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Oxidative phosphorylation system in rare types of mitochondrial diseases
Zdobinský, Tomáš ; Tesařová, Markéta (advisor) ; Pecinová, Alena (referee)
In their bioenergetic metabolism mammalian cells are primarily dependent on ATP production through the oxidative phosphorylation system (OXPHOS). Defects of OXPHOS function can lead to occurrence of mitochondrial disorders with different severity and diverse symptoms. Most severely affected are usually tissues with high energy demand which are also difficult to access for biochemical and other examinations. The aim of this thesis was mainly to characterize the effects of mutations in seven different genes (OPA1, DARS2, NDUFS8, NR2F1, HTRA2, MGME1, POLG) on bioenergetic metabolism and mitochondrial network structure of skin fibroblasts from eight different patients diagnosed with mitochondrial disorders. The main method used was measurement of oxygen uptake by permeabilized cells using highly sensitive polarography. Significant changes in fibroblast respiration of four patients were found. Changes in mitochondrial network morphology were found in two of those and two other patient cell lines compared to controls using fluorescent microscopy and different cultivating conditions. Skin fibroblasts are relatively easy to obtain and offer a number of benefits for both diagnostic and study purposes. The results of this work illustrate the possibilities of their use for validation of potential causal...
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Deletions in human mitochondrial DNA and causes of their formation
Zdobinský, Tomáš ; Tesařová, Markéta (advisor) ; Kazantsev, Dmitry (referee)
Mitochondria are organelles of eukaryotic cells that primarily provide energy metabolism, but also participate in metabolic processes such as biosynthesis of amino acids, heme groups, Fe-S clusters etc. Mitochondrial disorders represent heterogeneous group of diseases which can occur in both child and adult life. They affect various tissues and organs in different ways, most often manifesting themselves as disorders of nervous system, skeletal muscle, liver, kidneys or endocrine system. Mitochondrial DNA deletions contribute to pathogenesis of many of those diseases and they are a symptom of several defined syndromes. They most likely arise as a result of replication stalling resulting in a double strand break of DNA. This can be caused primarily by pathogenic changes in replication apparatus and nucleotide metabolism proteins. The aim of this work is to summarize the knowledge about mitochondria and structure and replication of their genome, but also to create a summary of the most important proteins whose mutation leads to mitochondrial diseases accompanied by deletions in mtDNA and to outline the mechanism by which they arise.
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