Molecular Biology and Genetics / Moleküler Biyoloji ve Genetik
Permanent URI for this collectionhttps://hdl.handle.net/11147/9
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Article Citation - WoS: 18Citation - Scopus: 21Intraperitoneal Mesenchymal Stem Cell Administration Ameliorates Allergic Rhinitis in the Murine Model(Springer Verlag, 2017) Işık, Sakine; Karaman, Meral; Adan, Aysun; Kıray, Müge; Bağrıyanık, Hüsnü Alper; Çağlayan Sözmen, Şule; Kozanoğlu, İlknur; Karaman, Özkan; Baran, Yusuf; Uzuner, NevinPrevious studies showed that bone marrow-derived mesenchymal stem cells (BMSCs) could ameliorate a variety of immune-mediated and inflammatory diseases due to their immunomodulatory and anti-inflammatory effects. In this study, we developed a mouse model of ovalbumin (OVA) induced allergic inflammation in the upper airways and evaluated the effects of the intraperitoneal administration of BMSCs on allergic inflammation. Twenty-five BALB/c mice were divided into five groups; group I (control group), group II (sensitized and challenged with OVA and treated with saline-placebo group), group III (sensitized and challenged with OVA and treated with 1 × 106 BMSCs), group IV (sensitized and challenged with OVA and treated with 2 × 106 BMSCs), and group V (sensitized and challenged with phosphate buffered saline (PBS) and treated with 1 × 106 BMSCs). Histopathological features (number of goblet cells, eosinophils and mast cells, basement membrane, epithelium thickness, and subepithelial smooth muscle thickness) of the upper and lower airways and BMSCs migration to nasal and lung tissue were evaluated using light and confocal microscopes. Levels of cytokines in the nasal lavage fluid and lung tissue supernatants were measured using enzyme-linked immunosorbent assay (ELISA). Confocal microscopic analysis showed that there was no significant amount of BMSCs in the nasal and lung tissues of group V. However, significant amount of BMSCs were observed in group III and IV. In OVA-induced AR groups (group II, III, and IV), histopathological findings of chronic asthma, such as elevated subepithelial smooth muscle thickness, epithelium thickness, and number of goblet and mast cells, were determined. Furthermore, the number of nasal goblet and eosinophil cells, histopathological findings of chronic asthma, and IL-4, IL-5, IL-13, and NO levels was significantly lower in both BMSCs-treated groups compared to the placebo group. Our findings indicated that histopathological findings of chronic asthma were also observed in mice upon AR induction. BMSCs migrated to the nasal and lung tissues following intraperitoneal delivery and ameliorated to the airway remodeling and airway inflammation both in the upper and lower airways via the inhibition of T helper (Th) 2 immune response in the murine model of AR.Article Citation - WoS: 62Citation - Scopus: 80Anti-Proliferative, Apoptotic and Signal Transduction Effects of Hesperidin in Non-Small Cell Lung Cancer Cells(Springer Verlag, 2015) Çinçin, Zeynep Birsu; Ünlü, Miray; Kıran, Bayram; Bireller, Elif Sinem; Baran, Yusuf; Çakmakoğlu, BediaPurpose: Hesperidin, a glycoside flavonoid, is thought to act as an anti-cancer agent, since it has been found to exhibit both pro-apoptotic and anti-proliferative effects in several cancer cell types. The mechanisms underlying hesperidin-induced growth arrest and apoptosis are, however, not well understood. Here, we aimed to investigate the anti-proliferative and apoptotic effects of hesperidin on non-small cell lung cancer (NSCLC) cells and to investigate the mechanisms involved. Methods: The anti-proliferative and apoptotic effects of hesperidin on two NSCLC-derived cell lines, A549 and NCI-H358, were determined using a WST-1 colorimetric assay, a LDH cytotoxicity assay, a Cell Death Detection assay, an AnnexinV-FITC assay, a caspase-3 assay and a JC-1 assay, respectively, all in a time- and dose-dependent manner. As a control, non-cancerous MRC-5 lung fibroblasts were included. Changes in whole genome gene expression profiles were assessed using an Illumina Human HT-12v4 beadchip microarray platform, and subsequent data analyses were performed using an Illumina Genome Studio and Ingenuity Pathway Analyser (IPA). Results: We found that after hesperidin treatment, A549 and NCI-H358 cells exhibited decreasing cell proliferation and increasing caspase-3 and other apoptosis-related activities, in conjunction with decreasing mitochondrial membrane potential activities, in a dose- and time-dependent manner. Through a GO analysis, by which changes in gene expression profiles were compared, we found that the FGF and NF-κB signal transduction pathways were most significantly affected in the hesperidin treated NCI-H358 and A549 NSCLC cells. Conclusions: Our results indicate that hesperidin elicits an in vitro growth inhibitory effect on NSCLC cells by modulating immune response-related pathways that affect apoptosis. When confirmed in vivo, hesperidin may serve as a novel anti-proliferative agent for non-small cell lung cancer.Article Citation - WoS: 33Citation - Scopus: 37Apoptotic Effects of Quercitrin on Dld-1 Colon Cancer Cell Line(Springer Verlag, 2014) Çinçin, Zeynep Birsu; Ünlü, Miray; Kıran, Bayram; Bireller, Elif Sinem; Baran, Yusuf; Çakmakoğlu, BediaQuercetin, which is the most abundant bioflavonoid compound, is mainly present in the glycoside form of quercitrin. Although different studies indicated that quercitrin is a potent antioxidant, the action of this compound is not well understood. In this study, we investigated whether quercitrin has apoptotic and antiproliferative effects in DLD-1 colon cancer cell lines. Time and dose dependent antiproliferative and apoptotic effects of quercitrin were subsequently determined by WST-1 cell proliferation assay, lactate dehydrogenase (LDH) cytotoxicity assay, detection of nucleosome enrichment factor, changes in caspase-3 activity, loss of mitochondrial membrane potential (MMP) and also the localization of phosphatidylserine (PS) in the plasma membrane. There were significant increases in caspase-3 activity, loss of MMP, and increases in the apoptotic cell population in response to quercitrin in DLD-1 colon cancer cells in a time- and dose-dependent manner. These results revealed that quercitrin has antiproliferative and apoptotic effects on colon cancer cells. Quercitrin activity supported with in vivo analyses could be a biomarker candicate for early colorectal carcinoma.Article Citation - WoS: 27Citation - Scopus: 27New Indication for Therapeutic Potential of an Old Well-Known Drug (propranolol) for Multiple Myeloma(Springer Verlag, 2013) Kozanoğlu, İlknur; Kartal Yandım, Melis; Çinçin, Zeynep Birsu; Özdoğu, Hakan; Çakmakoğlu, Bedia; Baran, YusufPurpose: Propranolol, a non-selective β-adrenergic receptor blocker, has been used for the treatment of the patients with hypertension for more than 50 years. There are several in vitro and in vivo evidences that β-adrenergic receptor antagonists inhibit proliferation and angiogenesis and also increase apoptosis in breast, skin, and colon cancers. The aim of this study was to investigate the cytotoxic and apoptotic effects of propranolol and the genes involved in propranolol-induced apoptosis in multiple myeloma cells. Methods: Time-dependent antiproliferation and apoptotic effects of propranolol were subsequently determined by MTT cell proliferation assay, changes in caspase-3 activity, loss of mitochondrial membrane potential (MMP), and also the localization of phosphatidylserine in the plasma membrane. Changes in expression levels of NF-ΚB pathway were examined by qRT-PCR array. Results: IC50 values of propranolol on U266 cells were calculated as 141, 100, and 75 μM after 24-, 48-, and 72-h propranolol exposure, respectively. There were significant increases in caspase-3 activity, loss of MMP, and increases in apoptotic cell population in response to propranolol in U266 cells in a time- and dose-dependent manner. There were increases in expression levels of BCL10, TRAF family members, interleukins, TLR1-4, TNFRSF10B, NF-κB, and the inhibitors of NF-κB genes, and significant decreases in expression levels of Bcl-2 in response to propranolol treatment were observed. Conclusion: These results revealed that propranolol has antiproliferative and apoptotic effects on multiple myeloma cells. Being supported with in vivo analyses, propranolol can be a good and economical way to treat multiple myeloma patients.Article Citation - WoS: 46Citation - Scopus: 53Therapeutic Potential of Targeting Ceramide/Glucosylceramide Pathway in Cancer(Springer Verlag, 2013) Kartal Yandım, Melis; Apohan, Elif; Baran, YusufSphingolipids including ceramides and its derivatives such as ceramide-1-phosphate, glucosylceramide (GlcCer), and sphingosine-1-phosphate are essential structural components of cell membranes. They now recognized as novel bioeffector molecules which control various aspects of cell growth, proliferation, apoptosis, and drug resistance. Ceramide, the central molecule of sphingolipid metabolism, generally mediates anti-proliferative responses such as inhibition of cell growth, induction of apoptosis, and/or modulation of senescence. There are two major classes of sphingolipids. One of them is glycosphingolipids which are synthesized from the hydrophobic molecule, ceramide. GlcCer, generated by glucosylceramide synthase (GCS) that transfers the glucose from UDP-glucose to ceramide, is an important glycosphingolipid metabolic intermediate. GCS regulates the balance between apoptotic ceramide and antiapoptotic GlcCer. Downregulation or inhibition of GCS results in increased apoptosis and decreased drug resistance. The mechanism underlying the drug resistance which develops with increased glucosylceramide expression is associated with P-glycoprotein. In various types of cancers, overexpression of GCS has been observed which renders GCS a good target for the treatment of cancer. This review summarizes our current knowledge on the structure and functions of glucosylceramide synthase and glucosylceramide and on the roles of glucosylceramide synthase in cancer therapy and drug resistance. © 2012 Springer-Verlag Berlin Heidelberg.Article Citation - WoS: 49Citation - Scopus: 50Targeting glucosylceramide synthase sensitizes imatinib-resistant chronic myeloid leukemia cells via endogenous ceramide accumulation(Springer Verlag, 2011) Baran, Yusuf; Bielawski, Jacek; Gündüz, Ufuk; Öğretmen, BesimPurpose: Drug resistance presents a major obstacle for the treatment of some patients with chronic myeloid leukemia (CML). Pro-apoptotic ceramide mediates imatinib-induced apoptosis, and metabolism of ceramide by glucosylceramide synthase (GCS) activity, converting ceramide to glucosyl ceramide, might contribute to imatinib resistance. In this study, we investigated the role of ceramide metabolism by GCS in the regulation of imatinib-induced apoptosis in drug-sensitive and drug-resistant K562 and K562/IMA-0.2 and K562/IMA-1 human CML cells, which exhibit about 2.3- and 19-fold imatinib resistance, respectively. Methods: Cytotoxic effects of PDMP and imatinib were determined by XTT cell proliferation assay. Expression levels of GCS were determined by RT-PCR and western blot. Intracellular ceramide levels were determined by LC-MS. Cell viability analyses was conducted by Trypan blue dye exclusion assay. Cell cycle and apoptosis analyses were examined by flow cytometry. Results: We first showed that mRNA and protein levels of GCS are increased in drug-resistant K562/IMA as compared to sensitive K562 cells. Next, forced expression of GCS in sensitive K562 cells conferred resistance to imatinib-induced apoptosis. In reciprocal experiments, targeting GCS using its known inhibitor, PDMP, enhanced ceramide accumulation and increased cell death in response to imatinib in K562/IMA cells. Conclusion: Our data suggest the involvement of GCS in resistance to imatinib-induced apoptosis, and that targeting GCS by PDMP increased imatinib-induced cell death in drug-sensitive and drug-resistant K562 cells via enhancing ceramide accumulation.Article Citation - WoS: 37Citation - Scopus: 40A Novel Mechanism of Dasatinib-Induced Apoptosis in Chronic Myeloid Leukemia; Ceramide Synthase and Ceramide Clearance Genes(Springer Verlag, 2011) Gencer, Emel Başak; Ural, Ali Uğur; Avcu, Ferit; Baran, YusufSphingolipids are bioeffector molecules that control various aspects of cell growth, proliferation, apoptosis, and drug resistance. Ceramides, the central molecule of sphingolipid metabolism, are inducer of apoptosis and inhibitors of proliferation. Sphingosine-1- phosphate (S1P) and glucosyleceramide, converted from ceramides by sphingosine kinase-1 (SK-1) and glucosyleceramide synthase (GCS) enzymes, respectively, inhibit apoptosis and develop resistance to chemotherapeutic drugs. In this study, we examined the therapeutic potentials of bioactive sphingolipids in chronic myeloid leukemia (CML) alone and in combination with dasatinib in addition to investigate the roles of ceramide-metabolizing genes in dasatinib-induced apoptosis. Cytotoxic effects of dasatinib, C8:ceramide, PDMP, and SK-1 inhibitor were determined by XTT cell proliferation assay. Changes in caspase-3 enzyme activity and mitochondrial membrane potential (MMP) were measured using caspase-3 colorimetric assay and JC-1 MMP detection kit. Expression levels of ceramide-metabolizing genes were examined by qRT-PCR. Application of ceramide analogs and inhibitors of ceramide clearance genes decreased cell proliferation and induced apoptosis. Targeting bioactive sphingolipids towards generation/accumulation of ceramides increased apoptotic effects of dasatinib, synergistically. It was shown for the first time that dasatinib induces apoptosis through downregulating expression levels of antiapoptotic SK-1 but not GCS, and upregulating expression levels of ceramide synthase (CerS) genes, especially CerS1, in K562 cells. On the other hand, dasatinib downregulates expression levels of both GCS and SK-1 and upregulate apoptotic CerS2, -5 and -6 genes in Meg-01 cells. Increasing endogenous ceramide levels and decreasing prosurvival lipids, S1P, and GC, can open the way of more effective treatment of CML.Article Citation - WoS: 42Citation - Scopus: 48The Roles of Bioactive Sphingolipids in Resveratrol-Induced Apoptosis in Hl60 Acute Myeloid Leukemia Cells(Springer Verlag, 2011) Çakır, Zeynep; Saydam, Güray; Şahin, Fahri; Baran, YusufPurpose Acute promyelocytic leukemia results from a translocation between 15 and 17 chromosomes that produce PML/RARa fusion protein. PML/RARa inhibits differentiation of myeloid precursor cells at stem cell level. Resveratrol is a phytoalexin that exerts cytotoxic effects on cancer cells. Ceramides have crucial roles in cell growth, proliferation, differentiation, drug resistance, and apoptosis. In this study, we examined the possible cytotoxic effects of resveratrol on acute myeloid leukemia cells and determined the roles of ceramide-metabolizing genes in resveratrol-induced apoptosis, in addition to investigating the possibility of increasing the sensitivity of HL60 cells to resveratrol by manipulating sphingolipids. Methods Cytotoxic effects of resveratrol, C8:ceramide, PDMP, and SK-1 inhibitor were determined by XTT cell proliferation assay. Changes in caspase-3 enzyme activity and mitochondrial membrane potential (MMP) were measured using caspase-3 colorimetric assay and JC-1 MMP detection kit. Expression levels of ceramide-metabolizing genes were examined by RT-PCR. Results The results revealed that manipulations of ceramide metabolism toward generation or accumulation of apoptotic ceramides increased apoptotic effects of resveratrol in HL60 cells, synergistically. More importantly, gene expression analyses revealed that resveratrol-induced apoptosis via increasing expression levels of ceramide generating genes and decreasing expression levels of antiapoptotic sphingosine kinase-1 and glucosylceramide synthase genes. Conclusion These results showed for the first time that increasing intracellular levels of ceramides by biochemical approaches has also increased sensitivity of HL60 cells to resveratrol. We also showed that resveratrol induces apoptosis through manipulating ceramide-metabolizing genes that resulted in the accumulation of ceramides in HL60 cells.Article Citation - WoS: 22Citation - Scopus: 28Caffeic Acid Phenethyl Ester Triggers Apoptosis Through Induction of Loss of Mitochondrial Membrane Potential in Ccrf-Cem Cells(Springer Verlag, 2011) Avcı, Çığır Biray; Gündüz, Cumhur; Baran, Yusuf; Şahin, Fahri; Yılmaz, Sunde; Doğan, Zeynep Özlem; Saydam, GürayPurpose CAPE (caffeic acid phenethyl ester) is one of the most valuable and investigated component of propolis which is composed by honeybees. In the current study, we aimed at examining apoptotic effects of CAPE on CCRF-CEM leukemic cells and at determining the roles of mitochondrial membrane potential (MMP) in cell death. Methods Trypan blue and XTT methods were used to evaluate the cytotoxicity. Apoptosis was examined by ELISA-based oligonucleotide and acridine orange/ethidium bromide dye techniques. Loss of mitochondrial membrane potential was evaluated using JC-1 dye by flow cytometric analysis and under fluorescent microscope. Results We detected the time-and dose-dependent increases in cytotoxic effect of CAPE on CCRF-CEM cells. ELISA and acridine orange/ethidium bromide results showed that apoptotic cell population increased significantly in CCRF-CEM cells exposed to increasing concentrations of CAPE. On the other hand, there was significant loss of MMP determined in response to CAPE in CCRF-CEM cells. Conclusion This in vitro data by being supported with clinical data may open the way of the potential use of CAPE for the treatment of leukemia.