• 동의생리병리학회지
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• 제21권1호
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• pp.190-198
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• 2007

Alzheimer's disease, a representative neurodegenerative disorder, is characterized by the presence of senile plaques and neurofibrillary tangles accompanied by neuronal damages. b-Amyloid peptide is considered to be responsible for the formation of senile plagues that accumulate in the brains of patients with Alzheimer's disease. There has been compelling evidence supporting that b-amyloid-induced cytotoxicity is mediated through generation of reactive oxygen species. In this study, we have investigated the possible protective effect of Rehmannia glutihosaagainst b-amyloid-induced oxidative ceil death in cultured human neuroblastoma SH-SY5Y cells. SH-SY5Y cells treated with b-amyloid underwent apoptotic death as determined by morphological features and positive in situterminal end-labeling (TUNEL staining). Rehmannia glutinosawater extract, wine, and vinegar pretreatments attenuated b-amyloid-induced cytotoxicity and apoptosis. Rehmannia glutinosa vinegar exhibited maximum protective effect by increasing the expression of anti-apoptotic protein, Bcl-2. in addition to oxidative stress, b-amyloid-treatment caused nitrosative stress via marked increase in the levels of nitric oxide, which was effectively blocked by Rehmannia glutinosa. To further explore the possible molecular mechanisms underlying the protective effect of Rehmannia glutinosa, we assessed the mRNA expression of cellular antioxidant enzymes. Treatment of Rehmannia glutinosa vinegar led to up-regulation of heme oxygemase-1 and catalase. These results suggest that Rehmannia glutinosa could modulate oxidative neuronal cell death caused by b-amyloid and may have preventive or therapeutic potential in the management of Alzheimer's disease. Particularly, Rehmannia glutinosa vinegar can augment cellular antioxidant capacity, there by exhibiting higher neuroprotective potential.

• 대한예방한의학회지
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• 제16권2호
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• pp.95-111
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• 2012

Objective : The water extract of Danguiyonghoihwan (DGYHW) has been traditionally used in treatment of tinnitus in Oriental Medicine. However, little is known about the mechanism by which DGYHW rescues auditory neuronal cells from injury damages. Therefore, in this study I effort to elucidate the mechanism of the cytoprotective effect of the DGYHW extract on glutamate-induced auditory sensorineuronal cell death. Methods : I determined the elevated cell viability by DGYHW extract on glutamate-induced auditory sensorineuronal cell death. Glutamate induced neuronal damage in oranotypic explant culture also, glutamate decreased cell viability on VOT-33 cells but pretreatment with DGYHW inhibited cell death. Results : One of the main mediator of glutamate-induced cytotoxicity was known to generation of reactive oxygen species (ROS). Pretreatment with DGYHW inhibited this ROS generation from glutamated-stimulated VOT-33 cells. Also, I identified that the ROS-induced DCF-DA green fluorescence is reduced by DGYHW pretreatment. The critical markers of apoptotic cell death were cleavages of procaspase-3 protease protein. So I checked the expression level and cleavage of procaspase-3 protease protein. Glutamate-treated VOT-33 cells were shown to have cleavage of procaspase-3 protease proteins and following reduction of expression of these proteins. But I found that pre-treatment with DGYHW protects glutamate-induced changes of biochemical marker protein, caspase-3. Conclusion : These findings indicated that DGYHW may prevent cell death from glutamate induced VOT-33 cell death by inhibiting the ROS generation and modulation of protein expressions in procaspase-3, catalase and Bcl-2.

• The Korean Journal of Physiology and Pharmacology
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• 제1권6호
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• pp.699-705
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• 1997

Although it is known that neuronal cell death during development occurs by apoptosis, the mechanisms underlying excitatory amino acid-induced neuronal cell death remain poorly understood. In this study we have examined the mechanism by which L-glutamate, an excitatory amino acid neurotransmitter, induces cell death in PC12 cell lines. To characterize cell death, we employed sandwich enzyme-linked immunosorbent assay(ELISA) method for cellular DNA fragmentation, DNA agarose gel electrophoresis and chromatin staining by acridine orange and ethidium bromide after treating the PC12 cells with L-glutamate. L-Glutamate caused dose-dependent cell death with a maximum at 24 hrs after the treatment. These cellular fragmentation was blocked by pretreatment of MK-801, a noncompetitive N-methyl-D-aspartic acid(NMDA) receptor antagonist, and nerve growth factor(NGF). Analysis of DNA integrity from L-glutamate-treated cells revealed cleavage of DNA into regular sized fragments, a biochemical hallmark of apoptosis. The PC12 cells that were induced to die by L-glutamate treatment exhibited classical chromatin condensation under the light microscopy after acridine orange and ethidium bromide staining. These results suggest that apoptosis is one of the key features that are involved in L-glutamate-induced excitotoxic cell death in PC12 cells, and these cell death are mediated by NMDA receptor and depend on NGF.

• 대한한의학회지
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• 제29권5호
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• pp.29-40
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• 2008

Objectives : It has been reported that Sophorae Subprostratae Radix (SSR) has a neuroprotective effect on cerebral ischemia in animals. In the present study, the authors investigated the neuroprotective effect of SSR on glutamate excitotoxicity. Glutamate excitotoxicity was induced by using NMDA, AMPA, and KA in PC12 cells and in organotypic hippocampal slice cultures. Methods :Methanolic extract of SSR was added at 0.5, 5, and 50 ${\mu}$g/ml to culture media for 24 hours. The effects of SSR were evaluated by measuring of cell viability, PI-stained neuronal cell death, TUNEL-positive cells, and MAP-2 immunoreactivity. Results : SSR increased PC12 cell viabilities significantly against AMPA-induced excitotoxicity, but not against NMDA-induced or KA-induced excitotoxicity. In organotypic hippocampal slice cultures damaged by NMDA-induced excitotoxicity, SSR attenuated neuronal cell death significantly in the CA1, CA3, and DG hippocampal regions and reduced TUNEL-positive cells significantly in CA1 and DG regions. In organotypic hippocampal slice cultures damaged by AMPA-induced excitotoxicity, SSR attenuated neuronal cell death and reduced TUNEL-positive cell numbers significantly in the CA1 and DG regions. In organotypic hippocampal slice cultures damaged by KA-induced excitotoxicity, SSR attenuated neuronal cell death significantly in CA3, but did not reduce TUNEL-positive cell numbers in CA1, CA3 or DG. In organotypic hippocampal slice cultures damaged by NMDA-induced excitotoxicity, SSR attenuated pyramidal neuron neurite retraction and degeneration in CA1. Conclusions : These results suggest that the neuroprotective effects of SSR are related to antagonistic effects on the NMDA and AMPA receptors of neuronal cells damaged by excitotoxicity and ischemia.

• The Korean Journal of Physiology and Pharmacology
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• 제10권4호
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• pp.167-172
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• 2006

In the present study, we developed a simple method to predict the neuronal cell death in the mouse hippocampus and striatum following transient global forebrain ischemia by evaluating both cerebral blood flow and the plasticity of the posterior communicating artery (PcomA). Male C57BL/6 mice were anesthetized with halothane and subjected to bilateral occlusion of the common carotid artery (BCCAO) for 30 min. The regional cerebral blood flow (rCBF) was measured by laser Doppler flowmetry. The plasticity of PcomA was visualized by intravascular perfusion of India ink solution. When animals had the residual cortical microperfusion less than 15% as well as the smaller PcomA whose diameter was less than one third compared with that of basilar artery, neuronal damage in the hippocampal subfields including CA1, CA2, and CA4, and in the striatum was consistently observed. Especially, when mice met these two criteria, marked neuronal damage was observed in CA2 subfield of the hippocampus. In contrast, after transient BCCAO, neuronal damage was consistently produced in the striatum, dependent more on the degree of rCBF reduction than on the plasticity of PcomA. The present study provided simple and highly reproducible criteria to induce the neuronal cell death in the vulnerable mice brain areas including the hippocampus and striatum after transient global forebrain ischemia.

• BMB Reports
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• 제46권7호
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• pp.370-375
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• 2013

Ischemia is characterized by oxidative stress and changes in the antioxidant defense system. Our recent in vitro study showed that 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride protects cortical astrocytes against oxidative stress. In the current study, we examined the effects of 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride on ischemia-induced neuronal damage in a gerbil ischemia/reperfusion models. Extensive neuronal death in the hippocampal CA1 area was observed 4 days after ischemia/reperfusion. Intraperitoneal injection of 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride (0.3 mg/kg body weight) significantly prevented neuronal death in the CA1 region of the hippocampus in response to transient forebrain ischemia. 2-Cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride administration reduced ischemia-induced increases in reactive oxygen species levels and malondialdehyde content. It also attenuated the associated reductions in glutathione level and superoxide dismutase, catalase, and glutathione peroxidase activities. Taken together, our results suggest that 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride protects against ischemia-induced neuronal damage by reducing oxidative stress through its antioxidant actions.

• 대한한의학회지
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• 제39권1호
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• pp.35-43
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• 2018

Objectives: In the brain, glutamate is the most important excitable neurotransmitter in physiological and pathological conditions. However, the high level of glutamate induces neuronal cell death due to exitotoxicity and oxidative stress. The present study investigated to evaluate a possible neuroprotective effect of furosin isolated from Euphorbia helioscopia against glutamate-induced HT22 cell death. Methods: Furosin was isolated from methanol extract of Euphorbia helioscopia and examined whether it protects glutamate-induced neuronal cell death. The cell viability was determined using Ez-Cytox assay. Anti-oxidative effect of furosin was determined by DPPH scavenging activities, and the levels of intracellular reactive oxygen species (ROS) were determined by the fluorescent intensity after staining the cells with $H_2DCFDA$. To evaluate apoptotic cell death, we performed nuclear staining and image-based cytometeric analysis. Results: The cell viability was significantly increased by treatement with furosin compared with the treatment with glutamate. Furosin showed a strong DPPH radical scavenging activity ($EC50=1.83{\mu}M$) and prevented the accumulation of intra cellular ROS. Finally, the presence of 50 and $100{\mu}M$ furosin significantly the percentage of apoptotic cells compared with glutamate treatment. Conclusion: The present study found that furosin is a potent neuroprotectant against glutamate-induced oxidative stress through inhibition of apoptotic cell death induced by glutamate. Therefore, the present study suggests that furosin as a bioactive compound of E. helioscopia can be a useful source to develop a drug for the treatment of neurodegenerative diseases and acute brain injuries.

• Biomolecules & Therapeutics
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• 제29권3호
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• pp.321-330
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• 2021

Oxidative stress plays a crucial role in the development of neuronal disorders including brain ischemic injury. Thioredoxin 1 (Trx1), a 12 kDa oxidoreductase, has anti-oxidant and anti-apoptotic functions in various cells. It has been highly implicated in brain ischemic injury. However, the protective mechanism of Trx1 against hippocampal neuronal cell death is not identified yet. Using a cell permeable Tat-Trx1 protein, protective mechanism of Trx1 against hydrogen peroxide-induced cell death was examined using HT-22 cells and an ischemic animal model. Transduced Tat-Trx1 markedly inhibited intracellular ROS levels, DNA fragmentation, and cell death in H2O2-treatment HT-22 cells. Tat-Trx1 also significantly inhibited phosphorylation of ASK1 and MAPKs in signaling pathways of HT-22 cells. In addition, Tat-Trx1 regulated expression levels of Akt, NF-κB, and apoptosis related proteins. In an ischemia animal model, Tat-Trx1 markedly protected hippocampal neuronal cell death and reduced astrocytes and microglia activation. These findings indicate that transduced Tat-Trx1 might be a potential therapeutic agent for treating ischemic injury.

• Biomolecules & Therapeutics
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• 제26권2호
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• pp.115-120
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• 2018

Chronic cerebral hypoperfusion (CCH), which is associated with onset of vascular dementia, causes cognitive impairment and neuropathological alterations in the brain. In the present study, we examined the neuroprotective effect of duloxetine (DXT), a potent and balanced serotonin/norepinephrine reuptake inhibitor, on CCH-induced neuronal damage in the hippocampal CA1 region using a rat model of permanent bilateral common carotid arteries occlusion. We found that treatment with 20 mg/kg DXT could attenuate the neuronal damage, the reduction of phosphorylations of mTOR and p70S6K as well as the elevations of $TNF-{\alpha}$ and $IL-1{\beta}$ levels in the hippocampal CA1 region at 28 days following CCH. These results indicate that DXT displays the neuroprotective effect against CCH-induced hippocampal neuronal death, and that neuroprotective effect of DXT may be closely related with the attenuations of CCH-induced decrease of mTOR/p70S6K signaling pathway as well as CCH-induced neuroinflammatory process.

• Natural Product Sciences
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• 제15권3호
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• pp.125-129
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• 2009

Neurodegenerative diseases such as Alzheimer's disease, stroke, and Parkinson's disease, are caused by neuronal cell death. Apoptosis, oxidative stress, inflammation, excitotoxicity or ischemia are discussed to play a role of neuronal cell death. In order to find the candidate of neuroprotective agent, neuroprotective activity of some medicinal plants was investigated with in vitro assay system using glutamate-induced neurotoxicity in primary cultures of rat cortical cells. The aqueous methanolic extracts of twenty-seven medicinal plants were evaluated the protective effects against glutamate-injured excitotoxicity in rat cortical cells at the concentration of 50 $\mu$g/ml and 100 $\mu$g/ml, respectively. Among them, extracts of Lonicera japonica, Taraxacum platycarpum, Polygonum aviculare, Gardenia jasminoides, Forsythia viridissima, Lygodium japonicum, Panax notoginseng, Akebia quinata, Anemarrhena asphodeloides and Phellodendron amurense showed significantly neuroprotective activities against glutamate-induced neurotoxicity in primary rat cortical cells.