• Journal of Ginseng Research
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• 제35권3호
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• pp.352-359
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• 2011

Korean red ginseng (KRG) is used worldwide as a popular traditional herbal medicine. KRG has shown beneficial effects on cardiovascular diseases, such as atherosclerosis, diabetes, and hypertension. Up-regulation of a cytoprotective protein, heme oxygenase (HO)-1, is considered to augment the cellular defense against various agents that may induce cytotoxic injury. In the present study, we demonstrate that KRG water extract induces HO-1 expression in human umbilical vein endothelial cells (HUVECs) and possible involvement of the anti-oxidant transcription factor nuclear factor-eythroid 2-related factor 2 (Nrf2). KRG-induced HO-1 expression was examined by western blots, reverse transcriptase polymerase chain reaction and immunofluorescence staining. Specific silencing of Nrf2 genes with Nrf2-siRNA in HUVECs abolished HO-1 expression. In addition, the HO inhibitor zinc protoporphyrin blunted the preventive effect of KRG on $H_2O_2$-induced cell death, as demonstrated by terminal transferase dUTP nick end labeling assay. Taken together, these results suggest that KRG may exert a vasculoprotective effect through Nrf2-mediated HO-1 induction in human endothelial cell by inhibition of cell death.

• Biomolecules & Therapeutics
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• 제24권5호
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• pp.510-516
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• 2016

Isoegomaketone (IK) was isolated from Perilla frutescens, which has been widely used as a food in Asian cuisine, and evaluated for its biological activity. We have already confirmed that IK induced the HO-1 expression via Nrf2 activation in RAW264.7 cells. In this study, we investigated the effect of IK on the mechanism of HO-1 expression. IK upregulated HO-1 mRNA and protein expression in a dose dependent manner. The level of HO-1 mRNA peaked at 4 h after $15{\mu}M$ IK treatment. To investigate the mechanisms of HO-1 expression modulation by IK, we used pharmacological inhibitors for the protein kinase C (PKC) family, PI3K, and p38 MAPK. IK-induced HO-1 mRNA expression was only suppressed by SB203580, a specific inhibitor of p38 MAPK. ROS scavengers (N-acetyl-L-cysteine, NAC, and glutathione, GSH) also blocked the IK-induced ROS production and HO-1 expression. Furthermore, both NAC and SB203580 suppressed the IK-induced Nrf2 activation. In addition, ROS scavengers suppressed other oxidative enzymes such as catalase (CAT), glutathione S-transferase (GST), and NADH quinone oxidoreductase (NQO-1) in IK-treated RAW264.7 cells. Taken together, it can be concluded that IK induced the HO-1 expression through the ROS/p38 MAPK/Nrf2 pathway in RAW264.7 cells.

• Biomolecules & Therapeutics
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• 제24권6호
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• pp.581-588
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• 2016

Lonchocarpine is a phenylpropanoid compound isolated from Abrus precatorius that has anti-bacterial, anti-inflammatory, antiproliferative, and antiepileptic activities. In the present study, we investigated the antioxidant effects of lonchocarpine in brain glial cells and analyzed its molecular mechanisms. We found that lonchocarpine suppressed reactive oxygen species (ROS) production and cell death in hydrogen peroxide-treated primary astrocytes. In addition, lonchocarpine increased the expression of anti-oxidant enzymes, such as heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), and manganese superoxide dismutase (MnSOD), which are all under the control of Nrf2/antioxidant response element (ARE) signaling. Further, mechanistic studies showed that lonchocarpine increases the nuclear translocation and DNA binding of Nrf2 to ARE as well as ARE-mediated transcriptional activities. Moreover, lonchocarpine increased the phosphorylation of AMP-activated protein kinase (AMPK) and three types of mitogen-activated protein kinases (MAPKs). By treating astrocytes with each signaling pathway-specific inhibitor, AMPK, c-jun N-terminal protein kinase (JNK), and p38 MAPK were identified to be involved in lonchocarpine-induced HO-1 expression and ARE-mediated transcriptional activities. Therefore, lonchocarpine may be a potential therapeutic agent for neurode-generative diseases that are associated with oxidative stress.

• 생명과학회지
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• 제33권10호
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• pp.776-782
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• 2023

Silymarin은 간 보호, 항산화, 항염, 항암 등 다양한 생리 활성을 나타내는 것으로 보고되었고, 본 연구에서는 산화 스트레스에 대한 항산화 잠재력과 그 기전을 세포 생존력 및 활성산소종 생성 분석과 Western blot 분석을 통해 RAW 264.7 세포에서 알아보고자 하였다. Silymarin은 세포 독성 없이 lipopolysaccharide(LPS)에 의해 자극된 세포 내 활성산소종을 농도 의존적으로 소거하였다. 그리고 항산화 효과를 보여주는 것으로 알려진 제2상 효소 중 하나인 heme oxygenase (HO)-1의 발현은 silymarin 처리에 의해 강하게 유도되었다. 또한 silymarin 처리는 항산화 효소의 전사인자인 nuclear factor-erythroid 2 p45-related factor (Nrf)-2의 발현을 유의미하게 유도하였고, 이는 HO-1 발현증가와 일치하였다. 세포내 산화와 환원 항상성 조절과 관련된 신호 전달물질인 mitogen activated protein kinase (MAPK)와 phosphoinositde 3-kinase (PI3K)의 인산화 정도 또한 Western blot으로 분석하였고, 그 결과 silymarin 은 p38 MAPK 인산화에 의해 HO-1 발현을 유도하는 것으로 나타났다. 그리고 tert-butyl hydroperoxide (t-BHP)를 이용하여 세포내 지질 과산화를 유도함으로써 silymarin에 의해 유도된 HO-1의 항산화 효과를 확인하였다. 그 결과 silymarin 처리에 의해 세포사멸이 유의적으로 억제되었고, p38의 선택적 저해제를 처리한 세포군에서는 t-BHP에 의해 유의적인 세포사멸이 발생하였다. 이 결과를 통해 silymarin은 Nrf-2/p38 MAPK 신호 전달 경로를 통해 HO-1의 발현을 유도하고, 이를 통해 항산화 효과를 높이는 것을 확인할 수 있었다.

• 대한한의학방제학회지
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• 제25권1호
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• pp.1-10
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• 2017

Objectives : The fruit of Prunus mume Siebold & Zucc. has been used as an alternative medicine and functional food in Korea and Japan for preventive and therapeutic purposes. However, its molecular actions and mechanism on anti-inflammatory activity have not been clearly investigated. The aim of this study was to clarify the anti-inflammatory activity of the ethanol extract of P. mume fruit (EEPM) in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells, and sought to understand the associated molecular mechanisms. Methods : Cytotoxicity was assessed by an MTT assay. The amount of nitric oxide (NO) production was determined by nitrite assay. The mRNA expression of inducible nitric oxide synthase (iNOS) was analyzed by RT-PCR. In addition, expression levels of iNOS, nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) protein were detected by Western blotting. Results : Our data indicated that EEPM inhibited NO production in LPS-stimulated RAW264.7 cells in a concentration-dependent manner. At the mRNA and protein levels, EEPM suppressed LPS-induced iNOS expression. On the other hand, EEPM markedly enhanced HO-1 expression, which was associated with an induction and nuclear translocation of Nrf2. Moreover, the inhibitory effect of EEPM against LPS‑induced NO production was significantly enhanced by hemin, a HO-1 inducer; however, EEPM's effect on the production of NO was abolished by zinc protoporphyrin IX, a HO-1 inhibitor. Conclusion : The results suggest that EEPM can act as a suppressor agent on NO production through an activation of Nrf2/HO-1 signaling pathway, and may be a promising candidate for the treatment of inflammatory diseases.

• 생약학회지
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• 제46권2호
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• pp.116-122
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• 2015

Keratinocytes are first barrier against outer challenges on skin. However, it is still largely unknown about effective protectors against ultraviolet B (UVB), and oxidative stress in human keratinocyte, HaCaT cells. Inducible heme oxygenase (HO)-1 acts against oxidants that are thought to play a role in the pathogenesis of skin disorders. Therefore, the purpose of this study was to evaluate the effect of Portulaca oleracea 70% EtOH extracts against hydrogen peroxide (H₂O₂)-induced oxidative stress in human keratinocytes, HaCaT cells. P. oleracea 70% EtOH extracts showed the potent protective effects on H₂O₂-induced toxicity by induced the expression of HO-1 in human keratinocyte, HaCaT cells. Furthermore, P. oleracea 70 % EtOH extracts caused the nuclear accumulation of nuclear factor E2-related factor 2 (Nrf2) in human keratinocytes, HaCaT cells. In addition, we found that treatment with c-Jun N-terminal kinase (JNK) inhibitor (SP600125) reduced P. oleracea 70% EtOH extracts-induced HO-1 expression, and JNK inhibitor (SP600125) also inhibited protective effects by P. oleracea 70% EtOH extracts. Therefore, these results suggest that P. oleracea 70 % EtOH extracts increases cellular resistance to H₂O₂-induced oxidative injury in human keratinocyte, HaCaT cells, presumably through JNK pathway-Nrf2-dependent HO-1 expression.

• Fisheries and Aquatic Sciences
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• 제26권1호
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• pp.35-47
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• 2023

Myelophycus caespitosus, a brown alga belonging to genus Myelophycus, has been traditionally used as a food and medicinal resource in Northeastern Asia. However, few studies have been conducted on its pharmacological activity. In this study, we evaluated whether methanol extract of M. caespitosus (MEMC) could protect against oxidative damage caused by hydrogen peroxide (H₂O₂) in C2C12 murine myoblasts. Our results revealed that MEMC could suppress H₂O₂-induced growth inhibition and DNA damage while blocking the production of reactive oxygen species. In H₂O₂-treated cells, cell cycle progression was halted at the G2/M phase, accompanied by changes in expression of key cell cycle regulators. However, these effects were attenuated by MEMC. In addition, we found that MEMC protected cells from induction of apoptosis associated with mitochondrial impairment caused by H₂O₂ treatment. Furthermore, MEMC enhanced the phosphorylation of nuclear factor-erythroid-2 related factor 2 (Nrf2) and expression and activity of heme oxygenase-1 (HO-1) in H₂O₂-treaetd C2C12 myoblasts. However, such anti-apoptotic and cytoprotective effects of MEMC were greatly abolished by HO-1 inhibitor, suggesting that MEMC could increase Nrf2-mediated activity of HO-1 to protect C2C12 myoblasts from oxidative stress.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2019년도 추계학술대회
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• pp.91-91
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• 2019

Hibiscus syriacus L. is widely distributed throughout Eastern and Southern Asia and its root bark has been used as a traditional remedy. Recently, the extracts of H. syriacus L. exerts anti-cancerous, anti-microbial, and anti-inflammatory activities. However, the effect of anthocyanin-rich fraction of H. syriacus L. petals (PS) has not been studied under excessive oxidative stress. In this study, we evaluated the cellular protective effect of PS in HaCaT human skin keratinocytes under hydrogen peroxide ($H_2O_2$)-induced oxidative stress conditions. PS at below $400{\mu}g/ml$ did not show any cell death; however, over $800{\mu}g/ml$ of PS gradually increased cell death. PS at below $400{\mu}g/ml$ significantly inhibited $H_2O_2$-induced apoptosis in HaCaT cells concomitant with downregulation of Bax and upregulation of pro-PARP and p-Bcl-2. Additionally, PS remarkably reversed $H_2O_2$-induced excessive reactive oxygen species (ROS) production and apoptosis, and also significantly inhibited mitochondrial ROS production concomitant with suppression of $H_2O_2$-induced mitochondrial depolarization. $H_2O_2$-mediated ratio of Bax to Bcl-2, and caspase-3 activation were markedly abolished in the presence of PS. Moreover, the inhibition of HO-1 function using zinc protoporphyrin, an HO-1 inhibitor, significantly attenuated the cellular protective effects of PS against $H_2O_2$, indicating the significance of HO-1 in PS mediated cytoprotective effect, which was mediated by activating nuclear factor erythroid 2-related factor-2 (Nrf2). Taken together, our results suggest that cytoprotective effect of PS in HaCaT keratinocytes against oxidative stress-induced apoptosis is mediated by inhibiting cellular and mitochondrial ROS production, which is downregulated by activating Nrf2/HO-1 axis.

• 동의생리병리학회지
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• 제28권1호
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• pp.63-68
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• 2014

The fruit of Tribulus terrestris L. (Zygophyllaceae) is an important source of traditional Korean and Chinese medicines. In this study, NNMBS223, consisting of the ethanol extract of T. terrestris, showed potent anti-inflammatory activities in RAW264.7 macrophages. We investigated the effect of NNMBS223 in suppressing the protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2 and production of iNOS-derived nitric oxide (NO), COX-2-derived prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-stimulated macrophages. In addition, NNMBS223 induced expression of heme oxygenase (HO)-1 through nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in macrophages. The effects of NNMBS223 on LPS-induced production of NO and PGE2 were partially reversed by the HO activity inhibitor tin protoporphyrin (SnPP). These findings suggest that Nrf2-dependent increases in expression of HO-1 induced by NNMBS223 conferred anti-inflammatory activities in LPS stimulated RAW264.7 macrophages.

• 생명과학회지
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• 제28권1호
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• pp.43-49
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• 2018

Chrysoeriol은 alfalfa에서 주로 발견되는, 식물계에 많이 분포하고 있는 flavone으로 전통의학에서 소화불량, 천식, 비뇨기계 이상의 치료에 사용되어 왔다. 최근의 연구에서는 항염증 효과가 있는 것으로 밝혀졌으나 항산화 효과에 대한 분석은 없었다. 본 연구에서는 chrysoeriol의 항산화 효과와 그 분자적 기전을 RAW 264.7 cell에서 세포생존율, reactive oxygen species (ROS)와 Western blot분석을 통해 알아보고자 하였다. Chrysoeriol은 lipopolysaccharide(LPS)에 의해 발생한 ROS를 세포독성없이 농도의존적으로 제거하였다. 그리고 항산화효과를 보이는 2상 효소 중 하나인 heme oxygenase (HO)-1의 발현을 강하게 유도하였고, 그와 동시에 전사인자인 Nrf2의 핵내 이동도 촉진하는 것으로 밝혀졌다. 특히, 산화스트레스에 대한 세포내 산화환원항상성 유지에 중요한 역할을 하고 있는 것으로 알려진 mitogen activated protein kinase (MAPK)와 phosphoinositide 3-kinase (PI3K)의 분석결과, chrysoeriol은 extracellular signal regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK)와 p38의 인산화를 통해 HO-1의 발현을 유도하는 것으로 나타났다. HO-1에 의한 항산화 효과를 확인하기 위하여 chrysoeriol을 전처리한 후 t-BHP에 의한 산화 스트레스에 세포를 노출시킨 결과, chrysoeriol 처리에 의해 세포사멸이 줄어드는 것을 확인하였고, HO-1의 유도제와 억제제의 처리에 따라 세포생존율 또한 조절되는 것을 확인할 수 있었다. 따라서, chrysoeriol은 HO-1의 발현을 유도하여 항산화 효과를 높이고 이것은 Nrf2/MAPK 신호전달 체계에 의한다는 것을 알 수 있었다.