• Food Science and Biotechnology
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• v.17 no.2
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• pp.343-348
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• 2008

In this study, antioxidant activity of methanol extract of Glycyrrhiza uralensis Fisch (GUE) against $H_2O_2$-induced DNA damage in human leucocytcs and cell death in PC12 cells was determined. The effect of GUE on $H_2O_2$-induced DNA damage in human leucocytcs was evaluated by the comet assay, where GUE ($1-50\;{\mu}g/mL$) was a dose dependent inhibitor of DNA damage induced by $H_2O_2$. The protective effect of GUE against $H_2O_2$-induced damage on PC12 cells was investigated by MTT reduction assay and lactate dehydrogenase release assay. A marked reduction in cell survival induced by $H_2O_2$ was significantly prevented by $1-50\;{\mu}g/mL$ of GUE. The enzyme activity of caspase-3 was elevated in $H_2O_2$-treated PC12 cells, while preincubation with GUE for 30 min inhibited $H_2O_2$-induced caspase-3 activation in a dose-dependent manner. In conclusion, GUE ameliorates $H_2O_2$-induced DNA damage in human leucocytes and has neuroprotective effect by preventing cell death in PC12 cell, suggesting that GU may be a potential candidate for novel therapeutic agents for neuronal diseases associated with oxidative stress.

• Journal of Ginseng Research
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• v.44 no.3
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• pp.490-495
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• 2020

Background: Ginsenoside Rk1, a saponin component isolated from heat-processed Panax ginseng Meyer, has been implicated in the regulation of antitumor and anti-inflammatory activities. Although our previous studies have demonstrated that ginsenoside Rg3 significantly attenuated the activation of NMDA receptors (NMDARs) in hippocampal neurons, the effects of ginsenosides Rg5 and Rk1, which are derived from heat-mediated dehydration of ginsenoside Rg3, on neuronal NMDARs have not yet been elucidated. Methods: We examined the regulation of NMDARs by ginsenosides Rg5 and Rk1 in cultured rat hippocampal neurons using fura-2-based calcium imaging and whole-cell patch-clamp recordings. Results: The results from our investigation showed that ginsenosides Rg3 and Rg5 inhibited NMDARs with similar potencies. However, ginsenoside Rk1 inhibited NMDARs most effectively among the five compounds (Rg3, Rg5, Rk1, Rg5/Rk1 mixture, and protopanaxadiol) tested in cultured hippocampal neurons. Its inhibition is independent of the NMDA- and glycine-binding sites, and its action seems to involve in an interaction with the polyamine-binding site of the NMDAR channel complex. Conclusion: Taken together, our results suggest that ginsenoside Rk1 might be a novel component contributable to the development of ginseng-based therapeutic treatments for neurodegenerative diseases.

• Journal of Oriental Neuropsychiatry
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• v.19 no.3
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• pp.117-127
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• 2008

Objective: Bee venom (BV) has been used for the treatment of inflammatory diseases such as rheumatoid arthritis and relief of pain in Oriental medicine. The two main components of BV are melittin and phospholipase A2 (PLA2). Of these, melittin, the major active ingredient of BV, has been reported to induce apoptosis and to possess anti tumor effects. Several studies have established that the agents inducing apoptosis in target organs suppress tumorigenesis. As the other component, PLA2 has been reported to induce neurite outgrowth in PC12 cells. However, there was no report about proliferative effect of BV in neuronal cells. In order to examine the effect of BV on glioma cell, human glioma cell line, U87 was used. Methods: Analysis of proliferation was confirmed by MTT assay. BV increased cell number through dose and duration dependent manner and these effects are apparent at a concentration of 10 ug/ml. To observe which signaling molecules will be activated by BV, phosphorylation of Akt, MAPK, PYK2 or CREB were examined by Western blot analysis. To study the long term effect of BV in U87 cells, the image of cells treated with BV for 4 days were obtained. Results: The phosphorylation levels of PYK2 and Akt were increased at 5 min after addition of 10 ug/ml of BV and sustained to 2 hours. On the other hand, phosphorylation of MAPK and CREB were increased at 5 min, maximum at 10 min, and returned to 30 min. These imply that BV may activate two different signaling pathways, PYK2/Akt and MAPK/CREB. BV treated cells showed increased neurite number and length. Conclusion: These results propose that BV may induce differentiation as well as proliferation of U87 cells through the activation of PYK2/ Akt and MAPK/ CREB.

• Biomolecules & Therapeutics
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• v.13 no.4
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• pp.256-262
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• 2005

We examined the signaling molecules involved in the 6-hydroxydopamine (6-OHDA)-induced neuronal cell death and increase in cellular glutathione (GSH) level in SK-N-SH cells. The 6-OH-DA-induced cell death was significantly prevented by the pretreatment with N-acetylcysteine (NAC), a thiol antioxidant, and BAPTA, an intracellular $Ca^{2+}$ chelator. Although 6-OHDA induced ERK phosphorylation, the pretreatment with PD98059, an ERK inhibitor, did not block 6-OHDA-induced cell death. In addition, the 6-OHDA-induced activation of caspase-3, a key signal for apoptosis, was blocked by the pretreatment with NAC and BAPTA. While the level of reactive oxygen species (ROS) was significantly increased in the 6-OHDA-treated cells, the cellular GSH level was not altered for the first 6-hr exposure to 6-OHDA, but after then, the level was significantly increased, which was also blocked by the pretreatment with NAC and BAPTA, but not by PD98059. Depletion of GSH by pretreating the cells with DL-buthionine-(S,R)-sulfoximine (BSO), a glutathione synthesis inhibitor, rather significantly potentiated the 6-OHDA-induced death. In contrast to the pretreatment with NAC, 6-OHDA-induced cell death was not prevented by the post-treatment with NAC 30 min after 6-OHDA treatment. The results indicate that the GSH level which is increased adaptively by the 6-OHDA-induced ROS and intracellular $Ca^{2+}$ is not enough to overcome the death signal mediated through ROS-$Ca^{2+}$ -caspase pathway.

• Biomolecules & Therapeutics
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• v.18 no.4
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• pp.454-462
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• 2010

The present study investigated the neuroprotective effect of Carpinus tschonoskii MAX and its intracellular protective mechanism on 6-hydroxydopamine (6-OHDA)-induced oxidative damage in PC12 cells. We found that pretreatment of PC12 cells with C. tschonoskii extract significantly inhibited the cell death induced by 6-OHDA in a dose dependent manner. C. tschonoskii extract decreased 6-OHDA-induced apoptotic events such as chromatin condensation, DNA fragmentation, the decrease of Bcl-2/Bax ratio, caspase-3 activation and PARP cleavage. C. tschonoskii extract also reduced generation of 6-OHDA-induced reactive oxygen species and nitric oxide. Furthermore, C. tschonoskii extract up-regulated the myocyte enhancer factor 2 D (MEF2D), a critical transcription factor for neuronal survival, and Akt activity, whereas it inhibited the activity of ERK1/2 and JNK. The results suggest that C. tschonoskii extract decreases 6-OHDA-induced oxidative stress and could prevent PC12 cell apoptosis induced by 6-OHDA via the up-regulation of MEF2D and Akt activity, and thus may have application in developing therapeutic agents for Parkinson's disease.

• Development and Reproduction
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• v.14 no.1
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• pp.35-41
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• 2010

Mammalian reproduction is regulated by a feedback circuit of the key reproductive hormones such as GnRH, gonadotropin and sex steroids on the hypothalamic-pituitary-gonadal axis. In particular, the onset of female puberty is triggered by gain of a pulsatile pattern and increment of GnRH secretion from hypothalamus. Previous studies including our own clearly demonstrated that genistein (GS), a phytoestrogenic isoflavone, altered the timing of puberty onset in female rats. However, the brain-specific actions of GS in female rats has not been explored yet. The present study was performed to examine the changes in the activities of GnRH neurons and their neural circuits by GS in female rats. Concerning the drug delivery route, intracerebroventricular (ICV) injection technique was employed to eliminate the unwanted actions on the extrabrain tissues which can be occurred if the testing drug is systemically administered. Adult female rats (PND 100, 210-230 g BW) were anaesthetized, treated with single dose of GS ($3.4{\mu}g$/animal), and sacrificed at 3 hrs post-injection. To determine the transcriptional changes of reproductive hormone-related genes in hypothalamus, total RNAs were extracted and applied to the semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). ICV infusion of GS significantly raised the transcriptional activities of enhanced at puberty1 (EAP-1, p<0.05), glutamic acid decarboxylase (GAD67, p<0.01) which are known to modulate GnRH secretion in the hypothalamus. However, GS infusion could not change the mRNA level of nitric oxide synthase 2 (NOS-2). GS administration significantly increased the mRNA levels of KiSS-1 (p<0.001), GPR54 (p<0.001), and GnRH (p<0.01) in the hypothalami, but decreased the mRNA levels of LH-$\beta$ (p<0.01) and FSH-$\beta$ (p<0.05) in the pituitaries. Taken together, the present study indicated that the acute exposure to GS could directly activate the hypothalamic GnRH modulating system, suggesting the GS's disrupting effects such as the early onset of puberty in immature female rats might be derived from premature activation of key reproduction related genes in hypothalamus-pituitary neuroendocrine circuit.

• Reproductive and Developmental Biology
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• v.29 no.2
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• pp.93-99
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• 2005

Human embryonic stem cells (hESCs) derived from the inner cell mass of blastocysts have the ability to renew themselves and to differentiate into cell types of all lineage. The present study was carried out to investigate whether the Wnt signaling pathway is related to maintaining self-renewal of hESCs. Glycogen Synthase Kinase 3 (GSK-3) inhibitor, BIO ((2'Z,3'E)-6-Bromoindirubin-3'-oxime) was treated to Miz-hES1 line for activation of Wnt signaling pathway. BIO-nontreated hESCs (control) and BID-treated hESCs were cultured for 5 days in the modified feeder-free system. During the culture of hESCs, differences were observed in the colony morphology between 2 groups. Controls were spread outwards whereas BIO-nontreated hESCs were clumped in the center and the differentiated cells were spreading outwards in the edges. The results of stem cell specific marker staining indicated that control were differentiated in large part whereas BIO-treated hESCs maintain self-renewal in the center of the colony. The results of lineage marker staining suggested that outer cells of the hESC colony were differentiated to the neuronal progenitor cells in both control and BIO-treated hESC. These results indicate that Wnt signaling is related to self-renewal in hESCs. In addition, control group showed higher composition of apoptotic cells $(23.76\%)$ than the BID-treated group $(5.59\%)$. These results indicate that BIO is effective on antapoptosis of hESCs.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.3
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• pp.279-286
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• 2016

Caffeic acid phenethyl ester (CAPE), derived from honeybee hives, is a bioactive compound with strong antioxidant activity. This study was designed to test the neuroprotective effect of CAPE in 3-nitropropionic acid (3NP)-induced striatal neurotoxicity, a chemical model of Huntington's disease (HD). Initially, to test CAPE's antioxidant activity, a 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid (ABTS) antioxidant assay was employed, and CAPE showed a strong direct radical-scavenging effect. In addition, CAPE provided protection from 3NP-induced neuronal cell death in cultured striatal neurons. Based on these observations, the in vivo therapeutic potential of CAPE in 3NP-induced HD was tested. For this purpose, male C57BL/6 mice were repeatedly given 3NP to induce HD-like pathogenesis, and 30 mg/kg of CAPE or vehicle (5% dimethyl sulfoxide and 95% peanut oil) was administered daily. CAPE did not cause changes in body weight, but it reduced mortality by 29%. In addition, compared to the vehicle-treated group, robustly reduced striatal damage was observed in the CAPE-treated animals, and the 3NP-induced behavioral deficits on the rotarod test were significantly rescued after the CAPE treatment. Furthermore, immunohistochemical data showed that immunoreactivity to glial fibrillary acidic protein (GFAP) and CD45, markers for astrocyte and microglia activation, respectively, were strikingly reduced. Combined, these data unequivocally indicate that CAPE has a strong antioxidant effect and can be used as a potential therapeutic agent against HD.

• The Korean Journal of Physiology and Pharmacology
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• v.20 no.2
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• pp.193-200
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• 2016

Sertraline, a selective serotonin reuptake inhibitor (SSRI), has been reported to lead to cardiac toxicity even at therapeutic doses including sudden cardiac death and ventricular arrhythmia. And in a SSRI-independent manner, sertraline has been known to inhibit various voltage-dependent channels, which play an important role in regulation of cardiovascular system. In the present study, we investigated the action of sertraline on Kv1.5, which is one of cardiac ion channels. The effect of sertraline on the cloned neuronal rat Kv1.5 channels stably expressed in Chinese hamster ovary cells was investigated using the whole-cell patch-clamp technique. Sertraline reduced Kv1.5 whole-cell currents in a reversible concentration-dependent manner, with an $IC_{50}$ value and a Hill coefficient of $0.71{\mu}M$ and 1.29, respectively. Sertraline accelerated the decay rate of inactivation of Kv1.5 currents without modifying the kinetics of current activation. The inhibition increased steeply between -20 and 0 mV, which corresponded with the voltage range for channel opening. In the voltage range positive to +10 mV, inhibition displayed a weak voltage dependence, consistent with an electrical distance ${\delta}$ of 0.16. Sertraline slowed the deactivation time course, resulting in a tail crossover phenomenon when the tail currents, recorded in the presence and absence of sertraline, were superimposed. Inhibition of Kv1.5 by sertraline was use-dependent. The present results suggest that sertraline acts on Kv1.5 currents as an open-channel blocker.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.1
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• pp.125-131
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• 2017

Status epilepticus is the most common serious neurological condition triggered by abnormal electrical activity, leading to severe and widespread cell loss in the brain. Lithium has been one of the main drugs used for the treatment of bipolar disorder for decades, and its anticonvulsant and neuroprotective properties have been described in several neurological disease models. However, the therapeutic mechanisms underlying lithium's actions remain poorly understood. The muscarinic receptor agonist pilocarpine is used to induce status epilepticus, which is followed by hippocampal damage. The present study was designed to investigate the effects of lithium post-treatment on seizure susceptibility and hippocampal neuropathological changes following pilocarpine-induced status epilepticus. Status epilepticus was induced by administration of pilocarpine hydrochloride (320 mg/kg, i.p.) in C57BL/6 mice at 8 weeks of age. Lithium (80 mg/kg, i.p.) was administered 15 minutes after the pilocarpine injection. After the lithium injection, status epilepticus onset time and mortality were recorded. Lithium significantly delayed the onset time of status epilepticus and reduced mortality compared to the vehicle-treated group. Moreover, lithium effectively blocked pilocarpine-induced neuronal death in the hippocampus as estimated by cresyl violet and Fluoro-Jade B staining. However, lithium did not reduce glial activation following pilocarpine-induced status epilepticus. These results suggest that lithium has a neuroprotective effect and would be useful in the treatment of neurological disorders, in particular status epilepticus.