• 생명과학회지
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• 제19권6호
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• pp.770-780
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• 2009

녹조류인 Spirulina platensis에서 추출하여 만든 새로운 광감작제와 660 nm의 다이오드 레이저를 이용한 광역학치료의 항암효과와 치료기전을 알아보았다. 세포 독성능은 MTT assay를 이용하였고, 세포사멸기전은 propidium iodide과 Hoechst 33342 염색법과 투과전자현미경으로 확인하였다. 또한 암세포가 이종 이식된 누드마우스 모델에서 광역학치료를 시행하여 항암효과를 확인하였다. 3종류의 클로로필 유도체 중 9-hydroxypheophorbide-a (9-HpbD-a)의 세포 독성능이 가장 우수하였고, 9-HpbD-a의 적정 레이저조사 시간은 30분 (3.2 J/$cm^{2}$), 광감작제를 투여하고 레이저조사시간까지의 배양시간은 최소 6시간 이상임을 확인하였다. 광역학치료의 세포사멸기전은 낮은 9-HpbD-a 농도에서 세포고사가 주된 세포사멸기전이었고, 높은 농도의 9-HpbD-a에서는 세포괴사에 의한 세포사멸이 주된 기전임을 확인하였다. 투과전자현미경 하에서도 같은 양상을 관찰하였다. 그리고 암세포가 이종 이식된 누드마우스 모델에서의 광역학치료는 제1군 정상대조군과 제2군 9-HpbD-a만을 투여한 종양조직모두 지속적인 종양의 성장(100% )을 보였고, 제3군인 레이저만을 종양조직에 조사한 실험군에서는 3 마리는 치료가 되지 않았고(75.0%), 1 마리는 재발(25.0%) 하였다. 제4군 광역학치료군에서 총16 마리의 종양에서 10 마리는 완치(62.5%), 4 마리는 재발(25.0%), 2 마리는 치유되지 않았음(12.5% )을 확인하였다. 9-HpbD-a와 660 nm 다이오드 레이저를 이용한 광역학치료는 유의한 항암효과를 나타내었고 9-HpbD-a를 이용한 광역학치료는 새로운 치료방법으로서 향후 암치료의 유용한 치료방법으로 기대된다.

• Tuberculosis and Respiratory Diseases
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• 제56권2호
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• pp.187-197
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• 2004

연구배경 : Nitric Oxide(NO)는 매우 다양한 생물학적 조절기능을 수행하는 분자로서 심혈관계, 신경계, 면역기능 등에 관여함은 물론 최근 세포 사에도 직 간접적으로 영향을 미치고 있음이 알려져 있다. NO의 이렇게 복잡한 생물학적 기능 수행은 reactive oxygen species(ROS), metal ions 및 단백질 등과 복잡한 상호작용에 의한 것이며 NO가 나타내는 생물학적 효과는 용량-의존적이며, 세포-특이적이라고 밝혀져 있다. NO는 간세포 및 현관내피세포에서는 아포프토시스를 억제하지만 종양세포 및 신경세포 등에서는 아포프토시스를 유도하는 것으로 보고되고 있다. NO는 여러 호흡기질환의 병태생리에도 관여하는 것으로 알려져 있는바 천식과 같은 염증성 기도 질환에서 호기 NO가 증가되어있는 반면 흡연자나 일차성 폐 고혈압 환자에서는 감소되어 있다고 보고되고 있다. 이러한 배경에서 NO가 폐 상피 세포의 세포 사에 미치는 영향과 신호전달 경로를 밝히기 위하여 본 연구를 시행하였다. 방 법 : 폐 상피 세포로는 A549 세포 주를, NO donor로서는 SNAP (S-nitroso-N-acetyl-penicillamine)과 SNP(sodium nitroprusside)를 사용하였다. 세포 독성 검사는 crystal violet assay를 이용하였고 아포프토시스 assay는 Hoechst 33342와 propium iodide(PI) 이중 염색 후 형광현미경을 이용하여 핵의 형태학적변화를 관찰함으로써 괴사(necrosis)와 감별하였다. 철에 의한 NO 유도성 세포 사 억제 효과를 관찰하기 위하여 RBC와 FeSO4를 이용하였다. NO 유도성세포사의 신호전달 경로에 bcl-2와 p53이 미치는 영향을 평가하기 위하여 bcl-2 과 발현 세포 주 (A549-bcl-2)와 p53 knock out 세포 주 (A549-E6)를 대상으로 세포독성을 비교하였고 p53 활성화는 Western blot을 이용하여 확인하였다. 결 과 : A549 세포 주에서 SNAP과 SNP 모두 농도-의존적 세포독성을 관찰할 수 있었다. 아포프토시스 assay에서 SNAP은 저 농도에서는 아포프토시스를, 고농도에서는 괴사를 유도함을 관찰하였고 SNP는 농도에 상관없이 세포사가 괴사의 형태를 나타냄을 확인하였다. 이는 SNP가 순순한 NO donor가 아니라 cyanide에 의한 세포독성의 결과라고 생각되며 고농도의 SNAP에 의한 괴사 유도는 peroxynitrite 생성에 의한 결과임을 시사한다. SNAP에 의한 세포 사는 RBC와 FeSO4등 철에 의해 억제됨을 확인하였고 bcl-2에 의해서 억제되었으며 p53을 활성화시키고 p53 knock out에 의해 차단되었다. 결 론 : 폐상피세포에서 NO는 저 농도에서는 아포프토시스를 고농도에서는 괴사에 의한 세포 사를 유도하며 철이 중요한 억제제이며 bcl-2 및 p53이 신호전달 경로에 있어서 중요한 역할을 담당하는 것으로 생각된다.

• 한국약용작물학회지
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• 제13권5호
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• pp.219-226
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• 2005

Sanguisorbae radix (SR) from Sanguisorba officinalis L. (Losaceae) is widely used in Korea and China due to its various pharmacological activity. The present study aims to investigate the effect of the methanol extract of SR on amyloid ${\beta}$ Protein(25-35) $(A{\beta}\;(25-35))$, a synthetic 25-35 amyloid peptide, -induced neurotoxicity using cultured rat cortical neurons. SR, over a concentration range of $10-50\;{\mu}g/ml$, inhibited the $A{\beta}$ (25-35) $(10\;{\mu}M)-induced$ neuronal cell death, as assessed by a 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay and the number of apoptotic nuclei, evidenced by Hoechst 33342 staining. Pretreatment of SR $(50\;{\mu}g/ml)$ inhibited $10\;{\mu}M\;A{\beta}$ (25-35)-induced} elevation of cytosolic calcium concentration $([Ca^{2+}]c)$, which was measured by a fluorescent dye, fluo-4 AM. SR $(10\;and\;50\;{\mu}g/ml)$ inhibited glutamate release into medium induced by $10\;{\mu}M\;A{\beta}(25-35)$, which was measured by HPLC, and generation of reactive oxygen species. These results suggest that SR prevents $A{\beta}$ (25-35)-induced neuronal cell damage in vitro.

• Natural Product Sciences
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• 제21권2호
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• pp.134-140
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• 2015

The present study aims to investigate the effect of methanol extract of Korean mistletoe (KM; Viscum album var. coloratum), on amyloid $\beta$ protein ($A\beta$) (25-35), a synthetic 25-35 amyloid peptide, -induced neurotoxicity in cultured rat cerebral cortical neurons and memory impairment in mice. Exposure of cultured neurons to $10{\mu}M$ $A\beta$ (25-35) for 24 h induced a neuronal cell death, which was measured by a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay and Hoechst 33342 staining. KM (10, 30 and $50{\mu}g/ml$) significantly inhibited the $A\beta$ (25-35)-induced apoptotic neuronal death. KM ($50{\mu}g/ml$) inhibited 10 μM Aβ (25-35)-induced elevation of intracellular calcium concentration ([Ca²⁺]_i), which was measured by a fluorescent dye, Fluo-4 AM. Glutamate release into medium and generation of reactive oxygen species (ROS) induced by $10{\mu}M$ $A\beta$ (25-35) were also inhibited by KM (10, 30 and $50{\mu}g/ml$). These results suggest that KM may mitigate the $A\beta$ (25-35)-induced neurotoxicity by interfering with the increase of [Ca²⁺]_i and then inhibiting glutamate release and generation of ROS in cultured neurons. In addition, orally administered KM (25 and 50 mg/kg, 7 days) significantly prevented memory impairment induced by intracerebroventricular injection of $A\beta$ (25-35) (8 nmol). Taken together, it is suggested that anti-dementia effect of KM is due to its neuroprotective effect against $A\beta$ (25-35)-induced neurotoxicity and that KM may have therapeutic role in prevention of the progression of Alzheimer's disease.

• 한국약용작물학회지
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• 제14권2호
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• pp.70-76
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• 2006

Paeoniae radix has been widely used for its anti-allergic, anti-inflammatory and analgesic effects, and demonstrated to have anticonvulsant, memory enhancing and anxiolytic activities. The present study was performed to examine the protective effect of methanol extract of Paeoniae radix (PR) from Paeoniae Japonica Miyabe et Takeda (Paeoniaceae) on hydrogen peroxide $(H_2O_2)-induced$ neurotoxicity using cultured rat cerebral cortical neuron. $H_2O_2$ produced a concentration-dependent reduction of neuronal viability, PR, over a concentration range of 10 to $100\;{\mu}g/ml$ showed concentration-dependent decrease of the $H_2O_2$$(100\;{\mu}M)-induced$ neuronal cell death, as assessed by a 3-[4,5-dimethylthiazol-2-yl]-2,5-di-phenyl-tetrazolium bromide assay and the number of apoptotic nuclei, evidenced by Hoechst 33342 staining. PR $(100\;{\mu}g/ml$ inhibited $100\;{\mu}M$ $H_2O_2-induced$ elevation of the cytosolic $Ca^{2+}$ concentration $([Ca^{2+}]_c)$, which was measured by a fluorescent dye, flue-4 AM. PR $(50\;{\mu}g/ml$ inhibited glutamate release into medium induced by $100\;{\mu}M$ $H_2O_2$, which was measured by HPLC, and generation of reactive oxygen species (ROS). These results suggest that PR may mitigate the $H_2O_2-induced$ neurotoxiciy by interfering with the increase of $[Ca^{2+}]_c$, and then inhibiting glutamate release and generation of ROS in cultured neurons.

• Nutrition Research and Practice
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• 제5권5호
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• pp.389-395
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• 2011

In the present study, a suitable drying method was developed for citrus press cakes (CPCs), which are produced as a by-product in citrus juice plants, and the protective effect of methanol extract of CPCs prepared by far-infrared radiation (FIR) drying against $H_2O_2$-induced DNA damage was evaluated versus that of freeze-dried CPCs. Methanol extract of FIR-dried CPCs exhibited comparatively good ROS scavenging activity versus the freeze-dried CPCs at the concentration of 100 ${\mu}g$/mL. The extract strongly enhanced the cell viability against $H_2O_2$-induced oxidative damage in Vero cells. Lipid peroxidation inhibitory activity of the extract from FIR-dried CPCs was comparable to that of the extract from freeze-dried CPCs. This sample also exhibited good protective effects against $H_2O_2$-mediated cell apoptosis as demonstrated by decreased apoptotic body formation in the nuclear staining with Hoechst 33342. In the comet assay, the CPC extracts exhibited strong inhibitory effects against $H_2O_2$-mediated DNA damage in a dose-dependent manner. Thus, this study demonstrated that FIR drying effectively preserves CPC as a functionally important natural antioxidant source and the FIR drying can be adapted for drying CPCs and is more economical for massive production than freeze drying.

• 한국약용작물학회지
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• 제15권2호
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• pp.105-111
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• 2007

The protective effect of ethanol extract of Korean mistletoe (KM; Viscum album coloratum) on hydrogen peroxide $(H_{2}O_{2})-induced$ neurotoxicity was examined in primary cultured rat cortical neurons. $H_{2}O_{2}$ reduced viability of cortical neurons in a concentration-dependent manner. The addition of KM, over a concentration range of 10 to 100 ${\mu}g/ml$, concentration-dependently prevented the $H_{2}O_{2}(100\;{\mu}M)-induced$ neuronal cell death, as assessed by a 3-[4,5-dimethylthiazol-2-yl]-2,5-di-phenyl-tetrazolium bromide (MTT) assay and Hoechst 33342 staining. KM significantly inhibited $H_{2}O_{2}-induced$ elevation of the cytosolic $Ca^{2+}$ concentration $([Ca^{2+}]_{c})$, which was measured by a fluorescent dye, fluo-4 AM. KM inhibited glutamate release into medium and generation of reactive oxygen species (ROS) induced by $H_{2}O_{2}$. These results suggest that KM may mitigate the $H_{2}O_{2}-induced$ neurotoxiciy by interfering with the increase of $[Ca^{2+}]_{c}$, and inhibiting glutamate release and generation of ROS in cultured neurons.

• ALGAE
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• 제27권3호
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• pp.205-213
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• 2012

The thermostability of antioxidant activity of dieckol, a phlorotannin isolated from brown seaweed Ecklonia cava was investigated. The thermostable antioxidant properties of dieckol were evaluated at 30, 60, and $90^{\circ}C$ for 7 days using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radical scavenging activities, and comparing its performance to that of ascorbic acid. The intracellular reactive oxygen species (ROS) scavenging activity and apoptotic body formation were investigated using DCF-DA assay and nuclear staining with Hoechst 33342, propidium iodide and flow cytometry. Dieckol treated at different temperatures during 7 days showed stable scavenging activities on towards DPPH and hydroxyl radicals. In addition, dieckol showed a stable protective effect against $H_2O_2$-induced apoptotic body formation in Vero cells. On the other hand, the radical scavenging activities and intracellular ROS scavenging activities of ascorbic acid, used as a positive control, were significantly decreased at $60^{\circ}C$ and $90^{\circ}C$ from on the 4th day and 3rd days, respectively. In conclusion, the results indicated that food grade antioxidant extracts containing dieckol derived from E. cava remain a stable during the temperatures encountered during the processing of food and cosmetics.

• Asian Pacific Journal of Cancer Prevention
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• 제16권17호
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• pp.7943-7957
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• 2015

Background and Aims: Colorectal cancer is one of the leading causes of death in the world. The aim of this study was to investigate the growth-suppression potentiality of a crude saponin extract (CSENS) prepared from medicinal herb, Nigella sativa, on human colon cancer cells, HCT116. Materials and Methods: HCT116 cells were subjected to increasing doses of CSENS for 24, 48 and 72 h, and then harvested and assayed for cell viability by WST-1. Flow cytometry analyses, cell death detection ELISA, fluorescent stains (Hoechst 33342 and acridine orange/ethidium bromide), DNA laddering and comet assays were carried out to confirm the apoptogenic effects of CSENS. Luciferase reporter gene assays, quantitative reverse transcription-polymerase chain reaction and Western blot analyses were performed to assess the impact of CAERS and CFEZO on the expression levels of key regulatory proteins in HCT116 cells. Results: The results demonstrated that CSENS inhibited proliferation and induced apoptosis. Apoptosis was confirmed by flow cytometry analyses, while CSENS-treated cells exhibited morphological hallmarks of apoptosis including cell shrinkage, irregularity in cellular shape, cellular detachment and chromatin condensation. Biochemical signs of apoptosis, such as DNA degradation, were observed by comet assay and gel electrophoresis. The pro-apoptotic effect of CSENS was caspase-3-independent and associated with increase of the Bax/Bcl-2 ratio. CSENS treatment down-regulated transcriptional and DNA-binding activities of NF-${\kappa}B$ and AP-1 proteins, associated with down-regulation of their target oncogenes, c-Myc, cyclin D1 and survivin. On the other hand, CSENS up-regulated transcriptional and DNA-binding activities of Nrf2 and expression of cytoprotective genes. In addition, CSENS modulated the expression levels of ERK1/2 MAPK, p53 and p21. Conclusions: These findings suggest that CSENS may be a valuable agent for treatment of colon cancer.

• 한국약용작물학회지
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• 제17권1호
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• pp.68-74
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• 2009

Vitis amurensis (VA; Vitaceae) has long been used in oriental herbal medicine. It has been reported that roots and seeds of VA have anti-inflammatory and antioxidant effects. In the present study, the protective effect of ethanol extract from stems and leaves of VA on hydrogen peroxide (${H_2}{O_2}$) (100 ${\mu}M$)-induced neuronal cell damage was examined in primary cultured rat cortical neurons. VA (10-100 ${\mu}g$/ml) concentration-dependently inhibited ${H_2}{O_2}$-induced apoptotic neuronal cell death measured by 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay and Hoechst 33342 staining. VA inhibited ${H_2}{O_2}$-induced elevation of intracellular $Ca^{2+}$ concentration (${[Ca^{2+}]}_i$) and generation of reactive oxygen species (ROS), which were measured by fluorescent dyes. Pretreatment of VA also prevented glutamate release into medium induced by 100 ${\mu}M$ ${H_2}{O_2}$, which was measured by HPLC. These results suggest that VA showed a neuroprotective effect on ${H_2}{O_2}$-induced neuronal cell death by interfering with ${H_2}{O_2}$-induced elevation of ${[Ca^{2+}]}_i$, glutamate release, and ROS generation. This has a significant meaning of finding a new pharmacological activity of stems and leaves of VA in the CNS.