• BMB Reports
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• v.44 no.8
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• pp.553-557
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• 2011

We previously reported that CDK2/Cyclin A can phosphorylate and activate the transcription factor NF-Y. In this study, we investigated a potential regulatory role for NF-Y in the transcription of Cyclin A and other cell cycle regulatory genes. Gel-shift assays demonstrate that NF-Y binds to CCAAT sequences in the Cyclin A promoter, as well as to those in the promoters of cell cycle G2 regulators such as CDC2, Cyclin B and CDC25C. Furthermore, expression of Cyclin A increases NF-Y's affinity for CCAAT sequences in the CDC2 promoter; however, Cyclin A's induction of CDC2 transcription is antagonized by p21, an inhibitor of CDK2/Cyclin A. These results suggest a model wherein NF-Y binds to and activates transcription from the Cyclin A promoter, increasing cellular levels of Cyclin A/CDK2 and potentiating NF-Y's capacity for transcriptional transactivation, and imply a positive feedback loop between NF-Y and Cyclin A/CDK2. Our findings are additionally indicative of a role for Cyclin A in activating Cyclin B/CDK1 through promoting NF-Y dependent transcription of Cyclin B and CDC2; NF-Y mediated crosstalk may therefore help to orchestrate cell-cycle progression.

• Pediatric Gastroenterology, Hepatology & Nutrition
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• v.15 no.1
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• pp.29-37
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• 2012

Purpose: To examine the prevalence of $Clostridium$ $difficile$ ($C.$ $difficile$) colonization (CDC) and potential neonatal determinants of CDC in hospitalized preterm infants. Methods: Fecal samples were serially collected within 72 h after birth and at 1, 2, and 4-6 weeks of age from preterm infants in the neonatal intensive care units (NICUs) of two different university hospitals. Total bacterial DNA was extracted from each fecal sample from 49 infants, and polymerase chain reaction (PCR) was performed with primers for the 16S gene of $C.$ $difficile$ and the toxin A and toxin B genes. The correlation between the results of $C.$ $difficile$ PCR assays and the clinical characteristics of the infants was analyzed. Results: The prevalence rates of CDC were 34.7, 37.2, 41.3, and 53.1% within 72 h after birth and at 1, 2, and 4.6 weeks of age, respectively. The toxin positivity rate was significantly higher in the infants with persistent CDC than in those with transient CDC (8/12 [66.7%] vs. 6/25 [24.5%] ($p$=0.001). Among the various neonatal factors, only the feeding method during the first week after birth was significantly associated with persistent CDC. Exclusive breast-milk feeding (EBMF) significantly decreased the risk of persistent CDC compared to formula or mixed feeding (adjusted odds ratio: 0.133, 95% confidence interval: 0.02-0.898, $p$=0.038). Conclusion: The prevalence of CDC increased with the duration of hospitalization in preterm infants in the NICU. EBMF during the first week after birth in hospitalized preterm infants may protect against persistent CDC.

• Journal of Life Science
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• v.23 no.6
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• pp.832-837
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• 2013

In this study, it was investigated the possible mechanisms by which glutamine deprivation exerts its anti-proliferative action in cultured human prostate carcinoma PC3 cells. Glutamine deprivation resulted in inhibition of growth and G2/M arrest of the cell cycle in a time-dependent manner without apoptosis induction, as determined by MTT assay, DAPI staining and flow cytometry analyses. The induction of G2/M arrest by glutamine deprivation was associated with the inhibition of expression of Cdc2, cyclin A and cyclin B1, and up-regulation of the expression of cyclin-dependent kinase (Cdk) inhibitor p21(WAF1/CIP1) in both transcriptional and translational levels. Moreover, glutamine deprivation increased the phosphorylation of checkpoint kinase (Chk)1 and Chk2; however, the levels of Cdc25C phosphorylation were decreased in response to glutamine deprivation in a time-dependent manner. Our data provide a first biochemical evidence that glutamine deprivation suppresses cell viability through G2/M phase arrest without induction of apoptosis in PC3 cells.

• BMB Reports
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• v.51 no.7
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• pp.350-355
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• 2018

HnRNPK is a multifunctional protein that participates in chromatin remodeling, transcription, RNA splicing, mRNA stability and translation. Here, we uncovered the function of hnRNPK in regulating the proliferation and differentiation of myoblasts. hnRNPK was mutated in the C2C12 myoblast cell line using the CRISPR/Cas9 system. A decreased proliferation rate was observed in hnRNPK-mutated cells, suggesting an impaired proliferation phenotype. Furthermore, increased G2/M phase, decreased S phase and increased sub-G1 phase cells were detected in the hnRNPK-mutated cell lines. The expression analysis of key cell cycle regulators indicated mRNA of Cyclin A2 was significantly increased in the mutant myoblasts compared to the control cells, while Cyclin B1, Cdc25b and Cdc25c were decreased sharply. In addition to the myoblast proliferation defect, the mutant cells exhibited defect in myotube formation. The myotube formation marker, myosin heavy chain (MHC), was decreased sharply in hnRNPK-mutated cells compared to control myoblasts during differentiation. The deficiency in hnRNPK also resulted in the repression of Myog expression, a key myogenic regulator during differentiation. Together, our data demonstrate that hnRNPK is required for myoblast proliferation and differentiation and may be an essential regulator of myoblast function.

• Environmental Mutagens and Carcinogens
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• v.25 no.2
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• pp.51-59
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• 2005

Overexpression of HSP25 delayed cell growth, increased the level of $p21^{waf}$, reduced the levels of cyclin D1, cylcin A and cdc2, and induced radioresistance in L929 cells. We demonstrated that extracellular regulated kinase (ERK) and MAP kinase/ERK kinase (MEK) expressions as well as their activation (phospho-forms) were inhibited by hsp25 overexpression. To confirm the relationship between ERK1/2 and hsp25-mediated radioresistance, ERK1 or ERK2 cDNA was transiently transfected into the hsp25 overexpressed cells and their radioresistance was examined. HSP25-mediated radioresistance was abolished by overexpression of ERK2, but not by overexpression of ERK1. Alteration of cell cycle distribution and cell cycle related protein expressions (cyclin D, cyclin A and cdc2) by hsp25 overexpression were also recovered by ERK2 cDNA transfection. Increase in Bc1-2 protein by hsp25 gene transfection was also reduced by subsequent ERK2 cDNA-transfection. In addition, HSP25 overexpression reduced reactive oxygen species (ROS) and increased expression of manganese superoxide dismutase (MnSOD) gene. Increased activation of NF-kB (IkB degradation) was also found in hsp25-overexpressed cells. Moreover, transfection of hsp25 antisense gene abrogated all the HSP25-mediated phenomena. To further elucidate the exact relationship between MnSOD induction and NF-kB activation, dominant negative $I-kB\alpha(I-kB\alpha-DN)$ construction was transfected to HSP25 overexpressed cells. $I-kB\alpha-DN$ inhibited HSP25 mediated MnSOD gene expression. In addition, HSP25 mediated radioresistance was blocked by $I-kB\alpha-DN$ transfection. Blockage of MnSOD with antisense oligonucleotides in HSP25 overexpressed cells, prevented apoptosis and returned the ERK1/2 activation to the control level. From the above results, we suggest for the first time that reduced oxidative damage by HSP25 was due to MnSOD-mediated down regulation of ERK1/2.

• Microbiology and Biotechnology Letters
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• v.41 no.4
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• pp.425-432
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• 2013

Typha orientalis, also known as bulrush or cattail, is a perennial herbaceous plant found in freshwater wetlands and has been widely used in constructed wetlands for wastewater treatment. Recent data has revealed that SH21B, a mixture composed of seven herbs including T. orientalis, exhibited an anti-adipogenic activity by the inhibition of the expression of adipogenic regulators. However, the anti-cancer effect of T. orientalis and its molecular mechanisms remain unclear. In this study, we evaluated the anti-cancer effect and its mechanism in the methanol extract of T. orientalis (METO) on human colon carcinoma HT29 cells. It was found that METO treatment showed cytotoxic activity in a dose-dependent manner, and induced G2/M cell cycle arrest and apoptosis in HT29 cells. The induction of G2/M arrest by METO was associated with the up-regulation of phospho-Cdc2 (Tyr15), an inactive form of Cdc2 and the down-regulation of Cdc25c phosphatase. METO also induced tumor suppressor p53 and cyclin-dependent kinase inhibitor p21 (WAF1/CIP1) expression. In addition, METO-induced apoptosis was characterized by the proteolytic activation of caspase-3, degradation of poly ADP ribose polymerase (PARP), and up-regulation of death receptor FAS and pro-apoptotic Bax expression. Collectively, these results indicate that the cell cycle inhibition and apoptosis induction of METO in HT29 cells allows for the possibility of its use in anti-cancer therapies.

• Journal of the Korean Society of Food Science and Nutrition
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• v.33 no.8
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• pp.1237-1245
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• 2004

Agaricus blazei Murill is a medicinal mushroom native to Brazil. It used to be a source of antitumor and immunoactive compounds and considered a health food in many countries. In the present study, it was examined the effects of water extract of A. blazei (WEAB) on the growth of human lung carcinoma cell line A549 in order to investigate the anti-proliferative mechanism by WEAB. Treatment of A549 cells to WEAB resulted in the growth inhibition, morphological change and induction of apoptotic cell death in a dose-dependent manner as measured by MTT assay and flow cytometric analysis. Flow cytometric analysis revealed that WEAB caused G2/M phase arrest of the cell cycle, which was associated with a down-regulation of cyclin A in both transcriptional and translational levels. WEAB treatment induced a marked up-regulation of cyclin-dependent kinase (Cdk) inhibitor p21, however, the levels of Cdk2, Cdc2, Wee1, Cdc25C and p53 expression were remained unchanged in WEAB treated cells. In addition, WEAB treatment inhibited the levels of cyclooxygenase (COX)-2 mRNA and protein without alteration of COX-l expression. Taken together, these findings suggest that WEAB may be a potential chemotherapeutic agent for the control of human lung carcinorma cells and further studies will be needed to identify the active compounds that confer the anti-cancer activity of WEAB. Once such compounds are identified, the mechanisms by which they exert their effects can begin to be characterized.

• Radiation Oncology Journal
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• v.21 no.4
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• pp.306-314
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• 2003

Purpose : In our Previous study, we have shown the main cel1 death pattern Induced by irradiation or protein tyrosine kinase (PTK) inhibitors in K562 human myeiogenous leukemic cell line. Death of the cells treated with irradiation alone was characterized by mitotic catastrophe and typical radiation-induced apoptosis was accelerated by herblmycin A (HMA). Both types of cell death were inhibited by genistein. In this study, we investigated the effects of HMA and genistein on cell cycle regulation and its correlation with the alterations of radiation-induced cell death. Materials and Methods: K562 cells In exponential growth phase were used for this study. The cells were Irradiated with 10 Gy using 6 MeV Linac (200-300 cGy/min). Immediately after irradiation, cells were treated with 250 nM of HMA or 25 $\mu$N of genistein. The distributions of cell cycle, the expressions of cell cycle-related protein, the activities of cyclin-dependent kinase, and the yield of senescence and differentiation were analyzed. Results: X-irradiated cells were arrested In the G2 phase of the cell cycle but unlike the p53-positive cells, they were not able to sustain the cell cycle arrest. An accumulation of cells in G2 phase of first ceil-cycle post-treatment and an increase of cyclin Bl were correlated with spontaneous, premature, chromosome condensation and mitotic catastrophe. HMA induced rapid G2 checkpoint abrogation and concomitant p53-independent Gl accumulation. HMA-induced cell cycle modifications correlated with the increase of CDK2 kinase activity, the decrease of the expressions of cyclins I and A and of CDK2 kinase activity, and the enhancement of radiation-induced apoptosis. Genistein maintained cells that were arrested in the G2-phase, decreased the expressions of cyclin Bl and cdc25c and cdc25C kinase activity, increased the expression of pl6, and sustained senescence and megakaryocytic differentiation. Conclusion: The effects of HMA and genistein on the radiation-induced cell death of KS62 cells were closely related to the cell cycle regulatory activities. In this study, we present a unique and reproducible model in which for investigating the mechanisms of various, radiation-induced, cancer cell death patterns. Further evaluation by using this model will provide a potent target for a new strategy of radiotherapy.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.11
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• pp.6419-6425
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• 2013

Background: Neoadjuvant chemotherapy (NACT) is a treatment modality whereby chemotherapy is used as the initial treatment of HNSCC in patients presenting with advanced cancer that cannot be treated by other means. It leads to shrinkage of tumours to an operable size without significant compromise to essential oro-facial organs of the patients. The molecular mechanisms behind shrinkage due to NACT is not well elucidated. Materials and Methods: Eleven pairs of primary HNSCCs and adjacent normal epithelium, before and after chemotherapy were screened for cell proliferation and apoptosis. This was followed by immunohistochemical analysis of some cell cycle (LIMD1, RBSP3, CDC25A, CCND1, cMYC, RB, pRB), DNA repair (MLH1, p53) and apoptosis (BAX, BCL2) associated proteins in the same set of samples. Results: Significant decrease in proliferation index and increase in apoptotic index was observed in post-therapy tumors compared to pre-therapy. Increase in the RB/pRB ratio, along with higher expression of RBSP3 and LIMD1 and lower expression of cMYC were observed in post-therapy tumours, while CCND1 and CDC25A remained unchanged. While MLH1 remained unchanged, p53 showed higher expression in post-therapy tumors, indicating inhibition of cell proliferation and induction of apoptosis. Increase in the BAX/BCL2 ratio was observed in post-therapy tumours, indicating up-regulation of apoptosis in response to therapy. Conclusions: Thus, modulation of the G1/S cell cycle regulatory proteins and apoptosis associated proteins might play an important role in tumour shrinkage due to NACT.

• Tribology and Lubricants
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• v.25 no.1
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• pp.20-24
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• 2009

TiC-derived carbon coatings have been synthesized at $600^{\circ}C$ temperature treatment with $H_2/Cl_2$ mixture gases. From Raman spectroscopy measurements, the modified layer was covered with carbon and the thick-ness of the layer was increased with increasing reaction time. And $I_D/I_G$ ratio was decreased with increasing reaction time. The superior tribological property was obtained from TiC reacted with $Cl_2$ gas for 2 hrs. And the tribological property measurements indicate that TiC-derived carbon layer has $0.9{\times}10_{-6}mm^3/Nm$ in wear coefficient and 0.13 in friction coefficient.