• 생명과학회지
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• 제24권9호
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• pp.1012-1018
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• 2014

제주조릿대(Sasa quelpaertensis Nakai)는 한라산에 넓게 분포되어 자생하는 식물로 최근 연구에서 항염증, 항당뇨, 항산화, 항암 효능을 가지는 것으로 알려져 있으나 대장암에서의 항암 효능 및 그에 따른 mechanism에 대해서는 명확히 밝혀지지 않았다. 본 연구에서는 인간 대장암 HT-29 세포를 대상으로 제주조릿대에 의한 항암작용과 기전에 대해 조사하였다. 제주조릿대에 의한 HT-29 세포의 증식 억제가 apoptosis 유도와 연관성이 있음을 DNA fragmentation와 flow cytometry 분석에 의한 sub-G1기의 세포빈도의 증가로 확인하였다. 제주조릿대에 의한 apoptosis 유발은 HT-29 세포의 S arrest 현상을 동반하였을 뿐만 아니라 발생한 산화질소의 증가와 anti-apoptotic factor인 IAP family (survivin, XIAP, cIAP-1, cIAP-2) 발현이 감소함으로써 촉진되었음을 확인할 수 있었다. 이러한 결과들은 제주조릿대가 대장암에 대한 치료제로서의 사용 가능성을 확인할 수 있었지만 이를 입증하기 위해서는 더 자세한 항암기전에 관한 연구가 진행되어야 한다고 사료된다.

• Nutrition Research and Practice
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• 제9권2호
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• pp.111-116
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• 2015

BACKGROUND/OBJECTIVES: Inonotus obliquus (I. obliquus, Chaga mushroom) has long been used as a folk medicine to treat cancer. In the present study, we examined whether or not ethanol extract of I. obliquus (EEIO) inhibits cell cycle progression in HT-29 human colon cancer cells, in addition to its mechanism of action. MATERIALS/METHODS: To examine the effects of Inonotus obliquus on the cell cycle progression and the molecular mechanism in colon cancer cells, HT-29 human colon cancer cells were cultured in the presence of $2.5-10{\mu}g/mL$ of EEIO, and analyzed the cell cycle arrest by flow cytometry and the cell cycle controlling protein expression by Western blotting. RESULTS: Treatment cells with $2.5-10{\mu}g/mL$ of EEIO reduced viable HT-29 cell numbers and DNA synthesis, increased the percentage of cells in $G_1$ phase, decreased protein expression of CDK2, CDK4, and cyclin D1, increased expression of p21, p27, and p53, and inhibited phosphorylation of Rb and E2F1 expression. Among I. obliquus fractions, fraction 2 (fractionated by dichloromethane from EEIO) showed the same effect as EEIO treatment on cell proliferation and cell cycle-related protein levels. CONCLUSIONS: These results demonstrate that fraction 2 is the major fraction that induces $G_1$ arrest and inhibits cell proliferation, suggesting I. obliquus could be used as a natural anti-cancer ingredient in the food and/or pharmaceutical industry.

• Food Science and Biotechnology
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• 제16권2호
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• pp.260-264
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• 2007

Gleditsiae Semen (GS) has been used in both Korea and China as herbal medicine for the treatment of cephalalgia, catharsis, and other diseases. However, the apoptosis of GS against human cancer cells has not previously been investigated. The primary objective of this study was to determine the mechanisms inherent in GS-induced cytotoxicity and apoptosis, using methanolic extract of GS (GSE) in HT-29 human colon carcinoma cells. We found that GSE induced cytotoxicity in HT-29 cells in a dose-dependent manner, and this effect was verified via a lactate dehydrogenase release assay and a colony formation assay. In particular, HT-29 cells showed extensive cell death when treated with $50\;{\mu}g/mL$ of GSE; the calculated $IC_{50}$ value was $20\;{\mu}g/mL$. It induced characteristic apoptotic signs in HT-29 cells, including chromatin condensation and DNA fragmentation, occurring within 6-24 hr when the cells were treated at a concentration of $50\;{\mu}g/mL$. Interestingly, we detected the activation of caspase-3 and -9, but not caspase-8, and apoptotic bodies in GSE-treated HT-29 cells. Collectively, our results indicate that GSE induces apoptosis via a mitochondria-mediated apoptotic pathway, and these findings may be significant with regard to the development of a new drug for the treatment of human colon carcinoma cells.

• Archives of Pharmacal Research
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• 제27권12호
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• pp.1253-1257
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• 2004

6-Methoxydihydrosanguinarine (6ME), a benzophenanthridine alkaloid derived from the methanol extracts of Hylomecon hylomeconoides, showed a dose-dependent effect at 1-10 ${\mu}M$ on causing apoptotic cell death in HT29 colon carcinoma cells $(IC_{50} = 5.0{\pm}0.2 {\mu}M)$. Treatment of HT-29 cells with 6ME resulted in the formation of internucleosomal DNA fragmentation. Treatment of the cells with 6ME caused activation of caspase-3, -8 and 9 protease and subsequent proteolytic cleavage of poly(ADP-ribose)polymerase. 6ME increased the expression of p53 and Bax and decreased the expression of Bid. These results indicate that p53 and proapoptotic Bcl-2 family proteins might participate in the antiproliferative activity of 6ME in HT29 cells.

• BMB Reports
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• 제43권11호
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• pp.750-755
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• 2010

Crude Orostachys japonicus polysaccharide extract (OJP) was prepared by hot steam extraction. Polysaccharides (OJPI) were separated from OJP by gel filtration chromatography and phenol-sulfuric acid assay. The average molecular weight of the OJPI was 30-50 kDa. The anti-proliferative effect of OJPI on HT-29 human colon cancer cells was investigated via morphology study, cell viability assay, apoptosis assay, cell cycle analysis, and cDNA microarray. OJPI inhibited proliferation and growth of HT29 cells and also stimulated apoptosis in a dose- and time-dependent manner. In cell cycle analysis, treatment with OJPI resulted in a marked increase of cells in the G0 (sub G1) and G2/M phases. To screen for genes involved in the induction of cell cycle arrest and apoptosis, the gene expression profiles of HT-29 cells treated with OJPI were examined by cDNA microarray, revealing that a number of genes were up- or down-regulated by OJPI. Whereas several genes involved in anti-apoptosis, cell proliferation and growth, and cell cycle regulation were down-regulated, expression levels of several genes involved in apoptosis, tumor suppression, and other signal transduction events were up-regulated. These results suggest that OJPI inhibits the growth of HT-29 human colon cancer cells by various apoptosis-aiding activities as well as apoptosis itself. Therefore, OJPI deserve further development as an effective agent exhibiting anticancer activity.

• Preventive Nutrition and Food Science
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• 제12권2호
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• pp.84-88
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• 2007

In this study, we investigated antimutagenic and anticancer activities of leaf mustard (LM, Brassica juncea) and leaf mustard kimchi (LMK) during their fermentation period. Methanol extracts were prepared from raw mustard, brined leaf mustard in 10% Gueun salt solution for 2 hrs, leaf mustard fermented at 15$^{\circ}C$ for 5 days after brined in 10% Guenun salt solution for 2 hrs (Fr-LM), fresh leaf mustard kimchi (Fresh-LMK) and optimally ripened leaf mustard kimchi fermented at 5$^{\circ}C$ for 30 days (OR-LMK). OR-LMK showed the strongest inhibitory activities against the mutagenicities induced by aflatoxin B1 in Salmonella Typhimurium TA100. LMs and LMKs inhibited the survival or growth of AGS human gastric adenocarcinoma cells and HT-29 human colon carcinoma cells in MTT assay and growth inhibition test. Among the extracts, OR-LMK and FR-LM exhibited strong antiproliferative effect against cancer cells, especially HT-29 cells. DAPI staining assay showed that OR-LMK induced apoptosis cell death of HT-29 cells in a dose-dependent manner. These results suggest that leaf mustards and leaf mustard kimchi have chemopreventive activities.

• 한국식품영양학회지
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• 제35권5호
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• pp.359-368
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• 2022

In this study, the quality characteristics of kimchi, such as its salinity, pH, and acidity, were measured and compared, and the HT-29 human colon cancer cells were used to show the anticancer effects of kimchi. The kimchi samples used herein included standard kimchi (SK), turnip kimchi (TK), and turnip-powder-added kimchi (TPK). The measured pH and acidity of TK and TPK showed no significant differences with those of SK. Compared to SK and TK, TPK had higher DPPH scavenging activity and higher total flavonoid content, confirming its antioxidant activity. The cancer cell growth inhibition rates of TK and TPK were significantly higher than that of SK. In HT-29 cells treated with TPK, the mRNA expression of Bcl-xL, an anti-apoptosis-related gene, was lower, and the mRNA expressions of the apoptosis-related genes Bax, Bad, and caspase-9 were higher. TPK showed significantly higher levels of mRNA expressions for the cell-cycle-related genes p53 and p21 than the other samples, in addition to suppression effects on cancer cell proliferation. Compared to SK, TK and TPK suppressed the growth of colon cancer cells and showed higher anticancer effects. Therefore, it is shown that kimchi with added turnip powder had high anticancer effects.

• 한국식품영양과학회지
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• 제44권3호
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• pp.338-346
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• 2015

본 연구에서는 최근 식생활의 서구화로 인해 발병률이 급증하고 있는 대장암의 진행을 억제하거나 감소시키고 인체 대장암 세포인 HT-29의 증식을 억제하며, 세포사멸을 유도하는 천연소재를 알아보기 위해서 flavonoid가 HT-29 인체 대장암 세포의 apoptosis 유도 및 기전에 미치는 영향을 알아보았다. MTT assay 결과 apigenin, rutin, naringenin, myricetin을 $100{\mu}M$ 농도로 처리하였을 때 62.71, 75.78, 74.24, 77.61%로 이 중 naringenin이 대장암 세포 성장에 억제 효과가 가장 높은 실험 결과를 나타내었다. Caspase-3 activity에서는 naringenin이 241.46%로 가장 높은 활성을 나타내었다. 이를 바탕으로 세포사멸과 관련된 유전자를 확인하고자 대장암 세포에 flavonoid인 apigenin, rutin, naringenin, myricetin에 $100{\mu}M$ 농도로 처리한 후 RTPCR을 실시한 결과, 세포사멸의 주요한 조절인자인 Bcl-2 family 단백질 중 Bcl-2는 rutin에 의해 감소되었고 Bax는 myricetin에 의해 증가하였으며, p53은 naringenin이 높게 발현되었다. 또한 western blotting을 통해 flavonoid인 apigenin, rutin, naringenin, myricetin에 $100{\mu}M$ 농도로 처리한 결과, Bcl-2 family 단백질과 더불어 세포사멸 조절에 중요한 역할을 하는 활성형인 cleaved caspase-3은 모두 증가하였고, 그중 myricetin이, PARP은 naringenin, E-cadherin은 rutin이 각각 높은 발현 양상을 나타내었다. 이번 실험 결과를 통해 flavonoid가 세포사멸의 주요한 조절 인자인 Bcl-2 family 단백질의 발현이나 caspase의 활성 등을 조절하여 암세포 사멸인자인 Bcl-2의 발현은 감소시키고 Bax, p53, PARP의 발현을 증가시키는 것을 통해 대장암 세포의 apoptosis를 유도하였다. 또한 암세포의 전이와 관련된 E-cadherin의 발현도 조절하는 것을 관찰하였다. 이상의 연구를 통해 flavonoid가 대장암 세포의 증식을 억제하는 효과가 있음을 확인하였으며, 세포사멸과 관련된 기전을 규명하였다. 이를 기초자료로 일상에서 쉽게 섭취할 수 있는 식품에 많이 존재하며 비교적 독성과 부작용이 적은 flavonoid를 이용한 천연 항암제 개발 가능성을 제시하였고, 추후 대장암의 암예방제 및 암치료제로 개발될 수 있도록 추가 연구 수행이 필요할 것으로 사료된다.

• Asian Pacific Journal of Cancer Prevention
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• 제16권6호
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• pp.2569-2574
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• 2015

Necroptosis, also known as "programmed necrosis", has emerged as a critical factor in a variety of pathological and physiological processes and is considered a cell type-specific tightly regulated process with mechanisms that may vary rather greatly due to the change of cell line. Here we used HT-29, a human colon cancer cell line, to establish a necroptosis model and elucidate associated mechanisms. We discovered that cobalt chloride, a reagent that could induce hypoxia-inducible $factor-1{\alpha}(HIF1{\alpha})$ expression and therefore mimic the hypoxic microenvironment of tumor tissue in some aspects induces necroptosis in HT-29 cells when caspase activity is compromised. On the other hand, apoptosis appears to be the predominant death form when caspases are functioning normally. HT-29 cells demonstrated significantly increased RIPK1, RIPK3 and MLKL expression in response to cobalt chloride plus z-VAD treatment, which was accompanied by drastically increased $IL1{\alpha}$ and IL6 expression, substantiating the notion that necrosis can induce profound immune reactions. The RIPK1 kinase inhibitor necrostatin-1 and the ROS scavenger NAC each could prevent necrosis in HT-29 cells and the efficiency was enhanced by combined treatment. Thus by building up a necroptosis model in human colon cancer cells, we uncovered that mechanically RIP kinases collaborate with ROS during necrosis promoted by cobalt chloride plus z-VAD, which leads to inflammation. Necroptosis may present a new target for therapeutic intervention in cancer cells that are resistant to apoptotic cell death.

• 약학회지
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• 제53권4호
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• pp.184-188
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• 2009

Fluorouracil (5-FU) is one of the most widely used chemotherapeutic drugs in the treatment of advanced colorectal cancer patients. Capsaicin (N-vanillyl-8-methyl-alpha-nonenamide), a spicy component of hot pepper, is a homovanillic acid derivative that preferentially induces cancer cells to undergo apoptosis. The purpose of the present study is to examine whether capsaicin enhances the anticancer effect of 5-fluorouracil in HT-29 human colon cancer cells by inducing apoptosis, and whether PPARgamma is involved in the capsaicin action in combination treatment with 5-FU. Treatment of the cells with either 5-FU or capsaicin alone for 48 h had little effect on the cell viability up to $50{\mu}M$ concentration, whereas co-treatment of the cells with capsaicin in the presence of 5-FU for 48 h significantly decreased the cell viability in a concentration-dependent manner. In addition, caspase-3 activity, a marker enzyme for apoptosis, was significantly increased by the combined treatment with 5-FU and capsaicin compared to the 5-FU or capsaicin alone treatment. Also, treatment with troglitazone, a peroxisome proliferator-activated receptor gamma ($PPAR{\gamma}$) agonist, further enhanced the effect of the combination treatment on the cell viability and caspase-3 activity, and bisphenol A diglycidyl ether (BADGE), a $PPAR{\gamma}$ antagonist, blocked the effect of the combination treatment. These results suggest that the combination treatment of HT-29 cells with 5-FU and capsaicin induces apoptotic cell death at relatively low concentration than each drug alone, and the combination treatment may be associated with the $PPAR{\gamma}$ pathway activation.