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Monitoring of mitochondria in living cells using fluorescence methods
Taliánová, Eliška ; Čejková, Darina (referee) ; Čmiel, Vratislav (advisor)
This bachelor thesis deals with the topic of monitoring mitochondria and evaluating their activity. The structure and function of cellular mitochondria are briefly discussed. Contains a list of commonly available fluorescent dyes. It investigates the possibilities of influencing mitochondria in vitro. It deals with the influence of mitochondrial membrane potential by means of various concentrations of glucose and subsequently evaluates mitochondrial activity using the dye JC-10.
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Sirtuin 3 and its function in cardiac metabolism
Procházka, Marek ; Horníková, Daniela (advisor) ; Koňaříková, Eliška (referee)
SIRT3 is a NAD+ -dependent deacetylase, that is abundant in the heart and essentially regulates cellular processes in cardiomyocytes. SIRT3 positively modulates most of the enzymes and proteins in the intermediate metabolism in mitochondria, which supply the necessary ATP energy for the heart muscle and that are centers of metabolism in cardiomyocytes. In the mitochondria SIRT3 inhibits the formation of ROS by activating an antioxidant system. SIRT3 has significant anti-apoptotic, anti-hypertrophic and anti-fibrotic cardioprotective effects. Its activation is based on several drugs and natural substances that could be a promising therapeutic approach to the treatment of cardiovascular diseases, which are currently the leading cause of death of more than a half of the European population. However, more studies are required for better understanding the processes in which SIRT3 is involved. The aim of this work is to summarize the function of SIRT3 in mitochondrial metabolism and cardiac physiology. Key words: sirtuin 3, cardioprotection, metabolism, heart, mitochondria
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Three-parent babies - new therapies of mitochondrial disorders
Helebrandtová, Veronika ; Pecinová, Alena (advisor) ; Ješina, Pavel (referee)
Mitochondria are essential parts of living cells, as they play a key role in cellular metabolism, especially in energy production. Due to their unique structure, the energy released during the oxidation of the substrates can be used to form the ATP. Mitochondria also contain their own DNA (mtDNA), which is maternally inherited and encodes catalytic subunits of oxidative phosphorylation complexes. Mitochondrial disorders of nuclear or mitochondrial origin, are common causes of inherited diseases and affect mainly the tissues with high energy requirements, such as heart or brain. Treatment of mitochondrial diseases is usually symptomatic and does not lead to complete recovery of the patient. As a result, new causal therapies, such as a gene therapy, are currently investigated. However, using this approach it is necessary to consider the origin of the mutation. Gene therapy of mitochondrial diseases of mtDNA origin is very complicated, therefore the new treatment strategy, mitochondrial replacement therapy, has been proposed. The principle of this technique is to prevent the transmission of mutated mtDNA from mother to offspring by transferring the nuclear genome of mother with mitochondrial disorder into donor's denucleated oocyte with healthy mitochondria. In this way, the child has genetic...
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Non-pharmacological approaches towards treatments of mitochondrial disorders
Kudrnovská, Barbora ; Mráček, Tomáš (advisor) ; Kalous, Martin (referee)
Mitochondrial diseases are characterized by a high degree of genotypic and phenotypic heterogeneity, but as a whole group, they represent one of the most common forms of inborn errors of metabolism. Treatment of these diseases is still very limited, despite massive body of ongoing research projects, which are focussed on new therapeutic approaches. Besides pharmacological methods, there are also non-pharmacological approaches such as certain diet regimens, physical activity or exposition to hypoxia. Due to their relatively generalized mechanism of action, these methods may offer benefits to a wider range of mitochondrial patients, despite different primary genetic defects. This work summarizes the existing knowledge about the use of exercise, ketogenic diet, and hypoxia application as possible therapeutic approaches towards the treatment of mitochondrial diseases. Keywords: mitochondria, mitochondrial diseases, therapy, exercise, ketogenic diet, hypoxia
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Mitochondrial dysfunctions in depression
Javorská, Jana ; Stuchlík, Aleš (advisor) ; Kolář, David (referee)
Depression (major depressive disorder; MDD) is a common and chronic psychiatric disorder. MDD causes a significant reduction in the quality of life, reduces the ability to work, and thus causes a burden on disabled patients, healthcare, and all of society. The pathophysiological features of depression are often ambiguous, and therefore current treatment for depression is insufficient. Chronic stress plays an important role in the pathogenesis of depression. It causes dysregulation of the hypothalamic-pituitary-adrenal axis (HPA axis) controlling cortisol release, imbalance of neurotransmitters and neuromodulators, and imbalance between pro- inflammatory and anti-inflammatory cytokines. Another neurobiological sign of depression is a disruption of energy metabolism in the brain, for which mitochondria are responsible. In addition to ATP, mitochondria also form reactive oxygen species (ROS). ROS are in excessive amounts during chronic stress and disrupt cellular energy. Therefore, substances related to mitochondrial energy and antioxidant effects are currently being investigated. Ketamine, dizocilpine and phencyclidine are particularly effective in mitochondrial metabolism. However, the clinical use of these substances is complicated by their hallucinogenic and neurotoxic effects. Antioxidants help...
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Monitoring of mitochondria in living cells using fluorescence methods
Taliánová, Eliška ; Čejková, Darina (referee) ; Čmiel, Vratislav (advisor)
This bachelor thesis deals with the topic of monitoring mitochondria and evaluating their activity. The structure and function of cellular mitochondria are briefly discussed. Contains a list of commonly available fluorescent dyes. It investigates the possibilities of influencing mitochondria in vitro. It deals with the influence of mitochondrial membrane potential by means of various concentrations of glucose and subsequently evaluates mitochondrial activity using the dye JC-10.
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Computational modelling of mechanical tests of animal cell
Orlová, Lucie ; Fuis, Vladimír (referee) ; Burša, Jiří (advisor)
Předkládaná diplomová práce se zabývá stavbou živých živočišných buněk a jejich odezvou na mechanické zatěžování. Zobecněným zaměřením práce je popis mechanického chování buňky nejenom ve fyziologickém, ale i v patologickém stavu. Výchozím předpokladem pro úspěšné řešení zadané úlohy je vysoce interdisciplinární přístup kombinující výpočtové přístupy mechaniky těles (v~tomto případě metodu konečných prvků) s lékařským výzkumem. Nejdůležitějším bodem při tvorbě výpočtového modelu, pomocí něhož je možné aproximovat chování živé buňky při zatížení, je zejména identifikace mechanicky významných komponent a~jejich materiálových parametrů. V tomto případě jsou jako mechanicky význačné identifikovány spojité součásti jádro, membrána a cytoplazma, které jsou nově propojeny s prvky diskrétními (mitochondriální sítí) v hybridním modelu, jehož platnost je ověřena pomocí experimentálních dat. Tento model slouží jako podklad k vyhodnocení míry vlivu mitochondrií na celkovou tuhost buňky.
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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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Bacterial proteins in the biogenesis of mitochondria of unicellular eukaryotes.
Petrů, Markéta
in English Formation of mitochondria by the conversion of a bacterial endosymbiont is the fundamental moment in the evolution of eukaryotes. An integral part of the organelle genesis was the displacement of the endosymbiont genes to host nucleus and simultaneous creation of new pathways for delivery of proteins synthesized now in the host cytoplasm. Resulting protein translocases are complexes combining original bacterial components and eukaryote-specific proteins. In addition to these novel protein import machines, some components of the original bacterial secretory pathways have remained in the organelle. While the function of a widely distributed mitochondrial homolog of YidC, Oxa1, is well understood, the role of infrequent components of Sec or Tat translocases has not yet been elucidated. So far, more attention has been paid to their abundant plastid homologs, which assemble photosynthetic complexes in the thylakoid membrane. In the thesis, the structure and function of prokaryotic YidC, Sec and Tat machineries and their eukaryotic homologs are described. By comparing both organelles of the endosymbiotic origin, the hypothesis is drawn on why these translocases have been more "evolutionary successful" in plastids than in mitochondria.
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Biochemical and molecular studies of cytochrome c oxidase and ATP synthase deficiencies
Fornůsková, Daniela ; Zeman, Jiří (advisor) ; Hyánek, Josef (referee) ; Stiborová, Marie (referee)
Mgr. Daniela Fornuskova PhD thesis Biochemical and molecular studies of cytochrome c oxidase and ATP synthase deficiencies ABSTRACT The mammalian organism fully depends on the oxidative phosphorylation system (OXPHOS) as the major energy (ATP) producer of the cell. Disturbances of OXPHOS may be caused by mutations in either mitochondrial DNA (mtDNA) or nuclear DNA (nDNA). One part of the thesis is focused on the role of early and late assembled nuclear-encoded structural subunits of cytochrome c oxidase (CcO) as well as Oxa1l, the human homologue of the yeast mitochondrial Oxa1 translocase, in the biogenesis and function of the human CcO complex using stable RNA interference of COX4, COX5A, COX6A1 and OXA1L, as well as expression of epitope-tagged Cox6a, Cox7a and Cox7b, in HEK (human embryonic kidney)- 293 cells. Our results indicate that, whereas nuclear- encoded CcO subunits Cox4 and Cox5a are required for the assembly of the functional CcO complex, the Cox6a subunit is required for the overall stability of the holoenzyme. In OXA1L knockdown HEK-293 cells, intriguingly, CcO activity and holoenzyme content were unaffected, although the inactivation of OXA1 in yeast was shown to cause complete absence of CcO activity. In addition, we compared OXPHOS protein deficiency patterns in mitochondria from skeletal...
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