• 생명과학회지
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• 제17권3호통권83호
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• pp.420-426
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• 2007

본 연구에서는 유전자 발현에 중요한 조절 역할을 하는 DNA 메틸화를 식이성 폴리페놀의 하나인 녹차의 대표적인 추출물 EGCG로 억제하였을 때 C2C12 myoblast 세포에 일어나는 현상을 살펴보았다. 그 결과 smooth muscle의 지표인 transgelin, smooth muscle ${\alpha}-actin$ mRNA와 단백질이 현저히 증가함을 보였고, 형태학적으로도 smooth muscle의 전형적인 모습을 보였다. 식이에 포함된 DNA 메틸화 억제제인 EGCG가 C2C12 myoblast를 smooth muscle로 분화를 유도하였으며, 암 예방 차원에서의 EGCG의 역할 외에 혈관질환과 같은 smooth muscle에 관련된 예방과 치료차원에서 EGCG의 역할이 있을 것으로 사료된다. 본 연구는 C2C12 myoblast를 smooth muscle로 유도하는 결정적인 신호전달 역할을 하는 유전자에 대한 연구는 수행하지 못하였다. 따라서 EGCG에 의해 변화되는 유전자에 대한 기전연구가 필요하다고 하겠다.

• Asian-Australasian Journal of Animal Sciences
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• 제20권7호
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• pp.1115-1119
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• 2007

This study was conducted to determine the difference between satellite cells (porcine) and myoblasts (C2C12) in their differentiation under the influence of 2, 4-thiazolidindion. C2C12 myoblast cells and porcine satellite cells (isolated from 10 d old $Landrace{\times}Duroc$ piglets) were grown to absolute confluency. Post confluent cells (day 0) were further exposed to adipogenic induction medium along with 2, 4-thiazolidindion ($8{\mu}M$) for 2 d. Thereafter, cells were exposed to 2, 4-thiazolidindion alone every 2 d till day 10 and analysed. The control was cultured in differentiation medium without any treatment. Increased (p<0.05) expression of transcriptional factors i.e. C/EBP-${\alpha}$ and PPAR-${\gamma}$ and transition of cells to adipocyte morphology was noticed from 2 d and 4 d onwards in satellite cells (Porcine) and myoblasts (C2C12) respectively. Myogenesis was observed to be suppressed completely in case of satellite cells compared to myoblasts in response to 2, 4-thiazolidindion. Pax-7 (transcriptional factor) appeared as a sole entity to satellite cells only, as it was not identified in case of myoblasts. Although both the cells were converting to adipoblasts, the degree of their conversion was different in response to 2, 4-thiazolidindion. Therefore, the hypothesis that satellite cells contribute various domains to the growing myoblasts appeared obscured and found to be dependent on the proliferative energy/or degree of fusion. However, it revealed satellite cells as currency to myoblasts/muscle.

• 대한한방내과학회지
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• 제29권1호
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• pp.243-257
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• 2008

Myostatin, a negative regulator of myogenesis, is shown to function by controlling the proliferation of myoblasts. In this study we show that myostatin is an inhibitor of myoblast differentiation and that this inhibition is mediated through Smad 3. To determine MyoD expression by Dodamtang treatment, we compared the expression pattern of $C_{2}C_{12}$ mouse myoblasts that constitutively express myostatin with control cells. In vitro, increasing concentrations of Dodamtang reversibly prevented the myogenic blockage of myoblasts by myostatin expression. ELISA assay, Western and confocal analysis indicated that treatment of Dodamtang to the low serum culture media increased the levels of MyoD leading to the inhibition of myogenic differentiation by myostatin. The stable transfection of $C_{2}C_{12}$ myoblasts with myostatin expressing constructs did rescue MyoD-induced myogenic differentiation. Consistent with this, the treatment of Dodamtang rescued the expression of a MyoD in $C_{2}C_{12}$ myoblasts treated with myostatin. Taken together, these results suggest that induction of MyoD by Dodamtang inhibits myostatin activity and expression via SMAD3 resulting in the rescue of the myoblasts to differentiate into myotubes. Thus we propose that myostatin action by Dodamtang plays a critical role in myogenic differentiation and that the muscular hyperplasia and hypertrophy seen in animals that blockage of functional myostatin is because of deregulated proliferation and differentiation of myoblasts.

• 대한한방내과학회지
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• 제43권3호
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• pp.344-362
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• 2022

Objective: In this study, the antioxidant activity of Ojayeonjong-hwan extracts was compared, and the following results were obtained. Methods: For hydrothermal and ethanol extracts, DPPH free radical and ABTS cationic radical erasing activity and reducing power using the FRAP method were compared, and the association between the antioxidant power of each extract and total phenol content was investigated. Significant results were obtained through in vitro apoptosis analysis through FFITC staining, mitochondrial membrane potential analysis, and ROS level measurement using C2C12 myoblastoma. Results: 1. In a comparison of DPPH free radical and ABTS cationic radical scavenging activity, water, and 70% ethanol extracts of Ojayeonjong-hwan (WEO and EEO) showed superior radical scavenging ability. 2. In the results of reducing power using the FRAP method, WEO and EEO showed antioxidant activity, which was shown to be dependent on the total phenol content contained in the extracts. 3. In comparison to the protective effect against H₂O₂-induced oxidative stress in C2C12 myoblasts, water extracts had no significant effect, but 70% ethanol extracts inhibited H₂O₂-mediated cytotoxicity in a concentration-dependent manner. 4. The cytotoxic protective effect of EEO against oxidative stress in C2C12 myoblasts was correlated with its inhibitory effects on H₂O₂-induced apoptosis and cell-cycle arrest. 5. In H₂O₂-treated C2C12 myoblasts, the apoptosis inhibitory effects of EEO were associated with the suppression of mitochondrial dysfunction and DNA damage. 6. The protective effects of EEO against H₂O₂-induced oxidative stress in C2C12 myoblasts were directly related to the inhibition of ROS generation. Conclusions: Ojayeonjong-hwan extracts all have protective potential against oxidative stress.

• 대한한의학방제학회지
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• 제29권4호
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• pp.181-189
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• 2021

Objectives : The calyx of Diospyros kaki Thunb. has usually been used to treat obstinate hiccup. This study investigated the protective effects of Diospyros kaki using C2C12 myoblasts of H₂O₂-induced oxidative stress. Methods : Cell viability and cytotoxicity were determined by MTT assay. The level of reactive oxygen species (ROS), malondialdehyde (MDA), and reduced glutathione (GSH) were measured by using optical properties. Results : The calyx of Diospyros kaki Thunb. extract showed no toxicity to C2C12 myoblasts until 20 ㎍/mL concentration and increased cell viability compared to the H₂O₂ treated group. The calyx of Diospyros kaki Thunb. extract inhibited the production of ROS and MDA at all concentrations. In addition, the calyx of Diospyros kaki Thunb. extract increased the concentration of GSH. Conclusion : This study provides that the calyx of Diospyros kaki Thunb. can be used as a potential material that exhibit antioxidative and protective effects on H₂O₂-induced oxidative stress in C2C12 Myoblasts.

• 한국식품영양과학회지
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• 제46권11호
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• pp.1408-1413
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• 2017

본 연구에서는 강황 열수 추출물의 항산화 활성 및 산화적 스트레스에 대한 보호 효과를 확인하고자 하였다. 항산화 활성을 확인하기 위해 총 페놀성 화합물 함량, 총 플라보노이드 함량 및 라디칼 소거능을 측정하였다. 강황 열수 추출물의 총 페놀성 화합물 및 플라보노이드 함량은 각각 $2,474.4{\pm}31.9mg$ GAE/100 g 및 $892.1{\pm}21.2mg$ CE/100 g으로 나타났다. 강황 열수 추출물의 라디칼 소거능은 DPPH 및 ABTS 라디칼을 이용하여 측정하였다. 실험 결과 DPPH 및 ABTS 라디칼 소거능을 $SC_{50}$ 값으로 계산하였을 때 각각 $188.5{\pm}3.0{\mu}g/mL$ 및 $92.0{\pm}0.9{\mu}g/mL$로 나타났다. 이러한 라디칼 소거능에는 강황 열수 추출물에 함유되어 있는 총 페놀성 화합물 및 플라보노이드가 영향을 미쳤을 것이라고 판단된다. C2C12 myoblast에 강황 열수 추출물을 처리하였을 때 $1,000{\mu}g/mL$ 농도까지 세포 독성이 나타나지 않음을 확인하였으며, 안전성이 확인된 $500{\mu}g/mL$ 농도까지 실험을 진행하였다. 강황 열수 추출물의 $H_2O_2$에 대한 보호 효과를 측정한 결과 강황 열수 추출물을 처리하였을 때 농도 의존적으로 보호 효과가 나타나는 것을 확인할 수 있었다. 또한, $H_2O_2$ 처리 후 DCF-DA 방법을 이용하여 세포 내 활성산소종(ROS) 수준을 측정한 결과 강황 열수 추출물을 처리하였을 때 세포 내 ROS 수준이 유의적으로 감소하는 것을 확인하였다. 이상의 결과로부터 강황 열수 추출물은 항산화 활성을 나타냈으며, C2C12 myoblast에 $H_2O_2$로 유도된 산화적 스트레스를 감소시키는 효과를 나타내는 것으로 생각된다.

• BMB Reports
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• 제53권11호
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• pp.605-610
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• 2020

Skeletal myogenesis is a complex process that is finely regulated by myogenic transcription factors. Recent studies have shown that saturated fatty acids (SFA) can suppress the activation of myogenic transcription factors and impair the myogenic differentiation of progenitor cells. Despite the increasing evidence of the roles of miRNAs in myogenesis, the targets and myogenic regulatory mechanisms of miRNAs are largely unknown, particularly when myogenesis is dysregulated by SFA deposition. This study examined the implications of SFA-induced miR-183-5p on the myogenic differentiation in C2C12 myoblasts. Long-chain SFA palmitic acid (PA) drastically reduced myogenic transcription factors, such as myoblast determination protein (MyoD), myogenin (MyoG), and myocyte enhancer factor 2C (MEF2C), and inhibited FHL1 expression and myogenic differentiation of C2C12 myoblasts, accompanied by the induction of miR-183-5p. The knockdown of FHL1 by siRNA inhibited myogenic differentiation of myoblasts. Interestingly, miR-183-5p inversely regulated the expression of FHL1, a crucial regulator of skeletal myogenesis, by targeting the 3'UTR of FHL1 mRNA. Furthermore, the transfection of miR-183-5p mimic suppressed the expression of MyoD, MyoG, MEF2C, and MyHC, and impaired the differentiation and myotube formation of myoblasts. Overall, this study highlights the role of miR-183-5p in myogenic differentiation through FHL1 repression and suggests a novel miRNA-mediated mechanism for myogenesis in a background of obesity.

• 대한한의학방제학회지
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• 제25권2호
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• pp.179-191
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• 2017

Objectives : Increased oxidative stress by reactive oxygen species (ROS) has been suggested as a major cause of muscle fatigue. Although several studies have demonstrated the various biological properties of Sophora flavescens Aiton, Glycyrrhiza uralensis Fischer and Dictamnus dasycarpus Turcz, but the antioxidative potentials have not been clearly demonstrated. The present study was designed to investigate the protective effects of their water and ethanol extract mixtures (medicinal herbal mixtures, MHMIXs) on hydrogen peroxide ($H_2O_2$)-induced cell damage and apoptosis in C2C12 myoblasts. Methods : Cytotoxicity was assessed by an MTT assay. Quantitative evaluation of apoptosis induction and ROS production was evaluated by flow cytometry analysis. Expression levels of apoptosis regulatory and DNA-damage proteins were detected by Western blotting. Result : The inhibition of $H_2O_2$-induced cell proliferation was effectively blocked in extracts of 3: 1: 1 (EMHMIXs-1) or 2: 2: 1 (EMHMIXs-2) of S. flavescens, G. uralensis and D. dasycarpus Turcz, ethanol extracts from various complex extracts in C2C12 myoblasts. EMHMIXs-1 and EMHMIXs-2 also effectively attenuated $H_2O_2$-induced C2C12 cell apoptosis, which was associated with the restoration of the upregulation of Bad and death receptor 4, and downregulation of XIAP and cIAP-1 induced by $H_2O_2$. In addition, these herbal mixtures significantly blocked the $H_2O_2$-induced ROS generation and phosphorylation of $p-{\gamma}H2A.X$, which suggests that they can prevent $H_2O_2$-induced cellular DNA damage. Conclusions : The results suggest that EMHMIXs-1 and EMHMIXs-2 could block the DAN damage and apoptosis of C2C12 myoblasts by oxidative stress through blocking ROS generation.

• 대한본초학회지
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• 제34권6호
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• pp.99-107
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• 2019

Objective : The present study was conducted to investigate the effects of Chaenomelis Fructus (CF) on the regulation of biogenesis in C2C12 mouse skeletal muscle cells. Methods : C2C12 myoblasts were differentiated into myotubes in 2% horse serum-containing medium for 5 days, and then treated with CF extract at different concentrations for 48 hr. The expression of muscle differentiation markers, myogenin and myosin heavy chain (MHC) and mitochondrial biogenesis-regulating factors, peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC1α), sirtuin1 (Sirt1), nuclear respiratory factor1 (NRF1) and transcription factor A, mitochondrial (TFAM), and the phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC) were determined in C2C12 myotubes by reverse transcriptase (RT)-polymerase chain reaction (RT-PCR) and western blot, respectively. The cellular glucose levels and total ATP contents were measured by cellular glucose uptake and ATP assays, respectively. Results : Treatment with CF extract (0.01, 0.02, and 0.05 mg/㎖) significantly increased the expression of MHC protein in C2C12 myotubes compared with non-treated cells. CF extract significantly increased the expression of PGC1α and TFAM in the myotubes. Also, CF extract significantly increased glucose uptake levels and ATP contents in the myotubes. Conclusion : CF extract can stimulate C2C12 myoblasts differentiation into myotubes and increase energy production through upregulation of the expression of mitochondrial biogenetic factors in C2C12 mouse skeletal muscle cell. This suggests that CF can help to improve skeletal muscle function with stimulation of the energy metabolism.

• Molecules and Cells
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• 제25권4호
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• pp.479-486
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• 2008

Mitogen-activated protein kinases (MAPKs) play an indispensable role in activation of the myogenic program, which is responsive to mechanical stimulation. Although there is accumulating evidence of mechanical force-mediated cellular responses, the role of MAPK in regulating the myogenic process in myoblasts exposed to cyclic stretch is unclear. Cyclic stretch induced the proliferation of C2C12 myoblasts and inhibited their differentiation into myotubes. In particular, it induced persistent phosphorylation of p38 kinase, and decreased the level of phosphorylation of extracellular-signal regulated kinase (ERK). Partial inhibition of p38 phosphorylation increased cellular levels of MyoD and p-ERK in stretched C2C12 cells, along with increased myotube formation. Treatment with $10{\mu}M$ PD98059 prevented myogenin expression in response to a low dose of SB203580 ($3{\mu}M$) in the stretched cells, suggesting that adequate ERK activation is also needed to allow the cells to differentiate into myotubes. These results suggest that cyclic stretch inhibits the myogenic differentiation of C2C12 cells by activating p38-mediated signaling and inhibiting ERK phosphorylation. We conclude that p38 kinase, not ERK, is the upstream signal transducer regulating cellular responses to mechanical stretch in skeletal muscle cells.