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Optimization of the method for viability testing in Haemonchus contortus
Mašátová, Eva ; Skálová, Lenka (advisor) ; Vokřál, Ivan (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Eva Mašátová Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Consultant: Mgr. Linh Thuy Nguyen Title of diploma thesis: Optimization of the method for viability testing in Haemonchus contortus Barber's pole worm (Haemonchus contortus) is one of the most widespread pathogenic nematodes of the Trichostrongylidae family parasitizing in the abomasum of small ruminants, mainly in sheep. The most common symptoms of haemonchosis are anaemia, lack of thriving and weight loss. Young animals can also die. A significant complication in the treatment of haemonchosis is the rapidly developing resistance of the barber's pole worm to the administered drugs, which has become a global problem in recent years. This work focused on testing the efficacy of benzimidazole anthelmintics (thiabendazole and albendazole) in three strains of barber's pole worm, which differ in their drug sensitivity. We worked with strains: ISE (Inbred Susceptible Edinburgh) strain - sensitive, IRE (Inbred Resistant Edinburgh) strain - resistant and WR (White-River) strain - multi- resistant. The egg hatch test (EHT) was used to monitor the drug effectiveness and thus to monitor the level of resistance. Eggs from individual strains were...
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The use of new ex vivo tests to find synthetic compounds with anthelmintic effect
Mičundová, Magdalena ; Skálová, Lenka (advisor) ; Szotáková, Barbora (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Magdalena Mičundová Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: The use of new ex vivo tests to find synthetic compounds with anthelmintic effect Haemonchus contortus is a parasitic nematode causing disease called haemonchosis in sheep and goats. This worldwide spread parasite feeds on the host's bloodstream, causing anemia and edema that can lead to death of the animal. Anthelmintic drugs are used for haemonchosis treatment, but recently their efficacy has decreased due to development of drug resistance in nematodes. Therefore, it is necessary to look for new drugs that will be effective in resistant nematode strains. In this work the anthelmintic effect of 13 new benzyloxyamide derivates (BLK) was studied in H. contortus eggs and adults. An egg hatch test, in which BLK8 was the most effective compound, was used first. A new method of bioluminiscence determination of adenosine triphosphate (ATP) was used in adults of H. contortus to monitor the effect of BLK analogues on viability of adult nematodes. The efficacy of BLK analogues was compared with the effect of classical anthelmintic levamisole. It is obvious from the results that several BLK analogues (especially...
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Determination of ovarian function parameters
Jará, Barbora ; Skálová, Lenka (advisor) ; Laštovičková, Lenka (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Thesis title: Determination of ovarian function parameters Supervisor: prof. RNDr. Lenka Skálová, Ph. D. Consultant: RNDr. Michala Vosátková, MBA Created by: Barbora Jará The levels of hormones that control of ovaria are important markers of development and health status of the female organism. Laboratory examination and correct interpretation of the results are important for proper diagnostic procedures. Among the hormones of ovarian function, the level of estradiol, progesterone, luteinizing and follicle-stimulating hormones is most often tested. Separation and immunochemical methods are used. For common clinical practice, immunochemical methods are particularly suitable due to their availability, relative accuracy, unpretentiousness, and speed of processing. The impossibility of standardizing individual methodological approaches represents the shortcoming of these methods. A series of examinations and interpretation of results can be performed using immunochemical methods only within one workplace. Keywords: estradiol, immunochemical methods, ECLIA, CMIA, RIA
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Optimization of a new analytical system for selected biochemical and immunochemical methods
Dvořáková, Kristýna ; Skálová, Lenka (advisor) ; Matysová, Ludmila (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Kristýna Dvořáková Thesis supervizor: prof. RNDr. Lenka Skálová, Ph.D. Consultant: Mgr. Martina Hrdinová Title of diploma thesis: Optimization of a new analytical system for selected biochemical and immunochemical methods Specific requirements for validation and verification of analytical methods according to the ČSN EN ISO 15189: 2013 standard must be met by every medical laboratory. The aim of the work was to meet the requirements of verification in the introduction of a new analytical system Cobas 8000 and also to compare the results with those obtained using older analyzer Cobas 6000 in the laboratory of clinical biochemistry. For the analysis of biochemical and immunochemical methods, we used diagnostic kits from the same supplier Roche s.r.o. We determined the selected clinic-biochemical markers by the basic methods using spectrophotometry, immunoturbidimetry and electrochemiluminiscence. We used certified calibration and control material in the process of method verification. The mixed patient serum was used for the evaluation of repeatability and comparability of the methods. We evaluated the verification and repeatability of methods using Microsoft Excel software. We compared the...
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The effect of thujone and piperitone on the expression of biotransformation enzymes in liver slices.
Štětková, Lenka ; Skálová, Lenka (advisor) ; Matoušková, Petra (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Lenka Štětková Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: The effect of thujone and piperitone on the expression of biotransformation enzymes in liver slices Monoterpenes are natural substances that are widely distributed in the essential oils of many plant species. Thujone and piperitone are found in certain plant species that can be part of dietary supplements, herbal preparations or used in foods as flavourings. The aim of this diploma thesis was to find out how thujone and piperitone affect the expression of selected biotransformation enzymes from the cytochrome P450 superfamily, the sulfotransferase and the UDP-glucuronosyltransferase superfamilies. Precision-cut liver slices obtained from the livers of 3 human donors were used for these experiments. Slices were incubated with thujone or piperitone for 24 hours at 10 μM and 50 μM concentrations. After incubation, the amount of mRNA and protein was quantified. The obtained results showed considerable interindividual variability. CYP1A2 gene expression was significantly increased in one donor sample after incubation with thujone (50 μM) and piperitone (10 μM, 50 μM). Piperitone (50 μM) increased the amount of mRNA of...
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The use of new ex vivo tests to find ferns with anthelmintic effect
Bednářová, Klára ; Skálová, Lenka (advisor) ; Szotáková, Barbora (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Klára Bednářová Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: The use of new ex vivo tests to find ferns with anthelmintic effect Haemonchus contortus is one of the worldwide spread nematode parasitizing in the gastrointestinal tract of small ruminants. The nematodes cause a disease called haemonchosis. A typical symptom of the disease is anemia resulting from the sucking of blood by nematodes. The disease also reduces productivity and may result in the death of the host. Only a few drugs are used for the treatment of haemonchosis, the number of newly introduced drugs is low. Nowadays, the emerging resistance to anthelmintics is a growing problem due to the high H. contortus adaptivity as well as the incorrect use of synthetic drugs. In addition, the synthetic drugs pollute the environment, which is why there is an effort to find herbal products with an anthelmintic effect. Anthelmintic activity has been demonstrated in some Asian and African ferns, so we decided to test the European fern species and evaluate their anthelmintic activity. Anthelmintic activity against nematodes, specifically H. contortus, was tested using egg hatch test and ATP bioluminescence assay in...
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Epigenetic changes induced by obesity and their heredity
Roušová, Lucie ; Skálová, Lenka (advisor) ; Boušová, Iva (referee)
Obesity, a chronic disease characterized by the deposition of excessive amounts of fat in the body, is the cause of the development of many other comorbidities. In recent years, this disease appears in ever younger people. Both genetic and environmental factors are involved in the development of obesity. The link between these two factors could be epigenetic modifications. Epigenetic changes are changes in a person's phenotype without changing their genotype. These changes, which include DNA methylation and histone modification, thus affect gene expression and function, without altering the DNA sequence. The aim of my bachelor thesis was to search out which epigenetic changes are associated with obesity and what genes are affected in this way. I focused on the changes that obesity causes, but also on the changes that cause obesity. I also tried to find information about whether these genome modifications are hereditary and how many generations are possibly affected by them. From the available information, it is clear that there are a lot of epigenetic modifications that are somehow associated with obesity. For example, the changes in proopiomelanocortin, tapsin or insulin like growth factor 2 genes have been detected. Some modifications are associated with paternal obesity (e.g. the...
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The effect of flubendazole on temozolomide treatment in glioblastoma cells
Dvořáková, Kateřina ; Skálová, Lenka (advisor) ; Pávek, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Kateřina Dvořáková Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: The effect of flubendazole on temozolomide treatment in glioblastoma cells Glioblastoma multiforme (GBM) is one of the most common and aggressive brain tumors in adults. Despite significant advances in treatment, GBM remains to have very poor prognosis and median survival of only 12 to 15 months after diagnosis. Standard treatment consists of maximal possible surgical resection followed by chemo-radiotherapy with antitumor drug temozolomide (TMZ), which use is problematic due to fast developing chemoresistance. The aim of this study was to examine the effect of potential anticancer drug, anthelmintic flubendazole (FLU) and effect of FLU in combination with TMZ on GBM cells. For this purpose, two GBM cell lines were used - A172 and T98G. In general FLU reduced cell proliferation more, especially in T98G cells. Moreover, the use of different combinations of TMZ + FLU showed even higher inhibitory effect on the viability of GBM cell lines. The combination of TMZ + FLU reduced the protein level of -tubulin and III-tubulin, simoultaneously interesting changes in STAT3 and EGFR expression, as well as lower...
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The effect of flubendazole and mebendazole on epithelial-mesenchymal transition.
Hochmalová, Martina ; Skálová, Lenka (advisor) ; Ambrož, Martin (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Martina Hochmalová Supervisor: prof. RNDr. Lenka Skálová, Ph.D. Consultant: RNDr. Věra Králová, Ph.D. Title of diploma thesis: The effect of flubendazole and mebendazole on the epithelial- mesenchymal transition. Epithelial-mesenchymal transition (EMT) is a process in which a non-motile epithelial cell switches to motile mesenchymal phenotype. This phenomenon is typical for multiple biological processes including cancer metastases. It could be induced by transforming growth factor-β (TGF-β) which is produced by cancer cells and which regulates its key transcription factors including SNAIL, ZEB and TWIST. EMT involves a series of defined events mediated by microtubules, including changes in cell shape and migration into the surrounding tissue. Based on these findings we suppose that the inhibition of tubulin polymerization could prevent the EMT process therefore cell migration and also cancer metastases. In our experiments, the effect of anthelminitic drugs flubendazole (FLU) and mebendazole (MBZ) on TGF-β-induced EMT in dysplastic oral keratinocytes was tested in vitro. Expression of molecules involved in EMT process was examined on microRNA and mRNA level using RT-PCR. The protein level of EMT...
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Tumor markers
Karmazínová, Simona ; Dršata, Jaroslav (advisor) ; Skálová, Lenka (referee)
Tumorous diseases are as old as life itself. They pose a serious social problem and everybody has encountered them at some time. Due to the polluted environment and modern way of living, tumorous diseases are becoming increasingly responsible for human deaths. Tumorous diseases can appear in any multicellular organism in some form. Traces of them have been identified in prehistoric people as well as in Egyptian mummies. The widespread word CANCER has its origin in ancient Greece. The first reports of laboratory evidence of tumorous diseases were published in scientific publications as early as the mid-19th century. This was the time of the onset of clinical biochemistry as a branch of science and medicine. Laboratory techniques did not exist then and testing was based on simple physico-chemical reactions. It was only in the second half of the 20th century that the study and examination of tumour markers enjoyed rapid development. Currently, the most common tumour can be determined by nearly any biochemical laboratory. The subject of my paper, "Tumour markers" is very broad - actually, each chapter would deserve to be an independent subject - and still, my thesis is far from a detailed analysis of each individual marker and methods of its determination. I aimed at providing a brief overview of the...
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