• Fisheries and Aquatic Sciences
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• v.20 no.3
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• pp.3.1-3.8
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• 2017

Seahorse, a syngnathidae fish, is one of the important organisms used in Chinese traditional medicine. Hippocampus abdominalis, a seahorse species successfully cultured in Korea, was validated for use in food by the Ministry of Food and Drug Safety in February 2016; however. the validation was restricted to 50% of the entire composition. Therefore, to use H. abdominalis as a food ingredient, H. abdominalis has to be prepared as a mixture by adding other materials. In this study, the effect of H. abdominalis on muscles was investigated to scientifically verify its potential bioactivity. In addition, the anti-fatigue activity of a mixture comprising H. abdominalis and red ginseng (RG) was evaluated to commercially utilize H. abdominalis in food industry. H. abdominalis was hydrolyzed using Alcalase, a protease, and the effect of H. abdominalis hydrolysate (HH) on the muscles was assessed in C2C12 myoblasts by measuring cell proliferation and glycogen content. In addition, the mixtures comprising HH and RG were prepared at different percentages of RG to HH (20, 30, 40, 50, 60, 70, and 80% RG), and the anti-fatigue activity of these mixtures against oxidative stress was assessed in C2C12 myoblasts. In C2C12 myoblasts, $H_2O_2$-induced oxidative stress caused a decrease in viability and physical fatigue-related biomarkers such as glycogen and ATP contents. However, treatment with RG and HH mixtures increased cell viability and the content of fatigue-related biomarkers. In particular, the 80% RG mixture showed an optimum effect on cell viability and ATP synthesis activity. In this study, all results indicated that HH had anti-fatigue activity at concentrations approved for use in food by the law in Korea. Especially, an 80% RG to HH mixture can be used in food for ameliorating fatigue.

• Journal of Korean Medicine for Obesity Research
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• v.23 no.2
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• pp.60-68
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• 2023

Objectives: This study was conducted to compare the effects of Hominis placenta (Jahage, J) and wild ginseng (SanSam, S) pharmacopuncture drugs on muscle differentiation and energy metabolism regulation in C2C12 myotubes. Methods: The C2C12 myoblasts were differentiated into myotubes for 5 days by replacing in medium containing 2% horse serum and then treated with J and S pharmacopuncture extract at different concentrations for 24 hr. The expression of myosin heavy chain and energy metabolism-regulating factors, myosin heavy chain (MHC), nuclear respiratory factor-1 (NRF-1), and proliferator-activated receptor γ coactivator-1 alpha (PGC-1α) were determined in C2C12 myotubes by western blot. Additionally, the phosphorylation of AMPK and the expression of mitochondrial biogenesis, including sirtuin 1 (SIRT1) were determined in the myotubes. Results: As a result, treatment with J and S pharmacopuncture extract at 0.1 and 1 mg/mL increased the MHC expression in C2C12 myotubes compared with non-treated cells, but only S pharmacopuncture was shown a significant and distinct increase in the expression. Expression of TFAM and NRF-1 was also shown significant increases in S and J pharmacopuncture in C2C12 myotubes compared to non-treated cells. The phosphorylation of AMPK and the expression of PGC-1α and SIRT1 showed increased expression in S and J pharmacopuncture compared to non-treated cells. The effect of low-dose of J pharmacopuncture on the phosphorylated adenosine monophosphate-activated protein kinase (AMPK) and PGC-1α expression was greater than that of S pharmacopuncture. Conclusions: In conclusion, both J and S pharmacopuncture promote muscle differentiation in C2C12 myoblasts into myotubes and energy metabolism through the AMPK/SIRT1 signaling pathway. This indicates that the pharmacopuncture with tonic herbal medicines can help to improve skeletal muscle function.

• Journal of Animal Reproduction and Biotechnology
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• v.37 no.1
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• pp.17-26
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• 2022

Alpha-linolenic acid is an important polyunsaturated fatty acid that exhibits anticancer, anti-inflammatory, and antioxidative effects. In this study, we investigated the protective effects of alpha-linolenic acid on the cell proliferation and differentiation of C2C12 cells under essential amino acid-deficient conditions. Different concentrations of alpha-linolenic acid and essential amino acids were added to the growth and differentiation media. The concentrations of 10 µM of alpha-linolenic acid and 2% essential amino acid were chosen for subsequent experiments. Supplementation with alpha-linolenic acid and essential amino acids improved the proliferation and differentiation of C2C12 cells and significantly increased the mRNA levels of catalase, superoxide dismutase, B-cell lymphoma-2, and beclin-1 as well as the protein levels of PPARγ coactivator-1α compared to those in the controls. Moreover, supplementation with alpha-linolenic acid and essential amino acids reduced the levels of phosphorylated H2A.X variant histone, Bcl-2-associated X, p53, and light chain 3 during C2C12 cell proliferation, and increased the expression levels of myogenic factors 4 (myogenin) and 5 during C2C12 cell differentiation. Overall, we determined that alpha-linolenic acid and essential amino acids maintained the cell proliferation and differentiation of C2C12 cells via their anti-oxidative, anti-apoptotic, and anti-autophagic effects.

• Journal of Ginseng Research
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• v.46 no.3
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• pp.444-453
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• 2022

Background: Compound K (CK) is among the protopanaxadiol (PPD)-type ginsenoside group, which produces multiple pharmacological effects. Herein, we examined the effects of CK on muscle atrophy under hyperlipidemic conditions along with its pro-myogenic effects. Further, the molecular pathways underlying the effects of CK on skeletal muscle have been justified. Methods: C2C12 myotubes were treated with palmitate and CK. C2C12 myoblasts were differentiated using CK for 4-5 days. For the in vivo experiments, CK was administered to mice fed on a high-fat diet for 8 weeks. The protein expression levels were analyzed using western blotting analysis. Target protein suppression was performed using small interfering (si) RNA transfection. Histological examination was performed using Jenner-Giemsa and H&E staining techniques. Results: CK treatment attenuated ER stress markers, such as eIF2a phosphorylation and CHOP expression and impaired myotube formation in palmitate-treated C2C12 myotubes and skeletal muscle of mice fed on HFD. CK treatment augmented AMPK along with autophagy markers in skeletal muscle cells in vitro and in vivo experiments. AMPK siRNA or 3-MA, an autophagy inhibitor, abrogated the impacts of CK in C2C12 myotubes. CK treatment augmented p38 and Akt phosphorylation, leading to an enhancement of C2C12 myogenesis. However, AMPK siRNA abolished the effects of CK in C2C12 myoblasts. Conclusion: These findings denote that CK prevents lipid-induced skeletal muscle apoptosis via AMPK/autophagy-mediated attenuation of ER stress and induction of myoblast differentiation. Therefore, we may suggest the use of CK as a potential therapeutic approach for treating muscle-wasting conditions associated with obesity.

• Journal of Life Science
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• v.24 no.9
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• pp.1019-1024
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• 2014

Skeletal muscle atrophy can be defined as a decrease in or a disease of the muscle tissue, or as a disorder of the nerves that control the muscle, through injury or lack of use. This condition is associated with reactive oxygen species (ROS), resulting in various muscular disorders. Exposure to ROS induces muscle atrophy through several biological factors, such as SOD1 and HSP70. We found that cymbidium root extract reduced the $H_2O_2$-induced viability loss in C2C12 myoblasts and inhibited apoptosis. In addition, we showed that the cymbidium root extract increased the expression of HSP70 and decreased the expression of SOD1 in the $H_2O_2$-induced C2C12 myoblasts. These results suggest that cymbidium root extract might have therapeutic value in reducing ROS-induced muscle atrophy.

• Food Science of Animal Resources
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• v.44 no.1
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• pp.132-145
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• 2024

Sarcopenia, the age-related muscle atrophy, is a serious concern as it is associated with frailty, reduced physical functions, and increased mortality risk. Protein supplementation is essential for preserving muscle mass, and horse meat can be an excellent source of proteins. Since sarcopenia occurs under conditions of oxidative stress, this study aimed to investigate the potential anti-muscle atrophy effect of horse meat hydrolysate using C2C12 cells. A horse meat hydrolysate less than 3 kDa (A4<3kDa) significantly increased the viability of C2C12 myoblasts against H₂O₂-induced cytotoxicity. Exposure of C2C12 myoblasts to lipopolysaccharide led to an elevation of cellular reactive oxygen species levels and mRNA expression of proinflammatory cytokines, including tumor necrosis factor-α and interleukin 6, and these effects were attenuated by A4<3kDa treatment. Additionally, A4<3kDa activated protein synthesis-related proteins through the protein kinase B/mechanistic target of rapamycin pathway, while decreasing the expression of activity and degradation-related proteins, such as Forkhead box O3, muscle RING finger protein-1, and Atrogin-1 in dexamethasone-treated C2C12 myotubes. Therefore, the natural material A4<3kDa has the potential of protecting against muscle atrophy, while further in vivo study is needed.

• Journal of Korean Medicine for Obesity Research
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• v.15 no.2
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• pp.93-103
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• 2015

Objectives: It was investigated the cytoprotective efficacies of isorhamnetin, a flavonoid originally derived from Hippophae rhamnoides L., against oxidative stress-induced apoptosis in C2C12 myoblasts. Methods: The effects of isorhamnetin on cell growth, apoptosis and reactive oxygen species (ROS) generation were evaluated by trypan blue dye exclusion assay, 4',6-diamidino-2-phenylindole staining and flow cytometry. The levels of apoptosis-regulatory and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway-related proteins, and caspase activities (caspase-3 and -9) were determined by Western blot analysis and colorimetric assay, respectively. Results: Our results revealed that treatment with isorhamnetin prior to hydrogen peroxide ($H_2O_2$) exposure significantly increased the C2C12 cell viability and, indicating that the exposure of C2C12 cells to isorhamnetin conferred a protective effect against oxidative stress. Isorhamnetin also effectively attenuated $H_2O_2$-induced apoptosis and ROS generation, which was associated with the restoration of the upregulation of Bax and downregulation of Bcl-2 induced by $H_2O_2$. In addition, $H_2O_2$ enhanced the activation of caspase-9 and -3, and degradation of poly (ADP-ribose)-polymerase, a typical substrate protein of activated caspase-3; however, these events were almost totally reversed by pretreatment with isorhamnetin. Moreover, isorhamnetin increased the levels of heme oxygenase-1, a potent antioxidant enzyme, associated with the induction of Nrf2. Conclusions: Our data indicated that isorhamnetin may potentially serve as an agent for the treatment and prevention of muscle disorders caused by oxidative stress.

• BMB Reports
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• v.53 no.5
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• pp.278-283
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• 2020

Muscle fibers are generally formed as multinucleated fibers that are differentiated from myoblasts. Several reports have identified transcription factors and proteins involved in the process of muscle differentiation, but the roles of microRNAs (miRNAs) in myogenesis remain unclear. Here, comparative analysis of the miRNA expression profiles in mouse myoblasts and gastrocnemius (GA) muscle uncovered miR-3074-3p as a novel miRNA showing markedly reduced expression in fully differentiated adult skeletal muscle. Interestingly, elevating miR-3074-3p promoted myogenesis in C2C12 cells, primary myoblasts, and HSMMs, resulting in increased mRNA expression of myogenic makers such as Myog and MyHC. Using a target prediction program, we identified Caveolin-1 (Cav1) as a target mRNA of miR-3074-3p and verified that miR-3074-3p directly interacts with the 3' untranslated region (UTR) of Cav1 mRNA. Consistent with the findings in miR-3074-3p-overexpressing myoblasts, knockdown of Cav1 promoted myogenesis in C2C12 cells and HSMMs. Taken together, our results suggest that miR-3074-3p acts a positive regulator of myogenic differentiation by targeting Cav1.

• Herbal Formula Science
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• v.28 no.1
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• pp.29-39
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• 2020

Objectives : Glucoraphanin is one of the well-known natural glucosinolates found in cruciferous plants. In the present study, we investigated the effects and molecular mechanism of glucoraphanin in dexamethasone-induced skeletal muscle atrophy in vitro model. Methods : The cytotoxic effects of glucoraphanin on C2C12 myoblasts or myotubes were evaluated by MTT assay. The glucoraphanin was evaluated effects in dexamethasone-induced skeletal muscle atrophy in C2C12 myotubes using a real-time PCR, western blots analysis, and immunofluorescence staining of myosin heavy chain. Result : Glucoraphanin had no cytotoxicity on both C2C12 myoblasts or myotubes. Dexamethasone markedly induced muscle atrophy by up-regulating muscle-specific ubiquitin E3 ligase markers, atrogin-1 and MuRF1, and down-regulating MyoD, a myogenic regulatory factor whereas co-treatment of glucoraphanin and dexamethasone dose-dependently inhibited it. Furthermore, decreased expressions of p-Akt, p-FOXO1, and p-FOXO3a induced by dexamethasone were reversed by co-treatment with glucoraphanin and dexamethasone. In addition, dexamethasone obviously reduced myotube diameters, while co-treatment of glucoraphanin and dexamethasone increased those to a similar level as control. Conclusions : These results show that glucoraphanin suppresses dexamethasone-induced muscle atrophy in C2C12 myotubes through activation of Akt/FOXO signaling pathway.

• Journal of Life Science
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• v.27 no.12
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• pp.1479-1485
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• 2017

Muscle atrophy due to aging, starvation, and various chronic diseases leads to a decrease in muscle fiber area and density due to reduced muscle protein synthesis and increased protein breakdown. This study investigated the effect of dexamethasone and hydrogen peroxide on the induction of muscle atrophy and expression of atrophy-related genes in differentiated C2C12 myotubes. C2C12 myoblasts were differentiated into myotubes in differentiation medium. During myoblast differentiation, muscle-specific transcription factors, such as myogenin, and MyoD expression increased. Differentiated C2C12 myotubes exposed to noncytotoxic levels of dexamethasone and hydrogen peroxide showed a decrease in myotube diameter, which was associated with up-regulation of muscle-specific ubiquitin ligases, such as muscle atrophy F-box (MAFbx)/atrogin-1 and muscle RING finger-1 (MuRF1), and down-regulation of myogenin and MyoD. These results demonstrated that dexamethasone and hydrogen peroxide induced atrophy through regulation of muscle-specific ubiquitin ligases and muscle-specific transcription factors in C2C12 myotubes. In this study, we confirmed the process of differentiation of C2C12 myoblasts into myotubes in in vitro experiments in the presence of atrophy. This muscle atrophy model of C2C12 cells induced by dexamethasone or hydrogen peroxide seems suited to studies of the mechanism of muscle atrophy suppression and to exploit the experiment for excavating new muscle atrophy.