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The study oxidative stress markers in animal model of chronic colitis in large intestine.
Hocková, Erika ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Crohn's disease and ulcerative colitis are both entities of inflammatory bowel disease (IBD). The nuclear transcription factor kappaB (NFκB) family consists of five different members, which are namely p65 (RelA), c-Rel, RelB, p50 and p52. NFκB has ability to promote the expression of various proinflammatory genes, to influence the course of mucosal inflammation and to mediate different cell-type specific effects. Bilirubin is an endogenous substance with anti-oxidative and anti-inflammatory effects, which influences the process of inflammation and has protective effect. This diploma thesis was aimed at conduction of methods of NFκB detection on paraffine tissue slices and observation of NFκB expression in large intestine walls of normobilirubin and hyperbilirubin rats (Gunn's variety) after chronic imunohistochemically- induced inflammation. Eight Gunn's variety rats (G+ ) and eight Wistar's variety rats (K+ ) were used in this study, all after long time administration of dextrane-sulfate (DSS). For seven following days, DSS (in concentration of 2%w/v) was added into drinkable water for these rats, followed by 14 days of clear drinkable water (without DSS) administered to rats. This three-weeks long cycle was repeated twice (3 cycles in total were completed). Six rats of both varieties (6 G+ and 6...
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The effect of atorvastatin on the expression of markers of the cascade transforming growth factor beta in experimental atherogenesis.
Frnochová, Lenka ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Lenka Frnochová The effect of atorvastatin on the expression of markers of the cascade transforming growth factor beta in experimental atherogenesis. Diploma thesis Charles Univerzity in Prague, Faculty of Pharmacy in Hradec Králové Pharmacy Background: The aim of this diploma thesis was to observed the expression of ALK-1 and VEGF in atherosclerotic plaques in ApoE and LDL receptor- deficient mice with regard to the use of atorvastatin. Methods: We used for research female C57BL/6J mice with double deficiency of apolipoprotein E and LDL receptor. Mice were 8 weeks old. Mice were divided into two groups: atorvastatin and control group. Both groups were fed cholesterol diet for 8 weeks and in atorvastatin group was also fed with atorvastatin. Then biochemical analysis of blood was performed. For detection of lipids in atherosclerotic lesions we used histological staining by oil red. Immunohistochemical analysis was performed in 1cm aortic sinus and part of aortic arch. The method used to detect the expression of ALK-1 and VEGF was the avidin-biotin method with detection using DAB. Results: Biochemical analysis showed that atorvastatin reduced levels of total cholesterol. After the staining by oil red atherosclerotic lesions were seen in both groups. Atorvastatin reduced the size of atherosclerotic...
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Sunitinib Effects on the Expression of ICAM-1 in Normotensive and Hypertensive Rats
Frnochová, Lenka ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Mgr. Lenka Frnochová Sunitinib effects on the expression of ICAM-1 in normotensive and hypertensive rats Thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Pharmacy Background: We quantified the expression of ICAM-1 in aorta of normotensive Wistar Kyoto rats and spontaneously hypertensive rats during use of sunitinib. For quantification we applied stereological and immunohistochemical methods. Methods: We used inbred male of Wistar Kyoto rats and spontaneously hypertensive rats for our research. Rats of both strains were divided into two groups - a sunitinib group and a control group. The experimental group was receiving drinking water with sunitinib of 10 mg/kg per day and rats in the control group was receiving only drinking water. Spontaneously hypertensive rats were receiving sunitinib for 8 weeks and after 5 days of regeneration they got sunitinib into the water back for another 8 weeks. Wistar Kyoto rats were receiving sunitinib in water for 8 weeks, but after 5 days of regeneration, these rats were receiving sunitinib only for 2 weeks due to toxicity and weight loss. Immunohistochemical analysis was applied in rat aorta. For detection of the ICAM-1 expression was used En Vision method with DAB detection. Results: Immunohistochemical staining showed that expression of...
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Sunitinib effects on endothelial expression of hemeoxygenase-I in normotensive rats
Svobodová, Barbora ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Barbora Svobodová Sunitinib effects on endothelial expression of hemeoxygenase-I in normotensive rats. Bachelor thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Medical Laboratory Technician Background: The aim of this bachelor thesis was to define and describe the expression of hemeoxygenase-1(HO-1) in the aortas of normotensive Wistar Kyoto (WKY) rats and evaluate possible effects of sunitinib treatment by means of immunohistochemical and stereological analysis. Methods: Two groups of WKY rats were used for the experiment (6 animals in each of them). Sunitinib was administered to the first group. The other one (the control one) received water only. Application of sunitinib proceeded by following scheme: 8 weeks of using/ 5 weeks rest period/ 2 weeks of using. After that we performed the immunohistochemical analysis on 60 preparations of rat's aortas which observed the expression of HO-1 by methodology En Vision with DAB detection. Then the stereological evaluation was performed. Results: The immunohistochemical analysis showed the expression of HO-1 in the endothelium only and it was noticed just in several animals especially from the group with sunitinib treatment. Conclusion: The immunohistochemical and stereological analysis demonstrated a significant increase of the...
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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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Evaluation of activity of potentional antimycotic substances through the use of microdilution broth method III
Zavřelová, Alena ; Konečná, Klára (advisor) ; Němečková, Ivana (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Title of thesis: Evalution of activity of potentional antimycotic substances through the use of microdilution broth method III Autor of thesis: Alena Zavřelová Supervisor: Mgr. Klára Konečná, Ph.D. Background: The aim of this thesis, due to the growing incidence of fungal diseases and the growing incidence of resistance, was to determine the activity of potentially antimycotic active substances. We tested newly synthesized compoused, structurally based on the substance of pyrazinamide which is significant antituberculotic. Methods: Testing of substances was carried out using a broth microdilution method on eight strains of pathogenic fungi consisting of five yeast strains: Candida albicans, Candida tropicalis, Candida krusei, Candida glabrata, Trichosporon asahii and three strains of filamentous fungi: Aspergillus fumigatus, Absidia corymbifera, Trichophyton mentagrophytes. Results: From the range of tested concentrations and from the total number of the thirty tested substances, the most noticeable antifungal aktivity was revealed in the substance of 5-methyl-6-(2-phenylhydrazinyl)pyrazine-2,3- dicarbonitrile. All strains, except Trichophyton mentagrophytes strain, showed a relatively...
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Optimalisation of methods for determination of CD44 receptor activation
Skalická, Nela ; Němečková, Ivana (advisor) ; Konečná, Klára (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Title of diploma thesis: Optimalisation of methods for determination of CD44 receptor activation. CD44 is a transmembrane glycoprotein present in most cell types which is involved in many physiological and pathological processes and it represents the main hyaluronic acid (HA) receptor. HA is a nonsulfated glycosaminoglycan that is present extensively in extracellular matrix, where it provides cell hydratation. HA is involved in cancer, affects cell migration, supports an angiogenesis and participates in wound healing. In this study, we focused on the optimization of methods for the evaluation of activation, the amount and cleavage of CD44 receptor in HT-29 cell line. We observed the total amount of the receptor and its cleaved domains using methods Western Blot, ELISA and flow cytometry. We used HA molecular weight 71 kDa and PMA (Phorbol- 12-Myristate-13-Acetate) for activation. Time-dependent changes appeared in the amount of total CD44 and its fragments after activation. We also observed cell death depending on the presence of HA and PMA. We demonstrated no significant effects of these substances on cell viability. We used fluorescent and confocal microscopy for visual evaluation of...
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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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Soluble endoglin effects on endothelial dysfunction markers in mouse aorta
Kalmanová, Lenka ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Title of Diploma Thesis: Soluble endoglin effects on endothelial dysfunction markers in mouse aorta Author of Diploma Thesis: Lenka Kalmanová Supervisor of Diploma Thesis: prof. PharmDr. Petr Nachtigal, Ph.D. Background: The aim of the presented diploma thesis was to identify and subsequently evaluate the expression of endothelial dysfunction markers (ICAM-1, VCAM-1, P- selectin) in aortas of two groups of transgenic mice. The difference between control and experimental group was in the level of human soluble endoglin. Methods: 12 months old male mice of transgenic strain CBAxC57BL/6J with inserted genetic information for the expression of human soluble endoglin were utilized for the experiment. The mice were divided into two groups - the experimental group (high levels of sEng) and the control group (low levels of sEng) by ELISA analysis of levels of sEng in plasma. Both groups of mice were fed with standard laboratory rodent diet. Western blot analysis of adhesion molecules ICAM-1, VCAM-1 and P-selectin in mouse aorta and subsequently ELISA analysis of plasma levels of sVCAM-1 molecule were performed. Total cholesterol and triacylglycerol levels were demonstrated by biochemical analysis....
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