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Detection of epithelial-mesenchymal transition markers (EMT) in cells in vitro
Špaček, Petr ; Szotáková, Barbora (advisor) ; Trejtnar, František (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Petr Špaček Supervisor: Doc. Ing. Barbora Szotáková, Ph.D. Consultant: RNDr. Věra Králová, Ph.D. Title of diploma thesis: Detection of epithelial-mesenchymal transition (EMT) markers in cells in vitro Epithelial-mesenchymal transition (EMT) is a process during which motile mesenchymal-like cells develop from non-motile parent epithelial cells. Physiologically, EMT plays important roles during embryonic development and wound healing. Loss of control over this mechanism can lead to fibrosis and cancer progression. Motile mesenchymal-like cells can pass through the basal lamina, get into the blood vessels and spread to distant tissues. Transition is regulated by EMT biomarkers. The biomarkers comprise wide spectrum of proteins, including cell surface proteins (E-cadherin, N-cadherin), cytoskeletal proteins (vimentin), microRNA (miR 200) and transcription factors (Snail, Twist). In this study, expression of EMT biomarkers was evaluated using RT-PCR and Western blotting. The ability to migrate was assessed using real-time analysis with the x-CELLigence system. Two known triggers of EMT, the StemXVivo™ EMT Inducing Media Supplement (IS) and TGF-β, were compared in human oral cancer cell lines DOK and...
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Effects of statins on Ras/Raf/ERK signaling in A431 cell line
Smutný, Tomáš ; Pávek, Petr (advisor) ; Trejtnar, František (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxikology Candidate: Tomáš Smutný Supervisor: Doc. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Effects of statins on Ras/Raf/ERK signaling in A431 cell line The ERK pathway is the signal transduction cascade, which plays an important role in different cellular functions e.g. cellular proliferation, differentiation, and survival. Its inappropriate activation is often presented in human cancers. The pathway consists of the many proteins, but the interest concentrates especially on members of the Ras subfamily (particularly H-Ras or K-Ras). Ras proteins are GTPases, which are significantly included in the oncogenesis of many human cancers. The key role in the Ras proteins function represents their posttranslation modifications with the lipid. The covalent binding of prenyl moiety to Ras protein facilitates its attachment to the plasmatic membrane, which allows its function. The prenyl groups are formed during the synthesis of the cholesterol as its precursors. The inhibition of the prenylation results in Ras proteins inactivation of the ERK signal transduction cascade and suppression of uncontrolled cell division where the activating mutations of Ras proteins are presented. Statins hypolipidemic...
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Study of Radiolabelling DOTA-Sargastrin and Determination the Stability of Labeled Product
Korec, David ; Lázníčková, Alice (advisor) ; Trejtnar, František (referee)
Doctoral thesis David Korec Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Biophysics and Physical Chemistry In this doctoral thesis was to find optimal conditions for radiolabelling of DOTA-peptide Sargastrin with metallic radionuclides 111 In or 177 Lu in such a quality that the labeled peptides can also be used at radiopharmacie department for biological experiments. Minimum required radiochemical purity for these experiments was 97%. Modified peptide-DOTA Sargastrin can be succesfully labeled with lutetium-177 with a radiochemical purity greater than 99% at pH 4.52, and heating at 80řC for 20 minutes. In the same way was DOTA- Sargastrin labeled with indium-111 with a maximum achieved radiochemical purity of 98.7%. It was found that a peptide labeled with lutetium showed slightly higher lipophilicity as its elution time was about 20 seconds longer than the elution time of the peptide labeled with indium. It is probably due to the different coordination prperties of both metals. In studying the stability it was shown that the radiochemical purity of 177 Lu-DOTA- Sargastrin remained greater than 99% up to 48 hours when stored in the fridge. The purity of 111 In-DOTA-Sargastrin withstand in the specified limit purity approximately 1.5hours after preparation. The...
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Study on Renal Accumulation of Receptor-specific Radiopeptides
Kubačková, Tereza ; Trejtnar, František (advisor) ; Lázníček, Milan (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate: Tereza Kubačková Consultant: doc. PharmDr. František Trejtnar, CSc. Title of Thesis: Study on renal accumulation of receptor-specific radiopeptides Radiolabeled peptides targeted to cholecystokinin/gastrin receptors are promising compounds for radiodiagnostics and radiotherapy of some malignancies. Shorter chain derivates of gastrin called minigastrins are particularly interesting. The use of these agents is limited by their nephrotoxicity caused by accumulation of the radiopeptides in the kidney. The aim of the study was to investigate accumulation rate and renal uptake mechanisms of new receptor specific peptides from the group of gastrin analogs using in vitro models. The investigation was aimed at a comparison of the renal uptake of DOTA-minigastrin 48 (DOTA-MG48) and DOTA-minigastin 11 (DOTA-MG11), both labeled with indium-111. In addition, a potential influence of the radiolabel on the renal accumulation was also investigated using DOTA- MG11 labeled with indium-111 or with lutecium-177. A comparison of the renal accumulation of the radiolabeled minigastrins with compounds from another group of receptor specific radiopeptides - somatostatin analogs such as 111 In-DOTA-NOC...
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In vitro evaluation of novel Toll-like receptor ligands I.
Hudáková, Kristína ; Trejtnar, František (advisor) ; Soukup, Ondřej (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Kristína Hudáková Supervisor: doc. PharmDr. František Trejtnar, CSc. Title of diploma thesis: In vitro evaluation of novel Toll-like receptor ligands I Vaccination against preventable infections prevents millions of deaths each year. Their immunity enhancing activity is strengthened by the presence of vaccine adjuvants. Development of vaccine adjuvants leads to improved safety profile and also can play a vital role in the research of new vaccines against pathogens against which the vaccines currently do not exist. The main aim of this diploma thesis was to verify the ability of rationally developed small molecule ligands to influence Toll-like receptors and thus their potential to be utilized as vaccine adjuvants. The assay was carried out using modified cell lines continually expressing the human TLR4 or TLR8 whose activation leads to production of secreted embryonic alkaline phosphatase. Ten analyzed substances labelled as DM 001 - DM 010 were examined for their agonistic and also antagonistic properties while interacting with the TLRs. Immunomodulatory activity of these tested samples was then determined by quantification of secreted alkaline phosphatase with the help of a colorimetric...
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Organic Cation Transporter 3 (OCT3/SLC22A3) and Multidrug and Toxin Extrusion 1 (MATE1/SLC47A1) Protein in the Placenta: Expression, Localization and Function
Ahmadimoghaddam, Davoud ; Štaud, František (advisor) ; Trejtnar, František (referee) ; Večeřa, Rostislav (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Candidate Mgr. Davoud Ahmadimoghaddam Supervisor Prof. PharmDr. František Štaud, Ph.D. Title of Doctoral Thesis Organic Cation Transporter 3 (OCT3/SLC22A3) and Multidrug and Toxin Extrusion 1 (MATE1/SLC47A1) Protein in the Placenta: Expression, Localization and Function. The aim of the present study was to investigate the expression, localization, and function of organic cation transporter 3 (OCT3, Slc22a3) and multidrug and toxin extrusion protein 1 (MATE1, Slc47a1) in the rat placenta. Using qRT-PCR, Western blotting and immunohistochemical techniques, we demonstrated abundant expression of OCT3 on the basolateral, i.e., fetus-facing side of the placenta, and MATE1 on the apical, i.e., maternal side of the placenta. To investigate the role of these transporters in the transplacental pharmacokinetics, the in situ method of dually perfused rat term placenta was employed in open- and closed-circuit arrangements; 1-methyl-4-phenylpyridinium (MPP+) was used as a model substrate of both OCT3 and MATE1. We provide evidence that OCT3 and MATE1 cause considerable asymmetry between maternal-to-fetal and fetal-to-maternal transport of MPP+ in favor of fetomaternal direction. Using closed- circuit...
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Preparation of Pharmacokinetic Dictionary
Dřevojánková, Barbora ; Trejtnar, František (advisor) ; Tilšer, Ivan (referee)
The thesis was prepared with the aim of to formulate a complete set of subject words that could serve as a basis for certain areas of the prepared explanatory dictionary on pharmacy. The aim was at least partially contribute to the integration of terminology in there fields of pharmacy. Our task was to analyze the pharmacokinetics and bioelimination of drugs, find source materials regarding these areas in the scientific literature and make a list of possible subject words. Afterwards it was necessary to create an optimized form of particular subject words that could be used as a basis for dictionary. This dictionary will also include other areas of pharmacy and will be intended for wide range of professional pharmacists, medicine doctors and students of these disciplines, or even other readers and general public. Prestigious foreign and domestic publications, especially textbooks on pharmacology published by academically recognized world prestigious authors were the sources for particular preparation and processing of definitions. From these resources, several alternative resolutions were prepared. Afterwards we selected the optimal one or there was created the best version of the subject word parallel following selected criteria. Criteria were firstly clarified of the language, the precision of...
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Viral hepatitis: novel insights and novel therapeutic interventions.
Davidovová, Eva ; Pávek, Petr (advisor) ; Trejtnar, František (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Bc. Eva Davidovová Supervisor: prof. PharmDr. Petr Pávek, Ph.D. Title of diploma thesis: Viral hepatitis: novel insights and novel therapeutic interventions. Viral hepatitis is a well-known worldwide problem. Hepatitis B virus (HBV) and hepatitis C virus (HCV) are characterized by the development of serious complications, especially with regard to the transition to the chronic stage of the disease, associated with the development of fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Interactions between viruses and host cells are quite complicated and not always fully understood. In general, the infection cycle of viruses is a multi-step process. A closer understanding of the entire life cycle of the virus is a major prerequisite for the invention of effective drugs. Viral hepatitis B and C have long been treated mainly with interferon alfa. Ribavirin was later added to HCV treatment and nucleoside / nucleotide analogs (NA) were introduced for HBV. Interferon was later pegylated to improve its properties. However, these drugs did not provide sufficient efficacy and were additionally associated with a number of side effects. It is precisely because of these disadvantages of the current...
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Effect of catechins on toxicity of selected drugs
Rudolfová, Petra ; Skálová, Lenka (advisor) ; Trejtnar, František (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Petra Rudolfová Supervisor: Prof. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: Effect of catechins on toxicity of selected drugs Catechins belong to the flavonoids and they are the main polyphenolic compounds of green tea. Catechins are considered to be very beneficial for the human organism. They have positive effect on the cardiovascular system and reduction of blood sugar levels. Also they have anticancer, antibacterial, antiviral, and antioxidant effects and prevent tooth decay. Catechins, however, also may inhibit or induce important biotransformation enzymes and thus can affect both desirable and undesirable effects of many drugs. In our study, in vitro effect of several of catechins (catechin, epicatechin, epigallocatechin, epicatechin-3-gallate and epigallocatechin gallate), and green tea extract Polyfenonu E on toxicity of drugs was tested. Doxorubicin and acetaminophen, and its toxic metabolite were chosen for our study. Experiments were conducted in intestinal tumor line HCT-8, and in the primary culture of isolated rat hepatocytes. For monitoring the effects of catechins, three different cell viability tests and measurements of reactive oxygen species (ROS) were...
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