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Conditions of prion propagation in cell cultures
Hobzová, Kristýna ; Janoušková, Olga (advisor) ; Rusina, Robert (referee)
Prion diseases are fatal neurodegenerative diseases that affect mammals, including humans, which are characterized by accumulation of pathologi- cal prion protein isoform (PrPTSE ) in the brain. The animals were commonly used for the prion disease research in the past but in recent years, the tissue cultures are being used as well. Tissue cultures have many advantages com- pared with animals. E.g. the possibility of a detailed study of the biochemical processes associated with prion diseases, and rapid and sensitive PrPTSE de- tecting method. However no reliable in vitro model was developed for human prion diseases so far. We focused on monitoring of transmission and propagation efficiency of different prion strains and on the influence of cultivation conditions on the transfer of the neuronal cell line CAD5, which is highly sensitive to prion infection. We confirmed the sensitivity of CAD5 cells to mouse-adapted scra- pie prion strains and we presented new facts about their ability to propagate mouse adapted prions of human strains and bovine spongiform encepha- lopathy. We have used CAD5 cell sensitivity to be infected with different prion strains in other parts of this work. In the second part, we focused on the cell sensitivity to prion infection and propagation of prion strains under different culture...
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Prion Diseases - a role of transition metal ions
Malová, Zuzana ; Lukeš, Ivan (advisor) ; Moško, Tibor (referee)
Prion diseases are widely spread diseases among mammalians. They are characterized by a change of prion protein structure PrPC (cellular, native state) to PrPSc (scrapie, structure typical for prion diseases). It is a change at secondary structure of protein from α-helical to β-sheet structure, which aggregates in brain. One possibility, why it happens is higher transitional metal ions concentration (Zn(II), Cu(II), Fe(III)) in the brain, which bind to PrPC and affects its structure. Aim of this thesis is a synthesis one of protentional drugs based on 2-aminothiazole and its modification to bind fluorescent or radiochemical markers and in future study of its possible interaction with transition metal ions. Key words: complexes; macrocyclic ligands; copper; prion diseases
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Conditions of prion propagation in cell cultures
Hobzová, Kristýna ; Janoušková, Olga (advisor) ; Rusina, Robert (referee)
Prion diseases are fatal neurodegenerative diseases that affect mammals, including humans, which are characterized by accumulation of pathologi- cal prion protein isoform (PrPTSE ) in the brain. The animals were commonly used for the prion disease research in the past but in recent years, the tissue cultures are being used as well. Tissue cultures have many advantages com- pared with animals. E.g. the possibility of a detailed study of the biochemical processes associated with prion diseases, and rapid and sensitive PrPTSE de- tecting method. However no reliable in vitro model was developed for human prion diseases so far. We focused on monitoring of transmission and propagation efficiency of different prion strains and on the influence of cultivation conditions on the transfer of the neuronal cell line CAD5, which is highly sensitive to prion infection. We confirmed the sensitivity of CAD5 cells to mouse-adapted scra- pie prion strains and we presented new facts about their ability to propagate mouse adapted prions of human strains and bovine spongiform encepha- lopathy. We have used CAD5 cell sensitivity to be infected with different prion strains in other parts of this work. In the second part, we focused on the cell sensitivity to prion infection and propagation of prion strains under different culture...
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Prionové proteiny a jejich interakce s těžkými kovy
Sedláčková, Eliška
Prion proteins are infectious glycoproteins that do not contain nucleic acids. Their specific function in the brain is not yet fully understood. Conversion of the cellular form of prion protein (PrPC) into the pathological isoform (PrPSc) is regarded as the cause of neurodegenerative diseases. PrPC is a glycoprotein that interacts with many divalent metal ions, particularly Cu2+ and Zn2+. Another protein having the ability to bind metals, is also metallothionein (MT), which consists of several specific forms. Regarding formation of neurodegenerative diseases is significant metallothionein isoform MT-3 occuring in the brain. One of the MT-3 functions in the brain is its participation in maintaining of the optimal concentration of metal ions. The aim of this study was to verify the presence of expression PrPC or MT-3 genes in E. coli bacterial cells. Furthermore, we investigated the effect of metal additions on E. coli bacterial culture expressing PrPC or MT-3 protein. In the end we focused on determining the total concentration of MT in E. coli bacterial cells expressing the PrPC or MT-3 protein. Any determination was compared to a standard E. coli BL21 (DE3) strain.
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Detekce prionových proteinů a jejich interakce s kovy a metalothioneinem
Cardová, Alžběta
Prion diseases are formed by a conformational change of prion-like protein (PrPC) with alfa-helix structure to the pathological isoform - prion (PrPSc) which acquires beta-sheet structure. PrPC physiological properties in the brain are insufficiently described but there is an assumption of its affinity to metal ions. Another protein with metal-binding ability is metallothionein (MT). Brain specific isoform of MT is called MT-III and it is assumed to participate in maintenance of metal ions concentration in the brain. Aim of this study was to prepare recombinant human PrPC in E. coli. Furthermore, this protein was used to detect interactions between metal ions (Cu, Zn), MT and PrPC by differential pulse voltammetry method. The final part was devoted to the MT-III determination in different genotypes of prion-infected and non-infectious mouse brain tissues.
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