• 대한한의학방제학회지
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• 제25권2호
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• pp.179-191
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• 2017

Objectives : Increased oxidative stress by reactive oxygen species (ROS) has been suggested as a major cause of muscle fatigue. Although several studies have demonstrated the various biological properties of Sophora flavescens Aiton, Glycyrrhiza uralensis Fischer and Dictamnus dasycarpus Turcz, but the antioxidative potentials have not been clearly demonstrated. The present study was designed to investigate the protective effects of their water and ethanol extract mixtures (medicinal herbal mixtures, MHMIXs) on hydrogen peroxide ($H_2O_2$)-induced cell damage and apoptosis in C2C12 myoblasts. Methods : Cytotoxicity was assessed by an MTT assay. Quantitative evaluation of apoptosis induction and ROS production was evaluated by flow cytometry analysis. Expression levels of apoptosis regulatory and DNA-damage proteins were detected by Western blotting. Result : The inhibition of $H_2O_2$-induced cell proliferation was effectively blocked in extracts of 3: 1: 1 (EMHMIXs-1) or 2: 2: 1 (EMHMIXs-2) of S. flavescens, G. uralensis and D. dasycarpus Turcz, ethanol extracts from various complex extracts in C2C12 myoblasts. EMHMIXs-1 and EMHMIXs-2 also effectively attenuated $H_2O_2$-induced C2C12 cell apoptosis, which was associated with the restoration of the upregulation of Bad and death receptor 4, and downregulation of XIAP and cIAP-1 induced by $H_2O_2$. In addition, these herbal mixtures significantly blocked the $H_2O_2$-induced ROS generation and phosphorylation of $p-{\gamma}H2A.X$, which suggests that they can prevent $H_2O_2$-induced cellular DNA damage. Conclusions : The results suggest that EMHMIXs-1 and EMHMIXs-2 could block the DAN damage and apoptosis of C2C12 myoblasts by oxidative stress through blocking ROS generation.

• Asian Pacific Journal of Cancer Prevention
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• 제16권2호
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• pp.465-469
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• 2015

Background: Orostachys japonicus A. Berger (A. Berger) is commonly used as a folk remedy for cancer therapy. However, the mechanisms of its anti-cancer activity are poorly investigated in human cancer cells. In this study, we investigated whether flavonoids extracted from Orostachys japonicus A. Berger (FEOJ) might have anticancer effects in human leukemia cells, focusing on cell death mechanisms. Materials and Methods: U937 human leukemic cancer cells were used. Results: FEOJ induced apoptosis in a dose-dependent manner in human U937 cancer cells. Flow cytometry revealed significant accumulation of cells with sub-G1 DNA content at the concentrations of $200{\mu}g/mL$ and $400{\mu}g/mL$. FEOJ-induced apoptosis was caspase-dependent through loss of mitochondrial membrane potential (MMP, ${\Delta}{\Psi}m$) in human U937 cancer cells, which might be associated with suppression of Bcl-2 and XIAP proteins. FEOJ induced the p38 MAPK signaling pathway, playing at least in part an important role in FEOJ-induced apoptosis. Conclusions: This study suggested that FEOJ may induce caspase-dependent apoptosis in human leukemic cells by regulating MMP (${\Delta}{\Psi}m$) through suppressing Bcl-2 and X-IAP. In addition, the results indicated that upstream p38 MAPK signaling regulates the apoptotic effect of FEOJ. This study provides evidence that FEOJ might have anti-cancer potential for human leukemic cells.

• Asian Pacific Journal of Cancer Prevention
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• 제14권10호
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• pp.6115-6120
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• 2013

Introduction: Some non-small cell lung cancer (NSCLC) tumor cells are insensitive to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) -based therapy. This study was conducted to examine the effect of embelin on the sensitivity of the A549 NSCLC cell line to TRAIL receptor2 (TRAILR2) monoclonal antibodies and to investigate the potential mechanisms. Materials and Methods: A549 cells were treated with embelin, TRAILR2 mAb or a combination of both. Cell viability was measured using ATPlite assay and apoptosis rates were determined by flow cytometry with AnnexinV-FITC and propidium iodide staining, with the expression levels of proteins analyzed by Western blotting. Results: The cell survival rate of separate treatments with 100 ng/ml TRAILR2 antibody or 25 uM embelin were $81.5{\pm}1.57%$ and $61.7{\pm}2.84%$, respectively. Their combined use markedly decreased cell viability in A549 cells to $28.1{\pm}1.97%$ (P<0.05). The general caspase inhibitor Z-VAD-FMK could inhibit the embelin-enhanced sensitivity of A549 cells to TRAILR2 mAb ($75.97{\pm}3.17%$)(P<0.05). Both flow cytometry and cell morphological analysis showed that embelin was able to increase TRAIL-induced apoptosis in A549 cells. Combined treatment with embelin and TRAILR2 mAb augmented the activation of initiator caspases and effector caspase. In addition, A549 cells showed increasing levels of TRAILR2 protein and decreasing levels of Bcl-2, survivin and c-FLIP following the treatment with embelin+TRAILR2 mAb. Conclusions: Embelin could enhance TRAIL-induced apoptosis in A549 cells. The synergistic effect of the combination treatment might be due to modulation of multiple components in the TRAIL receptor-mediated apoptotic signaling pathway, including TRAILR2, XIAP, survivin, Bcl-2 and c-FLIP.

• Journal of Microbiology and Biotechnology
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• 제28권6호
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• pp.1007-1021
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• 2018

Cancer represents one of the most significant threats to human health on a global scale. Hence, the development of effective cancer prevention strategies, as well as the discovery of novel therapeutic agents against cancer, is urgently required. In light of this challenge, this research aimed to evaluate the effects of several potent bioactive peptides and proteins contained in crocodile white blood cell extract (cWBC) against LU-1, LNCaP, PC-3, MCF-7, and CaCo-2 cancer cell lines. The results demonstrate that 25, 50, 100, and $200{\mu}g/ml$ cWBC exhibits a strong cytotoxic effect against all investigated cell lines ($IC_{50}$ $70.34-101.0{\mu}g/ml$), while showing no signs of cytotoxicity towards noncancerous Vero and HaCaT cells. Specifically, cWBC treatment caused a significant reduction in the cancerous cells' colony forming ability. A remarkable suppression of cancerous cell migration was observed after treatment with cWBC, indicating potent antimetastatic properties. The mechanism involved in the cancer cell cytotoxicity of cWBC may be related to apoptosis induction, as evidenced by typical apoptotic morphology features. Moreover, certain cWBC concentrations induced significant overproduction of ROS and significantly inhibited the $S-G_2/M$ transition in the cancer cell. The molecular mechanisms of cWBC in apoptosis induction were to decrease Bcl-2 and XIAP expression levels and increase the expression levels of caspase-3, caspase-8, and p53. These led to a decrease in the expression level of the cell cycle-associated gene cyclin-B1 and the arrest of cell population growth. Consequently, these findings demonstrate the prospect of the use of cWBC for cancer therapy.

• 한국식품영양과학회지
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• 제40권5호
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• pp.649-659
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• 2011

연구에서는 콩다닥냉이 추출물의 항암활성을 조사하기 위하여 잎 및 뿌리의 열수 추출물(WELVL 및 WELVR)이 HCT116 대장암세포의 증식 억제와 연관된 apoptosis 유도 기전에 관한 연구를 시도하였다. 본 연구의 결과에 의하면 HCT116 세포에 WELVL 및 WELVR을 처리하였을 경우에 유발되는 증식 억제 및 형태 변화는 apoptosis 유발과 밀접한 연관이 있었으며, 증식억제 및 apoptosis 유도 효과는 WELVL에 비하여 WELVR에서 높게 나타났다. 특히 WELVR에 의한 apoptosis 유발에는 FasL의 발현 증가를 통한 caspase-8의 활성화와 이로 인한 Bid 단백질의 단편화와 함께 Bcl-2 family의 발현 변화를 통한 mitochondria의 기능 이상과 이로 인한 caspase-9 및 -3의 활성화, 그리고 기질단백질들의 분해가 중요한 역할을 하는 것으로 나타났다. 또한 IAP family의 발현 감소로 인한 caspase의 활성 증가도 어느 정도 관여하는 것으로 생각되어진다. 따라서 WELVR 처리에 의하여 유발되는 apoptosis는 extrinsic pathway 및 intrinsic pathway를 모두 경유하는 multiple apoptotic pathway에 의하여 조절되는 것으로 생각되며, 이러한 결과들은 인체 암세포에서 콩다닥냉이의 항암작용을 이해하는데 중요한 자료가 될 것이고 나아가 콩다닥냉이 추출물을 포함한 그와 유사한 항암제 후보물질들의 연구 기초자료로서 사용될 수 있을 것으로 생각된다.

• 한국해양바이오학회지
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• 제1권2호
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• pp.63-70
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• 2006

본 연구에서는 U937 인체 백혈병 세포의 증식에 미치는 PTX-2의 영향을 조사한 결과, PTX-2의 처리에 따라 U937 세포는 처리 농도 및 처리 시간 의존적으로 심한 형태적 변형과 함께 증식이 억제되었다. 이러한 PTX-2 처리에 의한 U937 세포의 증식억제는 apoptosis 유발과 관련이 있었으며, 이를 DAPI staining에 의한 apoptotic body 형성, flow cytometry를 이용한 sub-G1 세포 빈도의 정량적 분석을 통하여 확인하였다. 이러한 PTX-2 처리에 의한 U937 세포의 apoptosis 유발은 Bcl-2 family에 속하는 anti-apoptotic 인자인 Bcl-$X_L$의 발현 감소 및 IAPs family에 속하는 유전자들의 선택적 발현 감소와 연관성이 있음을 알 수 있었다. 이상의 결과들은 인체 암세포에서 PTX-2의 항암작용을 이해하는데 중요한 자료가 될 것이고 나아가 PTX-2을 포함한 그와 유사한 항암제 후보물질들의 연구에 있어서 기초 자료로서 사용될 수 있을 것으로 생각된다.

• Journal of Ginseng Research
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• 제39권1호
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• pp.46-53
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• 2015

Background: Glucocorticoids (GCs) are commonly used in many chemotherapeutic protocols and play an important role in the normal regulation of bone remodeling. However, the prolonged use of GCs results in osteoporosis, which is partially due to apoptosis of osteoblasts and osteocytes. In this study, effects of Korean Red Ginseng (KRG) on GC-treated murine osteoblastic MC3T3-E1 cells and a GC-induced osteoporosis mouse model were investigated. Methods: MC3T3-E1 cells were exposed to dexamethasone (Dex) with or without KRG and cell viability was measured by the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Realtime polymerase chain reaction was performed to evaluate the apoptotic gene expression; osteogenic gene expression and alkaline phosphatase (ALP) activity were also measured. Western blotting was performed to evaluate the mitogen-activated protein kinase (MAPK) proteins. A GC-induced osteoporosis animal model was used for in vivo study. Results and conclusion: The MTT assay revealed that Korean Red Ginseng (KRG) prevents loss of cell viability caused by Dex-induced apoptosis in MC3T3E1 cells. Real-time polymerase chain reaction data showed that groups treated with both Dex and KRG exhibited lower mRNA levels of caspase-3 and -9, whereas the mRNA levels of Bcl2, IAPs, and XIAP increased. Moreover, groups treated with both Dex and KRG demonstrated increased mRNA levels of ALP, RUNX2, and bone morphogenic proteins as well as increased ALP activity in MC3T3-E1 cells, compared to cells treated with Dex only. In addition, KRG increased protein kinase B (AKT) phosphorylation and decreased c-Jun N-terminal kinase (JNK) phosphorylation. Moreover, microcomputed tomography analysis of the femurs showed that GC implantation caused trabecular bone loss. However, a significant reduction of bone loss was observed in the KRG-treated group. These results suggest that the molecular mechanism of KRG in the GC-induced apoptosis may lead to the development of therapeutic strategies to prevent and/or delay osteoporosis.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2019년도 춘계학술대회
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• pp.111-111
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• 2019

Glycyrrhizae radix is one of the most frequently prescribed ingredients in Oriental medicine, and G. radix extract has been shown to exert anti-cancer effects. However, the cellular and molecular mechanisms of apoptosis by G. radix are poorly defined. In the present study, it was examined the biochemical mechanisms of apoptosis by water extract of G. radix (WEGR) in human bladder T24 cancer cells. It was found that WEGR could inhibit the cell growth of T24 cells in a dose-dependent manner, which was associated with the induction of apoptotic cell death, as evidenced by the formation of apoptotic bodies, DNA fragmentation and increased populations of annexin-V positive cells. The induction of apoptotic cell death by WEGR was connected with an up-regulation of pro-apoptotic Bax protein expression and down-regulation of anti-apoptotic Bcl-2 and Bcl-xL proteins, and inhibition of apoptosis family proteins (XIAP, cIAP-1 and cIAP-2). In addition, apoptosis-inducing concentrations of WEGR induced the activation of caspase-9, an initiator caspase of the mitochondrial-mediated intrinsic pathway, and caspase-3, accompanied by proteolytic degradation of poly (ADP-ribose)-polymerase. WEGR also induced apoptosis via a death receptor-mediated extrinsic pathway by caspase-8 activation, resulting in the down-regulation of total Bid and suggesting the existence of cross-talk between the extrinsic and intrinsic pathways. Taken together, the present results suggest that WEGR may be a potential chemotherapeutic agent for the control of human bladder cancer cells.

• 생명과학회지
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• 제19권9호
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• pp.1314-1320
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• 2009

히스톤 디아세틸라이제 저해제(HDACI)는 최근에 새로운 미래형 항암제로 주목을 받고 있으며 다른 항암요법 및 치료제와의 병용치료에도 효과적으로 적용될 수 있을 것으로 기대하고 있다. HDACI은 다양한 조직기원의 암세포에서 증식억제 및 세포사멸 유도능이 시험되어 왔으나 비소세포폐암 세포에서 그 작용 및 기전이 명확히 조사된 바가 없다. 본 연구는 HDACI 중의 하나인 sodium butyrate (SB)를 비소세포폐암 세포주인 H460에 처리하여 세포생존율, 세포주기 분석, 세포사멸도를 평가하고, 이와 관련하여 세포사멸 관련 단백질, p53, ERK의 변화를 조사하고자 하였다. 3가지 다른 농도(2.5, 7.5, 20 mM)의 SB에 H460 세포가 48시간 노출되었을 때, 세포 생존율은 농도의존적으로 감소되었으나 7.5 mM 이상의 농도에서 대조군에 비해 유의한 생존을 감소를 보였고, 20 mM에서 생존을 50% 전후를 나타냈다. SB노출은 H460 세포의 사멸을 유발하였는데, 세포사의 유형은 아포토시스와 괴사가 동시에 발생함이 Annexin-V 분석으로 확인되었다. H460 세포에서 SB에 의해 유발되는 뚜렷한 세포주기의 변화양상은 G2/M기정지였으며, 이러한 세포주기의 지연현상으로 세포사멸이 초래되는 것으로 생각된다. SB처리는 아포토시스 발생관련 효소인 caspase-3과 caspase-7의 활성화를 유발하였으며, 이에 의한 PARP 단백 질의 분절화도 관찰되었다. 동시에, 항세포사별 단백질인 XIAP의 단백질 함량은 감소함을 보였다. SB 노출에 의한 세포주기의 G2/M기의 정지현상과 관련하여는 p53 단백질의 증가가 주목할 만한 하였다. SB의 H460세포에의 처리는 일반 ERK단백질의 함량 변화를 유도하지 않았으나, 인산화형의 ERK는 SB처리농도에 의존적으로 그 단백질 함량이 감소하였다. 이는 ERK가 비소세포폐암 세포인 H460에서 세포생존 및 유지와 관련된 단백질의 인산화에 계속적으로 관여하고 있다는 사실을 암시한다. 즉, SB의 처리는 ERK의 인산화를 유의하게 억제하는 기전과 관련이 있을 가능성이 높다고 추측된다. 향후 SB의 노출에 의한 PERK 감소 기전에 대한 연구가 추가적으로 진행되면 SB의 더욱 효율적인 암세포 사멸 유도 전략수립에 도움을 줄 수 있을 것이라 예상된다.