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Effect of soluble endoglin to endothelial cells in vitro
Klingová, Rebeka ; Fikrová, Petra (advisor) ; Doleželová, Eva (referee)
Charles Univerzity in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Rebeka Klingová Supervisor: PharmDr. Petra Fikrová, Ph.D. Title of diploma thesis: Effect of soluble endoglin to endothelial cells in vitro Aim: We determined the effects of soluble endoglin on endothelial cells, by the means of inflammatory markers. The aim of the study was to point out the possible association of the soluble endoglin with endothelial dysfunction. Methods: For our study we have selected the human endothelial cells from umbilical vein - HUVEC. We have influenced the cells with soluble endoglin in two concentrations and at two different time intervals. The results were evaluated in the statistical program, in which we have compared the control group with influenced cells and two concentrations between each other. The expressions of inflammatory markers were analyzed on the level of mRNA, using the real-time PCR method. Results: Significant changes in the expression of markers were observed on the vascular and intracellular adhesion molecule at both concentrations compared to the control group. Increased values of transcription were available also for cyclooxygenase 2 and decreased values for cadherin 5 compared to the control group. Conclusion: Changes in...
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Diagnostics of exanthematic childhood diseases
Slámová, Zuzana ; Doleželová, Eva (advisor) ; Prašnická, Alena (referee)
Zuzana Slámová Diagnostics of exanthematic childhood diseases Bachelor thesis Charles University, Faculty of Pharmacy in Hradec Králové Health bioanalytics Background: The aim of the bachelor thesis was to determine the morbidity, incidence and prevalence of selected exanthematic childhood diseases (measles, scarlet fever, rubella, fourth disease, fifth disease and chickenpox) in the period 2013 - 2016 at the Department of Clinical Microbiology and Immunology of Regional Hospital Liberec, a. s. including a comparison with occurance of diseases in the whole Czech Republic. Methods: The methodical part was focused on the methods which are the basis of specific analyzers at the Department of Clinical Microbiology and Immunology of the Regional Hospital Liberec, a. s. These methods are ELISA, CLIA, KFR, PCR and cultivation. Analyzers used to diagnose antibodies against selected diseases are Alegria® (ORGENTEC), LIAISON® XL (DiaSorin) and Chorus Trio (DIESSE). For the direct quick detection of the disease was used quantitative PCR (qPCR) method on the CFX96 Touch ™ Real-Time PCR Detection System (BioRad). Detection of bacterial agents was performed by cultivation. Results: The results were processed over the period 2013 - 2016 from data provided by the Department of Clinical Microbiology and Immunology...
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Progression of endothelial dysfunction in heart in a mouse model of atherosclerosis.
Filipová, Petra ; Němečková, Ivana (advisor) ; Doleželová, Eva (referee)
Charles University in Prague, Faculty od Pharmacy in Hradec Králové Department of Biological and Medical Sciences Progression of endothelial dysfunction in heart in a mouse model of atherosclerosis Author of diploma thesis: Petra Filipová Supervisor: RNDr. Ivana Němečková, Ph.D. Backround: Atherosclerosis is a disease when atherosclerotic lesions are formed in vessels. It represents the major cause of the heart failure and stroke. The endothelial dysfunction, which has been recorded in a large number of animals, is considered one of the main causes of atherosclerosis. In our work we have focused on the detection of the endothelial dysfunction markers in the heart of the ApoE/LDLR double knock-out mouse model. Methods: We detected changes in the expression of eNOS, p-eNOS and HO-1 in two-months and eight-months old ApoE/LDLR double knock-out mice, which were fed standard chow diet. For the semiquantitative evaluation of the observed markers we used the Western blot analysis of the homogenated left ventricle. Results: According to our results there is no difference in the expression of eNOS, p-eNOS and HO-1 in the studied mouse model of atherosclerosis in relation to age. This might mean, that the endothelial dysfunction in the coronary vessels of these mice doesn't develop. However, for further...
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Influence of resveratrol on bile flow.
Paclíková, Kristýna ; Doleželová, Eva (advisor) ; Cermanová, Jolana (referee)
Kristýna Paclíková Influence of resveratrol on bile flow Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Pharmacy Background: Protective effect of resveratrol is shown in many experimental models of cholestasis, but its effect on bile production in healthy individuals has not been studied yet. As the use of resveratrol is frequent in population, the aim of this thesis was to investigate the effect of resveratrol on the bile flow in healthy animals and the clarification of the mechanism of this effect. Methods: Wistar rats (n = 6, in each group, weighing 280 to 320 g) were divided into two groups: control group (Control) and the group of rats administered with resveratrol (10mg/kg/day, p.o.) for 28 days (RES). In vivo clearance study was performed to analyze bile production. Analysis of mRNA and protein expression of the transport proteins was performed by qRT-PCR and Western blot. Results: Resveratrol led to a significant increase in the cumulative bile flow. Analysis of mRNA and protein expression of the transport proteins revealed that bile flow is changed due to posttranscriptional induction of canalicular transporters Bsep and Mrp2. Simultaneously, resveratrol led to induction of Mrp4 efflux protein, expressed at the basolateral membrane of hepatocytes....
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Characterization of a synthetic leoligin derivative, with agonistic FXR and enhancing macrophage cholesterol efflux activity
Kovářová, Lenka ; Kovařík, Miroslav (advisor) ; Doleželová, Eva (referee)
Charles University, Faculty of Pharmacy in Hradec Králové, Department of Biological and Medical Sciences University of Vienna, Faculty of Life Sciences, Department of Pharmacognosy Candidate: Lenka Kovářová Supervisor: Pharmdr. Miroslav Kovařík, Ph.D. Consultant: Dr. Angela Ladurner Title of the diploma thesis: Characterization of a synthetic leoligin derivative, with agonistic FXR and enhancing macrophage cholesterol efflux activity Atherosclerosis is a pathologic multifactorial process triggering the development of cardiovascular diseases, which are the leading causes of death in the western world. The initial phase of atherosclerosis is characterized by the accumulation of lipid particles, mainly low-density lipoproteins (LDL) and macrophage-derived foam cells in large arteries, leading to the gradual thickening of the vessel wall. These progressive alterations elicit plaque formation, followed by rupture, thrombosis and finally can lead to a cardiovascular event. Reverse cholesterol transport is an important preventive mechanism, which ensures removal of excessive atherogenic lipoproteins from macrophages. This efflux is facilitated by ATP binding cassette transporters, mainly ABCA1 and ABCG1 and in part by scavenger receptor B1 (SR-B1). Several nuclear receptors, including PPARγ, LXRα and LXRβ...
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The effect of GLP-1 (glucagon-like peptide-1) analogues on rat hepatocytes in vitro
Krejčíková, Veronika ; Nachtigal, Petr (advisor) ; Doleželová, Eva (referee)
Bc. Veronika Krejčíková The effect of GLP-1 (glucagon-like peptide-1) analogues on rat hepatocytes in vitro. Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Specialist in Laboratory Methods Background: We observed the effect of liraglutide (LIRA; a GLP-1 analogue) on rat non-steatotic (ST-1) and steatotic (HF) rat hepatocytes in vitro. We also monitored its potential hepatoprotective effect on the cells damaged by acetaminophen (APAP). Methods: As a model for our experiments we used the primary cultures of rat hepatocytes isolated from the liver of male Wistar rats. For isolation of lean and fatty hepatocytes, we used rats fed by standard laboratory diet (10% energy from fat) and high fat diet (71% energy from fat) for 6 weeks. Hepatocytes were isolated by two-step collagenase perfusion. We used LIRA at concentrations of 1, 10 and 100 nmol/l on intact, ST-1 and HF hepatocytes; we also tested effect of LIRA on hepatocytes after exposure to 2.5; 3.75 and 5 mM APAP. After 24/20-hours incubation, we tested the activity of lactatdehydrogenase (LDH) in culture medium, the viability of hepatocytes, production of albumin, and level of MDA and ROS production. Morphological evaluation of cells was performed by using phase microscopy. Results: LIRA at tested concentrations...
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Western blot analysis of iNOS and HO-1 protein expression in aorta of mice with high soluble endoglin levels.
Tysoňová, Kristýna ; Rathouská, Jana (advisor) ; Doleželová, Eva (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Western blot analysis of iNOS and HO-1 protein expression in aorta of mice with high soluble endoglin levels Author of diploma thesis: Kristýna Tysoňová Supervisor: PharmDr. Jana Rathouská, Ph.D. Background: The aim of this thesis was to evaluate differences in the expression of iNOS and HO-1 (markers of nitration/oxidative stress) in the aorta of two groups of genetically modified mice on cholesterol diet that differed in levels of human soluble endoglin. Methods: For the study we used 6 months old female transgenic mice on a CBAxC57BL/6J strain backgroud with high level of human soluble endoglin. Using the ELISA analysis, soluble endoglin levels was determined and mice were divided into two groups according to the levels. The study group (Sol-Eng+ ) showed high levels of human soluble endoglin and their sibling mice with low (undetectable) plasma levels of the human endoglin were used as control group. Both groups were fed a special high fat and cholesterol diet for a period of 3 months. Biochemical analysis revealed total cholesterol levels. For detection and quantification of iNOS and HO-1 Western blot analysis was used. Results: After three months on cholesterol diet, the Sol-Eng+...
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C-reactive protein effects on aortic endothelium in spontaneously hypertensive rats
Kosán, Aleš ; Němečková, Ivana (advisor) ; Doleželová, Eva (referee)
C-REACTIVE PROTEIN EFFECTS ON AORTIC ENDOTHELIUM IN SPONTANEOUSLY HYPERTENSIVE RATS Diploma thesis Author: Aleš Kosán Study program: Pharmacy Charles University in Prague Faculty of Pharmacy in Hradec Králové Endothelial dysfunction and chronic inflammation are factors figuring in pathogenesis of metabolic syndrome. This paper is outlined as a pilot study, which represents basis for further research in the question of C-reactive protein influence on endothelium. The expression of four proteins previously associated with the development of endothelial dysfunction, was studied using the Western blot method: endothelin-1, heme oxygenase-1, endoglin and phosphorylated nitric oxide synthase. Analysis was carried out on the samples of homogenized rat aortas of transgenic spontaneously hypertensive rats with expression of human C-reactive protein. A standard spontaneously hypertensive rat strain was used as a control group. The detection was made by chemiluminescence substrate on the X-ray films and data were semiquantitatively evaluated by densitometry. We found increased expression in endoglin levels, valued at 186.3 % contrary to the control group. The expression of phosphorylated nitric oxide synthase was 325.6 % in comparison with the control group. The increase of expression was minimal (109.0 %) in...
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Role of Gq-coupled receptors in brown adipocytes
Čajková, Michaela ; Kovařík, Miroslav (advisor) ; Doleželová, Eva (referee)
Charles university in Prague, Pharmaceutical faculty in Hradci Králové, Department of biological and medical sciences Rheinische Friedrich-Wilhelms-University Bonn, Institute of Pharmacology and Toxicology Candidate: Michaela Čajková Supervisor: PharmDr. Miroslav Kovařík, Ph.D. Consultant: Dr. Linda Sarah Hoffmann Title of diploma thesis: Role of Gq-coupled receptors in brown adipocytes In my diploma thesis, we focused on four Gq-coupled receptors (F2R, LPHN1, α1DAR, TSHR) in brown adipocytes (BAs), which were identified in the screen as the highest expressed in immature and mature BAs. Our goal was to validate suggestion, that Thyroid stimulating hormone receptor (TSHR) plays a key role in differentiation of BAs and that F2R, LPHN1, α1D-AR might be important for BAs. In our study, we investigated gene expression of these four receptors in BAs, using analytical methodsquantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and Western blot. Results from analysis revealed, that expression of TSHR was increased in mature BAs, it means, that TSHR induce differentiation of BAs. The BAs transduced with short hairpin RNA (sh-RNA) against TSHR were less differentiated, this we proved also with Oil Red-O staining. Expression of adipocyte Protein 2 (aP2), peroxisome proliferator-activated...
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Alteration of transport proteins expression during intrahepatic cholestasis in rats
Kolouch, Jan ; Doleželová, Eva (advisor) ; Cermanová, Jolana (referee)
Jan Kolouch Alteration of transport proteins expression during intrahepatic cholestasis in rats Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Pharmacy Background: The aim of the diploma thesis was the verification of cholestatic liver injury by biochemical analysis of plasma and analysis of the main efflux and uptake hepatic transporters (Ntcp, Oatp1a4, Bsep, Mrp2, Mrp3, Mrp4) at the mRNA level. Methods: Wistar rats (n = 6, in each group; 280 - 320 g) were divided into two groups: Control group (LPS-K) and LPS group (lipopolysaccharide was administred at once i.p., 4mg/kg). Blood and liver samples were collected 12 hours after LPS administration. Biochemical analysis of plasma was performed by Cobas Integra ® 800 and GC/MS. Changes of mRNA expression of the transporters were evaluated by qRT-PCR. Results: In comparison to control group, LPS group showed significantly elevated plasmatic levels of the bile acids and also elevated activity of ALP and GMT. Plasmatic levels of bile acids were elevated to 818%, activity of ALP was elevated to 311% and activity of GMT to 2167%. Intrahepatic cholestasis led to the significant decrease of mRNA levels - Ntcp to 4.6%, Oatp1a4 to 2.2%, Mrp2 to 1.7% and Bsep to 25%. Conclusion: According to the results of this study, it is...
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