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    Analysis of tumor necrosis factor alpha-induced and nuclear factor kappa B-silenced LNCaP prostate cancer cells by RT-qPCR
    (Spandidos Publ Ltd, 2014) Gonen-Korkmaz, Ceren; Sevin, Gulnur; Gokce, Goksel; Arun, Mehmet Zuhuri; Yildirim, Gokce; Reel, Buket; Kaymak, Aysegul; Ogut, Deniz
    Prostate cancer is the second leading cause of morbidity and mortality in males in the Western world. In the present study, LNCaP, which is an androgen receptor-positive and androgen-responsive prostate cancer cell line derived from lymph node metastasis, and DU 145, which is an androgen receptor-negative prostate cancer cell line derived from brain metastasis, were investigated. TNF alpha treatment decreased p105 and p50 expression and R1881 treatment slightly decreased p105 expression but increased p50 expression with or without TNF alpha induction. As an aggressive prostate cancer cell line, DU145 transfected with six transmembrane protein of prostate (STAMP)1 or STAMP2 was also exposed to TNF alpha. Western blotting indicated that transfection with either STAMP gene caused a significant increase in NF kappa B expression following TNF alpha induction. In addition, following the treatment of LNCaP cells with TNF alpha, reverse transcription quantitative polymerase chain reaction (RT-qPCR) was performed with a panel of apoptosis-related gene primers. The apoptosis-related genes p53, p73, caspase 7 and caspase 9 showed statistically significant increases in expression levels while the expression levels of MDM2 and STAMP1 decreased following TNF alpha induction. Furthermore, LNCaP cells were transfected with a small interfering NF kappa B (siNF kappa B) construct for 1 and 4 days and induced with TNF alpha for the final 24 h. RT-qPCR amplifications were performed with apoptosis-related gene primers, including p53, caspases and STAMPs. However, no changes in the level of STAMP2 were observed between cells in the presence or absence of TNF alpha induction or between those transfected or not transfected with siNF kappa B; however, the level of STAMP1 was significantly decreased by TNF alpha induction, and significantly increased with siNF kappa B transfection. Silencing of the survival gene NF kappa B caused anti-apoptotic STAMPI expression to increase, which repressed p53, together with MDM2. NF kappa B silencing had varying effects on a panel of cancer regulatory genes. Therefore, the effective inhibition of NF kappa B may be critical in providing a targeted pathway for prostate cancer prevention.
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    Different Responses of RT-PCR Amplifications after Tumor Necrosis alpha (TNFa) Induction at Nuclear Factor kappa B (NFkB) Gene Silenced LNCaP Cells
    (Federation Amer Soc Exp Biol, 2013) Gonen-Korkmaz, Ceren; Sevin, Gulnur; Gokce, Goksel; Arun, Mehmet Z.; Yetik-Anacak, Gunay; Yildirim, Gokce; Reel, Buket; Ogut, Deniz; Kaymak, Aysegul
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    Introducing Basic Molecular Biology to Turkish Rural and Urban Primary School Children Via Hands-on PCR and Gel Electrophoresis Activities
    (Wiley-Blackwell, 2014) Selli, Cigdem; Yildirim, Gokce; Kaymak, Aysegul; Karacicek, Bilge; Ogut, Deniz; Gungor, Turkan; Erem, Erdem; Ege, Mehmet; Bumen, Nilay; Tosun, Metiner
    This study includes the results of a 2-day education project titled Molecular Biology Laboratory Summer School, MoBiLYO. The project was held at a University Research Center by scientists from Department of Pharmacology and graduate students. The project was composed of introductory lectures, model construction, DNA isolation, polymerase chain reaction (PCR), and gel electrophoresis. The participants were 13-year-old eighth-graders attending primary schools affiliated with Ministry of National Education in urban and rural areas of Izmir, Turkey. The purpose of this study was to introduce basic molecular biology concepts through individually performed experiments such as PCR and gel electrophoresis integrated with creative drama. The students were assessed at the beginning and the end of each project day via mini-tests, experimental and presentation skills evaluation forms. Data showed that students' knowledge about DNA structure and basic molecular biology techniques significantly increased. On the basis of experimental and presentational skills, there was no significant difference between kids from urban and rural schools or between public and boarding public schools, whereas the average score of girls was significantly higher than that of boys. In conclusion, individually performed experiments integrated with creative drama significantly increased students' perception of complex experimental procedures on basic molecular biology concepts. Data suggests that integration of these concepts into the science and technology curriculum of Turkish primary education may support the recruitment of future scientists who can handle rapidly developing genomic techniques that will affect our everyday life. (c) 2014 by The International Union of Biochemistry and Molecular Biology, 42(2):114-120, 2014
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    The Regulation of Matrix Metalloproteinase Expression and the Role of Discoidin Domain Receptor 1/2 Signalling in Zoledronate-treated PC3 Cells
    (Ivyspring Int Publ, 2015) Reel, Buket; Korkmaz, Ceren Gonen; Arun, Mehmet Zuhuri; Yildirim, Gokce; Ogut, Deniz; Kaymak, Aysegul; Micili, Serap Cilaker; Ergur, Bekir Ugur
    Discoidin Domain Receptors (DDR1/DDR2) are tyrosine kinase receptors which are activated by collagen. DDR signalling regulates cell migration, proliferation, apoptosis and matrix metalloproteinase (MMP) production. MMPs degrade extracellular matrix (ECM) and play essential role in tumor growth, invasion and metastasis. Nitrogen-containing bisphosphonates (N-BPs) which strongly inhibit osteoclastic activity are commonly used for osteoporosis treatment. They also have MMP inhibitory effect. In this study, we aimed to investigate the effects of zoledronate in PC3 cells and the possible role of DDR signalling and downstream pathways in these inhibitory effects. We studied messenger RNA (mRNA) and protein expressions of MMP-2,-9,-8, DDR1/DDR2 type I procollagen (TIP) and mRNA levels of PCA-1, MMP-13 and DDR-initiated signalling pathway players including K-Ras oncogene, ERK1, JNK1, p38, AKT-1 and BCLX in PC3 cells in the presence or absence of zoledronate (10-100 mu M) for 2-3 days. Zoledronate (100 mu M) down-regulated DDR1/DDR2, TIP mRNAs but did not change MMP-13 (collagenase-3) mRNA. However, zoledronate up-regulated MMP-8 (collagenase-2) mRNA. Zoledronate also inhibited mRNA expressions of K-Ras, ERK1, AKT-1, BCLX and PCA-1; but did not change JNK1, p38 mRNA levels. Zoledronate (100 mu M) supressed DDR1/DDR2, TIP expressions; and gelatinase (MMP-2/MMP-9) expressions/activities. Conversely, zoledronate up-regulated MMP-8 expression in PC3 cells. Zoledronate down-regulates MMP-2/-9 expressions in PC3 prostate cancer cells. DDR1/DDR2 signalling and DDR-initiated downstream Ras/Raf/ERK and PI3K/AKT pathways may at least partially responsible for MMP inhibitory effect of zoledronate.