Morphological and biochemical changes of lymphatic vessels in the soleus muscle of mice after hindlimb unloading.

INTRODUCTION/AIMS: Lymphatic vessels are responsible for the removal of metabolic waste from body tissues. They also play a crucial role in skeletal muscle functioning thorough their high-energy metabolism. In this study we investigated whether disuse muscle atrophy induced by hindlimb unloading is associated with an alteration in the number of lymphatic vessels and differential expression of lymphangiogenic factors in the soleus muscle. METHODS: Male C57BL/6 mice were subjected to tail suspension (TS) for 2 or 4 weeks to induce soleus muscle atrophy. After TS, lymphatic and blood capillaries in the soleus muscle were visualized and counted by double staining with LYVE-1 and CD31. The protein and mRNA levels of vascular endothelial growth factor (VEGF)-C, VEGF-D, and vascular endothelial growth factor receptor-3 were measured by Western blotting and real-time reverse transcript polymerase chain reaction, respectively. RESULTS: TS for 2 weeks resulted in a significant decrease in the number of blood capillaries compared with controls. However, there was no significant change in the number of lymphatic capillaries. By contrast, TS for 4 weeks resulted in a significant decrease in the number of lymphatic and blood capillaries. We observed a significant decrease in the mRNA levels of VEGF-C and VEGF-D in mice subjected to TS for 4 weeks. DISCUSSION: The decrease of intramuscular lymphatic vessels may a crucial role in the process of muscle atrophy.

Cessation of electrically-induced muscle contraction activates autophagy in cultured myotubes.

Exercise is known to improve skeletal muscle function. The mechanism involves muscle contraction-induced activation of the mTOR pathway, which plays a central role in protein synthesis. However, mTOR activation blocks autophagy, a recycling mechanism with a critical role in cellular maintenance/homeostasis. These two responses to muscle contraction look contradictory to the functional improvement of exercise. Herein, we investigate these paradoxical muscle responses in a series of active-inactive phases in a cultured myotube model receiving electrical stimulation to induce intermittent muscle contraction. Our model shows that (1) contractile activity induces mTOR activation and muscle hypertrophy but blocks autophagy, resulting in the accumulation of damaged proteins, while (2) cessation of muscle contraction rapidly activates autophagy, removing damaged protein, yet a prolonged inactive state results in muscle atrophy. Our findings provide new insights into muscle biology and suggest that not only muscle contraction, but also the subsequent cessation of contraction plays a substantial role for the improvement of skeletal muscle function.

Jump Exercise and Food Restriction on Bone Parameters in Young Female Rats.

We examined the effect of jump exercise on bone parameters in young female rats under food restriction. Seven-week-old female rats were divided into four groups: a sedentary and ad libitum feeding group (n = 10), a jump exercise and ad libitum feeding group (n = 9), a sedentary and 30% food restriction group (n = 9), and a jump exercise and 30% food restriction group (n = 10). The jump groups jumped 20 times/day, 5 times/week. The experiment lasted for 13 weeks. There were no interactions of jump exercise and food restriction on bone. Jump exercise under food restriction conditions induced higher bone strength, bone mineral content, bone area, bone mineral density (BMD), and cortical bone volume in young female rats, similar to rats under ad libitum feeding conditions. Bone strength parameters were not significantly different between ad libitum intake and food restriction with jump exercise training; however, BMD, bone size, and bone mass in the food restriction groups did not reach the levels of those in the ad libitum conditions group with jump exercise training. Neither jump exercise nor food restriction had a significant effect on serum estradiol or IGF-1. Our study reveals jump exercise attenuates loss of biomechanical properties and some bone sites with food restriction in young female rats.

Soy protein intake increased bone mineral density under nonenergy-deficiency conditions but decreased it under energy-deficiency conditions in young female rats.

Many young individuals attempt to lose too much weight because of a false body image, which induces low bone mineral density (BMD) resulting from energy restriction. In addition, a decrease in estrogen has been observed along with the decrease in BMD. Estrogen is responsible for maintaining bone mass, and soybeans contain high levels of isoflavones, which have estrogen-like effects. Thus, we hypothesized that soy protein prevents low BMD caused by energy deficiency in young female rats. The purpose of this study was to examine the effect of soy protein intake on bone loss by energy deficiency in young female rats. Female Sprague-Dawley rats (6 weeks old) were randomly divided into the following 4 experimental groups: ad libitum feeding and casein diet (AL-Cas); ad libitum feeding and soy diet (AL-Soy); 40% energy restriction and casein diet (ER-Cas); and 40% energy restriction and soy diet (ER-Soy). The experimental period was 10.5 weeks. The AL-soy group had significantly higher BMD of the femur than the AL-Cas group (AL-Cas = 156 ± 5 mg/cm2, AL-Soy = 165 ± 7 mg/cm2; P < .05). Meanwhile, the ER-Soy group had significantly lower BMD of the tibia, femur, and lumbar spine than the ER-Cas group (ER-Cas = 147 ± 7 mg/cm2, ER-Soy = 133 ± 10 mg/cm2; P < .01). These results show that compared with ad libitum control groups, soy protein resulted in higher BMD under nonenergy deficiency, but under energy-deficiency conditions, it resulted in lower BMD.

Involvement of PI3K/Akt/TOR pathway in stretch-induced hypertrophy of myotubes.

Skeletal muscle cells are hypertrophied by mechanical stresses, but the underlying molecular mechanisms are not fully understood. Two signaling pathways, phosphatidylinositol 3-kinase (PI3K)/Akt to target of rapamycin (TOR) and extracellular signal-regulated kinase kinase (MEK) to extracellular signal-regulated kinase (ERK), have been proposed to be involved in muscle hypertrophy. In this study we examined the involvement of these pathways in primary cultures of chick skeletal myotubes subjected to passive cyclic stretching for 72 hours, a time that was sufficient to induce significant hypertrophy in our preparations. Hypertrophy was largely suppressed by wortmannin or rapamycin, inhibitors of PI3K or mTOR, respectively. Furthermore, phosphorylation of Akt was enhanced by stretching and suppressed by wortmannin. The MEK inhibitor, U0126, exerted a minimal influence on stretch-induced hypertrophy. We found that cyclic stretching of myotubes activates the PI3K/Akt/TOR pathway, resulting in muscle hypertrophy. The MEK/ERK pathway may contribute negatively to spontaneous hypertrophy.

Repetitive stretch suppresses denervation-induced atrophy of soleus muscle in rats.

This study was conducted to examine whether stretch-related mechanical loading on skeletal muscle can suppress denervation-induced muscle atrophy, and if so, to depict the underlying molecular mechanism. Denervated rat soleus muscle was repetitively stretched (every 5 s for 15 min/day) for 2 weeks. Histochemical analysis showed that the cross-sectional area of denervated soleus muscle fibers with repetitive stretching was significantly larger than that of control denervated muscle (P<0.05). We then examined the involvement of the Akt/mammalian target of the rapamycin (mTOR) cascade in the suppressive effects of repetitive stretching on muscle atrophy. Repetitive stretching significantly increased the Akt, p70S6K, and 4E-BP1 phosphorylation in denervated soleus muscle compared to controls (P<0.05). Furthermore, repetitive stretching-induced suppression of muscle atrophy was fully inhibited by rapamycin, a potent inhibitor of mTOR. These results indicate that denervation-induced muscle atrophy is significantly suppressed by stretch-related mechanical loading of the muscle through upregulation of the Akt/mTOR signal pathway.