• Nutrition Research and Practice
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• 제13권4호
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• pp.279-285
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• 2019

BACKGROUND/OBJECTIVES: Excessive production of reactive oxygen species (ROS) such as hydroxyl (${\cdot}OH$), nitric oxide (NO), and hydrogen peroxide ($H_2O_2$) is reported to induce oxidative stress. ROS generated by oxidative stress can potentially damage glial cells in the nervous system. Cordyceps militaris (CM), a kind of natural herb widely found in East Asia. In this study, we investigated the free radical scavenging activity of the CM extract and its neuroprotective effects in $H_2O_2$-induced C6 glial cells. MATERIALS/METHODS: The ethanol extract of CM ($100-1,000{\mu}g/mL$) was used to measure DPPH, ${\cdot}OH$, and NO radical scavenging activities. In addition, hydrogen peroxide ($H_2O_2$)-induced C6 glial cells were treated with CM at $0.5-2.5{\mu}g/mL$ for measurement of cell viability, ROS production, and protein expression resulting from oxidative stress. RESULTS: The CM extract showed high scavenging activities against DPPH, ${\cdot}OH$, and NO radicals at concentration of $1,000{\mu}g/mL$. Treatment of CM with $H_2O_2$-induced oxidative stress in C6 glial cells significantly increased cell viability, and decreased ROS production. Cyclooxygenase-2 and inducible nitric oxide synthase protein expression was down-regulated in CM-treated groups. In addition, the protein expression level of phospho-p38 mitogen-activated protein kinase (p-p38 MAPK), phospho-c-Jun N-terminal kinase (p-JNK), and phospho-extracellular regulated protein kinases (p-ERK) in $H_2O_2$-induced C6 glial cells was down-regulated upon CM administration. CONCLUSION: CM exhibited radical scavenging activity and protective effect against $H_2O_2$ as indicated by the increased cell viability, decreased ROS production, down-regulation of inflammation-related proteins as well as p-p38, p-JNK, and p-ERK protein levels. Therefore, we suggest that CM could play the protective role from oxidative stress in glial cells.

• 대한한의학회지
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• 제22권3호
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• pp.1-10
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• 2001

Objectives : The water extract of Nueihyuljunbang (NHJB) has long been used for treatment of ischemic brain damage in Oriental Medicine. However, little is known about the mechanism by which the water extract of NHJB recovers brain cens from ischemic damage. Methods : To elucidate the protective mechanism on ischemic induced cytotoxicity, we investigated the regulation of lipopolysaccharide (LPS) and phorbol-12-myristate-13-acetate (PMA)-induced inducible nitric oxide synthase (iNOS) expression in C6 glial cells. Results : LPS combined PMA treatment for 72 hours in C6 glial cells markedly induced nitric oxide (NO), but treatment of the cells with the water extract of NHJB decreased dose-dependently nitrite formation. In addition, LPS combined PMA treatment for 72 hours induced severe celt death and lactate dehydrogenase (LDH) release in C6 glial cells. However, treatment of the celts with the water extract of NHJB did not induce significant change compared to control cells. Furthermore, the protective effects of the water extract of NHJB were mimicked by the treatment of NGMMA, a specific inhibitor of NOS. LPS combined PMA induced iNOS activation in C6 glial cells caused chromosomal condensation and fragmentation of the nuclei by caspase activation. The treatment of C6 glial cells with the water extract of NHJB might suppress apoptosis via caspase inhibition by regulation of iNOS expression. Conclusions : From the results, we suggest that the protective effects of the water extract of NHJB against ischemic brain damage may be mediated by regulation of iNOS during ischemic condition.

• 동의생리병리학회지
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• 제23권6호
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• pp.1368-1378
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• 2009

Unfolded protein response (UPR) is an important genomic response to endoplasmic reticulum (ER) stress. The ER response is characterized by changes in specific proteins, induction of ER chaperones and degradation of misfolded proteins. Also, the pathogenesis of several diseases like Alzheimer's disease, neuronal degenerative diseases, and diabetes reveal the role of ER stress as one of the causative mechanisms. Borneolum has been used for neuronal disease in oriental medicine. In the present study, the protective effect of borneolum on thapsigargin-induced apoptosis in rat C6 glial cells. Treatment with C6 glial cells with 5 uM thapsigargin caused the loss of cell viability, and morphological change, which was associated with the elevation of intracellular $Ca^{++}$ level, the increase in Grp78 and CHOP and cleavage of pro-caspase 12 Furthermore, thapsigargin induced Grp98, XBP1, and ATF4 protein expression in C6 glial cells. Borneolum reduced thapsigargin-induced apoptosis through ER pathways. In the ER pathway, borneolum attenuated thapsigargin-induced elevations in Grp78, CHOP, ATF4, and XBP1 as well as reductions in pro-caspase 12 levels. Also, our data showed that borneolum protected thapsigargin-induced cytotoxicity in astrocytes from rat (P3) brain. Taken together, our data suggest that borneolum is neuroprotective against thapsigargin-induced ER stress in C6 glial cells and astrocytes. Accordingly, borneolum may be therapeutically useful for the treatment of thapsigargin-induced apoptosis in central nervous system.

• 대한한방내과학회지
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• 제28권3호
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• pp.586-596
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• 2007

Objectives : This study aimed to investigate the underlying protective mechanism of Rhizoma Arisaematis(RA) on 3-NP-induced Cytotoxicity in rat C6 glial cells. Methods : We investigated treatment ofC6 cells with 20mM 3-NP and pretreatment with RA to cause loss of cell viability. and morphological change. which was associated with elevation of ROS level. increase in Bax/Bcl2 ratio and HIF-a protein expression Results : RA inhibited 3-NP-induced cell death in C6 glial cells and inhibited the changes of the : MMPT (mitochondria membrane potential transition) and inhibited the decrease of mitochondria complex II activity and 3-NP-induced ROS generation in C6 cells. And RA decreased the activity of HIF-a and Bax. and increased the activity of $Bcl_2$ in C6 glial cells Conclusions : RA markedly protects C6 glial cells from 3-NP-induced oxidative injury.

• 한국자원식물학회지
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• 제28권6호
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• pp.675-681
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• 2015

β amyloid protein (Aβ) plays a critical role in the pathogenesis of Alzheimer's disease (AD) and possibly in Aβ-induced mitochondrial dysfunction and oxidative stress. Aβ can directly cause reactive oxygen species (ROS) production. Overproduction of ROS is considered to be involved in the pathogenesis of neurodegeneration of AD. Here, we investigated 9 kinds of ramie (Boehmeria nivea, (L.) Gaud., BN; hereafter denoted as BN) for their protective action against oxidative stress in a cellular system using C6 glial cells. We observed loss of cell viability and high levels of ROS generation after treatment with hydrogen peroxide (H₂O₂) and Aβ_25-35. However, treatments with BN extracts led to an increase in cell viability and decrease in ROS production induced by H2O2 and Aβ25-35. In particular, the extracts of BN-01 (seobang variety from Seocheon) and BN-09 (local variety from Yeonggwang) showed excellent anti-oxidative properties. This indicates that BN extracts could prevent neurodegeneration by reducing oxidative stress in cells.

• Biomolecules & Therapeutics
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• 제24권3호
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• pp.338-345
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• 2016

Neurodegenerative diseases are often associated with oxidative damage in neuronal cells. This study was conducted to investigate the neuro-protective effect of methanolic (MeOH) extract of Perilla frutescens var. japonica and its one of the major compounds, rosmarinic acid, under oxidative stress induced by hydrogen peroxide ($H_2O_2$) in C6 glial cells. Exposure of C6 glial cells to $H_2O_2$ enhanced oxidative damage as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and thiobarbituric acid-reactive substance assays. The MeOH extract and rosmarinic acid prevented oxidative stress by increasing cell viability and inhibiting cellular lipid peroxidation. In addition, the MeOH extract and rosmarinic acid reduced $H_2O_2-indcued$ expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the transcriptional level. Moreover, iNOS and COX-2 protein expression was down-regulated in $H_2O_2-indcued$ C6 glial cells treated with the MeOH extract and rosmarinic acid. These findings suggest that P. frutescens var. japonica and rosmarinic acid could prevent the progression of neurodegenerative diseases through attenuation of neuronal oxidative stress.

• Nutrition Research and Practice
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• 제14권6호
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• pp.593-605
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• 2020

BACKGROUND/OBJECTIVES: Alzheimer's disease is common age-related neurodegenerative condition characterized by amyloid beta (Aβ) accumulation that leads cognitive impairment. In the present study, we investigated the protective effect of paeoniflorin (PF) against Aβ-induced neuroinflammation and the underlying mechanism in C6 glial cells. MATERIALS/METHODS: C6 glial cells were treated with PF and Aβ_25-35, and cell viability, nitric oxide (NO) production, and pro-inflammatory cytokine release were measured. Furthermore, the mechanism underlying the effect of PF on inflammatory responses and Aβ degradation was determined by Western blot. RESULTS: Aβ_25-35 significantly reduced cell viability, but this reduction was prevented by the pretreatment with PF. In addition, PF significantly inhibited Aβ_25-35-induced NO production in C6 glial cells. The secretion of interleukin (IL)-6, IL-1β, and tumor necrosis factor-alpha was also significantly reduced by PF. Further mechanistic studies indicated that PF suppressed the production of these pro-inflammatory cytokines by regulating the nuclear factor-kappa B (NF-κB) pathway. The protein levels of inducible NO synthase and cyclooxygenase-2 were downregulated and phosphorylation of NF-κB was blocked by PF. However, PF elevated the protein expression of inhibitor kappa B-alpha and those of Aβ degrading enzymes, insulin degrading enzyme and neprilysin. CONCLUSIONS: These findings indicate that PF exerts protective effects against Aβ-mediated neuroinflammation by inhibiting NF-κB signaling, and these effects were associated with the enhanced activity of Aβ degradation enzymes.

• 대한한방내과학회지
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• 제41권1호
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• pp.44-58
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• 2020

Objectives: This study was conducted to identify the protective effects of Chongmyunggongjin-dan (CMGJD) on Hydrogen peroxide (H₂O₂)-induced apoptosis mechanisms in C6 glial cells. Method: We used CMGJD after distilled water extraction, filtration, and lyophilization. The ROS scavenging effect was examined by fluorescence microscopy. Expression levels of proteins related to ROS generation were investigated by western blotting. Functional changes in organelles related to Reactive oxygen species (ROS) generation were investigated by immunoblotting and by verifying expression level of relevant enzymes. Results: The CMGJD extract protected the cells against H₂O₂-induced morphological changes and DNA fragmentation, inhibited the increase of Heme_oxygenase-1(HO^-1) and the decrease in catalase, protected against the loss of mitochondrial membrane potential, inhibited disturbances of lysosomal function, and induced an increase in peroxisomes. Conclusion: CMGJD was confirmed to have a protective effect on H₂O₂-induced C6 glial cell death possibly by blocking the pathways causing damage to subcellular organelles, such as mitochondria, lysosomes, and peroxisomes. We assume that CMGJD will be effective for the prevention and treatment of ischemic stroke in a clinical environment.

• Journal of Applied Biological Chemistry
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• 제63권2호
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• pp.137-145
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• 2020

Oxidative stress is one of the pathogenic mechanisms of various neurodegenerative diseases, such as Alzheimer's disease. Neuroglia, the most abundant cells in the brain, is thought to play an important role in the antioxidant defense system and neuronal metabolic support against neurotoxicity and oxidative stress. We investigated the protective effect of paeoniflorin (PF) against oxidative stress in C6 glial cells. Exposure of C6 glial cells to hydrogen peroxide (H₂O₂, 500 μM) significantly decreased cell viability and increased amounts of lactate dehydrogenase (LDH) release, indicating H₂O₂-induced cellular damage. However, treatment with PF significantly attenuated H₂O₂-induced cell death as shown by increased cell survival and decreased LDH release. The H₂O₂-stimulated reactive oxygen species production was also suppressed, and it may be associated with improvement of superoxide dismutase activity by treatment with PF. In addition, an increase in ratio of Bcl-2/Bax protein expression was observed after treatment with PF. In particular, the down-stream of the apoptotic signaling pathway was inhibited in the presence of PF, mostly by reduction of cleaved-poly ADP ribose polymerase, cleaved caspase-3, and -9 protein expression. Furthermore, H₂O₂-induced phosphorylation of c-Jun N-terminal kinase and extracellular signal-regulated kinase 1/2 was attenuated by treatment with PF. Taken together, neuroprotective effect of PF against oxidative stress probably result from the regulation of apoptotic pathway in C6 glial cells. In conclusion, our findings suggest that PF may be a potent therapeutic agent for neurodegenerative disorders.

• 대한한방내과학회지
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• 제21권3호
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• pp.467-475
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• 2000

The water extracts of Jagamcho-tang has been used for treatment of arrhythmia and palpitation in oriental traditional medicine. Brain is provided with blood flow by heart. Jagamcho-tang has been studied on ischemia and infarction in heart. However, little is known about the mechanism by which the water extracts of Jagamcho-tang rescues brain cells from ischemic damages. To elucidate the protective mechanism on ischemic induced cytotoxicity, the effects of Jagamcho-tang on ischemia induced cytotoxicity and generation of nitric oxide(NO) are investigated in C6 glioma cells. Jagamcho-tang induce NO in a dose dependent manner up to 2.5mg/ml in C6 glioma cells. The pretreatment of Jagamcho-tang protect sodium nitroprusside(SNP) (2mM) induced cytotoxicity. This effect of Jagamcho-tang is mimicked by treatment by pretreatment of SNP($100{\mu}M$), an exogenous NO donor. NG-monomethyl-L-arginine($N^{G}MMA$), a specific inhibitor of nitric oxide synthase (NOS), significantly blocks the protective effects of Jagamcho-tang on cell toxicity by ischemia. In addition, lipopolysaccharide(LPS) and phorhol 12 myristate 13-acetate(PMA) treatment for 72h in C6 glial cells markedly induce NO, but treatment of the cells with the water extracts of Jagamcho-tang decrease nitrite formation in a dose dependent manner. In addition, LPS and PMA treatment for 72h induce severe cell death and LDH release into medium in C6 glial cells. However treatment of the cells with the water extracts of Jagamcho-tang dose not induce significant changes compare to control cells. Furthermore, the protective effects of the water extracts of Jagamcho-tang is mimicked by treatment of $N^{G}MMA$. Taken together, I suggest that the protective effects of the water extracts of Jagamcho-tang against ischemic brain damages may be mediated by regulation of iNOS during ischemic condition.