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Study of the cytostatic effects of sulfur mustard
Krejčová, Martina ; Pourová, Jana (advisor) ; Vokřál, Ivan (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Martina Krejčová Supervisor: doc. PharmDr. Jana Pourová, Ph.D. Consultant: Mgr. Petr Jošt. Ph.D. Title of diploma thesis: Study of the cytostatic effects of sulfur mustard Sulfur mustard (HD) belongs to blistering agents used in chemical warfare. It is a bifunctional alkylating agent that covalently modifies DNA. The cytostatic effect of HD is characterized by halting cell division without cell death. The duration for which a cell can remain in this state depends on the extent of DNA damage, which can subsequently be converted into a cytotoxic effect leading to cell death. The aim of the study was to determine the concentration range of HD gas that exhibits cytostatic effects and to investigate the sensitivity of HaCaT skin keratinocyte cells synchronized in the G1, S, and G2/M phases of the cell cycle to alkylating damage induction following release from cell cycle blockage. The cytostatic effect was observed only at very low concentrations, in the range of units of µmol.l-1 . In our experiments, a concentration of 2 µmol.l-1 of HD prevented cell proliferation for 2 days. Lower, submicromolar concentrations, had a stimulatory effect on cell proliferation, while higher concentrations of HD...
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Context-dependent roles of cyclin-dependent kinase inhibitors (CDKi) in cancer
Doležalová, Šárka ; Janoštiak, Radoslav (advisor) ; Macůrek, Libor (referee)
Cell cycle progression is intricately regulated by many proteins. Among these proteins, cyclin- dependent kinase inhibitors (CKIs) play a key role, traditionally associated with cyclin- dependent kinase (CDK) inhibition and cell cycle arrest. However, recent research reveals that their function is much more complex in the context of cancer. CKI inhibitors are not mutated at all in many tumor types, and their expression is maintained or even increased, suggesting additional mechanisms by which they influence tumor pathogenesis. Post-translational modifications, especially phosphorylation, which affect their localization and stability, have a fundamental influence on their function. These modifications can then, for example, affect cell motility, apoptosis, or DNA repair. This can convert CKIs from tumor suppressors to tumor promoters or vice versa, strengthening their tumor suppressor properties. Key words: Cyclin-dependent kinases, inhibitors of cyclin-dependent kinases, cell cycle, cancer, post-translational modification
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Emergent properties of the G1/S network
Dražková, Jana ; Tomášek, Petr (referee) ; Palumbo,, Pasquale (advisor)
Tato práce se zabývá buněčným cyklem kvasinky Saccgaromyces cerevisiae. Oblastí našeho zájmu je přechod mezi G1 a S fází, kde je naším cílem identifikovat velikosti buňky v době počátku DNA replikace. Nejprve se věnujeme nedávno publikovanému matematickému modelu, který popisuje mechanismy vedoucí k S fázi. Práce poskytuje detailní popis tohoto modelu, stejně jako časový průběh některých důležitých proteinů či jejich sloučenin. Dále se zabýváme pravděpodobnostním modelem aktivace replikačních počátků DNA. Nově uvažujeme vliv šíření DNA replikace mezi sousedícími počátky a analyzujeme jeho důsledky. Poskytujeme také senzitivní analýzu kritické velikosti buňky vzhledem ke konstantám popisujícím dynamiku reakcí v modelu G1/S přechodu.
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Coherence-controlled holographic microscope in cell's life cycle research
Bartoníček, Jan ; Chmelík, Radim (referee) ; Uhlířová, Hana (advisor)
The subject of the bachelor thesis is live-cell imaging in a transmitted-light holographic microscope which was designed at the Institute of Physical Engineering BUT and comparing this imaging method with the phase-contrast microscopy. The first part is dedicated to a basic description of used imaging techniques and a cell biology. A description of an experiment preparation follows. In the part dedicated to a data analysis the method of dynamic phase differences is described and the method of growth monitoring is proposed. Both methods were used for the analysis of experiments which are described in the last part of this work. Experiments were focused on acquiring time-lapse data of a cell’s cycle and particularly the mitosis.
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Modelling of Cell Colony Dynamics
Bělehrádek, Stanislav ; Škutková, Helena (referee) ; Sedlář, Karel (advisor)
The content of the thesis is a description of intracellular processes responsible for cell cycle regulation and reactions of cells to external and internal stimuli. Thoroughly described are important signaling pathways with appropriate methods, which can be used to simulate them in silico. From these cellular processes, a cell cycle model is created and implemented in a tool programmed in C ++ with OpenGL used for visualization. The model is then tested for various cell processes including HeLa cells growth. Finally, the results are compared with the behavior of living cells.
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Cell cycle regulation and genome integrity protection in the early mammalian embryos
Knoblochová, Lucie ; Drutovič, David (advisor) ; Carr, Antony M. (referee) ; Fulková, Helena (referee)
(English version) Infertility is a major health problem, as it affects one in every six people worldwide (Njagi et al., 2023). One of the major reasons for infertility are aneuploidies, additions or losses of an entire or partial chromosome during cell division. Aneuploidies thus negatively influence cellular processes and potentially lead to developmental problems or embryo loss. It has been thought for a long time that aneuploidies arise mostly during oocyte development, and these mechanisms have been well studied. However, recent evidence has shown that aneuploidies arise also de novo after fertilisation and during the early embryonic development; but the molecular mechanisms of these abnormalities still remains elusive. Aneuploidies often originate during cell cycle division from unrepaired DNA damage in mitosis. DNA damage is sensed by DNA damage response (DDR) signalling pathways, which slow down or arrest cell cycle progression until it is resolved. An essential DDR factor during typical cell cycle progression is checkpoint kinase 1 (CHK1). However, the role of DDR factors in the early embryos, and especially CHK1, have not been well studied. Early embryonic development is regulated by maternal factors stored in the oocyte until the transcription of the embryonic genome begins. To study...
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The effect of synthetic modified mRNAs induced proliferation on pancreatic beta cells
Veľasová, Adriana ; Koblas, Tomáš (advisor) ; Bořek Dohalská, Lucie (referee)
Diabetes mellitus is a chronic disease caused by the loss of pancreatic beta cells due to autoimmune destruction or increased apoptosis. Beta-cell deficiency results in reduced insulin production, which plays an important role in glucose metabolism. The number of beta-cells in the body is one of the main factors that influence the development of this chronic disease. Therefore, it is necessary to find a way by which the number of beta-cells of the organism can be increased and thus the insulin production can be restored in a natural way without any need for the use of insulin infusions. However, the ability of beta-cells to divide decreases with age and is virtually nil in adulthood. The study of the cell cycle, especially the early and late cyclins and cyclin-dependent kinases, which act as cell cycle regulators, thus appears to be a promising way to restore natural insulin-producing tissues. In order to increase the number of beta cells entering the cell cycle, we focused on studying the effect of in vitro transcribed (IVT) mRNAs, encoding cyclins type D and cyclin dependent kinases 4 and 6 on stimulating cell division of isolated beta-cells. We found that transfection IVT mRNAs for type D cyclins in combination with cyclin-dependent kinases 4 and 6 significantly increased the proliferation of beta-cells...
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Coordination of growth and cell cycle progression in green algae
IVANOV, Ivan
Within the past century microalgae have gained importance both as model organisms in cell cycle research and as a biotechnological platform for the production of a variety of economically important compounds. This thesis examines the coordination of growth and cell cycle progression in green algae and attempts to explore the biotechnological relevance of some of the findings. Furthermore, the applicability of confocal Raman microscopy for both quantitative and qualitative analysis of storage biomolecules during the course of the cell cycle of Desmodesmus quadricauda is also investigated. Temperature and light shift experiments showed that there is no direct correlation between growth and cell cycle progression in D. quadricauda. Further analysis revealed that supraoptimal temperature has a profound effect on the cell cycle of Chlamydomonas reinhardtii causing a block in cell division, increase of cell size and over accumulation of starch. Starch production through supraoptimal temperature was successfully demonstrated in pilot scale experiments, however it was estimated that light availability within the culture poses a major limiting factor. Confocal Raman microscopy was successfully applied for the quantitative and qualitative analysis of storage biomolecules including starch, lipids, polyphosphates and guanine.
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The role of truncated PPM1D/Wip1 phosphatase in cancer
Martiníková, Andra-Stefania ; Macůrek, Libor (advisor) ; Souček, Pavel (referee) ; Mistrík, Martin (referee)
When encountering damage, the cells activate the DNA Damage Response (DDR) pathway and stop the cell cycle until the DNA is repaired. PPM1D/WIP1 phosphatase resumes the cell cycle after the damage has been repaired, by directly dephosphorylating DNA damage markers. The DDR pathway prevents genome instability or cancer development. Mutations in the Ppm1d gene encoding PPM1D result in an overstable and truncated protein observed both in cancer patients and in cancer cell lines. In this thesis, we used an "in-house" transgenic mouse model in which mutations in the exon 6 of the Ppm1d gene resulted in a truncated PPM1D protein. First, we observed high PPM1D levels and impaired DDR to gamma ionizing radiation (IR) in the mouse thymi having truncated PPM1D (Ppm1dT/+ ). We then bred the Ppm1dT/+ mice with the Trp53+/- heterozygote knock-out mice which are prone to thymic lymphoma. The Ppm1dT/+ Trp53+/- double-mutants had a higher frequency of developing IR-induced T-cell lymphomas, compared to the single Trp53+/- mutants. Moreover, truncated PPM1D leads to a defective cell cycle checkpoint activation in human non-transformed RPE cells (RPE1), which then proliferate despite the presence of DNA damage. RPE1 cells also display increased proliferation after replication stress. RPE1 or U2OS cells with...
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