|
|
Posttranslational modification of the adapter protein DAXX in the cellular response to genotoxic stress
Bražina, Jan ; Anděra, Ladislav (advisor) ; Černý, Jan (referee) ; Vodička, Pavel (referee)
Maintaining the chromosome continuity and complete genetic information in human cells is crucial for cell survival and the whole organism. It prevents life-threatening pathologies and preserves genetic continuity. However, cellular DNA is exposed to both endogenous and exogenous stress damaging its content and integrity. This stress activates mechanisms involving detection and repair of these damaged sites (DDR). One of the most serious types of DNA damage double-stranded breaks (DSB) occuring when both strands are severed. DSBs trigger wave of PTMs that regulate protein interactions, nuclear localization and catalytic activity of hundreds of proteins. Such modifications include acetylation, methylation, SUMOylation, ubiquitinylation and especially phosphorylation. The most important kinases involved in DDR kinases are ATM, ATR and DNA-PK. These kinases are activated immediately after the detection of the damaged area. DAXX (Death-associated protein 6) is an adapter and predominantly nuclear protein, which is involved in chromatin remodeling, gene expression modulation, antiviral response and depositing histone H3.3 variants into chromatin or telomeres. Daxx is essential for murine embryogenesis, since the homozygous deletion is lethal in E9.5-10. In 2006 a study mapping the substrates of kinases...
|
|
|
Novel mechanisms regulating immune tolerance and homeostasis in the intestine
Dobeš, Jan ; Filipp, Dominik (advisor) ; Černý, Jan (referee) ; Abramson, Jakub (referee)
Immune tolerance to host own tissues and cells is the fundamental attribute of properly working immune system. The repertoire of effector T-cells, which possess randomly generated antigen-specific receptors, is during their development shaped by central immune tolerance to retain only those specificities which do not recognize self-antigens. In addition, various mechanisms of peripheral tolerance keep in check potentially self reactive cells which escaped from the protective mechanism of central tolerance. Thus, a tight regulation of tolerance, operating at several anatomical places in the host body, collectively imposes immune homeostasis and well-being of the organism. The breach of central tolerance can have far reaching consequences, as demonstrated by mutations in Autoimmune regulator gene. These mutations lead to the development of severe autoimmune disease, comprising several clinical components, gastrointestinal associated symptoms including. We have shown, that in the absence of Autoimmune regulator, the occurrence of gastrointestinal symptoms is associated with the loss of thymic- mediated central tolerance to enteric α-defensins, essential antimicrobial peptides produced by intestinal Paneth cells. The loss of tolerance leads to the escape of defensin-specific T-cells to immune periphery...
|
|
|
Signaling effects of adenylate cyclase toxin action on phagocytes
Černý, Ondřej ; Šebo, Peter (advisor) ; Černý, Jan (referee) ; Dráber, Petr (referee)
The adenylate cyclase toxin (CyaA) plays a key role in the virulence of Bordetella pertussis. CyaA penetrates CR3-expressing phagocytes and catalyzes the uncontrolled conversion of cytosolic ATP to the key second messenger molecule cAMP. This paralyzes the capacity of neutrophils and macrophages to kill bacteria by oxidative burst and opsonophagocytic mechanisms. Here we show that CyaA suppresses the production of bactericidal reactive oxygen and nitrogen species in neutrophils and macrophages, respectively. The inhibition of reactive oxygen species (ROS) production is most-likely achieved by the combined PKA-dependent inhibition of PLC and Epac-dependent dysregulation of NADPH oxidase assembly. Activation of PKA or Epac interfered with fMLP-induced ROS production and the inhibition of PKA partially reversed the CyaA-mediated inhibition of ROS production. CyaA/cAMP signaling then inhibited DAG formation, while the PIP3 formation was not influenced. These results suggest that cAMP produced by CyaA influences the composition of target membranes. We further show here that cAMP signaling through the PKA pathway activates the tyrosine phosphatase SHP-1 and suppresses the production of reactive nitrogen species (RNS) in macrophages. Selective activation of PKA interfered with LPS- induced iNOS expression...
|
|
|
Characterization of the distribution and dynamics of the antigen-presenting cells using MHC II-EGFP knock-in mouse model
Pačes, Jan ; Černý, Jan (advisor) ; Tlaskalová - Hogenová, Helena (referee)
Results of recent studies indicate that dendritic cells are capable of transporting commensal intestinal bacteria into the mammary glands, which ultimately leads to their occurrence in breast milk. We have therefore decided to evaluate the phenotype of immunologically relevant antigen presenting cells (APCs) present in the mammary glands and the small intestine, respectively and perform a comparison study. We also studied plasticity of these populations during lactation. In situ immunodetection and flow cytometry methods were used to determine phenotype. We succeeded in optimising the methods for preparation of samples for flow cytometry and microscopy. We thoroughly tested protocols for 3D visualisation of APC populations and quantitative image analysis for correlation with flow cytometry, further optimization is nevertheless needed. We found out that during lactation large numbers of MHC II+ cells cluster around the alveoli and milk ducts. These cells are of a distinctly dendritic shape and their phenotype does not correspond to the APCs in the surrounding tissue. A pronounced increase of APC cells in the mammary glands between the fourth and sixth days of lactation was observed, with the majority of these cells expressing the CD103 antigen typical for cell populations of immune cells of the...
|
|
|
Progesterone influence on the maternal immune system in pregnancy
Škvorová, Anna ; Koucký, Michal (advisor) ; Černý, Jan (referee)
Pregnancy represents a major challenge to the maternal immune system. From an immunological point of view, a fetus is a semi-allograft. The mechanisms providing immunological paradox of fetal tolerance are still not well known and require further research. A complex network of immuno-endocrine interactions ensures fetal growth and development within the maternal uterus. The hormone playing an indispensable role in pregnancy is progesterone. The aim of this thesis is to summarize current knowledge of the effects of progesterone on the immune system in pregnancy and its mechanisms. Progesterone can affect target cells via the classical nuclear progesterone receptors, which act as transcription factors, or it can act using a variety of other ways, including non-genomic rapid signaling. Progesterone optimizes conditions for successful establishment and maintenance of pregnancy, changes the amount, localization and characteristics of immune cells and production of cytokines. It reduces the antigen-presenting capacity of dendritic cells, monocytes, and macrophages, suppresses NK cell cytotoxicity, supports the proliferation of uterine NK and dendritic cells, affects B cells and induces the formation of T regulatory cells and their recruitment into the fetal-maternal interface. The wide range of...
|
|
|
Mechanisms of specific immune response interactions with tumor cells.
Kaššák, Filip ; Hořejší, Václav (advisor) ; Černý, Jan (referee)
Interactions between the immune system and tumors have been among the highlights of present immunological research. An extensive body of new knowledge recently substantiated the long-presumed concept of cancer immunosurveillance. Immune system searches the organism for cells expressing tumor antigens or cellular stress signals and destroys them. T-cells, NK-cells and dendritic cells, as well as cytokine signaling and direct cell cytotoxicity play dominant role in this process. However, a fraction of nascent tumors can evade these mechanisms and create a dynamic equilibrium, gradually sculpting its phenotype by clonal selection. Eventually, tumor cells escape immune control by concealing themselves from recognition or by actively subjugating local immune response. This immunosubversion results in formation of immunosuppressive tumor microenvironment by recruiting protumorigenic cell populations, such as Treg cells, macrophages and myeloid derived suppressor cells. Soluble signaling molecules, as well as surface- expressed immune checkpoint molecules are exploited by tumor cells for inhibition of anti-tumor immunity. Highly effective therapeutic antibodies blocking these checkpoints have been developed for clinical use, with many more in current trials. Several other promising immunotherapeutic...
|
|
|
Antibody derivatives for the detection of human glutamate carboxypeptidase II
Bělousová, Nikola ; Bařinka, Cyril (advisor) ; Černý, Jan (referee)
Prostate cancer is the most common neoplasia in men. The therapy of progressed tumor is usually not efficient and early diagnosis is therefore crucial for successful treatment. Glutamate carboxypeptidase II (GCPII) is an established marker for prostate cancer imaging and therapy as the neoplastic transformation of prostate tissue is accompanied by the substantial increase of GCPII expression levels. Currently used GCPII-specific diagnostic and therapeutic reagents can be broadly categorized as small- molecule ligands or macromolecules. Antibodies are preferred macromolecules used in clinic. At the same time, however, protein engineering is regularly applied to modify natural antibodies to enhance their utility in biomedicine applications. This thesis summarizes the current knowledge about the GCPII structure and function and its role in diagnosis and therapy of prostate carcinoma. Powered by TCPDF (www.tcpdf.org)
|
|
|
Analysis of the resistence of B cell antigen receptor signaling to the inhibition of Src-family kinases
Borna, Šimon ; Brdička, Tomáš (advisor) ; Černý, Jan (referee)
Signalling through antigen specific receptors BCR and TCR is crucial for the development and the function of T cells and B cells. Although much is known about their signalling pathways a number of observations still remain to be clarified. In my thesis, I focused on the roles of Src-family kinases (SFKs) in the initiation of BCR- and TCR-mediated signalling. Several studies have suggested that in contrast to TCR signalling, BCR signal transduction could be initiated independently of SFKs or with only a minimal activity of these kinases. We used genetic approach to study the differences between TCR and BCR signalling apparatuses combined with inhibition of SFKs by pharmacological approach. Using this experimental set up, we show that the differences in the roles of SFKs and in the activities of SFKs needed for the initiation of BCR and TCR signalling are likely based on different composition or architecture of BCR and TCR. We further show that the SFK activity required for the initiation of TCR signalling is lower if ZAP-70 kinase is substituted with Syk kinase, which most likely reflects the different molecular mechanisms of Syk and ZAP-70 kinase activation. Key words: Src-family kinases, BCR receptor, TCR receptor, PP2, B cells, T cells, BCR signalling, TCR signalling.
|
|
|
mTOR mediated phosphorylation of Akt at Ser473 regulates cancer cell survival after severe hypertermia in PTEN-coordinated manner
Valentová, Iva ; Sadílková, Lenka (advisor) ; Černý, Jan (referee)
Hyperthermia (HT), a term used in general for temperatures higher than 42řC, induces cell stress response in dependence on its intensity, heating duration and target cell type (this is called a thermal dose). In vitro, HT causes changes in Ca2+ homeostasis as a consequence of endoplasmic reticulum stress and permeabilisation of the outer mitochondrial membrane. The release of cytochrom c further leads to activation of caspases that can result in cell death. In parallel, the presence of denatured proteins in the cell cytosol leads to the development of the so called unfolded protein response (UPR). UPR is characterized by increased expression and activation of molecular chaperones, increase in the intensity of proteosomal degradation and enhancement in autophagy. This process is accompanied by the attenuation of translation machinery, iniciation of DNA repair and activation of cytoprotective mechanisms leading to cell survival. This highly complex stress response mechanism is regulated mainly by the signalling pathways in the PI3K/Akt/mTOR axis. Cell survival is in the end fully dependent on the amount of energetic substrates, denatured proteins accumulation and on the level of irreversibly demaged DNA. If the termal dose is too high, the cell will ultimatively die. Hyperthermia is also one of the...
|
|
|
Historic causes and context of the present position of the Roman Catholic Church in the Czech society
Černý, Jan Karel ; Rychlík, Jan (advisor) ; Pehr, Michal (referee)
(in English) This thesis is focused on historic causes and context of contemporary position of the Roman Catholic Church in the Czech society. To chosen problematics I approach through a concept of spirituality which enables us to have a preview on other phenomena and processes that disrupted catholic religiosity. My thesis starts in the Enlightenment which has had a quite underlying ideological influence on the position churches all over the Europe, while I complemented this chapter by secularization aspects of Judeo-Christian tradition. The next chapter dedicating to 19. century is primarily focused on modernization processes (especially Czech nationalism) which could replace or suppressed traditional religiosity. The short 20. century is devided on formation of relationship between state, church and society (The First Republic and communist period) and simoustaneously in framework of this part it is introduced postmodern transformation of religion whose ,,offshoots" characterise present Czech spirituality. The Revolution and the beginning of the 90s are viewed from two approaches - from a dissolution of concensus and from adapted liminal theory. How it seems, the contemporary position of the Catholic Church is given profound processes - especially anti-catholic form of nationalism and a rate of...
|
guest ::
RSS feed.