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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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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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Detection of inflammation in heart in a mouse model of atherosclerosis
Dlouhá, Michaela ; Němečková, Ivana (advisor) ; Kovařík, Miroslav (referee)
Detection of heart inflammation in a mouse model of atherosclerosis Diploma thesis Author: Michaela Dlouhá Study program: Pharmacy Charles University in Prague Faculty of Pharmacy in Hradec Králové Background: Atherosclerosis is an inflammatory disease caused by deposition of blood lipids in the wall of arteries involving a wide range of inflammatory substances. Recent evidences suggest, that inflammation plays an irreplaceable role in the development and subsequent complications of this disease. Raising proinflammatory markers increase worsening of prognosis in patients with acute coronary syndrome, indepently of myocardial damage. Therefore, the aim of this study is to compare the changes in the expression of inflammatory markers in ApoE/LDLr-/- mouse model of atherosclerosis. Methods: By the Western blot method we monitored the expression of VCAM-1, ICAM-1 and P-selectin in the myocardium of 2 and 8 months old ApoE/LDLr-/- mice fed by standard diet. Our results show, that we did not detect any statistically significant differences in expression of observed markers. Results and conclusions: Based on the results we assume, that there is a deterioration of the inflammatory response in myocardial ApoE/LDLr-/- mice. However, for a more detailed description of coronary dysfunction in this model, we...
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Imunological and histological methods in Huntington's disease research
Jandurová, Patrícia ; Němečková, Ivana (advisor) ; Rathouská, Jana (referee)
Author: Patrícia Jandurová Title of thesis: Imunologické a histologické metody ve výzkumu Huntingtonovy nemoci Bachelor thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Study course: Medical laboratory technician Huntington's disease (HD) is a severe autosomal inherited neurodegenerative disorder that affects individuals regardless of gender. This disorder is caused by an expansion of CAG repeats in huntingtin gene, that is expressed in almost all tissues. Mutant protein causes a neuron degeneration in a brain of HD pacients which leads to a motor abnormalities and personality decay. The disease developes in the presence of a mutation in one of two huntingin alleles. The risk of transmission of disease for the direct descendants is 50%. This disease is very malign because of its late onset. An adequate therapy does not exist yet, but a lot of research teams focus on designig a suitable medical treatment. This work is devoted to the description of Huntington's disease and is focused on immunological and histological methods used in animal model experimental studies. Particular techniques are described and the results are supported with figures. Key words: Huntington's disease, CAG repeat, immunological and histological methods, animal model
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Study of membrane endoglin expression in cardiovascular system I
Večerková, Pavla ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Author: Večerková Pavla Supervisor: doc. PharmDr. Petr Nachtigal, Ph.D. Title of Diploma Thesis: Study of membrane endoglin expression in cardiovascular system I Background: We observed changes in the expression of endoglin in aorta and in soluble endoglin levels in blood in the apoE⁻/⁻/LDLr⁻/⁻ deficient two-month and eight-month old mice of C57BL/6J strain after administration of chow diet. Methods: We used the group of two-month and eight-month old apoE⁻/⁻/LDLr⁻/⁻ deficient mice of C57BL/6J strain, which were fed by standard chow diet. We performed immunohistochemical staining of aortic cuts of these mice and through ELISA analysis we were determined blood levels of soluble endoglin. Results: Endoglin expression was clearly visible on aortic endothelium with advanced atherosclerotic plaques. However, we did not observe significant differences in the staining pattern and intensity of the staining between two- month and eight-month old group of mice. Surprisingly, in both groups of mice we also found aortas without atherosclerotic changes. We did not observe significant changes in blood levels of soluble endoglin between two-month and eight-month old group of mice. Conclusions: The...
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Endothelial dysfunction markers and soluble endoglin
Pešlová, Lenka ; Nachtigal, Petr (advisor) ; Němečková, Ivana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Kralove Department of Biological and Medical Sciences Consultant: Doc. PharmDr. Petr Nachtigal, PhD. Student: Lenka Pešlová Title of Thesis: Endothelial dysfunction markers and soluble endoglin Background: To analyze possible changes in histological structure in myocardium of mice with low or high levels of soluble endoglin. We focused on description of general histology, observation of ligament changes and possible development of inflammatory reactions using immunohistochemical analysis. Method: In the study we used genetically modified female mice, strain CBAxC57BL/6J, with low or high levels of soluble endoglin (Sol-Eng+). Six-month- old female mice were fed for three months using a high fat rodent diet containing 1,25% of cholesterol and 40% of fat. Then their myocardia were resected. For identification of endoglin expression we used immunohistochemical analysis called En Vision-CD3-T-lymfocyty (rabbit anti mouse). In histological analysis we used hematoxylin-eosin and green trichrome staining. Results: Histological analysis did not reveal any pathological accumulation of collagen in any of animal myocardium we focused on. There were any changes in staining neither of nuclei or cytoplasm, nor any infiltration of leucocytes....
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C-reactive protein, cardiovascular diseases and endothelial dysfunction.
Bartoňková, Kristýna ; Němečková, Ivana (advisor) ; Rathouská, Jana (referee)
C-reactive protein, cardiovascular diseases and endothelial dysfunction Diploma thesis Author: Kristýna Bartoňková Study program: Pharmacy Charles University in Prague, Faculty of Pharmacy in Hradec Králové Background: Healthy endothelium significantly affects the vascular homeostasis which is damaged during the endothelial dysfunction. This dysfunction is linked to number of cardiovascular diseases. Endothelial dysfunction, together with the chronic inflammation, represents also basis of the metabolic syndrome. The aim of the thesis was to detect selected markers of endothelial dysfunction in recently published transgenic rat model of metabolic syndrome. Methods: In this thesis, following factors were evaluated: endothelial nitric oxide synthase, phosphorylated endothelial nitric oxide synthase, superoxide dismutase 3, cyclooxygenase 2, nuclear factor κB and forkhead box P3. Evaluation of their expression in the aorta homogenates was performed by method Western blot. The control group was composed of spontaneously hypertensive male rats, the evaluated group was made from transgenic spontaneously hypertensive male rats expressing human C-reactive protein. Then the detection of proteins was performed by chemiluminiscent method on X-ray films. All the data were evaluated by densitometry method....
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Effects of obesity and sleeve gastrectomy on fungiform papillae in a model of Wistar rat
Zadražilová, Jana ; Němečková, Ivana (advisor) ; Nachtigal, Petr (referee)
Jana Zadražilová Effects of obesity and sleeve gastrectomy on fungiform papillae in a model of Wistar rat Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Pharmacy Obesity has become a worldwide issue in which taste sensitivity and food preferences play crucial roles. The most effective treatment of morbidly obese patients is bariatric surgery. In our preliminary study, we tried to determine the effect of diet induced obesity and sleeve gastrectomy (SG) on the histology of rat tongue, especially how they affect the number of fungiform papillae (FP). Male Wistar rats were used in the experiment. After inducing obesity with a high-fat diet (HFD), some of the rats underwent SG, some had sham surgery and others stayed surgery-free to be considered as a control group, additionally, one group stayed on normal diet (ND) the entire duration of the experiment. After 16 weeks, all rats were sacrificed and histological examination of rat tongue was per- formed. The highest mean number of FP (5.11±1.45) was counted in diet induced obese rats that were fed a ND the last four weeks of the experiment. The lowest number of FP (3.38±1.45) was found in the experimental group of rats fed a HFD through- out the experiment, but underwent SG after 12 weeks of fattening. There was no...
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Optimalization of Leishmania antigen formulations for the diagnosis of canine leishmaniasis.
Danková, Barbora ; Němečková, Ivana (advisor) ; Vokřál, Ivan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Student: Barbora Danková Supervisor of diploma thesis: RNDr. Ivana Němečková, Ph.D. Specialized Supervisor: Prof. Anabela Cordeiro da Silva Title of diploma thesis: Optimalization of Leishmania antigen formulations for the diagnosis of canine leishmaniasis Leishmaniasis is a vector-borne disease caused by a protozoan parasite of the genus Leishmania and is prevalent in 98 countries worldwide. Reliable and accurate tests are necessary for laboratory diagnosis of Leishmania infection because of the wide spectrum of clinical characteristics and symptoms and high rate of asymptomatic infections that may occure. Serological diagnosis of canine leishmaniasis (CanL), especially enzyme-linked immunosorbent assay (ELISA), proved to be usefull tool. Data obtained from ELISA assay for the detection of antibodies against selected recombinant A, B, C, D, E and F proteins were evaluated in infected canine sera from Portugal and Brazil. This work presents the best formulation of recombinant proteins for serological diagnosis. We found that antigen C showed high level of sensitivity and specificity in recognition of positive and asymptomatic sera infected with CanL. Key words: canine leishmaniasis,...
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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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