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Epitranscriptomics and cardioprotective interventions
Benák, Daniel ; Hlaváčková, Markéta (advisor) ; Štengl, Milan (referee) ; Bohuslavová, Romana (referee)
1 Ischemic heart disease stands as the foremost global cause of mortality. Myocardial ischemia results in damage to cardiomyocytes which can further lead to impaired heart function. However, the extent of ischemic injury hinges not only on the intensity and duration of the ischemic stimulus but also on cardiac tolerance to ischemia. Therefore, it is extremely important to unravel the molecular basis of cardioprotective interventions such as adaptation to chronic hypoxia or fasting. We focused on the novel epitranscriptomic mechanisms around RNA modifications - N6 -methyladenosine (m6 A) and N6 ,2'-O-dimethyladenosine (m6 Am). Our findings revealed that while most epitranscriptomic modifiers displayed differential regulation in the heart following hypoxic adaptation and fasting, demethylases (ALKBH5 and FTO) were consistently upregulated after these cardioprotective interventions. Furthermore, we detected a discernible reduction in cardiac total RNA methylation levels after fasting. On the contrary, transcripts Nox4 and Hdac1, both of which play a role in the cytoprotective action of ketone bodies, exhibited increased methylation in hearts of fasting rats. Finally, inhibition of epitranscriptomic demethylases ALKBH5 and FTO decreased the hypoxic tolerance of adult rat primary cardiomyocytes isolated from...
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Thin Film Electrodes for Electrochromic Devices
Macalík, Michal ; Kadlec, Jaromír (referee) ; Nováková,, Sabina (referee) ; Sedlaříková, Marie (advisor)
The work deals with the deposition of layers for electrochromic device with different methods. Transparent electrically conductive layer of SnO2 was deposited by pyrolytic decomposition of peroxostannate solution. Hydrogen peroxide in starting solution contributes to the oxidation process of growth layers and to increase the electrical conductivity. Active electrochromic layer of WO3 was electrolytic deposited from the peroxotungstic acid solution. Optimal deposition time and the optimal annealing temperature of deposited layers were found. Passive electrochromic layer of V2O5 was deposited using dip-coating method from peroxovanadate solution. A contribution of solution diluted with distilled water was investigated. Found results were used to construct complete electrochromic device with polymer gel electrolyte.
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Printed transparent oxide conductors
Bartoš, Radim ; Pekárková, Jana (referee) ; Dzik, Petr (advisor)
Thin films of tin oxide doped by fluorine or antimony were prepared by spincoating technique and material printing technique. Plenty of inorganic and organic coumpouds were used as precursors of fluorine and antimony. Sheet resistence of films were measured by 4-probe method and film thickness were determined by contact profilometer. Films were analyzed by Xray diffraction (XRD) and scan electron microscopy (SEM).
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Exploring the role of opioid signaling in modulation of microglial function
Mali, Akash Shivling ; Novotný, Jiří (advisor) ; Svoboda, Petr (referee) ; Machová Urdzíková, Lucia (referee)
Microglial activation is the most important component of neuroinflammation. It appears that opioids may affect microglial M1/M2 polarization in different ways depending on the type of receptor employed. In addition to opioid receptors, Toll-like receptor 4 (TLR4) of the innate immune system can also be activated by some opioid ligands and thus elicit specific cellular responses. Although opioid receptors (ORs) are known to regulate neurotransmission in various peptidergic neurons, their potential role in modulation of microglial function remains largely unknown. In this study, we investigated the effects of OR agonists, namely DAMGO, DADLE, and U-50488, on polarization and metabolic modulation of C8-B4 microglial cells. Our findings have revealed that opioids effectively suppress lipopolysaccharide (LPS)-triggered M1 polarization and promote the M2 polarization state. This was evidenced by decreased phagocytic activity, decreased production of nitric oxide (NO), diminished expression of proinflammatory cytokines such as TNF-α, IL-1β, IL-6, IL-86, and IL-12 beta p40, along with an increased migration rate and elevated expression of anti-inflammatory markers such as IL-4, IL-10, IL-13 arginase 1, and CD206 in microglia compared to cells influenced by LPS. Furthermore, we have demonstrated that...
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The role of RNA demethylase FTO in differentiation, regulation of energy metabolism and sensitivity to streptozotocin of neuronal and glial cell models
Čočková, Zuzana
Fat mass and obesity associated (FTO) demethylase is responsible for erasure of the most abundant epitranscriptomic mark in eukaryotic mRNA, the N6-methyladenosine (m6A) residue. Together with other m6A erasers, writers (methyltransferases) and readers it forms an m6A regulatory pathway that controls the amount, location and biological effect of m6A. The dynamic regulation of the brain's m6A methylome during neurodevelopment is essential for maintaining cerebral functions. In addition, preclinical research suggests that the m6A regulatory pathway regulates energy balance in a tissue- and cell type-specific manner. The FTO gene has been associated with lifelong risks of obesity and metabolic syndrome as well as regulation of total body energy intake and expenditure. However, little is understood about the function of the m6A pathway in control of brain energy metabolism. That is of interest in pursuit of understanding Alzheimer's disease, as this illness is characterized by profound disruptions in cerebral energy metabolism and mounting evidence suggests that disrupted brain bioenergetics may play a role in the disease's early genesis, before the appearance of clinical symptoms. In the present thesis we aimed to investigate the role of FTO in models of two brain cell types, neurons and astrocytes....
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The role of RNA demethylase FTO in differentiation, regulation of energy metabolism and sensitivity to streptozotocin of neuronal and glial cell models
Čočková, Zuzana ; Novotný, Jiří (advisor) ; Horák, Martin (referee) ; Balík, Aleš (referee)
Fat mass and obesity associated (FTO) demethylase is responsible for erasure of the most abundant epitranscriptomic mark in eukaryotic mRNA, the N6-methyladenosine (m6A) residue. Together with other m6A erasers, writers (methyltransferases) and readers it forms an m6A regulatory pathway that controls the amount, location and biological effect of m6A. The dynamic regulation of the brain's m6A methylome during neurodevelopment is essential for maintaining cerebral functions. In addition, preclinical research suggests that the m6A regulatory pathway regulates energy balance in a tissue- and cell type-specific manner. The FTO gene has been associated with lifelong risks of obesity and metabolic syndrome as well as regulation of total body energy intake and expenditure. However, little is understood about the function of the m6A pathway in control of brain energy metabolism. That is of interest in pursuit of understanding Alzheimer's disease, as this illness is characterized by profound disruptions in cerebral energy metabolism and mounting evidence suggests that disrupted brain bioenergetics may play a role in the disease's early genesis, before the appearance of clinical symptoms. In the present thesis we aimed to investigate the role of FTO in models of two brain cell types, neurons and astrocytes....
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Modulation of RNA demethylase FTO function in SH-SY5Y cells: the effect on insulin signaling and mitochondrial respiration
Čočková, Zuzana ; Novotný, Jiří (advisor) ; Hlaváčková, Markéta (referee)
Aim of this thesis was to observe changes in oxidative metabolism and expression of important neuroenergetic proteins in human neuroblastoma cell line SH-SY5Y due to inhibition of FTO. FTO is a RNA demethylase that uses N6-methyladenosine as substrate. Differences in enzyme expression are connected to broad area of effects involving energy homeostasis. Mitochondria are cellular powerhouses, a key elements in production of energy and metabolic substrates, yet a source of potentially dangerous reactive oxygen species (ROS) and analogous reactive molecules. In order to better understand FTO purpose in neuronal energetic metabolism, we examined mitochondrial respiratory chain. Using high-resolution respirometry we were capable of observing impairment in mitochondrial respiration after FTO inhibition. There was considerable decline in endogenous respiration, maximal respiration rate and reserve capacity. In order to obtain more detailed view into mitochondrial respiration, expression levels of electron-transport complexes were quantified by Western blot technique. Slight reduction was identified in subunits of complex I and IV. However, the most prominent alteration was seen in complex II subunit. There were no differences in expression of complex III and ATP synthase subunits. Beside disrupted activity...
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Role of the FTO gene in the genetic determination of common multifactorial diseases
Dlouhá, Dana ; Hubáček, Jaroslav (advisor) ; Černá, Marie (referee) ; Rossmeislová, Lenka (referee)
Obesity is a risk factor for development of cardiovascular disease, diabetes type 2 and some cancers. Newly detected genetic risk factor for body weight is the FTO gene ("fat mass and obesity associated"). The aim of this thesis was determine 1) whether the presence of risk alleles correlate with BMI in Czech population and to determine 2) whether there is an association between variants in the FTO gene and risk of myocardial infarction/ acute coronary syndrome (MI/ ACS), 3) renal failure (ESRD), or 4) incidence of colorectal cancer (CRC). We analyzed polymorphisms rs17817449 (first intron) and rs17818902 (3rd intron) using by PCR-RFLP and then also RT PCR. We found an association of the first intron variant (but not the 3rd one) and BMI in Czech control population. We have detected an association of 1st intron SNP and BMI changes during the intervention study in obese children, but not in obese females. We found a correlation between the risk allele and increased risk of ACS (OR 1.49) in patients with MI. In patients with ESRD was detected association between the risk allele and the risk of disease (OR 1.37). We didn't confirmed the association between rs17817449 and the development of CRC. Representative selected groups of the Czech populations "MONICA" and "HAPPIE" were used as controls. One...
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The m6A pathway at the molecular level and its role in neurological diseases
Švendová, Aneta ; Černá, Barbora (advisor) ; Čočková, Zuzana (referee)
N6-methyladenosine is the most abundant modification in eukaryotic messenger RNA. This modification is reversible, thanks to a complex of methyltransferases and demethylases. The biological effects of m6 A are mediated through reader proteins. This complex mechanism of proteins contributes to many molecular processes such splicing, translation and transport. It also plays a role in many serious neurological diseases, such as Alzeheimer's disease, Parkinson's disease, major depressive disorder and attention deficit hyperactivity disorder. The purpose of this thesis is to describe the m6 A pathway, its regulation at the molecular level and to put it into context with neurological diseases of today. Key words: mRNA, mRNA metabolism, N6-methyladenosine, m6 A regulation, FTO, METTL, ALKBH5, neurodegenerative disorders, Alzheimer's disease, Parkinson's disease, major depressive disorder, ADHD
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The genetic background of obesity and its treatment with bariatric surgery
Lischková, Olga
Obesity is a frequent metabolic disease that causes many other health and socioeconomic complications. Obesity arises due to excessive energy intake and decrease in energy expenditure, which is a conseqence of contemporary lifestyle. Moreover, obesity has a strong genetic component. Common obesity is polygenic, multifactorial disease, in which individual genes interact with each other and with environmental factors. Genome-wide association studies, conducted between 2006-09, led to the discovery of dozens of gene loci that predispose individuals to obesity. The strongest signals were registered for polymorphisms in FTO (fat mass and obesity-associated) and near a gene MC4R (melanocortin 4 receptor). However, the contributions of these variations on the phenotype of obesity are very small, therefore, it is necessary to validate the results of such robust studies. It is very important to uncover the effects of genetic variants for understanding the molecular mechanisms of energy metabolism. The studies presented in this thesis refer about the impact of polymorphisms in selected genes on anthropometric and metabolic parameters of the patients of the Institute of Endocrinology and of healthy volunteers who underwent functional tests. Our cohort includes a representative sample of Czech children (COPAT...
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