Satoru Ato

Skeletal muscle mass and force decline with age, and the loss of muscle force precedes muscle atrophy. However, the underlying mechanisms remain unclear. Here, we investigated the role of the myosin co‐chaperone, uncoordinated mutant number‐45 myosin chaperone B (UNC45B), in regulating muscle mass and force. UNC45B expression decreased in mouse gastrocnemius muscle with age, particularly at 24 months old, and adeno‐associated virus vector‐mediated knockdown of Unc45b in 3‐month‐old mouse triceps surae muscle first reduced plantar flexor torque and then decreased gastrocnemius muscle mass. In addition, Unc45b knockdown in the triceps surae muscle resulted in lower bone mineral density. While maximum Ca ²⁺ ‐activated force in mechanically skinned fibers was not affected by Unc45b knockdown, Unc45b knockdown decreased the ratio of depolarization‐induced force to the maximum Ca ²⁺ ‐activated force. We established tamoxifen‐inducible skeletal muscle‐specific Unc45b knockout ( Unc45b imKO) mice to investigate whether the muscle atrophy and weakness due to the loss of Unc45b impacts metabolism and behavior. We found that Unc45b imKO reduced muscle mass and force at a whole‐body level, but did not influence systemic glucose tolerance, insulin sensitivity, or the respiratory exchange ratio. However, Unc45b imKO mice reduced the amount of deeper non‐rapid eye movement sleep, locomotor activity, and body temperature during the sleep phase. We conclude that UNC45B is essential for maintaining fast‐twitch muscle mass and muscle force. In addition, Unc45b deficiency‐mediated muscle loss is also associated with bone fragility, decreased body temperature, and impaired sleep quality.

Muscle contraction activates mTOR signaling, resulting in increased protein synthesis and muscle hypertrophy. Rapamycin-sensitive mTORC1 is important for cap-dependent translation, but the effects of suppressing mTORC1 function using rapamycin on myofibrillar protein synthesis caused by contraction remains unclear. We observed that the eIF4F complex is a translation initiator induced by contraction dependently on rapamycin-sensitive mTORC1. Myofibrillar protein translation increased by muscle contraction was insensitive to rapamycin.

Emerging evidence suggests the potential of rapamycin, an antibiotic from Streptomyces hygroscopicus that functions as a mechanistic target of rapamycin (mTOR) inhibitor, as a mimetic of caloric restriction (CR) for maintaining skeletal muscle health. Several studies showed that rapamycin administration (RAP) reduced appetite and energy intake. However, the physiological and molecular differences between RAP and CR in skeletal muscle are not fully understood. Here we observed the effects of 4 weeks of RAP administration and CR corresponding to the reduction in energy intake produced by RAP administration (PF, paired feeding) on fast glycolytic and slow oxidative muscle in young adult rats. We found that 4 weeks of RAP demonstrated low fast-glycolytic muscle mass with smaller type I and IIb/x myofiber size independent of the energy intake. In addition, PF improved the contractile function of the plantar flexor muscle, whereas RAP did not improve its function. The suppressing response of mTORC1 signaling to RAP is greater in slow-oxidative muscles than in fast-glycolytic muscles. In addition, systemic glucose tolerance was exacerbated by RAP, with reduced expression of Rictor and hexokinase in skeletal muscle. These observations imply that RAP may have a slight but significant negative impact and it obviously different to CR in young adult skeletal muscle.

The relationship between mechanistic target of rapamycin complex 1 (mTORC1) activation after resistance exercise and acetylcholine receptor (AChR) subunit gene expression remains largely unknown. Therefore, we aimed to investigate the effect of electrical stimulation‐induced intense muscle contraction, which mimics acute resistance exercise, on the mRNA expression of AChR genes and the signalling pathways involved in neuromuscular junction (NMJ) maintenance, such as mTORC1 and muscle‐specific kinase (MuSK). The gastrocnemius muscle of male adult Sprague–Dawley rats was isometrically exercised. Upon completion of muscle contraction, the rats were euthanized in the early (after 0, 1, 3, 6 or 24 h) and late (after 48 or 72 h) recovery phases and the gastrocnemius muscles were removed. Non‐exercised control animals were euthanized in the basal state (control group). In the early recovery phase, Agrn gene expression increased whereas LRP4 decreased without any change in the protein and gene expression of AChR gene subunits. In the late recovery phase, Agrn, Musk, Chrnb1, Chrnd and Chrne gene expression were altered and agrin and MuSK protein expression increased. Moreover, mTORC1 and protein kinase B/Akt‐histone deacetylase 4 (HDAC) were activated in the early phase but not in the late recovery phase. Furthermore, rapamycin, an inhibitor of mTORC1, did not disturb changes in AChR subunit gene expression after muscle contraction. However, rapamycin addition slightly increased AChR gene expression, while insulin did not impact it in rat L6 myotube. These results suggest that changes in the AChR subunits after muscle contraction are independent of the rapamycin‐sensitive mTORC1 pathway.

The purpose of this study was to investigate whether aging alters the effect of nutritional status on contraction-induced muscle protein metabolism. In an overnight fasted or fed states, the right gastrocnemius muscle of young (3 months) and aged (24 months) male C57BL/6J mice was isometrically contracted via percutaneous electrical stimulation. The left gastrocnemius muscle served as a control. In the fasted state, there were no differences in basal or contraction-induced muscle protein synthesis between young and old mice. However, in the fed state, basal muscle protein synthesis was greater in young mice, and contraction increased muscle protein synthesis only in young mice. In the fed state, although phosphorylation of 4E-BP1 was similarly increased by contraction in both ages, the increase in phosphorylation of p70S6K was greater in young mice. Our results indicate that aging impairs the ability to integrate signals from muscle contraction and nutrition, leading to aging-induced anabolic resistance to muscle contraction in the postprandial state.

Angiogenesis and muscle satellite cell (SC)-mediated myonuclear accretion are considered essential for the robust response of contraction-induced muscle hypertrophy. Moreover, both myonucleus and SCs are physically adjacent to capillaries and are the major sites for the expression of proangiogenic factors, such as VEGF, in the skeletal muscle. Thus, events involving the addition of new myonuclei via activation of SCs may play an important role in angiogenesis during muscle hypertrophy. However, the relevance among myonuclei number, capillary supply, and angiogenesis factor is not demonstrated. The Notch effector HeyL is specifically expressed in SCs in the skeletal muscle and is crucial for SC proliferation by inhibiting MyoD in overload-induced muscle hypertrophy. Here, we tested whether the addition of new myonuclei by SC in overloaded muscle is associated with angiogenic adaptation by reanalyzing skeletal muscle from HeyL-knockout (KO) mice, which show blunted responses of SC proliferation, myonucleus addition, and overload-induced muscle hypertrophy. Reanalysis confirmed blunted SC proliferation and myonuclear accretion in the plantaris muscle of HeyL-KO mice 9 wk after synergist ablation. Interestingly, the increase in capillary-to-fiber ratio observed in wild-type (WT) mice was impaired in HeyL-KO mice. In both WT and HeyL-KO mice, the expression of VEGFA and VEGFB was similarly increased in response to overload. In addition, the expression pattern of TSP-1, a negative regulator of angiogenesis, was also not changed between WT and HeyL-KO mice. Collectively, these results suggest that SCs activation-myonuclear accretion plays a crucial role in angiogenesis during overload-induced muscle hypertrophy via independent of angiogenesis regulators.

High-intensity muscle contractions (HiMCs) are known to increase c-Myc expression that is known to stimulate ribosome biogenesis and protein synthesis in most cells. However, although c-Myc mRNA transcription and c-Myc mRNA translation have been shown to be upregulated following resistance exercise concomitantly with increased ribosome biogenesis, this connection has not been tested directly. We investigated the effect of adeno-associated virus (AAV)-mediated c-Myc overexpression, with or without fasting or percutaneous electrical stimulation-induced HiMC, on ribosome biogenesis and protein synthesis in adult mouse skeletal muscles. AAV-mediated overexpression of c-Myc in mouse skeletal muscles for 2 wk increased the DNA polymerase subunit POL1 mRNA, 45S-pre-rRNA, total RNA, and muscle protein synthesis without altering mechanistic target of rapamycin complex 1 (mTORC1) signaling under both ad libitum and fasted conditions. RNA-sequencing (RNA-seq) analyses revealed that c-Myc overexpression mainly regulated ribosome biogenesis-related biological processes. The protein synthesis response to c-Myc overexpression mirrored the response with HiMC. No additional effect of combining c-Myc overexpression and HiMC was observed. Our results suggest that c-Myc overexpression is sufficient to stimulate skeletal muscle ribosome biogenesis and protein synthesis without activation of mTORC1. Therefore, the HiMC-induced increase in c-Myc may contribute to ribosome biogenesis and increased protein synthesis following HiMC. NEW & NOTEWORTHY Resistance exercise is known to increase c-Myc expression, which is known to stimulate ribosome biogenesis and protein synthesis in a variety of cells. However, whether the increase in c-Myc stimulates ribosome biogenesis and protein synthesis in skeletal muscles remains unknown. We found that c-Myc overexpression is sufficient to stimulate skeletal muscle ribosome biogenesis and protein synthesis without activation of mTORC1.

A high-fat diet is known to rapidly induce obesity, insulin resistance, and anabolic resistance to nutrition within a month. However, the influence of a short-term high-fat diet on the response of muscle protein synthesis to acute resistance exercise is unclear. We observed that a short-term high-fat diet causes obesity, insulin resistance, intramuscular lipid droplet accumulation, and anabolic resistance to resistance exercise specifically in type I and IIa fibers.

Skeletal muscle has numerous nuclei within a cell. The nucleus is considered as the central organelle for muscle protein synthesis (MPS). However, it is unclear whether myonuclear number is associated with MPS capacity within the individual muscle fibres. Therefore, the purpose of the present study was to reveal the relationship between myonuclear number per unit muscle fibre length and MPS under basal and conditions of elevated MPS by high-intensity muscle contraction (HiMC) using an in vivo nascent protein labelling technique (SUnSET) in rodents. We found that myonuclear number was positively correlated with MPS in individual muscle fibres in the basal condition. Similarly, ribosomal protein S6 (rpS6) content, which is a rough estimate of ribosome content, was positively correlated with MPS. However, myonuclear number was not associated with rpS6 content. In contrast to the basal condition, when MPS was increased by acute HiMC, no correlation was observed between myonuclear number and MPS, but the association between rpS6 and MPS was maintained. Importantly, these observations indicate that the number of nuclei in individual myofibers is related only to MPS at rest. However, the ribosome content in individual fibres is related to MPS of individual myofibers both at rest and following HiMC.