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인간 전립선암 PC-3 세포에서 Compound K에 의한 세포주기 조절 및 세포사멸 유전자 발현 변화
김광연,박광일,안순철,Kim, Kwang-Youn,Park, Kwang-Il,Ahn, Soon-Cheol 대한한의학방제학회 2021 大韓韓醫學方劑學會誌 Vol.29 No.4
Objectives : Previously, we reported that compound K isolated from fermented ginseng by Aspillus oryzae has a wide biochemical and pharmacological effect, including anti-cancer activity in prostate cancer PC-3 cells. Despite these findings, its signaling pathway and gene expression pattern are not clearly understood. Methods : To confirm the gene expression study of treated with compound K in PC-3 cells, a cDNA microarray chip composed of 44K human cDNA probes was used. MTT assay, western blot analysis, propidium iodide staining, and annexin V/propidium iodide staining were analyzed. Results : We confirmed the differences of gene expression profiles. Then, we analyzed with the cell cycle arrest, cell death and cell proliferation related genes using DAVID database. Conclusions : Our finding should be useful for understanding genome-wide expression patterns of compound K-mediated cell cycle arrest toward induction of cell death and be helpful for finding future cancer therapeutic targets for prostate cancer cells.
필발(Piper longum Linn.)로부터 전립선암 세포사멸물질 pipernonaline의 분리 및 동정
김광연(Kwang-Youn Kim),김윤진(Yun-Jin Kim),이완(Wan Lee),유선녕(Sun-Nyoung Yu),조효진(Hyo-Jin Cho),이선이(Sun-Yi Lee),이한승(Han-Seung Lee),손재학(Jae-Hak Sohn),오현철(Hyuncheol Oh),안순철(Soon-Cheol Ahn) 한국생명과학회 2009 생명과학회지 Vol.19 No.5
Prostate cancer has been a critical health problem due to an increase of prostate cancer-related deaths worldwide. Also, a frequent treatment option for prostate cancer is androgen ablation, but this treatment has a limited scope, especially for hormone-refractory cancer. There is an urgent need for the identification of alternative therapeutic strategies for prostate cancer. Previously, over one hundred species of dried-plant methanol extracts were tested for inhibitory effects on proliferation. One of them, Piper longum Linn. was selected based on its potent anti-proliferation effect. The dried root of P. longum Linn. was extracted with 100% methanol for 2-3 days and its extract was fractionated using chloroform. The chloroform layer was then subjected to column chromatography on silica gel, reverse phase-18 (RP-18) and Sephadex LH-20, in turn. Finally, the pure compound was obtained and identified as pipernonaline by NMR spectroscopic and physico-chemical analysis. In this study, anti-proliferation and cell cycle arrest effects of pipernonaline on human prostate cancer PC-3 cells were investigated using the MTT and PI staining, respectively. Our findings suggest that pipernonaline represents a dose-dependent growth inhibition pattern on PC-3 cells and, moreover, its growth inhibition is associated with sub-G1 and G0/G1 cell cycle accumulation in PC-3 cells. Also, these results provide an anticancer candidate for human prostate cancer.
Ro-Ui Kim(김로의),Soon-Cheol Ahn(안순철),Sun-Nyoung Yu(유선녕),Kwang-Youn Kim(김광연),Jong-Hwan Seong(성종환),Young-Guen Lee(이영근),Han-Soo Kim(김한수),Dong-Seob Kim(김동섭) 한국생명과학회 2011 생명과학회지 Vol.21 No.2
본 연구의 목적은 두유 curd를 형성하는 미생물을 분리하는 것이다. 두유 curd를 형성하는 미생물은 채소를 젖산균으로 발효시킨 전통적인 한국의 음식, 김치로부터 분리하였다. 분리 균주 196개 중 10개의 균주(strain No. 2-2-2, 2-15-2, 2-18-1, 2-19-2, 3-4-1, 3-4-2, 3-8-1, 3-8-3, 3-17-1, 4-39-5)가 단단한 두유 curd를 형성하였고 분자생물학적?생화학적 분석법에 의해 동정되었다. 분리균주로부터 추출한 genomic DNA는 16S rDNA 지역의 PCR 증폭을 위한 주형으로 사용하였다. GenBank 데이터로 16S rDNA 염기서열을 비교한 결과, 분리 균주들은 Leuconostoc mesenteroides group과 Lactobacillus sakei group으로 동정되었다. 두유 curd를 형성하는 균주들의 계통 발생학적 위치와 분류군은 neighbor-joining 방법을 이용하여 확인하였다. 또한, L. mesenteroides group은 생화학적 특성에 의해 L. mesenteroides subsp. dextranicum으로 동정되었다. 하지만 L. sakei group은 생화학적 특성 비교시 다양성을 보여 Lactobacillus sp.로 명명하였다. The purpose of this study was to isolate soy curd forming bacterial strains. Soy curd forming bacteria were isolated from Kimchi, a traditional Korean vegetable food that is fermented using lactic acid bacteria. Among 196 bacterial strains, ten isolates (strain No. 2-2-2, 2-15-2, 2-18-1, 2-19-2, 3-4-1, 3-4-2, 3-8-1, 3-8-3, 3-17-1, 4-39-5) formed firm soy curd. The isolated bacterial strains were identified by molecular biological and biochemical analyses. The genomic DNAs extracted from the isolated bacterial strains were used as a template for PCR amplification of 16S rDNA region. By comparing the results of the 16s rDNA sequences with GenBank data, the isolated strains were identified as Leuconostoc mesenteroides group and Lactobacillus sakei group. The phylogenetic position of soy curd forming strains and their related taxa were investigated using neighbor-joining method. L. mesenteroides group was further identified as L. mesenteroides subsp. dextranicum based on biochemical properties. L. sakei group was named Lactobacillus sp., because it showed a variety of biochemical properties.
장정현,김광연,안순철,권헌영,Chang, Jeong-Hyun,Kim, Kwang-Youn,Ahn, Soon-Cheol,Kwon, Heun-Young Korean Society of Life Science 2007 생명과학회지 Vol.17 No.8
Paclitaxel이 암세포에서 세포예정사를 유발할지라도, 아직 정확한 기전은 잘 알려져 있지 않다. 이에 본 연구에서는 HeLa $S_{3}$ 자궁암세포에서의 paclitaxel이 어떠한 영향을 미치는지 알아보고자 한다. 그리하여 방법으로는 세포독성검사, apoptotic cells의 형태학적 변화(DAPI 염색 ), western blot 분석법을 사용하여 수행하였다. 본 연구의 결과로 paclitaxel은 HeLa $S_{3}$ 세포에서 세포독성을 보이며 특히 paclitaxel의 $IC_{50}$ 값은 약 1 ${\mu}M$이며, paclitaxel 처리한 HeLa $S_{3}$ 세포에서 형태학적 변화(분절화)를 관찰하였고, flow cytometric 분석에서는 G2/M기가 차단되어 paclitaxel은 세포주기 특히 Sub-$G_{1}$기를 조절함을 알 수 있다. 그리고 Paclitaxel을 처리한 HeLa $S_{3}$ 세포에서는 PARP cleavage를 유발하였고 Bc1-2의 감소와도 관련되었다. Although paclitaxel induces apoptosis of cancer cells, its exact mechanism of action is not yet known. The present study has been performed to determine whether influence of paclitaxel in HeLa $S_{3}$ uterine cancer cells. Three assays were employed in this study: cell cytotoxicity, morphological assessments of apoptotic cells (DAPI staining assay), and western blot analysis. The results indicated that paclitaxel has cytotoxic effects in HeLa $S_{3}$ cells. Especially, the $IC_{50}$ value of paclitaxel was about 1 ${\mu}M$. And morphological changes (fragmentation) of cells were observed by paclitaxel in HeLa $S_{3}$ cells. The flow cytometric analysis of paclitaxel-treated cells indicated a block of G2/M phase. The results that pacli-taxel regulates the cell cycle, especially Sub-$G_{1}$ phase. Paclitaxel induces apoptosis of HeLa $S_{3}$ cells via PARP-dependent fashion, and this apoptosis is related to disappearance of Bcl-2 proteins.
Sang-Hun Kim(김상현),Kwang-Youn Kim(김광연),Sun-Nyoung Yu(유선녕),Seul-Ki Park(박슬기),In-Seok Kwak(곽인석),Moon-Soo Rhee(이문수),Byung-Ho Bang(방병호),Sung-Sik Chun(전성식),Soon-Cheol Ahn(안순철) 한국생명과학회 2012 생명과학회지 Vol.22 No.11
Pipernonaline은 후추나무과에 속하는 필발(Piper longum Linn.)의 유도체로서 전립선 암세포에 대한 항암활성이 보고되고 있다. 하지만 실제 암세포 내에서 생물학적 정보를 가진 수 많은 유전자들에 대한 발현이 어떻게 이루어지고 있는지 알려진 바가 없다. 본 연구에 사용된 microarray 분석은 동시에 수 만개 이상의 유전자 발현양상을 한번에 관찰할 수 있는 기술로서 특정 질병의 유전학적 특성과 기전 연구를 더 광범위하게 연구 할 수 있는 기술이다. 본 연구에서는 전립선 암세포인 PC-3 세포에 pipernonaline을 처리하여 cDNA microarray를 실시하였다. 이후, DAVID database를 이용하여 gene ontology의 Biological Process를 분석하여 세포사멸과 세포주기, 세포성장 및 증식에 관련된 유전자들을 우선적으로 분석하였다. 그 결과, 세포주기관련 256개, 세포사멸관련 197개, 세포성장 및 증식관련에 154개의 유전자가 확인 되었다. 이러한 결과는 pipernonaline은 전립선 암세포 내에 존재하는 생물학적 신호전달체계에 관련된 유전자 발현을 조절함으로써 항암활성을 나타내 것을 알 수 있었고, 이후 이러한 microarray의 추가적인 분석은 암세포 내 새로운 유전자의 탐색 및 메커니즘을 규명하는데 유용하게 사용할 수 있을 것으로 사료된다. It has been reported that pipernonaline isolated from Piper longum Linn. has a wide biochemical and pharmacological effect, including antitumor activity in prostate cancer PC-3 cells. However, its mechanism and expression pattern of many genes involved in biological functions are not clearly understood. To perform the gene expression study in PC-3 cells treated with pipernonaline, a cDNA microarray chip composed of 44,000 human cDNA probes was used. As a result, cell cycle-related genes, apoptosis-related genes, and cell proliferation/growth-related genes have been identified in gene ontology of the DAVID database. These results suggest that pipernonaline has antitumor activity by regulating the expression pattern of genes involved in biological signaling pathway in prostate cancer PC-3 cells. Further, additional analysis of these microarray data can be a useful tool to identify the mechanism and discovery of novel genes in cancer therapy.
이은혜,백수연,김광연,이슬기,김상찬,이형식,김영우,Lee, Eun Hye,Baek, Su Youn,Kim, Kwang-Youn,Lee, Seul-Gi,Kim, Sang Chan,Lee, Hyeong Sik,Kim, Young Woo 대한한의학방제학회 2016 大韓韓醫學方劑學會誌 Vol.24 No.3
Objectives : Rheum undulatum Linne and Glycyrriza uralensis Fischer are widely used herbal medicine. In this study, anti-oxidant and liver protective effects of R. undunlatum extract (RUE) and G. uralensis extract (GUE) were investigated in HepG2 cells, respectively. Oxidative stress and liver fibrosis were induced by arachidonic acid (AA) and iron, and CCl<sub>4</sub>.Methods : MTT assay was assessed for cell viability, and immunoblotting analysis was performed to detect expression of apoptosis related proteins. In addition, reactive oxygen species (ROS) and mitochondrial dysfunction were measured. In vivo, BALB/c mouse were orally administrated with the aqueous extract of 10 mg/kg RUE and 100 mg/kg GUE for 3 days and then, injected with CCl<sub>4</sub> 0.5 ml/kg body weight to induce acute liver damage. Serum ALT level was measured, and histological change was observed in Harris's hematoxylin and eosin stainResults : RUE and GUE pre-treatment increased relative cell viability in concentration dependent manner and altered the expression levels of apoptosis-related proteins such as procaspase 3, PARP and Bcl-xL. RUE and GUE also inhibited the mitochondrial dysfunction and excessive reactive oxygen species (ROS) production induced by AA and iron. In addition, RUE and GUE activated liver kinase B1 (LKB1), by increasing phosphorylation. Moreover, RUE and GUE treatment decreased liver injuries induced by CCl<sub>4</sub>, as evidenced by decreases in histological liver damage as well as serum alanine amino transferase (ALT) level.Conclusions : These data suggest that RUE and GUE has anti-oxidant and liver protective effects against AA and iron-induced oxidative stress and CCl<sub>4</sub>-induced liver injury.