National Repository of Grey Literature 69 records found  1 - 10nextend  jump to record: Search took 0.01 seconds. 
Immunogenicity of induced pluripotent stem cells (iPSC)
Tejklová, Tereza ; Drbal, Karel (advisor) ; Hájková, Michaela (referee)
Ectopic expression of several transcription factors into the somatic cells allows us to artificially dedifferentiate them into induced pluripotent stem cells (iPSC), which show great promise in regenerative medicine and personalized disease modelling, as well as diagnostic tools. Unique attribute of iPSC is the possibility of creating autologous cells for each patient, which could be used for transplantation without fear of immune rejection. However, cells differentiated from iPSC generally display decreased expression of MHC I glycoproteins, which leads to the activation of NK cells of innate immunity. T cells, the part of adaptive immunity, are activated after recognition of antigen peptide or foreign MHC I glycoproteins only in co-operation with costimulatory molecules, which are not usually expressed on iPSC. During dedifferentiation, cells keep the epigenetic profile of the source cell, which can result in the abnormal expression of genes within derived cell lines. Overall immunogenicity depends on the method of iPSC preparation, with respect to genomic stability. Another important factor is the immune environment of transplantation site as well as the tissue damage caused during transplantation. This results in the presentation of danger signals (DAMPs), which are then recognized by pattern...
Immunogenic cell death
Šímová, Michaela ; Drbal, Karel (advisor) ; Javorková, Eliška (referee)
According to the danger model, the immune system is activated by endogenous molecules known as danger-associated molecular patterns (DAMP) that are externalized from the interior of a dying cell to the cell surface or released into the extracellular space. Due to the loss of plasma membrane integrity a necrotic cell death as well as several types of proinflammatory programmed cell death are considered to be immunogenic, whereas apoptosis, on contrary, has been initially defined as a tolerogenic type of cell death. However, under certain circumstances, the immune response can be initiated by an apoptotic cell after exnternalization of DAMP molecules by newly described secretory pathways. This phenomenon was observed on tumor cells as a result of some widely used therapeutic modalities and is known as immunogenic cell death (ICD). Nomenclature of selected types of cell death is part of this thesis. The aim of this bachelor thesis is to provide an evidence of the experimental support for ICD theory during in vivo initiation of the immune response. I will evaluate the correlation between ICD and the induced exposure of DAMP molecules on the surface of tumor cells or their secretion to the extracellular space.
Detection of surface phenotype and chemosensitivity in bladder carcinoma cells in vitro
Šímová, Michaela ; Drbal, Karel (advisor) ; Vondálová Blanářová, Olga (referee)
Tumor malignancies are the second leading cause of death worldwide. One of the reasons for the failure of oncological treatment are the uniformly set clinical guidelines, which neglect the effect of high intertumoral heterogeneity. The in vitro chemosensitivity and resistance (CSRA) assays allow for the stratification of patients prior to therapy. Therefore, the CSRA are a long-considered method for personalization of components of chemotherapy regime. Nevertheless, none of them is being routinely used in clinical practice. Certain chemotherapeutics used for their cytotoxic and cytostatic effect are also able to induce so-called immunogenic cell death (ICD) of tumor cells and activate an anti-tumor immune response. Monitoring of changes in the expression of molecules associated with the regulation of the innate immune system on the surface of dying tumor cells would enable to predict the patient's ability to respond to treatment involving modern immunotherapeutics. The feasibility of CSRA using flow cytometry and microscopy is critically evaluated in this thesis on a model of bladder cancer. Simultaneously, the correlation of the immunogenic phenotype of tumor cells and their sensitivity to selected chemotherapeutics is discussed.
Fibroblast activation protein and local immunosuppression in glioblastoma
Ternerová, Nikola ; Stollinová Šromová, Lucie (advisor) ; Drbal, Karel (referee)
Glioblastomas (GBMs) are one of the most common malignant tumors in the central nervous system. The tumor microenvironment of GBMs contains malignant and non-malignant stromal cells, whose interactions contribute to several GBMs characteristics, including aberrant angiogenesis, high proliferation rate, and systemic and local immunosuppression. Fibroblast activation protein α (FAP) is a membrane serine protease that is sparsely expressed in healthy tissues but is upregulated in solid tumors, including GBMs. FAP can be expressed by both malignant and non- malignant stromal cells in the tumor microenvironment, and its expression in stromal cells has frequently been linked to impaired anti-tumor immune response. The role of FAP and FAP expressing stromal cells in the infiltration of immune subpopulations into the GBM microenvironment is still unclear. This diploma thesis aimed to develop a flow cytometry protocol for the analysis of immune cell subpopulations present within the tumor microenvironment in wild-type and FAP knockout mouse syngeneic glioblastoma model. Four methods combining mechanical and enzymatic dissociation were evaluated for their ability to preserve cell viability and expression of studied surface molecules using mouse non-tumorous and GBM tissue. The most suitable method for mouse...
Immunoscore in 3D tissue
Novák, Jaromír ; Drbal, Karel (advisor) ; Procházka, Jan (referee)
Solid tumors are complex structures comprising besides the cancer cells vasculature, extracellular matrix (ECM), soluble molecules and a plethora of various other cell types. These components form a so-called tumour microenvironment. From the numerous cell types that are part of tumor microenvironment, tumor infiltrating lymphocytes (TILs) play a major role in patient prognosis. Their presence is also of major importance with regard to new biological therapies based on immune checkpoint inhibitors. Crucial role of TILs is also reflected by the new approaches in cancer diagnostics namely by Immunoscore method (currently used in clinical settings). Immunoscore is based on localization and quantification of CD3+ and CD8+ TILs in thin histological sections of tumor tissue. The question remains to which extent the information obtained from 2D slices reflects the situation in tumor microenvironment considering its spatial heterogeneity. The development of new methodological approaches allowing evaluation of histological information in 3D is the key to answer this question. The theoretical part of this work first describes the heterogeneity of the tumor microenvironment and the role of immune cells within it. Then, the role of spatial heterogeneity and its possible influence on the histopathological...
Leukaemia associated immunophenotype in childhood acute leukaemias and its development during the course of disease
Podolská, Tereza ; Fišer, Karel (advisor) ; Drbal, Karel (referee)
Acute lymphoblastic leukaemia (ALL) is the most frequent childhood malignancy. One of the recent improvements in ALL treatment was the introduction of minimal residual disease (MRD) monitoring that enables risk stratification based treatment adaptation. The same MRD monitoring helps to choose relapse treatment, to guide indication for stem cell transplantation (SCT) and allows for a more personalized management of patients undergoing SCT. One of the main routes of MRD levels detection is characterisation of leukemic blasts using flow cytometry. However, flow cytometry is limited by its mainly manual expertise-based analysis. Such analysis is subjective and clearly insufficient for current complex data. While new computational tools are available for multidimensional flow cytometry data, there is an urgent need to test and adapt them for the use in clinical environment. The goal of this thesis is to detect immunophenotypes associated with leukaemia and their development by leveraging machine-assisted analysis of a set of diagnostic files selected based on information about more than three hundred thousand of multiparameter flow cytometry datasets. Advanced bioinformatic tools will help to detect blast and healthy haematopoietic populations, to derive their immunophenotypes and to identify individual...
Hematopoiesis in the models of zebrafish and medaka as a recipient for human HSC xenograft
Pravcová, Naďa ; Drbal, Karel (advisor) ; Svoboda, Ondřej (referee)
Danio rerio (zebrafish) and Oryzias latipes (medaka) have recently become popular model organisms to study hematopoiesis. These model organisms present several advantages in comparison to other commonly used models, the most common being Mus musculus (mouse). The advantages are shorter generation time, large offspring production, frequent spawning, external fertilization and development, the optical transparency of embryos amenable to genetic manipulation on the background of vast numbers of transgenic lines (mainly in zebrafish) and inbred strains (in medaka). Moreover, most of the mechanisms behind zebrafish and medaka hematopoiesis are conserved in higher vertebrates. Most importantly, the optical transparency in early development and in adult mutant transparent strains allows for observation of hematopoietic stem cell (HSC) development in vivo. Therefore, it is possible to generate humanized fish using xenotransplanted human HSCs for studies of the engraftment, differentiation, and trafficking of human HSC in vivo. Currently, the most popular organism for human HSC xenotransplantation is mice. This model system is not suitable for in vivo imaging of HSC engraftment. Moreover, a prior immunodepletion step is necessary. The process of immunodepletion includes genetic manipulation or irradiation...
Comparative structural analysis of Borreliella spp. virulent factors focusing on their surface topology conservation
Zdrha, Alois ; Drbal, Karel (advisor) ; Nunvář, Jaroslav (referee)
Gram-negative bacteria of genus Borreliella cause the nowadays spreading illness Lyme borreliosis. However, their classification as gram-negative bacteria is rather mislea- ding because they differ in their complex genome, pathogenic adaptation to their hosts and composition of their outer membrane. One species of borreliella can harbour up to 23 variable plasmids and one conserved chromosome and they still lack many crucial proteins needed for synthesis of key compounds. Hence, they have to obtain these compounds from the host. However, in order for borreliella to utilise the metabolic processes of the host, it has to first survive its immune response. Therefore borreliella contains many virulent factors including highly variable surface lipoproteins. The variability is a major obstacle to overcome when using their surface epitopes for detection and vaccine development because most of the dominant antigenic epitopes of borreliella are usually parts of the most variable regions of the lipoproteins. Nowadays, we can use many different algorithms which determine evolutionary conserved epitopes based on analysis of sequences of given lipoprotein, in order to find suitable targets for antibodies. In general, conserved epitopes are more suitable for vaccines, whereas more variable epitopes are better...
Characterization of T-cell clones from naïve and virtual memory compartment
Přibíková, Michaela ; Štěpánek, Ondřej (advisor) ; Drbal, Karel (referee)
Virtual memory (VM) CD8+ T cells represent a population of antigen-inexperienced T cells with an apparent memory phenotype. In lymphoreplete germ-free mice VM CD8+ T cells represent 10-20% of all peripheral CD8+ T cells. Their origin correlates with the levels of self-reactivity where the main factor that determinates the T-cell fate decision is the strength of homeostatic signals. In the first part of this thesis, we demonstrated that VM CD8+ T cells and naïve CD8+ T cells had distinct TCR repertoire and T-cell subsets contained different clonotypes. Moreover, 'VM clones' were enriched among VM T cells and were also present in naïve T cells. In contrast, 'naïve clones' were almost exclusively detected in naïve T cells. Next, we characterized the signaling of particular OVA-reactive TCRs from both naïve and VM subsets. We confirmed that 6 out of 8 tested TCRs were responsive to Kb-OVA. In the last part of the thesis, we developed and optimized a qPCR-based method for the relative quantification of specific T-cell clonotypes prior to and during the immune response. This method will serve as a tool for studying the biology of particular VM and naïve T-cell subsets and their role during the immune response. Keywords: T-cell receptor, homeostatic signaling, self-reactivity, virtual memory cells, T cells
Fibroblast activation protein and local immunosuppression in glioblastoma
Ternerová, Nikola ; Stollinová Šromová, Lucie (advisor) ; Drbal, Karel (referee)
Glioblastoma multiforme (GBM) is the most aggressive type of primary brain tumor. Current treatment includes surgical resection with following radio/chemotherapy, but prognosis of patients remains poor with median survival only about 15 months. GBM is characteristic for necrotic regions, abnormal vascularization and strong immunosuppression. Dynamic interactions of cancer cells, immune cells and other stromal cells in the tumor microenvironment can promote tumor growth and progression. Fibroblast activation protein α (FAP) is overexpressed by the cells in tumor tissue. FAP is important in angiogenesis, remodelation of extracellular matrix and immunomodulation in cancers. The role of FAP in the tumor microenvironment is the subject of recent research. The aim of the thesis was to prepare a syngeneic mouse model of glioblastoma with and without FAP expression, implement and optimalize the dissociation method for GBM tumor tissue and detect a variety of infiltrated immune cell populations in the GBM microenvironment by flow cytometry. Optimization of dissociation protocol for glioblastoma tissue was a crucial step for viable cell suspension required for cytometry study of immune cell populations. A combination of dissection by dissociator and enzymatic digestion with mild enzymes was found to be the...

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