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The role of microRNA in physiology and pathology
Janáková, Barbora ; Matoušková, Petra (advisor) ; Pávek, Petr (referee) ; Dostálová Merkerová, Michaela (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Barbora Hanousková Supervisor: doc. Ing. Petra Matoušková, Ph.D. Title of Doctoral Thesis: The role of microRNA in physiology and pathology MicroRNAs (miRNAs) are small single-stranded non-coding RNA molecules that play an important role in the regulation of gene expression. They are evolutionarily highly conserved and are present in the genome of all eukaryotic organisms, suggesting their importance in physiological processes. Due to the tissue specificity and their involvement in the pathogenesis of various diseases, miRNAs have been widely studied in connection with their potential use as specific and early biomarkers, or as a therapeutic target. The doctoral thesis, written in the form of an annotated set of publications, dealt with the study of miRNAs in various systems in vitro and in vivo, focusing on their role in adipogenesis and their use as biomarkers of pathological conditions. The introduction summarizes the theoretical information on epigenetic regulation with a more detailed focus on miRNAs, followed by comments on the author's individual publications and brief conclusions. In two studies using mouse models, several miRNAs were identified to have their expression profile...
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Novel bile acid derivatives as a promising therapeutic approach for liver and metabolic disorders
Štefela, Alžbeta ; Pávek, Petr (advisor) ; Vítek, Libor (referee) ; Juřica, Jan (referee)
IN ENGLISH LANGUAGE Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmacology and Toxicology Candidate: Mgr. Alžbeta Štefela Supervisor: Prof. PharmDr. Petr Pávek, PhD. Title of the doctoral thesis: Novel bile acid derivatives as promising therapeutic approach Bile acids (BAs) are amphipathic steroidal molecules that are traditionally known to facilitate intestinal digestion and absorption of lipids and fat-soluble substances. On top, the recent findings have revealed that they represent important signaling agents involved in the orchestration of lipid, glucose and energy metabolism and immune response. BAs exhibit these roles by activating intracellular nuclear receptors such as farnesoid X (FXR), pregnane X (PXR) vitamin D receptors. Furthermore, BAs act as endocrine signaling molecules and activate numerous biological cascades via a membrane G-protein-coupled receptor, termed TGR5. Therefore, the extensive modulation of BA scaffold underwent to identify compounds with specific targeting of above-mentioned receptors as a promising therapeutic approach for the treatment of various liver and metabolic disorders including cholestasis, biliary cirrhosis, nonalcoholic steatohepatitis or diabetes. The principal aim of this doctoral thesis was to investigate the structure...
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Novel bile acid derivatives as a promising therapeutic approach for liver and metabolic disorders
Štefela, Alžbeta ; Pávek, Petr (advisor) ; Vítek, Libor (referee) ; Juřica, Jan (referee)
IN ENGLISH LANGUAGE Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmacology and Toxicology Candidate: Mgr. Alžbeta Štefela Supervisor: Prof. PharmDr. Petr Pávek, PhD. Title of the doctoral thesis: Novel bile acid derivatives as promising therapeutic approach Bile acids (BAs) are amphipathic steroidal molecules that are traditionally known to facilitate intestinal digestion and absorption of lipids and fat-soluble substances. On top, the recent findings have revealed that they represent important signaling agents involved in the orchestration of lipid, glucose and energy metabolism and immune response. BAs exhibit these roles by activating intracellular nuclear receptors such as farnesoid X (FXR), pregnane X (PXR) vitamin D receptors. Furthermore, BAs act as endocrine signaling molecules and activate numerous biological cascades via a membrane G-protein-coupled receptor, termed TGR5. Therefore, the extensive modulation of BA scaffold underwent to identify compounds with specific targeting of above-mentioned receptors as a promising therapeutic approach for the treatment of various liver and metabolic disorders including cholestasis, biliary cirrhosis, nonalcoholic steatohepatitis or diabetes. The principal aim of this doctoral thesis was to investigate the structure...
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Influence of biotransformation and transport to the effects of chemotherapy and prognosis of breast cancer
Hubáčková, Miluše ; Souček, Pavel (advisor) ; Pávek, Petr (referee) ; Janatová, Markéta (referee)
Influence of biotransformation and transport to the effects of chemotherapy and prognosis of breast cancer Abstract: Introduction: The aim of this study was to evaluate the influence of biotransformation enzymes and enzymes of multidrug resistance for the prognosis and effects of chemotherapy of breast cancer patients. The first part dealed with the genetic factors and the influence of genetic polymorphisms in selected biotransformation genes and in ABCB1 gene and their influence in the formation and prognosis of breast cancer. In the second part, the expression profiles of these genes were analyzed as potential prognostic and predictive properties of a treatment's outcome. Materials and methods: Polymorphisms of biotransformation genes were determined using real-time PCR and polymorphisms of ABCB1 gene were determined using NanoChip Assay, which was performed in our cooperated department. In both cases, the polymorphisms were detected in DNA obtained from the blood of patients. The expression of genes was determined using quantitative real-time PCR in paired tumor and adjacent non-tumor tissue of breast cancer samples. The expression of NQO1 and p53 protein was assesed by immunohistochemistry in the tumor breast tissues. Results: The frequencies of the studied SNPs did not differ from the previous...
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Quantitative analysis of tyrosine kinase inhibitors employing liquid chromatography with mass spectrometry detection
Maier, Jan ; Pávek, Petr (advisor) ; Lochman, Lukáš (referee)
The submitted thesis is devoted to the quantitative analysis of tyrosine kinase inhibitors, specifically imatinib and nilotinib, by liquid chromatography-mass spectrometry method. The main purpose of developing this new method of analysis at the Department of Clinical Biochemistry and Diagnostics at the University Hospital in Hradec Králové was measuring and monitoring serum or plasma concentration levels of these drugs in patients with chronic myeloid leukemia, less often in patients with gastrointestinal stromal tumour. The main task during the elaboration of the thesis was to fully optimize and validate the method. Previously, this method for the analysis of tyrosine kinase inhibitors was routinely performed here by high-performance liquid chromatography with spectrophotometric (UV) detection. As part of the modernization of laboratory technology, they started to use high-performance liquid chromatography with mass spectrometry at the workplace. The analytes with their internal standards were obtained by a liquid-liquid extraction process. Then, samples were separated on a C18 reverse phase column using isocratic elution. Subsequently, both analytes were detected by a triple quadrupole tandem mass spectrometer with ESI ion source in a positive mode. As a part of the method validation was to...
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Pharmacological interventions of nuclear receptors in diabettes mellitus
Draský, Jakub ; Pávek, Petr (advisor) ; Bárta, Pavel (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Jakub Draský Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Pharmacological influence of nuclear receptors in diabetes mellitus therapy Nuclear receptors belong to the superfamily of transcription factors, their main functions include regulating the expression of target genes. In my work I focused mainly on the group of orphan receptors, namely the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR). A common feature of these receptors is their activation by a specific ligand. Both CAR and PXR have an essential function as biological sensors of hydrophobic xenobiotics when they induce enzymes I and II. phase of metabolism. They are also essential in the regulation of gluconeogenesis, insulin response, adipogenesis, cholesterol homeostasis, fatty acids, triglycerides and glycogen. The aim of this experimental work was to introduce a luciferase reporter assay method for two DNA constructs containing the promoter region of the PEPCK and CYP7A1 genes. We used the known agonist rifampicin and the antagonist SPA70 to activate/deactivate PXR. We used CITCO as a CAR receptor agonist. We first verified the functionality of the luciferase reporter gene assay...
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Viral hepatitis: novel insights and novel therapeutic interventions.
Davidovová, Eva ; Pávek, Petr (advisor) ; Trejtnar, František (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Bc. Eva Davidovová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Viral hepatitis: novel insights and novel therapeutic interventions. Viral hepatitis is a well-known worldwide problem. Hepatitis B virus (HBV) and hepatitis C virus (HCV) are characterized by the development of serious complications, especially with regard to the transition to the chronic stage of the disease, associated with the development of fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Interactions between viruses and host cells are quite complicated and not always fully understood. In general, the infection cycle of viruses is a multi-step process. A closer understanding of the entire life cycle of the virus is a major prerequisite for the invention of effective drugs. Viral hepatitis B and C have long been treated mainly with interferon alfa. Ribavirin was later added to HCV treatment and nucleoside / nucleotide analogs (NA) were introduced for HBV. Interferon was later pegylated to improve its properties. However, these drugs did not provide sufficient efficacy and were additionally associated with a number of side effects. It is precisely because of these disadvantages of the current...
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DNA methylation changes in oropharyngeal carcinoma
Birknerová, Natália ; Matoušková, Petra (advisor) ; Pávek, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Natália Birknerová Supervisor: doc. Ing. Petra Matoušková, Ph.D. Title of diploma thesis: DNA methylation changes in oropharyngeal cancer Oropharyngeal carcinoma (OPC) is a type of head and neck cancer (HNC) that represents the seventh most common malignancy worldwide. The vast majority (more than 90%) of cases are squamous cell carcinomas (SCC). OPC develops in the tissue of the tongue, tonsils, soft palate, and pharynx. In addition to traditional risk factors, human papillomavirus (HPV) has been identified as an additional independent risk factor for the development of these tumors. Epigenetic alterations refer to heritable changes in gene expression that occur without changes in the underlying DNA sequence and can contribute to carcinogenesis. They include DNA methylation, histone modification and non-coding RNAs effecting gene expression. This study aimed to investigate methylation levels of selected tumor-suppressor genes in oropharyngeal squamous cell carcinoma (OPSCC) in comparison to normal oropharyngeal tissue. DNA methylation levels of selected tumor-suppressor genes were analyzed using Methylation- Specific Multiplex Ligation-dependent Probe Amplification (MS-MLPA) in metastatic tumor...
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The changes in expression of long non-coding RNAs in hepatocellular carcinoma
Krhutová, Magdaléna ; Pávek, Petr (advisor) ; Matoušková, Petra (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Bc. Magdaléna Krhutová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: The changes in expression of long non-coding RNAs in hepatocellular carcinoma Hepatocellular carcinoma (HCC) is one of the highly prevalent cancers globally. A number of new cases of HCC and deaths rises every year. Molecular mechanisms of HCC are being intensively studied, yet they are not still fully understood. In addition to genetic alterations, epigenetics also plays an important role in HCC pathogenesis. Long non- coding RNAs (lncRNAs) are RNA molecules that are not capable of coding proteins, and their length is 200 nucleotides or more. Various studies have already revealed lncRNAs involved in tumorigenesis through binding to DNA, RNA and proteins. New studies also demonstrate significant changes in the expression of biotransformation enzymes in HCC, and interactions with microRNAs (miRNAs) and lncRNAs. This diploma thesis deals with the issue of long non-coding RNAs in relation to HCC. It summarizes the epidemiological situation, risk factors, and current possibilities of diagnosis and therapy of this disease. It also summarizes recently described genetic and epigenetic mechanisms contributing...
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The impact of apixaban on overall hemostatic potential.
Cablíková, Ladislava ; Pávek, Petr (advisor) ; Carazo Fernández, Alejandro (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Bc. Ladislava Cablíková Supervisors: Ass. Prof. Mojca Božič-Mijovski, Ph.D., prof. PharmDr. Petr Pávek, Ph.D., RNDr. Jana Nekvindová, Ph.D. Thesis title: The Impact of Apixaban on Overall Hemostatic Potential Disorders at certain levels of the complicated haemostatic system can lead to either bleeding or excessive blood coagulation. These pathological conditions are treated with anticoagulants, which aim to correct excessive coagulation. However, traditional anticoagulant therapy has many limitations, which initiated efforts to develop oral anticoagulants with a better profile. These new-generation anticoagulants are called DOAC - Direct Oral AntiCoagulans. Apixaban, as one of xabans, has predictable pharmacokinetics and pharmacodynamics and therefore does not require a routine laboratory monitoring of the treatment effect. Nevertheless, it still requires evaluation in urgent clinical situations. Standard coagulation screening assays, e.g., PT (prothrombin test) and APTT (activated partial thromboplastin test), do not fully reflect the actual status of the drug. Therefore, researchers aim is to find a relatively simple and fast hemostatic assay that would correlate with the actual condition...
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