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Ecotoxicity tests on cellular level
Procházková, Petra ; Vávrová, Milada (referee) ; Zlámalová Gargošová, Helena (advisor)
Endocrine disrupting compounds among significant environmental pollutants. They are dangerous both for humans and other organisms by affect their hormonal system. They enter the environment during production, use and disposal some chemicals, but they may be of natural origin. The aim of this work is to give a brief overview of possible methods of determination of the presence of ecotoxic substances in the environment, using the in vitro toxicity assays. These tests use cell cultures instead of test organisms. The work describes principles of ecotoxicity tests at the cellular level, such as cell viability assays, cell proliferation assays, assays based on the metabolic activity of cells or DNA microarrays. A significant part of this work is focused on the possibilities of determination of endocrine disruptors by selected assays. They are ligand binding assays, reporter gene assays (recombinant yeast assay, mammalian-based reporter gene assays) and cell proliferation assay called E-screen. Significance and rationale use of tests to prove the endocrine activities in environmental matrice is given through examples of studies in which the tests were used.
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Study of interactions of biomaterials with human cell cultures.
Maslonková, Ivana ; Kostovová, Iveta (referee) ; Márová, Ivana (advisor)
The bachelor thesis was focused on introducing of techniques of cultivation of human cells in vitro and optimization of their uses in variety of cytotoxicity tests. Further, in this work were studied also interactions of cells with biomaterials and prepared organic-inorganic nanoparticles. In the theoretical part the basic conditions of cell cultivation, the characteristics of keratinocytes as cell cultures, basic characteristics of particles and also short search about the most used tests of cytotoxicity on various cells systems were described. The experimental part was focused on the optimizing the preparation of silver particles, where maltose was chosen as the most appropriate reduction agent. The average size of these prepared particles was 35 nm. The possibility of preparation of combined organic-inorganic particles including liposome particles, PHB particles, alginate particles and chitosan particles was also tested in the thesis. Later, the silver nanoparticles were encapsulated into these organic-inorganic particles as an inorganic component. The liposome particles prepared by ultrasound were evaluated as the most stable of all the other prepared particles. Human keratinocytes were chosen as the cell cultures that were needed for the observation of the particle interactions with human cells. In this work basic methods of cell cultivation and passage were introduced. Conditions of experimental work in cell culture laboratory were setting and possible application of cell cultures to cytotoxicity testing by MTT was tested.
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Functionalization of gold nanoparticles for imaging
Jakubechová, Jana ; Pekárková, Jana (referee) ; Drbohlavová, Jana (advisor)
The diploma thesis deals with synthesis of gold nanoparticles and their surface functionalization suitable for in vitro imaging. In this view there are requirements for optical properties such as stability, monodispersity and no presence of cytotocxicity. In order to fulfill these demands the synthesis by Turkevich method with surface modification by glutathione and polytethylenglycol was performed. Analytical methods such as DLS, SEM and Zeta potential measurement were utilized to characterize the physical and chemical properties of synthesized gold nanoparticles. Finally, MTT assay was performed to evaluate toxicity of gold nanoparticles using HEK 293 cell line.
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Synthesis of spherical gold nanoparticles for biomedical applications
Gablech, Evelína ; Adam, Vojtěch (referee) ; Drbohlavová, Jana (advisor)
Tato práce se zabývá syntézou sférických zlatých nanočástic pro biomedicínské aplikace. Zlaté nanočástice byly syntetizovány ekologicky nezávadnými metodami, kterým je věnována i značná část rešerše. Cílem bylo nasyntetizovat stabilní koloidní zlaté nanočástice vhodné pro různé biomedicínké aplikace zejména, pro in vivo a in vitro zobrazovací metody, kterých přehled je take obsažen v teoretické části práce. Také byl proveden test cytotoxicity, jelikož částice mají být použitelné pro in vivo aplikace. Částice byly dale charakterizovány metodami SEM, DLS a UV-VIS.
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Natural drugs in cancer treatment and their cytotoxicity
Hájková, Tereza ; Raudenská, Martina (referee) ; Provazník, Ivo (advisor)
The thesis deals with the natural substances in context with the cancer disease. The natural substances have a positive effect on the human organism and they are able to influence the viability and the growth of the cancer cells. The main mechanical device is to influence the mechanisms needed to start the apoptosis of the cancer cells and stopping further proliferation. The cancer cell lines utilization in the cancer disease is discussed in the thesis too. The thesis states common methods of determining the natural substances cytotoxicity. For the experimental part of the thesis it was chosen the MTT test method and the xCELLigence system for monitoring in real time. The mechanical device of the tested substance capsaicin in application on the prostate cell lines, tumorous PC3 and nontumorous PNT1A influence will be observed within the experimental part of the thesis.
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Insights into the mechanism of action of bactericidal lipophosphonoxins
Panova, Natalya ; Zborníková, Eva ; Šimák, Ondřej ; Krásný, Libor ; Kolář, M. ; Látal, T. ; Seydlová, G. ; Rejman, Dominik
The advantages offered by antibiotics in the treatment of infectious diseases are endangered due to the increase in the number of antibiotic-resistant bacterial strains. This reduces the efficiency of antibiotic treatments and poses a serious health and economical problem. Currently, the need for novel antibiotics is becoming increasingly apparent. Recently, we discovered a series of compounds termed lipophosphonoxins exhibiting selective cytotoxicity towards gram-positive bacterial cells. The attempt to elucidate the mode of action of lipophosphonoxins is presented here.
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Cytotoxicity and secondary metabolites of cyanobacteria
HROUZEK, Pavel
Cyanobacteria are well-known producers of secondary metabolites of different chemical structures and a wide range of biological functions. In the present thesis, the cytotoxic activity of cyanobacteria, originating from different habitats, was studied in order to reveal whether cytotoxicity is an environmentally dependent characteristic. In addition, the data were compared with the toxicity of these extracts to the model invertebrate Artemia salina. The obtained data suggest that cytotoxic cyanobacteria are favoured under some conditions and thus more frequent in particular localities. The majority of the studied extracts and fractions exhibited cytotoxitity to the Sp/2 cell line not accompanied by toxicity to A. salina. Moreover, in most of the strains with both activities to A. salina and Sp/2 cells the toxic effect was caused by an identical fraction. This result suggests that the toxic effect of the cyanobacterial secondary metabolites is mostly affecting basal cell metabolism rather than targeting specific organisms. In one of studied strains, Cylindrospermum sp. C24/1989, novel lipopeptides puwainaphycin F and G have been detected, isolated and their structure and biological effect have been characterized. Both of these structures interfere with eucaryotic membranes and cause a Ca2+ leakage into the cell. Subsequently, an enhanced tyrosine phosohorylation and relocalization of f-actin in the cell was observed. Lastly, the correlation between metabolite production and the reconstructed phylogeny was studied in planktonic Dolichospermopsis strains. Most of the detected compounds were found to be randomly disspersed across the reconstructed phylogeny and thus cannot be considered as good chemotaxonomic markers. This result also hampers the possible detection of toxic cyanobacteria by morphological methods or molecular detection based on the 16SrDNA gene.
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Cytotoxicity of bentic and periphyton cyanobacteria
SKÁCELOVÁ, Kateřina
This study is focused on assessment of cytotoxic activity of 14 strains of benthic cyanobacteria against HeLa cell line. Extracts of the strains generating strong cytotoxicity were analyzed and fractionated with HPLC-MS, in order to determine active fractions. Mechanisms of effect on the cells were investigated using immunofluorescent staining. Two out of 14 studied strains exhibited strong cytotoxic activity. Extract of Cylindrospermum sp. CY OM contains active fraction inducting apoptosis and also a fraction affecting adhesion of cells. Activity of Anabaena oscillarioides HINDAK 1984/43 strain was caused by two cytotoxic fractions, in one of them a new linear peptide was discovered.
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Cytotoxicity of the filamentous cyanobacteria in relation to its ecology
TOMEK, Petr
The aims of this study was to screen a cytotoxic activity in 53 strains of the genera Nostoc and Anabaena and to compare occurence of active isolates within different ecological groups. Selected cytotoxic strains were subjected to HPLC/MS analysis to identify their extracts composition. Isolates with strong cytotoxic effect containing unknown substances were fractionated to obtain pure fraction of this compound. Finally, the selected strains toxicity dependence on time and concentration was evaluated.
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