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Preparation of expression vectors for the production of ZIKA and Dengue NS3 helicase
Daňhelová, Kateřina ; Konvalinka, Jan (advisor) ; Heidingsfeld, Olga (referee)
Zika and Dengue viruses have spread, due to globalisation, to all continents which lie at least in part in the subtropic and tropic climatic zones. This spread of these viruses is a reason of an increasing number of severe diseases caused by them. New drugs, which would be effective against these infections, could be an answer to this challenge. Various viral proteins, among them also viral helicase, which is the topic of this bachelor thesis, can be a suitable drug target. The task was to prepare expression constructs for production of recombinant helicases of Zika and Dengue viruses via the suitable bacterial strain Escherichia coli. Several constructs derived from plasmid pET-16b were prepared with inserted helicase of Zika and Dengue viruses. One of them was used for the preparation of recombinant purified helicase of Zika virus, that will be used for further research. [IN CZECH] Keywords: Flavivirus, Zika virus, Dengue virus, helicase, expression, purification, enzyme activity [IN CZECH]
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Expression of selected membrane transporters in placentas of pregnant women diagnosed for preterm rupture of membranes
Michalská, Martina ; Čečková, Martina (advisor) ; Jirkovský, Eduard (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Martina Michalská Supervisor: doc. PharmDr. Martina Čečková, Ph.D. Title of diploma thesis: Expression of selected membrane transporters in placentas of pregnant women diagnosed for preterm rupture of membranes Placenta is a key organ for pregnancy maintenance. One of its main functions is transport of compounds between mother and her fetus. The transplacental penetration is ensured due to membrane transporters that are present in the apical or basal side of trophoblast. Their expression level is affected by many physiological and pathological factors, among others it can be influenced by infection and inflamatory reaction. Inflammation is also one of the risk factors of preterm deliveries and it can be therefore assumed that these pathological states are accompanied by changes in expression of placental transporters. This study was performed using 51 placentas obtained from Faculty hospital in Hradec Králové from women who underwent preterm delivery and on 15 placentas delivered in term. The study employed quantitative RT-PCR approach. The mRNA expression of membrane transporters ABCB1, ABCG2, OATP1A2, OATP1B3, OATP2A1, OATP2B1, OATP3A1, OATP4A1 was assessed and the results were compared to...
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Immunogenetic and hormonal markers of predisposition to systemic rheumatic diseases particularly systemic lupus erythematosus
Fojtíková, Markéta ; Pavelka, Karel (advisor) ; Hrnčíř, Zbyněk (referee) ; Rovenský, Jozef (referee)
Fojtikova 2011 INTRODUCTION: Several factors like genetic susceptibility is required for systemic rheumatic diseases development. Immunomodulatory PRL effect supports autoimmunity. AIMS: 1. To detect the immunogenetic background (alleles HLA class I, II and microsatellite polymorphism of the transmembrane part exon 5 of MIC-A gene) of SLE and PsA. 2. To detect PRL serum and synovial fluid with regard to clinical and laboratory RA activity. 3. To find the role of the functional polymorphism -1149G/T SNP PRL of extrapituitary promoter of PRL gene in SLE, RA, PsA, SSc and inflammatory myopathies development. METHODS: Genetic analyses of pateints with SLE (n=156), RA (n=173), PsA (n=100), SSc (n=75), PM (n=47) a DM (n=68) and 123 healthy individuals: PCR-SSP (HLA clase I and II), PCR-fragment analysis (MIC-A) a PCR-RFLP (-1149 G/T SNP PRL). In 29 RA a 26 OA PRL serum and synovial fluid concentrations were detected using immunoradiometric assay. RESULTS: 1. The allele HLA-DRB1*03 (pc=0.008; OR 2.5) and haplotype HLA-DRB1*03-DQB1*0201 (pc <0.001; OR 4.54) were determined as risk immunogenetic markers for SLE in Czech population. In SLE versus controls allele MIC-A5.1 was increased (pc =0.005; OR 1.88). MIC-A5.1 together with HLA-DRB1*03 increases the risk for SLE development, pc <0.000001; OR 9.71....
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The effect of selected endocrine disruptors on cytochromes P450 1B1 and 3A1/2
Holecová, Jana
Many exogenous and endogenous compounds are referred to as endocrine disruptors (EDCs), as they interfere with natural synthesis, signaling and metabolism of endogenous hormones. Common exogenous endocrine disruptors are benzo(a)pyrene (BaP) and 17α-ethinylestradiol (EE2). Endogenous endocrine disruptor 17β-estradiol (E2) is frequently present in the environment as well. In this thesis, the effect of the mentioned EDCs and their combinations on gene and protein expression of CYP1B1, 3A1 and 3A2 in rat liver, kidney and lung was determined. Protein expression was studied using Western blot method and specific antibodies; gene expression was assessed by quantitative PCR. Moreover, the effect of tested EDCs and their combinations on BaP metabolism and CYP3A specific activity (measured as testosterone 6β-hydroxylation) were studied in liver microsomal samples. It was confirmed, that BaP significantly increases CYP1B1 expression in rat liver and lung both alone and together with EE2 or E2. Pretreatment of rat with E2 and BaP increases the ability of BaP to induce CYP1B1 expression. On the contrary, EE2, E2 and their combination decrease the CYP1B1gene expression. The rate of BaP metabolites formed in liver microsomal samples increases in rats pretreated with BaP and its combinations. In liver, there was...
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Changes in Expression of Membrane Molecules CD200R, CD95, CD95L, and Soluble CD200R Regulating inflammatory Responses in Patients Undergoing Cardiac Surgery
Holmannová, Drahomíra ; Krejsek, Jan (advisor) ; Brát, Radim (referee) ; Turánek, Jaroslav (referee)
Cardiac surgery is known to initiate a complex physiological response with the immune system activation (SIRS), neurohormonal response, metabolic changes, coagulopathies etc. SIRS is triggered by tissue injury, myocardial ischemia, reperfusion, use of anaesthesia, cardioplegia, extracorporeal circuit etc. Excessive immune system activation is associated with progression of SIRS, life-threatening multi-organ dysfunction (MOD), and increased morbidity/mortality in the postoperative period. The immune system response is regulated and terminated by both cellular and humoral regulatory and inhibitory mechanisms including changes in expression of in our study monitored molecules: CD200/CD200R, sCD200R and CD95/CD95L. Methods: The study included the measurement the expression of CD95, CD95L, CD200R, and sCD200R molecules in granulocyte and monocyte populations in blood samples of 30 patients who underwent heart surgery using CPB. Samples collected before surgery, after surgery, and in the postoperative period (1st , 3rd , 7th day) were analysed by flow cytometry and sCD200R by ELISA. Results: We discovered a significant increase in the percentage of granulocytes expressing inhibitory molecule CD200R (from 5% to 17.8%) instantly after surgery. It might be presumed that these cells are less susceptible to...
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Art group 12/15 Better Late than Never and its position in the Czech art after year 1989
Mayerová, Miroslava ; Bláha, Jaroslav (advisor) ; Čech, Viktor (referee)
Miroslava Mayerová Abstract Annotation Mayerová, Miroslava: Art group 12/15 and its position in the Czech art after year 1989. [Diploma thesis] Prague, 2019. Charles University in Prague, Faculty of education, Art education department. Title of the Diploma's Thesis: Art group 12/15 and its position in the Czech art after year 1989 ABSTRACT: The aim of the thesis is to describe and analyze the position of the Art Group 12/15 Better Late than never at Czech art scene after the year 1989, with an emphasis on generational changes and postmodern turn. In didactic part, the intention was to establish the awareness of the art group in the minds of students and, based on interdisciplinary ties, to repeat and broaden their knowledge of contexts. The theoretical part of the thesis is based on historical, art-historical and social science literature. To illustrate the time, atmosphere and position of Group 12/15 as precisely as possible, semi-structured and unstructured interviews with the group members themselves were used as a methodology. The diploma thesis shows that Group 12/15 and its generation are the bridge between the 2nd avant-garde, postmodern and contemporary art, maintaining the continuity of the Czech art scene. The didactic and practical part points out the importance of intergenerational discussion...
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MEDIATRE OF COMMUNICATION IN CONTEMPORARY PAINTING
Kopecká, Marie ; Pfeiffer, Jan (advisor) ; Kornatovský, Jiří (referee)
The content of this bachelor thesis is a comprehensive insight into the problematics of communication in art with the aim of application of specific theoretical principles into artistic experience with emphasis on the aesthetic education. Theoretical part of this thesis consists of two parts. First one is focusing on the historical development of the role of art in society, perception of art and starting points, which led artists to specific steps. The second part is focusing on the present. Here I mention the theoretical approaches to the issue of perception of art and visual communication, focus on what role the viewer plays in the process of communication and I analyze communication strategies of selected Czech contemporary artists. In the practical part I am dealing with approaches in my own art work, which is specific application of conclusions mentioned in the theoretical part. In this part I am focusing more on the process of my own work, on choices of particular themes, its specific rendition and following reflection with the aim of verification of particular effects of each medium, which should be integral part of every work in art. The didactic part contains theoretical starting points, which led to making didactic series and my own designed didactic series for the first degree of...
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Expression and purification of recombinant proteins of human Aichi virus and other kobuviruses
Ludvík, Tomáš ; Bouřa, Evžen (advisor) ; Petrvalská, Olívia (referee)
Viral 2Apro plays many important functions in the life cycle of virus. It is protease that is involved in the cleavage of the viral polyprotein in the VP1/2A region. The structure of protein contains active site, thanks to this the protein is able to bind various substrates in the host cell, which is essential for viral infection. With the help of 2Apro , the virus can bypass the host immune response and stop the production of key host proteins. Therefore, 2Apro is suitable target for antiviral intervention. In this work were prepared two recombinant proteins from Aichi virus and Coxsackievirus B3. These are viruses of the family Picornaviridae, which belong among the human pathogens that in some cases cause serious diseases. Aichi virus is the cause of gastroenteritis and Coxsackievirus B3 in the worst cases causes dilated cardiomyopathy. Cardiomyopathy leads to poor heart function due to it is enlargement and can lead to heart failure. (In Czech) Key words: recombinant proteins, 2Apro , Aichi virus, Coxsackievirus B3
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Effect of tyrosine kinase inhibitor cabozantinib and cytotoxic alkaloid ellipticine on expression and activity of cytochromes P450 1A1, 1A2 and 1B1
Měkotová, Barbora ; Dračínská, Helena (advisor) ; Jeřábek, Petr (referee)
In recent years, tyrosine kinase inhibitors have been more and more used for the targeted cancer therapy, due to their ability to disrupt intracellular signalling pathways associated with the development of tumours. Cabozantinib is the tyrosine kinase inhibitor which has been approved for the treatment of thyroid cancer and it is also effective against several other types of cancer. However, multiple drugs combination is often used in anticancer therapy, which may result in their cytochrome P450-mediated interactions. Although this may affect the therapeutic effect of the drugs and cause adverse effects on the organism, very little is known about the effect of cabozantinib on biotransformation enzymes. Therefore, the effect of cabozantinib not only alone but also in combination with the known cytostatic ellipticine on the expression and the activity of cytochromes P450 1A1, 1A2 and 1B1 in rat liver and kidney in vivo was studied in this work. The gene expression was determined by quantitative PCR, the amount of protein was studied by Western blotting and consecutive immunodetection. The enzyme activity was studied using specific marker reactions, 7-ethoxyresorufin O-deethylation for CYP1A1, 7-methoxyresorufin O-demethylation for CYP1A2 and 17β-estradiol 4-hydroxylation for CYP1B1. Our results...
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Vliv ZnO nanočástic na obsah fenolických látek v Solanum lycopersicum
Zelinka, Radim
Zinc oxide nanoparticles are highly variable materials capable of producing positive and negative responses to their effects on plants. Negative effects are often associated with their excessive concentration, which can cause oxidative stress in plants.Conversely, suitable dosages lead to positive manifestations of growth and improvement in the properties studied. ZnO nanoparticles provide plants with a more acceptable form of zinc that is well accessible and easier to use. However, information about their pos-sible toxic effect in combination with another stress factor is lack. The aim of this work was to compare and evaluate the effects of zinc in the form of nanoparticles (ZnO NPs) with the effects of zinc bound in chelate bonding (Zn-EDTA), in combination with UV-C stress factor in plants Solanum lycoperscicum L. Results of this pilot experiment show that both ZnO and Zn-EDTA nanoparticles of 10.95 mg Zn2+ /l in combination with UV-C radiation have a negative effect on vitality and induce oxidative stress in treated plants. The decisive effect on the toxic effect was based on the concentration used and the form of zinc applied. Differences were expres-sed in expression levels of genes affected by oxidative stress. The results obtained from the expression analysis of the genes made it possible to observe the individual transcripts used to respond to oxidative stress and also to provide information on the possible sy-nergistic effect of UV-C radiation and zinc in both studied forms (Zn-EDTA and ZnO NPs).
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