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Study of effect of carcinogenic benzo[a]pyrene on cytochromes P450 and cytochrome b5 expression in organs of the rat
Blecha, Tomáš ; Moserová, Michaela (advisor) ; Dračínská, Helena (referee)
Polycyclic aromatic hydrocarbons are a large group of organic compounds which consist solely of carbon and hydrogen atoms, two or more fused aromatic rings. They represent group of persistent organic pollutants (POPs) present in all components and fields of the environment. Benzo[a]pyrene is characteristic compound of polycyclic aromatic hydrocarbons formed by five fused aromatic rings. Benzo[a]pyrene is procarcinogen with genotoxic effects, which is metabolically activated by variety of enzyme systems such as cytochrome P450 and epoxide hydrolase to reactive metabolite of BaP-7,8-diol-9,10-epoxide (BPDE) and 9-hydroxy-4,5-epoxy-BaP. These reactive metabolites can form covalent DNA adducts. In the present work, we studied the influence of carcinogenic benzo[a]pyrene on the expression of cytochrome P450 (CYP) 1A1, 1A2 and cytochrome b5 in livers, kidneys and lungs of laboratory rats. Total RNA was isolated and afterwards converted into cDNA with the participation of random primers. Using polymerase chain reaction in real time (RT-PCR) the relatively gene expression of CYP1A1, CYP1A2 and cytochrome b5 was quantified in the organs of rats treated with BaP and control (untreated) animals to a reference gene (β-actin). It was found that benzo[a]pyrene significantly increases expression of CYP1A1 and...
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Forensic analysis of epigenetic factors and mRNA
Andreasová, Karolina ; Šimková, Halina (advisor) ; Zachová, Markéta (referee)
In last years, forensic genetics focused on a research of epigenetic factors and mRNA. It turned out that their analysis provides valuable information that can be obtained from a small amount of a biological trace and which included e.g. estimation of circumstances of death, age estimation, discrimination within monozygotic twins, identification of tissues and body fluids, sample authentication, determination of paternal allele, etc. In this thesis, it is discussed methods of epigenetic factors and mRNA analysis and their potential application in future forensic practice.
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Role onkogenní mikroRNA-155 a proto-onkogenu MYB u chronické lymfatické leukémie
Vargová, Karina ; Stopka, Tomáš (advisor) ; Móciková, Heidi (referee) ; Trka, Jan (referee)
(EN) Chronic lymphocytic leukemia (B-CLL) represents a disease of mature-like B-cells. Due to failed apoptosis but also due to enhanced proliferative signals, the leukemic B-cells accumulate in the peripheral blood, bone marrow, lymph nodes and spleen. The clinical course of B-CLL is very heterogeneous; in some patients B-CLL progresses very rapidly into an aggressive form. Such patients need therapy sooner while in other patients with indolent B-CLL the onset of therapy takes years. Several standard prognostic and disease progression markers are used for disease staging and monitoring, however a reliable marker that will suggest when to start therapy is unknown. Expression of small, non-coding microRNAs is often deregulated and represent important prognostic markers in variety of cancers including leukemia. Hence in our study we concentrated to miR-155, an important molecule regulating differentiation of hematopoietic cells, inflammation process and antibody production. Its aberrant expression was described in Hodgkin`s as well as in non-Hodgkin`s lymphoma, including indolent lymphoproliferations like B- CLL. Our results confirmed elevated levels of both, primary miR-155 transcript and mature form of miR-155 in our B-CLL patient samples (N=239). The aberrant expression of miR-155 in B-CLL samples...
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Messenger RNA stability and microRNA activity in mouse oocytes
Flemr, Matyáš ; Svoboda, Petr (advisor) ; Motlík, Jan (referee) ; Hampl, Aleš (referee)
The oocyte-to-zygote transition represents the only physiological event in mammalian life cycle, during which a differentiated cell is reprogrammed to become pluripotent. For its most part, the reprogramming relies on the accurate post-transcriptional control of maternally deposited mRNAs. Therefore, understanding the mechanisms of post-transcriptional regulation in the oocyte will help improve our knowledge of cell reprogramming. Short non- coding microRNAs have recently emerged as an important class of post-transcriptional regulators in a wide range of cellular and developmental processes. MicroRNAs repress their mRNA targets via recruitment of deadenylation and decapping complexes, which typically accumulate in cytoplasmic Processing bodies (P-bodies). The presented work uncovers an unexpected feature of the microRNA pathway which is found to be suppressed in fully-grown mouse oocytes and through the entire process of oocyte-to-zygote transition. This finding is consistent with the observation that microRNA-related P-bodies disassemble early during oocyte growth and are absent in fully-grown oocytes. Some of the proteins normally associated with P-bodies localize to the oocyte cortex. At the final stage of oocyte growth, these proteins, together with other RNA-binding factors, form subcortical...
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Analysis of selected gene transcripts in the rat myocardium adaptated to chronic hypoxia
Kašparová, Dita ; Žurmanová, Jitka (advisor) ; Nováková, Olga (referee)
Dita Kašparová Chronická hypoxie a exprese genů 4 Abstract Adaptation to chronic hypoxia (CH) is characterized by a variety of functional changes in order to maintain metabolic and energy homeostasis. It has been known for many years that both humans and animals indigenous or adapted to high-altitude hypoxia are more tolerant to an acute ischemic injury of the heart. Cardioprotective mechanisms activated by adaptive responses to chronic hypoxia can be the result of altered transcriptional regulations in left ventricles. Here we report results from the gene expression profiling of adaptive responses in three models of chronically hypoxic heart. Adult male Wistar rats were exposed for 21 days to either continuous normobaric hypoxia (CCH; 10% O2) or CCH interrupted daily by 1-hour reoxygenation (RCH) or CCH interrupted daily by 16-hour (CIH). Cardiprotective effect of CCH adaptation is abolished by brief daily reoxygenation, RCH adaptation. In the present study, we aimed to determine myocardial mRNA expression of 19 candidate genes divided into three important groups: i) Hypoxia inducible factor (HIF1α) and its prolyl and asparaginyl hyroxylases (PHDs and FIH respectively, ii) Creatine kinase (CK) isoenzymes which play important role in energy homeostases of heart and iii) the group of main enzymatic...
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The role of microRNAs in chronic lymphocytic leukemia
Moravec, Martin ; Macůrková, Marie (referee) ; Hájek, Miroslav (advisor)
2 Abstract MicroRNAs (miRs, miRNAs) are recently discovered molecules (19-25 nucleotides long) that regulate gene expression at post-transcriptional level by either blocking protein synthesis or mRNA degradation. As a part of gene silencing mechanism, miRNAs are involved in cellular processes, such as apoptosis, cell proliferation, development and viral defence. miRNAs have been intensely studied in connection to disease pathogenesis. Chronic lymphocytic leukemia (CLL) is the most common leukemia in Western countries affecting mostly elderly people. In my work I focus on explanation of miRNA functions and their contributions to chronic lymphocytic leukemia (CLL). I describe previously published data about miRNA-15, miRNA-16, miRNA-143, miRNA-145 and miRNA-155 in connection to this disease. Based on recent reports, I also discuss the potential role of miRNA-326 in CLL pathogenesis.
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The effect of cancerogenic azo dye Sudan I on expression of biotransformation enzymes
Hejduková, Žaneta ; Dračínská, Helena (referee) ; Svášková, Dagmar (advisor)
Sudan I is a widely used azo dye which has the ability to cause carcinomas in organs and tissues of experimental animals. During reactions catalyzed by microsomal monooxygenase enzyme systems or cytoplasmic biotransformation enzymes, Sudan I is oxidized to reactive metabolites that covalently bind to nucleic acids and cause their damage. Sudan I can also be metabolized by reduction, e. g. by a DT-diaphorase enzyme (NQO1). Reduction of Sudan I is considered to be a detoxification reaction. In this work, the in vivo action of Sudan I is examined in terms of its ability to induce an expression of the biotransformation enzyme DT-diaphorase in tissues of rats treated with the azo dye. The aim of this work was to quantify the degree of NQO1 induction at mRNA level. After the isolation of total RNA from organs of rats treated with Sudan I, the RNA was converted to cDNA by reverse transcription using random hexamers as primers. Using specific probes, the abundance of mRNA for the enzyme NQO1 in the organs of treated rats was quantified by "real-time" PCR, relatively to the control gene with a constant expression (β-actin). Through comparing thus determined amounts of mRNA in individual organs of treated and untreated rats, it has been found that Sudan I had caused a significant increase in the expression...
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