• 동의생리병리학회지
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• 제19권1호
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• pp.234-241
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• 2005

Galla Rhois is a nest of parasitic bug, Mellaphis chinensis Bell, in Rhus chinensis Mill. Galla Rhois has been used for the therapy of diarrhea, peptic ulcer, hemauria, etc., that showed various antiinflammatory activity, and other biological properties. We studied the effect of Galla Rhois water extract(GRWE). The cytotoxic activity of GRWE in HL-60 cells was increased in a concentration-dependent manner. GRWE was cytotoxic to HL-60 cells, with $IC_50$ of $100{\mu}g/m{\ell}$. Treatment of GRWE to HL-60 cells showed the fragmentation of DNA in a concentration manner, suggesting that these cells underwent apoptosis. In addition, the flow cytometric analysis revealed GRWE concentration-dependently increased apoptotic cells with hypodiploid DNA content and arrested G1 phase of cell cycle. These results indicate that GRWE may have a possibility of potential anticancer activities. Treatment of HL-60 cells with GRWE was induced activation of caspase-3, caspase-8 and proteolytic cleavage of poly(ADP-ribose) polymerase. Also, caspase-3 was directly activated via caspase-8 activation. GRWE also caused the release of cytochrome c from mitochondria into the cytosol. GRWE-induced cytochrome c release was mediated by caspase-8-dependent cleavage of Bid and Bax translocation. These results suggest that caspase-8 mediates caspase-3 activation and cytochrome c release during GRWE-induced apoptosis in HL-60 cells.

• 한국식품영양과학회지
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• 제37권9호
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• pp.1114-1119
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• 2008

ECCG는 녹차 카테킨의 주요 성분으로 항산화작용으로 인한 항암작용이 보고되고 있다. 본 연구는 EGCG가 전이성이 강한 인체 유방암 세포인 MDA-MB-231의 세포사멸에도 영향을 주는지를 알아보고자 하였다. 인체 유방암 세포 배양액에 EGCG를 0, 5, 10, $20\;{\mu}M$로 첨가시켜, 세포사멸과 관련된 단백질들의 단백질과 mRNA 발현, caspase-3 활성을 관찰하였다. EGCG 첨가 농도가 $5\;{\mu}M$ 이상부터 세포사멸을 억제하는 단백질인 bcl-2의 단백질과 mRNA 발현이 감소하였으며, 세포사멸을 유도하는 단백질인 bax의 단백질과 mRNA 발현은 유의적으로 증가하여 결과적으로 EGCG 첨가에 따라 bcl-2/bax의 비율이 유의적으로 감소하였다. 또한 세포사멸의 마지막 단계인 caspase-3의 활성은 EGCG 농도가 증가할수록 유의적으로 증가하였다. 본 연구 결과를 종합해 보면 전이성이 강한 인체 유방암 세포 MDA-MB-231에서 EGCG는 암세포에서 bcl-2의 발현은 억제시키고 bax의 발현은 증가시키며, caspase-3의 활성을 증가시켜 세포사멸을 유도하는 것으로 확인하였다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권18호
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• pp.7919-7923
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• 2014

20(S)-Protopanaxadiol (PPD), a ginsenoside isolated from Pananx quinquefolium L., has been shown to inhibit growth and proliferation in several cancer cell lines. The aim of this study was to evaluate its anticancer activity in human breast cancer cells. MCF-7 cells were incubated with different concentrations of 20(S)-PPD and cytotoxicity was evaluated by MTT assay. Occurrence of apoptosis was detected by DAPI and Annexin V-FITC/PI double staining. Mitochondrial membrane potential was measured with Rhodamine 123. The Bcl-2 and Bax expression were determined by Western blot analysis. Caspase activity was measured by colorimetric assay. 20(S)-PPD dose-dependently inhibited cell proliferation in MCF-7 cells, with an $IC_{50}$ value of $33.3{\mu}M$ at 24h. MCF-7 cells treated with 20(S)-PPD presented typical apoptosis, as observed by morphological analysis in cell stained with DAPI. The percentages of annexin V-FITC positive cells were 8.92%, 17.8%, 24.5% and 30.5% in MCF-7 cells treated with 0, 15, 30 and $60{\mu}M$ of 20(S)-PPD, respectively. Moreover, 20(S)-PPD could induce mitochondrial membrane potential loss, up-regulate Bax expression and down-regulate Bcl-2 expression. These events paralleled activation of caspase-9, -3 and PARP cleavage. Apoptosis induced by 20(S)-PPD was blocked by z-VAD-fmk, a pan-caspase inhibitor, suggesting induction of caspase-mediated apoptotic cell death. In conclusion, the 20(S)-PPD investigated is able to inhibit cell proliferation and to induce cancer cell death by a caspase-mediated apoptosis pathway.

• Asian Pacific Journal of Cancer Prevention
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• 제17권7호
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• pp.3289-3294
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• 2016

Naringin, a bioflavonoid found in Citrus seeds, inhibits proliferation of cancer cells. The objectives of this study were to investigate the mode and mechanism(s) of hepatocellular carcinoma HepG2 cell death induced by naringin. The cytotoxicity of naringin towards HepG2 cells proved dose-dependent, measured by MTT assay. Naringin-treated HepG2 cells underwent apoptosis also in a concentration related manner, determined by annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI) employing flow cytometry. Mitochondrial transmembrane potential (MTP) measured using 3,3'-dihexyloxacarbocyanine iodide ($DiOC_6$) and flow cytometer was reduced concentration-dependently, which indicated influence on the mitochondrial signaling pathway. Caspase-3, -8 and -9 activities were enhanced as evidenced by colorimetric detection of para-nitroaniline tagged with a substrate for each caspase. Thus, the extrinsic and intrinsic pathways were linked in human naringin-treated HepG2 cell apoptosis. The expression levels of pro-apoptotic Bax and Bak proteins were increased whereas that of the anti-apoptotic Bcl-xL protein was decreased, confirming the involvement of the mitochondrial pathway by immunoblotting. There was an increased expression of truncated Bid (tBid), which indicated caspase-8 proteolysis activity in Bid cleavage as its substrate in the extrinsic pathway. In conclusion, naringin induces human hepatocellular carcinoma HepG2 cell apoptosis via mitochondria-mediated activation of caspase-9 and caspase-8-mediated proteolysis of Bid. Naringin anticancer activity warrants further investigation for application in medical treatment.

• Molecules and Cells
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• 제39권2호
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• pp.119-128
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• 2016

Angelica polymorpha Maxim root extract (APRE) is a popular herbal medicine used for treating stomachache, abdominal pain, stomach ulcers, and rheumatism; however the effect of APRE on cancer cells has not yet been explored. Here, we examined APRE cytotoxicity seen on target neuroblastoma cells (NB) using cell viability assays, DAPI visualization of fragmented DNA, and Western blotting analysis of candidate signaling pathways involved in proliferation and apoptosis. We demonstrated that APRE reduced cell viability in NB to a greater extent than in fibroblast cells. In addition, we found that APRE could inhibit the three classes of MAPK proteins and could also down-regulate the PI3K/AKT/GSK-$3{\beta}$ activity all being relevant for proliferation and survival. APRE could also up-regulate Bax expression and down-regulate Bcl-2 and Mcl-1. With APRE treatment, depolarization of mitochondria membrane potential and activation of caspase-3 was demonstrated in the SH-SY5Y cells. We could not found increased activity of death receptor and caspase-8 as markers of the extrinsic apoptosis pathway for the APRE treated cells. In presence of a caspase-3 siRNA and a pan-caspase inhibitor, APRE could not reduce the viability of NB cells to a significant degree. So we predicted that with APRE, the intrinsic pathway was solely responsible for inducing apoptosis as we also showed that the non-caspase autophagy pathway or ER stress-ROS mediated pathways were not involved. These findings demonstrate that an intrinsic mitochondria-mediated apoptosis pathway mediates the apoptotic effects of APRE on SH-SY5Y cells, and that APRE shows promise as a novel agent for neuroblastoma therapy.

• Animal cells and systems
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• 제13권4호
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• pp.399-403
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• 2009

$p19^{ras}$ is an alternative splicing variant of the proto-oncogene c-H-ras pre-mRNA of $p21^{ras}$. In contrast to $p21^{ras}$, $p19^{ras}$ does not have a C-terminal CAAX motif that targets the plasma membrane and is localized to both the cytoplasm and nucleus. We found that $p19^{ras}$ activated the transcriptional activity of $p73{\beta}$ through protein-protein interactions in the nucleus. p73 is known to play an important role in cellular damage responses such as apoptosis. Although p73 is a structural and functional homologue of p53, p73-mediated apoptosis has not yet been clearly elucidated. In this study, we demonstrate that the interaction between $p19^{ras}$ and $p73{\beta}$ accelerated $p73{\beta}$-induced apoptosis through a caspase-3 dependent pathway. Treatment with DEVD-CHO, a caspase inhibitor, also strengthened $p73{\beta}$-mediated apoptosis through a caspase-3 dependent pathway. Furthermore, the enhanced transcriptional activity of endogenous $p73{\beta}$ by treatment with Taxol was amplified by $p19^{ras}$ overexpression, which markedly increased caspase-3 dependent apoptosis in the p53-null SAOS2 cancer cell line. Our findings indicate a functional linkage between $p19^{ras}$ and p73 in caspase-3 mediated apoptosis of cancer cells.

• 대한한방부인과학회지
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• 제19권4호
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• pp.47-60
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• 2006

Purpose : This study was conducted to investigate the inhibitory effects of Scutellaria barbata D. D on on the cell proliferation of HeLa Cells. Methods : Human uterine cervical carcinoma HeLa cells were cultured in the 1%, 5% and 10% concentration of Scutellaria barbata D. D on solution for 24, 48 and 72 hours for the direct inhibitory effects of Scutellaria barbata D. D on. Then we examined the effect of Scutellaria barbata D. D on solution on the cell proliferation inhibition by XTT assay. DNA fragmentation, MAP kinase activity and caspase activity by FACS analysis in HeLa cells. Results : We found that the proliferation of HeLa cells was significantly decreased in Scutellaria barbata D. D on solution containing groups comparing with a control group in a concentration-dependant manner. When HeLa cells were cultivated for 24 hours with 5% Scutellaria barbata D. D on solution containing group, the percentage of HeLa cells with activated caspase was the highest. Scutellaria barbata D. D on solution reduced the MAP kinase activity of HeLa cells comparing with the control group. By the XTT assay, the cell's activity was decreased in 5% and 10% Scutellaria barbata D. D on solution containing groups in 24 and 72 hours cultivation and 10% group in 48 hours. DNA fragmentation and caspase-3 activity of HeLa cells, however, were changed insignificantly. Conclusion : From this study we could suggest that Scutellaria barbata D. D on is available to the inhibition and apoptosis of human cervical carcinoma cell line, HeLa cells in vitro.

• 대한약학회:학술대회논문집
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• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2-2
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• pp.166.2-166.2
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• 2003

Garlic (Allium sativum) has been used as a general food and a remedy in Oriental for a long time. Since garlic compounds have been also shown to inhibit growth of tumors and to modulate the activity of carcinogenesis, the effects of allicin on growth and survival in human gastric epithelial cells were evaluated by cell viability, cell cycle analysis and DNA fragmentation. Protein levels of cytochrome C, Bcl-xL, Bax and AIF were detected by Western blotting. Effects of recombinant VacA on caspase proteases activity were also determined. (omitted)

• 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.

• 한국약용작물학회지
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• 제17권2호
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• pp.145-150
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• 2009

This study investigated the protective roles and mechanism of magnolol, from the stem bark of Magnolia officinalis against potential neurotoxin 3-hydroxykynurenine (3-HK)-induced neuronal cell death. For the evaluation of protective role of magnolol, we examined cell viability, apoptotic nuclei, change of mitochondrial membrane potential and caspase activity in human neuroblastoma SH-SY5Y cells. It was found that 3-HK induces neuronal cell death in the human neuroblastoma SH-SY5Y cell line. The reduced cell viability produced characteristic features such as cell shrinkages, plasma membrane blebbing, chromatin condensation, and nuclear fragmentation. The cells treated with 3-HK showed an increase in the concentration of reactive oxygen species (ROS) as well as in caspase activity. In addition, both are involved in the 3-HK-induced apoptosis. Magnolol attenuated the cell viability reduction by 3-HK in both a dose- and time-dependent manner. Optical microscopy showed that magnolol inhibited the cell morphological features in the 3-HK-treated cells. Furthermore, the increase in the ROS concentration and the caspase activities by 3-HK were also attenuated by magnolol. These results showed that magnolol has a protective effect on the 3-HK induced cell death by inhibiting ROS production and caspase activity.