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The role of protein kinase C and its targets in cardioprotection
Holzerová, Kristýna ; Hlaváčková, Markéta (advisor) ; Alán, Lukáš (referee) ; Vízek, Martin (referee)
The mortality of cardiovascular diseases remains high and it likely tends to increase in the future. Although many ways how to increase the resistance against myocardial ischemia- reperfusion damage have been described, few of them were transferred into clinical practice. Cardioprotective effect of chronic hypoxia has been described during 60s of the last century. Its detailed mechanism has not been elucidated, but a number of components has been identified. One of these components presents protein kinase C (PKC). The role of PKC was described in detail in the mechanism of ischemic preconditioning, but its involvement in the mechanism of cardioprotection induced by chronic hypoxia remains unclear. One reason is the amount of PKC isoforms, which have often contradictory effects, and the diversity of hypoxic models used. The most frequently mentioned isoforms in connection with cardioprotection are PKCδ and PKCε. The aim of my thesis was to analyze changes in these PKC isoforms at two different cardioprotective models of hypoxia - intermittent hypobaric (IHH) and continuous normobaric hypoxia (CNH). We also examined the target proteins of PKCδ and PKCε after the adaptation to IHH, which could be involved in the mechanism of cardioprotection. These included proteins associated with apoptosis and...
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Adenosine receptors and transporters in rat myocardium: the effect of adaptation to chronic hypoxia
Neumannová, Kateřina ; Novotný, Jiří (advisor) ; Hlaváčková, Markéta (referee)
2. Abstract Adaptation to chronic hypoxia is in addition to ischemic preconditioning one of the two known cardioprotective mechanisms. The precise molecular basis of these processes is still not fully explained. There are some studies that suggest the possible involvement of the adenosinergic signaling system in this adaptation. In this work, we focused on the characterization of the adenosinergic system in the myocardium of rats adapted to two regimens of chronic hypoxia - a protective continuous normobaric hypoxia (CNH) and non-protective intermittent hypoxia (INH/R, 23 h hypoxia and 1 h normoxia). Initially, we compared the total amount of adenosine receptors in samples from different groups of adapted animals. We discovered changes mainly at A2B receptor, which increased at CNH and declined in INH/R. This result suggests the possible involvement of A2B receptors in cardioprotection afforded by adaptation to chronic hypoxia. Furthermore, we investigated the distribution of various types of adenosine receptors and transporters in the plasma membrane of cardiac cells. We observed that A2A and A3 localize in membrane microdomains together with membrane enzyme CD73 that produces adenosine in the extracellular space by degrading AMP. A1 and A2B receptors similarly as nucleoside transporters ENT1, ENT2 and...
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The role of pro- and anti-inflammatory factors in adaptation of heart to hypoxia
Benák, Daniel ; Hlaváčková, Markéta (advisor) ; Holzerová, Kristýna (referee)
Myocardial hypoxia induces several physiological changes on cellular and molecular level in order to keep homeostasis in low oxygen conditions. Essential role in this adaptive response is played by secretory regulatory proteins called cytokines. Differences in concentrations of pro-inflammatory and anti-inflammatory cytokines during hypoxic conditions in the myocardium may significantly influence the fate of the heart tissue, i.e. contribute to its injury or protection. This knowledge is used in experimental cardioprotective strategies - adaptation to chronic hypoxia, ischemic preconditioning, postconditioning and remote conditioning - all of which affect the cytokine levels. The main pro-inflammatory cytokines are TNF-α, IL-1 and cytokine family IL-6. The principal anti-inflammatory cytokine is IL-10. This thesis aims to summarize the effects of pro- and anti-inflammatory cytokines in adaptation of heart to hypoxia and to cover their pathological and protective impact on the myocardium.
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HPLC method devolopment for betacarotene determination in nutraceuticals
Hlaváčková, Markéta ; Šatínský, Dalibor (advisor) ; Chocholoušová Havlíková, Lucie (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Bc. Markéta Hlaváčková Consultant: Doc. RNDr. Dalibor Šatínský, Ph.D. Diploma Thesis Title: Development of HPLC Method for the Determination of Betacarotene in Nutritional Supplements A new HPLC method was optimized and developed for the determination of betacarotene contents in nutritional supplements Betakaroten Farmax, GS Betakaroten FORTE (GreenSwan pharmaceuticals), Walmark Betakaroten, Bioaktivní karoten (Pharma Nord Denmark), Beta karoten Max, BETAVID + LUTEIN (Naturvita), Karovit (Vitabalans Oy), SELZINK PLUS (PRO.MED.CS), Pupalkový olej (Aromatica). The method is based on using Ascentis Express C18 column (30 x 4.6 mm; 2.7 µm) and UV detection at 450 nm. There was used isocratic elution of the mobile phase 100% methanol at a flow-rate of 1.5 ml/min. Column oven temperature was set at 60 řC during the measurement. The retention time of betacarotene under the optimized and validated conditions was 2.185 min. Keywords: betacarotene, HPLC, Betakaroten Farmax, GS Betakaroten FORTE (GreenSwan pharmaceuticals), Walmark Betakaroten, Bioaktivní karoten (Pharma Nord Denmark), Beta karoten Max, BETAVID + LUTEIN (Naturvita), Karovit (Vitabalans Oy), SELZINK PLUS (PRO.MED.CS), Pupalkový olej...
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Role of protein kinase C isoforms in cardioprotective mechanism of chronic hypoxia
Hlaváčková, Markéta ; Novák, František (advisor) ; Kopecký, Jan (referee) ; Novotný, Jiří (referee)
Cardiovascular diseases, particularly acute myocardial infarction, are one of the leading causes of death in developed countries. It is well known that adaptation to chronic intermittent hypobaric hypoxia (IHH) confers long-lasting cardiac protection against acute ischemia/reperfusion injury. Protein kinase C (PKC) appears to play a role in its cardioprotective mechanism since the administration of general PKC inhibitor completely abolished the improvement of ischemic tolerance in IHH hearts. However, the involvement of individual PKC isoforms remains unclear. Therefore, the primary aim of this study was to investigate the potential involvement of PKCδ and PKCε, the most prevalent PKC isoforms in rat heart, in the mechanism of IHH-induced cardioprotection. We showed that IHH up- regulated PKCδ protein in left ventricle, enhanced its phosphorylation on Ser643 and increased its co-localization with markers of mitochondrial and sarcolemmal membranes. PKCδ subcellular redistribution induced by IHH as well as the infarct size-limiting effect of IHH was reversed by acute treatment with PKCδ inhibitor rottlerin. These data support the view that PKCδ plays a significant role in IHH-induced cardioprotection. On the other hand, adaptation to IHH decreased the PKCε total protein level without affecting its...
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The role of mitochondria in cardioprotective effect induced by hypoxia in rat
Lomnický, Matouš ; Žurmanová, Jitka (advisor) ; Hlaváčková, Markéta (referee)
Aerobic organisms need sufficient oxygen supply to maintain homeostasis. These organisms are frequently exposed in hypoxic environments naturally, and also occur in hypoxic states in various pathological conditions. Cardioprotective effect of hypoxia had been recognised more than 30 years ago; and later on, cardioprotective effects of ischemic preconditioning were discovered. Long term exposure to hypobaric hypoxia activates cardioprotective mechanisms, which lower the aftermathes of short term ischemia of myocardia and the effects of further health complications. The core of protective mechanisms has not yet been fully clarified. This work deals with the significance of mitochondria on cardioprotection during hypobaric hypoxia adaptation. This work describes physiological adaptive processes on selected animals on natural hypoxic conditions and also molecular mechanisms, examined on experimental models. Molecular mechanisms of the origins of cardioprotective effects discovered so far, mainly indicate PKC signal pathways through thyrosine kinase and mitogenes of activated kinase and also indicate an activation of sarcKATP-channels and mitoKATP-channels. Opening of these channels can protect mitochondria against a Ca2+ overload, or can lead to an increase in mitochondrial capacity which is possibly connected...
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Autophagy in the heart
Šprláková, Katarína ; Hlaváčková, Markéta (advisor) ; Tomšů, Eva (referee)
Currently, it is growing evidence that autophagy is involved in the prevention of various diseases, which of course also includes heart diseases. This thesis is therefore aimed at clarifying the role of autophagy in the heart, especially during ischemia and subsequent reperfusion. Autophagy is a physiological cellular process by which the cell maintains homeostasis by eliminating long-lived proteins and damaged organelles. The role of autophagy during ischemia/reperfusion in the heart is complex. Predominantly it functions as a pro-survival pathway, because it protects the heart from ischemia or hypoxia. However, when triggered over, which happens during reperfusion, it may lead to cell death. In the heart autophagy is activated in response to various stimuli, such as decrease in ATP and subsequent activation of AMPK, protein Bnip3, reactive oxygen and nitrogen species, the opening of mitochondrial permeability transition pore, endoplasmic reticulum stress or unfolded protein response.
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The role of protein kinase C alpha in individual cancer cell invasiveness
Szabadosová, Emília ; Brábek, Jan (advisor) ; Hlaváčková, Markéta (referee)
Protein kinase C alpha was one of the first identified isoenzyme of PKC family. Since then PKC has been shown to be important in various signal cascades. One of the best known role is in tumor invasiveness and in development of metastases. A role of protein kinase C alpha was pointed out in regulation of Rho/ROCK pathway in amoeboid invasiveness and also Raf/MAPK signaling cascade of mesenchymal movement. ERM proteins, which act in cancer invasiveness, are regulated by protein kinase C too.
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HPLC method devolopment for betacarotene determination in nutraceuticals
Hlaváčková, Markéta ; Šatínský, Dalibor (advisor) ; Chocholoušová Havlíková, Lucie (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Bc. Markéta Hlaváčková Consultant: Doc. RNDr. Dalibor Šatínský, Ph.D. Diploma Thesis Title: Development of HPLC Method for the Determination of Betacarotene in Nutritional Supplements A new HPLC method was optimized and developed for the determination of betacarotene contents in nutritional supplements Betakaroten Farmax, GS Betakaroten FORTE (GreenSwan pharmaceuticals), Walmark Betakaroten, Bioaktivní karoten (Pharma Nord Denmark), Beta karoten Max, BETAVID + LUTEIN (Naturvita), Karovit (Vitabalans Oy), SELZINK PLUS (PRO.MED.CS), Pupalkový olej (Aromatica). The method is based on using Ascentis Express C18 column (30 x 4.6 mm; 2.7 µm) and UV detection at 450 nm. There was used isocratic elution of the mobile phase 100% methanol at a flow-rate of 1.5 ml/min. Column oven temperature was set at 60 řC during the measurement. The retention time of betacarotene under the optimized and validated conditions was 2.185 min. Keywords: betacarotene, HPLC, Betakaroten Farmax, GS Betakaroten FORTE (GreenSwan pharmaceuticals), Walmark Betakaroten, Bioaktivní karoten (Pharma Nord Denmark), Beta karoten Max, BETAVID + LUTEIN (Naturvita), Karovit (Vitabalans Oy), SELZINK PLUS (PRO.MED.CS), Pupalkový olej...
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