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Membrane Properties of Glial Cells after Ischemia in vivo
Pivoňková, Helena ; Anděrová, Miroslava (advisor) ; Vyklický, Ladislav (referee) ; Vlachová, Viktorie (referee)
In my PhD thesis, I aimed to characterize the membrane properties of astrocytes and NG2 glia during the acute and chronic phases of the ischemic injury in the central nervous system with a focus on the expression and activity of K+ ion channels. Two ischemia models in rats were used for the experiments - global cerebral ischemia 5 causing selective neuronal loss and astrogliosis in the CA1 region of the hippocampus, and incubation of spinal cord segments in a solution with high K+ concentration. We found that hippocampal astrocytes depolarize starting 3 days after ischemia, and demonstrate an increase in the inward rectification of their whole cell currents 1 month after ischemia. The Kir4.1 channel expression in the hippocampal tissue was downregulated starting 3 days after ischemia, while the expression of Kir2.1, Kir5.1 and TREK1 channels was strongly upregulated in reactive astrocytes 1 month after ischemia. NG2 glia displayed a significant increase in the outwardly rectifying delayed and A-type K+ (KDR and KA) currents 2 hours and 3 days after ischemia, and a decrease in the inwardly rectifying K+ currents 3 days after ischemia, a typical current pattern of proliferating cells. Complex astrocytes in the spinal cord pre-incubated in a solution with high [K+] showed the largest changes in their membrane...
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Cell therapy in animal models - preclinical studies
Juhásová, Jana ; Motlík, Jan (advisor) ; Grim, Miloš (referee) ; Jendelová, Pavla (referee)
The progress of cell therapy can be greatly facilitated by using suitable experimental models. It is essential to verify the clinical usefulness of new healing procedures obtained in studies on laboratory animals by using a large animal model. One of suitable models well acceptable in medical community is undoubtedly the miniature pig, which resembles humans in terms of physiology and body proportions. This PhD thesis presents the summary of our experimental studies relating to possible exploitation of mesenchymal and neural stem cells in the healing of locomotive apparatus and neural tissue disorders in humans or animals. The first part of the thesis briefly describes the current issue of cell therapy and animal models, mesenchymal cells and/or their combination with new types of scaffolds, neurogenesis, neural stem cells and their potential application in therapy of spinal cord injury. The second part is focused on the goals and methodology, the individual publications being listed in the third part. Our experiments with iatrogenic physeal defect in rabbits, which served as a model of the occurrence of valgous deformation in the clinical practice, showed the positive preventive and therapeutical effects of a new type of scaffolds seeded with allogeneic mesenchymal stem cells in animals without...
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Effect of intravenously applicated lactoferine on early faze; of myocardial ischemia II. Biochemical examinations
Písaříková, Veronika ; Semecký, Vladimír (advisor) ; Mladěnka, Přemysl (referee)
Lactoferrin (Lf) is an 80-kDa iron-binding glykoprotein. Lf is present physiologically in exocrine secretions, eg. tears, saliva, milk, sinovial fluid, seminal fluid and in the secondary granules of neutophils. The precise function of Lf in organism is considered to be very complex and it is still a hot subject of scientific disputation. Lf was documented to act as antimicrobial, antiinflamatory and antitumoral agent. Another of its properties is ability to inhibit hydroxyl radical formation via Fenton reaction. Afinity of Lf to iron is about 260times higher than that of transferrin. Isoproterenol (ISO) has been used as a model compound to induce infarct-like lesions in the rat and various other animal species. Administration of necrotic dose of ISO caused ischaemia followed by damage of the myocardium. Ischaemia alters iron homeostasis and redox-active free iron, which catalyses ROS-generation. The only possibility for myocardial tissue recovery in acute moycardial infarction represents the reperfusion of ischemic myocardium. But the whole process is associated with damage of myocardium due to burst of hydroxyl radical catalyzed by free iron (Fenton reaction). In our experiment we studied the effects of Lf in a catecholamine model of myocardial injury. We had four experimental groups divided according to...
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Cardioptrotective role of transcription factor HIF isoforms in rats
Bučinská, Ivana ; Žurmanová, Jitka (advisor) ; Pavlínková, Gabriela (referee)
Adaptation to chronic hypoxia is characterized by a variety of functional changes in order to maintain metabolic and energy homeostasis. It has been known for many years that both humans and animals indigenous or adapted to high-altitude hypoxia are more tolerant to an acute ischemic injury of the heart. HIF1α is found as a primary transcriptional regulator of adaptive response to hypoxia in all tissues, whereas HIF2α is more tissue specific. The activity of HIFα is regualted by prolyl hydoxylases (EGLN). The EGLN1 was shown to be more efficient in HIF1α hydroxylation than in HIF2α. As well as the EGLN3 is more specific for HIF2α . Under hypoxic conditions, HIF activates the transcription of about 70 genes. Hence, HIF1α and HIF2α can play an essential role in pathophysiology of ischemic heart disease. The aim of our study was to determine myocardial expression of HIFs and its regulatory hydroxylases in adult male Wistar rats adapted to continuous normobaric hypoxia (H; FIO2 = 0.1) for 3 weeks. Another two groups of rats were exposed to normobaric hypoxia intermittently for either 8 h/day (INH) or 23 h/day (RH) during the 3-week adaptation period. While H induces protective cardiac phenotype, the later regimen (RH) does not. The protein expression of HIFs, PHDs and FIH were then determined by...
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Myocardial beta-adrenergic signaling during adaptation of rats to chronic hypoxia
Hahnová, Klára ; Novotný, Jiří (advisor) ; Ostašov, Pavel (referee)
Endogenous cardiac protection against acute ischemia/reperfusion injury can by increased by cardiac adaptation to various forms of chronic hypoxia. Chronic hypoxia induces a large variety of adaptive changes in the myocardium that could be considered as protective, but the exact mechanism of increased ischemic tolerance is unknown. Different studies suggest that catecholamine release and their effect on -adrenergic signaling after adaptation to chronic hypoxia contributes to cardioprotection. In this study we focused on characterization of -adrenergic receptors ( -ARs) in the myocardium of rats after adaptation to three different hypoxic conditions: 1. intermittent normobaric hypoxia - INH/R (23 h hypoxia, 1 h reoxygenation), 2. intermittent normobaric hypoxia - INH (8 h hypoxia, 16 h normoxia), 3. continuous normobaric hypoxia - CNH (24 h hypoxia). We compared how each hypoxic model affects the total number of -adrenergic receptors and proportion of individual subtypes ( 1-and 2-ARs) in the left and right ventricles compared control normoxic rats. The INH model had apparently no effect on -ARs in either ventricles. On the other hand, adaptation to INH/R and CNH was accompanied by a significant decrease (by about 25%) in the total number of -adrenergic receptors in the right ventricles. Our present...
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Analysis of selected gene transcripts in the rat myocardium adaptated to chronic hypoxia
Kašparová, Dita ; Žurmanová, Jitka (advisor) ; Nováková, Olga (referee)
Dita Kašparová Chronická hypoxie a exprese genů 4 Abstract Adaptation to chronic hypoxia (CH) is characterized by a variety of functional changes in order to maintain metabolic and energy homeostasis. It has been known for many years that both humans and animals indigenous or adapted to high-altitude hypoxia are more tolerant to an acute ischemic injury of the heart. Cardioprotective mechanisms activated by adaptive responses to chronic hypoxia can be the result of altered transcriptional regulations in left ventricles. Here we report results from the gene expression profiling of adaptive responses in three models of chronically hypoxic heart. Adult male Wistar rats were exposed for 21 days to either continuous normobaric hypoxia (CCH; 10% O2) or CCH interrupted daily by 1-hour reoxygenation (RCH) or CCH interrupted daily by 16-hour (CIH). Cardiprotective effect of CCH adaptation is abolished by brief daily reoxygenation, RCH adaptation. In the present study, we aimed to determine myocardial mRNA expression of 19 candidate genes divided into three important groups: i) Hypoxia inducible factor (HIF1α) and its prolyl and asparaginyl hyroxylases (PHDs and FIH respectively, ii) Creatine kinase (CK) isoenzymes which play important role in energy homeostases of heart and iii) the group of main enzymatic...
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Study of Electrophysiological Function of the Heart in Experimental Cardiology
Ronzhina, Marina ; Tyšler, Milan (referee) ; Halámek, Josef (referee) ; Kolářová, Jana (advisor)
Srdeční poruchy, jejichž příkladem je ischemie myokardu, infarkt, hypertrofie levé komory a myokarditida, jsou v experimentální kardiologii obvykle studovány na modelu izolovaného srdce. Kritéria pro detekci srdečních poruch však nejsou pro zvířecí modely standardizována, což komplikuje srovnání a interpretaci výsledků různých experimentálních studií. Obzvlášť složitá situace nastává při současném výskytu několika patologických jevů, jejichž vzájemná součinnost komplikuje rozpoznání jejich individuálních účinků. Korektní posouzení stavu srdce vyžaduje také zohlednění mnoha faktorů spojených s akvizicí dat. Tato práce je věnována kvantitativnímu hodnocení elektrofyziologických změn způsobených globální ischemií myokardu. Vliv ischemie byl hodnocen pro fyziologická srdce a srdce se zvětšenou levou komorou a dále pro srdce nabarvená napěťově-citlivým barvivem di-4-ANEPPS. Přestože jsou oba fenomény často zastoupeny v animálních studiích, nebyl dosud popsán jejich vliv na manifestaci ischemie v elektrogramech (EG), ani nebyl kvantifikován jejich vliv na přesnost detekčních algoritmů pro identifikaci ischemie. Práce shrnuje kvantitativní změny srdeční funkce vyvolané ischemií (v normálních podmínkách, při hypertrofii levé komory, a při administraci barviva) založené na hodnocení EG a VKG parametrů. Dále práce obsahuje rozbor důležitých aspektů akvizice záznamů, jako je umístění snímacích elektrod, způsob výpočtu deskriptorů z EG a VKG (s použitím výsledků manuálního rozměření záznamů, nebo bez něj) a identifikace okamžiku vývoje ischemie v preparátu. Nedílnou součást práce tvoří návrh, realizace a ověření metod pro automatickou detekci ischemie v experimentálních záznamech. Výsledky práce dokazují, že dosažení opakovatelných a věrohodných výsledků je podmíněno zohledněním všech výše uvedených faktorů souvisejících jak se stavem srdce, tak s metodikou záznamu a analýzy dat.
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Antioxidative enzymes in heart
Kletečková, Lenka ; Hloušková, Patricie (advisor) ; Rychlíková, Jana (referee)
Cardiovascular diseases are one of the most frequently causes of morbidity and mortality in developed countries. An increase of oxidative stress is implicated by progression of heart diseases. Antioxidative enzymes protect the organism against them. The superoxide dismutases, the peroxiredoxins, the glutathione peroxidases and the catalase are advised to be used as the main antioxidative enzymes. The review is focused on the role of these antioxidative enzymes in the cardiovascular system. Antioxidative enzymes prevent the lipoperoxidation of biological membranes, the proteins inactivation, the DNA mutation and the apoptosis. It is well-known, that antioxidative enzymes positively influence a homeostasis of the vascular endothelium, prevent a burst of the cardiovascular disease and they are important for physiological functions, the morphology and the contractility of heart. The effect of antioxidative enzymes on the cardiovascular system is very complicated process and it still has not clarified all aspects of their action. A reseach of these enzymes is essential for understading the mechanisms of pathophysiological changes, which proceed into the cardiovascular system during the atherosclerosis, the myocardial infarction, the ischemic- reperfusion injury of the tissue and the calcification and...
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The role of lipids and ROS in cardioprotective mechanism of chronic hypoxia
Balková, Patricie
The role of lipids and ROS in cardioprotective mechanism of chronic hypoxia Cardiovascular diseases, mainly ischemic heart disease is one of the most frequently cause of morbidity and mortality in developed countries. Therefore effective protection of the heart against ischemia and reperfusion injury is the crucial aim of experimental and clinical cardiology. One of the main streams of cardiovascular research is looking for possibilities of natural heart resistance augmentation. Adaptation to chronic hypoxia is one possibility how to protect the heart against ischemia-reperfusion injury. Chronic hypoxia increases resistance of the myocardium to acute deficiency of oxygen leading to vetricular arrythmias, postischemic contractile dysfunction and necrotic changes in the tissue. Recently, it has been shown that reactive oxygen species (ROS) play an important role in the cardioprotective mechanism of chronic hypoxia. It is known that oxidative stress has a harmful effect in acute ischemia-reperfusion however ROS generated during the adaptation to hypobaric intermittent chronic hypoxia play a role in the induction of cardioprotection. In this study, we demonstrated that adaptation of adult rats to chronic hypoxia increased the activity and protein abundance of manganese superoxide dismutase (MnSOD) in the...
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The role of lipids and ROS in cardioprotective mechanism of chronic hypoxia
Balková, Patricie ; Nováková, Olga (advisor) ; Drahota, Zdeněk (referee) ; Šmíd, František (referee)
The role of lipids and ROS in cardioprotective mechanism of chronic hypoxia Cardiovascular diseases, mainly ischemic heart disease is one of the most frequently cause of morbidity and mortality in developed countries. Therefore effective protection of the heart against ischemia and reperfusion injury is the crucial aim of experimental and clinical cardiology. One of the main streams of cardiovascular research is looking for possibilities of natural heart resistance augmentation. Adaptation to chronic hypoxia is one possibility how to protect the heart against ischemia-reperfusion injury. Chronic hypoxia increases resistance of the myocardium to acute deficiency of oxygen leading to vetricular arrythmias, postischemic contractile dysfunction and necrotic changes in the tissue. Recently, it has been shown that reactive oxygen species (ROS) play an important role in the cardioprotective mechanism of chronic hypoxia. It is known that oxidative stress has a harmful effect in acute ischemia-reperfusion however ROS generated during the adaptation to hypobaric intermittent chronic hypoxia play a role in the induction of cardioprotection. In this study, we demonstrated that adaptation of adult rats to chronic hypoxia increased the activity and protein abundance of manganese superoxide dismutase (MnSOD) in the...
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