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Ames test in the drug development
Klaučová, Martina ; Pávek, Petr (advisor) ; Konečná, Klára (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Martina Klaučová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Consultant: PharmDr. Ivona Pávková, Ph.D. Diploma thesis title: Ames test in the drug development Background: Thesis objective is the determination of potential genotoxicity of newly developed drugs within primary testing and the introduction of the Ames microfluctuation test which can be used in common laboratory conditions. Methods: I used commercially supplied kit based on the principles of Ames test which detects reverse mutation through colour changes of the samples using bacterial strains S. typhimurium. At first I had to study literary sources and then I could design the procedures of the Ames microfluctuation test, preparation of the chemicals and storage of the strains which are optimal for all laboratories. Results: The drug samples T6445 and T6447 with 30 µM concentration tested by metabolic activation S9 on bacterial strain ST TA 98 show genotoxicity. The sample UOCHB1 with 30 µM concentration tested without activation shows possible genotoxicity on both strains ST TA 98 and ST TA 100. Other samples do not show any toxicity. I used 3 different procedures during the designation of assay. The most suitable version of the...
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Evaluation of activity of potentional antibiotic substances through the use of microdilution broth method II
Kopečná, Klára ; Konečná, Klára (advisor) ; Jílek, Petr (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of biological and medical sciences Study program: Pharmacy Autor: Klára Kopečná Supervisor: Mgr. Klára Konečná, Ph.D. Title of diploma thesis: Evaluation of activity of potencional antibiotic substances through the use of microdilution broth method II Background: To evaluate the antibacterial activity of the compounds synthesized by the Department of Pharmaceutical Chemistry and Drug control, Faculty of Pharmacy in Hradec Králové, Charles University in Prague under the leadership of prof. PharmDr. Martin Doležal, Ph.D. Methods: Using broth microdilution method was tested antibacterial activity at eight bacterial strains, namely Staphylococcus aureus, Staphylococcus aureus methicilin resistant, Staphylococcus epidermidis, Enterococcus sp., Escherichia coli, Klebsiella pneumoniae, Klebsiella pneumoniae ESBL positive and Pseudomonas aeruginosa. Results: Test substances were divided into 5 groups according to similarities in the chemical structure. The best antibacterial activity was observed in the group of derivatives of N-benzyl- 6-(alkylamino)pyrazine-2-carboxamide. Conclusion: Antibacterial activity was detected in 10 of 24 tested compounds. From active substances, all but one danced only Gram-positive bacteria. Among...
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Identification of DNA sequence binding C/EBP and C/EBPb proteins involved in RANKL expression
Dušek, Lukáš ; Křivčíková, Lucie (advisor) ; Konečná, Klára (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Author: Lukáš Dušek Supervisor: Ing. Lucie Křivčíková, Asist. Vid Mlakar, B. Sc. microbial., Ph.D. Thesis title: Identification of DNA sequence binding C/EBP and C/EBPb proteins involved in RANKL expression Bone thinning - osteoporosis - is an increasing problem of human health worldwide. Johnell and Kanis estimated that in 2000 more than 9.0 million of osteoporotic fractures occurred - this means about 3 fractures from osteoporosis every second [1]. Because the decrease in bone mineral density (BMD) is associated with age and with the aging of the world population, the issue of bone health and especially osteoporosis is expected to emerge in its urgency. Discovery of osteoprotegerin (OPG) / receptor activator of nuclear factor kappa-B (RANK) / RANK-ligand (RANKL) pathway in late 90' gave scientists a fruitful target for future investigations about bone metabolism and revealed interesting interconnections with cardiovascular diseases, immunity and cancer survival and targeting. This study had one experimental and two theoretical goals: The experimental and primary goal was to identify transcription factors and DNA binding sites in -662 to -438 region upstream of RANKL transcription start...
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Determination the mechanism of entry F. tularensis into B lymphocytes
Hadámková, Barbora ; Konečná, Klára (advisor) ; Janďourek, Ondřej (referee)
Barbora Hadámková Determination the mechanism of entry F. tularensis into B lymphocytes Diploma thesis Charles University, Faculty Of Pharmacy in Hradec Králové Study program: Pharmacy Background: Besides processing the research with basics knowledge of the problem, the main aim of the study was the analysis of mechanism of entrance of intracellular bacteria Francisella tularensis into B cells. Methods: The B cells, which we obtained through peritoneal lavage from mice Balb/c, we blocked using antibodies individual complement receptors, B cell receptor and Fcƴ receptor. The population of the cells was infected by bacteria F. tularensis LVS/GFP opsonized by complement and/or by antibodies. Using flow cytometry we measured the percentage of infection of individual subpopulations of B cell B1a, B1b and B2 and we evaluated the influence of blocking and opsonization on the infection. Results: From the measured data, we can say that the percentage of infected B cells after infection by F. tularensis opsonized by complement is increased. This increase was more distinct in subtype of B cells B1b and B2. On the other hand, the opsonization F. tularensis by antibodies did not affect the infection. We also found out, that blocking of Fcƴ receptor has decrease the infection, if we used for infection of B cells...
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Susceptibility profile of biofilms of non-albicans Candida spp. to echinocandins
Pešková, Pavla ; Křivčíková, Lucie (advisor) ; Konečná, Klára (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Kralové Department of Biological and Medical Sciences Candidate: Pavla Pešková Supervisor: Ing. Lucie Křivčíková Diploma Thesis: Susceptibility profile of biofilms of non-albicans Candida spp. to echinocandins Yeasts of the genus Candida are one of the most frequent human fungal pathogens. Infections caused by them are related to a specific form of growth - biofilm (BF), which has increased their resistance to antifungal treatment. Since bloodstream infections caused by non-albicans Candida species are increasing, it is important to focus on their susceptibility characteristics. The main aim of our experiment was to examine the susceptibility profiles of BF produced by rare non-albicans Candida species to echinocandins. We tested 3 species of the genus Candida - C. lusitaniae, C. guilliermondii and C. krusei and 3 different echinocandins - anidulafungin (AND), caspofungin (CAS) and micafungin (MFG). Echinocandins have unique mechanisms of action. They inhibit the function of the enzyme β-1,3-glucan synthase. Disruption of its function leads to inhibition of β-1,3-glucan production, damage of fungal cell wall and loss of viability of the cell. In experimental part we used YNB medium and RPMI 1640 medium to grow Candida species BF and...
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Mycotoxins and human health
Roudná, Iva ; Konečná, Klára (advisor) ; Jílek, Petr (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Roudná Iva Supervisior: Mgr. Konečná Klára, Ph.D. Adviser: RNDr. Klaban Vladimír Title of the bachelor thesis: Mycotoxins and human health This bachelor thesis deals with all aspects of the mycotoxin biosynthesis and with the negative effect of mycotoxins not only on the human organism but also on domestic and farm animals from the point of view of animal husbandry. In the first part of the thesis mycotoxins such as contaminant substances in the human environment are introduced and defined and possible misuse of mycotoxins in terrorist attacks is also mentioned. This is followed by a general description of micromycetes and their significance not only as producers of dangerous mycotoxins, but also the positive use of micromycetes in food production, modern biotechnologies, in biological plant protection as well as in the sewage treatment are also discussed. A great attention is also given to the pathological effects of micromycetes because they can cause various different mycotic and allergic disorders. The following chapter deals with the sources of mycotoxins. The thesis particularly deals with the three most important genera of mycotoxin-producing moulds: Aspergillus,...
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Expression of SWC7 molecules on the pig B lymphocytes.
Fikrová, Petra ; Jílek, Petr (advisor) ; Konečná, Klára (referee)
The aim of this graduation thesis is to clarify the expression profile of the SWC7 molecule on B cells in swine and eventually to resolve its function during maturation. At present it is possible to define the maturation pathways of porcine B lymphocytes in the periphery on the base of the expression CD2 and CD21 cell surface molecules. The flow cytometry was chosen as a central experimental method for detection of B lymphocytes carrying SWC7 molecule on their surface. The ability of individual B cell subpopulation to produce antibodies was also measured by ELISA. Principal phenotype characterization showed that the SWC7 molecule is found only on the surface of IgM+ and CD21+ B cells. These may be CD2 positive or negative. Based on this finding, the individual subpopulations of IgM+ CD21+ B cells differing in expression of SWC7 and CD2 molecules were purified by flow cytometry. Subpopulations were thereafter cultivated and subsequently analyzed for alterations in their phenotype. Media from cultivated cells were also analyzed for presence of porcine IgM, IgG and IgA antibodies by ELISA. Results indicate that the expression of SWC7 molecule on the surface of B cells may be considered as a maturation marker because loss of SWC7 molecules from the cell surface is irreversible and because only SWC7- B cells are...
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Allergies, medications and their complications, in this context,
Březinová, Lenka ; Jílek, Petr (advisor) ; Konečná, Klára (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Lenka Březinová Supervisor: PharmDr. Petr Jílek, CSc. Title of bachelor thesis: Allergies, their complications and drugs in this context, early 21st century An allergy is one of the main health problems in major modern society. Although allergy is known for centuries, dramatic growth of incidence has been noticed in last decades. Discovery of immunoglobulin E as a carrier of direct hypersensitivity was an important progress in allergology. Allergic reactions are divided into six classes, which can diffuse each other. Clinical symptoms are predestined by mediators, which come from different cells. The best known is histamine, then leukotrienes, eicosanoids etc. Allergy can be complicated by rhinoconjuctivitis, chronic sinusitis, nasal polyps, middle ear inflammation, atopic eczema, and asthma or life threatening anaphylactic shock. Allergic symptoms are inhibited by antihistamines. There are two generation of these basic drugs. The next option to treat the allergy is the immunotherapy, which is the only one who has potential to avert or change process of allergic disease. However this treatment is very expensive. Allergy is worldwide epidemic with fast growing occurrence in...
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Current methods in diagnostics, treatment and prevention of infection caused by methicilin resistant Staphylococcus aureus (MRSA)
Korejsová, Erika ; Konečná, Klára (advisor) ; Vejsová, Marcela (referee)
Candidate: Erika Korejsová Supervisor: Mgr. Klára Konečná, Ph.D. Consultant: MUDr. Marie Smolíková Title: Current methods in diagnostics, treatment and prevention of infection caused by methicilin resistant Staphylococcus aureus (MRSA) Bachelor thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Study field: Medical Bioanalytics Staphylococcus aureus is a bacterium that is found in the nasal mucosa of healthy people (20-40%) and skin without causing disease. Under certain circumstances, such as damage or injury of skin, mucous membranes and in immuno-incompetent individuals, the MRSA infection can originate. Therefore, hospitalized patients are much more susceptible to this infection. The biggest dangers represent MRSA (methicillin-resistant Staphylococcus aureus) strains. These strains of Staphylococcus aureus are mostly multi-resistant, where in addition to resistance to oxacillin (methicillin), resistence to other antibiotik occurs as well. Treatment of these patients is problematic, because MRSA strains are extensively resistant and persist in the organism and in the environment. Their eradication is not simple. The patient's blood reinfection often occurs due to low levels of antibiotics. It means a considerable financial...
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