• Asian Pacific Journal of Cancer Prevention
• /
• 제13권5호
• /
• pp.2175-2178
• /
• 2012

Purpose: Definite diagnosis of follicular thyroid carcinoma (FTC) is based on the presence of capsular or vascular invasion. To date, no reliable and practical method has been introduced to discriminate this malignant neoplasm from follicular thyroid adenoma (FTA) in fine needle aspiration biopsy material. Matrix metalloproteinase-2 (MMP-2), by degrading extracellular matrix, and caspase-3, by induction of apoptosis, have been shown to play important roles in carcinogenesis and aggressive behavior in many tumor types. The aim of this study was to examine expression of MMP-2 and caspase-3 in thyroid follicular neoplasms and to determine their usefulness for differential diagnosis. Method: Sixty FTAs and 41 FTCs were analysed immunohistochemically for MMP-2 and caspase-3. Result: MMP-2 was positive in 4 FTCs (9.8%), but in none of FTAs, with statistical significance (p= 0.025). Caspase-3 was positive in 30 (50%) of FTAs and in 27 (65.9%) of FTCs. Conclusion: Our results show MMP-2 expression only in FTCs and suggest that this protein may be a useful marker to confirm diagnosis of FTC versus FTA with 100% specificity and 100% predictive value of a positive test. We failed to show any differential diagnostic value for caspase-3 in thyroid follicular neoplasms.

• BMB Reports
• /
• 제45권9호
• /
• pp.526-531
• /
• 2012

Many malignant tumors become resistant to tumor necrosis factor-alpha (TNF-${\alpha}$)-induced cell death during carcinogenesis. In the present study, we examined whether parkin acts as a tumor suppressor in HeLa cells, a human cervical cancer cell line resistant to TNF-${\alpha}$-induced cell death. TNF-${\alpha}$-treatment alone did not affect HeLa cell viability. However, expression of parkin restored TNF-${\alpha}$-induced apoptosis in HeLa cells. Increased cell death was due to the activation of the apoptotic pathway. Expression of parkin in TNF-${\alpha}$-treated HeLa cells stimulated cleavage of the pro-apoptotic proteins caspase-8, -9, -3, -7 and poly ADP ribose polymerase (PARP). In addition, parkin expression resulted in decreased expression of the caspase inhibitory protein, survivin. These results suggest that parkin acts as a tumor suppressor in human cervical cancer cells by modulating survivin expression and caspase activity. We propose that this pathway is a novel molecular mechanism by which parkin functions as a tumor suppressor.

• Natural Product Sciences
• /
• 제22권4호
• /
• pp.293-298
• /
• 2016

Plant-derived triterpenoids commonly possesses biological properties such as anti-inflammatory, antimicrobial, anti-viral and anti-cancer. Luvunga scandens is one of the plant that produced triterpenoids. The aims of the study was to analyze cell cycle profile and to determine the expression of p53 unregulated modulator of apoptosis (PUMA), caspase-8 and caspase-9 genes at mRNA level in MCF-7 cell line treated with two triterpenoids, flindissol (1) and 3-oxotirucalla-7,24-dien-21-oic-acid (2) isolated from L. scandens. The compounds were tested for cell cycle analysis using flow cytometer and mRNA expression level using quantitative RT-PCR. The number of MCF-7 cells population which distributed in Sub G1 phase after treated with compound 1 and 2 were 7.7 and 9.3% respectively. The evaluation of the expression of genes showed that both compounds exhibited high level of expression of PUMA, caspase-8 and caspase-9 as normalized to ${\beta}-actin$ via activation of those genes. In summary, the isolated compounds of L. scandens plant showed promising anticancer properties in MCF-7 cell lines.

• Asian Pacific Journal of Cancer Prevention
• /
• 제16권5호
• /
• pp.1833-1837
• /
• 2015

An aristolactam-type alkaloid, isolated from Orophea enterocarpa, is enterocarpam-III (10-amino-2,3,4,6-tetramethoxyphenanthrene-1-carboxylic acid lactam). It is cytotoxic to various human and murine cancer cell lines; however, the molecular mechanisms remain unclear. The aims of this study were to investigate cytotoxic effects on and mechanism (s) of human cancer cell death in human hepatocellular carcinoma HepG2 and human invasive breast cancer MDA-MB-231 cells compared to normal murine fibroblast NIH3T3 cells. Cell viability was determined by MTT assay to determine $IC_{10}$, $IC_{20}$ and $IC_{50}$ levels, reactive oxygen species (ROS) production with 2',7'-dichlorohydrofluorescein diacetate and the caspase-3, -8 and -9 activities using specific chromogenic (p-nitroaniline) tetrapeptide substrates, viz., DEVD-NA, IETD-NA and LEHD-NA and employing a microplate reader. Mitochondrial transmembrane potential (MTP) was measured by staining with 3, 3'-dihexyloxacarbocyanine iodide ($DiOC_6$) and using flow cytometry. The compound was cytotoxic to HepG2 and MDA-MB-231 cells with the $IC_{50}$ levels of $26.0{\pm}4.45$ and $51.3{\pm}2.05{\mu}M$, respectively. For murine normal fibroblast NIH3T3 cells, the $IC_{50}$ concentration was $81.3{\pm}10.1{\mu}M$. ROS production was reduced in a dose-response manner in HepG2 cells. The caspase-9 and -3 activities increased in a concentration-dependent manner, whereas caspase-8 activity did not alter, indicating the intrinsic pathway activation. Enterocarpam-III decreased the mitochondrial transmembrane potential (MTP) dose-dependently in HepG2 cells, suggesting that the compound induced HepG2 cell apoptosis via the mitochondrial pathway. In conclusion, enterocarpam-III inhibited HepG2 and MDA-MB-231 cell proliferation and induced human HepG2 cells to undergo apoptosis via the intrinsic (mitochondrial) pathway and induction of caspase-9 activity.

• BMB Reports
• /
• 제48권9호
• /
• pp.531-536
• /
• 2015

Gecko proteins have long been used as anti-tumor agents in oriental medicine, without any scientific background. Although anti-tumor effects of Gecko proteins on several cancers were recently reported, their effect on bladder cancer has not been investigated. Thus, we explored the anti-tumor effect of Gecko proteins and its cellular mechanisms in human bladder cancer 5637 cells. Gecko proteins significantly reduced the viability of 5637 cells without any cytotoxic effect on normal cells. These proteins increased the Annexin-V staining and the amount of condensed chromatin, demonstrating that the Gecko proteinsinduced cell death was caused by apoptosis. Gecko proteins suppressed Akt activation, and the overexpression of constitutively active form of myristoylated Akt prevented Gecko proteins-induced death of 5637 cells. Furthermore, Gecko proteins activated caspase 9 and caspase 3/7. Taken together, our data demonstrated that Gecko proteins suppressed the Akt pathway and activated the intrinsic caspase pathway, leading to the apoptosis of bladder cancer cells. [BMB Reports 2015; 48(9): 531-536]

• 대한한의학방제학회지
• /
• 제10권2호
• /
• pp.213-223
• /
• 2002

The fruiting bodies of Isaria japonica have been traditionally used in Korea to treat cancer. An apoptosis-inducing compound, 4-Acetyl-12, 13-epoxyl -9-trichothecene-3, I5-diol (AETD), was isolated from the methanol extract of fruiting bodies of Isaria japonica Yasuda by bioassay -guided fractionation. The apoptosis of murine bladder cancer cell line (NBT-Ⅱ) by the compound was accessed by propidium iodide staining flow cytometric analysis, and apoptosis-inducing activity at $IC_{50}$ concentration (5 nmol／L) was further confirmed by a nuclear morphological change, a ladder pattern of DNA fragmentation, and an activation of caspase-3. These results indicate that AETD induces apoptosis of NBT-Ⅱ cells via expression of caspase-3.

• 생명과학회지
• /
• 제29권12호
• /
• pp.1393-1400
• /
• 2019

본 연구에서는 사람의 제대정맥 내피세포에서 고농도 당에 의해 유도되는 세포사멸과 연관된 미토콘드리아의 기능적 지표 변화에 미치는 diazoxide의 효과를 관찰하였다. 고농도 당에 노출된 내피세포에서 세포사멸이 시간에 따라 증가하였고, caspase 3와 8, 9의 활성 증가가 동반되었다. Caspase 3와 9의 억제제들이 세포사멸을 감소시킨 반면 caspase 8의 억제제는 효과가 없었다. 고농도 당에 노출된 세포에서 미토콘드리아 막전위의 탈분극과 막투과도의 증가, 치토크롬 C (cytochrome C)의 유리가 동반됨을 관찰할 수 있었다. Diazoxide는 고농도 당에 의한 미토콘드리아 의존성 세포사멸 신호의 활성화를 억제하였다. Diazoxide의 이러한 효과들은 미토콘드리아막의 ATP-억제성 칼륨통로 차단제인 5-hydroxydecanoate에 의해 차단되었다. 이상의 결과들을 종합하면 diazoxide가 미토콘드리아막의 ATP-억제성 칼륨통로 개방을 통해 미토콘드리아 의존성 세포사멸 신호기작의 활성화를 차단하여 고농도 당에 의해 유도되는 세포사멸을 억제하는 것으로 사료된다.

• Asian Pacific Journal of Cancer Prevention
• /
• 제17권3호
• /
• pp.1499-1506
• /
• 2016

Background: Crocus sativus and its major constituent crocin are well established to have anti-cancer properties in breast cancer cells (MCF-7). However the role of C. sativus extract (CSE) and crocin on caspase signaling mediated MCF-7 cell death at molecular level is remains unclear. In this study, we tried to unravel role of CSE and crocin on caspase mediated MCF-7 cells death and their in vivo preclinical toxicity profiling and immune stimulatory effect. Materials and Methods: CSE extract was fractionated by HPLC and crocin was isolated and characterized by NMR, IR, and MS. MCF-7 cells were treated with both CSE and crocin and expression of Bcl-2 and Bax was assessed after 24 and 36 hours. Furthermore, caspase 3, caspase 8 and caspase 9 expression was determined by Western blotting after 24 hours of treatment. DNA fragmentation analysis was performed for genotoxicity of CSE and crocin in MCF-7 cells. The in vivo toxicity profile of CSE (300 mg/kg of b.wt) was investigated in normal Swiss albino mice. In addition, peritoneal macrophages were collected from crocin (1, 1.5 and 2 mg/kg body weight) treated mice and analyzed for ex vivo yeast phagocytosis. Results: Immunoblot analysis revealed that there was time dependent decline in anti-apoptotic Bcl-2 with simultaneous upregulation of Bax in CSE and crocin treated MCF-7 cells. Further CSE and crocin treatment downregulated caspase 8 and 9 and cleaved the caspase 3 after 24 hours. Both CSE and crocin elicited considerable DNA damage in MCF-7 cells at each concentration tested. In vivo toxicity profile by histological studies revealed no observable histopathologic differences in the liver, kidney, spleen, lungs and heart in CSE treated and untreated groups. Crocin treatment elicited significant dose and time dependent ex vivo yeast phagocytosis by peritoneal macrophages. Conclusions: Our study delineated involvement of pro-apoptotic and caspase mediated MCF-7 cell death by CSE and crocin at the molecular level accompanied with extensive DNA damage. Further we found that normal swiss albino mice can tolerate the maximum dose of CSE. Crocin enhanced ex vivo macrophage yeast phagocytic ability.

• 한국식품영양과학회지
• /
• 제41권7호
• /
• pp.901-906
• /
• 2012

본 연구에서는 넓패 메탄올 추출물의 농도별 처리가 인체 자궁암 세포 HeLa의 세포사멸에 미치는 영향을 확인하기 위하여 세포독성 측정, Hoechst 33258 staining, flow cytometry 분석을 통하여 세포사멸을 확인하였다. 넓패 메탄올 추출물 처리 시 HeLa 세포에서 농도 의존적으로 세포의 증식을 억제하였으며, 또한 넓패 메탄올 추출물은 농도 의존적으로 핵을 응축하고 apoptotic bodies을 생성하였다. 유세포 분석을 통하여 apoptosis를 측정한 결과, 넓패 메탄올 추출물의 농도가 증가함에 따라 유의적으로 apoptotic 세포가 증가하였다. Western blot을 통해 PARP 단백질의 절단 현상을 분석한 결과, 넓패 메탄올 추출물의 처리 농도와 시간에 따라 PARP 단백질의 절단 현상이 증가하였다. 또한 넓패 메탄올 추출물은 caspase-8, caspase-9 및 caspase-3 활성을 농도와 시간에 따라 증가시켰으며, caspase 저해제인 z-VAD-fmk로 처리 시 넓패 메탄올 추출물에 의한 세포사멸이 유의적으로 감소되어 넓패 메탄올 추출물에 의한 HeLa 세포의 apoptosis 유도에 caspase가 중요한 역할을 하고 있음을 확인하였다. 따라서 넓패 메탄올 추출물은 HeLa 자궁암 세포의 apoptosis를 유도하는 것으로 나타나 넓패의 항암효과 가능성을 제시하였다.

• Nutrition Research and Practice
• /
• 제8권2호
• /
• pp.132-137
• /
• 2014

BACKGROUND/OBJECTIVES: In this study, the apoptogenic activity and mechanisms of cell death induced by hexane extract of aged black garlic (HEABG) were investigated in human leukemic U937 cells. MATERIALS/METHODS: Cytotoxicity was evaluated by MTT (3-(4, 5-dimethyl-thiazol-2-yl)-2, 5-diphenyl tetrazoliumbromide) assay. Apoptosis was detected using 4,6-diamidino-2-phenyllindile (DAPI) staining, agarose gel electrophoresis and flow cytometry. The protein levels were determined by Western blot analysis. Caspase activity was measured using a colorimetric assay. RESULTS: Exposure to HEABG was found to result in a concentration- and time-dependent growth inhibition by induction of apoptosis, which was associated with an up-regulation of death receptor 4 and Fas legend, and an increase in the ratio of Bax/Bcl-2 protein expression. Apoptosis-inducing concentrations of HEABG induced the activation of caspase-9, an initiator caspase of the mitochodrial mediated intrinsic pathway, and caspase-3, accompanied by proteolytic degradation of poly(ADP-ribose)-polymerase. HEABG also induced apoptosis via a death receptor mediated extrinsic pathway by caspase-8 activation, resulting in the truncation of Bid, and suggesting the existence of cross-talk between the extrinsic and intrinsic pathways. However, pre-treatment of U937 cells with the caspase-3 inhibitor, z-DEVD-fmk, significantly blocked the HEABG-induced apoptosis of these cells, and increased the survival rate of HEABG-treated cells, confirming that HEABG-induced apoptosis is mediated through activation of caspase cascade. CONCLUSIONS: Based on the overall results, we suggest that HEABG reduces leukemic cell growth by inducing caspase-dependent apoptosis through both intrinsic and extrinsic pathways, implying its potential therapeutic value in the treatment of leukemia.