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Type II secretion system in gram negative bacteria.
Klápšťová, Veronika ; Doležal, Pavel (advisor) ; Mach, Jan (referee)
This thesis describes the structure and the assembly of the type II secretion system which is found in some gram negative bacteria such as Vibrio cholerae, Pseudomonas aeruginosa, Legionella pneumophila or Klebsiella oxytoca and which transports their exoproteins across the outer membrane. The thesis also presents some important substrates translocated by this system - toxins, enzymes and proteins involved in bacterial metabolism or motility - and hypothetical mechanism of the secretion. Conclusion is focused on the discovery of the secretion system in mitochondria of some protists and its importance for the research of the evolution of eukaryotic organisms.
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Changes in oxidative phosphorylation during development of cellular senescence
Zima, Michal ; Hodný, Zdeněk (advisor) ; Kašparová, Dita (referee)
Cellular senescence represents a state of permanent cell cycle arrest. It is considered to be an active response of the cell to various extrinsic and intrinsic types of stress, which are damaged and/or uncapped telomeres, activation of certain oncogenes, DNA damage and effects of several cytokines. This thesis describes current mechanisms which may result in establishment of senescence phenotype, putting those facts in association with changes in oxidative phosphorylation. In thesis are also mentioned features of senescence cells and their impact on the neighborhood. Special attention is focused on the role of reactive oxygen species in promotion of cellular senescence, mechanisms of their elevation, the role of NADPH oxidases and the inhibition of mitochondrial oxidative phosphorylation complexes by activity of cytokine signaling pathways STAT3 and TGFbeta. Key words: cellular senescence, reactive oxygen species, cytokines, mitochondria, oxidative phosphorylation chain, NADPH oxidases, Signal Transducer and Activator of Transcription 3 (STAT3), TGF-β, DNA damage response (DDR)
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The role of mitochondrial creatine kinase and hexokinase in cardioprotective mechanisms induced by chronic hypoxia
Wasková, Petra ; Žurmanová, Jitka (advisor) ; Drahota, Zdeněk (referee) ; Nováková, Olga (referee)
IN ENGLISH The ischemia-reperfusion (I/R) injury, which is a consequence of myocardial infarction, represents a major cause of death worldwide. One of the most effective cardioprotective interventions increasing the resistance of hearts to the I/R injury is the adaptation to a chronic hypoxia (CH). However, the molecular mechanisms of CH are still not well understood. The most important factors responsible for the I/R injury are reactive oxygen species (ROS) produced by complexes I and III within the mitochondrial electron transport chain. Potential candidates maintaining ROS at a low level are mitochondrial creatine kinase (mtCK) and two hexokinase isoforms (HK1 and HK2). These enzymes highly support the mitochondrial oxidative phosphorylation by increasing the availability of ADP for complex V of the respiratory chain. In addition, the HK binding to mitochondria inhibits binding of the pro- apoptotic protein BAX, thereby protecting cardiac cells against apoptosis. Besides the mitochondrial CK isoform, there are two cytosolic CK (CKM and CKB) present in cardiomyocytes that help to maintain energy homeostasis. Based on the known anatomical and physiological differences between the left (LV) and the right (RV) ventricles, the first study focused on the comparing ventricles in terms of the energy...
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Effects of selected biologically active natural compounds on mammalian energy metabolism
Seďová, Karolína ; Flachs, Pavel (advisor) ; Doleželová, Šárka (referee)
Obesity is associated with many diseases, particularly type 2 diabetes, hypertension, dyslipidemia , hyperglycemia and cardiovascular diseases. If a patient experiences at least a trio of these diseases are diagnosed metabolic syndrome. The prevalence of obesity and related diseases has increased at an alarming rate in recent years and has become a worldwide health problem. Cardiovascular disease is the leading cause of death worldwide. Some minor components which are commonly found in our diet (eg some plant metabolites), directly or indirectly affect energy metabolism of our body and thus can reduce or even prevent developing health complications associated with the metabolic syndrome. This thesis, aims to clearly summarize the findings and results of scientific studies on selected plant metabolites occurring in extracts of specific plants that have preventive or therapeutic effects on obesity and other components of the metabolic syndrome. This work describes the use of these substances in the pharmaceutical industry and the food industry, the primary natural resources , and mechanisms of action , including effective doses. Powered by TCPDF (www.tcpdf.org)
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The effect of endothelial cell proliferation on susceptibility to mitochondrially controlled apoptosis and oxidative stress.
Blecha, Jan ; Novák, Petr (advisor) ; Plecitá, Lydie (referee)
Mitochondria are multifunctional organelles playing a key role in energy metabolism and cell death induction. Mitochondria, and specifically their respiratory chain, are also the main producers of reactive oxygen species (ROS) in cells. Metabolism can be affected by the state of cellular proliferation and certain ROS-inducing agents have an antiangiogenic effect based on the preferential elimination of proliferating endothelial cells (EC). Therefore, in this work we investigated, whether mitochondria could be responsible for different sensitivity of proliferation and confluent EC to cell death. We mainly focused on systems that regulate ROS level and apoptosis: respiratory chain (ROS production), antioxidant defense (ROS detoxification) and Bcl-2 family of proteins (apoptosis regulation). First, we treated EC with functional and nonfunctional respiratory chain with various oxidative stress- and apoptosis-inducing agents and determined ROS production and susceptibility to apoptosis in proliferating and confluent cells. Our results show that functional respiratory chain greatly increases the susceptibility of proliferating cells to ROS induction and apoptosis, whereas in qiescent cells it protects against cell death. Given these findings, we assessed the activity of respiratory chain in proliferating...
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Effects of antidepressants and depressive disorders on mitochondrial functions
Hroudová, Jana ; Fišar, Zdeněk (advisor)
Mood disorders are serious diseases. Nevertheless, their pathophysiology is not sufficiently clarified. Biological markers that would facilitate the diagnosis or successful prediction of pharmacotherapy are still being sought. The aim of the study was to find out whether mitochondrial functions are affected by antidepressants, mood stabilizers and depression. Our research is based on recent hypotheses of mood disorders, the advanced monoamine hypothesis, the neurotrophic hypothesis, and the mitochondrial dysfunction hypothesis. We assume that impaired function of mitochondria leads to neuronal damage and can be related to the origin of mood disorders. Effects of antidepressants and mood stabilizers on mitochondrial functions can be related to their therapeutic or side effects. In vitro effects of pharmacologically different antidepressants and mood stabilizers on the activities of mitochondrial enzymes were measured in mitochondria isolated from pig brains (in vitro model). Activity of monoamine oxidase (MAO) isoforms was determined radiochemically, activities of other mitochondrial enzymes were measured spectrophotometrically. Overall activity of the system of oxidative phosphorylation was measured electrochemically using high- resolution respirometry. Methods were modified to measure the same...
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Yeast retrograde signaling pathway between mitochondria and the nucleus
Podholová, Kristýna ; Palková, Zdena (advisor) ; Fraiberk, Martin (referee)
Retrograde signaling pathway is the pathway between mitochondria and nucleus. This pathway helps Saccharomyces cerevisiae to cope with worsening of conditions of life, such as depletion of rich nutrient sources and necessity of use poor resources, reduction of mitochondrial membrane potential, or loss of mitochondrial DNA causing disturbances in the citric acid cycle. Most of these conditions are associated with aging yeast populations. Key retrograde pathway proteins include RTG transcription factors Rtg1p/Rtg3p and cytoplasmic protein Rtg2p. Retrograde pathway is upregulated by several positive and negative regulators including the TOR pathway, which negatively regulates retrograde pathway. The retrograde pathway target genes include genes coding for tricarboxylic cycle enzymes, peroxisomal enzymes, transporters and other enzymes of anaplerotic pathways. Retrograde response help cells to modify their metabolism so that they are able to overcome unfavorable environmental conditions in which they live. Powered by TCPDF (www.tcpdf.org)
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System of oxidative phosphorylation and methods for its functional analysis
Daňhelovská, Tereza ; Tesařová, Markéta (advisor) ; Míčová, Petra (referee)
The system of oxidative phosphorylation (OXPHOS) is main and essential source of energy (ATP) in eukaryotic cells. It is complex process situated in the inner mitochondrial membrane in which 4 enzymes of the respiratory chain, 2 mobile carriers and the ATP synthase participate. Defect in any part of OXPHOS may lead to the mitochondrial disorders. An incidency of the mitochondrial disorders is estimated to be 1:5000 and mitochondrial disorders are the most common inherent metabolic diseases. Clinically, it is extremely heterogeneous group of disorders affecting primarily tissue with high energy demand, for example brain, hearth and muscle. With regard to broad spectrum of clinical symptoms and relatively poor genotype-phenotype correlation of mitochondrial genetic defects, it is preferred to perform complete laboratory diagnostic tests including several biochemical and molecular genetic approaches. This bachelor thesis summarizes actual information about mechanism of OXPHOS and describes three major approaches to detect its functionality. Three major approaches are: spectrophotometric measurement of individual OXPHOS enzyme activities, measuring capacity of the mitochondrial energy generating system using various radioactive labeled substrates and measuring of oxygen consumption by polarography. To...
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Expression of selected defects of oxidative phosphorylation system in cultivated fibroblasts
Marková, Michaela ; Hansíková, Hana (advisor) ; Kalous, Martin (referee)
AAbbssttrraacctt:: The mammalian organism is entirely dependent on ATP production by oxidative phosphorylation system (OXPHOS) on the inner mitochondrial membrane. OXPHOS is composed of respiratory chain complexes I-IV, ATP synthase and also include two electron transporters cytochrome c and coenzyme Q. Disorders of mitochondrial energy metabolism caused by OXPHOS defects are characterized by extreme heterogeneity of clinical symptoms, variability of tissues affected and the severity of the defect at the level of individual tissues. The mitochondrial disorders are not always clearly expressed at the level of available tissue or most easily available cultured fibroblasts and/or currently available methods are not capable to detect the defects on the fibroblasts level. The aim of this master thesis was to identify by biochemical methods, especially by high sensitive polarography, OXPHOS disorders in cultured fibroblasts. Cell lines from 10 patients with isolated (SURF21, SCO1 ND1, ND5) or combined defects of OXPHOS complexes whose biochemical defect was confirmed in muscle tissue as well as 14 patients with non- mitochondrial diseases (8 patients with Huntington disease, 6 patients with disorder of sulphur amino acids metabolism) were analysed. Furthermore impact of various cultivation conditions on OXPHOS...
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Imaging of fluorescently labelled mitochondria using Biplane_FPALM microscopy
Dostál, Marek ; Plášek, Jaromír (advisor) ; Valenta, Jan (referee)
Title: Imaging of fluorescently labelled mitochondria using Biplane FPALM microscopy Author: Bc. Marek Dostál Department: Fyzikální ústav UK Supervisor: prof. RNDr. Jaromír Plášek, CSc. Consultants: RNDr. Petr Ježek, DrSc. Mgr. Hana Engstová, Ph.D. Abstract: In this thesis the results are presented of a search for suitable PCFP markers for a visualization inner and outer mitochondrial membranes by unique biplane FPALM microscope. We participated in debugging the measuring software for the purpose of 3D vizualization mitochondrial nets and determine their parameters. For this purpose were developed two methods. The first one can be applied to determine the parametrs of outer membranes and the second one is applicable to determine parametrs of inner membranes. On smaller statistical file we managed to confirm dependence quality of mitochondrial net on the cell culture conditions. Under stress conditions e.g. hypoxia the quality of mitochondrial net is changed. Keywords: mitochondria, Biplane FPALM, GFP, 3D visualization.
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