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On purinoceptors in the urinary bladder of hte rat: altered effects of purinergic P1A1 receptors in cystitis
Veselá, Renata ; Wsól, Vladimír (advisor) ; Soukup, Ondřej (referee)
This project was focused on studies of the location and function of selected purinergic receptors in the rat urinary bladder. Its special concerns are on adenosine P1A1 purinoceptors and on their expression in healthy and inflamed urinary bladders of the rat. It was found that both P2X1 and P1A1 receptors are expressed in urinary bladder. The quantity of P1A1 receptors is decreased in the inflamed bladder. Functional studies based on an agonist stimulation (adenosine) showed importance of P1A1 receptors for relaxation of the tissue. During the cystitis this effect was reduced. Electric field stimulation (EFS) pointed out the difference between the response to a stimulus affecting just postsynaptic receptors (adenosine stimulation) and responses to nerve stimulation including both post- and presynaptic effects.
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Studies on carbonyl reductases 1 and 3 variants with emphasis on S-nitrosoglutathione as substrate and inactivator
Hartmanová, Tereza ; Wsól, Vladimír (advisor) ; Kvasničková, Eva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Tereza Hartmanová Supervisor: Dr. Claudia Staab, Dr. Hans-Jörg Martin, Prof. Ing. Vladimír Wsól, Ph.D. Title of the diploma thesis: Studies on carbonyl reductases 1 and 3 variants with emphasis on S-nitrosoglutathione as substrate and inactivator Human CBR1 and CBR3 (carbonyl reductases 1 and 3) are monomeric, NADPH-dependent enzymes belonging to the short chain dehydrogenase/reductase superfamily. Although they are highly similar at the amino acid level (72 % identity) the enzymes exhibit considerable differences in substrate specificity. The CBR1 substrate spectrum is well described, including a variety of compounds e.g. the endogenous indol isatin, S-nitrosoglutathione (GSNO), prostaglandins, quinones, and many carbonyl group bearing xenobiotics. In contrast, CBR3 shows a distinct and much narrower range of substrates and its role is still not fully clarified. Nevertheless, the dissimilar substrate spectra strongly indicate that CBR1 and CBR3 play different metabolic roles. In the present study, the catalytic properties of CBR1 towards the latest CBR1 substrate described, GSNO, were investigated. CBR3 was assessed for potential GSNO-reducing activity, but no in vitro activity was...
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The Role of Reductases in Cancer.
Škarydová, Lucie ; Wsól, Vladimír (advisor) ; Hodek, Petr (referee) ; Bílková, Zuzana (referee)
Only a small attention was paid for long time to reducing enzymes, but today it is clear that these are an important part of the endogenous metabolism and also the phase I biotransformation of xenobiotics. The significant group of reducing enzymes are carbonyl reductases that belong to two superfamilies - short chain dehydrogenases/reductases (SDR) and aldo-keto reductases (AKR). Their role in cancer is now intensively studied and their functions in cancer it is possible to divide into two main sections. It is known that carbonyl reductases play a substantial role in hormone-dependent cancers as prostate, breast or endometrial cancer. Active estrogens or androgens are important growth factors for these cancers because they evoke increasing of cell proliferation so that elevated possibility of mutations of important genes and development of cancer. Carbonyl reductases along with other enzymes (e.g. aromatase) participate in formation of these active sex hormones in extragonadal tissues, so an inhibition of such enzymes may be a target of anticancer therapy of hormone-dependent cancers. It is necessary to determine which enzymes are essential for the formation of active sex hormones in particular types of cancers. Besides hormone-dependent cancers, carbonyl reductases play also role in cancers that...
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Scheme " Heath city"
Valčíková, Šárka ; Wsól, Vladimír (advisor) ; Kvasničková, Eva (referee)
1028 inhabitants of the town Kroměříž were investigated in the project "Healthy Town" in the years 2005, 2006 and 2007. Using acquired data, some epidemiological characteristics of region's inhabitants were evaluated and important experiences were put into preventive clinical practice. The experiences are following: 1. The project "Healthy Town" is not a pilot study. Contact centers are attended by citizens who pay attention to active interest of their health. In this population women and senior citizens dominate, many of them have health problems and the incidence of cardiovascular factors considerably exceeds "the average population". The study has shown that 37% of them are overweight and 13% are obese. 16% of inhabitants have hypertension of over 140/90 mm Hg. 57% suffer from lipid metabolism failure and 14% suffer from hyperglycemia. 2. Then we observed a relationship between BMI index and investigated high risk markers (systolic and diastolic pressure, serum concentration of total cholesterol, LDL cholesterol, HDL cholesterol, triglycerides and glucose). We have proved a significant relation between the degree of overweight and values of blood pressure and concentration of triglycerides, and on the other hand we have discovered a statistically insignificant relation between total cholesterol...
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Purification and characterization of membrane-bounded reductases from human liver
Pelcová, Vendula ; Wsól, Vladimír (advisor) ; Kvasničková, Eva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Vendula Pelcová Supervisor: Prof. Ing. Vladimír Wsól, Ph.D. Title of rigorous thesis: Purification and characterization of membrane-bound reductase from human liver. This study was focused on implementation of experimental techniques for purification of human liver microsomal fraction. There were chosen three basic steps of purification process. As a first step, ion-exchange chromatography was chosen followed by two purification steps on the base of gel filtration. Before purification the microsomal fraction had to be solubilized to release proteins from membrane bounds and conditions had to be optimized. Single fractions were subsequently incubated with specific substrate of reductases oracin and the quantity of metabolite 11-dihydroracin was assessed by achiral HPLC analysis. For the most active fractions the SDS-PAGE analysis was done to set size of unknown enzyme. By use of antibody against 11beta-HSD1 was axcleded the presence of this reductase in all analyses fractions. By use of described purification techniques we were successful to concentrate fraction with unknown human liver reductase.
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Purification and characterization of membrane-bound reductase / / human liver tissue
Pelcová, Vendula ; Wsól, Vladimír (advisor) ; Kvasničková, Eva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Vendula Pelcová Supervisor: Prof. Ing. Vladimír Wsól, Ph.D. Title of rigorous thesis: Purification and characterization of membrane-bound reductase from human liver. This study was focused on implementation of experimental techniques for purification of human liver microsomal fraction. There were chosen three basic steps of purification process. As a first step, ion-exchange chromatography was chosen followed by two purification steps on the base of gel filtration. Before purification the microsomal fraction had to be solubilized to release proteins from membrane bounds and conditions had to be optimized. Single fractions were subsequently incubated with specific substrate of reductases oracin and the quantity of metabolite 11-dihydroracin was assessed by achiral HPLC analysis. For the most active fractions the SDS-PAGE analysis was done to set size of unknown enzyme. By use of antibody against 11beta-HSD1 was axcleded the presence of this reductase in all analyses fractions. By use of described purification techniques we were successful to concentrate fraction with unknown human liver reductase.
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Versatile use of liquid chromatography and mass spectrometry in drug metabolism studies
Suchanová, Bohumila ; Wsól, Vladimír (advisor) ; Solich, Petr (referee) ; Lemr, Karel (referee)
Human organism has always been exposed to a vast array of chemicals encountered in the environment. Chemical revolution has significantly influenced biological evolution of humans leading to serious unpredictable toxicities. In response to continual chemical stress they have developed a variety of enzymes to transform these xenobiotics. Xenobiotics are mostly highly lipophilic and cannot readily be excreted from the body without metabolism to more hydrophilic, water-soluble metabolites. Not only environmental chemicals represent xenobiotics but also drugs, dietary components etc. Biotransformation studies play an important role in the drug discovery and development process. Usually data from drug metabolism is required before a new substance can advance towards the development stages of a new therapeutic agent. Data on metabolism is frequently used to optimize drug candidates, suggest more active compounds or support toxicology studies. The increased flux of new chemical entities into drug discovery has placed an increased need for fast and reliable information on the metabolism of these substances. Liquid chromatography coupled with mass spectrometry can meet demands for rapid drugs and metabolites analysis imposed by modern drug discovery strategies. This dissertation thesis presents an evidence...
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Development and applications of affinity carrier for isolation of human carbonyl-reducing enzymes
Andrýs, Rudolf ; Wsól, Vladimír (advisor) ; Šebela, Marek (referee) ; Šatínský, Dalibor (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Rudolf Andrýs Supervisor: Prof. Ing. Vladimír Wsól, Ph.D Title of dissertation thesis: DEVELOPMENT AND APPLICATION OF AFFINITY CARRIER FOR ISOLATION OF HUMAN CARBONYL-REDUCING ENZYMES For several millennia the human medicine is based on application of small bioactive molecules that are administered in the form of plant extracts or synthetic compounds. However, their use in modern medicine is not possible without a detailed understanding of their biochemical effects and identification of their molecular targets. Chemical proteomics based on the specific recognition between the bioactive molecule and the target molecule is currently the most widely used techniques for identification of molecular targets of small molecules. Compared to conventional biochemical methods (e.g. 2D electrophoresis), chemical proteomics represents particularly sensitive and very selective technique that enable successful identification of biomolecules from complex biological samples that are naturally presented in very small concentrations. Carbonyl- reducing enzymes, which play an important role in physiology due to their involvement in metabolism of various endogenous (e.g. prostaglandins, steroid...
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