• Journal of Ginseng Research
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• v.47 no.6
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• pp.726-734
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• 2023

Background: Skeletal muscles play a key role in physical activity and energy metabolism. The loss of skeletal muscle mass can cause problems related to metabolism and physical activity. Studies are being conducted to prevent such diseases by increasing the mass and regeneration capacity of muscles. Ginsenoside Rg5 has been reported to exhibit a broad range of pharmacological activities. However, studies on the effects of Rg5 on muscle differentiation and growth are scarce. Methods: To investigate the effects of Rg5 on myogenesis, C2C12 myoblasts were induced to differentiate with Rg5, followed by immunoblotting, immunostaining, and qRT-PCR for myogenic markers and promyogenic signaling (p38MAPK). Immunoprecipitation confirmed that Rg5 increased the interaction between MyoD and E2A via p38MAPK. To investigate the effects of Rg5 on prevention of muscle mass loss, C2C12 myotubes were treated with dexamethasone to induce muscle atrophy. Immunoblotting, immunostaining, and qRT-PCR were performed for myogenic markers, Akt/mTOR signaling for protein synthesis, and atrophy-related genes (Atrogin-1 and MuRF1). Results: Rg5 promoted C2C12 myoblast differentiation through phosphorylation of p38MAPK and MyoD/E2A heterodimerization. Furthermore, Rg5 stimulated C2C12 myotube hypertrophy via phosphorylation of Akt/mTOR. Phosphorylation of Akt induces FoxO3a phosphorylation, which reduces the expression of Atrogin-1 and MuRF1. Conclusion: This study provides an understanding of how Rg5 promotes myogenesis and hypertrophy and prevents dexamethasone-induced muscle atrophy. The study is the first, to the best of our knowledge, to show that Rg5 promotes muscle regeneration and to suggest that Rg5 can be used for therapeutic intervention of muscle weakness and atrophy, including cancer cachexia.

• Journal of Nutrition and Health
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• v.56 no.6
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• pp.602-614
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• 2023

Purpose: The balance between synthesis and degradation of proteins plays a critical role in the maintenance of skeletal muscle mass. Mitochondrial dysfunction has been closely associated with skeletal muscle atrophy caused by aging, cancer, and chemotherapy. Polygalacin D is a saponin derivative isolated from Platycodon grandiflorum (Jacq.) A. DC. This study aimed to investigate the effects of polygalacin D on myoblast differentiation and muscle atrophy in association with mitochondrial function in in vitro and in zebrafish models in vivo. Methods: C2C12 myoblasts were cultured in differentiation media containing different concentrations of polygalacin D, followed by the immunostaining of the myotubes with myosin heavy chain (MHC). The mRNA expression of markers related to myogenesis, muscle atrophy, and mitochondrial function was determined by real-time quantitative reverse transcription polymerase chain reaction. Wild type AB^* zebrafish (Danio rerio) embryos were treated with 5-fluorouracil, leucovorin, and irinotecan (FOLFIRI) with or without polygalacin D, and immunostained to detect slow and fast types of muscle fibers. The Tg(Xla.Eef1a1:mitoEGFP) zebrafish expressing mitochondria-targeted green fluorescent protein was used to monitor mitochondrial morphology. Results: The exposure of C2C12 myotubes to 0.1 ng/mL of polygalacin D increased the formation of MHC-positive multinucleated myotubes (≥ 8 nuclei) compared with the control. Polygalacin D significantly increased the expression of MHC isoforms (Myh1, Myh2, Myh4, and Myh7) involved in myoblast differentiation while it decreased the expression of atrophic markers including muscle RING-finger protein-1 (MuRF1), mothers against decapentaplegic homolog (Smad)2, and Smad3. In addition, polygalacin D promoted peroxisome proliferator-activated receptor-gamma coactivator (Pgc1α) expression and reduced the level of mitochondrial fission regulators such as dynamin-1-like protein (Drp1) and mitochondrial fission 1 (Fis1). In a zebrafish model of FOLFIRI-induced muscle atrophy, polygalacin D improved not only mitochondrial dysfunction but also slow and fast muscle fiber atrophy. Conclusion: These results demonstrated that polygalacin D promotes myogenesis and alleviates chemotherapy-induced muscle atrophy by improving mitochondrial function. Thus, polygalacin D could be useful as nutrition support to prevent and ameliorate muscle wasting and weakness.

• Journal of Nutrition and Health
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• v.57 no.1
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• pp.53-64
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• 2024

Purpose: Mitochondria play a crucial role in preserving skeletal muscle mass, and damage to mitochondria leads to muscle mass loss. This study investigated the effects of oxypeucedanin hydrate, a furanocoumarin isolated from Angelica dahurica radix, on myogenesis and mitochondrial function in vitro and in zebrafish models. Methods: C2C12 myotubes cultured in media containing 0.1, 1, 10, or 100 ng/mL oxypeucedanin hydrate were immunostained with myosin heavy chain (MHC), and then multinucleated MHC-positive cells were counted. The expressions of markers related to muscle differentiation, muscle protein degradation, and mitochondrial function were determined by quantitative reverse transcription polymerase chain reaction. To investigate the effects of oxypeucedanin hydrate on mitochondrial dysfunction, Tg(Xla.Eef1a1:mito-EGFP) zebrafish embryos were treated with 5-fluorouracil, leucovorin, and irinotecan (FOLFIRI) with or without oxypeucedanin hydrate and analyzed for mito-EGFP intensity and mitochondrial length. Results: Oxypeucedanin hydrate significantly increased MHC-positive multinucleated myotubes (≥ 3 nuclei) and increased the expression of the myogenic marker myosin heavy chain 4. However, it decreased the expressions of muscle-specific RING finger protein 1 and muscle atrophy f-box (markers of muscle protein degradation). Furthermore, oxypeucedanin hydrate enhanced the expressions of markers of mitochondrial biogenesis (peroxisome proliferator-activated receptor-gamma coactivator 1 alpha, transcription factor a mitochondrial, succinate dehydrogenase complex flavoprotein subunit A, and cytochrome c oxidase subunit 1) and mitochondrial fusion (optic atrophy 1). However, it reduced the expression of dynamin-related protein 1 (a mitochondrial fission regulator). Consistently, oxypeucedanin hydrate reduced FOLFIRI-induced mitochondrial dysfunction in the skeletal muscles of zebrafish embryos. Conclusion: The study indicates that oxypeucedanin hydrate promotes myogenesis by improving mitochondrial function, and thus, suggests oxypeucedanin hydrate has potential use as a nutritional supplement that improves muscle mass and function.

• Journal of Physiology & Pathology in Korean Medicine
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• v.22 no.2
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• pp.340-345
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• 2008

Extract of Acanthopanax senticosus has recently been demonstrated to possess significant antidiabetic potential, in accordance with the traditional use of this plant as an antidiabetic natural health product. The present study evaluated the effects of fermented extract (FE) of this plant on glucose-stimulated insulin secretion, glucose uptake, and streptozotocin-induced type 1 diabetes model. A 3 h pretreatment with FE prevented $IL-1{\beta}$ and $IFN-{\gamma}$ toxicity in isolated rat islets. However, it did not affect insulin-stimulated glucose uptake in C2C12 myotubes. In addition, pretreatment of mice with FE blocked the destruction of streptozotocin-induced islets and the development of type 1 diabetes. FE reduced blood glucose level, increased insulin secretion, and improved glucose tolerance in streptozotocin-treated mice, whereas nonfermented extract (NFE) had moderate effects. Immunohistochemical staining for insulin clearly showed that pretreatment with FE blocked the STZ-induced islets destruction and restored the number of islet cells that secreted insulin to the level of the control. Although the active principles and their mechanisms of action remain to be identified, FE may nevertheless represent a novel complementary therapy and a source of novel therapeutic agents against type 1 diabetes mellitus.

• Journal of Ginseng Research
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• v.42 no.1
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• pp.116-121
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• 2018

Background: Black ginseng (BG) has greatly enhanced pharmacological activities relative to white or red ginseng. However, the effect and molecular mechanism of BG on muscle growth has not yet been examined. In this study, we investigated whether BG could regulate myoblast differentiation and myotube hypertrophy. Methods: BG-treated C2C12 myoblasts were differentiated, followed by immunoblotting for myogenic regulators, immunostaining for a muscle marker, myosin heavy chain or immunoprecipitation analysis for myogenic transcription factors. Results: BG treatment of C2C12 cells resulted in the activation of Akt, thereby enhancing hetero-dimerization of MyoD and E proteins, which in turn promoted muscle-specific gene expression and myoblast differentiation. BG-treated myoblasts formed larger multinucleated myotubes with increased diameter and thickness, accompanied by enhanced Akt/mTOR/p70S6K activation. Furthermore, the BG treatment of human rhabdomyosarcoma cells restored myogenic differentiation. Conclusion: BG enhances myoblast differentiation and myotube hypertrophy by activating Akt/mTOR/p70S6k axis. Thus, our study demonstrates that BG has promising potential to treat or prevent muscle loss related to aging or other pathological conditions, such as diabetes.

• Biomedical Science Letters
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• v.13 no.4
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• pp.251-261
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• 2007

A myofiber of skeletal muscle is composed of myofibrils, sarcolemma (plasma membrane), and constameres, which anchor the myofibrils to the sarcolemma. Achvillin is a recently identified F-actin binding muscle protein, co-isolates with dystrophin and caveolin-3 in low-density sarcolemma of striated muscle, and colocalizes with dystrophin at costameres, the specialized adhesion sites in muscle. Archvillin also binds to nebulin and localizes at myofibrillar Z-discs, the lateral boundaries of the sarcomere in muscle. However other roles of archvillin on the dynamics of myofibrillogenesis remain to be defined. The goal of this study is, by using siRNA-mediated gene silencing technique, to investigate the effect of archvillin on the dynamics of myofibrillogenesis in cell culture of a mouse skeletal myogenic cell line (C2C12), where presumptive myoblasts withdraw from the cell cycle, fuse, undergo de novo myofibrillogenesis, and differentiate into mature myotubes. The roles of archvillin in the assembly and maintenance of myofibril and during the progression of myofibrillogenesis induced in skeletal myoblast following gene silencing in the cell culture were investigated. Fluorescence microscopy demonstrated that the distribution of archvillin was changed along the course of myofibril assembly with nebulin, vinculin and F-actin and then located at Z-lines with nebulin. Fluorescence microscopy demonstrated that knockdown of mouse archvillin expression led to an impaired assembly of new myofibrillar clusters and delayed fusion and myofibrillogenesis although the mouse archvillin siRNA did not affect those expressions of archvillin binding proteins, such as nebulin and F-actin. This result is corresponded with that of RT-PCR and western blots. When the perturbed archvillin was rescued by co-transfection with GFP or Red tagged human archvillin construct, the inhibited cell fusion and myotube formation was recovered. By using siRNA technique, archvillin was found to be involved in early stage of myofibrillogenesis. Therefore, the current data suggest the idea that archvillin plays critical roles on cell fusion and dynamic myofibril assembly.

• Journal of Life Science
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• v.33 no.12
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• pp.1036-1045
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• 2023

Sarcopenia, a condition characterized by the insidious loss of skeletal muscle mass and strength, represents a significant and growing healthcare challenge, impacting the mobility and quality of life of aging populations worldwide. This study investigated the therapeutic potential of soybean leaf extract (SL) for dexamethasone (Dexa)-induced muscle atrophy in vitro and in an in vivo model. In vitro experiments showed that SL significantly alleviated Dexa-induced atrophy in C2C12 myotube cells, as evidenced by preserved myotube morphology, density, and size. Moreover, SL treatment significantly reduced the mRNA and protein levels of muscle RING-finger protein-1 (MuRF1) and muscle atrophy F-box (MAFbx), key factors regulating muscle atrophy. In a Dexa-induced atrophy mouse model, SL administration significantly inhibited Dexa-induced weight loss and muscle wasting, preserving the mass of the gastrocnemius and tibialis anterior muscles. Furthermore, mice treated with SL exhibited significant improvements in muscle function compared to their counterparts suffering from Dexa-induced muscle atrophy, as evidenced by a notable increase in grip strength and extended endurance on treadmill tests. Moreover, SL suppressed the expression of muscle atrophy-related proteins in skeletal muscle, highlighting its protective role against Dexa-induced muscle atrophy. These results suggest that SL has potential as a natural treatment for muscle-wasting conditions, such as sarcopenia.

• Molecules and Cells
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• v.38 no.4
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• pp.356-361
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• 2015

Mitochondrial dysfunction is associated with insulin resistance and diabetes. We previously showed that retinoid X receptor ${\alpha}$ ($RXR{\alpha}$) played an important role in transcriptional regulation of oxidative phosphorylation (OXPHOS) genes in cells with mitochondrial dysfunction caused by mitochondrial DNA mutation. In this study, we investigated whether mitochondrial dysfunction induced by incubation with OXPHOS inhibitors affects insulin receptor substrate 1 (IRS1) mRNA and protein levels and whether $RXR{\alpha}$ activation or overexpression can restore IRS1 expression. Both IRS1 and $RXR{\alpha}$ protein levels were significantly reduced when C2C12 myotubes were treated with the OXPHOS complex inhibitors, rotenone and antimycin A. The addition of $RXR{\alpha}$ agonists, 9-cis retinoic acid (9cRA) and LG1506, increased IRS1 transcription and protein levels and restored mitochondrial function, which ultimately improved insulin signaling. $RXR{\alpha}$ overexpression also increased IRS1 transcription and mitochondrial function. Because $RXR{\alpha}$ overexpression, knock-down, or activation by LG1506 regulated IRS1 transcription mostly independently of mitochondrial function, it is likely that $RXR{\alpha}$ directly regulates IRS1 transcription. Consistent with the hypothesis, we showed that $RXR{\alpha}$ bound to the IRS1 promoter as a heterodimer with peroxisome proliferator-activated receptor ${\delta}$ ($PPAR{\delta}$). These results suggest that $RXR{\alpha}$ overexpression or activation alleviates insulin resistance by increasing IRS1 expression.