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Öğe A disintegrin-like metalloproteinase with thrombospondin motif 8 expression analysis in OUMS-27 chondrosarcoma cells before and after insulin administration(Natl Inst Science Communication-Niscair, 2016) Erden, Gonul; Akyol, Sumeyya; Comertoglu, Ismail; Altuntas, Aynur; Cakmak, Ozlem; Ugurcu, Veli; Yukselten, YunusA disintegrin-like and metalloproteinase with thrombospondin motif 8 (ADAMTS8) is a secreted protease with anti-angiogenic properties. It inhibits vascular endothelial growth factor (VEGF) induced angiogenesis and suppresses fibroblast growth factor 2 (FGF-2) induced vascularization. Angiogenesis and extracellular matrix degradation are the key events in tumor progression, and ADAMTS8 is also known to be a member of the aggrecanases family. In the present study, we investigated the expression levels of ADAMTS8 in chondrosarcoma cells to elucidate the effect of insulin on the tumor cells in terms of ADAMTS production. The OUMS-27 cells were cultured and separately exposed to 10 mu mol/mL insulin up to 11 days in Dulbecco's modified Eagle's medium. After specific time limits (days 1, 3, 7, and 11), the culture was terminated and RNA was isolated using TRIzol reagent and converted to cDNA. The expression levels of ADAMTS8 were evaluated by qRT-PCR. The ADAMTS8 expression in OUMS-27 cells exhibited about 4-fold decrease following insulin treatment on day 11. Statistically significant differences were noted between the control and day 1 (P = 0.008), day 7 (P = 0.047) and day 11 (P = 0.008) groups. The effect of insulin on chondrosarcoma cells in terms of ADAMTS8 expression has not been reported earlier. The decrease in ADAMTS8 expression could be considered as a novel finding that has the potential to explain some pathophysiological mechanisms of tumor cells. Furthermore, the finding could also shed some light on the relationships between matrix degradation and insulin treatment in vitro.Öğe ADAMTS12 Depletion by Insulin in OUMS-27 Human Chondrosarcoma Cells(Turkish League Against Rheumatism, 2015) Altuntas, Aynur; Akyol, Sumeyya; Adam, Bahattin; Cakmak, Ozlem; Ugurcu, Veli; Erden, Gonul; Yukselten, YunusObjectives: In this study, we aim to investigate the association between articular damage in diabetes and a disintegrin-like and metalloproteinase with thrombospondin motifs 12 (ADAMTS12) at gene expression and protein levels. Materials and methods: OUMS-27 human chondrosarcoma cells were used to investigate how ADAMTS12 levels changed in vitro condition in presence and absence of insulin. The study included three groups of cells treated with 10 mu g/mL of insulin, and a control group. Cells were incubated with insulin in medium for one day, three days, and seven days. The effects of insulin on ADAMTS12 were investigated at both gene expression and protein levels. The relationships between the variables were tested by Mann-Whitney U test. Results: ADAMTS12 expression was significantly lower in the groups treated with insulin medium for one day and seven day periods (p=0.008 and p=0.008, respectively) compared to the control group. No significant difference was detected in the expression level between the groups kept in insulin medium for three days and the control group (p=0.55). In addition, protein amounts of the groups exposed to insulin medium for one, three, and seven day periods were lower. Conclusion: Insulin reduces the amount of ADAMTS12 which causes delayed recovery of cartilage tissue in the OUMS-27 cell lines utilized in our study for their chondrocytic properties. This reduction due to insulin treatment may contribute to recovery of cartilage tissue.Öğe Effect of insulin on the mRNA expression of procollagen N-proteinases in chondrosarcoma OUMS-27 cells(Spandidos Publ Ltd, 2015) Akyol, Sumeyya; Comertoglu, Ismail; Firat, Ridvan; Cakmak, Ozlem; Yukselten, Yunus; Erden, Gonul; Ugurcu, VeliChondrosarcoma is one of the most common bone tumors, and at present, there is no non-invasive treatment option for this cancer. The chondrosarcoma OUMS-27 cell line produces proteoglycan and type II, IX, and XI collagens, which constitutes cartilage tissue. A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) proteases are a group of secreted proteases, which include the procollagen N-proteinases ADAMTS-2, -3 and -14. These procollagen N-proteinases perform a role in the processing of procollagens to collagen and the maturation of type I collagen. The present study aimed to improve the understanding of the causes of metastasis, local invasion and resistance to chemo- and radiotherapy in chondrosarcoma, as well as the effect of insulin on cancer cells. The present study was designed to reveal the effects of insulin on procollagen N-proteinases in chondrosarcoma OUMS-27 cells. The cells were cultured in Dulbecco's modified Eagle's medium (DMEM) alone or in DMEM containing 10 mu g/ml insulin. The medium was changed every other day for 11 days. The cells were harvested on days 1, 3, 7 and 11, and total RNA isolation was performed immediately following harvesting. The expression levels of ADAMTS2, ADAMTS3 and ADAMTS14 mRNA were estimated by reverse transcription-quantitative polymerase chain reaction using appropriate primers. ADAMTS2 mRNA expression was found to be decreased on day 7 (P=0.028) and increased at day 11 compared with the control group (P=0.016). The increase in mRNA concentration at day 11 was significantly different compared to the concentrations on days 3 (P=0.047) and 7 (P=0.008). The expression of ADAMTS3 mRNA decreased immediately subsequent to insulin induction on day 1 compared with the control group (P=0.008). The most evident decrease in mRNA concentration was seen at day 7 subsequent to insulin induction (P=0.008). The present results demonstrated that ADAMTS2 and ADAMTS3 may perform a role in the invasion and metastasis of tumors, and may also possess proteolytic activity that results in the breakdown of the extracellular matrix (ECM). Insulin itself can modulate the biosynthesis of ECM macromolecules that are altered in diabetes through various pathways.Öğe Evidence for the Control of Aggrecanases by Insulin and Glucose in Alzheimer's Disease(Kure Iletisim Grubu A S, 2014) Akyol, Sumeyya; Ugurcu, Veil; Cakmak, Ozlem; Altuntas, Aynur; Yukselten, Yunus; Akyol, Omer; Sunguroglu, AsumanObjective: Alzheimer's disease (AD) is a progressive and irreversible central nervous system disease, which slowly destroys cognitive skills and memory, and eventually even the ability to handle the simplest tasks. The initiation and progression of AD is a poorly understood complex process. Here, we have investigated possible biological mechanisms that could be responsible for the increased risk for diminished brain function associated with diabetes in AD. Method: The U87 cell line (human primary glioblastoma cell line) was cultured in Dulbecco's modified Eagle's medium. Cells were incubated with insulin (10 mu g/ml), low glucose (11 mM, 2 mg/ml) and high glucose (55 mM, 10 mg/ml) for 48 hours. Cells were harvested and protein isolations were performed. Primary anti-ADAMTS5, anti-IL-33, anti-NF kappa B, and anti-GAPDH antibodies were used to detect corresponding proteins and to measure band densities in Western membranes using a specific program. Results: Western blot analysis showed ADAMTS5 protein decreases in insulin-applied U87 cells. High glucose application led to a notable increase in ADAMTS5 levels in cells, while low glucose application caused a moderate increase in ADAMTS5 levels. An apparent induction of IL-33 protein was observed in high glucose-applied cells, while a moderate decrease was noted in the low-glucose applied group. Insulin administration led to a decrease in IL-33 levels. Immunoreaction of NF kappa B with corresponding antibody was found to be sharply decreased in insulin-applied cells while low and high glucose application led to a moderate decrease in NF kappa B. Conclusion: This is the first reported study that has investigated both aggrecanases and inflammation mediators in the same experimental setup with U87 cells and interpreted the results in the various aspects of AD pathophysiology related to diabetes and hyperglycemia. Our findings suggest that insulin and glucose may have important functions in the synthesis of ADAMTS, IL-33, and NF kappa B through undefined mechanism(s). Further investigations dealing with all aggrecanases and other class of ADAMTS enzymes should be carried out together with the above-mentioned parameters with the collaboration of molecular biology, genetics, immunology, and other related disciplines in order to elaborate the pathophysiological importance of ADAMTS enzymes and inflammation mediators in AD.Öğe Hydrogen Peroxide-Induced Oxidative Damage in Human Chondrocytes: The Prophylactic Effects of Hypericum Perforatum Linn Extract on Deoxyribonucleic Acid Damage, Apoptosis and Matrix Remodeling by a Disintegrin-Like and Metalloproteinase With Thrombospondin Motifs Proteinases(Turkish League Against Rheumatism, 2014) Akyol, Sumeyya; Yukselten, Yunus; Cakmak, Ozlem; Ugurcu, Veli; Altuntas, Aynur; Gurler, Mukaddes; Akyol, OmerObjectives: This in vitro study aimed to examine the protective roles of Hypericum perforatum Linn (HPL) extract on cell viability, DNA damage, apoptosis and a disintegrin-like and metalloproteinase with thrombospondin motifs (ADAMTS) proteins in chondrocytes induced by hydrogen peroxide (H2O2), as a model of chondrocytes subjected to reactive oxygen species attack in rheumatoid arthritis and osteoarthritis. Materials and methods: Human chondrosarcoma cell line (OUMS-27) was used. Cells were incubated with different concentrations of methanolic extract (100, 400, and 750 mu g/ml) of HPL for 36 hours, and then treated with 0.7 mM H2O2 for two hours. Trypan blue was used for evaluation of cell viability, while DNA damage was evaluated by alkaline Comet assay. Caspase-1, ADAMTS5, ADAMTS9, and glyceraldehyde-3-phosphate dehydrogenase proteins were analyzed by Western blot. Results: In vitro H2O2 treatment decreased OUMS-27 cell viability. Cells pretreated with HPL at concentration of 400 mu g/mL were best protected from H2O2 toxicity. Compared to 100 mu g/ml concentration, pretreatment of cells with 750 or 400 mu g/ml of HPL generated more protection against H2O2-induced DNA damage. Hydrogen peroxide application to the cells led to a slight increase in Caspase-1 expression, which shows no apoptosis. The most prominent increase in Caspase-1 level was shown in cells treated with 400 mu g/ml of HPL extract. There was an increase in ADAMTS9 and a decrease in ADAMTS5 levels upon H2O2 administration. Pretreatment with HPL led to more decrease in ADAMTS5 level, indicating the protection of extracellular matrix attacked by these proteinases in cartilage tissue. Conclusion: It can be concluded that HPL has a potential to reverse the negative effects and processes induced by H2O2 in OUMS-27 cells and it can protect the surrounding cartilage area of chondrocytes from oxidative damage, which is suggested to be one of the main molecular factors accused for progression of rheumatoid arthritis and osteoarthritis.Öğe Isolation of Human Glioblastoma Multiforme Primary Cells and analysis of CD133+(Federation Amer Soc Exp Biol, 2015) Yukselten, Yunus; Yildiz, Dilara Akcora; Demircan, Kadir; Sunguroglu, Asuman; Ugur, Hasan[Abstract Not Available]Öğe PRKCA, PTPN1 and ZYX Are Possible Markers to Target CD133+GBM Stem Cells(Federation Amer Soc Exp Biol, 2016) Ozkanca, Seyma; Bal, Merve Gulsen; Yukselten, Yunus; Yildiz, Dilara Akcora; Ugur, Hasan Caglar; Demircan, Kadir; Sunguroglu, Asuman[Abstract Not Available]Öğe PXN and TRIO are elevated in CD133+GSCs transcriptomically(Federation Amer Soc Exp Biol, 2016) Sunguroglu, Asuman; Bunsuz, Merve; Yukselten, Yunus; Bal, Merve Gulsen; Ozkanca, Seyma; Yildiz, Dilara Akcora; Demircan, Kadir[Abstract Not Available]Öğe The Role of IL33 in Glioblastoma Multiforme(Federation Amer Soc Exp Biol, 2015) Demircan, Kadir; Yukselten, Yunus; Bal, Merve; Bunsuz, Merve; Yildiz, Dilara Akcora; Sunguroglu, Asuman; Ugur, Hasan[Abstract Not Available]
