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Modern mathematical methods in the research of immune reconstitutions and immunodeficiencies
Stuchlý, Jan ; Kalina, Tomáš (advisor) ; Šinkora, Jiří (referee) ; Štěpánek, Ondřej (referee)
In this thesis we present new analytical and integrative approaches for broad spectrum of applications of flow cytometry. CVID Large (88 individuals) cohort of CVID patients was immunopheno- typed by flow cytometry. The "probability binning" algorithm was used to au- tomatically assess the immunophenotype which presented unprecedented stabi- lity. We were able to define sub-group of CVID patients with highly activated immunophenotype showing hallmarks of immunosenescence and common clinical characteristics (thrombocytopenia, lung fibrosis and bronchiectasis). The severity of clinical complications correlated quantitatively with the immunophenotype. Analysis of cellular proteome We have designed technics which allow for highly multiplexed (>1000 of antibodies) analysis of human proteome using af- finity proteomics. We analyzed the changes of proteome of human cell lines and characterized the proteome of acute leukemias. The yet undescribed stability of proteome with respect the sub-cellular localization is shown and theoretical and practical background for antibody standardization and validation for the use in a nity proteomics is presented. Topological analysis of cytometry data Multiparametric flow cytometry allows for description of complex topological relationships of the cells in di erent stages of...
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Role of Bardet-Biedl syndrome (BBS) protein complex in T cells
Niederlová, Veronika ; Štěpánek, Ondřej (advisor) ; Černý, Jan (referee)
BBSome is a protein complex crucial for trafficking of specific cargoes to the primary cilium. Although primary cilia are typically not present in cells of haematopoietic origin, such as T cells, recent research has revealed striking parallels between the primary cilium and the immunological synapse. Amongst other similarities, both structures are supposed to use the same transport machinery involving Rab8 and IFT20, the close interaction partners of BBSome. The first goal of this thesis was to investigate the role of BBSome in the biology of T cells. Using RT-qPCR, we have shown that BBSome subunits are expressed in lymphoid tissues and T cells. Studies of localization of BBSome subunits in Jurkat cell line and primary OT-I T cells revealed that the subunits have distinct localization patterns with BBS4 localizing to the centrosome and BBS1, BBS5, and BBip10 having dispersed localization. After the contact with an antigen presenting cell, BBS4 re-localizes to the immunological synapse. Mutations in BBSome encoding genes cause Bardet-Biedl syndrome (BBS), a rare ciliopathy presenting with multiorganic symptoms. The second goal of this thesis was to examine the associations between BBS and the immune system. Examination of medical records of more than 450 BBS patients revealed that autoimmune...
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Prague advertisment in the interwar period
Štěpánek, Ondřej ; Jakubec, Ivan (advisor) ; Vošahlíková, Pavla (referee)
In this diploma thesis Prague Advertisement in the Interwar Period its author Ondřej Štěpánek deals with a so far almost neglected topic - advertisement in Prague in years 1918 -1939. After outlining the development of advertisement in these two decades in the whole newly constituted state, the author focuses on regional specifics in the capital city that were set by local regulations. He mainly addressed the legality of individual advertising means, especially those in public spaces, and he also described producers of advertisement in Prague. The author concludes that advertisement made a large progress in the respective period, particularly during the late 1920s and the first half of the 1930s. Advertising companies had to deal with many restrictions and prohibitions brought by the new legislation. This diploma thesis provides new insight in the research of the advertisement history, especially in the area of the development of advertising agencies.
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Regulatory CD8+ T lymphocytes
Chadimová, Tereza ; Štěpánek, Ondřej (advisor) ; Grobárová, Valéria (referee)
Regulatory T cells are essential for the maintenance of peripheral immune tolerance, preventing alloimmunity and pathogen-induced immunopathology. While regulatory CD4+ T lymphocytes have been studied for over two decades, the existence of regulatory T lymphocytes in the CD8+ compartment is still controversial. Through the years, multiple CD8+ T lymphocytes subpopulations have been reported to exhibit regulatory activity, yet none has been canonically recognized. The aim of this thesis is to review current knowledge of these putative CD8+ regulatory lymphocytes and evaluate whether or not these subsets can be established as regulatory T cells. The emphasis of this thesis will be put on the phenotypic characteristics, regulatory activity and inhibitory mechanisms of these cells. Keywords: CD8+ Regulatory T lymphocytes, immunological self-tolerance, immunosuppression
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Homeostatic memory and virtual memory CD8+ T cells
Přibíková, Michaela ; Štěpánek, Ondřej (advisor) ; Krulová, Magdaléna (referee)
This work sums up current knowledge of so-called homeostatic proliferation-induced memory CD8+ T cells (HP) and virtual memory CD8+ T cells. These populations do not represent true immunological memory because they are generated in the absence of a cognate foreign antigen. However, both HP-memory and virtual memory T cells share some phenotypical and functional features with true memory T cells, including the ability provide rapid immune response during infection. HP-memory T cells are generated via homeostatic proliferation during experimentally induced lymphopenia. Virtual memory T cells might arise via homeostatic proliferation during neonatal or age-related periods of lymphopenia, however, they can be generated also in healthy lymphoreplete hosts. Based on detail analysis of these two populations, I concluded that HP-memory CD8+ T cells and virtual memory CD8+ T cells most likely use identical differentiation program and represent the same T cell population. Keywords: homeostatic proliferation, lymphopenia, CD8+ T cells, immunological memory, virtual memory cells
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Synthesis of biologically active compounds with quinazoline scaffold
Štěpánek, Ondřej
This thesis concludes my contribution to research of HIV-1 capsid assembly inhibitors. It has been shown that 2,4-disubstituted quinazoline derivatives are able to inhibit this process both, in competitive biochemical assay based on the AlphaScreen technology as well as in tissue cultures. The main objective of the work was to prepare the aforementioned quinazolines, to design and prepare new candidates with higher activity based on results of biochemical tests, and also to try to increase the solubility of otherwise poorly soluble compounds. Disubstituted quinazolines are relatively easily accessible from the commercially available anthranilic acid derivatives. These are converted to the corresponding quinazolin- 4(3H)-ones by the condensation reactions. In this work, two methods were used for the preparation of quinazolin-4(3H)-ones: reaction of acyl chlorides with aromatic anthranilamides provided 2-arylamidobenzamides whose subsequent cyclisation under basic conditions led to derivatives of 2-arylquinazolin-4(3H)-one; reaction of anthranilic acid esters with aromatic nitriles, which afforded desired quinazolin-4(3H)-one in one reaction step. Chlorination of 2-arylquinazolin-4(3H)-ones using POCl3 then led to 2-aryl-4- chloroderivatives as key intermediates. Nucleophilic substitution of...
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Synthesis of biologically active compounds with quinazoline scaffold
Štěpánek, Ondřej ; Kotora, Martin (advisor) ; Hájíček, Josef (referee) ; Moravcová, Jitka (referee)
This thesis concludes my contribution to research of HIV-1 capsid assembly inhibitors. It has been shown that 2,4-disubstituted quinazoline derivatives are able to inhibit this process both, in competitive biochemical assay based on the AlphaScreen technology as well as in tissue cultures. The main objective of the work was to prepare the aforementioned quinazolines, to design and prepare new candidates with higher activity based on results of biochemical tests, and also to try to increase the solubility of otherwise poorly soluble compounds. Disubstituted quinazolines are relatively easily accessible from the commercially available anthranilic acid derivatives. These are converted to the corresponding quinazolin- 4(3H)-ones by the condensation reactions. In this work, two methods were used for the preparation of quinazolin-4(3H)-ones: reaction of acyl chlorides with aromatic anthranilamides provided 2-arylamidobenzamides whose subsequent cyclisation under basic conditions led to derivatives of 2-arylquinazolin-4(3H)-one; reaction of anthranilic acid esters with aromatic nitriles, which afforded desired quinazolin-4(3H)-one in one reaction step. Chlorination of 2-arylquinazolin-4(3H)-ones using POCl3 then led to 2-aryl-4- chloroderivatives as key intermediates. Nucleophilic substitution of...
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The Industry Business of Prague Municipality in the first half of 20th Century. The Industrial Factories of Capital City Prague, ltd.
Štěpánek, Ondřej ; Jakubec, Ivan (advisor) ; Drnek, Kryštof (referee)
This work describes the industrial activities of the Prague municipality in building industry in the first half of the twentieth century, especially in the years 1921-1948, during which the company "Industrial factories of the Capital City of Prague" operated successfully in which the municipality owned half of the shares. The main objective is to monitor the activities of the company running the lime and brick factories and a sawmill. I focused on its development and significance. This thesis is based on information from the Prague City Archives, from the fund of Industrial Factories. The research illustrates the quantities of produced and sold building materials, then also important buildings in which the company was involved as a supplier. The main contribution of this work is the "re-discovery" of an interesting company which significantly supported the building industry in Prague, supplied the building material not only to municipal buildings and was a competitor for cartels.
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