Biyomühendislik Ana Bilim Dalı Tez Koleksiyonu

Permanent URI for this collectionhttps://hdl.handle.net/20.500.12573/417

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Advancing machine learning analysis of non-coding RNA: A novel approach of negative sequence generation
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2024) Orhan, Mehmet Emin; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Many supervised machine learning models have been developed for the classification and identification of non-coding RNA (ncRNA) sequences. These models play a significant role in the diagnosis and treatment of various diseases. During such analyses, positive learning datasets typically consist of known ncRNA examples, some of which may even be confirmed with strong experimental evidence. However, there is no database of validated negative sequences for ncRNA classes or standardized methodologies for generating high quality negative samples. To overcome this challenge, a new method for generating negative data called the NeRNA (Negative RNA) method has been developed in this study. NeRNA generates negative sequences using known ncRNA sequences and their octal representations, similar with frame shift mutations found in biology but without base deletions or insertions. In this thesis, the NeRNA method was tested separately with four different ncRNA datasets, including microRNA (miRNA), transfer RNA (tRNA), long non-coding RNA (lncRNA), and circular RNA (circRNA). Additionally, a species-specific case study was conducted to demonstrate and compare the performance of the study's miRNA predictions. The results of 1000-fold cross-validation on machine learning algorithms such as Decision Trees, Naive Bayes, Random Forest classifiers, and deep learning algorithms like Multilayer Perceptrons, Convolutional Neural Networks, and Simple Feedforward Neural Networks showed that models developed using datasets generated by NeRNA exhibited significantly high prediction performance. NeRNA has been published as an easy-to-use, updatable, and modifiable KNIME workflow, along with example datasets and required extensions that can be downloaded and utilized. NeRNA is designed specifically as a powerful tool for RNA sequence data analysis.
BBSome regulates ARL13B-dependent joint elongation of two distinct cilia in Caenorhabditis elegans
(Abdullah Gül Üniversitesi / Fen Bilimleri Enstitüsü, 2023) Turan, Merve Gül; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Cilia or flagella are interchangeably used to refer to the hair-like organelles extending from the cell surface to communicate with environmental signals or triggers. Cilium, the singular form of cilia, and its components are well-conserved structures throughout evolution and are divided into motile and primary cilium. The primary cilia of different cells are seen to form joint cilia by extending in parallel. For instance, PHA and PHB primary cilia in C. elegans protrude from the ends of the dendrite but extend parallel to one another and intersect in the middle portion of the cilia, reaching the same length. Nevertheless, the molecular mechanisms underlying how parallel cilia get similar lengths remain mysterious. In this thesis, we used C. elagans as a model organism to examine the molecular mechanism associated with the cilia direction. We generated various single, double, and triple mutants to examine PHA and PHB cilia for phenotype and length. We found that a Joubert syndrome protein, ARL13B, is required for determining cilia direction in PHA & PHB cilia and ASE & ASI cilia.
A BIOMIMETIC APPROACH FOR SMALL DIAMETER VESSELS: BILAYERED VASCULAR GRAFTS MADE of ALGINATE and POLY(-CAPROLACTONE)
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2019) GÜRDAP, Seda; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Cardiovascular diseases (CVDs) still remain one of the leading causes of morbidity and mortality across the world. A typical symptom of CVDs is the vascular occlusion. There are many strategies for treatment such as angioplasty, stent application and bypass grafting. Although synthetic blood vessels are successfully used in bypassing of the medium (>10 and<6) and large sized (10 mm) vessels, they have high failure problem for the replacement of small diameter ( 6 mm) vessel because of early thrombosis formation. Tissue engineering, mimicking the structural, mechanical and cell growth characteristics of the native vessels is a promising treatment method for CVDs. In this study, it was aimed to fabricate a bilayered vascular scaffold by combining thermally induced phase separation and electrospinning methods. First, alginate porous layer was produced as the inner layer with the average pore diameter of approximately 100 μm to enable endothelial cell attachment and proliferation. Then, the inner layer was covered with electrospun polycaprolactone (PCL) membrane to strength the endurance of vascular graft. The mechanical test showed that the bilayered vascular scaffold has a close mechanical characteristic to native vessels with elastic modulus of 2 .45 1.7 MPa and estimate burst pressure of 0,18 MPa. Also, heparin was chemically immobilized to scaffold to elongate the release time, which can result in reduced thrombosis. In addition, cross-linked scaffold lost 21% of its mass for 6 weeks showed the moderate degradation level that can support the neotisue formation via cell migration to the scaffold, while the scaffold is synergistically degraded. According to the results, the materials prepared by biomimetical approach revealed that they have a great potential to be used as a synthetic vascular graft.
Biosynthesis of high value-added carotenoids by engineered microorganisms
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2024) Arslansoy, Nuriye; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Carotenoids are pigment molecules that play an important role in coloring plants, algae, and other organisms. These molecules exhibit various biological activities such as anticancer, antiviral and antioxidant activities. They have a huge market size and are mainly used in the food, feed, and cosmetic industries. The current supply chain for carotenoids is mostly relied on the extraction from plants and/or chemical synthesis for certain carotenoids. However, these strategies have various bottlenecks and disadvantages such as being affected by climate change, more difficult and costly extraction processes, and environmental issues. These can be overcome with microbial biosynthesis, which not only addresses the previous problems but also provides advantages of producing in a short time and scale-up for industrial production. In this research, we aimed to biosynthesize the high value-added carotenoids by engineered microorganisms. The genome of a native producer of zeaxanthin diglucoside, identified as endophytic Pseudomonas sp. 102515, was first edited by CRISPR-Cas9 to knock out zeaxanthin glucosyltransferase (CrtX), lycopene β-cyclase (CrtY) and beta-carotene hydroxylase (CrtZ). This led to ΔcrtX, ΔcrtY and ΔcrtZ mutant strains of Pseudomonas sp. 102515. On the other hand, overexpression plasmids carrying crtW, CaZEP and CaZEP-CaCCSm40 genes were constructed and transformed to ΔcrtX mutant to synthesize astaxanthin, violaxanthin and capsanthin/capsorubin. HPLC analysis of extracts from mutant strains and overexpression strains revealed that all the engineered strains produced the corresponding carotenoids such as zeaxanthin, β-carotene, and lycopene. Thus, this study paved the way for the biosynthesis of valuable carotenoids in the engineered endophytic bacteria
Centella asiatica extract containing bilayered electrospun wound dressing
(Abdullah Gül Üniversitesi, 2019) KOÇ, NURAY; AGÜ, Yaşam ve Doğa Bilimleri Fakültesi, Biyomühendislik Bölümü; KOÇ, NURAY
Innovative and bioactive wound dressings prepared by electrospinning mimicking the native structure of the extracellular matrix (ECM) have gained significant interest as an alternative to conventional wound care applications. In this study, bilayered wound dressing material was produced by sequential electrospinning of quaternized poly(4- vinyl pyridine) (upper layer) on the Centella Asiatica (CA) extract containing electrospun poly(D, L-lactide-co-glycolide) (PLGA)/poly(3-hydroxybutyrate-co-3- hydroxy valerate) (PHBV) blend membrane (lower layer). Scanning electron microscopy (SEM) was utilized to show a uniform and bead-free fiber structure of electrospun membranes. The average diameter of CA extract containing electrospun PLGA/PHBV blend membrane was calculated 0.471±0.11 µm, whereas the average fiber diameter of electrospun poly(Q-VP) membranes was in the range of 0.460±0.057 µm. Chemical, thermal, mechanical properties, and adsorption capacity of electrospun membranes, as well as the cumulative release of CA from the electrospun PLGA/PHBV membrane, were investigated. Viability, adhesion, and attachment of human fibroblast cells on the electrospun membranes on pre-set days were evaluated by the colorimetric CellTiter 96® Aqueous One Solution Cell Proliferation Assay (MTS assay) and SEM. Results revealed that CA loaded bilayered electrospun wound dressing showed promoted attachment and proliferation of fibroblasts. Hence, it can be concluded that CA extract containing bilayered electrospun wound dressing prepared in this study has a promising potential for wound healing applications.
CHARACTERIZATION OF NOVEL CILIARY GENE TMEM145
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2020) PİR, Mustafa Samet; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Cilia and flagella are highly conserved, microtubule based cellular structures which are found in most of the organisms. They have variety of functions from enabling movement in protozoa to signal transduction in multi cellular organisms. Defects in the structure or the function of cilia in human cause a broad range of diseases called ciliopathies. These defects in cilia are caused by mutations on ciliary genes and some non-ciliary genes that affect function of cilia. Therefore, there is a constant need for new ciliary genes to be identified which may help reveal the molecular basis of ciliopathies. We have identified C15A7.2, a GPCR protein in Caenorhabditis elegans as a ciliary gene which is an ortholog of human TMEM145 gene. We have investigated the function of C15A7.2 encoding protein TMEM-145 and found decrease in the speed of intraflagellar transport system in C15A7.2 mutant. We have not observed any structural defect in neither single nor various double mutants, implying that TMEM-145 is not required for ciliogenesis. Having localized exclusively in cilia, TMEM-145 is required to be further investigated.
Cloning of 32-mer MaSP1 Gene ınto pBbB6c plasmid vector and transformation to Escherichia coli neb 10-beta
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2023) Benk, Ruveyda; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
The main purpose of my thesis was to clone Masp1 spider silk protein encoding gene from dragline type spider into E.coli NEB 10-beta organism. The recombinant microbial production of spider silk protein and converting it into a fiber format would ultimately produce a biomaterial also called as biosteel with high toughness and elasticity whereas low density compared to Kevlar, steel and carbon fiber. For this purpose, the gene encoding the dragline spider protein (MaSP1) was cloned into E. coli NEB 10-beta using recombinant molecular methods. First, 8-mer MaSP1 was synthesized and cloned via pGSI high copy cloning vector by sticky end cutting with restriction enzymes of KpnI,Kpn2I followed by heat-shock transformation into E.coli. Second, we performed restriction of the 8-mer plasmid by NheI and Kpn2I to extract the 8-mer. Later, the restriction was performed by SpeI and Kpn2I to obtain linearized pGSI containing 8-mer Masp1. A ligation was applied to merge 8-mer and pGSI plasmid carrying 8-mer Masp1 to achieve 16-mer Masp1 containing pGSI. Again, this plasmid was heat-shock transformed into E.coli. Following the same restriction 32-mer Masp1 containing pGSI plasmid was achieved. Finally, 32-mer Masp1 fragment was cut from pGSI cloning vector and ligated to pBbB6c low copy expression plasmid followed by electroporation into E.coli. The band size of 32-mer Masp1 gene was aligned between 3 kb and 5 kb which is an agreement with the calculated size of 32-mer Masp1 gene. Future studies should focus on the expression of Masp1 and the efficient production of this valuable recombinant protein under bioreactor conditions with cutting edge bioprocessing techniques.
Determination of probiotic properties of lactic acid bacteria isolated from traditional food products
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2023) Yiğit, Mehmet Burak; 0000-0002-6777-6773; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Probiotics are microorganisms that live in our bodies and positively affect health when consumed regularly. One of the ways to have a healthy body is to have a healthy microbiota. Because of that, the importance given to the consumption of probiotic foods among the public is increasing. Since probiotics are especially abundant in fermented and traditional foods, consuming these foods is vital to have a healthy microflora. In this thesis, probiotic potentials of bacteria isolated from tarhana, einkorn sourdough, Turkish and Bulgarian-type boza and pickled beetroot foods were investigated, and obtained results were discussed. Based on acid and bile salt tolerance tests, MRS ES-2-3-7-11-12- 17, MRS PT-2-14-16, MRS N-1, MRS EB-3, MRS T-1, M17 N-2 -3-4 showed higher viability in acidic environments (pH 2.0 and 3.0) than the control groups, M17 N-3-4 and M17 TB-1-2 strains showed higher viability at 0.3% and 0.5% bile salt conditions than other strains. For 10 strains which are selected for further tests, in the adhesion to Caco2 cells, MRS ES-3, MRS N-1, MRS T-2, M17 BB-7, M17 N-2 and M17 N-3 showed over 35% adhesion, especially, MRS N-1 and M17 N-2 showed over 85% adhesion to Caco-2 cells. For the antimicrobial activity test, ES-3 strain showed limited effect on S. aureus ATCC 6538 and K. pneumoniae ATCC 4352 pathogens, while other strains showed no inhibitory effect on pathogens. Finally, according to the results of 16S rRNA sequencing, it was determined that MRS ES-3, ES-7, PT-14 strains belonged to L. plantarum, MRS ES-11 strain belonged to L. brevis, M17 BB-7 strain belonged to E. faecium and M17 TB-2 strain belonged to E. durans species.
DEVELOPMENT OF A KIDNEY TARGETTED NANOCARRIER FORMULATION AGAINST CISPLATIN INDUCED NEPHROTOXICITY
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2019) ÇAKIR, Şerife; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Chitosan is a natural polymer which is less toxic in the body than other synthetic polymers. Chitosan sodium tripolyphosphate (TPP) nanoparticles produced by ionic gelation method are known to be good drug delivery agents for human body tissues, such as brain and kidney. In this study, to eliminate nephrotoxicity in kidney caused by cisplatin, an anti-cancer drug, chitosan-TPP nanoparticles loaded with gene silencing siRNAs were used. In vitro studies have been tested in human kidney cell line Hek293 cells and the toxic effects of nanoparticles were found to be low compared to 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide and 2,3- Bis-(2-Methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide, disodium salt cell cytotoxicity results. The penetration of nanoparticles into the cell was confirmed by fluorescence microscopy and flow cytometry. In vivo studies have shown that nanoparticle injected with siRNA loaded into rats of type 6-8 week in the Balb-c type. Control and siRNA-loaded chitosan nanoparticles cisplatin-treated mice group were used for the animal experiment. After cisplatin injections, creatinine and BUN assays were performed to detect the level changing by nanoparticles after cisplatin-induced siRNA delivery. GAPDH is a control gene and PKCδ, P53, OCT1, OCT2 and γGT genes have important roles in kidney proximal tubule cells. In this study, mRNA levels of these genes were also examined by quantitative PCR. While siRNAs were decreased in the first days of injection, this effect was lost in the following days. Thus, the silencing potential of each siRNA is variable. However, this variability shows a significant change in the study.
DEVELOPMENT OF A LOW-COST MICROFLUIDIC SYSTEM TO DETECT IMMUNOMAGNETICALLY CAPTURED LEUKEMIA CELLS
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2020) Akar, Ünal; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Sensör teknolojileri fiziksel özellikleri ölçülebilir sinyallere dönüştürmek için kullanılır. Özellikle yarı iletken teknolojisindeki gelişmelerle birlikte, mikroakışkan olarak bilinen yeni bir teknoloji ortaya çıktı. Mikroakışkanlar zamandan ve maliyetten tasarruf sağlayan ileri bir teknolojidir. Kimya, biyoloji, bilgi teknolojisi, optik vb. gibi farklı kullanım alanlarına sahiptir. Akut lenfoblastik lösemi kötü huylu bir kan kanseridir, özellikle B öncüllü akut lenfoblastik lösemi çocukluk döneminde çok tehlikelidir. Eğer tedaviden sonra vücut içinde az bir miktar da olsa kanser hücresi kalırsa, doktorlar bu hücreleri fiziksel ya da diğer semptomları inceleyerek tespit edemeyebilirler, bu hücreler kanserin tekrar etmesine sebep olabilir bu duruma Minimal Kalıntı Hastalığı (MRD) denir. MRD, akım sitometrisi ve genetik çalışmalar ile teşhis edilebilir. Bu tür tedavilerin de kendi sınırlamaları vardır; örneğin pahalı olması ve eğitilmiş bir personele ihtiyaç duyulması gibi. Bu proje ile amacımız lösemi hücrelerinin yüzeyindeki belirteçleri antikor ile yakalamak (CD10, CD19 ve CD45) için düşük maliyetli bir mikroakışkan sistem geliştirmektir. Yakalama için antikor kaplı manyetik boncuklar kullanılmıştır. Son adımda; manyetik ayırma işleminden sonra lösemi hücreleri altın kareler üzerinde sabitlenmiştir. Böylece, hastaların tedavi süresince verdikleri dönütleri anlamak için hücreler sayılabilmiştir.
Development of breast cancer targeted, multifunctional cross-linked micelle nanocarriers
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2023) Bayram, Nazende Nur; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
In this thesis, we developed two different micelle-based nanocarriers, which are pH-responsive and core cross-linked micelle (CCMs), and specifically target HER2 receptor on breast cancer cells. Intracellularly degradable and stabilized micelles were prepared by core cross-linking and RAFT polymerization in the presence of an acid-sensitive cross-linker. Poly(OEGMA) and poly(SBMA) were used as shell parts of these micelles in order to compare the effect of hydrophilic coatings on nanocarrier characteristics. In the first design, we applied drug conjugation (Doxorubicin) with a cleavable linker while in the second design, we used the encapsulation method for drug loading. Targeted micelles were obtained by coupling of HER2-specific peptides (VSSTQDFP and LTVSPWY) and antibody (Herceptin) to POEGMA and poly (SBMA) based CCMs, respectively. These nanocarriers are designed to be stable in blood circulation but cleavable intracellulary to achieve controlled drug release. Nanocarriers were characterized structurally by FTIR and 1H-NMR spectroscopies for all synthesis and conjugation steps. Moreover, nanocarriers and drug-loaded formulations were investigated by Zetasizer, Nanosight, and TEM/SEM analysis. The results showed that designed nanocarriers have a very high potential for HER2-specific targeted drug release for the treatment of breast cancer. This thesis holds significant importance due to its successful demonstration of two distinct systems exhibiting high stability, pH sensitivity, and high selectivity for HER2-targeted therapy of breast cancer.
Discovering new pathogenic variants by In silico analysis
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2022) Zorluer, Ziya Furkan; 0000-0003-2206-6695; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Inherited diseases are health problems caused by one or more abnormalities in the genome. It can be caused by changes in a single gene (monogenic) or multiple genes (polygenic), or by a damage on chromosomes. Genetic variation is the differences in the DNA sequences that can be observed within a species or in alleles. Evaluation of genetic variants, together with reported phenotypic or pathogenic annotations from non-human organisms, facilitates the comparison of these variants with their human counterparts. In this work, we combined pathogenic and phenotypic annotations with variants, and these phenotypic orthologous variants from seven organisms can provide clues to the functional consequences of human genetic variants.
The effect of hedgehog inhibition and autophagy modulation on the proliferation and survival of AML cell lines /
(2019) ŞANSAÇAR, MERVE; AGÜ, Yaşam ve Doğa Bilimleri Fakültesi, Biyomühendislik Bölümü; ŞANSAÇAR, MERVE
Acute myeloid leukemia (AML) is a heterogeneous hematopoietic malignancy which occurs as a result of many chromosomal abnormalities such as translocations, deletions or insertions. Aberrant signaling pathways such as PI3K/AKT/mTOR, Notch and Hedgehog pathway have a role in the pathogenesis of AML. Hedgehog pathway (Hh) is a conserved signalling pathway that is important during embryogenesis. It crosstalks with other pathways and regulate autophagy, a cellular degradation and organelle turnover process. Several studies suggested that autophagy modulation could act as an escape mechanism in AML. Given the role of autophagy and Hh in AML, understanding the relationship between autophagy and Hh pathway is important to overcome the leukemic growth. Hence, we checked the effect of Hh inhibition using GANT61 on MOLM-13 and CMK cells using MTT cell viability assay. GANT61 led to a decrease in the both MOLM-13 and CMK cells. After that, we sought to understand the effect of autophagy modulation on CMK and MOLM-13 cell lines and we have found that autophagy inhibitors, NH4CI, Chloroquine(CQ), Hydroxychloroquine and Nocodazole lead to a decrease in the proliferation of CMK and MOLM-13 cell lines. However, PP242, an autophagy activator, had no effect on the proliferation of CMK and MOLM-13 cell lines. Combination treatment of autophagy modulators and GANT61 had a synergistic effect on MOLM-13 but not on CMK. GANT61 treatment increased autophagy in AML cell lines that were correlated with an increase in the expression of LC3B-II detected by western blotting. Also, combination treatment with nocodazole and GANT61 elevated that increase in LC3B-II in both MOLM-13 and CMK cell lines. AKT protein expression changed depending on the type of treatment and cell lines. In conclusion, targeting of Hh and autophagy is a promising therapy against MOLM-13 cell line but not against CMK.
The effect of histone deacetylase inhibition and autophagy modulation on the cholangiocarcinoma cells
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2022) Yenigül, Münevver; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Cholangiocarcinoma (CCA), also known as biliary tract cancer, is a heterogeneous group of malignancies formed by the differentiation of epithelial cells in the biliary tract. CCA is the second most common primary liver tumor and it has both an increasing rate and high mortality worldwide with its late diagnosis, refractory type, and aggressiveness. The effects of autophagy modulators and HDAC inhibitors in CCA are not fully known. This study is proposed a novel treatment approach with the combinational therapy of autophagy and HDAC inhibitors for CCA patients. In results obtained with alone HDACis, alone autophagy modulators, and combinations of HDACis and autophagy modulators, Nocodazole from autophagy modulators and MS-275 and Romidepsin from HDAC inhibitors showed a better synergistic effect on the TFK-1 and EGI-1 cell lines of the cholangiocarcinoma. In cell cycle analysis of the combination, was achieved arrest at the S phase and G2/M phase. In conclusion, this study highlights the important combination of HDAC inhibitors and autophagy modulators, which is a promising therapy in CCA.
THE EFFECT OF HISTONE DEACETYLASE INHIBITORS ON PTEN/PI3K/AKT/mTOR PATHWAY AND THE CARCINOGENESIS OF CHEMORESISTANT CHOLANGIOCARCINOMA AT A MOLECULAR LEVEL
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2021) Helin, Sağır; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Kolanjiokarsinom (CCA) agresif bir adenokarsinomdur ve ikinci en sık görülen birincil karaciğer tümörüdür. CCA gelişiminin kesin etiyolojisi hala net olarak tanımlanmamıştır. Mevcut kemoterapötik tedaviler, çoklu ilaç direnci nedeniyle etkili olmadığından, kemorezistant CCA yaygındır. Histon deasetilaz inhibitörleri (HDACis); umut verici antikanser karakteri göstermektedir ve HDAC işlevindeki düzensizlikler otofaji için önemli olan ve kemo-dirençli CCA'da bulunan yolaklar ile ilişkilidir, örneğin PTEN/PI3K/AKT/mTOR. Bu nedenle, sisplatine dirençli CCA hücre hatları ürettik ve SAHA, MS-275 ve Romidepsin yoluyla HDAC inhibisyonunun ve Nocodazol ve Klorokin ile otofaji inhibisyonunun etkisini kontrol ettik. Romidepsin ve Nocodazol'ün kombinasyon tedavisi sisplatine dirençli hücrelerin proliferasyonunu azalttı. Apoptotik analiz yapıldı ve sonuçlar erken apoptotik ve apoptotik hücre ölümündeki artışı kanıtladı. Ayrıca, hücre döngüsü analizi sonuçları, hücre döngüsünde durdurulma göstermiştir. Western blotlama ile PTEN, Histon H3 ve Asetillenmiş H3 protein ekspresyonlarını kontrol ettik. Sonuçlar, PTEN ekspresyon seviyesi ile HDAC inhibisyonu arasındaki olası ilişkiyi gösteriyordu. Direnç durumunda PTEN lokalizasyonu çok önemli olduğundan, immünofloresan boyama gerçekleştirdik ve hem hassas hem de sisplatine dirençli hücrelerde PTEN'in yerini tespit ettik. Sonuçlar, sisplatine dirençli hücrelerinde sitoplazmaya PTEN translokasyonunu gösteriyordu. Sonuç olarak, HDAC ve otofaji inhibisyonunun kombinasyon tedavisi, kemorezistan kolanjiokarsinomaya karşı umut verici bir tedavidir.
Identification of single domain antibodies against SARS-CoV-2 omicron variant via protein-protein docking approaches
(Abdullah Gül Üniversitesi, Fen Bilimleri Enstitüsü, 2022) İlmek, Özkan; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Omicron, became the dominant variant in 2022 in terms of spreading rate, has managed to evade from an immune system of patients due to its unique mutations. Single domain antibodies (sdAb) which are functionally important parts of conventional antibodies are commonly used for diagnosis and treatment. Although there are many sdAbs developed to combat coronavirus in recent years, their effectiveness against Omicron variant has not been sufficiently tested and the effect of mutations regarding neutralization level is not clear. In this study, structure modelling of 850 sdAb sequences obtained from previous studies were generated using AlphaFold 2 and effectiveness of these sdAbs against Omicron variant was tested via protein-protein docking approach. In the docking process, within a realistic approach, missing residues were completed into Spike protein PDB structures, and Spike protein homotrimer structure in closed state conformation was used. Finally, top 1000 and top 100 scores are determined as a threshold value for different protein-protein docking scoring functions such as HDOCK, PRODIGY and Bluues. sdAbs that have successful results for Omicron variant were listed. There were 4 sdAbs which exceed the threshold values after 2 different docking experiments against the Omicron variant. The scripting codes and methodological approach developed within this thesis can be used against new SARS-CoV-2 variants that may emerge in the future or other diseases.
IDENTIFICATION OF SURFACE PROTEOME OF B CELL ACUTE LYMPHOBLASTIC LEUKEMIA CELL LINE
(Abdullah Gül Üniversitesi / Fen Bilimleri Enstitüsü / Biyomühendislik Ana Bilim Dalı, 2022) Boyvat, Dudu; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
B-cell acute lymphoblastic leukemia is characterized by over and uncontrolled expression of B lymphocytes. B-ALL may occur as a result of aberrant cytosolic signal transduction and molecular abnormalities such as gene mutations, abnormal protein interactions, and an un-arrested cell cycle. Due to these abnormalities, surface proteins that compromised one-third of the proteome show different expressions compared to the healthy cells. These differences are currently in use for diagnostic and treatment approaches. Here, we aimed to isolate and identify the surface proteins of the CCRF-SB cell line to identify new, additional possible target antigens with the mass spectrometrybased proteomics approach using two different surface protein isolation strategies. The surface proteins of CCRF-SB cells were isolated with the surface biotinylation method and N-linked glycoprotein enrichment methods. With the biotinylation method, we isolated 782 proteins with 1% FDR. Gene Ontology Cellular Compartment analysis showed that 467 of these isolated proteins are annotated as ‘Membrane’. 263 of those proteins are annotated as ‘Extracellular Space’. These isolated cell surface proteins include HLA protein complexes and well-known CD19 surface markers. With the Nlinked glycosylation enrichment method 229 protein identified with 1% FDR rate. Gene Ontology Cellular Compartment analysis showed that 155 of these isolated proteins are annotated as ‘Membrane’, 132 of those proteins are annotated as ‘Extracellular Space’. Both methods identified different proteins from each other. This result showed that to map the surfaceome of CCRF-SB cell line, it is required to combine these two enrichment methods.
Immunomagnetic separation and immobilization of leukemia cells using cell surface markers /
(Abdullah Gül Üniversitesi, 2017) GERÇEK, TAYYİBE; AGÜ, Yaşam ve Doğa Bilimleri Fakültesi, Biyomühendislik Bölümü; GERÇEK, TAYYİBE
Acute Lympoblastic Leukemia, shortly ALL, especially B-precursor Acute Lymphoblastic Leukemia is the most common blood malignancies among childhood cancers. There are different kinds of leukemia treatments but after those therapies, there is likelihood of relapsing the cancer in years because of remained cancer cells in the patient’s body. Nevertheless remained cancer cells in the body after therapy cannot be seen in the routine clinical follow-up tests. Those kind of diseases like leukemia are named as Minimal Residual Disease (MRD). Currently, there are only two ways to detect Minimal Residual Disease. These are flow cytometry and real-time quantified polymerase chain reaction (RQ-PCR). However many different labs have these Flow Cytometry and RQ-PCR, both devices should be calibrated to detect MRD cells. Nowadays there is a consensus about requiring MRD detection but still it is discussed by the authorities about when and how it should be done. The ultimate aim of this project is to produce a chip that can detect MRD cells. With these work we try to capture leukemic cells by using magnetic beads in nano and microsize. These magnetic beads are coated with CD19 and CD45 antibody markers that are in the membrane of leukemia cells. Next step after capturing them with magnetic beads is to immobilize them onto a surface. Golden surfaces are used and functionalized by antibodies. Thus an immunosandwich structure is occured and the cells are immobilized.
Influence of 3D bioprinting parameters on printability and mechanical behavior of the PCL scaffolds
(Abdullah Gül Üniversitesi / Fen Bilimleri Enstitüsü, 2023) Saniye Aylin, Ceylan; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı
Polycaprolactone (PCL) is a synthetic polymer that exhibits desirable properties such as biodegradability, tolerable mechanical properties, and biocompatibility for a diverse range of tissue engineering applications. In this study, we analyzed the effects of polymer concentration (10%, 25%, 50% and 75% w/v), solvent effect (dichloromethane, chloroform and acetic acid), and device parameters (pressure, speed, nozzle-surface distance, nozzle gauge, infill density) on printed scaffolds fabricated through 3D Bioprinting. Scanning electron microscopy (SEM) and optical microscopy were used to assess printability, and uniaxial tensile testing was performed to evaluate mechanical behavior. The aim of this study was to investigate the effects of different printing speeds (5 mm/s, 10 mm/s, and 15 mm/s) on the mechanical properties of PCL_DCM and PCL_CF scaffolds. The scaffolds printed at the lowest speed exhibited the highest ultimate tensile strength (UTS) values. Scaffolds printed at 5 mm/s with the highest printing pressure (480 kPa) demonstrated a remarkably high Young's modulus of 39.69 MPa and a UTS value of 6.4 for PCL_DCM, as well as Young's modulus of 26.80 MPa and a UTS value of 6.3 MPa for PCL_CF. Additionally, we investigated the influence of polymer concentrations (50% and 75%) and infill densities (50%, 70%, and 90%). The results showed that increasing the infill density and using a lower concentration (50%) led to improvements in Young's modulus and UTS values for both PCL_DCM and PCL_CF scaffolds. These results highlight the importance of carefully controlling printing parameters to optimize the mechanical properties of the printed scaffolds.
The inhibition of autophagy and hedgehog pathway leads to a decrease in the proliferation of cholangiocarcinoma
(Abdullah Gül Üniversitesi, 2019) AKTAŞ, NİHAN; AGÜ, Fen Bilimleri Enstitüsü, Biyomühendislik Ana Bilim Dalı; AKTAŞ, NİHAN
Cholangiocarcinoma (CCA) is the second most common liver cancer type. The median survival rate of CCA patients is really low. Aberrant signaling pathways such as PI3K/AKT/mTOR pathway could be main drivers in CCA pathogenesis. Hedgehog (Hh) pathway is also dysregulated in several carcinomas including CCA. It regulates and crosstalks with autophagy, which is a lysosomal degradation process. There is no study showing the crosstalk between Hh pathway and autophagy in the context of CCA. Since both autophagy and Hh pathways are dysregulated in CCA, better understanding of how they crosstalk with each other and contribute to CCA pathogenesis is important. Considering this crosstalk between Hh pathway and autophagy, we conducted a combination treatment comprising Hh and autophagy pathway inhibitors in EGI-1 and TFK-1 CCA cell lines. In our study, we firstly checked anti-proliferative effects of Hh pathway inhibitor, GANT61, and different autophagy blockers using MTT and Annexin V assay and cell cycle analysis. After determination of IC30 of GANT61 (15 uM), chloroquine (25 uM for TFK-1 and 50 uM for EGI-1), and nocodazole (0.2 uM for EGI-1 and 0.4 uM for TFK-1), we conducted combination experiments. When we inhibit Hh pathway with targeting different steps of autophagy, we observed that proliferation of both EGI-1 and TFK-1 cells decreased compared to single treatments. After that, we checked the expression of autophagy-related LC3B protein and Akt, a negative regulator of autophagy, using western blotting after single treatments and combinational treatments. Based on the change in LC3B and Akt expression, we also concluded that, inhibition of autophagy with Hh pathway either induce or inhibit autophagy depends on the administered treatments. This study highlights the importance of deciphering the exact mechanisms that control autophagy in CCA, thus leading to better treatment.

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