The effect of microgravity on skeletal muscles has so far been examined in rat and mice only after short-term (5–20 day) spaceflights. The mice drawer system (MDS) program, sponsored by Italian Space Agency, for the first time aimed to investigate the consequences of long-term (91 days) exposure to microgravity in mice within the International Space Station. Muscle atrophy was present indistinctly in all fiber types of the slow-twitch soleus muscle, but was only slightly greater than that observed after 20 days of spaceflight. Myosin heavy chain analysis indicated a concomitant slow-to-fast transition of soleus. In addition, spaceflight induced translocation of sarcolemmal nitric oxide synthase-1 (NOS1) into the cytosol in soleus but not in the fast-twitch extensor digitorum longus (EDL) muscle. Most of the sarcolemmal ion channel subunits were up-regulated, more in soleus than EDL, whereas Ca 2+ -activated K + channels were down-regulated, consistent with the phenotype transition. Gene expression of the atrophy-related ubiquitin-ligases was up-regulated in both spaceflown soleus and EDL muscles, whereas autophagy genes were in the control range. Muscle-specific IGF-1 and interleukin-6 were down-regulated in soleus but up-regulated in EDL. Also, various stress-related genes were up-regulated in spaceflown EDL, not in soleus. Altogether, these results suggest that EDL muscle may resist to microgravity-induced atrophy by activating compensatory and protective pathways. Our study shows the extended sensitivity of antigravity soleus muscle after prolonged exposition to microgravity, suggests possible mechanisms accounting for the resistance of EDL, and individuates some molecular targets for the development of countermeasures.

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Effects of heat stress, mechanical stretching or a combination of both on the expression of heat shock proteins (HSPs) and total protein level were studied in a culture system. Rat skeletal muscle cells (L6) were cultured on flexible-bottomed culture plates. They were subjected to one of the four following conditions: (1) 97 h incubation at 37 degrees C, (2) 1 h incubation at 41 degrees C followed by 96 h incubation at 37 degrees C, (3) 1 h incubation at 37 degrees C followed by 96 h cyclic stretching (18% of initial length, 2-s stretch and 4-s release) at 37 degrees C or (4) 1 h incubation at 41 degrees C followed by 96 h cyclic stretching at 37 degrees C. The expression of HSP72 and HSP90 and total protein was determined in the crude homogenates, supernatant and pellets. Cellular protein concentrations in the homogenates and pellets were increased by heat stress and/or mechanical stress (stretch). A cumulative effect of the combination of heating and stretch on the protein concentration in the homogenates and in the pellets was noted. The expressions of HSP72 and HSP90 in the pellets were also increased by heat stress and/or stretch. However, HSP90 in the supernatant did not change following heat stress and/or stretch. The regulation of HSP72 and HSP90 expression in skeletal muscle cells may be closely related to total protein, the abundance of which is also stimulated by mechanical and heat stresses. These observations suggest strongly that heating and passive stretch of muscle may be useful as a means of increasing muscle mass, not only in athletes but also in patients during rehabilitation.

. Changes in PKB/Akt and calcineurin signaling during recovery in atrophied soleus muscle induced by unloading.

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.

Abstract:The effects of heat-stress on proliferative potential in vivo were studied in rat skeletal muscle. Male Wistar rats (7-weeks-old) were divided into two groups: control (n = 24) and heatstressed (n = 24). Rats in the experimental group were exposed to environmental heat-stress (41°C for 60 min) in a heat chamber without anesthesia. The soleus muscles were dissected 1, 7, and 14 days after the heat exposure. The wet and dry weights of soleus muscle relative to body weight in the heat-stressed group were significantly higher than controls 7 days after the exposure (10.1% and 17.5%, respectively, p < 0.05). The distribution of 5-bromo-2′-deoxyuridine and proliferating cell nuclear antigen-positive nuclei, that are the indicators for the cell proliferation, were increased by 2.2 and 5.1 times, respectively 1 day after heating (p < 0.05). The expressions of heat shock protein 72 (58.0%) and phosphorylated p70S6 kinase (52.3%) were increased 1 day following heat exposure (p < 0.05). These results suggest that heat-stress could promote the cell proliferation and induce muscular hypertrophy.

Results suggested that loading plays an important role in the activation of the regenerating potential of injured skeletal muscle.

induced activation of AMPK negatively regulates myotube hypertrophy through the HSP72-mediated pathway in C2C12 skeletal muscle cells. Am J Physiol Endocrinol Metab 306: E344 -E354, 2014. First published December 17, 2013; doi:10.1152/ajpendo.00495.2013.-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 AICARtreated myotubes in those without treatment. AICAR-induced suppression of myotube hypertrophy and HSP72 expression was attenuated in the siRNA-mediated AMPK␣ knockdown myotubes. AICAR increased microRNA (miR)-1, a modulator of HSP72, and the increase of miR-1 was not induced in AMPK␣ knockdown condition. Furthermore, siRNA-mediated HSP72 knockdown blocked AICARinduced inhibition of myotube hypertrophy. AICAR upregulated the gene expression of muscle Ring-finger 1, and this alteration was suppressed in either AMPK␣ or HSP72 knockdown myotubes. The phosphorylation of p70 S6 kinase Thr 389 was downregulated by AICAR, whereas this was attenuated in AMPK␣, 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. microRNA; heat shock transcription factor 1; muscle rRing-finger 1; atrogin-1; acetyl-CoA carboxylase SKELETAL MUSCLE HAS A GREATER CAPACITY to adapt to various stimuli. Increased loading, such as resistance training and mechanical stretching, stimulates protein synthesis and reduces protein degradation, thereby inducing muscle hypertrophy (10). On the other hand, decrease of use, such as immobilization, denervation, aging, and/or various pathological conditions, attenuates protein synthesis and increases protein degradation, resulting in atrophy (12,32). Although the process of skeletal muscle adaptation to hypertrophic and atrophic stimuli has been studied, the molecular mechanism involved in this process is not fully understood yet.5=-AMP-activated protein kinase (AMPK) is well known as a sensor for cellular energy status and metabolic stress, such as muscle contraction, fasting, hypoxia, ischemia, and/or oxidative and osmotic stresses and as a signaling intermediary that controls the use of glucose and fatty acids in skeletal muscle (8,14,15). In addition, several studies in the past decade have suggested that AMPK plays an important rol...