• Journal of Life Science
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• v.34 no.1
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• pp.37-47
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• 2024

FUN14 domain-containing protein 1 (FUNDC1), an outer mitochondrial membrane protein, contributes to removal of damaged mitochondria through mitophagy. In this study, to elucidate the role of the FUNDC1 in the amyloid beta peptide (A_β)-induced neuropathy, changes in the degree of mitochondrial dysfunction and cell injury caused by A_β treatment were examined in the HT-22 neuronal cells in which the FUNDC1 expression was transiently silenced or overexpressed. We found that A_β treatment causes a time-dependent decrease of the FUNDC1 expression. In the A_β-treated cells, there were a drop in MTT reduction ability, depletion of cellular ATP, disruption of mitochondrial membrane potential, stimulation of cellular ROS production, and increased mitochondrial Ca²⁺ load. Activation of caspase-3 and induction of apoptotic cell death were also observed. Transient silencing of the FUNDC1 expression by transfection with the FUNDC1 small interfering RNA per se caused mitochondrial dysfunction and apoptotic cell death like the effect of A_β treatment. Conversely, in cells in which the FUNDC1 was transiently overexpressed by FUNDC1-Myc transfection, overexpression itself had no effect on the mitochondrial functional integrity and cell survival but showed a significant prevention effect against mitochondrial and cell injury caused by A_β treatment. Overall, these results suggest that the FUNDC1 is importantly involved in the A_β-induced mitochondrial dysfunction and cell injury in the HT-22 neuronal cells.

• Biomolecules & Therapeutics
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• v.27 no.2
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• pp.145-151
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• 2019

Methamphetamine (METH) acts strongly on the nervous system and damages neurons and is known to cause neurodegenerative diseases such as Alzheimer's and Parkinson's. Flavonoids, polyphenolic compounds present in green tea, red wine and several fruits exhibit antioxidant properties that protect neurons from oxidative damage and promote neuronal survival. Especially, epicatechin (EC) is a powerful flavonoid with antibacterial, antiviral, antitumor and antimutagenic effects as well as antioxidant effects. We therefore investigated whether EC could prevent METH-induced neurotoxicity using HT22 hippocampal neuronal cells. EC reduced METH-induced cell death of HT22 cells. In addition, we observed that EC abrogated the activation of ERK, p38 and inhibited the expression of CHOP and DR4. EC also reduced METH-induced ROS accumulation and MMP. These results suggest that EC may protect HT22 hippocampal neurons against METH-induced cell death by reducing ER stress and mitochondrial damage.

• Korean Journal of Acupuncture
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• v.38 no.1
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• pp.32-42
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• 2021

Objectives : In this study, we investigated the neuroprotective effects of ethanol extract of Lonicera japonica flower buds (EELJ) on glutamate-induced neurotoxicity in mouse hippocampus-derived neuronal HT22 cells. Methods : After analyzing the cytoprotective effect of EELJ on glutamate in HT22 cells, the inhibitory effect of apoptosis was studied using flow cytometry. In order to analyze the antioxidant efficacy of EELJ, the levels of reactive oxygen species (ROS) and glutathione (GSH) were investigated, and the effects on the activities of superoxide dismutase (SOD) and catalase (CAT) were also analyzed. Furthermore, the effect of EELJ on the expression of apoptosis regulators such as Bax and Bcl-2 in glutamate-treated HT22 cells was investigated. Results : According the current results, pretreatment with EELJ significantly reduced glutamate-induced loss of cell viability and release of lactate dehydrogenase. EELJ also markedly attenuated glutamate-induced generation of intracellular ROS, which was associated with increased levels of GSH, and activity of SOD and CAT in glutamate-stimulated HT22 cells. In addition, EELJ was strikingly inhibited glutamate-induced apoptosis in HT22 cells. Furthermore, the expression of pro-apoptotic Bax was increased and the expression of anti-apoptotic Bcl-2 was decreased in glutamate-treated HT22 cells, while in the presence of EELJ, their expressions were maintained at the control levels. Conclusions : These findings indicate that EELJ protects glutamate-induced cytotoxicity in HT22 hippocampal neurons through antioxidant activity. Therefore, although identification of biologically active substances of EELJ and re-evaluation through animal experiments is necessary, this natural substance is a promising candidate for further research in preventing and treating oxidative stress-mediated neurodegenerative diseases.

• The Korea Journal of Herbology
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• v.22 no.2
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• pp.65-72
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• 2007

Objectives : In this study, we investigate that Euphorbiae lathyridis Semen extract contributes to growth inhibitory effect and anti-cancer activity on the HT-29 human colon cancer cells. Methods : Euphorbiae lathyridis Semen was extracted from the Semen of the plant using 80% Methanol. The Euphorbiae lathyridis Semen extract was treated to different concentrations for 24 hr, 4Shr or 72hr. Growth inhibitory effect was analyzed by measuring FACS study and MTT assay. Cell apoptosis was confirmed by surveying caspases cascades activation using Westem blot. Results : Exposure to Euphorbiae lathyridis Semen extract (0.4mg/ml) results in an inhibitory effect on cell growth in HT-29 cells. Growth inhibition by Euphorbiae lathyridis Semen extract in HT-29 cells was related with the inhibition of proliferation and induction of apoptosis. The Euphorbiae lathyridis Semen extract induces DNA fragmentation in HT-29 cells. Furthermore, Euphorbiae lathyridis Semen extract induces cell apoptosis through the activation of caspases-3, caspase-9 and PARP cleavage. Conclusion : Euphorbiae lathyridis Semen extract induces apoptosis in human colon cancer cells, therefore, we suggest that Euphorbiae lathyridis Semen extract can be used as a novel class of anti-cancer drugs.

• The Korean Journal of Physiology and Pharmacology
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• v.22 no.5
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• pp.585-595
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• 2018

Amitriptyline, a tricyclic antidepressant, is commonly used to treat depression and neuropathic pain, but its mechanism is still unclear. We tested the effect of amitriptyline on 5-hydroxytryptamine 3 ($5-HT_3$) receptor currents and studied its blocking mechanism because the clinical applications of amitriptyline overlapped with $5-HT_3$ receptor therapeutic potentials. Using a whole-cell voltage clamp method, we recorded the currents of the $5-HT_3$ receptor when 5-HT was applied alone or co-applied with amitriptyline in cultured NCB-20 neuroblastoma cells known to express $5-HT_3$ receptors. To elucidate the mechanism of amitriptyline, we simulated the $5-HT_3$ receptor currents using Berkeley $Madonna^{(R)}$ software and calculated the rate constants of the agonist binding and receptor transition steps. The $5-HT_3$ receptor currents were inhibited by amitriptyline in a concentration-dependent, voltage-independent manner, and a competitive mode. Amitriptyline accelerated the desensitization of the $5-HT_3$ receptor. When amitriptyline was applied before 5-HT treatment, the currents rose slowly until the end of 5-HT treatment. When amitriptyline was co-applied with 5-HT, currents rose and decayed rapidly. Peak current amplitudes were decreased in both applications. All macroscopic currents recorded in whole cell voltage clamping experiments were reproduced by simulation and the changes of rate constants by amitriptyline were correlated with macroscopic current recording data. These results suggest that amitriptyline blocks the $5-HT_3$ receptor by close and open state blocking mechanisms, in a competitive manner. We could expand an understanding of pharmacological mechanisms of amitriptyline related to the modulation of a $5-HT_3$ receptor, a potential target of neurologic and psychiatric diseases through this study.

• Molecules and Cells
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• v.45 no.4
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• pp.216-230
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• 2022

SERTA domain-containing protein 1 (Sertad1) is upregulated in the models of DNA damage and Alzheimer's disease, contributing to neuronal death. However, the role and mechanism of Sertad1 in ischemic/hypoxic neurological injury remain unclear. In the present study, our results showed that the expression of Sertad1 was upregulated in a mouse middle cerebral artery occlusion and reperfusion model and in HT22 cells after oxygen-glucose deprivation/reoxygenation (OGD/R). Sertad1 knockdown significantly ameliorated ischemia-induced brain infarct volume, neurological deficits and neuronal apoptosis. In addition, it significantly ameliorated the OGD/R-induced inhibition of cell viability and apoptotic cell death in HT22 cells. Sertad1 knockdown significantly inhibited the ischemic/hypoxic-induced expression of p-Rb, B-Myb, and Bim in vivo and in vitro. However, Sertad1 overexpression significantly exacerbated the OGD/R-induced inhibition of cell viability and apoptotic cell death and p-Rb, B-Myb, and Bim expression in HT22 cells. In further studies, we demonstrated that Sertad1 directly binds to CDK4 and the CDK4 inhibitor ON123300 restores the effects of Sertad1 overexpression on OGD/R-induced apoptotic cell death and p-Rb, B-Myb, and Bim expression in HT22 cells. These results suggested that Sertad1 contributed to ischemic/hypoxic neurological injury by activating the CDK4/p-Rb pathway.

• Korean Journal of Pharmacognosy
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• v.51 no.1
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• pp.30-35
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• 2020

We previously reported that caffeic acid isolated from Lonicera japonica showed potent neuroprotective activities against glutamate injured neuronal cell death in primary cortical cells. In this study, we tried to confirm the neuroprotective activity in glutamate injured HT22 cells and elucidate mechanisms of neuroprotective action of caffeic acid. We used glutamate induced HT22 cell death as a bioassay system. The compound decreased reactive oxygen species increased by high concentration of glutamate treatment in HT22 cells. Also, Ca²⁺ concentration was decreased by this compound. This compound made mitochondrial membrane potential maintain to normal condition. This also affected anti-oxidative enzymes and glutathione contents. Treatment of this compound increased not only glutathione reductase and peroxidase to the control level and also amount of glutathione, an endogeneous antioxidant. These experimental results showed that caffeic acid isolated from L. japonica exerted potent neuroprotective activity through the anti-oxidative pathway.

• Korean Journal of Pharmacognosy
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• v.50 no.2
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• pp.118-123
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• 2019

Glutamate acts as an important neurotransmitter in brain. However, high concentration of glutamate showed an excitatory neurotoxicity and resulted to neuronal cell death. Neuronal cell death is known for one of the reason of Alzheimer's disease, a neurodegenerative disease. We tried to find neuroprotective medicinal plants by neuroprotection activity against glutamate injured HT22 cells as a model system. In the course of bioscreening of various medicinal plants, Taraxacum platycarpum extract showed significant neuroprotective activity. We tried to elucidate mechanisms of neuroprotective activity. T. platycarpum extract reduced ROS and intracellular $Ca^{2+}$ concentration increased by glutamate induced neurotoxicity. In addition, mitochondrial membrane potential was restored to the control level. Also, glutathione level, glutathione reductase and glutathione peroxidase activity were increased by T. platycarpum extract treatment. These data suggested that T. platycarpum showed neuroprotective activity via antioxidative activity.

• Korean Journal of Pharmacognosy
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• v.52 no.1
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• pp.19-25
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• 2021

We previously reported that lonicerin isolated from Lonicera japonica methanolic extract had potent neuro-protective activities in neuronal cell death injured by excessive glutamate. In this study, we tried to confirm the neuroprotective activities of L. japonica extract and lonicerin in glutamate injured HT22 cells and establish mechanisms of neuroprotective action of lonicerin. We used HT22 cell death injured by glutamate as a bioassay system. The compound decreased reactive oxygen species increased by excessive glutamate treatment in HT22 cells. Also, Ca2+ concentration was decreased by lonicerin treatment. This compound made mitochondrial membrane potential maintain to normal condition. Lonicerin also increased not only glutathione reductase but also peroxidase to the control level. And this compound increased amount of glutathione, an endogenous antioxidant. These results indicated that lonicerin isolated from L. japonica showed potent neuroprotective activity through the anti-oxidative pathway.

• Korean Journal of Pharmacognosy
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• v.53 no.1
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• pp.1-7
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• 2022

In the previous study, we reported that luteolin isolated from Lonicera japonica methanolic extract had potent neuroprotective activities in neuronal cell death injured by excessive glutamate. In this study, we tried to confirm the neuroprotective activities of luteolin in glutamate injured HT22 cells and establish mechanisms of neuroprotective action of luteolin. We used HT22 cell death injured by glutamate as a bioassay system. Luteolin decreased reactive oxygen species increased by excessive glutamate treatment in HT22 cells. Also, Ca²⁺ concentration was decreased by luteolin treatment. Luteolin made mitochondrial membrane potential maintain to normal condition. It also increased not only glutathione reductase but also peroxidase to the control level. And it increased amount of glutathione, an endogenous antioxidant. These results suggested that luteolin isolated from L. japonica showed potent neuroprotective activity through the anti-oxidative pathway.