• Asian Pacific Journal of Cancer Prevention
• /
• 제17권3호
• /
• pp.1299-1308
• /
• 2016

Our previous studies clearly demonstrated that a combination of WP 631 and Epo B has higher activity against ovarian cancer cells than either of these compounds used separately. In order to fully understand the exact mechanism of action in combination, we assessed effects on the cell cycle of SKOV-3 cells. We evaluated three control points essential for WP 631 and Epo B action to determine which cell cycle-regulating proteins (CDK1/cyclin B complex, EpCAM or HMGB1) mediate activity. The effects of the drug on the cell cycle were measured based on the nuclear DNA content using flow cytometry. Expression of cell cycle-regulating genes was analyzed using real-time PCR. It was discovered that WP 631, at the tested concentration, did not affect the SKOV-3 cell cycle. Epo B caused significant G2/M arrest, whereas the drug combination induced stronger apoptosis and lower mitotic arrest than Epo B alone. This is very important information from the point of view of the fight against cancer, as, while mitotic arrest in Epo B-treated cells could be overcame after DNA damage repair, apoptosis which occurs after mitotic slippage in combination-treated cells is irreversible. It clearly explains the higher activity of the drug combination in comparison to Epo B alone. Epo B acts via the CDK1/cyclin B complex and has the ability to inhibit CDK1, which may be a promising strategy for ovarian cancer treatment in the future. The drug combination diminishes EpCAM and HMGB1 expression to a greater degree than either WP 631 and Epo B alone. Owing to the fact that the high expression of these two proteins is a poor prognostic factor for ovarian cancer, a decrease in their expression, observed in our studies, may result in improved efficacy of cancer therapy. The presented findings show that the combination of WP 631 and Epo B is a better therapeutic option than either of these drugs alone.

• 한국환경성돌연변이발암원학회지
• /
• 제21권2호
• /
• pp.82-88
• /
• 2001

Recently, there has been significant progress in understanding the control process of the cell division cycle. To investigate the influence of toxic substances on the cell cycle, the effect of benzo(a)pyrene (BAP) and mitomycine C (MMC) on synchronized HeLa cells was analyzed during the cell cycle. To synchronize the HeLa cells, 10$^{6}$ cells were grown for 1 day and then treated with 1 mM hydroxyurea for 14 h. The arrested cells were then allowed to proceed through their cell cycle by removing the hydroxyurea and resupplying a fresh medium. The arrested cells in the G1/S transition then proceeded to the S phase after 4 h, the G2/M phase after 8h, and the G1 phase after 12 h, subsequent to the resupply of a fresh medium. In the untreated HeLa cells, the p34$^{cdc2}$ kinase activity, measured using a p34$^{cdc2}$ specific peptide, peaked after 8h (G2/M) and then declined after 12 h (G1). However, treatment with 30 $\mu$M BAP delayed the peak of the p34$^{cdc2}$ kinase activity. The amount of p34$^{cdc2}$ remained unchanged in the untreated, BAP-, and MMC-treated cells throughout the cell cycle. The cyclin B level peaked after 8 h in the untreated cells, yet peaked after 10-12 h in the BAP-treated cells. There was no significant change in the cyclin B level in the MMC-treated cells.

• Animal Bioscience
• /
• 제35권7호
• /
• pp.1010-1020
• /
• 2022

Objective: The experiment was conducted to evaluate the effects of maternal undernutrition during late pregnancy on the expressions of genes involved in growth and development in ovine fetal perirenal brown adipose tissue (BAT). Methods: Eighteen ewes with singleton fetuses were allocated to three groups at day 90 of pregnancy: restricted group 1 (RG1, 0.33 MJ metabolisable energy [ME]/kg body weight [BW]^0.75/d, n = 6), restricted group 2 (RG2, 0.18 MJ ME/kg BW^0.75/d, n = 6), and a control group (CG, ad libitum, 0.67 MJ ME/kg BW^0.75/d, n = 6). The fetuses were removed at day 140 of pregnancy. All data were analyzed by using the analysis of variance procedure. Results: The perirenal fat weight (p = 0.0077) and perirenal fat growth rate (p = 0.0074) were reduced in RG2 compared to CG. In fetal perirenal BAT, the protein level of uncoupling protein 1 (UCP1) (p = 0.0001) was lower in RG1 and RG2 compared with CG and UCP1 mRNA expression (p = 0.0265) was decreased in RG2. The protein level of myogenic factor 5 (Myf5) was also decreased in RG2 (p = 0.0001). In addition, mRNA expressions of CyclinA (p = 0.0109), CyclinB (p = 0.0019), CyclinD (p = 0.0015), cyclin-dependent kinase 1 (CDK1) (p = 0.0001), E2F transcription factor 1 (E2F1) (p = 0.0323), E2F4 (p = 0.0101), and E2F5 (p = 0.0018) were lower in RG1 and RG2. There were decreased protein expression of peroxisome proliferator-activated receptor-γ (PPARγ) (p = 0.0043) and mRNA expression of CCAAT/enhancer-binding protein-α (C/EBPα) (p = 0.0307) in RG2 and decreased PPARγ mRNA expression (p = 0.0008) and C/EBPα protein expression (p = 0.0015) in both RG2 and RG1. Furthermore, mRNA expression of bone morphogenetic protein 4 (BMP4) (p = 0.0083) and BMP7 (p = 0.0330) decreased in RG2 and peroxisome proliferator-activated receptor co-activator-1α (PGC-1α) reduced in RG2 and RG1. Conclusion: Our observations support that repression of regulatory factors promoting differentiation and development results in the inhibition of BAT maturation in fetal perirenal fat during late pregnancy with maternal undernutrition.

• 한방안이비인후피부과학회지
• /
• 제32권4호
• /
• pp.1-12
• /
• 2019

Objectives : The purpose of this study was to identify the anticancer effect of biological substances of ethylacetate(EtOAc) fraction from Orostachys japonicus(OJEF), their effect on human melanoma A375 cells and the related molecular mechanisms. Methods : The MTS assay was used to confirm the inhibition of cancer cell proliferation in A375 cells. And the $MUSE^{TM}$ analyzer was used to determine the ability of OJEF to induce cell cycle arrest. Western blotting was used to determine the changes in protein expression in A375 cells after treatment with OJEF. Results : OJEF showed cytotoxicity to A375 cells. And cell cycle arrest occurred in G1 phase and G2/M phase owing to inhibition of CDK1, cyclin B1, CDK4, and cyclin D, which are related to cell cycle regulation and cell division control. Conclusion : OJEF is effective in regulating cell cycle of human melanoma cells and thus can be a good theraputic agent to treat patients with melanoma.

• 대한한의학방제학회지
• /
• 제15권1호
• /
• pp.213-228
• /
• 2007

Objectives : The purpose of this report was to investigate the chemotherapeutic effect of Bujeonghangamtang against cancer cells. Materials and Methods : Various cancer cell lines including PANC-1, C6 glioma, SH-SY5Y, HepG2, and MCF-7 cells, were used. Apoptosis was determined by DAPI nuclei staining and flow cytometry in PANC-1 cells treated with 1 mg/ml Bujeonghangamtang for 48 hr. Expression of cell cycle arrest mediators including, cdc2p34 and cyclin B1 proteins were measured by Western blot analysis. Mitochondrial membrane potential was measured by fluorescence staining with JC-1, rhodamine 123. Result : Bujeonghangamtang induced the apoptosis of PANC-1, which was characterized as nucleic acid and genomic DNA fragmentation, chromatin condensation, and sub-G0/G1 fraction of cell cycle increase. but not C6 glioma, SH-SY5Y, HepG2, and MCF-7 cells. PANC-1 cells were markedly sensitive to Bujeonghangamtang. Treatment with Bujeonghangamtang resulted in the decreased expression of cdc2p34 and cyclin B1. Treatment with Bujeonghangamtang also increased the ROS production and induced mitochondrial dysfunction. Conclusion : Bujeonghangamtang exerted cytotoxicity against human Pancreatic cancer cells via cell cycle arrest-mediated apoptotic signaling including ROS production and mitochondrial dysfunction. Our data suggest that Bujeonghangamtang may be an important modulator of chemosensitivity of cancer cells against anticancer chemotherapeutic agents.

• Asian Pacific Journal of Cancer Prevention
• /
• 제15권22호
• /
• pp.9835-9839
• /
• 2014

Fisetin is an effective compound extracted from lacquer which has been used in the treatment of various diseases. Preliminary data indicate that it also exerts specific anti-cancer effects. However, the manner in which fisetin regulates cancer growth remains unknown. In this study, we elucidated interference of fisetin with targets of the nuclear factor ${\kappa}B$ signal transduction pathway activated by Epstein-Barr virus encoding latent membrane protein 1 (LMP1)in nasopharyngeal carcinoma (NPC) cells, Results showed that fisetin inhibited the survival rate of CNE-LMP1 cells and NF-${\kappa}B$ activation caused by LMP1. Fisetin also suppressed nuclear translocation of NF-${\kappa}B$ (p65) and $I{\kappa}B{\alpha}$ phosphorylation, while inhibiting CyclinD1, all key targets of the NF-${\kappa}B$ signal transduction pathway. It was suggested that interference effects of fisetin with signal transduction activated by LMP1 encoded by the Epstein-Barr virus may play an important role in its anticancer potential.

• 대한임상검사과학회지
• /
• 제38권1호
• /
• pp.72-81
• /
• 2006

Wee1 is a kinase regulator of the M-phase promoting factor (MPF; a complex of cdc2 and cyclin B1). The present study was undertaken to determine the role(s) of wee1 in the early stages of mouse ovarian follicles. The expression of wee1 and the correlated cell-cycle components, namely cdc2, cyclin B1, and cdc25C, were evaluated by immunohistochemistry. In addition, the expression of Tyr15-phosphorylated cdc2 (cdc2-p) was also examined to determine whether wee1 kinase phosphorylates cdc2 existed. Each component except cdc25C was found cytoplasmic in the oocytes at all stages of follicles, while cdc25C was not detected in primordial follicles. It was found primarily in ovarian somatic cells and to a small extent in granulosa cells of the growing follicles. To further confirm the expression of cell-cycle components in the primordial follicular oocytes, day1 ovaries were enzymatically and mechanically dissociated, then oocytes were isolated from somatic including pre-granulosa cells, and we confirmed that cdc2-p was expressed in oocytes of primordial follicles. From the results of the present study, we concluded wee1, without the counteracting cdc25C, would cause meiotic arrest of oocytes by the inhibitory phosphorylation of cdc2. The expression of all these proteins in the granulosa cells of growing follicles may regulate their mitosis concurrently with the growth of oocytes and follicles.

• International Journal of Oral Biology
• /
• 제44권2호
• /
• pp.43-49
• /
• 2019

Tannic acid (TA) is a water-soluble polyphenol compound found in various herbal plants. We investigated the chemopreventive effects of TA on FaDu hypopharyngeal squamous carcinoma cells. In an 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, TA showed dose-dependent cytotoxicity with a half maximal inhibitory concentration (IC50) of 50 ?M. Cell cycle analysis and immunofluorescence imaging demonstrated that under low-dose ($25{\mu}M$) treatment, FaDu cells were arrested in G2/M phase, and as the dose of TA was increased, apoptosis was induced with the increase of cell population at sub-G1 phase. The expressions of various cyclins, including cyclin D1 and cyclin-dependent kinases (CDK-1 and CDK-2), were down-regulated at low doses of TA, whereas apoptotic effectors such as cleaved caspase 3, cleaved caspase 7, and poly (ADP-ribose) polymerase (PARP) were expressed in a dose-dependent manner in Western blotting. In addition, TA-induced apoptosis of FaDu cells might be mediated by the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase pathway, with the upregulation of p-AKT/p-PKB (phosphorylated protein kinase B) and p-ERK. Overall, our data support the hypothesis that TA is a potential candidate agent for the treatment of hypopharyngeal cancer.

• 한국미생물·생명공학회지
• /
• 제47권1호
• /
• pp.34-42
• /
• 2019

Apigenin, a common natural product that is found in many plants and vegetables, has been reported to have many biological activities, including antioxidative, anti-inflammatory, and anticancer effects. The triple-negative breast carcinoma cell line MDA-MB-231 is known to be highly invasive and resistant to chemotherapy. In this study, we investigated the anticancer effect of apigenin on human MDA-MB-231 cells. First, the cytotoxicity of apigenin toward MDA-MB-231 cells was analyzed by MTT assay. Then, the cell cycle and apoptotic effects of apigenin were examined, and the molecular mechanism underlying its anticancer activity was explored. Apigenin inhibited the growth of the cells in a dose-dependent manner, correlating with the cell cycle arrest at the G2-M phase as well as an increase of early apoptosis. The cell-cycle inhibitory effect was highly associated with the increased expression of p21 and decreased expression of CDK6, cyclin D1, and cyclin B1. The induction of apoptosis by apigenin was associated with the upregulated expression of cleaved PARP and cleaved caspase-3, -7, and -9.

• Journal of Microbiology and Biotechnology
• /
• 제25권3호
• /
• pp.413-417
• /
• 2015

Recently, we isolated HY253, a novel decahydrofluorene analog with a molecular structure of 7,8a-divinyl-2,4a,4b,5,6,7,8,8a,9,9a-decahydro-1H-fluorene-2,4a,4b,9a-tetraol from the roots of Aralia continentalis, which is known as Dokwhal (獨活), a traditional medicinal herb. Moreover, we previously reported its cytotoxic activity on cancer cell proliferation in human lung cancer A549 and cervical cancer HeLa cells. The current study aimed to evaluate its detailed molecular mechanisms in cell cycle arrest and apoptotic induction in human hepatocellular carcinoma HepG2 cells. Flow cytometric analysis of HepG2 cells treated with $60{\mu}M$ HY253 revealed appreciable cell cycle arrest at the G1 phase via inhibition of Rb phosphorylation and down-regulation of cyclin D1. Furthermore, using western blots, we found that up-regulation of cyclin-dependent kinase inhibitors, such as p21^CIP1 and p27^KIP1, was associated with this G₁ phase arrest. Moreover, TUNEL assay and immunoblottings revealed apoptotic induction in HepG2 cells treated with $60{\mu}M$ HY253 for 24 h, which is associated with cytochrome c release from mitochondria, via down-regulation of anti-apoptotic Bcl-2 protein, which in turn resulted in activation of caspase-9 and -3, and proteolytic cleavage of poly(ADP-ribose) polymerase (PARP). Accordingly, we suggest that HY253 may be a potent chemotherapeutic hit compound for treating human liver cancer cells via up-regulation and activation of the p53 gene.