• Journal of Microbiology and Biotechnology
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• 제21권9호
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• pp.921-929
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• 2011

Stimulation of AMP-activated protein kinase (AMPK) signaling followed by increase of glucose uptake in L6 myotubes were studied with organic solvent extract of Malva verticillata (MV) seeds. Ethanol extract of M. verticillata seeds (MVE) significantly increased the phosphorylation level of AMPK, acetyl-CoA carboxylase (ACC), and glucose uptake in L6 myotube cells. The MVE was fractionated with n-hexane (MVE-H), chloroform (MVE-C), ethylacetate (MVE-E), n-butanol (MVE-B), and water (MVE-W). MVE-H (150 ${\mu}g$/ml) showed the highest phosphorylating activity and increased glucose uptake by 2.3-fold. Oral administration of MVE-H (40 mg/kg) for 4 weeks to type 2 diabetic (db/db) mice reduced non-fasting and fasting blood glucose levels by 17.1% and 23.3%, respectively. Phosphorylation levels of AMPK and ACC in the soleus muscle and liver tissue of db/db mice were significantly increased by the administration of MVE-H. MVE-H was further fractionated using preparative HPLC to identify the AMPK-activating compounds. The NMR and GC-MS analyses revealed that ${\beta}$-sitosterol was a major effective compound in MVE-H. Phosphorylation levels of AMPK and ACC, and glucose uptake were significantly increased by the treatment of MVE-S (${\beta}$-sitosterol) isolated from M. verticillata to L6 cells, and these effects were attenuated by an AMPK inhibitor (Compound C) pretreatment. These results, taken together, demonstrate that increased glucose uptake in L6 myotubes by MVE-H treatment is mainly accomplished through the activation of AMPK. Our finding suggests that the extract isolated from M. verticillata seed would be beneficial for the treatment of metabolic disease including type 2 diabetes and hyperlipidemia.

• Environmental Analysis Health and Toxicology
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• 제20권1호
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• pp.75-85
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• 2005

This study was aimed to know the effect of cadmium chloride (CdCl₂) on glucose transport in L6 myotube and its action mechanism. CdCl₂ increased the 2-deoxy- (l-3H)-D-glucose (2-DOG) uptake 1.9 and 2.4 fold at 10 and 25 μM respectively. To investigate the stimulating-mechanism of glucose transport induced by CdCl₂, the wortmannin and PD98059 were used as PI3K (phosphatidylinositol 3-kinase) inhibitor and MAPK inhibitor respectively, which did not affect 2-DOG uptake. This fact suggests that CdCl₂ induced 2-DOG uptake may not be concerned to the insulin signalling pathway. Whereas nifedipine, a calcium channel blocker, and trifluoperazine, a calmodulin inhibitor, were found to inhibit the 2-DOG uptake stimulted by CdCl₂. In addition, we also measured the ROS (reactive oxygen species) production and GSH level in L6 myotube to investigate the correlation between the glucose uptake and ROS. CdCl₂(25 μM) increased ROS generation approximately 1.5 fold and changed the cellular GSH level, but GSSG/GSH ratio remained unchanged. CdCl₂ stimulated 2-DOG uptake and ROS generation were inhibited by N-acetylcystein. And BSO pretreatment, a potent inhibitor of γ-GCS, resulted in the dramatic decrease of 2-DOG uptake and also the increase of the sensitivity to cadmium cytotoxicity. The obtained results suggest that CdCl₂-stimulated glucose uptake might be based on the activation of HMP shunt as an antioxidant defense mechanism of the cells.

• 한국진공학회:학술대회논문집
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• 한국진공학회 2015년도 제49회 하계 정기학술대회 초록집
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• pp.158-158
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• 2015

Myoblast are myogenic precursors that proliferate, activate, and differentiate on muscle injury to sustain the regenerative capacity of skeletal muscle; The neuronal isoform of nitric oxide synthase (nNOS, termed also NOS-I) is expressed in normal adult skeletal muscle, suggesting important functions for Nitric oxide (NO) in muscle biology1,2,3. However, the expression and subcellular localization of NO in muscle development and myoblast differentiation are largely unknown. In this study, we examined effects of the nitric oxide generated by a microwave plasma torch, on proliferation/differentiation of rat myoblastic L6 cells. Experimental data pertaining to nitric oxide production are presented in terms of the oxygen input in units of cubic centimetres per minute. The various levels of nitric oxide are observed depending on the flow rate of nitrogen gas, the ratio of oxygen gas, and the microwave power4. In order to evaluate the potential of nitric oxide as an activator of cell differentiation, we applied nitric oxide generated from the microwave plasma torch to L6 skeletal muscles. Differentiation of L6 cells into myotubes was significantly enhanced the differentiation after nitric oxide treatment. Nitric oxide treatment also increase the expression of myogenesis marker proteins and mRNA level, such as myogenin and myosin heavy chain (MHC), as well as cyclic guanosine monophosphate (cGMP), However during the myotube differentiation we found that NO activate oxidative stress signaling erks expression. Therefore, these results establish a role of NO and cGMP in regulating myoblast differentiation and elucidate their mechanism of action, providing a direct link with oxidative stress signalling, which is a key player in myogenesis. Based on these findings, nitric oxide generated by plasma can be used as a possible activator of cell differentiation and tissue regeneration.

• The Korean Journal of Physiology and Pharmacology
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• 제23권6호
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• pp.539-547
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• 2019

Anoctamin 5 (ANO5)/TMEM16E belongs to a member of the ANO/TMEM16 family member of anion channels. However, it is a matter of debate whether ANO5 functions as a genuine plasma membrane chloride channel. It has been recognized that mutations in the ANO5 gene cause many skeletal muscle diseases such as limb girdle muscular dystrophy type 2L (LGMD2L) and Miyoshi muscular dystrophy type 3 (MMD3) in human. However, the molecular mechanisms of the skeletal myopathies caused by ANO5 defects are poorly understood. To understand the role of ANO5 in skeletal muscle development and function, we silenced the ANO5 gene in C2C12 myoblasts and evaluated whether it impairs myogenesis and myotube function. ANO5 knockdown (ANO5-KD) by shRNA resulted in clustered or aggregated nuclei at the body of myotubes without affecting differentiation or myotube formation. Nuclear positioning defect of ANO5-KD myotubes was accompanied with reduced expression of Kif5b protein, a kinesin-related motor protein that controls nuclear transport during myogenesis. ANO5-KD impaired depolarization-induced $[Ca2^{+}]_i$ transient and reduced sarcoplasmic reticulum (SR) $Ca^{2+}$ storage. ANO5-KD resulted in reduced protein expression of the dihydropyridine receptor (DHPR) and SR $Ca^{2+}-ATPase$ subtype 1. In addition, ANO5-KD compromised co-localization between DHPR and ryanodine receptor subtype 1. It is concluded that ANO5-KD causes nuclear positioning defect by reduction of Kif5b expression, and compromises $Ca^{2+}$ signaling by downregulating the expression of DHPR and SERCA proteins.

• 생명과학회지
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• 제23권9호
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• pp.1163-1169
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• 2013

인슐린은 근육세포 표면으로 포도당 수송체 4(glucose transporter 4, GLUT4)를 유도하여 혈액 속의 포도당을 세포 내로 유입시키도록 작용한다고 알려져 있다. Fagopyritol은 인슐린과 유사한 작용을 하는 것으로 알려져 있으므로, 본 연구에서는 혈당강하 효과가 있다고 알려진 fagopyritol을 랫드의 근육세포주(L6GLUT4myc 세포)에 처리하여, 아직 명확하게 밝혀지지 않은 fagopyritol의 혈당강하 기전을 규명하고자 수행하였다. Fagopyritol의 혈당강하 기전을 규명하기 위하여 근원세포(myoblast)와 근관세포(myotube)에 fagopyritol을 처리하여 액틴 필라멘트의 구조와 GLUT4에 미치는 영향을 분석하였다. Fagopyritol을 myoblast에 처리하였을 때, GLUT4가 처리군에서 대조군과 비교하여 유의 있게 원형질막 쪽으로 유도되는 것을 확인하였고, 액틴 필라멘트의 구조가 재조정되면서 GLUT4의 이동을 돕는 것으로 생각된다. 또한 fagopyritol이 인슐린과 유사한 작용 경로를 가지는지 확인하기 위하여, 인슐린 작용 경로에서 중요한 역할을 하는 것으로 알려진 phosphatidylinositol 3-kinase (PI3K)의 억제제인 LY294002를 fagopyritol과 함께 처리하였을 때 GLUT4가 원형질막 쪽으로 유도되지 않는 것을 확인하였다. Fagopyritol을 myotube에 처리하였을 때, myoblast에 처리하였을 때와 유사한 결과를 나타내었다. 이러한 결과를 종합하면 fagopyritol이 인슐린과 유사한 작용을 하여 액틴 필라멘트의 구조 변경과 GLUT4의 이동을 촉진시키는 것으로 사료된다.

• Journal of Applied Biological Chemistry
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• 제60권4호
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• pp.373-381
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• 2017

Hippocampus abdominalis, the big belly sea horse, is widely known for its medicinal value in Chinese folk medicine. In this study, extract obtained by proteolytic degradation of this species was investigated for its effects on skeletal muscle development, both in vitro and in vivo. Muscle cell lines ($C_2C_{12}$ and $L_6$) treated with the bioactive peptide did not have any detrimental effects on the cell viability, which was above 80%. Optical microscopy analysis on the morphology of the sea horse extract (SHE)-treated cells showed enhanced differentiating ability with myotube formation. Moreover, cells incubated with the hydrolysate displayed decreased proliferation rate, as recorded by the electric cell substrate impedance sensing system, thereby supporting enhanced differentiation. For a period of 12 weeks, mice models were fed with SHE and simultaneously subjected to treadmill exercise, which increased the expression of Myogenin, a key myogenic regulatory factor. In addition, there was an increase in the expression of AMPK- and Cytochrome C, both of which are important in mitochondrial biogenesis. Thus, the SHE from Hippocampus abdominalis can be a promising candidate as protein supplement aiding muscle development.

• Journal of Microbiology and Biotechnology
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• 제33권7호
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• pp.915-925
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• 2023

Sarcopenia is defined as loss of muscle mass and strength due to aging. Recent studies show that sarcopenia may improve via the gut-muscle axis, suggesting that gut health may affect muscle phenotypes. In this study, we aimed to investigate the ability of Lactobacillus rhamnosus JY02 as a probiotic strain isolated from kimchi to alleviate sarcopenia. L. rhamnosus JY02-conditioned medium (CM) reduced dexamethasone (DEX)-induced myotube diameter atrophy and expression of muscle degradation markers (MuRF1 and atrogin-1) in C2C12 cells. The amelioration of sarcopenia was investigated by measuring body composition (lean mass), hand grip strength, myofibril size (using histological analysis), and mRNA and protein expression of muscle-related factors in a DEX-induced mouse model. The results of these analyses showed that L. rhamnosus JY02 supplementation promoted the production of muscle-enhancement markers (MHC Iβ, MHC IIα, and Myo-D) and reduced both the production of muscle degradation markers and the symptoms of muscle atrophy (loss of lean mass and muscle strength). We also found decreased levels of pro-inflammatory cytokines (IL-6, IFN- γ) and increased levels of anti-inflammatory cytokines (IL-10) in the serum of DEX+JY02-administered mice compared to those in DEX-treated mice. Overall, these results suggest that L. rhamnosus JY02 is a potent probiotic supplement that prevents sarcopenia by suppressing muscle atrophy.

• The Korean Journal of Physiology and Pharmacology
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• 제22권6호
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• pp.697-703
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• 2018

Myoblast fusion depends on mitochondrial integrity and intracellular $Ca^{2+}$ signaling regulated by various ion channels. In this study, we investigated the ionic currents associated with $[Ca^{2+}]_i$ regulation in normal and mitochondrial DNA-depleted(${\rho}0$) L6 myoblasts. The ${\rho}0$ myoblasts showed impaired myotube formation. The inwardly rectifying $K^+$ current ($I_{Kir}$) was largely decreased with reduced expression of KIR2.1, whereas the voltage-operated $Ca^{2+}$ channel and $Ca^{2+}$-activated $K^+$ channel currents were intact. Sustained inhibition of mitochondrial electron transport by antimycin A treatment (24 h) also decreased the $I_{Kir}$. The ${\rho}0$ myoblasts showed depolarized resting membrane potential and higher basal $[Ca^{2+}]_i$. Our results demonstrated the specific downregulation of $I_{Kir}$ by dysfunctional mitochondria. The resultant depolarization and altered $Ca^{2+}$ signaling might be associated with impaired myoblast fusion in ${\rho}0$ myoblasts.

• 한국식품영양과학회지
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• 제46권1호
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• pp.39-45
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• 2017

본 연구에서는 생산지역별 감초의 물질 표준화를 위해 주요지표성분 중 glycyrrhizic acid를 지표물질로 선정하고 정량분석을 위해 분석법 검증을 수행하였다. 감초의 glycyrrhizic acid의 분석법 검증은 특이성, 직선성, 검출 및 정량한계, 정확성, 정밀성을 구하여 확립하였다. 확립된 분석 방법으로 시험한 국내외 생산지역별 감초의 glycyrrhizic acid 정량 분석 결과 국내산 중에서는 제천산이, 수입산 중에서는 우즈베키스탄산이 각각 16.98, 54.22 mg/g으로 높은 함량을 나타내었다. 또한, 현대인의 심각한 대사질환 중 하나인 당뇨병 예방 및 개선에 효과적인 기능성 천연물 소재를 발굴하고 항당뇨 효과 검증을 위해 생산지역별 감초 에탄올 추출물의 ${\alpha}-glucosidase$ 저해 활성 및 glucose uptake 활성을 평가하였다. ${\alpha}-Glucosidase$ 저해 활성은 국내외 5종 중 제천, 곡성지역의 감초에서 무처리군 대비 높은 저해 활성을 보였고, 우즈베키스탄산은 유의적인 활성이 나타나지 않았다. Glucose uptake 활성의 경우 인슐린 처리를 한 대조군 대비 제천, 영주, 우즈베키스탄 지역의 감초에서 각각 24.19, 27.18, 26.92% 유의적으로 증가함을 확인하였다. 따라서 본 연구의 지표성분 분석, 분석법 검증 결과 및 생리활성 평가는 생산지역별 감초의 물질 표준화 및 품질 평가의 기초자료를 제공하는 데 큰 의미가 있으며, 감초가 대사질환인 당뇨 예방 및 개선에 효과적인 생약식품 소재로의 개발 가능성을 확보하였다. ${\alpha}-Glucosidase$ 저해 활성 및 glucose uptake 활성 평가뿐만 아니라 감초에 함유된 많은 물질연구를 통해 알려지지 않은 활성에 관한 계속된 추후 연구가 필요할 것으로 보인다.