• Toxicological Research
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• 제35권2호
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• pp.167-179
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• 2019

Ovarian cancer is the fifth main cause of pre-senescent death in women. Although chemotherapy is generally an efficient treatment, its side effects and the occurrence of chemotherapeutic resistance have prompted the need for alternative treatments. In this study, ${\alpha}$-mangostin and apigenin were evaluated as possible anticancer alternatives to the chemotherapeutic drug doxorubicin, used herein as a positive control. The ovarian adenocarcinoma cell line SKOV-3 (ATCC No. HTB77) was used as model ovarian cancer cells, whereas the skin fibroblast line CCD-986Sk (ATCC No. CRL-1947) and lung fibroblast line WI-38 (ATCC No. CCL-75) were used as model untransformed cells. Apigenin and doxorubicin inhibited the growth of SKOV-3 cells in a dose- and time-dependent manner. After 72 hr exposure, doxorubicin was mostly toxic to SKOV-3 cells, whereas apigenin was toxic to SKOV-3 cells but not CCD-986Sk and WI-38 cells. ${\alpha}$-Mangostin was more toxic to SKOV-3 cells than to CCD-986Sk cells. A lower cell density, cell shrinkage, and more unattached (floating round) cells were observed in all treated SKOV-3 cells, but the greatest effects were observed with ${\alpha}$-mangostin. With regard to programmed cell death, apigenin caused early apoptosis within 24 hr, whereas ${\alpha}$-mangostin and doxorubicin caused late apoptosis and necrosis after 72 hr of exposure. Caspase-3 activity was significantly increased in ${\alpha}$-mangostin-treated SKOV-3 cells after 12 hr of exposure, whereas only caspase-9 activity was significantly increased in apigenin-treated SKOV-3 cells at 24 hr. Both ${\alpha}$-mangostin and apigenin arrested the cell cycle at the $G_2/M$ phase, but after 24 and 48 hr, respectively. Significant upregulation of BCL2 (apoptosis-associated gene) and COX2 (inflammation-associated gene) transcripts was observed in apigenin- and ${\alpha}$-mangostin-treated SKOV-3 cells, respectively. ${\alpha}$-Mangostin and apigenin are therefore alternative options for SKOV-3 cell inhibition, with apigenin causing rapid early apoptosis related to the intrinsic apoptotic pathway, and ${\alpha}$-mangostin likely being involved with inflammation.

• Asian Pacific Journal of Cancer Prevention
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• 제16권6호
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• pp.2453-2458
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• 2015

Background: X-linked inhibitor of apoptosis protein (XIAP) associated factor 1 (XAF1) exhibits aberrantly low or absent expression in various human malignancies, closely associated with anti-apoptosis and overgrowth of cancer cells. However, limited attention has been directed towards the contribution of XAF1 to invasion, apoptosis, and cisplatin (DDP)-resistance of epithelial ovarian cancer (EOC) cells. This study aimed to evaluate the potential effects of XAF1 on invasion, cell cycle, apoptosis, and cisplatin-resistance by overexpressing XAF1 in SKOV-3 and SKOV-3/DDP cells. Methods and Results: The pEGFP-C1-XAF1 plasmid was transfected into SKOV-3 and SKOV-3/DDP cells, and the expression of XAF1 at both mRNA and protein levels was analyzed by reverse transcription-PCR and Western blotting. Overexpression of XAF1 suppressed XIAP expression in both SKOV-3 and SKOV-3/DDP cells. Transwell invasion assays demonstrated that XAF1 exerted a strong anti-invasive effect in XAF1-overexpressing cells. Moreover, flow cytometry analysis revealed that XAF1 overexpression arrested the cell cycle at G0/G1 phase, and cell apoptosis analysis showed that overexpression of XAF1 enhanced apoptosis of SKOV-3 and SKOV-3/DDP cells apparently by activating caspase-9 and caspase-3. Furthermore, MTT assay confirmed a dose-dependent inhibitory effect of cisplatin in the tested tumor cells, and overexpression of XAF1 increased the sensitivity of SKOV-3 and SKOV-3/DDP cells to cisplatin-mediated antiproliferative effects. Conclusions: In summary, our data indicated that overexpression of XAF1 could suppress XIAP expression, inhibit invasion, arrest cell cycle, promote apoptosis, and confer cisplatin-sensitivity in SKOV-3 and SKOV-3/DDP cells. Therefore, XAF1 may be further assessed as a potential target for the treatment of both cisplatin-resistant and non-resistant EOCs.

• 생명과학회지
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• 제33권2호
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• pp.129-137
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• 2023

활성산소는 세포의 다양한 생리활성에 중요한 역할을 수행한다. 본 연구에서는 리소포스파티드산에 의해 유도되는 SKOV-3 세포의 이동을 조절하는 신호전달 기전 연구를 수행하였다. IGF-1 및 LPA는 처리시간 그리고 용량 의존적으로 SKOV-3 세포의 이동을 촉진시켰으며, 리소포스파티드산은 이에 따라 Akt의 인산화도 촉진하였다. 리소포스파티드산에 의한 세포이동은 리소포스파티드산 수용체 억제제에 의해 길항되었으나 IGF-1에 의한 세포이동에는 영향이 없었다. PI3K 및 ROCK의 억제는 리소포스파티드산에 의한 세포의 이동을 길항하였으나 MAPK 억제제에 의해서는 길항되지 않았다. 리소포스파티드산에 의해 형성되는 활성산소는 PI3K 및 ROCK의 억제제에 의해 길항되었으며 활성산소를 킬레이트화하면 리소포스파티드산에 의한 세포의 이동이 억제되었다. 또한 리간드에 의해 활성산소를 형성하는 NOX를 억제하면 리소포스파티드산에 의한 세포의 이동도 억제 되는 것이 관찰되었다. Rictor 및 Akt1의 발현을 억제하면 활성산소 및 세포의 이동이 저해되었으나 Raptor 및 Akt2의 발현조절은 모두 영향이 없는 것으로 관찰되었다. 마지막으로 우성활성화형태인 Akt1의 과발현은 리소포스파티드산의 자극이 없어도 SKOV-3 세포의 이동을 촉진하는 것으로 관찰되었다. 이러한 결과들을 바탕으로 리소포스파티드산은 mTORC2/Akt1/NOX 신호전달 기전을 통해 활성산소를 형성하고 SKOV-3 난소암세포의 이동을 촉진한다는 것을 제안한다.

• Asian Pacific Journal of Cancer Prevention
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• 제14권12호
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• pp.7197-7201
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• 2013

Aim: To investigate the effects of diallyl trisulfide (DT) on apoptosis of cisplatin (DDP)-resistant human epithelial ovarian cancer SKOV-3 cells (SKOV-3/DDP), and the role of p53 upregulated modulator of apoptosis (PUMA). Methods: SKOV-3/DDP cells were randomly divided into control, DT, DPP and DPP+DT groups, which were treated with DT or combined DT and DDP. All cells were incubated for 48 h. and apoptosis rates were assessed by flow cytometry. mRNA and protein expression of PUMA, Bax and Bcl-2 was determined by RT-PCR and Western blot assays, respectively. Results: Compared with control group, the apoptosis rates of SKOV-3/DDP cells in DT groups were obviously increased, with dose-dependence (P < 0.05), the mRNA and protein expressions of PUMA, Bax also being up-regulated (P < 0.05), while those of Bcl-2 were down-regulated (P < 0.05). Compared with DT groups, the apoptosis rate in the DDP+DT group was significantly increased (P < 0.05). After knockdown of PUMA with specific siRNA, the apoptosis rate of SKOV-3/DDP cells was obviously decreased (P < 0.05). Conclusion: DT can promote the apoptosis of SKOV-3/DDP cells with PUMA playing a critical role.

• Asian Pacific Journal of Cancer Prevention
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• 제15권20호
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• pp.8595-8601
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• 2014

Background: miR-200a expression is frequently altered in numerous cancers. The aim of the present study was to determine the role of microRNA-200a in advanced ovarian carcinomas. Materials and Methods: We measured miR-200a expression in 72 matched normal ovarian tissues and advanced ovarian carcinomas, and also two ovarian carcinoma cell lines (SKOV3 and SKOV3.ip1 - the latter being more invasive and metastatic than the parental SKOV3) by stem-loop real-time RT-PCR based on TaqMan microRNA assay using U6 as a reference. Levels of miR-200a expression were compared by disease stage, tumor grade, histology, and lymph node involvement. To evaluate the role of microRNA-200a, cell proliferation and invasion of SKOV-3 and SKOV-3.ip1 were analyzed with miR-200a inhibitor/mimic transfected cells. Results: Of 72 paired samples, 65 cancer tissues overexpressed microRNA-200a greater than two fold in comparison with matched normal epithelium. Specifically, patients with lymph node metastasis showed significant elevation. The level correlated with clinicopathological features, including high tumor grade, late disease stage, most notably with lymph node metastasis, but not with tumor histology. In addition, SKOV-3.ip1 cells also overexpressed miR-200a compared with SKOV-3, and miR-200a inhibitor transfected SKOV-3.ip1 cells showed significant reduction in cellular proliferation and invasion, while a miR-200a mimic stimulated the opposite behavior. Conclusions: We provide definitive evidence that miR-200a is up-regulated in a significant proportion of advanced ovarian carcinomas, and that elevated miR-200a expression facilitates tumor progression. Our findings support the notion that miR-200a is an onco-microRNA for ovarian cancer, and elevation is a useful potential diagnostic indicator. This study also provides a solid basis for further functional analysis of miR-200a in advanced ovarian cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제15권2호
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• pp.797-802
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• 2014

We screened a small molecular library that was designed and independently synthesized in vitro and found a new drug (MY-03-01) that is active against ovarian cancer. We established that MY-03-01 effectively inhibited SKOV-3 cell survival in a dose-dependent manner, based on cell viability rates, and that it not only induced SKOV-3 apoptosis by itself, but also did so synergistically with paclitaxel. Secondly, when MY-03-01 was applied at $40{\mu}M$, its hemolytic activity was less than 10%, compared with the control, and there was almost no damage to nor mal cells at this concentration. In addition, we used DAPI staining and flow cytometry to show that MY-03-01 could significantly induce apoptosis of SKOV-3 cells. Finally, we found that MY-03-01 likely induced SKOV-3 apoptosis by activating caspase3 and caspase9 through the mitochondrial pathway.

• Journal of Ginseng Research
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• 제44권1호
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• pp.96-104
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• 2020

Objectives: Oleanolic acid, a minor element of ginsenosides, and its derivatives have been shown to have cytotoxicity against some tumor cells. The impact of cytotoxic effect of oleanolic acid 3-acetate on ovarian cancer SKOV3 cells and endometrial cancer HEC-1A cells were examined both in vivo and in vitro to explore the underlying mechanisms. Methods: Cytotoxic effects of oleanolic acid 3-acetate were assessed by cell viability, phosphatidylserine exposure on the cell surface, mitochondrial release of cytochrome C, nuclear translocation of apoptosis-inducing factor, depolarization of mitochondrial transmembrane potential (∆Ψ_m), and generation of reactive oxygen species (ROS). In vivo inhibition of tumor growth was also assessed with xenografts in immunocompromised mice. Results: Oleanolic acid 3-acetate exhibited potent cytotoxicity toward SKOV3 and HEC-1A cells by decreasing cell viability in a concentration-dependent manner. Importantly, oleanolic acid 3-acetate effectively suppressed the growth of SKOV3 cell tumor xenografts in immunocompromised mice. Furthermore, oleanolic acid 3-acetate induced apoptotic cell death as revealed by loss of ∆Ψ_m, release of cytochrome c, and nuclear translocation of apoptosis-inducing factor with a concomitant activation of many proapoptotic cellular components including poly(ADP-ribose) polymerase, Bcl-2, and caspases-8, caspase-3, and caspase-7. Oleanolic acid 3-acetate, however, caused a decrease in ROS production, suggesting the involvement of an ROS-independent pathway in oleanolic acid 3-acetate-induced apoptosis in SKOV3 and HEC-1A cells. Conclusion: These findings support the notion that oleanolic acid 3-acetate could be used as a potent anticancer supplementary agent against ovarian and endometrial cancer. Oleanolic acid 3-acetate exerts its proapoptotic effects through a rather unique molecular mechanism that involves an unconventional ROS-independent but mitochondria-mediated pathway.

• 대한임상검사과학회지
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• 제47권4호
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• pp.298-305
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• 2015

최근 종양을 극복하고자 하는 새로운 접근 방법가운데 하나로, 종양세포내에 발현되는 줄기세포 전사인자들(Oct4, Sox2, KLF4 and Lin28)을 억제하여 종양을 치료하는 연구들이 증가하고 있다. 본 실험은 미분화 전사인자를 표적(조절)하는 microRNA-126을 이용하여 난소종양세포들(6종: HSC832(t)c, Ovcar3, Skov3, PA-1, TOV21G and Tov112D)들 생존과 성장에 어떠한 생물학적 변화를 유도하는지 연구하였다. Scramble과 microRNA-126를 난소종양세포들에 처리 후 세포모양 관찰결과 Skov3를 제외한 난소 종양세포들에서 형태학적 모양 변성과 부유현상을 관찰하였다. CCK-8을 이용한 세포분열능 분석에서 Skov3를 제외한 난소 종양세포들의 분열능력이 점차적으로 감소되는 것을 확인하였다. 특히 Tov112D, Tov21G and PA-1에서 각 시간대별로 뚜렷한 세포분열 능력 감소를 확인할 수 있었다. RT-PCR결과 미분화 전사인자들(Sox2, Lin28)의 발현감소를 확인할 수 있었다. 이러한 결과들은 microRNA-126이 다양한 난소 종양세포들을 표적하여 세포분열능과 사멸을 유도할 수 있는 가역적 환경(유전자 발현조절)을 제공함과 동시에 임상 치료에 대한 분자생물학적 단서를 제공할 수 있을 것이다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권8호
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• pp.3363-3368
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• 2014

Background: Curcumin, a phenolic compound extracted from the rhizomes of Curcuma longa, has shown cytotoxic effects against a variety of cancers. The aim of this study was to identify potential microRNA (miRNA) mediators of the anticancer effects of curcumin in ovarian cancer cells. Materials and Methods: SKOV3 ovarian cancer cells were treated with curcumin ($10-60{\mu}M$) and miR-9 expression, cell proliferation, and apoptosis were assessed. The effects of miR-9 depletion on curcumin-mediated growth suppression were also examined. Phosphorylation of Akt and forkhead box protein O1 (FOXO1) was measured in cells with miR-9 overexpression or curcumin treatment. Results: Curcumin caused a significant and dose-dependent increase of miR-9 expression in SKOV3 cells, while significantly impeding cell proliferation and stimulating apoptosis. Depletion of miR-9 significantly (p<0.05) attenuated the growth-suppressive effects of curcumin on SKOV3 cells, coupled with reduced percentages of apoptotic cells. In contrast, overexpression of miR-9 significantly enhanced the cleavage of caspase-3 and poly(ADP-ribose) polymerase and promoted apoptotic death in SKOV3 cells. Western blot analysis showed that both miR-9 overexpression and curcumin similarly caused a significant (p<0.05) decline in the phosphorylation of Akt and FOXO1, compared to untreated cells. Conclusions: The present study provided evidence that curcumin exerts its cytotoxic effects against SKOV3 ovarian cancer cells largely through upregulation of miR-9 and subsequent modulation of Akt/FOXO1 axis. Further studies are needed to identify direct targets of miR-9 that mediate the anticancer effects of curcumin in ovarian cancer cells.

• 생명과학회지
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• 제22권12호
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• pp.1621-1627
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• 2012

세포의 이동은 성장, 면역 작용, 그리고 혈관 신생 등 많은 생리현상에 중요한 역할을 한다. 또한 염증 및 종양 세포 침윤 등의 다양한 병리적 현상과도 밀접한 연관이 있다. 본 연구에서는 lysophosphatidic acid (LPA)는 활성산소의 생성을 통해 SKOV-3 난소암세포의 이동을 조절한다는 것을 관찰하였다. 먼저, 난소 암세포인 SKOV-3에서 LPA에 의한 세포의 이동이 강하게 일어남을 확인하였다. LPA에 의한 SKOV-3 세포의 이동은 phosphatidylinositol 3-kinase (PI3K)/Akt 신호전달체계를 저해시키는 약물에 의해서 완벽히 억제됨을 확인하였으나 ERK 신호전달체계를 저해시키는 약물에 의해서는 전혀 영향을 받지 않았다. 그리고 SKOV-3 세포에서 LPA에 의한 활성산소 형성이 시간에 따라 강하게 일어남을 확인하였다. 더욱이 LPA에 의한 활성산소 형성도 PI3K 또는 Akt의 저해제에 의해서 완벽히 억제됨을 확인하였으나 ERK 신호전달을 억제하였을 때는 거의 영향을 받지 않았다. SKOV-3 세포에서 LPA에 의해 생성된 활성산소는 diphenylene idonium (DPI, $10{\mu}M$), apocyanin (Apo, $10{\mu}M$)과 같은 NADPH oxidase 억제제를 전 처리하였을 때 활성산소가 형성되지 못함을 관찰하였다. 그러나 xanthine oxidase (allopurinol, Allo, $10{\mu}M$), cyclooxygenase (indomethacin, Indo, $10{\mu}M$), 또는 mitochondrial respiratory chain complex I (rotenone, Rot, $10{\mu}M$)를 억제하였을 때는 LPA에 의한 활성산소 형성에 영향을 주지 못함을 확인하였다. 마지막으로 활성산소 억제제인 N-acetylcysteine (NAC, $10{\mu}M$)에 의해서 LPA에 의한 암세포의 이동이 억제됨을 관찰하였다. 이와 더불어 LPA에 의한 SKOV-3 세포의 이동도 NADPH oxidase 억제에 의해 저해가 됨을 확인하였다. 이러한 연구결과로 보아 LPA에 의한 활성산소의 형성에는 PI3K/Akt/NADPH oxidase 신호전달체계가 중추적인 역할을 하며 이를 통해 암세포의 이동을 조절한다는 것을 알 수 있었다.