• Journal of Photoscience
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• 제9권2호
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• pp.509-511
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• 2002

In this study, we investigated apoptotic cell death induced by photodynamic therapy using 5-aminolaevulinic acid (ALA-PDT) in human promyelocytic leukemia cells (HL-60). ALA-PDT induced apoptosis in HL-60 cells as confirmed by DNA agarose gel electrophoresis and nuclear staining with Hoechst 33342. The apoptotic cell death was inhibited by addition of broad-spectrum caspase inhibitor Z-Asp-CH$_2$-DCB, indicating that the apoptotic cell death was induced in a caspase-dependent manner. Actually, western blotting analysis revealed that caspase-3 was processed as early as 1.5 h after ALA-PDT. Cytoplasmic cytochrome c released from mitochondria was detected by western blotting. However, inhibitor of caspase-9, a cysteine protease located in the downstream of cytochrome c release, was not able to reduce the apoptotic cell death. Therefore, the mitochondrial apoptotic pathway was not involved in the ALA-PDT-induced apoptosis. On the other hand, it was found that ALA-PDT-induced apoptosis was clearly inhibited by pretreatment of caspase-8 inhibitor. These data suggest that caspase-8-mediated apoptotic pathway is important in ALA-PDT-induced cell death.

• Molecules and Cells
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• 제38권4호
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• pp.349-355
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• 2015

EphA7 has been implicated in the regulation of apoptotic cell death in neural epithelial cells. In this report, we provide evidence that EphA7 interacts with caspase-8 to induce apoptotic cell signaling. First, a pull-down assay using biotinylated ephrinA5-Fc showed that EphA7 co-precipitated with wild type caspase-8 or catalytically inactive caspase-8 mutant. Second, co-transfection of EphA7 with caspase-8 significantly increased the number of cleaved caspase-3 positive apoptotic cells under an experimental condition where transfection of EphA7 or caspase-8 alone did not affect cell viability or apoptosis. EphA4 also had a causative role in inducing apoptotic cell death with caspase-8, whereas EphA8 did not. Third, caspase-8 catalytic activity was essential for the apoptotic signaling cascade, whereas tyrosine kinase activity of the EphA4 receptor was not. Interestingly, we found that kinase-inactive EphA4 was well co-localized at the plasma membrane with catalytically inactive caspase-8, suggesting that an interaction between these mutant proteins was more stable. Finally, we observed that the extracellular region of the EphA7 receptor was critical for interacting with caspase-8, whereas the cytoplasmic region of EphA7 was not. Therefore, we propose that Eph receptors physically associate with a transmembrane protein to form an apoptotic signaling complex and that this unidentified receptor-like protein acts as a biochemical linker between the Eph receptor and caspase-8.

• Asian Pacific Journal of Cancer Prevention
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• 제15권15호
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• pp.6075-6080
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• 2014

Lung cancer is the leading cause of cancer-related death worldwide. Here we investigated the antitumor effect and mechanism of Zhejiang (Huzhou and Jiande) saffron against lung cancer cell lines, A549 and H446. Using high performance liquid chromatography (HPLC), the contents of crocin I and II were determined. In vitro, MTT assay and annexin-V FITC/PI staining showed cell proliferation activity and apoptosis to be changed in a dose- and time-dependent manner. The inhibition effect of Jiande saffron was the strongest. In vivo, when mice were orally administered saffron extracts at dose of 100mg/kg/d for 28 days, xenograft tumor size was reduced, and ELISA and Western blotting analysis of caspase-3, -8 and -9 exhibited stronger expression and activity than in the control. In summary, saffron from Zhejiang has significant antitumor effects in vitro and in vivo through caspase-8-caspase-9-caspase-3 mediated cell apoptosis. It thus appears to have more potential as a therapeutic agent.

• 생물정신의학
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• 제5권2호
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• pp.219-226
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• 1998

Cytosine arabinoside(AraC) inhibits DNA synthesis and ${\beta}$-DNA polymerase, an enzyme involved in DNA repair. This, a potent antimitotic agent, is clinically used as an anticancer drug with side effect of severe neurotoxicity. Earlier reports suggested that inhibition of neuronal survival by AraC in sympathetic neuron may be due to the inhibition of a 2'-deoxycytidine-dependent process that is independent of DNA synthesis or repair and AraC induced a signal that is triggers a cascade of new mRNA and protein synthesis, leading to apoptotic cell death in cultured cerebellar granule cells. The present study would suggest whether caspase family(ICE/CED-3-like protease) involved in AraC-induced apoptosis pathway of PC12 cells. It was observed that treatment of PC12 cells with AraC led to decrease of viability by MTT assay and morphology changes, which did not suggest that AraC induced apoptosis in PC12 cells. The mRNA of caspase-1/caspase-3 were expressed in PC12 cells constitutively, and AraC did not activate caspase family. These results suggest that caspase-1/caspase-3 may not be required for AraC-induced cell death pathway in PC12 cells.

• Toxicological Research
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• 제17권3호
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• pp.215-223
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• 2001

카드뮴의 주요한 표적장기이며 카드뮴이 만성 및 급성 폭로시 축적되는 가장 중요한 장기인 간의 세포독성을 Hepalclc7세포에서 caspases및 Bax단백질의 활성과 발현 그리고 미토콘드리아 세포막 전위 변화(MPT) 등을 조사하여 다음과 같은 결과를 얻었다. 1. 카드뮴은 농도의존적으로 간세포인 Hepalclc7 세포의 생존율을 감소시켰다. 2. 카드뮴을 농도별로 처리하였을 때 100 M 이상의 농도에서 세포사멸의 특징중의 하나인 DNA분절현상을 확인하였다. 3. 카드뮴 처리 후 caspase-3, caspase-8, caspase-9 의 활성변화를 조사한 결과 caspase-3,-9 pretease 활성이 시간이 경과함에 따라 증가하였다. 4. 카드뮴 처리 후 cytochrome c가 세포질내로 방출되었고 이는 caspase-9 proteas의 활성화를 유도하였다. 5. 카드뮴 처리 후 Bax가 세포질에서 미토콘드리아로 이동하여 cytochrome c의 세포질내로의 방출에 관여하였다. 6. 카드뮴 처리시 미토콘드리아 세포막 전위차의 감소를 JC-1 형광염색을 통하여 확인하였다. 이상의 결과는 카드뮴에 의한 Hepalclc7 세포사멸의 신호전달기전은 세포질내에 있는 Bax가 미토콘드리아로 이동, cytochrome c의 세포질내로의 방출, 그리고 caspase-3, 9 pretease 활성화를 의해서 매개되는 것으로 판단된다. 또한 Bax 단백질의 발현변화가 미토콘드리아의 기능변화에 기여하였으리라 사료된다.

• 생명과학회지
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• 제34권2호
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• pp.94-104
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• 2024

β-lapachone은 다양한 유형의 질병을 치료하기 위해 남미 및 중미 지역의 전통 의학에서 널리 사용되어 온 Tabebuia vellanedae의 껍질에서 분리된 천연 퀴논 화합물의 일종이다. β-lapachone은 여러 유형의 암세포에서 강력한 항암 활성을 갖는 것으로 보고되었지만, 간세포암종 세포의 증식에 대한 효과는 아직 불분명하다. 따라서 본 연구에서는 β-lapachone 인간 간세포암종 Hep3B 세포의 증식에 미치는 영향을 조사하였으며, 본 연구의 결과에 의하면, β-lapachone 처리에 의한 Hep3B 세포의 세포생존율 감소는 세포사멸 유도와 밀접한 관련이 있었다. 또한, β-lapachone이 처리된 Hep3B 세포에서는 항세포사멸 인자인 Bcl-2의 발현이 감소한 반면, 세포사멸 유도 인자인 Bax의 발현은 증가하였으며, 이는 caspase cascade의 활성 증가와 연관성이 있었다. 그러나 pan-caspase 억제제가 존재하는 경우 β-lapachone에 의해 유발된 세포사멸은 약화되었으며, 이는 β-lapachone에 의한 세포사멸 유도가 caspase 의존적인 현상임을 의미한다. 아울러, β-lapachone의 처리는 ERK 경로를 활성화시키면서 PI3K/Akt 경로의 활성을 억제하였으며, β-lapachone 유도 세포사멸에 ERK 억제제의 효과는 미미했지만, PI3K 억제제는 β-lapachone에 의해 유도된 세포사멸을 유의하게 증가시켰다. 비록 생체 내 동물 모델에서의 확인이 필요하지만, 본 연구의 결과는 간세포암종 세포에서 β-lapa-chone의 항암 활성을 이해하는 데 유용한 자료로 활용될 것이다.

• 대한한의학회지
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• 제23권3호
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• pp.145-163
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• 2002

Objectives: The purpose of this investigation was to evaluate the effects of Woohwangcheongsim-won and to study the mechanism for neuronal death protection in hypoxia with Embryonic day 20 (E20) cortical cells of a rat (Sprague Dawley). Methods: E20 cortical cells were dissociated in neurobasal media and grown for 14 days in vitro (DIV). On 14 DIV, Woohwangcheongsim-won was added to the culture media for 24 hrs or 72 hrs. On 17 DIV, cells were given a hypoxic shock and further incubated in normoxia for another three days. On 20 DIV, Woohwangcheongsim-won's effects for neuronal death protection were evaluated by LDH assay, propidium iodide stain and phospho-H2AX immunostain and the mechanisms were studied by Bcl-2, Bak, Bax, caspase family, PKCα, ca1pain I. Results & Conclusions : 1. This study indicated that Woohwangcheongsim-won's effects for neuronal death protection in hypoxia were confirmed by LDH assay, propidium iodide stain and phospho-H2AX immunostain in culture method of Embryonic day 20(E20) cortical neuroblasts. 2. Woohwangcheongsim-won's mechanisms for neuronal death protection in hypoxia are to reduce the membrane damage fraction, to restrain DNA truncate, to restrain inflow of cytochrome c into cellularity caused by Bak diminution, to reduce the caspase cascade intiator caspase-8 and the effector caspase-3, to reduce the calpain I activity and to increase PKCand its activity in the membrane fraction. (J Korean Oriental Moo 2002;23(3):145～163)

• 한국수산과학회지
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• 제44권3호
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• pp.216-224
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• 2011

Capsosiphon fulvescens is well-known green sea algae that, in recent years, has been proposed as a potential anticancer drug. In this study, we found that C. fulvescens glycoprotein (Cf-GP) had pro-apoptotic effects on human gastric carcinoma cells. By SDS-PAGE, we confirmed that C. fulvescens extract contained a glycoprotein. Using H33342 staining, we found that the Cf-GP caused cell death in a does-dependent manner, while an MTS assay showed decreased cellular viability due to induction of apoptosis. To determine the effect of Cf-GP on apoptosis-related cellular events, cells were treated with Cf-GP and the expression of several apoptosis-related protein was determined by Western blotting. Our results indicate that Cf-GP activated both a caspase cascade and PARP, which is a substrate of caspase-3, caspase-8 and the Bcl-2 family proteins. In addition, we assessed caspase-3, and -8 activation and annexin V staining. Our results revealed a cell cycle arrest, itself leading to an increased percentage of sub-G1 cells. Our findings indicate that Cf-GP may be a source of bio-functional material with therapeutic effects on human gastrointestinal cancer.

• 생약학회지
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• 제48권4호
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• pp.273-279
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• 2017

The aim of this study was to investigate the mechanisms underlying apoptosis induced by a broussochalcone B (BCB) from Broussonetia papyrifera in HepG2 cells. The results showed that BCB treatment for 24 hr significantly inhibited cell viability in a dose-dependent manner, and induced apoptosis in HepG2 cells. More so, BCB treatment triggered the cleavage of caspase-8, -9, -3, poly (ADP-ribose) polymerase (PARP), increase of Bax level, and decrease of Bcl-2 expression. A general caspase inhibitor (z-VAD-fmk) blocked BCB-induced cell death. Furthermore, BCB treatment caused reactive oxygen species (ROS) production in a dose-dependent manner. In addition, an antioxidant N-acetylcysteine (NAC) blocked BCB-induced ROS production and cell death. Therefore, these results indicate that BCB-induced apoptosis is mediated by a caspase dependent pathway and ROS production in HepG2 cells.

• Journal of Microbiology and Biotechnology
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• 제19권3호
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• pp.286-290
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• 2009

In the course of screening for substances inhibiting apoptosis of U937 human leukemia cells induced by etoposide ($10\;{\mu}g/ml$), Forsythiae fructus, which showed a high level of inhibition, was selected. The regulating compounds were purified from the ethyl acetate extract by silica gel column chromatography and HPLC. The active substance was purified and identified as rengyolone by spectroscopic methods. This compound showed inhibitory activity on caspase-3 induction, a major protease of the apoptosis cascade, with an $IC_{50}$ value of $38.96\;{\mu}M$ after 8 h of etoposide treatment in U937 cells. The expression level of caspase-3 and poly(ADP-ribose) polymerase (PARP) were dose-dependently inhibited by the compound, suggesting that rengyolone inhibits etoposide-induced apoptosis via downregulation of caspases.