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The role of PCr/CK shuttle in adult rat myocardium under normoxic hypoxic conditions
Honcová, Lada ; Žurmanová, Jitka (advisor) ; Novotný, Jiří (referee)
The creatine kinase (CK) is an important enzyme of cell energy metabolism in excitable tissue. It occurs in four isoforms. Two cytosolic isoforms are functional in mono and hetero dimers and two mitochondrial isoforms reach tetramer and octamer forms. Its primary function is the regeneration of ATP close to ATPases and phosphocreatine pool from creatine and ATP, which gives its posphate in places of acute requirements of high energy demand. Dysfunction of CK is connected with heart, muscle and neurological diseases and CK is often used as a clinical indicator. This work is focused to the role of CK in energy metabolism of hypoxia adapted myocardium. CK thaks to production of ADP in mitochondria decreases a membrane potential as well as production of reactive oxygen species (ROS). ROS cause most of damage during ischemic heart disease and infarct of myocardium. That`s why cardioprotective effects and CK itself during hypoxia are investigated.
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Localization of cytosolic isoforms of creatine kinase and hexokinase in hypertrophied heart
Heleš, Mário ; Žurmanová, Jitka (advisor) ; Mrózková, Petra (referee)
Hypertrophy of the heart is tightly bound to the metabolic adaptations and a cellular remodeling. An important and dynamic system contributing to the maintenance of energy homeostasis is the creatine kinase system (CK). The microcompartmentalization of CK isoforms maintains the flux of ATP between energy production and consumption sites and ensures the effectiveness of the CK system. Changes in expression and activity of CK isoforms during hypertrophy are already well described - to extend this knowledge, this thesis quantified changes in association of cytosolic CK isoforms and sarcomeres. Another essential system, maintaining homeostasis in overloaded heart is composed of the hexokinase (HK) isoforms, located also in cytosol and in mitochondrial compartment. HK1 is associated with mitochondria under physiological conditions, maintaining mitochondrial membrane potential, while HK2 is located mainly in the cytosol. Under stress conditions translocation of HK2 into mitochondrial membrane occurs, which increases the direct supply of ADP to complex V of the respiratory chain and decreases the probability of apoptosis activation. We analyzed association of individual HK isoforms with mitochondria within the second aim of this thesis. Third aim of the thesis was to characterize changes in the CK and M...
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The role of PCr/CK shuttle in adult rat myocardium under normoxic hypoxic conditions
Honcová, Lada ; Žurmanová, Jitka (advisor) ; Novotný, Jiří (referee)
The creatine kinase (CK) is an important enzyme of cell energy metabolism in excitable tissue. It occurs in four isoforms. Two cytosolic isoforms are functional in mono and hetero dimers and two mitochondrial isoforms reach tetramer and octamer forms. Its primary function is the regeneration of ATP close to ATPases and phosphocreatine pool from creatine and ATP, which gives its posphate in places of acute requirements of high energy demand. Dysfunction of CK is connected with heart, muscle and neurological diseases and CK is often used as a clinical indicator. This work is focused to the role of CK in energy metabolism of hypoxia adapted myocardium. CK thaks to production of ADP in mitochondria decreases a membrane potential as well as production of reactive oxygen species (ROS). ROS cause most of damage during ischemic heart disease and infarct of myocardium. That`s why cardioprotective effects and CK itself during hypoxia are investigated.
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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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Úloha regulačních proteinů pro pohyblivost rybích spermií
DZYUBA, Viktoriya
The investigation of the energetic aspects of spermatozoon motility implementation (Chapter 2) was carried out using demembranated spermatozoa of taxonomically distant fish species (common carp and sterlet). Special attention was given to the functioning of ATP regeneration systems: adenylate kinase (AK), and creatine kinase (CK). It was shown for the first time that the phosphocreatine/CK system is present in sterlet spermatozoa and plays an essential role in ATP regeneration. Spermatozoa of carp and sterlet were shown to have similar systems for ATP regeneration, while the efficacy of the studied systems differs in these species. The low baseline activity of CK in carp and AK in sterlet suggest these to be the source of the most pronounced effects of their inhibition on energy supply for flagella movement in the respective species. The presence of a maturational process during the post-testicular transit of sperm in sturgeon was recently ascertained in our laboratory (Chapter 3). This discovery prompted investigation of the factors that regulate this process including the involvement of proteolysis regulators and prooxidant-antioxidant system. As a result of this study (Chapter 3.3), we found that there was no significant difference between proteolytic profiles of seminal fluids (SF) of testicular sperm (TS) and Wolffian duct sperm (WS). It suggests that the majority of proteases present in SF of mature sperm originate in the testis. Measure of amidase and anti-proteolytic activities in the SF of sterlet sperm showed significant decrease in activities as the sperm passed through the kidneys and Wolffian ducts. Considering our observation that trypsin inhibition during in vitro TS maturation blocked the maturation process (Chapter 3.1), and based on zymography, amidase and anti-proteolytic activity determination, we think that the decrease in anti-proteolytic activity of spermatozoa surroundings during their post-testicular transit could be needed to prepare them for maturation. The present study showed that maturation of sturgeon spermatozoa and different times of storage in Wolffian ducts (in vivo storage), are accompanied by significant alterations in motility parameters as well as in SF redox balance (Chapter 4.1). A high level of TBA-reactive substances (TBARS) and a high activity of antioxidant enzymes were found in immature TS compared to those in WS. The high activity of the enzymatic antioxidant system (AOS) allows TS to cope with the deleterious effects of excessive reactive oxygen species production and to retain the ability to become motile after post-testicular transit, or after in vitro maturation. The increase in TBARS content during in vivo storage was not accompanied by a corresponding increase in activity of AOS. We suggest that extended time in the Wolffian ducts resulted in sperm oxidative damage resulting from inadequate AOS efficacy and, finally, in decreases in motility parameters. Short-term hypothermic in vitro storage of sterlet sperm resulted in a significant decrease in motility and velocity without changes of AOS activity (Chapter 4.2). It means that AOS of sterlet sperm is effective enough to prevent the development of oxidative stress during short-term storage. Short period of tench sperm exposure to hypotonic conditions was shown to induce oxidative stress and, as a result, sperm quality decline (Chapter 4.3). The combined results of the study concerning the regulation of sperm prooxidant-antioxidant status (Chapter 4) during spermatozoa maturation, motility activation and sperm in vivo and in vitro storage may confirm a dual role of reactive oxygen species (regulatory or damaging depending from the levels of their formation and elimination) in fish sperm physiology.
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