• 대한한방부인과학회지
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• 제18권3호
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• pp.77-91
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• 2005

Purpose : Trichosanthis Radix is traditional medical herb which has been shown to inhibit tumor cell proliferation. In this study, the effects of Trichosanthis Radix extract were investigated on inducing growth inhibition and apoptosis of human uterine cervical carcinoma cells. Methods : Human uterine cervical carcinoma cells line, ME-180, was used for the study. The cells were treated with varying concentrations of Trichosanthis Radix extract. Cell growth and inhibitory rate were measured by MTT assay. Apoptosis induction was detected by fluorescence microscopy, DNA ladder formation and flow cytometry. Results : Trichosanthis Radix extract inhibited the growth of human uterine cervical carcinoma cells in a dose-dependent manner. It induced ME-180 cells to undergo apoptosis including fragmented nuclei and nucleosome-sized DNA fragmentation. Flow cytometric analysis showed the increasing rate of apoptotic cells by Trichosanthis Radix extract. Reduction of mitochondrial membrane potential and increase in caspase-3 activity and were found in ME-180 cells treated with Trichosanthis Radix extract. Conclusion : Our data suggest that Trichosanthis Radix extract inhibit the growth and proliferation of ME-180 cells by apoptotic induction and facilitates its activity via caspase-3 activation initiated by depolarization of mitochondria.

• 생명과학회지
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• 제25권12호
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• pp.1384-1392
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• 2015

적작약, 사간, 치자, 적복령, 승마 및 백출 등 6가지의 한약재로 구성된 사간탕은 동의보감에서 위완옹(胃脘癰)을 치료하는 처방으로 알려져 있으나, 항암 효능에 대한 구체적인 연구는 전혀 이루어진 바 없다. 본 연구에서는 사간탕의 항암활성 연구의 일환으로 AGS 인체 위암세포의 증식에 미치는 영향을 조사하였다. 본 연구의 결과에 의하면 사간탕 추출물 처리 농도의 증가에 따라 AGS 위암세포의 증식 및 생존율이 억제되었으며, 이는 apoptosis 유발에 의한 것임을 염색질 응축, DNA 단편화 및 annexin-V 염색 등을 통하여 확인하였다. 사간탕 추출물 처리에 의한 apoptosis 유발에는 pro-apoptotic Fas 단백질의 발현 증가 및 anti-apoptotic Bcl-2 발현의 감소와 mitochondrial membrane potential의 소실이 동반되었다. 아울러 사간탕 추출물이 처리된 AGS 위암세포에서 extrinsic 및 intrinsic apoptosis 경로 활성의 개시에 중요한 caspase-8 및 -9 뿐만 아니라 effector caspase인 caspase-3의 활성도 증가하였으며, 활성화된 caspase-3의 기질 단백질인 PARP의 단편화도 관찰되었다. 그러나 pan-caspase inhibitor의 선처리에 의한 caspase 활성을 차단하였을 경우, 사간탕 추출물 처리에 의한 염색질 응축 및 DNA 단편화 현상이 관찰되지 않았으며, apoptosis 유발 및 증식억제 효과도 유의적으로 억제되었다. 따라서 사간탕 추출물 처리에 의한 AGS 위암세포의 apoptosis 유발은 extrinsic 및 intrinsic apoptosis 경로가 동시 활성을 통한 caspase 의존적인 과정을 통하여 이루어지고 있음을 알 수 있었으며, 그 과정은 아마도 pro-apoptotic Bid의 truncation이 관여할 것으로 추정된다. 이상의 결과는 향후 in vivo 모델을 이용한 사간탕 추출물의 항암활성 조사 및 사간탕 추출물 내 주요 생리활성 물질의 탐색 등을 위한 유용한 자료로 사용될 것이다.

• 동의생리병리학회지
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• 제20권2호
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• pp.358-364
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• 2006

To investigate the possible mechanism of Drynariae Rhizoma extracts as a candidate of anti-cancer drug, I examined the effects of Drynariae Rhizoma extracts on the apoptosis of U937 cell line. MTT assay, flow cytometric analysis, SDS-polyacrylamide gel electrophoresis, Western blot analysis, and RT-PCR were performed. Drynariae Rhizoma extracts treatment reduced the cell viablilty of U937 cells in a dose-dependent manner, which was associated with induction of apoptotic cell death. Drynariae Rhizoma extracts treatment also reduced the levels of Bcl-xL anti-apoptotic protein expression and increased the levels of caspase-3, p53, pro-apoptotic protein, in U937 cells. RT-PCR data revealed that the level of bcl-2, bcl-xL mRNA expressions decreased in a dose-dependent manner. These findings suggest that Drynariae Rhizoma extracts may have induction of apoptotic cell death via regulation of several growth regulatory gene products. The abbreviations used are: FBS, fetal bovine serum; PBS, phosphate buffered saline; PI, propidium iodide; OD, optical density; DiOC6, 3,3-dihexyloxa carbcyanine iodide; MTT, 3 [4-5-dimethylthiazol-2-yl] -2-diphenyltetrazolium bromide.

• Toxicological Research
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• 제23권3호
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• pp.215-221
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• 2007

Although sanguinarine, a benzophenanthridine alkaloid, possesses anti-cancer properties against several cancer cell lines, the molecular mechanisms by which it inhibits cell growth and induces apoptosis have not been clearly understood. In order to further explore the critical events leading to apoptosis in sanguinarine-treated MCF-7 human breast carcinoma cells, the following effects of sanguinarine on components of the mitochondrial apoptotic pathway were examined: generation of reactive oxygen species (ROS), alteration of the mitochondrial membrane potential (MMP), and the expression changes of Bcl-2 family proteins. We show that sanguinarine-induced apoptosis is accompanied by the generation of intracellular ROS and disruption of MMP as well as an increase in pro-apoptotic Bax expression and a decrease of anti-apoptotic Bcl-2 and Bcl-xL expression. The quenching of ROS generation with N-acetyl-L-cysteine, the ROS scavenger, protected the sanguinarine-elicited ROS generation, mitochondrial dysfunction, modulation of Bcl-2 family proteins, and apoptosis. Based on these results, we propose that the cellular ROS generation plays a pivotal role in the initiation of sanguinarine-triggered apoptotic death.

• Toxicological Research
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• 제32권3호
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• pp.225-230
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• 2016

Zingiber officinale Roscoe has been widely used as a folk medicine to treat various diseases, including cancer. This study aims to re-examine the therapeutic potential of co-administration of natural products and cancer chemotherapeutics. Candidate material for this project, ${\alpha}$-zingiberene, was extracted from Zingiber officinale Roscoe, and ${\alpha}$-zingiberene makes up $35.02{\pm}0.30%$ of its total essential oil. ${\alpha}$-Zingiberene showed low $IC_{50}$ values, $60.6{\pm}3.6$, $46.2{\pm}0.6$, $172.0{\pm}6.6$, $80.3{\pm}6.6$ (${\mu}g/mL$) in HeLa, SiHa, MCF-7 and HL-60 cells each. These values are a little bit higher than $IC_{50}$ values of general essential oil in those cells. The treatment of ${\alpha}$-zingiberene produced nucleosomal DNA fragmentation in SiHa cells, and the percentage of sub-diploid cells increased in a concentration-dependent manner in SiHa cells, hallmark features of apoptosis. Mitochondrial cytochrome c activation and an in vitro caspase-3 activity assay demonstrated that the activation of caspases accompanies the apoptotic effect of ${\alpha}$-zingiberene, which mediates cell death. These results suggest that the apoptotic effect of ${\alpha}$-zingiberene on SiHa cells may converge caspase-3 activation through the release of mitochondrial cytochrome c into cytoplasm. It is considered that anti-proliferative effect of ${\alpha}$-zingiberene is a result of apoptotic effects, and ${\alpha}$-zingiberene is worth furthermore study to develop it as cancer chemotherapeutics.

• Korean Journal of Acupuncture
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• 제29권2호
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• pp.271-289
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• 2012

Objectives : Ganoderma lucidum(Ganoderma or lingzhi, 靈芝) is a well-known oriental medical mushroom containing many bioactive compounds. The possible mechanisms involved in its effects on cancer cells remain to be elucidated. In the present study, the anti-proliferative and apoptotic activities of the G. lucidum ethanol extract(GEE), in AGS human gastric cancer cells were investigated. Methods : It was found that exposure of AGS cells to GEE resulted in the growth inhibition in a dose and time dependent manner as measured by trypan blue count and MTT assay. The anti-proliferative effect of GEE treatment in AGS cells was associated with morphological changes and formation of apoptotic bodies, and the flow cytometry analysis confirmed that GEE treatment increased the populations of apoptotic-sub G1 phase. Growth inhibition and apoptosis of AGS cells by GEE were connected with a concentration and time-dependent up-regulation of tumour necrosis factor-related apoptosis-inducing ligand(TRAIL) expression. Results : The levels of XIAP and survivin expression, members of IAP family proteins, were gradually down-regulated by GEE treatment. However other members of IAP family proteins such as cIAP-1 and cIAP-2 remained unchanged in GEE-treated AGS cells. GEE treatment also induced the proteolytic activation of caspase-3, caspase-8 and caspase-9 and a concomitant degradation of poly(ADP-ribose) polymerase(PARP) protein, a caspase-3 substrate protein. Additionally, GEE-induced apoptosis was associated with the inhibition of Akt activation in a concentration and time-dependent manner, and pre-treatment with LY294002, a phosphoinositide 3-kinase(PI3K)/Akt inhibitor, significantly increased GEE-induced growth inhibition and apoptosis. Conclusions : Therefore, G. lucidum has a strong potential as a therapeutic agent for preventing cancers such as gastric cancer cells.

• Journal of Microbiology and Biotechnology
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• 제16권8호
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• pp.1262-1268
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• 2006

In the search for a novel cytotoxic substance from medicinal plants, HY53 ($C_{17}H_{32}O_2N_2$; molecular weight 296) was isolated from the leaves of Pata de Vaca (Bauhinia forficata). The growth of the HepG2 cells was inhibited in a dose-dependent manner when treated with 0.07 to 0.40 mM HY53 for 24 h (IC$_{50}$: 0.13 mM). Furthermore, nuclear DAPI staining revealed the typical nuclear features of apoptosis in the HepG2 cells exposed to 0.27 mM HY53, whereas a flow cytometric analysis of the HepG2 cells using propidium iodide showed that the apoptotic cell population increased gradually from 8% at 0 mM to 23% at 0.14 mM and 45% at 0.40 mM after being exposed to each concentration of HY53 for 24 h. Moreover, a TUNEL assay also exhibited the apoptotic induction of the HepG2 cells treated with HY53. To obtain further information on the HY53-induced apoptosis, the expression level of certain apoptosis-associated proteins was examined using a Western blot analysis. Treatment of the HepG2 cells with HY53 resulted in the activation of caspase-3, and subsequent proteolytic cleavage of poly(ADP-ribose) polymerase (PARP). Consequently, the results confirmed that the apoptosis in the HepG2 cells was induced by HY53 and the involvement of caspase-3-mediated PARP cleavage in the apoptotic process.

• Clinical and Experimental Reproductive Medicine
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• 제29권3호
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• pp.195-200
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• 2002

Objective : To investigate the expression of apoptosis related proteins and apoptotic cells on the human ovarian follicles. Materials and Methods: Thirty five Formalin-fixed paraffin-embedded human ovarian tissue blocks were selected from the surgical pathology files of the department of pathology, College of Medicine, Yonsei University, for the period from 1996 to 1998. All specimen were from premenopausal women aged from $32{\sim}45$. Ovarian tissues were collected from the patients performing hysterectomy for benign uterine diseases. Immunohistochemical staining was performed for the detection of DNA fragmented cell, Bcl-2, Bax, Fas and Fas-ligand. Results: Bcl-2 and bax were not expressed on the surrounding cells and oocyte of the primary, primordial and preantral follicles. Fas and Fas-ligand (Fas-L) were not expressed on the surrounding cells on the primordial and primary follicles. But expressed on the surrounding granulosa cells and oocyte in the primordial and primary follicles. In the healthy follicles, Bcl-2 was expressed on the granulosa cells, however, Bax was not expressed. DNA fragmented cells were expressed on the inner granulosa cell layer of atretic follicles. Conclusion: Fas, Fas-ligand, and Bax may be responsible for the follicular atresia and Bcl-2 may be involved in the follicular survival in the human ovary.

• Molecules and Cells
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• 제37권12호
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• pp.857-864
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• 2014

Astrocyte elevated gene-1 (AEG-1) is a recently discovered oncogene that has been reported to be highly expressed in various types of malignant tumors, including renal cell carcinoma. However, the precise role of AEG-1 in renal cancer cell proliferation and apoptosis has not been clarified. In this study, we transfected the renal cancer cell line Caki-1 with a plasmid expressing AEG-1 short hairpin RNA (shRNA) and obtained cell colonies with stable knockdown of AEG-1. We found that AEG-1 down-regulation inhibited cell proliferation and colony formation and arrested cell cycle progression at the sub-G1 and G0/G1 phase. Western blot analysis indicated that the expression of proliferating cell nuclear antigen (PCNA), cyclin D1 and cyclin E were significantly reduced following AEG-1 down-regulation. In addition, AEG-1 knockdown led to the appearance of apoptotic bodies in renal cancer cells, and the ratio of apoptotic cells significantly increased. Expression of the antiapoptotic factor Bcl-2 was dramatically reduced, whereas the pro-apoptotic factors Bax, caspase-3 and poly (ADPribose) polymerase (PARP) were significantly activated. Finally, AEG-1 knockdown in Caki-1 cells remarkably suppressed cell proliferation and enhanced cell apoptosis in response to 5-fluorouracil (5-FU) treatment, suggesting that AEG-1 inhibition sensitizes Caki-1 cells to 5-FU. Taken together, our data suggest that AEG-1 plays an important role in renal cancer formation and development and may be a potential target for future gene therapy for renal cell carcinoma.

• 대한임상검사과학회지
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• 제39권2호
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• pp.68-75
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• 2007

약 100년 전에 박테리아가 암을 억제한다는 보고를 바탕으로 다양한 미생물이 항암효과를 가지는 백신 개발에 이용되거나 또는 미생물의 세포 밖 독소 단백질을 찾아내고 있다. Psuedomonas aeruginosa exotoxin A(ETA)는 암세포에서 세포성장을 억제하고 세포 죽음을 유발하는 것으로 알려져 있다. 하지만 ETA가 세포 자멸사를 유도하는 정확한 기전은 아직 알려져 있지 않다. 따라서 본 연구에서는 세포자멸사의 유도를 확인하기 위해 K562 cell을 이용하여 세포의 형태학적 변화, 세포독성, Annexin-V binding assay 그리고 세포주기를 분석하였으며, 그 결과로 ETA는 K-562세포에서의 세포증식과 성장을 억제하였고, 세포자멸사 기작을 통한 K-562 암세포의 사멸을 일으켰음을 관찰하였다. 또한 flow cytometric analysis에서는 ETA가 세포주기 중 특히 sub-G1 기를 정지시키는 것으로 나타났다. 본 연구는 ETA가 인간 백혈병 K-562 암세포의 세포성장을 억제하고 sub-G1 기를 정지시킴으로서 세포자멸사를 유도하고 있음을 확인하였다.