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Cell surface CD47 expression in cancer stem cell-targeted tumor therapy
Kuzmík, Ján ; Drbal, Karel (advisor) ; Černý, Jan (referee)
CD47 is a transmembrane glycoprotein with a high expression in both, healthy and cancer (stem) cells. Level of the CD47 expression is negatively correlated with survival of cancer patients. Binding of CD47 to SIRPα, localized on a phagocyte, triggers intracellular signaling cascade. The final effect of this cascade is dephosphorylation of nonmuscle myosin-IIA, which disrupts its function and accumulation to phagocytic synapse. The blockage of CD47-SIRPα signaling pathway in a presence of the pro-phagocytic signal induces phagocytosis of cancer cells. Afterwards, phagocytes can serve as the antigen presenting cells and prime T cell response. Role of CD47-SIRPα signaling pathway in immunity has established this pathway as a target of cancer therapy testing. Preclinical research has identified a positive therapeutic effect of blocking this signaling pathway. Nowadays, the first phase of clinical trials is being conducted. The most prevalent approach of blocking CD47-SIRPα signaling pathway in therapy is the use of anti-CD47 blocking monoclonal antibodies, which cause mild anemia. However, alternative approaches of blocking this pathway are also being developed. In this bachelor thesis, I have summarized the research related to the blockage of CD47-SIRPα signaling pathway as a cancer therapy.
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The Role of Heat Shock Proteins in Pathogenesis of Placental Insufficinecy
Slabá, Kristýna ; Hromadníková, Ilona (advisor) ; Černý, Jan (referee)
Heat shock proteins (Hsp) are highly conserved proteins that are part of the universal stress response of the cell. Their main function is to protect cells against structural and functional damage. Organisms exposed to different forms of stress, such as e.g. a lack of nutrients or water, hypoxia, infection or inflammation, demonstrated an increased gene expression of these proteins. Pregnancy complications cause stress conditions for maternal and fetal organism, which may result in an increased gene expression of Hsp. In my thesis, I examined the concentration of extracellular mRNA for five different heat shock proteins (Hsp27, Hsp60, Hsp70, Hsp90, HspBP1) in the plasma of pregnant women and wheather this concentration is affected by possible pregnancy complications (preeclampsia, fetal growth restriction and gestational hypertension). I also investigated a possible correlation between mRNA plasma concentration for Hsp and pulsatility index values (PI) obtained by Doppler ultrasound. This research should help to invent a new predictive method for pregnancy complications, based on a detection of specific biomarkers in the first trimester of pregnancy. The research was conducted on plasma samples obtained from peripheral blood of pregnant women, whose collection was performed during clinical manifestations of...
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Chimeric antigen receptors in the treatment of hematological malignacies
Fellnerová, Adéla ; Filipp, Dominik (advisor) ; Černý, Jan (referee)
Chimeric antigen receptors (CARs) are artificial molecules composed of an antibody derived antigen recognition domain which is fused with the signal transduction domain derived from the physiological TCR. CAR technology used to transduce patients T-cells and endow them with the specificity to a certain surface antigen, has been a major breakthrough in cancer immunotherapy in the last decade. This strategy has been most successful for treating hematologic malignancies. Various CAR approaches and applications are currently tested mainly in the United States where many clinical trials have been launched. In contrast, in the Czech Republic, there are only a few teams focused on this topic with no clinical trials going on. During my work on this diploma thesis and in close collaboration with MUDr. Pavel Otáhal, PhD., who is working on implementation of CAR technology into the Czech clinics for the treatment of B-cell malignancies, individual functional CARs were prepared and tested. CAR expressing Jurkat T-cell lines were generated using a lentiviral vector transduction system. CAR functionality was determined by two different assays. We have shown that individual CARs are able to recognize the B-cell lineage specific antigens CD19 and CD20 and significantly up-regulate the activation molecule CD69 upon...
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Analysis of Memrane Proteins of Pathogenic Bacterim Francisella Tularensis
Schmidt, Monika ; Szotáková, Barbora (advisor) ; Černý, Jan (referee) ; Konopásek, Ivo (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate Mgr. Monika Schmidt Supervisor Doc. Ing. Barbora Szotáková, Ph.D. Title of Doctoral Thesis Analysis of membrane proteins of pathogenic bacterium Francisella tularensis Bacterium Francisella tularensis is highly infectious pathogen causing disease tularaemia. Due to the lack of standardization and little protection against highly virulent strains, the only vaccine developed against this pathogen is not allowed for clinical usage. Conserved hypothetical lipoprotein homological to thiol/disulfide oxidoreductase (DsbA) was recently described as essential virulence factor of Francisella tularensis. The dsbA gene deletion led to attenuation of the strain and development of immunoprotection. The DsbA protein sequence revealed the presence of carboxy-terminal DsbA_Com1-like domain harbouring the catalytic active site C-X-X-C and cis-proline and domain amino-terminal to FKBP type peptidyl-prolyl isomerase. This work was focused on functional a substrate characterization of DsbA protein. The functional analysis of this protein showed both the importance of the active site, cis-proline and the FKBP_N domain for the thiol/disulphide oxidoreductase activity. Further, this work also revealed the in...
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Activation of the initiation caspases and regulation of their activity
Votavová, Barbora ; Anděra, Ladislav (advisor) ; Černý, Jan (referee)
Caspases are the key proteins participating in both activation and execution of apoptosis. Extrinsic or intrinsic apoptotic signaling leads to sequential activation of the initiation and execution caspases. Activation of initiator caspases is mediated by their processing in multiprotein complexes and activated initiator caspases then specifically cleave and thus activate the effector caspases. These then cleave a number of structural and functional proteins, which consequently leads to cellular selfdestruction and its breakdown to apoptotic bodies. Considering the fundamental significance of the initiation of apoptosis, the activation as well as the activity of initiator (but also effector) caspases is strictly regulated at several levels. Primary the intensity and character of the recieved signal is crucial for the effective formation of the caspase activation complex. Then the concentration of intracellular ionts, nucleotides and various proteins (proteins from the Bcl-2 family, inhibitors of apoptosis (IAPs), heat shock proteins,…) can also highly influence individual steps of caspase activation. Caspases themself can be posttranslationally modified (phosphorylated, ubiquitylated,…) and their activity can be either suppressed or also enhanced. All these processes form complex regulatory network serving...
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Emerging role of Toll-like receptors in central tolerance
Súkeníková, Lenka ; Filipp, Dominik (advisor) ; Černý, Jan (referee)
Medullary thymic epithelial cells (mTECs) provide a specific thymic microenvironment for the processes associated with T cell development. Their irreplaceable function is the expression of specific set of antigens which are expressed only in peripheral tissues, called tissue restricted antigens (TRAs). Such expression, often referred to as promiscuous, was originally described with the discovery of transcriptional regulator Aire. Aire, which has a potential to interact with many other transcription factors, also binds to DNA, and thus can alter the general pattern of cellular gene expression. T cells exhibiting a strong affinity to TRAs expressed on mTECs are removed from the thymus by negative selection or their development is deviated to regulatory T cell (Treg) lineage. Studies on mice and humans confirmed the critical role of Aire protein in the establishment of the central tolerance. Inactivating mutations in Aire gene cause deficiency in TRA expression, failure to remove and the escape of self-specific T cells from the thymus to periphery, and in turn, autoimmunity. Experimental evidence points to the key role of NF-κB signaling pathway in mTECs development. The very same pathway is regulated also by Toll-like receptors (TLRs), which recognize evolutionary conserved structures derived from...
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Cell-mediated peripheral tolerance in lymph nodes
Brabec, Tomáš ; Filipp, Dominik (advisor) ; Černý, Jan (referee)
Tolerance of immune system to body-self constituents is a crucial issue for immunologists to solve. While the mechanisms of central tolerance are now described to well extent, antigen-specific tolerance mechanisms on immunological periphery are just beginning to be revealed, characterized and appreciated. Recently, novel models of peripheral tolerance emerged. Particularly, a model based mostly on lymph node stromal cells could be of profound importance, since it provides answers to some fundamental questions in tolerance immunology. So far, no review paper highlighting these newly discovered roles of lymph node stromal cells was published. Therefore, in this study we summarize data covering this topic, published up-to-date. Further, this text provides a basic overview of lymph node functional anatomy. To better illustrate the topic, we also show some experimental evidence demonstrating lymph node architecture and the localization of extrathymic Aire-expressing cells, one of the lymph node-resident populations, recently implicated in peripheral tolerance maintenance. Powered by TCPDF (www.tcpdf.org)
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Appearing and Salvation. Subjectivity in The Material Phenomenology of Michel Henry
Černý, Jan ; Novotný, Karel (advisor) ; Švec, Ondřej (referee) ; Karul, Róbert (referee)
The doctoral thesis examines the possibilities of phenomenological philosophy to engage in the question of salvation in a Christian sense with reference to Michel Henry's material phenomenology. Henry's last three books signified a turn towards Christianity within his work and related the tension of two basic modes of appearing, assumed by his phenomenology, to the question of the life and death of a human. Material phenomenology strongly exposed the subjective pole of appearing and made subjectivity the stage for the story of human salvation. The thesis examines both the general concept of subjectivity in material phenomenology and the particular concept of a divine and human subject in the last three books of Michel Henry. The thesis follows the way Michel Henry creates the phenomenology of the inner- divine life; it examines the movement of a human subject from the inside of the divine life to the transcendence of the world, and then its return through the "second birth" to the divine life being displayed within itself; it enquires in what sense the divine and human subject are incarnated subjects; it demonstrates how the human subject is becoming a subject understanding the word of a divine life which speaks both within itself and in the Scripture. The thesis pays attention to the relation of a...
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Aire-expressing cells in immune peripheral tissues
Vobořil, Matouš ; Filipp, Dominik (advisor) ; Černý, Jan (referee)
5 Abstract Tolerance to "self" is the fundamental property of the immune system and its breakdown can lead to autoimmune diseases. In order to eliminate self-reactive T- cells during their development in thymus (central tolerance), Aire promotes the expression of peripheral self-antigens in medullary thymic epithelial cells (mTECs). Recently, Aire was suggested to fulfil a similar function in rare lymph node and spleen cells (peripheral tolerance). However, the detection, characterization and function of these extrathymic Aire-expressing cells is still obscure. The main objective of presented thesis was to investigate if Aire positive cells are also present in other lymphoid as well as non-lymphoid tissues. Using two independent mouse transgenic models we identified the Aire-reporter expressing cells in several lymphoid tissues such as Peyer's patches, spleen and bone marrow as well as in one non-lymphoid organ, the lungs. We show here that based on the expression of B220, EpCAM and CD11c markers these heterogenic cells consist of at least five phenotypically distinct subpopulations, and with the exception of those from lungs, all of them are strictly of hematopoietic origin. This study also demonstrates that Aire on protein level is predominantly expressed by one of these subpopulations with CD45+ MHCII+...
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Transmembrane adaptor proteins and their roles in immunoreceptor signaling
Borna, Šimon ; Hořejší, Václav (advisor) ; Černý, Jan (referee)
Transmembrane adaptor proteins are group of proteins, which lack intrinsic enzymatic activity, but they share similar structure. They play essential role in signal transduction, because they are able to recruit cytoplasmic proteins to proximity of plasma membrane and thus trigger signal pathways. In this review I mainly focused on these, which were not reviewed for a long time, or were recently discovered and I am trying to summarise current knowledge about roles of these proteins in leukocyte signalling. Key words: transmembrane adaptor proteins, receptor signaling, immunoreceptors
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