have recently been published on experiments using a tibial nerve transection technique in mice. In this chapter, we demonstrate and explain the procedures used to transect the sciatic and tibial nerves in mice.

Effects of heat stress on muscle mass and the expression levels of heat shock proteins and lysosomal cathepsin L in soleus muscle of young and aged mice.

Effects of heat stress on skeletal muscle mass in young and aged mice were investigated. Young (7-week) and aged (106-week) male C57BL/6J mice were randomly assigned to control and heat-stressed groups in each age. Mice in heat-stressed group were exposed to heat stress (41 °C for 60 min) in an incubator without anesthesia. Seven days after the exposure, soleus muscles were dissected from both hindlimbs. Protein content and the relative composition of Type II fibers in aged soleus were lower than those in young muscle. In aged soleus, higher baseline expression levels of HSP25, HSP72, and cathepsin L were observed compared with those in young muscle (p < 0.05). However, there were no significant differences in the expression levels of phosphorylated p70 S6 kinase (p-p70S6K), calpain 1, and calpain 2 of soleus between two age groups. A significant increase in muscle mass of both age groups was induced by heat stress (p < 0.05). Heat stress also upregulated the expressions of HSP25, HSP72, and p-p70S6K in both ages (p < 0.05). On the other hand, a significant decrease in cathepsin L expression by heating was observed in aged soleus, but not in young (p < 0.05). Both the percentage of Type I fibers and the expression of calpains in both age groups were unchanged following heat stress. Heat stress-associated downregulation of cathepsin L may be attributed to the upregulation of HSP72, which stabilizes lysosomal membranes (p < 0.05). Upregulations of HSP25, HSP72, and p-p70S6K and/or the downregulation of cathepsin L may play a role in heat stress-associated muscle hypertrophy in aged soleus muscle.

Responses of neuromuscular properties to unloading and potential countermeasures during space exploration missions.

We reviewed the responses of the neuromuscular properties of mainly the soleus and possible mechanisms. Sensory nervous activity in response to passive shortening and/or active contraction, associated with plantar-flexion or dorsi-flexion of the ankle joints, may play an essential role in the regulation of muscle properties. Passive shortening of the muscle fibers and sarcomeres inhibits the development of tension, electromyogram (EMG), and afferent neurogram. Remodeling of the sarcomeres, which decreases the total sarcomere number in a single muscle fiber causing recovery of the length in each sarcomere, is induced in the soleus following chronic unloading. Although EMG activity and tension development in each sarcomere are increased, the total tension produced by the whole muscle is still less owing to the lower sarcomere number. Therefore, muscle atrophy continues to progress. Moreover, walking or slow running by rear-foot strike landing with the application of greater ground reaction force, which stimulates soleus mobilization, could be an effective countermeasure. Periodic, but not chronic, passive stretching of the soleus may also be effective.

Methylglyoxal reduces molecular responsiveness to 4 weeks of endurance exercise in mouse plantaris muscle.

Endurance exercise triggers skeletal muscle adaptations, including enhanced insulin signaling, glucose metabolism, and mitochondrial biogenesis. However, exercise-induced skeletal muscle adaptations may not occur in some cases, a condition known as exercise resistance. Methylglyoxal (MG) is a highly reactive dicarbonyl metabolite and has detrimental effects on the body such as causing diabetic complications, mitochondrial dysfunction, and inflammation. This study aimed to clarify the effect of methylglyoxal on skeletal muscle molecular adaptations following endurance exercise. Mice were randomly divided into four groups (n = 12/group): sedentary control group, voluntary exercise group, MG-treated group, and MG-treated with voluntary exercise group. Mice in the voluntary exercise group were housed in a cage with a running wheel, whereas mice in the MG-treated groups received drinking water containing 1% MG. Four weeks of voluntary exercise induced several molecular adaptations in the plantaris muscle, including increased expression of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC1α), mitochondria complex proteins, Toll-like receptor 4 (TLR4), 72-kDa heat shock protein (HSP72), hexokinase II, and glyoxalase 1; this also enhanced insulin-stimulated Akt Ser473 phosphorylation and citrate synthase activity. However, these adaptations were suppressed with MG treatment. In the soleus muscle, the exercise-induced increases in the expression of TLR4, HSP72, and advanced glycation end products receptor 1 were inhibited with MG treatment. These findings suggest that MG is a factor that inhibits endurance exercise-induced molecular responses including mitochondrial adaptations, insulin signaling activation, and the upregulation of several proteins related to mitochondrial biogenesis, glucose handling, and glycation in primarily fast-twitch skeletal muscle.NEW & NOTEWORTHY This study investigated the effect of methylglyoxal, which is a highly reactive carbonyl metabolite and has detrimental effects on the body, on skeletal muscle adaptations following endurance exercise. Evidences from this study show that methylglyoxal is a factor deteriorating responsiveness to endurance exercise in primarily fast-twitch skeletal muscle. The findings contribute to understand the internal factors that should be focused to maximize the exercise effects.

Fine-Tuning of Piezo1 Expression and Activity Ensures Efficient Myoblast Fusion during Skeletal Myogenesis.

Mechanical stimuli, such as stretch and resistance training, are essential in regulating the growth and functioning of skeletal muscles. However, the molecular mechanisms involved in sensing mechanical stress during muscle formation remain unclear. Here, we investigated the role of the mechanosensitive ion channel Piezo1 during myogenic progression of both fast and slow muscle satellite cells. We found that Piezo1 level increases during myogenic differentiation and direct manipulation of Piezo1 in muscle stem cells alters the myogenic progression. Indeed, Piezo1 knockdown suppresses myoblast fusion, leading to smaller myotubes. Such an event is accompanied by significant downregulation of the fusogenic protein Myomaker. In parallel, while Piezo1 knockdown also lowers Ca2+ influx in response to stretch, Piezo1 activation increases Ca2+ influx in response to stretch and enhances myoblasts fusion. These findings may help understand molecular defects present in some muscle diseases. Our study shows that Piezo1 is essential for terminal muscle differentiation acting on myoblast fusion, suggesting that Piezo1 deregulation may have implications in muscle aging and degenerative diseases, including muscular dystrophies and neuromuscular disorders.

Possible role of NF-ĸB signals in heat stress-associated increase in protein content of cultured C2C12 cells.

Heat stress is one of the hypertrophic stimuli on mammalian skeletal muscle. Nuclear factor-κB (NF-κB) signaling plays an important role in the regulation of skeletal muscle mass. However, the effects of heat stress on NF-κB signaling in skeletal muscle cells remain unclear. Effects of heat stress and/or administration of BAY11-7082, an inhibitor of NF-κB, on NF-κB signals and protein content of skeletal muscle were studied by using cell culture system. Differentiated mouse myoblasts (C2C12) were subjected to either (1) control (cultured at 37°C without BAY11-7082), (2) heat stress at 41°C for 60 min, (3) BAY11-7082 administration (1.25 µM) or (4) heat stress combined with BAY11-7082 administration. Heat shock protein 72 (HSP72) was upregulated by heat stress with or without administration of BAY11-7082. The increase in inhibitor of κBα (IκBα), which regulates the phosphorylation of NF-κB, and the decrease in phosphorylated NF-κB were also induced by administration of BAY11-7082 and/or heat stress. Protein content in C2C12 cells was increased by the administration of BAY11-7082 with a semi-logarithm fashion. Significant increases in the protein content of C2C12 cells were observed 48 h following heating with or without administration of BAY11-7082. These observations suggest that heat stress might increase muscle protein through the downregulation of NF-κB signaling. Inhibition of NF-κB induced by application of heat stress might be one of the hypertrophic stimuli on skeletal muscle cells.

Responses of muscle mass, strength and gene transcripts to long-term heat stress in healthy human subjects.

The present study was performed to investigate the effects of long-term heat stress on mass, strength and gene expression profile of human skeletal muscles without exercise training. Eight healthy men were subjected to 10-week application of heat stress, which was performed for the quadriceps muscles for 8 h/day and 4 days/week by using a heat- and steam-generating sheet. Maximum isometric force during knee extension of the heated leg significantly increased after heat stress (~5.8%, P < 0.05). Mean cross-sectional areas (CSAs) of vastus lateralis (VL, ~2.7%) and rectus femoris (~6.1%) muscles, as well as fiber CSA (8.3%) in VL, in the heated leg were also significantly increased (P < 0.05). Statistical analysis of microarrays (SAM) revealed that 10 weeks of heat stress increased the transcript level of 925 genes and decreased that of 1,300 genes, and gene function clustering analysis (Database for Annotation, Visualization and Integrated Discovery: DAVID) showed that these regulated transcripts stemmed from diverse functional categories. Transcript level of ubiquinol-cytochrome c reductase binding protein (UQCRB) was significantly increased by 10 weeks of heat stress (~3.0 folds). UQCRB is classified as one of the oxidative phosphorylation-associated genes, suggesting that heat stress can stimulate ATP synthesis. These results suggested that long-term application of heat stress could be effective in increasing the muscle strength associated with hypertrophy without exercise training.

[A case of laminopathy with the mutation of LMNA gene identified by the exome analysis of disease-related genes].

Laminopathy, caused by mutations in the LMNA gene, include a variety of diseases, such as Emery-Dreifuss muscular dystrophy. A Japanese woman developed progressive muscle weakness, muscle atrophy and joint contractures of upper and lower limbs after the age of two years old. She had restrictive respiratory dysfunction, and developed both supraventricular and ventricular arrhythmias after the fourth decade of life. At 55 years old, she had tracheostomy, required mechanical ventilation and was implanted with the implantable cardioverter defibrillator. The serum level of creatine kinase was within normal range. Electromyography showed polyphasic or large motor unit potentials and reduced interference pattern, while relatively normal recruitment. The exome analysis of disease-related genes revealed a heterozygous pathogenic variant c.1072G>A (p.E358K) in the LMNA gene, which contributed to the diagnosis of laminopathy.

A possible role of NF-kappaB and HSP72 in skeletal muscle hypertrophy induced by heat stress in rats.

Effects of heat stress on phosphorylated nuclear factor-kappaB (phospho-NF-kappaB) and tumor necrosis factor alpha (TNFalpha) contents in skeletal muscles were studied. Male Wistar rats (7-week-old) were randomly assigned to control and heat-stressed groups. Rats in heat-stressed group were exposed to heat stress (42 degrees C for 60 min) in an incubator without anesthesia. Soleus muscles were dissected and weighted 1, 3, and 7 days after the heat exposure. Significant increases in the wet weight and protein content of soleus were observed 7 days following the exposure (p < 0.05). Heat stress also induced the up-regulation of heat shock protein 72 (HSP72), IkappaBalpha (inhibitor of NF-kappaB) and the increase in the relative population of Pax7-positive satellite cells to total muscle nuclei before the increase in muscle mass. The content levels of phospho-NF-kappaB and TNFalpha were significantly decreased 1 and 3 days after heat stress, respectively (p < 0.05). A negative correlation between HSP72 and phospho-NF-kappaB contents was observed 1 day after the heat exposure. These observations suggest that the decrease in NF-kappaB signaling may play a part of a role in heat stress-associated muscle hypertrophy.

[A case of autosomal recessive hypomyelinating leukodystrophy without GJA12 mutation presenting a novel phenotype].

A 50-year-old woman, who had consanguineous parents, developed gait disturbance at age 3, and revealed nystagmus, cerebellar ataxia, peripheral neuropathy, and spastic tetraparesis. She admitted to our hospital at age 14, and the symptoms progressed very slowly. MRI of this case at age 45 showed a remarkable, diffuse hypomyelination of the cerebrum. Her older sister who already deceased at age 16 showed neurological symptoms similar to this case. The patient was found to have no proteolipid protein-1 gene duplications and deletions and base substitution. Her symptoms were considered to be different from those of typical HLD2, 3, 4 and 5. She carried no GJA12 mutations. These facts suggested that this disease is a novel, autosomal recessive hypomyelinating leukodystrophy.