• Clinical and Experimental Pediatrics
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• 제49권4호
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• pp.439-445
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• 2006

목 적 : ${\alpha}$-Galactosylceramide (GalCer)로 자극한 자연살상 T 세포(NKT)는 CD1d 및 T 세포 수용체(T cell receptor) 의존적으로 일부 백혈병에서 항암효과를 발현하나, CD1d음성인 신경모세포종에서는 세포독성을 유도할 수 없다. 이들 NKT세포의 활성화 시 분비되는 많은 양의 사이토카인의 직접적인 항암효과에 대해서는 소수의 보고가 있다. 본 연구에서는 hCD1d/${\alpha}$-GalCer tetramer로 NKT세포를 자극하여 얻은 상청액(supernatant)을 이용하여 NKT세포에 의한 신경모세포종의 치료적 접근의 가능성을 알아보았다. 방 법 : 신경모세포종 세포 주를 IMDM 배지에 배양하였고, NKT세포에서 분비되는 사이토카인 양은 cytometric bead array (CBA)분석으로 측정하였다. 세포 생존율은 calcein-AM 형광물질을 이용하여 digital image microscopy scanning (DIMSCAN)으로 측정하였고 특이 세포고사(specific apoptosis)는 annexin V and 7-AAD 염색 후 유식세포분석기를 통하여 산출하였다. 결 과 : 활성화된 NKT세포는 많은 양의 IL-2, IL-4, INF-${\gamma}$와 TNF-${\alpha}$을 분비하였다. NKT 자극 후 얻어진 상청액은 8개의 신경모세포종 세포 주 중 4개에서 의미있는 세포독성을 나타냈으며, 그 기전은 annexin-V 염색이나 pancaspase 억제제의 전 처치 실험으로 세포고사을 통하여 유도됨을 알 수 있었다. 그리고 이들 세포고사 유도는 anti-TNF-${\alpha}$, anti-IFN-${\gamma}$ 중화항체의 단독투여 시 현저히 감소하였고 동시투여 시에는 완전하게 억제되었다. 결 론 : NKT 세포의 활성화에 의해 분비된 IFN-${\gamma}$와 TNF${\alpha}$가 일부 신경모세포종 세포 주에서 협동적 세포 독성을 유도하였다.

• 자연과학논문집
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• 제9권1호
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• pp.25-29
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• 1997

Glucocorticoid에 의한 항염증 작용의 second messenger로 생각되어지는 annexin superfamily중 하나인 37 kDa의 단백질, lipocortin-1의 작용기작을 이해할 목적으로 in vivo에서 protein-protein interaction을 인식하는 yeast-based genetic assay인 yeast two assay를 통하여 lipocortin-1과 결합하는 단백질 유전자를 분리하여 조사하였다. 이 방법으로 실험을 수행한 결과 분리된 유전자가 human serine proteinase 유전자와 homology가 있는 것으로 밝혀졌다.

• 대한자기공명의과학회:학술대회논문집
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• 대한자기공명의과학회 2004년도 제9차 학술대회 초록집
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• pp.23-32
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• 2004

The chemotherapy sensitive Lewis lung carcinoma (LLC) and chemotherapy resistant Lewis lung carcinoma (CR-LLC) tumors concurrently implanted in mice, and compare these findings with histological macroscopic observations against 3D reconstruction of Fluorescence Molecular Tomography (FMT) preformed in vivo on the same animals. For the 3D image reconstruction we used 32 laser source images, a flat image and 3D surface rendering that confused for 3D Fluorescence Molecular Imaging (FMI). A minimum of ten tissue sections were analyzed per tumor for quantification of the TUNEL-positive cells, cell-associated Cy5.5-Annexin and vessel-associated Alexa Fluor-Lectin. These are useful apoptosis and angiogenesis markers, and they serve as validation experiments to data obtained in vivousing a Cy5.5-Annexin V conjugate injected intravenously in chemotherapy-treated animals carrying the tumors studied histologically. We detected higher levels of apoptosis and corresponding higher levels of Cy5.5 fluorescence in the LLC vs. the CR-LLC tumors according to tissue depth and these findings confirm that in vivo staining with the Cy5.5-Annexing conjugate correlates well with in vitro TUNEL staining and is consistent with the higher apoptotic index expected from the LLC line. There appeared to be 1.38% more apoptosis for LLC than CR-LLC. Consequently there is good correlation between the histology results and in vivo fluorescence-mediated optical imaging. In conclusion the apoptotic images of 3D FMI were validated by microscopic histological image analysis. This is a significant result for the continuous progress of fluorescence 3D imaging research.

• Asian Pacific Journal of Cancer Prevention
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• 제16권13호
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• pp.5265-5271
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• 2015

Background: The effects of plant products on cancer cells has become a field of major importance. Many substancesmay induce apoptosis in anti-cancer treatment. Pectins, a family of complex polysaccharides, and their degradation products may for exasmple exert apoptotic effects in cancer cells. Apples and citrus fruits are the main sources of pectin which can be applied for anti-cancer research. The present study concerned an intact form of pectic-oligoshaccharide named pectic acid (poly galactronic acid). Materials and Methods: Inhibition of cell proliferation assays (MTT), light microscopy, fluorescence microscopy (acridin orange/ethidium bromide), DNA fragmentation tests, cell cycle analysis, annexin PI and Western blotting methods were applied to evaluate apoptosis. Results: The results indicated that pectic acid inhibited cell growth and reduced cell attachment after 24h incubation. This did not appear to be due to necrosis, since morphological features of apoptosis were detected with AO/EB staining and cell cycling was blocked in the sub-G1 phase. Annexin/PI and DNA fragmentation findings indicated that apoptosis frequency increased after 24h incubation with pectic acid. In addition, the data showed pectic acid induced caspase-dependent apoptosis. Conclusions: These data indicate that apple pectic acid without any modification could trigger apoptosis in MDA-MB-231 human breast cancer cells and has potential to improve cancer treatment as a natural product.

• The Korean journal of internal medicine
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• 제34권1호
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• pp.146-155
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• 2019

Background/Aims: Indoxyl sulfate (IS) is a uremic toxin and an important causative factor in the progression of chronic kidney disease. Recently, paricalcitol (19-nor-1,25-dihydroxyvitamin D2) was shown to exhibit protective effects in kidney injury. Here, we investigated the effects of paricalcitol treatment on IS-induced renal tubular injury. Methods: The fluorescent dye 2',7'-dichlorofluorescein diacetate was used to measure intracellular reactive oxygen species (ROS) following IS administration in human renal proximal tubular epithelial (HK-2) cells. The effects of IS on cell viability were determined using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays and levels of apoptosis-related proteins (Bcl-2-associated protein X [Bax] and B-cell lymphoma 2 [Bcl-2]), nuclear $factor-{\kappa}B$ ($NF-{\kappa}B$) p65, and phosphorylation of mitogen-activated protein kinase (MAPK) and protein kinase B (Akt) were determined by semiquantitative immunoblotting. The promoter activity of $NF-{\kappa}B$ was measured by luciferase assays and apoptosis was determined by f low cytometry of cells stained with f luorescein isothiocyanate-conjugated Annexin V protein. Results: IS treatment increased ROS production, decreased cell viability and induced apoptosis in HK-2 cells. IS treatment increased the expression of apoptosis-related protein Bax, decreased Bcl-2 expression, and activated phosphorylation of MAPK, $NF-{\kappa}B$ p65, and Akt. In contrast, paricalcitol treatment decreased Bax expression, increased Bcl-2 expression, and inhibited phosphorylation of MAPK, $NF-{\kappa}B$ p65, and Akt in HK-2 cells. $NF-{\kappa}B$ promoter activity was increased following IS, administration and was counteracted by pretreatment with paricalcitol. Additionally, flow cytometry analysis revealed that IS-induced apoptosis was attenuated by paricalcitol treatment, which resulted in decreased numbers of fluorescein isothiocyanate-conjugated Annexin V positive cells. Conclusions: Treatment with paricalcitol inhibited IS-induced apoptosis by regulating MAPK, $NF-{\kappa}B$, and Akt signaling pathway in HK-2 cells.

• KSBB Journal
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• 제32권2호
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• pp.83-89
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• 2017

Gamma (${\gamma}$)-oryzanol is a substance abundant in rice, which is widely cultivated in Asian countries. In this study, we evaluated the effect of ${\gamma}$-oryzanol treatment on proliferation and apoptosis of AGS human gastric carcinoma cells. AGS cells were treated with ${\gamma}$-oryzanol for 72 h in a dose dependent manner. Treatment of ${\gamma}$-oryzanol (50, 100, and $200{\mu}g/mL$) resulted in decreased AGS cell proliferation and increased number of cells in the sub-G1 population. Additionally, apoptotic cells were investigated by annexin V staining and mitochondrial membrane potential assays. Our results indicated that ${\gamma}$-oryzanol treatment increased the number of annexin V-positive cells and depolarized cells. This demonstrated that ${\gamma}$-oryzanol is effective for the induction of apoptosis in AGS cells. We next examined the expression of promising anticancer drug target molecules, including PTEN and HSP90. We found that treatment of ${\gamma}$-oryzanol induced the expression of PTEN in AGS cells. Under the same treatment conditions, ${\gamma}$-oryzanol reduced the expression of HSP90 in AGS cells. These results suggest that ${\gamma}$-oryzanol-induced apoptosis was accompanied by changes in regulation of PTEN and HSP90 in AGS cells. Taken together, ${\gamma}$-oryzanol could be used as a functional substance for the prevention of gastric cancer.

• Journal of Ginseng Research
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• 제45권1호
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• pp.126-133
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• 2021

Background: 20(S)-protopanaxadiol (20(S)-PPD), one of the aglycone derivatives of major ginsenosides, has been shown to have an anticancer activity toward a variety of cancers. This study was initiated with an attempt to evaluate its anti-cancer activity toward human endometrial cancer by cell and xenograft mouse models. Methods: Human endometrial cancer (HEC)-1A cells were incubated with different 20(S)-PPD concentrations. 20(S)-PPD cytotoxicity was evaluated using MTT assay. Apoptosis was detected using the annexin V binding assay and cell cycle analysis. Cleaved poly (ADP-ribose) polymerase (PARP) and activated caspase-9 were assessed using western blotting. HEC-1A cell tumor xenografts in athymic mice were generated by inoculating HEC-1A cells into the flank of BALB/c female mice and explored to validate 20(S)-PPD anti-endometrial cancer toxicity. Results: 20(S)-PPD inhibited HEC-1A cell proliferation in a dose-dependent manner with an IC50 value of 3.5 μM at 24 h. HEC-1A cells morphologically changed after 20(S)-PPD treatment, bearing resemblance to Taxol-treated cells. Annexin V-positive cell percentages were 0%, 10.8%, and 58.1% in HEC-1A cells when treated with 0, 2.5, and 5 μM of 20(S)-PPD, respectively, for 24 h. 20(S)-PPD subcutaneously injected into the HEC-1A cell xenograft-bearing mice three times a week for 17 days manifested tumor growth inhibition by as much as 18% at a dose of 80 mg/kg, which sharply contrasted to controls that showed an approximately 2.4-fold tumor volume increase. These events paralleled caspase-9 activation and PARP cleavage. Conclusion: 20(S)-PPD inhibits endometrial cancer cell proliferation by inducing cell death via a caspase-mediated apoptosis pathway. Therefore, the 20(S)-PPD-like ginsenosides are endowed with ample structural information that could be utilized to develop other ginsenoside-based anticancer agents.

• 생명과학회지
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• 제26권9호
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• pp.1015-1021
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• 2016

Pyrimidine 유도체의 일종인 5-fluorouracil (5-FU)은 광범위하게 사용되는 항암제의 일종으로, thymidylate synthase의 활성을 억제시켜 핵산의 합성 및 대사기능 자애 유발 물질이다. 본 연구에서는 Ewing′s 육종 CHP-100 세포에서 5-FU의 증식억제와 연관된 기전 해석으로 시도하였다. 본 연구의 결과에 의하면, 5-FU 처리 시간의 경과에 따른 CHP-100 세포의 증식억제가 세포주기 G1 arrest 유발에 따른 것임을 알 수 있었다. 5-FU에 의한 CHP-100 세포의 G1 arrest는 retinoblastoma protein (pRB)의 탈인산화에 따른 전사인자 E2F-1 및 E2F-4와의 결합 촉진과 연관성이 있었다. 비록 5-FU 처리가 cyclin-dependent kinases의 발현에는 크게 영향을 주지 않았으나, 정상배지에서 배양된 대조군에 비하여 cyclin A 및 B의 발현이 5-FU 처리 시간 의존적으로 억제되었다. 또한 5-FU에 의한 CHP-100 세포의 G1 arrest는 apoptosis 유도와 연관이 있음을 핵 내 염색질의 응축에 따른 apoptotic body의 형성증가, poly (ADP-ribose) polymerase의 단편화 및 annexin V 염색 등을 통하여 확인하였다. 아울러 5-FU는 pro-apoptotic Bax 단백질의 발현 증가 및 anti-apoptotic Bcl-2의 발현 감소를 통한 mitochondrial membrane potential의 소실을 촉진시켰으며, 이로 인하여 미토콘드리아에서 세포질로의 cytochrome c 유리가 증가시켰음을 알 수 있었다. 따라서 본 연구의 결과는 5-FU에 의한 CHP-100 세포의 증식억제와 연관된 G1 arrest 및 apoptosis 유도에는 pRB의 인산화 억제 및 미토콘드리아 기능의 손상이 최소한 관여하고 있음을 의미하는 것이다.

• Natural Product Sciences
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• 제25권1호
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• pp.23-27
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• 2019

From the pericarps of Litsea japonica (Thunb.) Jussieu, eighteen butanolide derivatives (1 - 18) were evaluated for their cytotoxic activity against HeLa, HL-60, and MCF-7 cells. Compounds 1-9 with 2-alkylidene-3-hydroxy-4-methylbutanolides structure exhibited cytotoxic activities against cancer-cell lines. Among them, compound 8 (litsenolide $D_2$) exhibited the most potent cytotoxicity against the tested cell lines, including HeLa, HL-60, and MCF-7, with $IC_{50}$ values of $17.6{\pm}1.3$, $4.2{\pm}0.2$, and $12.8{\pm}0.0{\mu}M$, respectively. Compound 8 induced apoptosis in a dose-dependent manner. Annexin V/Propidium Iodide (PI) double staining confirmed that 8 effectively induced apoptosis in MCF-7 cells. To the best of our knowledge, we have reported cytotoxic activity of butanolides from L. japonica against these cancer-cell lines for the first time.

• 생명과학회지
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• 제24권1호
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• pp.92-97
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• 2014

Cordycepin은 Cordyceps militaris에서 처음 유래된 nucleoside adenosine 유도체의 일종으로 면역증강 및 항암활성을 포함한 다양한 약리 기능이 있는 것으로 알려져 있다. 본 연구에서는 LNCap 인체 전립선 암세포 모델을 이용하여 cordycepin에 의한 항암활성 기전을 연구하였다. Cordycepin 처리에 따라 LNCap 세포는 처리 농도 의존적으로 증식이 억제되었으며, 이는 apoptosis 유발과 연관성이 있음을 poly ADP-ribose polymerase의 단편화 현상과 Annexin V 염색에 의한 정량적 분석으로 확인하였다. Cordycepin 처리에 따른 flow cytometric analysis 결과로서 cordycepin이 세포주기 G2/M기 정체 현상을 유발하였음을 알 수 있었으며, 이는 cyclin B1 및 cyclin A의 발현 감소와 연관성이 있었다. 또한 cordycepin이 처리된 LNCap 세포에서 cyclin-dependent kinase (CDK) inhibitor p21Waf1/Cip1의 발현이 증가되었지만, CDK2, CDC2 및 Cdc25C의 발현에는 큰 영향을 미치지 않았으며, cordycepin에 의하여 증가된 p21 단백질은 CDK2 및 CDC2와의 복합체를 형성하고 있었다. 본 연구의 결과는 LNCap 전립선 암세포에서 cordycepin에 의한 G2/M 및 apoptosis 유발은 p53 비존적인 CDK inhibitor p21의 발현 증가가 중요한 역할을 하고 있음을 보여주는 것이다.