• Asian Pacific Journal of Cancer Prevention
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• 제15권12호
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• pp.5071-5076
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• 2014

Cholangiocarcinoma (CCA) is one of the aggressive cancers with a very poor prognosis. Several efforts have been made to identify and develop new agents for prevention and treatment of this deadly disease. In the present study, we examined the anticancer effect of luteolin on human CCA, KKU-M156 cells. Sulforhodamine B assays showed that luteolin had potent cytotoxicity on CCA cells with IC50 values of $10.5{\pm}5.0$ and $8.7{\pm}3.5{\mu}M$ at 24 and 48 h, respectively. Treatment with luteolin also caused a concentration-dependent decline in colony forming ability. Consistent with growth inhibitory effects, luteolin arrested cell cycle progression at the G2/M phase in a dose-dependent manner as assessed by flow cytometry analysis. Protein expression of cyclin A and Cdc25A was down-regulated after luteolin treatment, supporting the arrest of cells at the G2/M boundary. Besides evident G2/M arrest, luteolin induced apoptosis of KKU-M156 cells, demonstrated by a distinct sub-G1 apoptotic peak and fluorescent dye staining. A decrease in the level of anti-apoptotic Bcl-2 protein was implicated in luteolin-induced apoptosis. We further investigated the effect of luteolin on JAK/STAT3, which is an important pathway involved in the development of CCA. The results showed that interleukin-6 (IL-6)-induced JAK/STAT3 activation in KKU-M156 cells was suppressed by treatment with luteolin. Treatment with a specific JAK inhibitor, AG490, and luteolin diminished IL-6-stimulated CCA cell migration as assessed by wound healing assay. These data revealed anticancer activity of luteolin against CCA so the agent might have potential for CCA prevention and therapy.

• Animal cells and systems
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• 제11권1호
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• pp.9-15
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• 2007

The naphthoquinone analog (2,3-dichloro-6,9-dihydroxy-1,4-naphtoquinone, NA) has an inhibitory effect on cdc25A protein phosphatase in vitro, which is responsible for G1/S transition during cell cycle. However, the exact mechanism inducing the growth inhibition is not understood. In this study, we investigated the regulatory mechanisms of growth arrest induced by NA, as a new potent inhibitor of cdc25A phosphatase, in human hepatocarcinoma SK-hep-1 cells. We found that NA induced the G1 arrest by perturbation of protein tyrosine dephosphorylation of Cdk2, which may be resulting from inhibition of cdc25A phosphatase. In addition, p21 was expressed in a p53-independent manner and participated in the NA-induced G1 arrest by inhibiting Cdk2 activity. Although the exact mechanism is not known, the p21 expression might be related to MAPK activation. From these results, we suggest that NA induces G1 arrest via inhibition of cdc25A and induction of p53-independent p21 expression in SK-Hep-1 cells.

• Molecules and Cells
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• 제39권9호
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• pp.699-704
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• 2016

Developmentally regulated GTP-binding protein 2 (DRG2) plays an important role in cell growth. Here we explored the linkage between DRG2 and G2/M phase checkpoint function in cell cycle progression. We observed that knockdown of DRG2 in HeLa cells affected growth in a wound-healing assay, and tumorigenicity in nude mice xenografts. Flow cytometry assays and [$^3H$] incorporation assays indicated that G2/M phase arrest was responsible for the decreased proliferation of these cells. Knockdown of DRG2 elicited down-regulation of the major mitotic promoting factor, the cyclin B1/Cdk1 complex, but upregulation of the cell cycle arresting proteins, Wee1, Myt1, and p21. These findings identify a novel role of DRG2 in G2/M progression.

• Radiation Oncology Journal
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• 제30권2호
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• pp.78-87
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• 2012

Purpose: Troglitazone (TRO) is a peroxisome proliferator-activated receptor ${\gamma}$ ($PPAR{\gamma}$) agonist. TRO has antiproliferative activity on many kinds of cancer cells via G1 arrest. TRO also increases $Cu^{2+}/Zn^{2+}$-superoxide dismutase (CuZnSOD) and catalase. Cell cycle, and SOD and catalase may affect on radiation sensitivity. We investigated the effect of TRO on radiation sensitivity in cancer cells in vitro. Materials and Methods: Three human cervix cancer cell lines (HeLa, Me180, and SiHa) were used. The protein expressions of SOD and catalase, and catalase activities were measured at 2-10 ${\mu}M$ of TRO for 24 hours. Cell cycle was evaluated with flow cytometry. Reactive oxygen species (ROS) was measured using 2',7'-dichlorofluorescin diacetate. Cell survival by radiation was measured with clonogenic assay. Results: By 5 ${\mu}M$ TRO for 24 hours, the mRNA, protein expression and activity of catalase were increased in all three cell lines. G0-G1 phase cells were increased in HeLa and Me180 by 5 ${\mu}M$ TRO for 24 hours, but those were not increased in SiHa. By pretreatment with 5 ${\mu}M$ TRO radiation sensitivity was increased in HeLa and Me180, but it was decreased in SiHa. In Me180, with 2 ${\mu}M$ TRO which increased catalase but not increased G0-G1 cells, radiosensitization was not observed. ROS produced by radiation was decreased with TRO. Conclusion: TRO increases radiation sensitivity through G0-G1 arrest or decreases radiation sensitivity through catalase-mediated ROS scavenging according to TRO dose or cell types. The change of radiation sensitivity by combined with TRO is not dependent on the PPAR ${\gamma}$ expression level.

• 대한한의학회지
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• 제20권2호
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• pp.88-97
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• 1999

To characterize the effects of Gilgyung-Tang(GGT) on cellular proliferation and viability of normal lung fibroblast cells, we examined the cell cycle progression and cell cycle-related gene expression in T3891 using a flow cytometry and a quantitative RT-PCR analysis. 1. The significant surpression effect of cellular proliferations of GGT was observed in proportion to a certain concentration and time. 2. GGT was identified to induce apoptotic death of damaged cells by treatment with a DNA-damage agent and etoposide, while it stimulated the recovery of cellular viability of normal cells. 3 The significant reductions of mRNA expression of PCAN, c-Fos treated by GGT were observed. 4. The significant inductions of mRNA expression of p53, CDKN1. Gadd45 treated by GGT were observed. 5. The apoptosis caused by the reduction of Bcl-2 genes was significant and the Bax genes were increased. but the amount of Fas genes were not changed. These results strongly suggest that GGT triggers arrest of the cell cycle at G1 phase, and thus causes an inhibition of cellular proliferation of human normal lung cells through the transcriptional up-regulation of cell cycle inhibitory genes and down-regulation of induction of cell cycle stimulating genes respectably.

• 한국생명과학회:학술대회논문집
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• 한국생명과학회 2008년도 제49회 학술심포지움 및 국제학술대회
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• pp.90.2-90.2
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• 2008

• 생명과학회지
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• 제16권1호
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• pp.12-16
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• 2006

항암유전자 Brca1의 변이는 유방암 및 난소암에 대한 감수성을 증가시키며, Brca1은 DNA손상신호후 세포주기 조절에 필수적인 역할을 한다. 연구결과, Brca1이 세포주기 S기와 G2/M 조절점에서 중요한 역할을 담당함이 밝혀졌다. 그러나, Brca1의 spindle checkpoint 관여여부는 알려져 있지 않다. 본 연구에서는 spindle checkpoint를 활성화시키는 nocodazole를 처리하여 야생형, $p53^{-/-}$ 그리고 $p53^{-/-}\;Brca1^{-/-}$ 세포주의 세포주기 변화를 조사하였다. 야생형과 $p53^{-/-}$ 세포주는 신속한 mitosis기 정지가 나타난 반면, $p53^{-/-}\;Brca1^{-/-}$ 세포주의 경우 모든 세포가 M기에서 정지하지 않았다. Double-thymidine block 기법에 의한 세포주기 동조화후 nocodazole 처리시에도 $p53^{-/-}\;Brca1^{-/-}$ 세포주에서는 일부세포가 M기 조절점을 통과하여 계속 G1기로 진행하였다. 형태학적 분석에서도 nocodazole 함유배지에서 계속 증식하는 세포형태가 관찰되었다. 이와 같은 결과들은 Brca1이 spindle checkpoint가 정상적으로 작동하는데 중요한 역할을 담당한다는 것을 의미하고 있다.

• 자연과학논문집
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• 제9권1호
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• pp.1-7
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• 1997

칼슘/calmodulin-의존적 단백질 인산화 효소 II (CaMK-II)는 세포의 여러 기능을 조절하는 다양한 단백질들을 인산화시키는 효소이다. 세포 내부의 칼슘의 농도는 세포의 주기에 따라 변하므로 CaMK-II의 활성도 역시 세포주기에 따라 변하는 지를 조사함으로 세포주기에서의 CaMK-II의 역할을 알아보려 하였다. NIH3T3 세포를 CaMK-II의 활성도에는 전혀 영향을 주지 않는 여러 가지 약제로 처리하여 세포주기상의 특정한 시점에 동일하게 정지시킨 후, 세포내의 CaMK-II 활성도를 합성 펩타이드기질을 이용하여 측정하였다. 또한 일정 시점으로부터 동조화된 세포내의 CaMK-II의 활성도의 변화를 측정하여 한 세포주기 동안 효소의 활성도 변화의 양상을 조사하였다. 세포주기상 각각 G0, G1, G1/S, G2/M기에 정지된 세포내의 CaMK-II 총활성도는 대조군과 차이가 없었으나 M기에서는 낮았다. 그러나 자가인산화에 의한 CaMK-II의 칼슘-비의존성 활성도는 M기에서 가장 높았다. 이러한 양상은 G1기에서부터 동조화된 세포내 CaMK-II의 칼슘-비의존성 활성도 변화 양상과도 일치하였다. CaMK-II의 생리학적 의미를 지닌 활성도는 인산화에 의한 calcium-비의존성 활성도임을 비추어 볼 때 M기에서 CaMK-II가 세포분열의 과정에서 중요한 기능을 하고 있음을 보여주고 있다.

• Journal of Microbiology and Biotechnology
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• 제28권4호
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• pp.542-550
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• 2018

Heat shock protein 90 (Hsp90) is treated as a molecular therapeutic target for the prevention and treatment of cancer. Geldanamycin (GA) was the first identified natural Hsp90 inhibitor, but hepatotoxicity has limited its clinical application. Nevertheless, a new GA analog (WK-88-1) with the non-benzoquinone skeleton, obtained from genetically engineered Streptomyces hygroscopicus, was found to have anticancer activity against two human breast cancer cell lines. WK-88-1 produced concentration-dependent inhibition of cell proliferation, cell cycle arrest, and apoptosis in estrogen receptor (ER)-positive MCF-7 and ER-negative MDA-MB-231 cell lines. Detailed analysis showed that WK-88-1 downregulated some key cell cycle molecules (CDK1 and cyclin B1) and lead to $G_2/M$ cell cycle arrest. Further studies also showed that WK-88-1 could induce human breast cancer cell apoptosis by downregulating Hsp90 client proteins (Akt, p-Akt, IKK, c-Raf, and Bcl-2), decreasing the ATP level, increasing reactive oxygen species production, and lowering the mitochondrial membrane potential. Meanwhile, we discovered that WK-88-1 significantly decreased the levels of Her-2 and $ER-{\alpha}$ in MCF-7 cells but not in MDA-MB-231 cells. In addition, WK-88-1 significantly increased caspase-3, -8, and -9 activities and the cleavage of PARP in a concentration-dependent manner (with the exception of caspase-3 and PARP in MCF-7 cells). Taken together, our preliminary results suggest that WK-88-1 has the potential to play a role in breast cancer therapy.

• The Korean Journal of Physiology and Pharmacology
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• 제19권5호
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• pp.421-426
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• 2015

The increased potential for vascular smooth muscle cell (VSMC) growth is a key abnormality in the development of atherosclerosis and post-angioplasty restenosis. Abnormally high activity of platelet-derived growth factor (PDGF) is believed to play a central role in the etiology of these pathophysiological situations. Here, we investigated the anti-proliferative effects and possible mechanism(s) of murrayafoline A, a carbazole alkaloid isolated from Glycosmis stenocarpa Guillamin (Rutaceae), on PDGF-BB-stimulated VSMCs. Murrayafoline A inhibited the PDGF-BB-stimulated proliferation of VSMCs in a concentration-dependent manner, as measured using a non-radioactive colorimetric WST-1 assay and direct cell counting. Furthermore, murrayafoline A suppressed the PDGF-BB-stimulated progression through $G_0/G_1$ to S phase of the cell cycle, as measured by [$^3H$]-thymidine incorporation assay and cell cycle progression analysis. This anti-proliferative action of murrayafoline A, arresting cell cycle progression at $G_0/G_1$ phase in PDGF-BB-stimulated VSMCs, was mediated via down-regulation of the expression of cyclin D1, cyclin E, cyclin-dependent kinase (CDK)2, CDK4, and proliferating cell nuclear antigen (PCNA), and the phosphorylation of retinoblastoma protein (pRb). These results indicate that murrayafoline A may be useful in preventing the progression of vascular complications such as restenosis after percutaneous transluminal coronary angioplasty and atherosclerosis.