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Kanser tedavisi amaçlı nano parçacıkların sentezi ve değerlendirilmesi
Nanotechnology-based drug delivery systems have emerged as promising tools in cancer therapy, offering improved drug targeting, reduced systemic toxicity, and enhanced therapeutic efficiency. Among various nanomaterials, iron oxide nanoparticles (IONPs) are widely recognized for their biocompatibility, magnetic responsiveness, and surface modification potential. In this study, quercetin-loaded iron oxide nanoparticles (Q-IONPs) were synthesized and evaluated for their physicochemical properties and cytotoxicity against the MCF-7 breast cancer cell line. The nanoparticles were prepared using the co-precipitation method by mixing ferrous sulfate (FeSO₄) and ferric chloride (FeCl₃), followed by the addition of sodium hydroxide (NaOH) to form iron oxide nanoparticles. Quercetin was subsequently incorporated to enhance the formulation's therapeutic efficacy. Particle size analysis revealed an average hydrodynamic diameter of 394.2 ± 6.5 nm, confirming the formation of nanosized particles. Preliminary pH monitoring suggested good stability of the nanoparticles under near-physiological conditions. The cytotoxicity of Q-IONPs will be further assessed against MCF-7 cells using the MTT assay to evaluate their potential as anticancer agents. The outcome of this research is expected to provide valuable insights into the development of quercetin-based iron oxide nanoparticles as a promising nanocarrier system for targeted cancer therapy.
FGD-pet görüntüleme kullanarak alzheimer hastaliğinin radyomik analizi
Alzheimer's disease (AD) demands more advanced and personalized diagnostic methods. This dissertation proposes a radiomics-driven framework using FDG-PET to enhance the diagnosis, staging, and risk evaluation of AD. By integrating image-derived features and machine learning, we aim to facilitate early, non-invasive, and individualized assessment. We developed a fully automated brain radiomics platform capable of distinguishing between CN, MCI, and AD individuals. The hippocampus, entorhinal cortex, and amygdala emerged as key discriminative regions. Our simplified FDG-PET model achieved an AUC of 0.853 for predicting amyloid positivity, a critical pathological hallmark of AD, using features from the hippocampus, inferior parietal lobule, and isthmus cingulate. The platform was further extended for ApoE4 genotype prediction, achieving an AUC of 0.945 with features extracted from the hippocampus, amygdala, thalamus, and pars orbitalis. We also investigated hippocampus-amygdala connectivity, identifying robust biomarkers such as Shape Mesh Volume and GLDM Small Dependence Low Gray Level Emphasis (AUC = 0.88). Subregional analysis of hippocampal and amygdaloid structures revealed additional radiomic features, including GLRLM Long Run Emphasis and GLDM Small Dependence Emphasis, which differentiate AD from MCI and CN, indicating early microstructural and metabolic changes prior to visible atrophy. Our findings establish FDG-PET radiomics as a reliable, non-invasive imaging biomarker for diagnosing and monitoring AD. The proposed framework enables risk stratification and supports clinical decision-making, paving the way for preventive and personalized AD management.
Yumuşak kısım yapısı ve sert kısım miktarının poliüretanürelerin yüzey ve yığın özellikleri üzerindeki etkisi
Due to their interesting combination of bulk and surface properties segmented thermoplastic polyurethanes (TPU), polyureas and polyurethaneureas (TPUU) find wide range of applications in many diverse fields. One of the emerging applications of TPUUs include their usage as biomaterials in blood contacting applications. This is mainly due to the possibility of designing and synthesizing TPUUs with controlled bulk and surface properties that possess very good blood and tissue compatibility. [1] The main goals of this study were the investigation of the influence of; (i) soft segment (SS) structure, (ii) SS molecular weight and (iii) hard segment (HS) content on the morphology and bulk and surface properties of TPUUs. In addition, another aim was to investigate the relationship between the surface energies of copolymers and their resistance to biofilm formation. A large number of segmented thermoplastic poly(urethaneurea)s (TPUU), polyurethanes (TPU) and polyureas (PU) based on five different SS were synthesized by using the conventional two step "prepolymer" polymerization method (or in one step if the polymer was non-chain extended) and characterized. Bis(4-isocyanatocyclohexyl)methane (HMDI) was used as the diisocyanate and 2-methyl-1,5-diaminopentane (MDAP) or 1,4 butanediol (BD) was used as the chain extender. Five different SS which were used in this study included poly(ethylene oxide) glycol (PEO), poly(propylene oxide) glycol (PPO), poly(tetramethylene oxide) glycol (PTMO), aminopropyl and hydroxyhexyl terminated polydimethylsiloxane (PDMS) and a hydroxy terminated polyfluoroether (PFE) oligomer (Fluorolink E10-H®). HS contents of the copolymers were generally kept constant at 20 and 30% by weight with a few exception. To investigate the influence of the molecular weight, SS oligomers with two different molecular weights of 1,000 and 2,000 g/mol were used during the synthesis. The bulk and surface properties of all polymers were characterized by using a large number of techniques, which included; ATR-FTIR (Attenuated Total Reflection Fourier Transform Infrared), DSC (Differential Scanning Calorimetry), SAXS (Small Angle X-ray Scattering) and stress strain analysis, AFM (Atomic Force Microscopy), XPS (X-ray Photoelectron Spectroscopy) and static water contact angle measurements. Selected copolymers with different surface energies were also tested for biofilm formation. Results obtained indicated extensive biofilm formation on all samples regardless of their surface properties. Characterization results showed that polyether based copolymers, especially those with PEO SS, displayed poor microphase separation when compared with PDMS and PFE based copolymers. This is expected since the polyethers used in this study have higher solubility parameters and ether groups can form hydrogen bonding with urethane and urea groups, which is not possible in case of PDMS and PFE. Surface characterization by contact angle measurements showed dramatic differences depending on the SS structure and molecular weight. As indicated by XPS studies, in general the soft segments tend to migrate to the polymer surface and affect the hydrophobicity and hydrophilicity of the copolymer depending on their solubility parameters. The extent of biofilm formation was not affected by the surface energy differences within the copolymers. Many alternatives for the bacterium sticking mechanism on surfaces have been proposed in the literature. [2] Some studies suggest that bacterium tend to stick on hydrophilic surfaces[3], while other studies suggest that bacterium tend to stick on hydrophobic surfaces [2]. It should also be noted that the sticking mechanism of every strain of bacterium is different [4], which makes this concept even more challenging to explain. Therefore our study is a contribution to the literature on this topic and suggests that the sticking mechanism of Staphylococcus aureus 700698 strain of bacterium is not affected by the surface of the polymer being hydrophilic or hydrophobic.
CRISPR-Cas9 aracılığı ile kromatin modifiye eden genlerin insan hücre yeniden programlanmasında incelenmesi
Reprogramming of somatic cells to pluripotency via four transcription factors, Oct4, Sox2, Klf4, and c-Myc (OSKM) involves extensive remodeling of the epigenome which is carried out by a number of chromatin modifiers. A number of these genes have been studied in the context of reprogramming and defined as barriers or essential regulators of this process. However, established protocols limit a comprehensive examination due to the susceptibility of shRNA vectors to proviral silencing which accompanies reprogramming, and the unavailability of human knock-out cell lines. Therefore, we hypothesized that pooled CRISPR-Cas9 knock-out libraries can enable systematic investigation of chromatin modifiers and reveal their role in different steps of reprogramming by enabling permanent gene modifications. To this end, we constructed a CRISPR-Cas9 knock-out library targeting the 247 chromatin modifiers including chromatin remodelers, writers, readers and erasers of histone and DNA modifications. We utilized this library to generate heterogeneous cell populations harboring putative chromatin modifier knock-outs, which were then reprogrammed with the delivery of OSKM. FACS-based enrichment of TRA-1-60 positive cells, and low copy exogenous retroviral EGFP silencing was utilized to collect emerging iPSCs at different time points of reprogramming. The abundance of CRISPR guide RNAs in isolated cells were determined by next generation sequencing. Analysis of relative enrichment of specific guide RNAs confirmed the majority of the known reprogramming regulators and identified novel stage specific barriers and essential regulators for human somatic cell reprogramming. These include PAXIP1, BRD2 and USP22 as barriers, and CHAF1A, KDM8 and PRMT5 as essential regulators. In addition, functional analysis based on class of genes revealed that various MLL complexes act as barriers, and BAD, SWI/SNF, and MOZ/MORF complexes act as essential regulators for reprogramming. Based on the screen results, stage specific combinatorial use of small molecule inhibitors enabled accelerated and more synchronous efficient reprogramming. Taken together this study establishes roadblocks of reprogramming through chromatin landscape, and can guide more efficient reprogramming strategies to be formulated.
Medulloblastoma'da kromatin değiştirici enzimlerin fonksiyonlarının incelenmesi
Medulloblastoma is the most common pediatric brain cancer and it consists of four main molecular subgroups, which are Wingless (WNT), Sonic Hedgehog (SHH), Group 3 and Group 4. These subgroups have different transcriptional, cytogenetic and mutational spectra, making medulloblastoma a complex disease. Current therapy protocols for medulloblastoma include surgical resection, craniospinal irradiation and chemotherapy. Although current treatments result in high overall survival, many survivors experience severe neurological disorders. Therefore, targeted and more efficient therapies are necessary for the treatment of medulloblastoma. Genomic studies revealed that several chromatin modifying enzymes (CMEs) such as histone methyltransferases (HMTs), histone demethylases (HDMs), histone acetyltransferases (HATs) and histone deacetylases (HDACs) are mutated and/or differentially expressed in medulloblastoma. However, the roles of CMEs in the initiation and progression of medulloblastomas are ill-defined. To this end, we mainly focused on the epigenetics of medulloblastoma to develop novel and more effective therapeutic options. Firstly, we investigate the roles of chromatin modifiers in medulloblastoma by utilizing a chemical library targeting different chromatin modifying enzymes. Our screen revealed some potential inhibitors, namely GSK-J4, IOX-1, Belinostat, Vorinostat, 5-Azacytidine, ML324, SGC0946, Trichostatin A and Chaetocin, that induced cell death in medulloblastoma cells significantly. To investigate whether these drugs are specific to cancer cells, we tested selected drugs in a dose-dependent manner in non-malignant BJ Fibroblasts and revealed that they were relatively non-toxic to normal cells. Since the roles of histone demethylases are barely defined in medulloblastoma, we focused on KDM6A and KDM6B, which are targets of GSK-J4 and IOX-1. To this end, we targeted KDM6A and KDM6B by shRNAs and examined whether their loss changed the growth rate medulloblastoma cells. However, we did not observe any changes in the proliferation rate of MB cells upon KDM6A and/or KDM6B loss. Although many medulloblastoma tumors respond to chemotherapy well, there are still many medulloblastoma patients who do not respond to standard of care chemotherapy. As it has been identified that many chromatin modifiers are mutated and/or differentially expressed in the most aggressive and metastatic subgroups (Group 3 and Group 4), we speculated whether chromatin modifiers play role in resistance of medulloblastoma cells to chemotherapeutic agents. From those, vincristine is the main chemotherapeutic agent used in different treatment protocols, we decided to establish a vincristine-resistant medulloblastoma cell line by dose-escalation method to elucidate the function of CMEs in therapy resistance. We performed another drug screen in parental and newly established vincristine-resistant cells and identified that 8 different HDAC inhibitors (Trichostatin A, Rocilinostat, CXD101, Tubastatin A HCl, Belinostat, Romidepsin, PCI-24781 and Mocetinostat), 2 histone demethylase inhibitors (IOX-1 and KDOBA67), 2 kinase inhibitors (5-Iodotubercidin, SGI-1776), MAZ1805 (Halofuginol) and MAZ1392 induced cell death both in parental and vincristine-resistant cells significantly. Besides, we performed combination treatment with epigenetic drugs and vincristine and discovered that A-395, CBP/BRD4, GSK-J5, GSK343, GSK864, LLY-507, OF-1, SGC-CBP30, SRT1720 and UNC2400 induced cell death of resistant populations when they were combined with vincristine.
Nakil edilen adacıkların vücuttan reddedilmesini önlemek amacıyla bağışıklık sisteminden ayrı tutulabilen bir bölgesel mikroortam dizayn edilmesi
The development of tolerance induction approaches via immune engineering that can restore and/or replace non-functional tissues and organs represents the leading front of emerging regenerative therapies. Type-1 diabetes (T1D) is an organ-specific autoimmune disease characterized by destruction of pancreatic β cells, which are insulin-secreting cells by autoreactive T cells and other immune cells. Loss of β-cell, thus insulin, makes patients dependent on exogenous insulin or to overcome this need; transplantation of pancreas or intact islets. One of the promising approaches used for treatment of diabetes is the transplantation of islets; however, it also comes with its pitfalls. First and most obvious one is the limitation of donor source. In addition, transplantation of islets or entire pancreas requires suppression of the immune system to prevent graft rejection. This suppression is achieved through immune suppressor drugs, which leaves the body defenseless against infections and increases its susceptibility to other complications such as cancer. Considering all these limitations, immunotherapeutic strategies have focused on restoring immunologic self-tolerance, thus removing the problem at it roots; preventing β cell destruction by patients's own immune system. The main focus of this approach is regulatory T cells (Tregs), which are essential cells in suppression of autoreactive immune responses and maintenance of self-tolerance. Stellate cells (SCs) have various effects on the immune system such as recruitment of Tregs and induction of T cell apoptosis. Besides, they can promote vascularization, secreting vascular endothelium growth factor (VEGF). Some chemokines are also key modulators in recruitment of Tregs. Macrophage-derived chemokine or C-C motif chemokine ligand 22 (MDC/CCL22) is one of the novel chemokines used for Treg recruitment by binding to CCR4 receptor on their membrane. In this study, we designed an immune privileged microenvironment around implantable insulin secreting islets site to provide local graft tolerance and to overcome limitations associated with donor cells. We focused on achieving local immunosuppression through overexpression of CCL22 proteins by SCs, which recruited immunosuppressive Tregs. We prepared insulin-secreting pseudoislets through aggregation of mouse insulinoma 6 (MIN6) cells as a model system to mimic naïve islet morphology. Our results demonstrated that transfected SCs can secrete CCL22 and recruit a population of Tregs towards the implant in vivo. This study is promising to provide fundamental understanding of the SC-islet interaction, ligand synthesis and transport from stellate cells at the graft site for ensuring local immune tolerance to target Type I diabetes. Our results also establish a new paradigm for creating tolerable grafts for other chronic diseases such as diabetes, anemia, cancer, CNS diseases and advance the science of graft tolerance.
K-Ras proteini ve onkojenik mutasyonları üzerine kapsamlı bir çalışma: Dinamik bir bakış açısı
K-Ras is the most frequently mutated protein in human cancers driving cancer initiation, progression and drug resistance, directly leading to nearly a million deaths per year. Sadly, there are still no drugs in that directly target mutant K-Ras in the clinic. Recent studies utilizing dynamics information show promising results for selectively targeting it. However, despite extensive characterization, the regulatory mechanisms of K-Ras dynamics remain elusive. Since protein function is related to its dynamics, understanding these mechanisms can present novel opportunities for identifying target sites on mutant K-Ras surface. In this work, we investigate the regulation mechanisms of K-Ras dynamics and the effects of nucleotide binding and mutations on these mechanisms using extensive molecular dynamics (MD) simulations. We applied different MD simulation data analysis techniques to compare the dynamic characteristics of both active and inactive forms of wild-type K-RasWT and mutant K-RasG12D, the most recurrent mutant in cancer patients. Our results on K-RasWT showed excellent agreement with experimental data and served as a reference point for K-RasG12D analysis. Then, we demonstrated how G12D mutation induces structural and conformational changes that result in characteristic correlated motions in active K-RasG12D. Moreover, we developed a novel conditional time-delayed correlations (CTC) based approach to predict causal relationships in regulation of K-Ras dynamics. CTC analysis identified the regulatory sites that control K-Ras dynamics. Finally, we identified a novel drug target pocket in active K-RasG12D and screened a small molecule library against it using docking techniques. Our study draws a complete picture of the regulation of K-Ras dynamics. We anticipate that the identified regulatory sites on active K-RasG12D can present novel opportunities for direct targeting of K-RasG12D in future drug discovery efforts.
PRISM'in geçmiş CAPRI turları ile değerlendirilmesi
Proteins are key elements of a cell to perform the wide range of molecular and cellular activity. Proteins perform their function through binding to other proteins, DNA, RNA, and small molecules. Therefore, predicting how a protein interacts with its binding partners is one of the most important objectives of structural biology. As a result of the improvements in experimental structure determination methods, the number of individual protein structures in PDB has increased vastly. However, the number of complex structures does not increase as fast as the individual proteins which creates demand for new approaches to predict complexes from the individual protein structures. Currently, there are a lot of computational approaches to predict the complex structures. Critical Assessment of Prediction of Interactions (CAPRI) is a well-known community-wide experiment with the purpose of establishing a routine which allows testing the performance of several different docking algorithms created. The success rate of predicted structures is verified by the several evaluation criteria determined by the CAPRI association. Those criteria are interface and ligand rmsds (I-rmsd, L-rmsd), native residue contacts and number of clashes. In this study, I assessed the performance of PRISM (Protein Interaction by Structural Matching) using CAPRI evaluation criteria. The main objectives were to determine how much PRISM is successful in predicting the complex structures of available CAPRI targets and to force the limits of PRISM by stretching the parameters of PRISM. PRISM could not predict the correct complex structures for 33% of targets majority of which correspond to homodimers and enzyme/inhibitor complexes. The results also indicate that considering just the structures with the negative energy score results in the loss of 36% of successful predictions implying a problem in the scoring function. For further testing, RosettaDock was used as an alternative scoring function. Both scoring approaches yielded a correlation. To increase the success rate, some parameters of PRISM were changed however, no significant improvement has been achieved.
Kalp destek cihazlarının tasarımı ve eniyileştirilmesi: iHeart VAD ve iATVA
Heart failure, some congenital heart defects, and some circulatory disorders are diseases which are treated or alleviated with mechanical circulatory support devices. Even though the ideal therapy for heart failure is a cardiac transplant, due to the donor shortage, Left Ventricular Assist Devices has emerged. As a mechanical pump, an LVAD pumps the blood from the left ventricle to the aorta for supporting the heart. Similarly, for the treatment of Fontan patients with a univentricular heart, some experimental mechanical circulatory support devices have been proposed in the literature. These devices are called Fontan Ventricle Assist Device (FVAD). This work comprised the design, development, and experiments of İstanbul Heart VAD (iHeart VAD) and integrated Aortic-Turbine Venous-Assist (iATVA) which are a novel centrifugal LVAD and a novel FVAD (a centrifugal turbine-pump couple), respectively. Both devices were designed following the turbomachinery theory. For the iHeart VAD, this work includes computer simulations, hydraulics tests, blood tests and acute animal tests. For the iATVA, this work focuses on hydraulic tests and cardiovascular mock-loop tests. For both devices, chronical animal tests are planned for the near future.
Doku mühendisliği ve rejeneratif tıp uygulamaları için biyoçözünür iskele yapılarının tasarımı, üretimi ve karakterizasyonu
Biocompatible and biodegradable scaffolds for tissue engineering and regenerative medicine applications were developed. Bone and skin tissue engineering applications were focused throughout the study. Biomaterials such as hydroxyapatite (HA), chitosan (CS) and poly(lactic acid) (PLA) were used to fabricate scaffolds for potential bone regeneration applications. HA was synthesized through different production routes via sol-gel process. The process was critically analyzed and evaluated for synthesis of pure HA. Nano-composite scaffolds of CS/HA were fabricated through simple solvent casting method. Selection of proper solvent media (formic acid, a green solvent) produced stable and well defined dispersion of HA that resulted in intercalated structural composites. Furthermore, 3D PLA scaffolds were developed through fused deposition modeling and CS/HA were incorporated in these scaffolds. Formic acid enabled embedding of CS/HA through chemical etching of PLA surfaces. These scaffolds were analyzed in-vitro by seeding U2-OS human osteosarcoma cells. Scaffolds modified with CS/HA showed higher cell proliferation as compared to unmodified ones. The scaffolds developed in this study show great potential for use as substrates for bone tissue engineering applications. For skin tissue engineering applications, polycaprolactone (PCL) and the blend of a polyurethane (PU) prepared through stoichiometric reaction of PCL (Mn=2000 g/mol) and 1,6-hexamethylene diisocyanate (HDI) and silk fibroin (SF) were used. PCL was synthesized through ring opening polymerization of Ɛ-caprolactone in the presence of T-9 catalyst and 1,2-bis(3-aminopropoxy)ethane as initiator. SF was obtained by degumming of silk cocoons through a novel process developed in this study and subsequent dissolution. Nano-fibrous scaffolds of PCL and PU with and without different amounts of SF were prepared through a "green" electrospinning process in formic acid, which is a low toxicity Q3C class 3 solvent, rather than commonly used class 2 solvents such as tetrahydrofuran (THF) and dimethylformamide (DMF). Successful addition of SF and its effect on these scaffolds was analyzed through various characterization techniques. In-vitro analysis was done by seeding Human BJ and NIH/3T3-Mouse embryo fibroblast cells on PCL, PCL/SF and PU, PU/SF scaffolds respectively. ATP and MTT cytotoxicity analyses revealed significant enhanced proliferation of fibroblast cells as compared to pristine PCL and PU scaffolds. We believe these novel composite scaffolds with good mechanical properties and bioactivity can be used for skin tissue engineering applications.
Memeli hücrelerde protokadherinlerin rollerinin araştırılması
Cancer is one of the leading causes of morbidity and mortality around the globe. It is a complex disease characterized by hyperproliferation, genetic instability causing cumulative mutations, and at advanced stages spread to secondary sites, through a process known as metastasis. Metastasis is directly dependent on cancer cells' ability to migrate and, for an intervention in metastasis, it is pertinent to understand the molecular details of cell migration. In the current thesis, we attempted to understand the roles of a cell surface protein protocadherin-7 (PCDH7) in cell migration. Previous work from our group has shown PCDH7 as a cell-cycle regulated membrane protein. In current thesis, we characterized the effects of perturbation of PCDH7 expression on cell migration of mammalian cells in culture, and its role in the cortical migration of neuronal progenitors during embryonic development using in-utero electrophoresis. We also characterized its sub-cellular localization using fixed sample imaging and live-cell imaging, which highlighted its association with cellular compartments directly involved in cell migration dynamics. To further characterize molecular mechanisms behind PCDH7's role in cell migration, we used a quantitative proteomics approach to compare global changes upon PCDH7's knockout which outlines a plethora of upregulated and downregulated proteins. Many of these proteins have previously been shown to be associated with cellular compartments involved in cell migration or have a molecular function characterized in the regulation of cell migration. Taken together, our work outlines a critical role of PCDH7 in cell migration, association with migration-associated subcellular compartments, and possible molecular players in its role. Our work is a step towards recognizing PCDH7 as a promising therapeutic target for control of cancer metastasis.
Lazer işleme ile geliştirilen mikroakışkan çip içerisinde su damlalarının boyutlarına bağlı sınıflandırılması ve anjiyojenez çalışmalarına yönelik mikroakışkan çip tasarımı
We demonstrate an autonomous, high throughput and rigorous mechanism for sorting of droplets with different dimensions over inclined (10 degree), shallow (700 nm) and narrow (22 and 30 µm) guiding tracks defined by laser micromachining. We fabricated a microfluidic device containing two independent T-junctions and inlets for droplet generation allowing broad range of size and speed tuning as well as droplet merging before entering the Hele-Shaw channel hosting the guiding track. In the first part, we investigate partial guiding of different sizes of droplets under same host liquid flow conditions based on the three forces acting on the droplets namely drag force, frictional force and confinement force. For a bigger droplet, drag force being a quadratic function of droplet diameter dominates the confinement force causing the droplets leaving the inclined track earlier covering less vertical distance and guided partially. Secondly, as the speed of the droplet is increased by increasing the channel flow rate while keeping its size constant, higher drag force is exerted on the droplet once again causing partial guiding. Finally, we demonstrate sorting of smaller guided droplets coflowing with bigger unguided droplets as a result of merging from two inlets in the common tapered region just before entering the Hele-Shaw channel. For all the experiments, we have considered two chip designs with different guiding track widths (22 µm and 30 µm) and found that the droplets undergo stronger guiding for the case of wider track because of the higher confinement force. All experimental results are correlated with analytical model results incorporating droplet size, speed, interfacial tension, contact angle and realistic droplet shape by finite volume method. Microfluidic technology combined with tissue engineering has significantly increased the progress in cell biology and helped in understanding the physiological and pathophysiological transitions. PDMS due to its bio-compatibility, optical transparency and elasticity is widely used in fabrication of microfluidic devices for rapid prototyping to understand the 3D physiological microenvironment. We designed a novel microfluidic chip that incorporates a collagen hydrogel scaffold for 3D cell growth and enables controlled diffusion of medium through it. Thus, it is a convenient design that incorporates a hydrogel that mimicks the cellular microenvironment. Our design consists three parallel microchannels in contact with each other. By the unique height design, the hydrogel solution stays in the defined region and takes the shape of the collagen loading channel due to capillary action and surface tension effect. Angiogenesis; the formation of new capillaries from existing ones is mediated by endothelial cells. The hydrogel provides a porous 3D support which mediates the exchange of O2 and nutrients. We studied the response of endothelial cells by stimulating them with vascular endothelial growth factor (VEGF) that diffuses through the 3D hydrogel scaffold from biochemical channel to the cell channel inside a microfluidic chip. Our novel design provides a favorable microenvironment for Endothelial cells to grow sprouts under the influence of stimuli by having a wide contact area between the collagen loading channel and the side channels. We found that our design works for studying sprouting angiogenesis and can be used for mimicking 3D microenvironments. We also designed another microfluidic chip that will be used as an indicator of the angiogenic potential of a tumor. It incorporates cells trapping wells in one of the side channels for trapping tumor cells and can be grown to become tumor spheroids in a chip. The side channel enables the formation of spheroids from injected tumor cells on the chip. This design is different from the previous design in a sense that no stimulus in introduced directly in the chip rather the growth factors and molecules secreted by tumor spheroids in its microenvironment will trigger endothelial sprouts. Different type of tumors secrete different growth factors and molecules. By using our designed microfluidic chip the angiogenic tendency of different tumors spheroids can be studied.
Renshaw hücrelerinin sinaptik fizyolojisinin incelenmesi ve bu hücrelerin motor nöron hasarındaki rolü
Among the circuits of interneurons within the spinal cord, the Renshaw's inhibitory circuit is known to directly feedback on alpha-motoneurons. Inhibitory Renshaw cells (RCs) are located in the ventral horn of the spinal cord. Their unique features such as receiving direct cholinergic input from alpha-motoneurons and their ability for burst-firing provide a suitable model for studying circuits of the spinal cord. Although RC circuitry has been investigated for more than 70 years, its exact function has yet to be discovered due to the limitation of techniques. Therefore, the aim of this thesis was to develop a reliable method to investigate the characteristics of recurrent inhibition in healthy people and pathophysiology of RCs in patients who suffer from amyotrophic lateral sclerosis (ALS). After optimizing the stimulation parameters of the motor axons in a reproducible manner, we recorded single motor units and analyzed the individual sets of action potentials using probability as well as frequency-based analysis. For this purpose, the largest motor axons in the tibial nerve were electrically stimulated to activate RCs antidromically and the smallest motor units innervating soleus muscle were recorded using intramuscular electromyography in healthy people and ALS patients. The frequency methods indicated that the duration of the recurrent inhibition was between 30 to 55 ms in healthy people depending on the background firing rate of single motor units which were inversely proportional. Moreover, we found an evidence to support a longer inhibition on smaller diameter alpha-motoneurons compared to larger ones. On the other hand, the efficiency of the RCs was extremely reduced in the lumbar-affected ALS patients, i.e. the recurrent inhibition duration was about 11 ms, one-third to one-fifth of the healthy people. These findings indicate a high correlation with the controlled animal studies, and that this method can be used as a reliable tool to shed more light on the functions and dysfunctions of the RC system in humans. Moreover, the outcome from this thesis provides evidence about the involvement of last-order interneurons, e.g. RCs, in the development of movement disorders, such as ALS.
Silisyum kıvrılan dağıtılmış geribildirimli yapıların tümleşik fotonik incelemeri: Biyolojik ligandlar için örnek bir düzen
Halka çınlaçları, dağıtılmış Bragg yansıtıcıları (DBY), Mach-Zehnder girişimölçerleri (MZG) ile bunların ardışık bağlanmış sürümleri gibi tümleşik fotonik yapılar virüs, protein, biyoimleyici, deoksiribonükleik asit (DNA), ve mikroribonükleik asit (mikroRNA) gibi örneklerin biyoalgılanması yaygın olarak çalışılmış, ve başarıyla denenmiştir. Biz biyoalgılama uygulamaları için yeni bir tür yalıtkan-üzeri-silisyum (YÜS) kıvrımlı dağıtılmış geribildirim (KDG), antisimetrik KDG (AKDG), ile simetrik KDG (SKDG) yapılarını öneriyoruz. Bağdaşmak için taşıyıcı dalgakılavuzu gerektiren halka çınlaçlarının tersine, bu tektaş yapıları tasarlamak için bir enine elektrik (EE) kutuplu silisyum dalgakılavuzu kullanılmıştır. Bu tasarımların yapıtaşı kıvrımlı halka yansıtıcıdır (KHY). İzgesel kip ayrımı gösteren, KDG yapısını oluşturmak için 3 tane KHY ardışık olarak kullanılmıştır. AKDG yapısı için 4 tane içiçe geçmiş KHY kullanılmıştır. AKDG yapısı izgesinde, C = 0.09 bağlaşım katsayısı için izgesel kip ayrımı; 0.27 < C < 0.51 için elektromanyetik irkiltilmiş saydamlık (EIS)-benzeri tepeler görülmüştür. SKDG yapısı 5 tane içiçe geçmiş KHY kullanılmıştır. SKDG izgesi, C = 0.24 için kip ayrımlı Fano çınlaması; 0.78 < C < 0.94 için elektromanyetik irkiltilmiş saydamlık (EIS)-benzeri tepeler gösterir. SKDG yapısının Fano çizgibiçimli deneysel izgesi 26 dBm sönümleme oranı ve 368 dBm/nm eğim oranı gösterir. Bir çınlaç algılayıcının öz algılama sınırı λ/QS olarak gösterilebilir; burada λ boş uzay dalgaboyu, Q çınlacın nitelik katsayısı, S duyarlılıktır. KDG yapısının izgesel ayrılmış tepeleri ile, AKDG, ve SKDG yapılarının elektromanyetik irkiltilmiş saydamlık (EIS)-benzeri tepeleri Q = 50000 düzeyinde nitelik katsayısı gösterir; öyle ki 1550 nm dalgaboyundaki biyomolekül soğurması katılmadığında, λ = 1550 nm dalgaboyunda, S = 50 nm/RIU duyarlılık değerinde, silisyum fotonik yapılarımızın kuramsal öz sınırı 0.0006 [RIU] olarak bulunur. Var olan tümleşik fotonik devre yapılarına ek olarak, bizim sunduğumuz yeni tektaş yalıtkan-üzeri-silisyum (YÜS) fotonik yapılar biyoalgılama uygulamalarında ümit vermektedir.
Uyarana duyarlı polimerlerin kendiliğinden yapılanarak oluşturduğu yapılar ve bu yapıların kontrollü salınım uygulamaları
Self-assembled nanostructures of stimuli-responsive polymers combine the advantages of responding to environmental changes with specific properties of the nano-systems. Effect of various physicochemical factors on self-assembly process is important in designing nano-systems for controlled release applications. This thesis reports the formation and controlled release applications of hydrogen-bonded (H-bonded) self-assembled structures (layer-by-layer (LbL) films or fibers) consisting of a thermo-responsive polymer, poly(2-ethyl 2-oxazoline) (PEOX), and pH-responsive molecules, tannic acid (TA) or malonic acid (MA). pH-responsive H-bonded LbL assemblies of PEOX and TA restructure in acidic phosphate buffer into pH-responsive fibers. Since H-bonds between PEOX and TA are stable in acidic solutions, the restructuring was attributed to hydrophobicity increase of PEOX in time by H2PO4- ions. A systematic investigation of the influence of Hofmeister series of salts on the properties of PEOX/TA multilayers was presented. pH stability of the films increased and the film thickness decreased with the anion type in Hofmeister series in direction from kosmotropic (dehydration of polymer chains) to chaotropic anions (direct binding of ion on the chain). PEOX/TA multilayers as controlled release platforms were demonstrated by using Rhodamine 6G dye molecule. Release was pH and temperature-responsive depending on the ionization of TA and cloud point temperature (Tcp) of PEOX. Effect of chemical structure of H-accepting polymer on release was studied by forming poly(N-isopropylacrylamide) (PNIPAM)/TA multilayers which exhibited faster release due to more hydrophobic structure. Hollow fibers of PEOX/MA formed in aqueous solutions by slow self-assembly. At pH2, hollow fibers formed by H-bonding while at pH4, electrostatic bridging of PEOX chains by MA was dominant. Fiber formation was temperature-responsive depending on Tcp of PEOX. The effect of three salts (Na2CO3, NaCl, NaSCN) on fiber formation was studied. The potential of fibers in controlled release applications was demonstrated by the release of Ciprofloxacin, a broad-spectrum antibiotic, at pH7.4.
Elektrotitreşim yoluyla kademeli sürtünme değişimi
Rendering tactile effects on a touch screen via electrovibration has many potential applications. However, our knowledge on tactile perception of change in friction and the underlying contact mechanics are both very limited. In this thesis, we investigate the tactile perception and the contact mechanics for a step change in friction under electrovibration during a relative sliding between a finger and the surface of a capacitive touch screen. First, we conduct magnitude estimation experiments to investigate the role of normal force and sliding velocity on the perceived tactile intensity for a step increase and decrease in friction, called rising friction (RF) and falling friction (FF). To investigate the contact mechanics involved in RF and FF, we then measure the frictional force, the apparent contact area, and the strains acting on the fingerpad during sliding at a constant velocity under three different normal loads using a custom-made experimental setup. The results show that the participants perceived RF stronger than FF, and both the normal force and sliding velocity significantly influenced their perception. These results are supported by our mechanical measurements; the relative change in friction, the apparent contact area, and the strain in the sliding direction were all higher for RF than those for FF, especially for low normal forces. Taken together, our results suggest that different contact mechanics take place during RF and FF due to the viscoelastic behavior of fingerpad skin, and those differences influence our tactile perception of a step change in friction.
Epigenetik yeniden programlama ile MLL-AF9 lösemilerini hedefleyen yeni tedavi yöntemleri
Leukemia is a highly complex disorder of blood and bone marrow and characterized by inhibition of differentiation during hematopoiesis which leads to abnormal cell proliferation. Mixed lineage leukemia (MLL) is a form of acute leukemia that represents poor prognosis and due to chromosomal translocation, resulting in a hyperactive MLL fusion protein. MLL leukemias are largely based on epigenetic irregulations rather than genomic instability. A transcription factor (AF9) that fuses with MLL plays a role in the uncontrolled growth of acute monocytic leukemia. Chromatin modifying enzymes are aberrantly expressed in leukemias and targeting these regulators such as the Lysine-specific demethylase (LSD1) and Histone deacetylase (HDAC) has been considered as a novel treatment modality. In MLL-AF9 leukemias, recruitment of these enzymes due to translocation results in the activation of several genes that inhibit differentiation and lead uncontrolled cell proliferation. In this study, novel compounds that synthesized to inhibit LSD1, HDAC6, and both LSD1& HDAC6 (Dual Inhibitor) are characterized. We showed that the compounds inhibit target enzymes by in vitro enzyme activity tests, we also showed that the compounds engage with the target enzymes in the cell by cellular thermal shift assay (CETSA). The toxic effects of the compounds on leukemia cells were shown with ATP-dependent cell viability tests. Increased histone methylation and acetylation levels after inhibition of target enzymes were indicated with Western blot. Increased expression of downstream genes due to enzyme inhibition was showed with RT-qPCR. The synergistic effect of the inhibitors with the therapeutic drugs used in the clinic was investigated. We showed that the death response could be increased with a combination of Doxorubicin and our epigenetic inhibitors. Finally, RNA-seq analysis revealed the molecular mechanism of Doxorubicin synergy. Defects in epigenetic pathways involving increased expression levels or abnormal patterns of activity are one of the key drivers of cell proliferation in cancer. Targeting MLL-AF9 is an attractive strategy for therapeutic intervention in patients with this genetic translocation. Since our inhibitors are more effective on MLL-AF9 leukemias, we hope that our study will result in the characterization of targeted drugs for use in leukemia therapy.
Self-assembled whispering gallery mode, random and distributed feedback biolasers
Ever since their first demonstration by Theodore Maiman in 1960, lasers have enormous influence in our life. Laser light is coherent in space and time, directional and its spectrum is tunable. With their unique emission properties, lasers have a broad range of applications including barcode readers, blu-ray players, high-resolution imaging, and fiber-optic communication. Moreover, their use is emerging in biomedical imaging, diagnosis, and surgery. Commonly, lasers are fabricated using artificial or inorganic materials, however, these lasers are not proper to interface with living systems such as cells and tissues. As a solution, in this research, we fabricated biomaterial-based lasers (biolasers) made of biocompatible materials of proteins and biopolymers. Motivating from the coffee-stain effect, we developed unconventional cavities for biolasers using self-assembly methods. First, we demonstrated whispering gallery mode biolasers using silk fibroin protein and synthetic biopolymers. No tools, components, and/or human intervention are needed after the construction process is initiated. In the first section, we showed the transition of 2D coffee stain into 3D hollow spheroid structure and utilized them spheroid lasers. Next, we used the suppression of the coffee stain effect to fabricate disk lasers that are flexible, physically transient, and can be directly integrated on a variety of substrates. Second, we explained the use of self-assembled spatially localized feedback random lasers utilizing cracks. Directional random lasers with high Q-factors were presented using a wide variety of materials including fluorescent proteins, silk fibroin, silica nanoparticles, and synthetic biopolymers. Finally, we stated single transverse mode all protein lasers utilizing distributed feedback cavities and the emission was adjusted according to the cavity design. Our findings introduce an innovative way of fabricating biocompatible and biodegradable lasers. This research generates the foundation of the advances of biolasers that are implantable to living systems.
Evre kaydırmalı çınlaç boşalım izgegözlem yöntemi ve sündürülmüş optik lif temelli doğrusal çınlaç algılayıcısı
This dissertation presents the linear optical cavity sensor based on fiber Bragg grat- ings and tapered optical fiber. The sensing and detection approach used in this thesis is phase shift cavity ring down spectroscopy. It is because of linear nature of the cavity and detection mechanism applied, that the whole study is named as "linear cavity tapered fiber sensor using phase shift cavity ring down spectroscopy". First part of the thesis is dedicated to the study of experimental investigations that are available in the literature for the purpose of optical sensing. Among the reviewed techniques are the surface plasmon resonance (SPR), resonant wavelength shift, direct absorption spectroscopy, cavity ring down spectroscopy (CRDS) and its vari- ants like cavity enhanced absorption spectroscopy (CEAS). Then these techniques are sorted out depending upon the advantages that they have on each other and limitations that put constraint on their performance. For example, surface plasmon resonance is a single pass approach, which is its major constraint. But multi-pass method dubbed as resonant wavelength shift, although it is sensitive enough but suf-fers from laser intensity changes. On the other hand, cavity ring down spectroscopy, is free from such errors but it is expensive and complicated technique. The sensing technique called phase shift cavity ring down spectroscopy not only combines the advantages of CRDS and resonant wavelength shift but also provide solutions to their short comings. That is, it is economically feasible and not affected by the noise related to laser intensity. In the manuscript under study, mathematical foundation of the phase shift cavity ring down spectroscopy is also given due attention. In addition to this, fabrication of fiber Bragg gratings, their character- ization, tapered optical fiber and its evanescent field are the topics which are also discussed in the same thesis.In the second part of the thesis, an experimental investigation has been reported by using tapered fiber based cavity. Here we employed a tapered fiber-based linear fiber cavity for the demonstration of a highly sensitive sensor for sucrose concentration in water using PS-CRDS. This, linear fiber cavity has a small cavity length (∼1.25 m) that enables the observation and tracking of individual cavity modes in transmission and phase spectra recorded during laser sweeps. Hence, it eliminates the need for Pound–Drever–Hall locking of the laser source to the cavity resonance and thus provide a simpler experimental scheme. The analysis of the recorded data sets was performed to track the changes in phase shifts observed only when the laser wavelength is in resonance with the cavity modes. Such a mode-tracking PS-CRDS approach reveals limit of detection values less than around 400 µM, better than the performance of previously demonstrated PS-CRDS sucrose concentration sensors employing fiber loop resonators. The sensitivity of our sensor critically depends on the fiber taper diameter and can reach up to around 6◦/1mM Suc. for 3.2 µm taper diameter at 6 MHz modulation frequency using fiber Bragg gratings (FBGs) with reflectivities around 86%. This value can be further increased by employing FBGs with higher reflectivities or fiber tapers with smaller diameters provided that the cavity loss due to water absorption is compensated with an amplifier. It is well known fact common sources of optical losses are scattering, optical components, splicing or water absorption (for liquid phase measurements). These losses become more prominent at small taper waist diameters. So, by com- pensating these losses, working at the taper diameter lesser than 2 µm is possible. Last section of the thesis explains the working of the system for above said idea. Basically, to compensate for the optical losses, a booster optical amplifier (BOA) was added in to the previously designed simple PS-CRDS experimental scheme. Af- ter carefully adjusting the gain, this optical amplifier, provides enhanced ring down time and more round trips of light into the cavity. Which ultimately manifests itself in the form of improved sensitivity and detection limit. Some exemplary measurements performed with the amplified PS-CRDS showed the sensitivity of 4.84◦/1mM Suc. for 2 µm taper diameter at 2 MHz modulation fre- quency. Higher sensitivity can also be achieved by adjusting booster optical amplifier at higher gains and using even more thinner tapers. The thesis ends with the con- cluding remarks and future directions. It highlights the potential applications of PS-CRDS and suitable candidates to replace tapered optical fiber as a sensing head. Implementation of these changes further enhance the performance of sensing device.
Optoelektronik nöral arayüzleri için yeni biyouyumlu kuantum noktaları ve nanoteknolojik birleşimler
Colloidal semiconductor quantum dots offer high potency in the realm of targetable and remotely addressable biointerfaces due to their low-cost solution-processability, wide spectral tunability and high photostability. But most popular and well-studied quantum dots contain cadmium and lead, which are known to be highly toxic and carcinogenic for biomedical applications. For that reason, the search for appropriate biocompatible quantum dots and their utilization is an important challenge in the development of quantum dot based bioelectronic devices. This thesis addresses efficient neural interfaces based on nanoengineered assemblies of novel and biocompatible quantum dots. In the first part of the thesis, we synthesized type-II indium phosphide/zinc oxide core/shell quantum dots for the first time. These quantum dots were incorporated into photoelectrode structure and induced bioelectrical current that triggered the firing of a single neural cell at 4 μW mm−2, 26-fold lower than the ocular safety limit for continuous exposure to visible light. Next, we inspired by the nonradiative energy transfer used by photosynthetic systems and developed quantum funnels based on indium-based rainbow quantum dots that are assembled in a graded energy profile. The novel nanoengineered assembly facilitated optical neuromodulation of a single cell and enhanced bioelectric current 215% per unit absorbance in comparison with the control sample. In the second part of the thesis, we reported colloidal aluminum antimonide quantum dots for the first time. The synthesis was performed via controlled reaction of aluminum chloride and antimony bis(trimethylsilyl)amide in the presence of superhydride. These quantum dots showed excitonic transitions in the UV-A region and tunable band-edge emission in the blue spectral range (quantum yield of up to 18%). Among all III-V quantum dots, aluminum antimonide quantum dots showed the brightest core emission in the blue spectral region. Finally, we used aluminum antimonide quantum dots as biointerfacing layer in a photovoltaic device for neural stimulation. This type of neural interface generated capacitive bioelectrical current with a rise time of ~55 μs and with a magnitude over 600 μA.cm-2, pointing out the most sensitive quantum dot based capacitive biointerface. This biointerface did not exhibit any toxic effect on the cells and demonstrated operational lifetime of one year in aqueous environment. These findings show that novel cadmium-free quantum dots can induce a biocompatible and effective biological junction and introduce a new route in the use of quantum dots in optoelectronic device architectures for light-triggered bioelectrical devices for applications such as artificial retinal prostheses.
Ses lateralizasyonunda kulaklar arası zaman ve seviye ipuçlarının entegrasyonunun çeşitlilik kombinasyonu perspekti ne dayalı olarak modellenmesi
A sound source with non-zero azimuth leads to interaural time and level differences. Studies on the hearing system imply that these cues are encoded in different parts of the brain, but combined to produce a single lateralization percept. According to the duplex theory of sound lateralization, ITD and ILD play a more significant role, respectively, in low- and high-frequency stimulations. The current study investigates the relative weights of these two cues based on the error rate data obtained from two major two-alternative forced-choice experiments. In the experiments, ITD and ILD values were extracted from a generic head-related transfer function (HRTF). In experiment I, three different sounds containing either only low frequencies, or only high frequencies, or both were lateralized by imposing these ITD and ILD values. The subjects' lateralization error rate data indicate that the performance of the binaural hearing system based on ITD and ILD cues undergo a linear combination based on the maximum ratio combining approach. In experiment II, a complex sound consisting of two low- and seven high-frequency harmonic tones was used as test sound. Each stimulus consisted of a reference sound having the ITD-ILD pair of a certain azimuth and a probe sound that was different from the reference sound in either its ITD or in its ILD or in both. Subjects indicated the direction of the shift they perceived in the lateralization of sound. The error rate data showed that the weights of the two cues change with the azimuth of the sound source. Based on the SNR values obtained from the error rate data, it was shown that the two pieces of spatial information supplied by ITD and ILD are combined in the brain, but not additively for shifts in lateral positions around small azimuths. The integration model of the ILD and ITD cues was defined by a diversity combination approach incorporating the observed directional bias with inward preference