• 대한한방내과학회지
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• 제35권1호
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• pp.79-91
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• 2014

Objectives : To investigate the anti-cancer effect of Palbohoichoon-tang (PBHCT) extracts. Methods : The cell viability was assessed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MMT) assay and cell morphological changes were microscopically analyzed after staining with $10{\mu}M$ 2-[4-amidinophenyl]-6-indolecarbamidine dihydrochloride (DAPI) and TUNEL. We also analyzed expression of Bcl2, $Bcl_{xL}$, Bax, procaspase-3, procaspase-9, and procyclic acidic repetitive protein (PARP) by western blot method. Results : Observations showed that PBHCT induced the apoptotic cell death proved by increased sub-G1 phase cell population, apoptotic body formation and chromatin condensation. Western blot analysis of total cell lysates revealed that the PBHCT induced cleavage of caspase-9, caspase-3 and poly (ADP-ribose) polymerase (PARP). In addition, PBHCT dose-dependently increased the activity of caspase-9, caspase-3 and PARP-1. Furthermore, PBHCT reduced anti-apoptotic Bcl2, $Bcl_{xL}$ expression which contributed to the loss of mitochondrial membrane potential and the activations of caspase-9 and caspase-3. Conclusions : These findings suggest that PBHCT exerts anti-cancer effects on human neuroblastoma SH-SY5Y cells by inducing apoptotic death via down-regulation of anti-apoptotic proteins such as Bcl2 and $Bcl_{xL}$, up-regulation of pro-apoptotic proteins such as Bax, and activation of caspase cascades and PARP-1.

• Toxicological Research
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• 제35권1호
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• pp.83-91
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• 2019

Nanoparticles (NPs) have been recognized as both useful tools and potentially toxic materials in various industrial and medicinal fields. Previously, we found that zinc oxide (ZnO) NPs that are neurotoxic to human dopaminergic neuroblastoma SH-SY5Y cells are mediated by lipoxygenase (LOX), not cyclooxygenase-2 (COX-2). Here, we examined whether human bone marrow-derived mesenchymal stem cells (MSCs), which are different from neuroblastoma cells, might exhibit COX-2- and/or LOX-dependent cytotoxicity of ZnO NPs. Additionally, changes in annexin V expression, caspase-3/7 activity, and mitochondrial membrane potential (MMP) induced by ZnO NPs and ZnO were compared at 12 hr and 24 hr after exposure using flow cytometry. Cytotoxicity was measured based on lactate dehydrogenase activity and confirmed by trypan blue staining. Rescue studies were executed using zinc or iron chelators. ZnO NPs and ZnO showed similar dose-dependent and significant cytotoxic effects at concentrations ${\geq}15{\mu}g/mL$, in accordance with annexin V expression, caspase-3/7 activity, and MMP results. Human MSCs exhibited both COX-2 and LOX-mediated cytotoxicity after exposure to ZnO NPs, which was different from human neuroblastoma cells. Zinc and iron chelators significantly attenuated ZnO NPs-induced toxicity. Conclusively, these results suggest that ZnO NPs exhibit both COX-2- and LOX-mediated apoptosis by the participation of mitochondrial dysfunction in human MSC cultures.

• Journal of Microbiology and Biotechnology
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• 제23권2호
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• pp.274-277
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• 2013

The unique properties of graphene have earned much interest in the fields of materials science and condensedmatter physics in recent years. However, the biological applications of graphene remain largely unexplored. In this study, we investigate the cell culture conditions, which are exposed to graphene onto glass and $SiO_2$/Si using human nerve cell line, SH-SY5Y. Cell viability was 84% when cultured on glass and $SiO_2$/Si coated with graphene as compared to culturing on polystyrene surface. Fluorescence data showed that the presence of graphene did not influence cell morphology. These findings suggest that graphene may be used for biological applications.

• 대한본초학회지
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• 제30권1호
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• pp.67-75
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• 2015

Objectives : The aim of this study is to examine the neuroprotective effect of wheat bran extract (WBE) against ${\beta}$-amyloid ($A{\beta}$)-induced apoptotic cell death in SH-SY5Y human neuroblastoma cells and memory impairment in triple transgenic animal model's of Alzheimer's disease (3xTg AD mice). Methods : In SH-SY5Y cells, MTT assay and TUNEL staining were conducted to evaluate the protective effect of WBE against $A{\beta}_{25-35}$-induced neurotoxicity and apoptosis. Alterations in mitochondrial transmembrane potential (MMP), expression of proapoptotic Bax and antiapoptotic Bcl-2 proteins, cleavage of PARP, and brain-derived neurotrophic factor (BDNF) levels were analyzed to elucidate the neuroprotective mechanism of WBE. To further investigate the memory enhancing effect of WBE, Morris water maze test was performed in 3xTg AD mice. Results : In SH-SY5Y cells, WBE protected against $A{\beta}_{25-35}$-caused cytotoxicity and apoptosis as shown by the restoration of cell viability in MTT assay and inhibition of DNA fragmentation in TUNEL staining. $A{\beta}_{25-35}$-induced apoptotic signals such as dissipation of MMP, decreased Bcl-2/Bax ratio, and cleavage of PARP were suppressed by WBE. Moreover, WBE up-regulated the protein levels of BDNF, which seemed to be mediated by activation of cAMP response element-binding protein (CREB). In 3xTg AD mice, oral administration of WBE attenuated learning and memory deficit as verified by reduced mean escape latency in water maze test. Conclusions : WBE protects neuronal cells from $A{\beta}_{25-35}$-induced apoptotic cell death and restores learning and memory impairments in 3xTg AD mice. These findings suggest that WBE exhibit neuroprotective potential for the management of AD.

• Journal of Nutrition and Health
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• 제54권6호
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• pp.618-630
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• 2021

본 연구에서는 생강을 효소처리하여 수용화율을 높인 후 고초균을 이용하여 발효시킨 생강발효물 (FGEs)을 제조하고, 이를 SH-SY5Y에 6-OHDA와 함께 처리하여 세포 보호 효과 및 AChE 저해 활성을 평가하였다. 그 결과, 6-OHDA로 자극된 신경세포에서 FGE를 처리한 모든 실험군에서 세포 생존율이 증가하고 LDH 농도가 감소하였다. 6-OHDA로 유발된 세포자멸사를 억제할 수 있는지 확인하기 위해 핵의 형태학적인 변화 및 caspase-3 활성을 확인하였다. FGE를 처리한 모든 실험군에서 핵의 손상 및 apoptotic body의 감소와 caspase-3 억제 활성을 확인할 수 있었다. 또한 FGE는 AChE의 활성을 유의적으로 감소시켰다. 시료 간의 활성 차이를 비교하였을 때, 생강에 효소 처리 후 고초균으로 발효한 추출물군 (E/BKG와 E/BCG)의 신경세포 보호 활성이 효소 처리하지 않은 발효생강군 (BKG와 BCG) 보다 유의적으로 크게 나타났다. 그러나 발효에 사용된 고초균 2종 간의 활성은 유사하였으며 처리군 간 유의적인 차이를 보이지 않았다. 본 연구 결과로부터 효소처리하여 수용화율을 높여 고초균으로 발효한 FGE는 신경세포 보호 및 AChE 저해 효과를 나타내어 향후 신경질환 연구를 위한 기초자료 제공 및 고부가가치 식품소재 개발에 이용 가능할 것으로 판단된다.

• 한국약용작물학회지
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• 제17권5호
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• pp.341-345
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• 2009

Cerebral ischemia results from a transient or permanent reduction in cerebral blood flow that decreases oxygen and glucose supply. When the cellular oxygen supply is reduced to critical level, damage to cells and induction of cell death are occurred by excitotoxicity, oxidative stress and inflammation. Ischemia remains one of the leading causes of death, but there is no effective treatment that might protect neurons gainst ischemia by interrupting the cascade of cell death. In this study, human neuroblastoma SH-SY5Y cells are exposed to oxygen and glucose deprivation (OGD) followed by reoxgenation. OGD can mimic the acute restriction of metabolite and oxygen supply caused by ischemia and is widely used as a model of ischemic conditions. SH-SY5Y cells are treated samples at the commencement of OGD to achieve different final concentrations, and cell viabilities were quantified using the measurement of flow cytometry analysis. Of those tested, the extracts of Polygala tenuifolia (roots), Dictamnus dasycarpus (barks), Polygala tenuifolia (roots), Eucommia ulmoides (branches), Eucommia ulmoides (barks), Poria cocos (whole), Sophora flavescens (roots) showed neuroprotective effects, with $EC_{50}$ values of $4.5{\pm}0.6$, $7.9{\pm}1.5$, $10.5{\pm}0.7$, $18.4{\pm}1.9$, $19.6{\pm}0.3$, $21.6{\pm}1.9$, and $30.7{\pm}3.9{\mu}g/m{\ell}$, respectively.

• BMB Reports
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• 제52권7호
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• pp.439-444
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• 2019

Although hypoxic/ischemic injury is thought to contribute to the incidence of Alzheimer's disease (AD), the molecular mechanism that determines the relationship between hypoxia-induced ${\beta}$-amyloid ($A{\beta}$) generation and development of AD is not yet known. We have now investigated the protective effects of N,4,5-trimethylthiazol-2-amine hydrochloride (KHG26702), a novel thiazole derivative, on oxygen-glucose deprivation (OGD)-reoxygenation (OGD-R)-induced $A{\beta}$ production in SH-SY5Y human neuroblastoma cells. Pretreatment of these cells with KHG26702 significantly attenuated OGD-R-induced production of reactive oxygen species and elevation of levels of malondialdehyde, prostaglandin $E_2$, interleukin 6 and glutathione, as well as superoxide dismutase activity. KHG26702 also reduced OGD-R-induced expression of the apoptotic protein caspase-3, the apoptosis regulator Bcl-2, and the autophagy protein becn-1. Finally, KHG26702 reduced OGD-R-induced $A{\beta}$ production and cleavage of amyloid precursor protein, by inhibiting secretase activity and suppressing the autophagic pathway. Although supporting data from in vivo studies are required, our results indicate that KHG26702 may prevent neuronal cell damage from OGD-R-induced toxicity.

• BMB Reports
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• 제37권2호
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• pp.185-191
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• 2004

Tissue transglutaminase (tTGase) regulates various biological processes, including extracellular matrix organization, cellular differentiation, and apoptosis. Here we report the protective role of tTGase in the cell death that is induced by the tumor necrosis factor $\alpha$ (TNF-$\alpha$) and ceramide, a product of the TNF-$\alpha$ signaling pathway, in human neuroblastoma SH-SY5Y cells. Treatment with retinoic acid (RA) induced the differentiation of the neuroblastoma cells with the formation of extended neurites. Immunostaining and Western blot analysis showed the tTGase expression by RA treatment. TNF-$\alpha$ or $C_2$ ceramide, a cell permeable ceramide analog, induced cell death in normal cells, but cell death was largely inhibited by the RA treatment. The inhibition of tTGase by the tTGase inhibitors, monodansylcadaverine and cystamine, eliminated the protective role of RA-treatment in the cell death that is caused by TNF-$\alpha$ or $C_2$-ceramide. In addition, the co-treatment of TNF-$\alpha$ and cycloheximide ecreased the protein level of tTGase and cell viability in the RA-treated cells, supporting the role of tTGase in the protection of cell death. DNA fragmentation was also induced by the co-treatment of TNF-$\alpha$ and cycloheximide. These results suggest that tTGase expressed by RA treatment plays an important role in the protection of cell death caused by TNF-$\alpha$ and ceramide.

• 대한한방부인과학회지
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• 제23권3호
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• pp.1-13
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• 2010

Purpose: These studies were undertaken to evaluate the anti-oxidative and neuroprotective effects of Gamisoyo-san(GMSYS). Materials and Methods: We studied the antioxidant effects of GMSYS by assessing the DPPH free radical and the ABTS radical cation inhibition activities, the total polyphenolic contents(TPC). To evaluate the effects of GMSYS in the human neuroblastoma cells, we measured the cell viabilities in SH-SY5Y cells treated with GMSYS. Then we observed the protective effects of GMSYS against 6-OHDA induced neurotoxicity in SH-SY5Y cells. To confirm the neuroprotective effects of GMSYS in the primary culture of mesencephalic dopaminergic cells, we counted the TH-immunopositive cells and measured the NO and TNF-$\alpha$ after the treatment of GMSYS and 6-OHDA. Results: The DPPH free radical and the ABTS radical cation inhibition activities were increased in a dose dependent manner and the IC50 were $133.60{\mu}g/m{\ell}$ and $106.20{\mu}g/m{\ell}$, respectively. The TPC was 0.78%. There were no differences between the various concentrations of GMSYS and the control in the cell viability of SH-SY5Y cells. The neuroprotective effects of GMSYS were shown in the co-treatment group at the low concentrations of $25{\mu}g/m{\ell}$ and the post-treatment group at all concentrations. After the treatment of GMSYS and 6-OHDA in the primary culture of dopaminergic cells, the TH-immunopositive cells were significantly increased in $0.2{\mu}g/m{\ell}$ of GMSYS than the 6-OHDA group. The NO and TNF-$\alpha$ were significantly decreased in $0.2{\mu}g/m{\ell}$ of GMSYS than the 6-OHDA group. Conclusions: This study shows that GMSYS has the antioxidant and neuroprotective effects, especially in the mesencephalic dopaminergic cells. We suggest that GMSYS could be useful for the treatment of postmenopausal depression related with the degeneration of dopamine neuron.

• Molecules and Cells
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• 제37권3호
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• pp.226-233
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• 2014

Excessive reactive oxygen species (ROS) generated from abnormal cellular process lead to various human diseases such as inflammation, ischemia, and Parkinson's disease (PD). Sensitive to apoptosis gene (SAG), a RING-FINGER protein, has anti-apoptotic activity and anti-oxidant activity. In this study, we investigate whether Tat-SAG, fused with a Tat domain, could protect SH-SY5Y neuroblastoma cells against 1-methyl-4-phenylpyridinium ($MPP^+$) and dopaminergic (DA) neurons in the substantia nigra (SN) against 1-methyl-4-phenyl-1,2,3,6-tetra-hydropyridine (MPTP) toxicity. Western blot and immunohistochemical analysis showed that, unlike SAG, Tat-SAG transduced efficiently into SH-SY5Y cells and into the brain, respectively. Tat-SAG remarkably suppressed ROS generation, DNA damage, and the progression of apoptosis, caused by $MPP^+$ in SH-SY5Y cells. Also, immunohistochemical data using a tyrosine hydroxylase antibody and cresyl violet staining demonstrated that Tat-SAG obviously protected DA neurons in the SN against MPTP toxicity in a PD mouse model. Tat-SAG-treated mice showed significant enhanced motor activities, compared to SAG- or Tat-treated mice. Therefore, our results suggest that Tat-SAG has potential as a therapeutic agent against ROS-related diseases such as PD.