Repeated bouts of resistance exercise in rats alter mechanistic target of rapamycin complex 1 activity and ribosomal capacity but not muscle protein synthesis.

NEW FINDINGS: What is the central question of this study? Is muscle protein synthesis (MPS) additionally activated following exercise when ribosomal capacity is increased after repeated bouts of resistance exercise (RE)? What is the main finding and its importance? Skeletal muscles with increased ribosome content through repeated RE bouts showed sufficient activation of MPS with lower mechanistic target of rapamycin complex 1 signalling. Thus, repeated bouts of RE possibly change the translational capacity and efficiency to optimize translation activation following RE. ABSTRACT: Resistance exercise (RE) activates ribosome biogenesis and increases ribosome content in skeletal muscles. However, it is unclear whether the increase in ribosome content subsequently causes an increase in RE-induced activation of muscle protein synthesis (MPS). Thus, this study aimed to investigate the relationship between ribosome content and MPS after exercise using a rat RE model. Male Sprague-Dawley rats were categorized into three groups (n = 6 for each group): sedentary (SED) and RE trained with one bout (1B) or three bouts (3B). The RE stimulus was applied to the right gastrocnemius muscle by transcutaneous electrical stimulation under isoflurane anaesthesia. The 3B group underwent stimulation every other day. Our results revealed that 6 h after the last bout of RE, muscles in the 3B group showed an increase in total RNA and 18S+28S rRNA content per muscle weight compared with the SED and 1B groups. In both the 1B and 3B groups, MPS, estimated by puromycin incorporation in proteins, was higher than that in the SED group 6 h after exercise; however, no significant difference was observed between the 1B and 3B groups. In the 1B and 3B groups, phosphorylated p70S6K at Thr-389 increased, indicating mechanistic target of rapamycin complex 1 (mTORC1) activity. p70S6K phosphorylation level was lower in the 3B group than in the 1B group. Finally, protein synthesis per ribosome (indicator of translation efficiency) was lower in the 3B group than in the 1B group. Thus, three bouts of RE changed the ribosome content and mTORC1 activation, but not the degree of RE-induced global MPS activation.

Repeated bouts of resistance exercise attenuate mitogen-activated protein-kinase signal responses in rat skeletal muscle.

Resistance exercise training induces skeletal muscle hypertrophy, but repeated bouts gradually attenuate this anabolic effect. Attenuation of mechanistic target of rapamycin complex 1 (mTORC1) activation by repetitive resistance exercise is involved in this process, but the mechanism leading to inactivation of mTORC1 remains unclear. In this study, we investigated repetition-dependent changes in mitogen-activated protein kinases (MAPKs) and the 90-kDa ribosomal S6 kinase (p90RSK), upstream regulators of mTORC1, in a rat resistance-exercise model. Resistance exercise was associated with increased phosphorylation of 70-kDa ribosomal protein S6 kinase (Thr389), but its magnitude was decreased with repeated bouts. Additionally, phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 (Thr202/Tyr204) and p38 MAPK (Thr180/Tyr182), which are MAPKs, decreased with repeated bouts. A similar result was also observed for p90RSK phosphorylation (Thr573). These observations indicate that repeated bouts desensitized ERK1/2 and p38 MAPK, subsequently attenuating p90RSK phosphorylation. This reduction in p90RSK phosphorylation may have been partly responsible for the blunting of mTORC1 activation by resistance exercise.

Intramuscular injection of mesenchymal stem cells augments basal muscle protein synthesis after bouts of resistance exercise in male mice.

Skeletal muscle mass is critical for activities of daily living. Resistance training maintains or increases muscle mass, and various strategies maximize the training adaptation. Mesenchymal stem cells (MSCs) are multipotent cells with differential potency in skeletal muscle cells and the capacity to secrete growth factors. However, little is known regarding the effect of intramuscular injection of MSCs on basal muscle protein synthesis and catabolic systems after resistance training. Here, we measured changes in basal muscle protein synthesis, the ubiquitin-proteasome system, and autophagy-lysosome system-related factors after bouts of resistance exercise by intramuscular injection of MSCs. Mice performed three bouts of resistance exercise (each consisting of 50 maximal isometric contractions elicited by electrical stimulation) on the right gastrocnemius muscle every 48 h, and immediately after the first bout, mice were intramuscularly injected with either MSCs (2.0 × 106 cells) labeled with green fluorescence protein (GFP) or vehicle only placebo. Seventy-two hours after the third exercise bout, GFP was detected only in the muscle injected with MSCs with concomitant elevation of muscle protein synthesis. The injection of MSCs also increased protein ubiquitination. These results suggest that the intramuscular injection of MSCs augmented muscle protein turnover at the basal state after consecutive resistance exercise.

Cooling-promoted myogenic differentiation of murine bone marrow mesenchymal stem cells through TRPM8 activation in vitro.

TRPM8 agonist has been reported to promote osteogenic differentiation of mesenchymal stem cells (MSCs), therefore we evaluated whether cooling-induced activation of TRPM8 promotes myogenic differentiation of MSCs. We used 5-azacytidine as a myogenic differentiation inducer in murine bone marrow-derived MSCs. Addition of menthol, a TRPM8 agonist, to the differentiation induction medium significantly, increased the percentage of MyoD-positive cells, a specific marker of myogenic differentiation. We performed intracellular Ca2+ imaging experiments using fura-2 to confirm TRPM8 activation by cooling stimulation. The results confirmed that intracellular Ca2+ concentration ([Ca2+ ]i) increases due to TRPM8 activation, and TRPM8 antagonist inhibits increase in [Ca2+ ]i at medium temperatures below 19°C. We also examined the effect of cooling exposure time on myogenic differentiation of MSCs using an external cooling stimulus set at 17°C. The results showed that 60 min of cooling had an acceleratory effect on differentiation (2.18 ± 0.27 times). We observed that the TRPM8 antagonist counteracted the differentiation-promoting effect of the cooling. These results suggest that TRPM8 might modulate the multiple differentiation pathways of MSCs, and that cooling is an effective way of activating TRPM8, which regulates MSCs differentiation in vitro.

Resistance training rejuvenates aging skin by reducing circulating inflammatory factors and enhancing dermal extracellular matrices.

Aerobic training (AT) is suggested to be an effective anti-aging strategy for skin aging. However, the respective effects of resistance training (RT) have not been studied. Therefore, we compared the effects of AT and RT on skin aging in a 16-week intervention in 61 healthy sedentary middle-aged Japanese women. Data from 56 women were available for analysis. Both interventions significantly improved skin elasticity and upper dermal structure, and RT also improved dermal thickness. After the training intervention, expression of dermal extracellular matrix-related genes was increased in normal human primary dermal fibroblasts. AT and RT had different effects on circulating levels of factors, such as cytokines, hormones in serum, and metabolites, and RT increased dermal biglycan (BGN). To our knowledge, this is the first report to show different effects of AT and RT on skin aging and identify the key factors involved in RT-induced skin rejuvenation.

Relationship between protein intake and resistance training-induced muscle hypertrophy in middle-aged women: A pilot study.

OBJECTIVE: The aim of this study was to observe the relationship of protein intake at each meal and daily total with change in lean tissue mass with progressive resistance exercise training (RET) in healthy middle-aged women. METHODS: Twenty-two healthy Japanese women were recruited from Shiga Prefecture, Japan, and a supervised whole body RET program was conducted twice a week for 16 wk. The dietary intake was assessed using 3-d dietary records. Dual-energy x-ray absorptiometry was used to measure the whole body lean soft tissue mass (WLTM). Multiple regression analysis was performed to examine the relationship between the protein intake and RET-induced changes in the WLTM after adjusting for age, sleep quality, physical activity, and energy intake. RESULTS: The 16-wk RET program caused a significant gain in the WLTM (1.46 ± 0.45%, P = 0.004). Multiple regression analysis showed that the baseline protein intake at breakfast was negatively associated with the percent change in the WLTM (ß = -1.598; P = 0.022). Additionally, the percent change (ß = 0.624; P = 0.018) in protein intake at breakfast was positively associated with the percent change in WLTM. CONCLUSION: Increasing protein intake at breakfast may contribute to RET-induced muscle hypertrophy in middle-aged women, especially among those who habitually consume low-protein levels at breakfast. However, future studies with larger sample sizes are required to confirm the importance of protein intake at breakfast.

Magnetization of mesenchymal stem cells using magnetic liposomes enhances their retention and immunomodulatory efficacy in mouse inflamed skeletal muscle.

Sarcopenia, an age-related reduction in skeletal muscle mass and strength, is mainly caused by chronic inflammation. Because mesenchymal stem cells (MSCs) have the capacity to both promote myogenic cell differentiation and suppress inflammation, they are a promising candidate for sarcopenia treatment. In this study, to achieve the long-term retention of MSCs in skeletal muscle, we prepared magnetized MSCs using magnetic anionic liposome/atelocollagen complexes that we had previously developed, and evaluated their retention efficiency and immunomodulatory effects in mouse inflamed skeletal muscle. Mouse MSCs were efficiently magnetized by incubation with magnetic anionic liposome/atelocollagen complexes for 30 min under a magnetic field. The magnetized MSCs differentiated normally into osteoblasts and adipocytes. Additionally, non-magnetized MSCs and magnetized MSCs increased IL-6 and inducible nitric oxide synthase mRNA expression and decreased TNF-α and IL-1ß mRNA expression in C2C12 mouse skeletal muscle myotubes through paracrine effects. Moreover, magnetized MSCs were significantly retained in cell culture plates and mouse skeletal muscle after their local injection in the presence of a magnetic field. Furthermore, magnetized MSCs significantly increased IL-6 and IL-10 mRNA expression and decreased TNF-α and IL-1ß mRNA expression in inflamed skeletal muscle. These results suggest that magnetized MSCs may be useful for effective sarcopenia treatment.

Intramuscular injection of mesenchymal stem cells activates anabolic and catabolic systems in mouse skeletal muscle.

Skeletal muscle mass is critical for good quality of life. Mesenchymal stem cells (MSCs) are multipotent stem cells distributed across various tissues. They are characterized by the capacity to secrete growth factors and differentiate into skeletal muscle cells. These capabilities suggest that MSCs might be beneficial for muscle growth. Nevertheless, little is known regarding the effects on muscle protein anabolic and catabolic systems of intramuscular injection of MSCs into skeletal muscle. Therefore, in the present study, we measured changes in mechanistic target of rapamycin complex 1 (mTORC1) signaling, the ubiquitin-proteasome system, and autophagy-lysosome system-related factors after a single intramuscular injection of MSCs with green fluorescence protein (GFP) into mouse muscles. The intramuscularly-injected MSCs were retained in the gastrocnemius muscle for 7 days after the injection, indicated by detection of GFP and expression of platelet-derived growth factor receptor-alpha. The injection of MSCs increased the expression of satellite cell-related genes, activated mTORC1 signaling and muscle protein synthesis, and increased protein ubiquitination and autophagosome formation (indicated by the expression of microtubule-associated protein 1 light chain 3-II). These results suggest that the intramuscular injection of MSCs activated muscle anabolic and catabolic systems and accelerated muscle protein turnover.

The effect of repeated bouts of electrical stimulation-induced muscle contractions on proteolytic signaling in rat skeletal muscle.

Mechanistic target of rapamycin complex 1 (mTORC1) plays a central role in muscle protein synthesis and repeated bouts of resistance exercise (RE) blunt mTORC1 activation. However, the changes in the proteolytic signaling when recurrent RE bouts attenuate mTORC1 activation are unclear. Using a RE model of electrically stimulated rat skeletal muscle, this study aimed to clarify the effect of repeated RE bouts on acute proteolytic signaling, particularly the calpain, autophagy-lysosome, and ubiquitin-proteasome pathway. p70S6K and rpS6 phosphorylation, indicators of mTORC1 activity, were attenuated by repeated RE bouts. Calpain 3 protein was decreased at 6 h post-RE in all exercised groups regardless of the bout number. Microtubule-associated protein 1 light chain 3 beta-II, an indicator of autophagosome formation, was increased at 3 h and repeated RE bouts increased at 6 h, post-RE. Ubiquitinated proteins were increased following RE, but these increases were independent of the number of RE bouts. These results suggest that the magnitude of autophagosome formation was increased following RE when mTORC1 activity was attenuated with repeated bouts of RE.

The distribution of eukaryotic initiation factor 4E after bouts of resistance exercise is altered by shortening of recovery periods.

Insufficient duration of recovery between resistance exercise bouts reduces the effects of exercise training, but the influence on muscle anabolic responses is not fully understood. Here, we investigated the changes in the distribution of eukaryotic initiation factor (eIF) 4E, a key regulator of translation initiation, and related factors in mouse skeletal muscle after three successive bouts of resistance exercise with three durations of recovery periods (72 h: conventional, 24 h: shorter, and 8 h: excessively shorter). Bouts of resistance exercise dissociated eIF4E from eIF4E binding protein 1, with the magnitude increasing with shorter recovery. Whereas bouts of resistance exercise with 72 h recovery increased the association of eIF4E and eIF4G, those with shorter recovery did not. Similar results were observed in muscle protein synthesis. These results suggest that insufficient recovery inhibited the association of eIF4E and eIF4G, which might cause attenuation of protein synthesis activation after bouts of resistance exercise.

Apple polyphenols induce browning of white adipose tissue.

We and others have shown that apple polyphenols decrease adipose tissue mass. To better understand the underlying mechanisms and to expand clinical applicability, we herein examine whether apple polyphenols induce adipose thermogenic adaptations (browning) and prevent diet-induced obesity and related insulin resistance. In mice fed a standard diet, daily apple polyphenol consumption induced thermogenic adaptations in inguinal white adipose tissue (iWAT), based on increases in the expression of brown/beige adipocyte selective genes (Ucp1, Cidea, Tbx1, Cd137) and protein content of uncoupling protein 1 and mitochondrial oxidative phosphorylation enzymes. Among the upstream regulatory factors of browning, fibroblast growth factor 21 (FGF21) and peroxisome proliferator-activated receptor gamma coactivator 1 α (PGC-1α) levels were concomitantly up-regulated by apple polyphenols. In the primary cell culture experiment, the results did not support a direct action of apple polyphenols on beige adipogenesis. Instead, apple polyphenols increased tyrosine hydroxylase (a rate-limiting enzyme of catecholamine synthesis) in iWAT, which activates the adipocyte thermogenic program possibly via intratissue cellular communications. In high-fat fed mice, apple polyphenols induced beige adipocyte development in iWAT, reduced fat accumulation, and increased glucose disposal rates in the glucose and insulin tolerance tests. Taken together, dietary administration of apple polyphenols induced beige adipocyte development in iWAT possibly via activation/induction of the peripheral catecholamine synthesis-FGF21-PGC-1α cascade. Results from diet-induced obese mice indicate that apple polyphenols have therapeutic potential for obesity and related metabolic disorders.

The Effect of Leucine-Enriched Essential Amino Acid Supplementation on Anabolic and Catabolic Signaling in Human Skeletal Muscle after Acute Resistance Exercise: A Randomized, Double-Blind, Placebo-Controlled, Parallel-Group Comparison Trial.

Resistance exercise transiently activates anabolic and catabolic systems in skeletal muscle. Leucine-enriched essential amino acids (LEAAs) are reported to stimulate the muscle anabolic response at a lower dose than whey protein. However, little is known regarding the effect of LEAA supplementation on the resistance exercise-induced responses of the anabolic and catabolic systems. Here, we conducted a randomized, double-blind, placebo-controlled, parallel-group comparison trial to investigate the effect of LEAA supplementation on mechanistic target of rapamycin complex 1 (mTORC1), the ubiquitin-proteasome system and inflammatory cytokines after a single bout of resistance exercise in young men. A total of 20 healthy young male subjects were supplemented with either 5 g of LEAA or placebo, and then they performed 10 reps in three sets of leg extensions and leg curls (70% one-repetition maximum). LEAA supplementation augmented the phosphorylation of mTORSer2448 (+77.1%, p < 0.05), p70S6KThr389 (+1067.4%, p < 0.05), rpS6Ser240/244 (+171.3%, p < 0.05) and 4EBP1Thr37/46 (+33.4%, p < 0.05) after resistance exercise. However, LEAA supplementation did not change the response of the ubiquitinated proteins, MuRF-1 and Atrogin-1 expression. Additionally, the mRNA expression of IL-1ß and IL-6 did not change. These data indicated that LEAA supplementation augments the effect of resistance exercise by enhancing mTORC1 signal activation after exercise.

Consecutive bouts of electrical stimulation-induced contractions alter ribosome biogenesis in rat skeletal muscle.

Ribosome biogenesis has been implicated in resistance exercise training (RET)-induced skeletal muscle hypertrophy. However, it is unclear how increasing bouts of RET affects ribosome content and biogenesis. This was investigated in the present study using simulated RET where rat skeletal muscle is subjected to increasing bouts of electrical stimulation. Sprague-Dawley rats were randomly assigned to the following seven groups: sedentary for 5 days (SED) or 6 wk (SED_6w), resistance-exercise trained with 1 bout (1B), 2 bouts (2B), 3 bouts (3B), 6 bouts (6B), and 18 bouts (18B). RET was simulated on the right gastrocnemius muscle by transcutaneous electric stimulation under isoflurane anesthesia, and a RET bout was given 3 times a week. Rats in 1B, 2B, and 3B groups showed increased 45S precursor (pre-) rRNA and 18S+28S rRNA content per muscle weight and elevated mRNA levels of c-myc and upstream binding factor (UBF). Increases in phosphorylated UBF and total cyclin D1 protein level were observed 48 h after RET; the former increased as a function of RET duration. In 3B, 6B, and 18B groups, the 18S+28S rRNA content per muscle weight was kept unchanged, and 45S pre-rRNA, cyclin D1, and phosphorylated UBF levels in 18B were lower than those in 3B. These results suggest that RET activates ribosome biogenesis and increases ribosome content through modulation of UBF and cyclin D1 activity at its early phase. Additional bouts of RET may not lead to a further increase in ribosome content per muscle weight through possibly the attenuation of transcription process. NEW & NOTEWORTHY Ribosome biogenesis has been implicated in resistance exercise training-induced skeletal muscle hypertrophy. However, it remains unclear how this is influenced by the volume of repeated bouts of resistance exercise training. Using resistance exercise training model with rat skeletal muscle, we provide evidence that ribosome biogenesis is stimulated by the initial few bouts of resistance exercise training with no additional effect of further increase in the exercise bout.

Influence of shortened recovery between resistance exercise sessions on muscle-hypertrophic effect in rat skeletal muscle.

Resistance exercise training induces muscle hypertrophy, and recovery between sessions is one of the major determinants of this effect. However, the effect of the recovery period between sessions on muscle hypertrophy following resistance exercise training remains unclear. To elucidate the effect of recovery period on hypertrophy, in the present study, we investigated changes in protein degradation systems and hypertrophic responses in rat skeletal muscle to resistance training with variable recovery periods. In the conventional recovery group (exercised every 72 h) and a shorter recovery group (exercised every 24 h), 18 bouts of resistance exercise consisting of 50 repetitions of a 3-sec maximal isometric contraction caused muscle hypertrophy and slight activation of muscle protein degradation systems. By contrast, in an excessively shorter recovery group (exercised every 8 h), 18 bouts of resistance exercise did not cause hypertrophy and markedly activated protein degradation systems, accompanied by inflammatory responses. These observations indicate that excessive shortening of recovery between sessions does not cause skeletal muscle hypertrophy, likely due to the activation of proteolysis induced by inflammatory responses to resistance exercise training.

Influence of past injurious exercise on fiber type-specific acute anabolic response to resistance exercise in skeletal muscle.

We investigated the influence of past injurious exercise on anabolic response of skeletal muscle fibers to resistance exercise (RE). Wistar rats were divided into exercise (E) and exercise-after-injury (I-E) groups. At age 10 wk, the right gastrocnemius muscle in each rat in the I-E group was subjected to strenuous eccentric contractions. Subsequently, RE was imposed on the same muscle of each rat at 14 wk of age in both groups. Peak joint torque and total force generation per body mass during RE were similar between the groups. Muscle protein synthesis (MPS) in the I-E group was higher than that in the E group 6 h after RE. Furthermore, levels of phospho-p70S6 kinase (Thr389) and phospho-ribosomal protein S6 (phospho-rpS6) (Ser240/244), a downstream target of p70S6 kinase, were higher in the I-E group than in the E group. For the anabolic response in each fiber type, the I-E group showed a higher MPS response in type IIb, IIa, and I fibers and a higher phospho-rpS6 response in type IIx, IIa, and I fibers than the E group. In the I-E group, the relative content of myosin heavy chain (MHC) IIa was higher and that of MHC IIb was lower than those in the E group. In addition, type IIa fibers showed a lower MPS response to RE than type IIb fibers in the I-E group. In conclusion, the past injurious exercise enhanced the MPS and phospho-rpS6 responses in type IIb, IIa, and I fibers and type IIx, IIa, and I fibers, respectively. NEW & NOTEWORTHY Past injurious exercise increased the muscle protein synthesis (MPS) response and mammalian target of rapamycin complex 1 (mTORC1) signaling activation to resistance exercise. In the responses of each fiber type, the past injurious exercise increased the MPS and phosphorylation ribosomal protein (Ser240/244) responses in type IIb, IIa, and I fibers and type IIx, IIa, and I fibers, respectively.

Past injurious exercise attenuates activation of primary calcium-dependent injury pathways in skeletal muscle during subsequent exercise.

Past contraction-induced skeletal muscle injury reduces the degree of subsequent injury; this phenomenon is called the "repeated bout effect (RBE)." This study addresses the mechanisms underlying the RBE, focusing on primary calcium-dependent injury pathways. Wistar rats were subdivided into single injury (SI) and repeated injury (RI) groups. At age 10 weeks, the right gastrocnemius muscle in each rat in the RI group was subjected to strenuous eccentric contractions (ECs). Subsequently, mild ECs were imposed on the same muscle of each rat at 14 weeks of age in both groups. One day after the exercise, the RI group showed a lower strength deficit than did the SI group, and neither group manifested any increase in membrane permeability. The concentration of protein carbonyls and activation of total calpain increased after ECs given at the age of 14 weeks. Nonetheless, these increases were lower in the RI group than in the SI group. Furthermore, calcium-dependent autolysis of calpain-1 and calpain-3 in the RI group was diminished as compared with that in the SI group. Although peak ankle joint torque and total force generation during ECs at the age of 14 weeks were similar between the two groups, phosphorylation of JNK (Thr183 /Tyr185 ), an indicator of mechanical stress applied to a muscle, was lower in the RI group than in the SI group. These findings suggest that activation of the primary calcium-dependent injury pathways is attenuated by past injurious exercise, and mechanical stress applied to muscle fibers during ECs may decrease in the RBE.

Relationship between exercise volume and muscle protein synthesis in a rat model of resistance exercise.

Resistance exercise (RE) volume is recognized as an important factor that stimulates muscle protein synthesis (MPS) and is considered, at least in part, to be involved in the mammalian target of rapamycin complex 1 (mTORC1)-associated signaling. However, the effects of relatively high-volume RE on mTORC1 and MPS remain unclear. In the present study, we used an animal model of RE to investigate the relationship between RE volume and MPS. Male Sprague-Dawley rats were subjected to RE, and muscle samples were obtained 6 h after performing 1, 3, 5, 10, or 20 sets of RE. Although 1 set of RE did not increase MPS [measured by the surface sensing of translation (SUnSET) method], multiple sets (3, 5, 10, and 20 sets) significantly increased MPS. However, the increase in MPS reached a plateau after 3 or 5 sets of RE, and no further increase in MPS was observed with additional RE sets. In contrast to the MPS response, we observed that p70S6K phosphorylation at Thr389, a marker of mTORC1 activity, and Ser240/244 phosphorylation of rpS6, a downstream target of p70S6K, gradually increased with higher RE volume. The above results suggest that the relationship between RE volume and MPS was not linear. Thus the increase in MPS with increasing RE volume saturates before p70S6K phosphorylation, suggesting a threshold effect for the relationship between p70S6K activation and MPS.NEW & NOTEWORTHY The aim of this study was to investigate the relationship between resistance exercise (RE) volume and muscle protein synthesis. We found that the relationship between RE volume and p70S6K phosphorylation was almost linear, but the increase in muscle protein synthesis began to plateau after approximately five sets of RE.

Repeated bouts of resistance exercise with short recovery periods activates mTOR signaling, but not protein synthesis, in mouse skeletal muscle.

The recovery period between bouts of exercise is one of the major factors influencing the effects of resistance exercise, in addition to exercise intensity and volume. However, the effects of shortening the recovery time between bouts of resistance exercise on subsequent protein synthesis remain unclear. In this study, we investigated the consequences of shortening the recovery time between bouts of resistance exercise on protein synthesis and related processes in mouse skeletal muscles. Eighteen male C57BL/6J mice were randomly subjected to three bouts of resistance exercise with 72 (72H), 24 (24H), or 8 h (8H) of recovery periods between bouts. Resistance exercise, consisting of five sets of 3 s × 10 isometric contractions with 3 min rest between sets, was elicited on the right tibialis anterior muscle via percutaneous electrical stimulation on the deep peroneal nerve under isoflurane anesthesia. The left muscle served as an internal control. Six hours after the third bout of exercise, protein synthesis was found to be activated in the 72H and 24H groups, but not in the 8H group. Phosphorylation of p70S6K at Thr 389, a marker of mammalian target of rapamycin (mTOR) signaling, was increased in all groups, with the 8H group showing the highest magnitude. In contrast, protein carbonylation was observed only in mice in the 8H group. These results suggest that repeated bouts of resistance exercise with 8 h of recovery periods do not effectively increase the levels of muscle protein synthesis despite activation of the mTOR signaling pathway, which likely involves oxidative stress.

Preventive effects of electrical stimulation on inflammation-induced muscle mitochondrial dysfunction.

Cachexia is a complex metabolic syndrome associated with underlying chronic diseases and is characterized by the overexpression of pro-inflammatory cytokines such as tumor necrosis factor alpha (TNF-α), which impair muscle oxidative metabolism. We hypothesized that electrical stimulation (ES) would prevent decrement in muscle oxidative metabolism by suppressing the phosphorylation of p38 MAPK, a critical regulator of inflammatory response. Therefore, the purpose of the present study was to verify the effects of ES on inflammatory-induced decrement of oxidative metabolism in mice tibialis anterior muscles. ICR mice were randomly divided into three groups: control, lipopolysaccharide (LPS) injection for 4days, and LPS injection plus ES (LPS+ES). Cachexia was induced in the animals in the LPS groups via LPS injection (10mg/kg body weight/day, i.p.) during the intervention period. The animals in the LPS+ES group were stimulated electrically (carrier frequency, 2500Hz; modulation frequency, 100Hz; duration, 240s/day; type of contraction, isometric) during the intervention period. LPS injection resulted in decreased body and muscle wet weight and increased expression of TNF-α in plasma and skeletal muscle. In addition, LPS injection decreased indicators of mitochondrial function such as succinate dehydrogenase (SDH) and citrate synthase (CS) activity as well as the expression of PGC-1ɑ, and increased the phosphorylation of p38 MAPK. On the other hand, the intervention of ES attenuated the changes in muscle wet weight, SDH activity, CS activity, p38 MAPK, and PGC-1ɑ. These results suggest that ES could prevent decrement in muscle oxidative metabolism induced by pro-inflammatory cytokines in cachexia.