• 동의생리병리학회지
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• 제17권6호
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• pp.1383-1392
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• 2003

Apoptosis is a morphologically and biochemically district form of cell death that occurs in many different cell types in a wide variety of organisms. Albizzia julibrissin belonging the family Leguminosae has been used for the treatment of contusion, sore throat, amnesia, and insomnia in oriental traditional medicine. This study investigates whether the water extract of A. julibrissin induce apoptotic cell death in Jurkat T-acute lymphoblastic leukemia (ALL) cells. Jurkat cells were increased inhibitions of cell viability in a concentration-dependent manner by A. julibrissin. This herbal medicine also caused apoptosis as measured by cell morphology and DNA fragmentation. The capability of A. julibrissin to induce apoptosis was associated with proteolytic cleavage of specific target proteins such as poly (ADP-ribose)polymerase (PARP) and beta-catenin proteins suggesting the possible involvement of caspases. Our result showed that Bcl-2 and Bax protein levels were not changed in all A. julibrissin-treated groups compared to control group. These results suggest that A. julibrissin-mediated apoptosis is independent with Bcl-2 related signaling pathway in this cells. The purpose of the present study is also to investigate the Effect of A. julibrissin on cell cycle progression. Our results showed that G1 checkpoint related gene products (cyclin D1, cyclin dependent kinase 4, retinoblastoma, E2F1) were decreased in their protein levels in a dose-dependent manners after treatment of the extract. These results indicate that the increase of apoptotic cell death by A. julibrissin may be due to the inhibition of cell cycle progression in wild type p53-lacking Jurkat cells.

• Journal of Microbiology and Biotechnology
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• 제14권5호
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• pp.978-984
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• 2004

In the course of screening for a novel inhibitor of CDC2, HY558-1 was isolated from a culture broth of Penicillium minioluteum F558. Moreover, it was found that HY558-1 had an effect on both the cell cycle regulation and apoptosis of human cervical adenocarcinoma HeLa cells. A flow cytometric analysis of HeLa cells revealed appreciable cell cycle arrest at the G1 and G2/M phases following treatment with HY558-1. Furthermore, DNA fragmentation due to apoptosis was observed in HeLa cells treated with HY558-1. To obtain further information on the cell cycle arrest and apoptotic induction induced by HY558-1, the expression of certain cell cycle and apoptosis-associated proteins was examined using a Western blot analysis. The results revealed that HY558-1 inhibited the phosphorylation of pRb and decreased the expression levels of CDK2, CDC2, and cyclin A in the cell cycle progression. It was also shown that the level of $p21^{WAF1/CIP1}$ was increased in HeLa cells treated with 0.52 mM of HY558-1. Accordingly, HY558-1 was found to inhibit the proliferation of HeLa cells through the induction of G1 phase arrest by inhibiting pRb phosphorylation via an upregulation of $p21^{WAF1/CIP1}$, and G2/M phase arrest by directly inhibiting CDC2 and cyclin A. Moreover, HeLa cells treated with 0.52 mM of HY558-1 exhibited apoptotic induction associated with the cleavage of Bid and release of cytochrome c from mitochondria into the cytosol. Subsequent investigation of the activation of caspase-3 and cleavage of poly (ADP-ribose) polymerase (PARP) suggested that the mitochondrial pathway was primarily involved in the HY558-1-induced apoptosis in HeLa cells.

• Applied Biological Chemistry
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• 제51권2호
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• pp.129-135
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• 2008

Berberine은 전통적인 중의약재로 이용되어지는 isoquinoline alkaloid로 황련, 황백과 같은 식물에서 주로 추출되며, 약리효과로는 항암, 항염, 항균과 같은 다양한 효과를 나타내는 것으로 알려져 있다. 그러나 간암세포에서 berberine의 산화적 스트레스에 의한 세포사멸기전에 대해서는 아직 밝혀진 바 없다. 따라서 본 연구는 사람의 간암세포인 HepG2 세포에서 berberien의 세포사멸기전에 reactive oxygen species(ROS)와 MAP kinase의 연관성을 조사하였다. Berberine은 HepG2 세포에서 처리 시간과 농도에 의존적으로 세포독성효과를 보였으며, $LD_{50}$은 berberine(50 ${\mu}M$) 처리 후 48시간에서 관찰 되었고, 세포고사의 특징인 핵의 응축 및 분절, DNA의 분절이 확인되었다. 또한 berberine에 의해 caspase-3, p53, p38 그리고 Bax의 발현이 현저하게 증가된 반면, anti-apoptotic 신호기전인 Bc1-2의 발현은 감소되었다. 이와 더불어 세포 내 nitric oxide(NO)와 ROS의 생성도 증가되었다. 본 연구 결과 HepG2 세포에서 berberine은 산화적 스트레스인 ROS와 NO의 생성을 유발하고 p38 MAP kinase와 p53의 인산화를 유도하였으며 미토콘드리아에서 Bcl-2의 감소와 bax의 증가, caspase-3의 활성을 경유하여 DNA의 손상을 통한 세포고사가 이루어지는 것을 확인 하였다.

• BMB Reports
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• 제53권11호
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• pp.582-587
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• 2020

It is well known that oxidative stress participates in neuronal cell death caused production of reactive oxygen species (ROS). The increased ROS is a major contributor to the development of ischemic injury. Indoleamine 2,3-dioxygenase 1 (IDO-1) is involved in the kynurenine pathway in tryptophan metabolism and plays a role as an anti-oxidant. However, whether IDO-1 would inhibit hippocampal cell death is poorly known. Therefore, we explored the effects of cell permeable Tat-IDO-1 protein against oxidative stress-induced HT-22 cells and in a cerebral ischemia/reperfusion injury model. Transduced Tat-IDO-1 reduced cell death, ROS production, and DNA fragmentation and inhibited mitogen-activated protein kinases (MAPKs) activation in H₂O₂ exposed HT-22 cells. In the cerebral ischemia/reperfusion injury model, Tat-IDO-1 transduced into the brain and passing by means of the blood-brain barrier (BBB) significantly prevented hippocampal neuronal cell death. These results suggest that Tat-IDO-1 may present an alternative strategy to improve from the ischemic injury.

• Molecules and Cells
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• 제24권1호
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• pp.95-104
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• 2007

The mechanism of acacetin-induced apoptosis of human breast cancer MCF-7 cells was investigated. Acacetin caused 50% growth inhibition ($IC_{50}$) of MCF-7 cells at $26.4{\pm}0.7{\mu}M$ over 24 h in the MTT assay. Apoptosis was characterized by DNA fragmentation and an increase of sub-G1 cells and involved activation of caspase-7 and PARP (poly-ADP-ribose polymerase). Maximum caspase 7 activity was observed with $100{\mu}M$ acacetin for 24 h. Caspase 8 and 9 activation cascades mediated the activation of caspase 7. Acacetin caused a reduction of Bcl-2 expression leading to an increase of the Bax:Bcl-2 ratio. It also caused a loss of mitochondrial membrane potential that induced release of cytochrome c and apoptosis inducing factor (AIF) into the cytoplasm, enhancing ROS generation and subsequently resulting in apoptosis. Pretreatment of cells with N-acetylcysteine (NAC) reduced ROS generation and cell growth inhibition, and pretreatment with NAC or a caspase 8 inhibitor (Z-IETD-FMK) inhibited the acacetin-induced loss of mitochondrial membrane potential and release of cytochrome c and AIF. Stress-activated protein kinase/c-Jun $NH_4$-terminal kinase 1/2 (SAPK/JNK1/2) and c-Jun were activated by acacetin but extracellular-regulated kinase 1/2 (Erk1/2) nor p38 mitogen-activated protein kinase (MAPK) were not. Our results show that acacetin-induced apoptosis of MCF-7 cells is mediated by caspase activation cascades, ROS generation, mitochondria-mediated cell death signaling and the SAPK/JNK1/2-c-Jun signaling pathway, activated by acacetin-induced ROS generation.

• Clinical and Experimental Reproductive Medicine
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• 제31권2호
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• pp.83-94
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• 2004

Objective: To investigate whether GnRH-agonist (GnRH-Ag) using in IVF-ET affects apoptosis of human granulosa-luteal cells and expression of peripheral benzodiazepine receptor (PBR) protein involved in the apoptosis of the cells. Methods: Granulosa-luteal cells obtained during oocyte retrieval were cultured and treated with $10^{-5}M$ GnRH-Ag. Apoptosis of the cells by the treatment was confirmed using DNA fragmentation analysis 24 h after culture. The presence of PBR protein within the cells was examined by immunofluorescence staining and the expression of the protein was analyzed by Western blotting. In addition, it was measured for progesterone and nitric oxide (NO) produced by granulosa-luteal cells after GnRH-Ag treatment. To evaluate the relationship between NO production and PBR expression, sodium nitroprusside (SNP) as a NO donor was added in media and investigated the expression of PBR protein by Western blotting. Results: Apoptosis increased in the granulosa-luteal cells 24 h after GnRH-Ag treatment, whereas the expression of PBR protein significantly decreased. Furthermore, the production of progesterone and nitric oxide (NO) by the cells significantly fell from 12 h after the treatment. In the results of Western blotting after SNP treatment, the expression of PBR protein increased in the treatment with SNP alone to the granulosa-luteal cells, but was suppressed in the treatment with GnRH-Ag and SNP. Additionally, the staining result of PBR protein in the cells showed the even distribution of it through the cell. Conclusion: These results demonstrate that GnRH-Ag treatment induces apoptosis, decreasing expression of PBR protein and NO production in human granulosa-luteal cells. The present study suggests that one of the apoptosis mechanism of human granulosa-luteal cells by GnRH-Ag might be a signal transduction pathway via NO and PBR.

• 한국식품영양과학회지
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• 제39권8호
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• pp.1119-1125
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• 2010

본 연구에서는 resveratrol을 전립선 암 치료제로의 활용 가능성을 조사하기 위하여 primary 인체 전립선 암세포에 대한 resveratrol의 성장억제 효과 및 그 기전에 대하여 조사 하였다. Resveratrol은 RC-58T/h/SA#4 세포에서 농도 및 시간에 의존적으로 세포의 증식을 억제하였으며, $IC_{50}$ 값은 암세포인 RC-58T/h/SA#4, LNCaP, PC-3에서는 각각 245, 320, $340\;{\mu}M$, 전립선 정상세포인 RWPE-1에서는 $982\;{\mu}M$로 나타나 정상세포에서보다는 암세포에서 그 독성이 크게 나타났다. 또한 resveratrol에 의해 유도된 세포 사멸은 핵 응축, sub-G1 함량 증가 및 DNA 분절 현상이 나타나 apoptosis를 유도함을 알 수 있었다. Resveatrol은 caspase-8, -9 및 effector casapse-3 활성을 농도 의존적으로 증가시켰으며, caspase 저해제인 z-VAD-fmk로 caspase의 처리 시 resveratrol에 의한 apoptosis 유도 현상이 유의적으로 감소되어 resveratrol에 의한 RC-58T/h/SA#4 세포의 apoptosis 유도에 caspase가 중요한 역할을 하고 있음을 확인하였다. Resveratrol에 의해 anti-apoptotic 인자인 Bcl-2 및 Bid 단백질의 발현은 감소하였으나, pro-apoptotic 인자인 Bax 단백질 발현은 변화가 없었다. 따라서 본 연구는 resveratrol이 RC-58T/h/SA#4세포에서 caspase 의존형 미토콘드리아 경로에 의해 유도되며, resveratrol은 전립선암 치료제로서 사용 가능성을 시사한다.

• 대한한의학방제학회지
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• 제25권1호
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• pp.51-68
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• 2017

Objectives : Oxidative stress due to excessive accumulation of reactive oxygen species (ROS) is one of the risk factors for the development of several chronic diseases, including neurodegenerative diseases. Methods : In the present study, we investigated the protective effects of cheongnoemyeongsin-hwan (CNMSH) against oxidative stress‑induced cellular damage and elucidated the underlying mechanisms in neuronal-derived SH-SY5Y cells. Results : Our results revealed that treatment with CNMSH prior to hydrogen peroxide (H2O2) exposure significantly increased the SH-SY5Y cell viability, indicating that the exposure of the SH-SY5Y cells to CNMSH conferred a protective effect against oxidative stress. CNMSH also effectively attenuated H2O2‑induced comet tail formation, and decreased the phosphorylation levels of the histone ${\gamma}H2AX$, as well as the number of apoptotic bodies and Annexin V‑positive cells. In addition, CNMSH exhibited scavenging activity against intracellular ROS generation and restored the mitochondria membrane potential (MMP) loss that were induced by H2O2, suggesting that CNMSH prevents H2O2‑induced DNA damage and cell apoptosis. Moreover, H2O2 enhanced the cleavage of caspase-3 and degradation of poly (ADP-ribose)-polymerase, a typical substrate protein of activated caspase-3, as well as DNA fragmentation; however, these events were almost totally reversed by pretreatment with CNMSH. Furthermore, CNMSH increased the levels of heme oxygenase-1 (HO-1), which is a potent antioxidant enzyme, associated with the induction of nuclear factor-erythroid 2-related factor 2 (Nrf2). According to our data, CNMSH is able to protect SH-SY5Y cells from H2O2-induced apoptosis throughout blocking cellular damage related to oxidative stress through a mechanism that would affect ROS elimination and activating Nrf2/HO-1 signaling pathway. Conclusions : Therefore, we believed that CNMSH may potentially serve as an agent for the treatment and prevention of neurodegenerative diseases caused by oxidative stress.

• 생명과학회지
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• 제21권6호
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• pp.851-857
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• 2011

Sulforaphane (SFN)은 브로콜리와 다른 십자화과 채소로부터 분리해 낸 이소시아네이트 구조를 갖는 물질이다. 최근 연구에서는 다양한 암세포에서 SFN이 세포주기를 억제하여 증식저해와 세포사멸을 유도한다고 알려져 있다. 본 연구에서는 연골암 세포주 HTB 94를 이용하여 연골암세포에서의 SFN의 세포사멸 분자적 기전을 연구하였다. SFN은 처리농도 의존적으로 연골암세포의 증식억제와 세포사멸을 유도하는 것을 세포형태변화 및 핵염색, MTT 실험을 통하여 확인하였다. Apoptotic 세포는 DNA 단편전기영동 분석을 통한 분절 DNA 확인 및 유세포분석기를 이용한 subG1 구간의 세포분석을 이용하여 확인하였다. 연골암세포주 HTB-94에 SFN 처리 시 p53 단백질의 발현증가 및 pro-caspase-3의 분해를 통한 caspase-3의 활성을 증가를 Western blot analysis를 통하여 확인하였다. 본 연구결과를 종합해 볼 때, 연골암세포주 HTB-94에 SFN을 처리시 유도되어지는 세포사멸은 p53과 caspase-3 의존적인 신호조절경로를 통하여 조절되는 것을 확인하였다. 본 연구결과는 SFN의 증식억제 효과 및 연골암세포 세포사멸 유도기전을 이용하여 향후 연골암세포의 치료약제 개발에 도움이 될 것으로 기대한다.

• 대한한방내과학회지
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• 제28권3호
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• pp.473-491
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• 2007

Objectives : This study was designed to investigate the antiproliferative activity of the water extract of Samgibopae-tang (SGBPT) in NCI-H460 and A549 non-small-cell lung cancer cell lines Methods : In this study, we measured the subsistence, form of NCI-H460 and A549 non-small-cell lung cancer cell by hemocytometer and DAPI staining. In each cell, we analyzed DNA fragmentation. reverse transcription-polymerase chain reaction and measured activity of caspase-3, caspase-8 and caspase-9. Results and Conclusions : We found that exposure of A549 cells to SGBPT resulted in growth inhibition in a dose-dependent manner. butSGBPT did not affect the growth of NCI-H460 cells. The antiproliferative effect by SGBPT treatment in A549 cells was associated with morphological changes. SGBPT treatment partially induced the expression of DR5 cells and the expression of Faswas markedly increased in both transcriptional and translational levels in A549 cells. SGBPT treatment partially induced the expression of Bcl-2, Bcl-XL and the expression of Bid was markedly decreased in translational levels in A549 cells. However, SGBPT treatment did not affect the expression of IAP family in A549 orNCI-H460 cells. SGBPT treatment partially induced the expression of caspase-3, caspase-8, caspase-9 activity which markedly increased in a dose-dependent manners in A549 cells. The fragmental development of PARP and ${\beta}$-catenin protein was observed in A549 cells by SGBPT treatment. SGBPT treatment induced the expression of PLC-${\gamma}1$ protein which decreased in A549 cells. SGBPT treatment partially induced the expression of DFF45/ICAD which markedly increased in a dose-dependent manner in A549 cells. Taken together. these findings suggested that SGBPT-induced inhibition of human lung carcinoma did not affect NCI-H460 cell growth. However, SGBPT-induced inhibition of human lung carcinoma A549 cell growth was associated with the induction of death receptor and mitochondrial pathway. The results provided important new insights into the possible molecular mechanisms of the anti-cancer activity of SGBPT.