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Detections of opioid receptors in plasma membranes. Binding assays with specific radioligands
Višněvská, Kateřina ; Hudeček, Jiří (advisor) ; Wilhelm, Jiří (referee)
Morphin is an important opioid drug which is used in medicine as a highly potent analgesic in treatment of both acute and chronic pain, post-traumatic shock and diarrhea. However, in treatment of chronic pain, the serious problem is represented by step-by-step development of decreased sensitivity of physiological response to morphine exposure (tolerance) and, subsequently, the psychological as well as physical dependence on this drug. Opioids interact with three types of opioid receptors: μ-OR, δ-OR and κ-OR. One of primary goals of many pharmaceutical companies is an effort to design more specific and simultaneously potent OR agonists, which would exhibit highly efficacious analgesic effects, but would not develop fast physical dependence in conditions of the long-term usage. In our work, the primary attention was devoted to μ-OR because these receptors are primarily responsible for physiological effect of morphine and genesis of tolerance and drug dependence. We studied the binding of specific agonist [3 H]DAMGO to plasma membranes (PM) isolated from rat brain cortex of rats adapted to increasing doses of morphine (10-50 mg/kg) for 10 days. PM isolated by centrifugation in Percoll density gradient from morphine-adapted rats exhibited the decrease of maximum binding capacity Bmax of this...
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Assay of fluorescent products in erytrocytes of patients with Alzheimer' s disease
Kohutiar, Matej ; Wilhelm, Jiří (advisor)
Free radicals are highly reactive species with one unpaired electron in orbital. Reactive oxygen and nitrogen species count among important biologic molecules of radical nature. It is very important to fix a concentration of free radicals in cell on non-toxic limits. Products of radical damage are cumulated extra or intracellulary and they are main components of lipofuscin-like pigments. Lipofuscin-like pigments contains in their molecular structure fluorofores, so they are good substrates for fluorescent analysis. Alzheimer's disease is a very actual social and economical problem. Etiology of Alzheimer's disease is still unknown. Histologically, the characteristic presence of Alzheimer's disease is a senil plaques of amyloide ?. ROS and RNS diffuse through hematoencephalic barrier in vessel's lumen and attacks red blood cells. Radical damage of erythrocytes is associated with an increase of concentration of oxidative stress products in cytosol. Sample for fluorescent analysis has been prepared from a red-blood cell extract from 30 patients and 8 healthy controls. Fluorescent spectra of healthy controls have emission maxima in area 327-343nm. In compare with controls, spectra of pacients were more heterogenous in area upon 380nm. The study of oxidative cell damage is important for understanding of...
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Tissue factor in acute coronary syndromes
Malý, Martin ; Hampl, Václav (advisor) ; Wilhelm, Jiří (referee) ; Vízek, Martin (referee) ; Málek, Ivan (referee)
In this review are summarized essentials of the physiology and the pathophysiology of TF in terms of the literature sources. But still, until today, there are many questions to be answered: the source and the level of TF in healthy subjects, the method how to measure the "trombogenic" potential of TF because its role in many biological processes, the "normal" levels, the possible impact of risk factors and possibility of risk stratification of "healthy" subjects according to the levels of TF like marker of the blood with procoagulant potential. Answering these questions and impact of the blocking of TF as a therapeutical approach in acute coronary syndromes is a challenging field of investigation in blood coagulation. Powered by TCPDF (www.tcpdf.org)
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The study of fluorescent products in the mitochondria after an attack by free radicals
Ivica, Joško ; Wilhelm, Jiří (advisor) ; Entlicher, Gustav (referee) ; Drahota, Zdeněk (referee)
Lipofuscin-like pigments are products of reactions involving free radical attack onto molecules with nucleophilic groups. They can be for med, for example, in the reactions between lipid peroxidation decomposition products, such as aldehydes, and amino-group containing compounds, e.g. phospholipids, peptides. Owing to their intrinsic fluorescent properties LFP can be easily measured. LFP are relatively stable and therefore have been successfully used as robust markers of oxidative damage. We undertook the metabolomic studies, where fluorescent LFP were first analysed spectrofluorimetrically by using tridimensional and differential fluorescence spectral arrays. After that, certain LFP were analysed by means of high performance liquid chromatography, in order to resolve the mixture of compounds into distinct fractions. For this purpose we used LFP prepared after isolated heart itochondria had been exposed in vitro to oxidative stress initiated by various triggers. LFP were also analysed during early development in rat brain, which is accompanied by transient increase in oxygen concentration, and in erythrocytes from patients with Alzheimer's disease. We developed HPLC methods for qualitative analysis of LFP of different origin. This analysis unfolded that LFP indeed consist of many chromatographically...
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Detections of opioid receptors in plasma membranes. Binding assays with specific radioligands
Višněvská, Kateřina ; Hudeček, Jiří (advisor) ; Wilhelm, Jiří (referee)
Morphin is an important opioid drug which is used in medicine as a highly potent analgesic in treatment of both acute and chronic pain, post-traumatic shock and diarrhea. However, in treatment of chronic pain, the serious problem is represented by step-by-step development of decreased sensitivity of physiological response to morphine exposure (tolerance) and, subsequently, the psychological as well as physical dependence on this drug. Opioids interact with three types of opioid receptors: μ-OR, δ-OR and κ-OR. One of primary goals of many pharmaceutical companies is an effort to design more specific and simultaneously potent OR agonists, which would exhibit highly efficacious analgesic effects, but would not develop fast physical dependence in conditions of the long-term usage. In our work, the primary attention was devoted to μ-OR because these receptors are primarily responsible for physiological effect of morphine and genesis of tolerance and drug dependence. We studied the binding of specific agonist [3 H]DAMGO to plasma membranes (PM) isolated from rat brain cortex of rats adapted to increasing doses of morphine (10-50 mg/kg) for 10 days. PM isolated by centrifugation in Percoll density gradient from morphine-adapted rats exhibited the decrease of maximum binding capacity Bmax of this...
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The role of arachidonic acid metabolites via cytochrome P-450 in the pathogenesis of hypertension
Čertíková-Chábová, Věra ; Tesař, Vladimír (advisor) ; Wilhelm, Jiří (referee) ; Štípek, Stanislav (referee)
Background: 20-HETE and EETs, metabolites derived from arachidonic acid by cytochrome P-450 (CYP), are part of the effector mechanisms in the renin-angiotenzin-aldosterone system. They regulate vasoconstriction/ vasodilation and natriuresis and thus contribute to the regulation of blood pressure. Hypertensive rats transgenic for mouse Re-2 renin gene (TGR) are a good model for hypertension caused by a single gene and very suitable for the study of the role20-HETE and EETs in vivo. Hypothesis: Abnormal production and/or activity of 20-HETE and EETs contributes to the development and or/maintenance of high blood pressure in TGR. Materials and methods: Hypertensive TGR heterozygous males of various ages weres studied. Normotensive HanSD animals (the same genetic background without Ren-2 gene) were used as controls. Three complex studies were performed: Study 1: Chronic inhibition of CYP with non-selective inhibitors 1- aminobenzotriazol (ABT) and CoCl2 in young prehypertensive and adult hypertensive animals and respective controls. Study 2: Acute experiments with selective inhibitors N-metyl-sulfonyl- 12,12-dibromododec-11- enamide (DDMS) and N-metylsulfonyl-6-(2- propargyloxyfenyl)-hexamide MS-PPOH. Study 3: Chronic selective inhibition of 20-HETE production by DDMS and selective inhibition of EETs...
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Tissue factor in acute coronary syndromes
Malý, Martin ; Hampl, Václav (advisor) ; Wilhelm, Jiří (referee) ; Vízek, Martin (referee) ; Málek, Ivan (referee)
In this review are summarized essentials of the physiology and the pathophysiology of TF in terms of the literature sources. But still, until today, there are many questions to be answered: the source and the level of TF in healthy subjects, the method how to measure the "trombogenic" potential of TF because its role in many biological processes, the "normal" levels, the possible impact of risk factors and possibility of risk stratification of "healthy" subjects according to the levels of TF like marker of the blood with procoagulant potential. Answering these questions and impact of the blocking of TF as a therapeutical approach in acute coronary syndromes is a challenging field of investigation in blood coagulation. Powered by TCPDF (www.tcpdf.org)
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The effect of free radicals on ageing
Lukšanová, Hana ; Wilhelm, Jiří (advisor) ; Entlicher, Gustav (referee) ; Macháčková, Ivana (referee)
Cílem této studie bylo ověřit platnost Harmanovy teorie stárnutí, založené na působen í volných ra dikálů, na nových modelech stárnutí rostlin a kvasinek, jelikož tyto organizmy byly nedávno přijaty mezinárodní komunitou jako uznávané modely pro studium mechanizmu stárnutí na molekulární úrovni. Vzhledem k tomu, že v Harmanově teorii stárnutí má centrální význam endogenní produkce volných radikálů, zaměřila jsem se ve své studii na sledování časového průbě hu vzniku jejich koncových produktů . Paralelně jsem sledovala změny antioxid a ční ch systémů, které rozhodují o rozsahu poškození vyvolaném volnými radikály. V poslední době se ukazuje, že procesů stárnutí se účastní i reaktivní slo u čen iny dusíku, a proto jsem zaměři l a pozornost i na mechanizmy zahrnující účast těchto sloučenin. Jako modelové organizmy jsem použila fazol (Phaseo/us vu/garis L.), trasgenní rostliny tabáku (Nicotiana tabacum L. ) s pozměněnou koncentrací cytokininů, husen íček rolní (Arabidopsis tha/iana) a kvasinky (Saccharomyces cerevisiae). Powered by TCPDF (www.tcpdf.org)
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