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    Analysis of tumor necrosis factor ?-induced and nuclear factor ?B-silenced LNCap prostate cancer cells by RT-qPCR
    (Spandidos Publications, 2014) Gonen-Korkmaz C.; Sevin G.; Gokce G.; Arun M.Z.; Yildirim G.; Reel B.; Kaymak A.; Ogut D.
    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 DU145, which is an androgen receptor-negative prostate cancer cell line derived from brain metastasis, were investigated. TNF? treatment decreased p105 and p50 expression and R1881 treatment slightly decreased p105 expression but increased p50 expression with or without TNF? 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?. Western blotting indicated that transfection with either STAMP gene caused a significant increase in NF?B expression following TNF? induction. In addition, following the treatment of LNCaP cells with TNF?, 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? induction. Furthermore, LNCaP cells were transfected with a small interfering NF?B (siNF?B) construct for 1 and 4 days and induced with TNF? 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? induction or between those transfected or not transfected with siNF?B; however, the level of STAMP1 was significantly decreased by TNF? induction, and significantly increased with siNF?B transfection. Silencing of the survival gene NF?B caused anti-apoptotic STAMP1 expression to increase, which repressed p53, together with MDM2. NF?B silencing had varying effects on a panel of cancer regulatory genes. Therefore, the effective inhibition of NF?B may be critical in providing a targeted pathway for prostate cancer prevention. © 2014, Spandidos Publications. All right reserved.
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    Doxycycline down-regulates matrix metalloproteinase expression and inhibits NF-?b signalling in LPS-induced PC3 cells
    (Via Medica, 2016) Ogut D.; Reel B.; Korkmaz C.G.; Arun M.Z.; Micili S.C.; Ergur B.U.
    Introduction. Matrix metalloproteinase enzymes (MMPs) play important role in inflammation, malignant cell proliferation, invasion and angiogenesis by mediating extracellular matrix degradation. Doxycycline, a synthetic tetracycline, behaves as a MMP inhibitor at a subantimicrobial dose and inhibits tumor cell proliferation, invasion and angiogenesis. The aberrant activity of nuclear factor kappa B (NF-?B) causes activation of MMPs and thereby proliferation and invasion of cancer cells. The aim of this study was to investigate the effects of doxycycline on the expression of MMPs in lipopolysaccharide (LPS)-induced PC3 human prostate cancer cells and the possible role of NF-?B signaling. Material and methods. PC3 cells were incubated with LPS (0.5 µg/mL) for 24 h in the presence or absence of doxycycline (5 µg/mL). The effects of LPS and doxycycline on the expressions of MMP-2, MMP-8, MMP-9, MMP-10, NF-?B/p65, I?B-?, p-I?B-?, IKK-ß were examined by Western blotting and immunohistochemistry in PC3 cells. Furthermore, relative proteinase activities of MMP-2 and MMP-9 were determined by gelatin zymography. Results. LPS increased expression and activity of MMP-9 and expression of MMP-8, MMP-10, NF-?B/p65, p-I?B-?, IKK-ß and doxycycline down-regulated its effects with the exception of MMP-10 expression. The expression of MMP-2 and I?B-a was affected by neither LPS nor doxycycline. Conclusions. Our findings indicate that doxycycline inhibits the expression of various MMPs and NF-?B signaling may play a role in the regulation of MMPs expression in LPS-induced PC3 human prostate cancer cells. ©Polish Society for Histochemistry and Cytochemistry.