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Congenital disorders of glycosylation and their impact on cellular and energetic metabolism
Rychtárová, Lucie ; Hansíková, Hana (advisor) ; Hodek, Petr (referee) ; Pecinová, Alena (referee)
Inherited disorders of glycosylation (CDG) are a large group of more than 160 types of metabolic disorders caused by genetic defects that lead to impaired glycan biosynthesis and modification. The lipid dolichol plays an essential role in glycan biosynthesis. Glycans play a key role in the function and structure of proteins and lipids and their deficiency leads to severe clinical symptoms. CDG usually manifests in childhood as a multisystem disorder. Families thus face a serious health problem due to the progressive and highly variable nature of the disease, the unfavorable prognosis and, with few exceptions, the unavailability of treatment. Currently, we still do not have a sufficient range of methods to recognize rare types of CDG and our knowledge of the pathophysiology of CDG is still limited. The first aim of this work was to optimize the method of determination of dolichol isoforms and to study them in physiology and pathology. The second aim of the work was to investigate the bioenergetic status and overall metabolism in the most common type of CDG - deficiency of phosphomannomutase 2 (PMM2-CDG), and in CDG caused by a defect in dolichol biosynthesis. The distribution of urinary dolichol isoforms in the population was characterized using an optimized method. The dolichol isoforms profile...
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The role of mitochondrial energy metabolism in cell senescence
Zima, Michal ; Trnka, Jan (advisor) ; Pecinová, Alena (referee)
Cellular senescence represents status, when the cells cease to divide and remain in permanent cell cycle arrest. Senescence is considered to be an active response of the cell to various extrinsic and intrinsic types of stress such as certain oncogene activation, exposing to several cytokines or drugs and damaged and/or uncapped telomeres. Senescent cells are characterised by extensive modification of gene expression, flattened and enlargement of cellular body. Hypothetically, altered gene expression may lead also to increase of certain surface proteins expression. Such protein can be L1 cell adhesive molecule (L1CAM), which is expressed heterogeneously within the population. This Thesis describes current knowledge of cellular senescence, the mechanism, which may result in establishment of senescence phenotype, and also the characteristic markers of senescence. Thesis also puts together the heterogeneity of L1CAM expression in A375 senescent cells with oxygen consumption rate and extracellular acidification rate performed by Seahorse XFe24 metabolic analyser. Therefore, ells were sorted according to their levels of expressing L1CAM onto low and high L1CAM expressing subpopulations. Obtained data show potential correlation between the rate of L1CAM expression in A375 cells and the metabolic rate. Key...
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Analysis of testing for the presence of alkohol and other addictive substances during the performance of profession in railroad traffic in the Czech Republic in the years 2008 - 2012
Pecínová, Alena ; Šejvl, Jaroslav (advisor) ; Kuda, Aleš (referee)
The aim is to map the field of testing for the presence of alcohol and other addictive substances during the performance of profession in railway traffic, comparison of results of testing for the presence of alcohol and other substance during the performance of profession in railway transport in the period 2008 - 2012 in companies: Czech Railways, joint stock company, ČD Cargo, a joint stock company and the Railway Infrastructure Administration, state organization and comparison of practical measures with Guidance on managing safety risks related to the Influence of Alcohol, Drugs and/or Psychoactive Medication issued by the International Union of Railways. The work is focused on the guidelines of listed companies, testing methods for the presence of alcohol and other addictive substances listed companies, and the Czech Republic legislation dealing with this issue. There were used for the processing "Guidance on managing safety risks related to the Influence of Alcohol, Drugs and/or Psychoactive Medication", policy documents and regulations, data from annual reports and safety and health at work reports of listed companies in period 2008 - 2012, and information provided by the Department of the system of safety rail operation, which analyses incidents in all these three railway companies. All three...
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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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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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Molecular bases of sensitivity to electron transport chain inhibition-induced cell death
Blecha, Jan ; Rohlena, Jakub (advisor) ; Brábek, Jan (referee) ; Pecinová, Alena (referee)
1 Abstract in English Mitochondrial electron transport chain (ETC) targeting shows a great promise in cancer therapy. However, why modern ETC-targeted compounds are tolerated on the organismal level and what are the molecular reasons for this tolerance remains unclear. Most somatic cells are in a non-proliferative state, and features associated with the ETC in quiescence might therefore contribute to specificity. Thus, we investigated the ETC status and the role of two major consequences of ETC blockade, reactive oxygen species (ROS) generation and inhibition of ATP production, in cell death induction in breast cancer cells and in proliferating and quiescent non-transformed cells. First, we characterised the effect of a newly developed ETC inhibitor mitochondria- targeted tamoxifen (MitoTam) in in vitro and in vivo tumour models of breast cancer with varying status of the Her2 oncogene. We document that Her2high cells and tumours have increased assembly of respiratory supercomplexes (SCs) and increased complex I-driven respiration in vitro and in vivo. They are also highly sensitive to MitoTam. Unlike the parental compound tamoxifen, MitoTam efficiently suppressed experimental Her2high tumours without systemic toxicity. Mechanistically, MitoTam inhibits complex I- driven respiration and disrupts respiratory...
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Proposal of addiction interventions for employees of Railway Infrastructure Administration (RIA) Needs analysis and implementation possibilities of preventive measures
Pecínová, Alena ; Popov, Petr (advisor) ; Barták, Miroslav (referee)
The aim of this thesis is to analyze the needs of employer in the field of addictology- side. Secondary targets include: a) analysis of training needs of employees enrolled in the employer's training program for the design of educational-preventive project intended for these RIA (Railway Infrastructure Agency) employees; b) analysis of UIC recommendations addictological interventions aimed for management of safety risks associated with the use of alcohol and other substance abuse at work; c) analysis of implementation possibilities of the recommendations according to the previous point. An analysis of documents - Czech legislation, recommendations UIC - focused on measures regulating the use of alcohol and other substance abuse at work and addictological interventions proposed by UIC. Knowledge questionnaire for employees participated in employer's training program was prepared. Respondents were not informed in advance about the offer to participate in the research, they weren't informed about the addictology knowledge test. During at about 60 minutes there were completed 3100 answers from 31 respondents without training with 40 - 70% correct results (in average 53.4 %). There were differences in correct answers in some parts of questionnaire based on information knowledge of respondents. On the...
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Proposal of addiction interventions for employees of Railway Infrastructure Administration (RIA) Needs analysis and implementation possibilities of preventive measures
Pecínová, Alena ; Popov, Petr (advisor) ; Kuda, Aleš (referee)
The aim of this thesis is to analyze the needs of employer in the field of addictology-side. Secondary targets include: a) analysis of training needs of employees enrolled in the employer's training program for the design of educational-preventive project intended for these RIA (Railway Infrastructure Agency) employees; b) analysis of UIC recommendations addictological interventions aimed for management of safety risks associated with the use of alcohol and other substance abuse at work; c) analysis of implementation possibilities of the recommendations according to the previous point. An analysis of documents - Czech legislation, recommendations UIC - focused on measures regulating the use of alcohol and other substance abuse at work and addictological interventions proposed by UIC. Knowledge questionnaire for employees participated in employer's training program was prepared. Respondents were not informed in advance about the offer to participate in the research, they weren't informed about the addictology knowledge test. During at about 60 minutes there were completed 3100 answers from 31 respondents without training with 40 - 70% correct results (in average 53.4 %). There were differences in correct answers in some parts of questionnaire based on information knowledge of respondents. UIC...
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The role of mitochondrial energy metabolism in cell senescence
Zima, Michal ; Trnka, Jan (advisor) ; Pecinová, Alena (referee)
Cellular senescence represents status, when the cells cease to divide and remain in permanent cell cycle arrest. Senescence is considered to be an active response of the cell to various extrinsic and intrinsic types of stress such as certain oncogene activation, exposing to several cytokines or drugs and damaged and/or uncapped telomeres. Senescent cells are characterised by extensive modification of gene expression, flattened and enlargement of cellular body. Hypothetically, altered gene expression may lead also to increase of certain surface proteins expression. Such protein can be L1 cell adhesive molecule (L1CAM), which is expressed heterogeneously within the population. This Thesis describes current knowledge of cellular senescence, the mechanism, which may result in establishment of senescence phenotype, and also the characteristic markers of senescence. Thesis also puts together the heterogeneity of L1CAM expression in A375 senescent cells with oxygen consumption rate and extracellular acidification rate performed by Seahorse XFe24 metabolic analyser. Therefore, ells were sorted according to their levels of expressing L1CAM onto low and high L1CAM expressing subpopulations. Obtained data show potential correlation between the rate of L1CAM expression in A375 cells and the metabolic rate. Key...
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Analysis of testing for the presence of alkohol and other addictive substances during the performance of profession in railroad traffic in the Czech Republic in the years 2008 - 2012
Pecínová, Alena ; Šejvl, Jaroslav (advisor) ; Kuda, Aleš (referee)
The aim is to map the field of testing for the presence of alcohol and other addictive substances during the performance of profession in railway traffic, comparison of results of testing for the presence of alcohol and other substance during the performance of profession in railway transport in the period 2008 - 2012 in companies: Czech Railways, joint stock company, ČD Cargo, a joint stock company and the Railway Infrastructure Administration, state organization and comparison of practical measures with Guidance on managing safety risks related to the Influence of Alcohol, Drugs and/or Psychoactive Medication issued by the International Union of Railways. The work is focused on the guidelines of listed companies, testing methods for the presence of alcohol and other addictive substances listed companies, and the Czech Republic legislation dealing with this issue. There were used for the processing "Guidance on managing safety risks related to the Influence of Alcohol, Drugs and/or Psychoactive Medication", policy documents and regulations, data from annual reports and safety and health at work reports of listed companies in period 2008 - 2012, and information provided by the Department of the system of safety rail operation, which analyses incidents in all these three railway companies. All three...
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