Master Degree / Yüksek Lisans Tezleri
Permanent URI for this collectionhttps://hdl.handle.net/11147/3008
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Master Thesis Investigating Oncogenic Role of Sema6d in Breast Cancer Cells(Izmir Institute of Technology, 2019) Günyüz, Zehra Elif; Yalçın Özuysal, Özden; Yalçın Özuysal, Özden; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyBreast cancer, the most commonly diagnosed cancer type and the leading cause of cancer-associated deaths, is the major health issue among women worldwide. In many cancer types, the expression of the semaphorins and their receptors such as plexins and neuropilins are dysregulated. SEMA6D is a member of class-6 family transmembrane semaphorin proteins and acts through Plexin-A1 receptor. It was previously shown that overexpression of SEMA6D in breast cancer cell line MCF-7 leads to a reduction in proliferation and an increase in migration. On the other hand, in the MDA-MB-231 breast cancer cell line, overexpression of SEMA6D had no significant effect on proliferation but enhanced migration. In this study, we aimed to analyze the effects of SEMA6D overexpression in normal breast cell line MCF10A and investigate the invasive behavior and transformation capacity of SEMA6D overexpressing breast cancer cell lines. We demonstrated that overexpression of SEMA6D leads to elevated proliferation, viability and migration in MCF10A cells, whereas it did not trigger their anchorage-independent growth. On the other hand, MDA-MB-231 and MCF7 cells stably expressing SEMA6D showed reduced colony formation in the soft-agar assay. Furthermore, the invasiveness of MDA-MB-231 cells was elevated with SEMA6D overexpression, whereas SEMA6D overexpression did not stimulate the invasiveness of MCF-7 cells through matrigel microenvironment, whereas slightly trigger invasion through bone microenvironment. In conclusion, SEMA6D overexpression has cell-specific effects on breast cancer. The exact role of SEMA6D in breast cancer development remains undefined and must be further investigated.Master Thesis Determination of Therapeutic Potential of Luteolin for Acute Lymphoblastic Leukemia Cells(Izmir Institute of Technology, 2019) Gürler, Sevim Beyza; Baran, Yusuf; Baran, Yusuf; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyAcute lymphoblastic leukemia (ALL) is a hematologic malignancy characterized by increased level of immature lymphoblasts in bone marrow and peripheral blood. The developments of lymphoblasts are genetically/epigenetically inhibited. One of the most common genetic abnormalities in ALL is BCR/ABL translocation which regulates the several pathways related to proliferation, anti-apoptotic and drug resistance through its aberrant tyrosine kinase activity. Although the current treatment strategies include targeting BCR/ABL via tyrosine kinase inhibitors; complete remission, overall survival and mortality of Ph+ ALL patients are still worse as compared to Ph- ALL patients. Therefore, new strategies combined with current treatments are needed for Ph+ ALL patients who are qualified as high risk group of ALL. Different studies showed thatluteolin has anti-cancer and anti-tumor effects on wide range cancer types including breast, colon, lung cancer except ALL in both in vitro and in vivo. In this study, the dose and time dependent cytotoxic, apoptotic and cytostatic effects of luteolin on Philadelphia chromosome +ALL cells were determined for the first time. Besides, the effect of luteolin on cell growth and proliferation of two different healthy cell lines was shown. Moreover, the effect of luteolin on bioactive sphingolipids genes which regulate the several pathways including cell proliferation, apoptosis, drug resistance and senescence in cell was determined in Ph positive ALL cells for the first time. As a consequence, luteolin has cytotoxic, apoptotic and cytostatic effects on Ph positive ALL cells and bioactive sphingolipids genes are regulated in this therapeutic potential by luteolin.Master Thesis Determination of Therapeutic Potential of Apigenin on Acute Lymphoblastic Leukemia Cells(Izmir Institute of Technology, 2019) Uzuner, Erez; Baran, Yusuf; Baran, Yusuf; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyAcute lymphoblastic leukemia (ALL) is a hematological disorder initiating from blood-forming cells of bone marrow. ALL is characterized by the Philadelphia chromosome (Ph) arisen from a translocation between chromosome 9 and 22. This chromosome encodes BCR-ABL oncogene that is a driver regulator. BCR-ABL based studies improved tyrosine kinase inhibitors (TKI) including imatinib, dasatinib, nilotinib, and ponatinib to eliminate this disease. However, the studies on Ph+ ALL patients showed that bioactive sphingolipids have crucial roles in the elimination of the positive effects of these drugs by activating the proliferation-associated pathways, inhibition of apoptosis and increasing drug resistance of the cells treated with these drugs. In this study, therapeutic potential of apigenin, which is a natural flavonoid obtained from celery, parsley and chamomile was investigated on Ph+ ALL cell line, SD-1, and non-cancerous lung cell line Beas-2B. The cytotoxic effects of apigenin on SD-1 and Beas-2B cells were determined by MTT cell proliferation assay. The cell viability analyses on SD-1 cells were conducted by Trypan blue dye exclusion assay following apigenin treatment. Cell cycle and apoptosis analyses including Annexin V/PI-dual staining and JC-1 dye-based mitochondrial membrane potential were examined by flow cytometry. Expression levels of bioactive sphingolipids were determined by RT-PCR and western blot. The cytotoxic analyses indicated that apigenin selectively inhibits the expression of SD-1 cells whereas the IC50 value of apigenin for SD-1 cells has the anti-apoptotic roles in Beas-2B cells. SD-1 cells experience cell death via apoptosis-related pathways and apigenin might arrest the cells at G2/M phases. Indeed, the changes in the expression levels of bioactive sphingolipids genes indicated their roles in apigenin-induced apoptosis in SD-1 cells. This study investigated the cytotoxic and apoptotic effects of apigenin on SD-1 cells and the roles of apigenin in bioactive sphingolipid metabolism for the first time.Master Thesis Analysis of Tnfrsf10b-As Long-Noncoding Rna's Effects on Various Cancer Cell Properties(Izmir Institute of Technology, 2019) Alkan, Ayşe Hale; Akgül, Bünyamin; Akgül, Bünyamin; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyLong noncoding RNAs (lncRNAs) being longer than 200 nucleotides constitute a different class of RNA molecules. Several studies indicated that they have regulatory role in cellular processes including cancer development. Some of them have exclusively high expression in particular cancer types and regulate certain cancer cell properties. This renders them potential biomarker or therapeutic target in cancer. In this study, effects of a candidate lncRNA TNFRSF10B-AS and lncCAMTA1 on cancer cell properties were investigated. Candidate lncRNAs from Doxorubicin, Fas mAB, TNF-alpha and Cisplatin treated HeLa cell line were chosen and their expression level was measured in different cell lines including healthy (BEAS2B and MCF10A), metastatic (H1299 and MDA-MB- 231) and non-metastatic cell lines (A549 and MCF-7) by qPCR. From a few candidates lncCAMTA1 and TNFRSF10B-AS were selected for further analysis. qPCR results obtained from comparison of different cancer cell lines showed that their expression differs at least in one comparison of cell lines. TNFRSF10B-AS silencing decreased proliferation of HeLa cells. lncCAMTA1 was silenced or overexpressed in HeLa cells but phenotypic effect couldn’t be detected by apoptosis and cell proliferation assay. Additionally, phenotypic effect also couldn’t be observed in other cell lines when TNFRSF10B-AS was silenced.Master Thesis Assesment of Genes That Play Role in Phenolic Compound Degradation in Olive Orchard Microbiome(Izmir Institute of Technology, 2019) Acar, Ertuğrul; Soyer, Ferda; Soyer Dönmez, Ferda; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyThe olive tree (Olea Europea L.) is one of the most important fruit trees in Mediterranean countries. Its products, olive oil and table olives, are important components of the Mediterranean diet and widely consumed all around the World. Olives and virgin oil provide a rich source for phenolic compounds. The plant phenolics are secondary metabolites, and possesses several common biological and chemical properties. In this study, microorganisms were collected from soil, olive leaves, fruits, and Olive fruit fly larva and Olive mill wastewater (OMWW). They were characterized by 16S rRNA analysis. The microorganisms that were tolerant to phenolic compounds were selected in order to seek which genes were associated with the phenolic compound degradation. The genes related to the degradation of the selected organisms were identified by Sanger Sequencing and the level of phenol-degrading gene expression were aimed to be compared by using qPCR and Droplet Digital PCR (ddPCR). Microorganisms which degrade phenolic compounds can be harnessed for the purpose of bioremediation. However, the number of defined phenolic compound degrading microorganisms is still low in the literature. For this reason, many different microorganisms were used at the same time for bioremediation. Investigation of olive orchard microorganisms and phenolic-degrading genes might benefit bioremediation in the future. In this study, 8 different bacterial strains were identified and characterized from olive orchards. After that, their phenol hydroxylase and catechol 1,2 dioxygenase genes tried to be sequenced with primers designed by using of reference strains in NCBI database.Master Thesis Assessment of Liposomal Formulations and Biological Activities of Eggplant Glycoalkaloids(Izmir Institute of Technology, 2019) Tatlıdil, Engin; Frary, Anne; Tatlıdil, Engin; Frary, Anne; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyBillions of dollars are spent every year in the world for cancer treatments and research. In recent years, bioactive compounds are being tested as promising therapeutics. Among these compounds, eggplant glycoalkaloids: solasonine and solamargine are known to be effective against skin cancer and diseases. However, these compounds are water insoluble. This reduces transdermal drug delivery and the efficacy of solasonine and solamargine. Nanocarriers are used for transdermal drug delivery of water insoluble molecules. In this study, liposomes were used as nanocarriers to increase drug delivery of solasonine and solamargine. In the first stage of the study, empty liposomes produced from four different lecithin types with hydration temperatures of 40°C, 45°C and 50°C were evaluated according to their physical, chemical stability and drug loading capacity criteria at three different storage temperatures (4°C, 25°C, 37°C). The liposome formulation which was most suitable for the continuation of the study was determined. In the second stage of the study, solasonine and solamargine loaded liposomes were produced according to the formulation determined in the first stage and these liposomes were evaluated according to their physical, chemical stability, zeta potentials and drug leakage rate criteria for 3 months and it was determined that the drug loaded formulation was stable during the monitoring process. Furthermore, the release profiles of the drugs in different release media were determined and also the efficacy of the free and encapsulated states of solasonine and solamargine were tested in HaCaT and SCC-25 cell lines and IC50 values were determined.Master Thesis Determination of Therapeutic Effects of Multifunctional Micelle-Based Nanocarriers on Breast Cancer Cells(Izmir Institute of Technology, 2019) Ulu, Gizem Tuğçe; Baran, Yusuf; Ulu, Gizem Tuğçe; Baran, Yusuf; 01.01. Units Affiliated to the Rectorate; 04.03. Department of Molecular Biology and Genetics; 01. Izmir Institute of Technology; 04. Faculty of ScienceBreast cancer is the most common and frequent cause of death among women composed to all types of cancer. Current treatment protocols do not provide complete cure or selective drug delivery while targeted therapy can provide an important avenue for successful treatment of breast cancer. In this study, therapeutic effects of drug-conjugated nanocarrier system with enhanced stability and double moiety pH-sensitivity on breast cancer (SKBR-3- HER-2- positive), normal breast epithelial (MCF-10A, HER-2-negative) and chronic myeloid leukemia (K562, HER-2-negative) cells were determined. With this approach, SKBR-3 cells were targeted by single nanocarriers having selectivity with unused peptide ligand (HER-2), stability with cross-linking of core moiety, and cleavage by two sites of pHeffect and drug release properties. After physicochemical characterization of micellebased nanocarriers, cytotoxic, apoptotic and cytostatic effects of doxorubicin conjugated micelles were determined. Doxorubicin conjugated micelles with HER-2 peptide (DOX-HER-2-NCs) had more cytotoxic effects on HER-2 positive cells. Additionally, intracellular amounts of doxorubicin is higher in SKBR-3 cells with applied DOX-HER-2-NCs as determined by fluorescence imaging. The apoptosis rate was increased on SKBR-3 at 50% cell growth inhibition (IC50) as determined by Annexin-V/Propidium iodide double staining. However, there was not any significant change in loss of mitochondrial membrane potential. Additionally, DOX-HER-2-NCs resulted in cell cycle arrest at G2/M-phase in response to IC50 value. Besides, protein level of Bcl-2 did not change while protein level of Bax and Caspase-3 were increased as determined by Western Blotting. This project provides novel and more effective treatment of breast cancer by using multifunctional properties of nanocarriers.Master Thesis The Effect of Glibenclamide in Lipopolysaccharide Stimulated Brain Microvascular Endothelial Cells(Izmir Institute of Technology, 2019) Cihankaya, Hilal; Tosun, Çiğdem; Tosun, Çiğdem; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyEndothelial cells are essential components of the blood brain barrier (BBB) that regulate the exchange of solutes between the vasculature and the brain parenchyma. The integrity of the BBB is disrupted after central nervous system (CNS) injuries, and it has been associated with the Sur1-Trpm4 channels. Once these channels are opened, Na+ flows into the cells causing edema and cell death. To mimic CNS injuries in vitro, lipopolysaccharide (LPS) was used as an endotoxin to initiate proinflammatory mediators to increase endothelial permeability, and glibenclamide was used as an antagonist of Sur1-Trpm4 channels to reduce the inflammatory response and to maintain the structural integrity of BBB proteins. To demonstrate the role of glibenclamide following LPS stimulation, we determined the cytotoxicity of LPS in bEnd.3 cells by cleaved caspase-3 expression and propidium iodide staining. We also investigated the protective effect of glibenclamide on NF-B translocation, and BBB proteins; collagen IV (COL IV) and zonula occludens 1 (ZO-1) in LPS stimulated bEnd.3 cells. Our results revealed that 1g/ml LPS exposure was sufficient for NF-B nuclear translocation, along with a statistically significant decrease in the expressions of COL IV and ZO-1 proteins, and a significant increase in cell death. We also demonstrated that glibenclamide was able to restore the expressions of COL IV and ZO-1, significantly reduce NF-B translocation, and cell death. Taken together, LPS induction in bEnd.3 cells can be used to investigate endothelial cell dysfunction due to inflammation in stroke and trauma models and glibenclamide can be used as a protective drug to inhibit LPS stimulated inflammatory response, cell death and disruptions in the structures of key BBB proteins.Master Thesis Investigation of Autophagy Related Markers in Brain Tissue of Early-Onset Tay-Sachs Disease Mouse Models(Izmir Institute of Technology, 2018) Ateş, Nurselin; Ateş, Nurselin; Seyrantepe, Volkan; Seyrantepe, Volkan; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyTay-Sachs disease is an autosomal recessively inherited lysosomal storage disorder that mainly affects the central nervous system. It is caused by mutations on the HEXA gene encoding α-subunit of β-Hexosaminidase A enzyme. The enzyme normally catalyses GM2 to GM3 conversion but when it is absent or dysfunctional the GM2 degradation is interrupted. Progressive accumulation of the undegraded GM2 ganglioside in neurons causes neurodegeneration and eventual death for the patient. The Hexa-/- mice generated as Tay-Sachs model was nearly normal and a bypass mechanism mediated by a sialidase was discovered. Neu3 sialidase involvement in ganglioside degradation in the Tay-Sachs disease pathology was reported and the Hexa-/-Neu3-/- mice was observed to mimic the neuropathologic and clinical phenotype of the disease. Therefore, it can be used as early-onset-Tay-Sachs disease mouse model. Lysosomal storage diseases have been reported as disorders of autophagy as the lysosomal accumulation expected to affects the autophagical-lysosomal pathway in one way or another. In the concept of our study comparative analysis of WT, Hexa -/- ,Neu3 -/- and Hexa -/- Neu3 -/- mice provided the information that early-onset Tay-Sachs disease model exhibit impairment in autophagic flux and secondary accumulation of autophagic components. The effect of abnormal GM2 and this secondary accumulation on apoptotic regulators and trigger factors were also investigated. In the light of our study, impairment in autophagic flux, increased oxidative stress and ER-stress are involved in the disease pathology of early-onset Tay-Sachs disease mouse model.Master Thesis Expression of Aquaporin 1, 3 and 4 in T Cell Activation and Apoptosis(Izmir Institute of Technology, 2018) Gelmez, Ayşe Bengisu; Nalbant Aldanmaz, Ayten; Nalbant Aldanmaz, Ayten; 04.03. Department of Molecular Biology and Genetics; 04. Faculty of Science; 01. Izmir Institute of TechnologyAquaporins (AQPs) are membrane proteins responsible for transporting water, some gases and small solutes such as CO2 and glycerol. Until now, it has been shown that AQP1, 3 and 5 expressed in both B and T lymphocytes of mice, regulate cell volume. However, aquaporin expression involved in activation, proliferation, and differentiation as well as apoptosis of T cells are not well known yet. The goal of this study is to detect the expression level of AQP1, AQP3, and AQP4 in activated and apoptotic T cells. In order to do that, two types of T cells cultured in both condition were utilized. Peripheral Blood Mononuclear Cells (PBMCs) were isolated from Human peripheral blood drawn from healthy donors by ficoll density gradient Centrifugation method. Naive CD4+ T cells were sorted from PBMC. The stimulants generating Th17 were chosen for activation and differentiation of naïve CD4 T cells. Jurkat cell line as a second cell type were activated by PMA/Ionomycin as well as treated by camptothecin for apoptotic processing. Th17 and Jurkat cell cultures were analysed by flow cytometry to measure the rate of both activation and apoptosis. Western Blot was performed to identify expression of AQP 1, 3 and 4. We found a significance between increased expression level of AQP1, 3, and 4 in activated T cells as well as decreased expression level of each three AQPs in apoptotic T cell populations. According to our findings, tested aquaporin proteins may play roles in T cell activation, differentiation, and apoptosis. The scientific significance of this research is that it can fill the gaps about these three functional processes of T cells. Besides, all findings can contribute to treatment of many autoimmune disease like MS which Th17 cells involve in pathogeny.