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The content of components of ATP synthasome in different rat tissues and in patients with defects in ATP synthase
Mikulová, Tereza ; Houštěk, Josef (advisor) ; Kalous, Martin (referee)
The complexes of oxidative phosphorylation (OXPHOS) are situated in the inner mitochondrial membrane in higher structural and functional complexes, so-called supercomplexes, which facilitates substrate channeling. ATP synthase is also able to organize in higher structures. In mammalian mitochondria, ATP synthase is usually present in a dimeric form. There is evidence of its trimerization and even tetramerization. Furthermore, it seems that ATP synthase catalyzing the phosphorylation of ADP to ATP, adenine nucleotide translocator (ANT) ensuring the exchange of ADP for newly synthesized ATP across the inner mitochondrial membrane and phosphate carrier (PiC) allowing the import of inorganic phosphate (Pi) into the matrix of mitochondria are assembled in a supercomplex called ATP synthasome. This association among the components of phosphorylative apparatus seems to increase the efficiency of processes leading to the ATP synthesis. First, we studied amounts of the components of phosphorylative apparatus in connection with various ATP synthase contents among mitochondria isolated from nine rat tissues. Mitochondrial proteins were separated by denaturing electrophoresis (SDS-PAGE) and their content was analyzed using specific antibodies. In agreement with our expectations, the highest content of...
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Possibilities of UCP2 (uncoupling protein 2) induction in rat hepatocytes in in vivo conditions
Bolehovská, Radka ; Červinková, Zuzana (advisor) ; Kalous, Martin (referee) ; Novotný, Dalibor (referee)
Possibilities of UCP2 (uncoupling protein 2) induction in rat hepatocytes in in vivo conditions Introduction Uncoupling protein 2, discovered in 1997, is the first described homologue of uncoupling protein 1. Uncoupling proteins increase the permeability of inner mitochondrial membrane for protons, decrease the efficiency of energy conversion, inhibit the ATP synthesis and stimulate energy release in form of heat. Uncoupling proteins also increase the substrate oxidation and reduce production of reactive oxygen species in mitochondria. Objective, material and method The aim of this study was to establish and optimised the quntitative real-time PCR for detection of UCP2 mRNA expression kinetics in rat liver tissue. The present study was conducted to assess the effects of acute and chronic treatment with triiodothyronine and the effect of partial hepatectomy on liver uncoupling protein 2 mRNA levels in male Wistar rats. Results Intraperitoneal injection of one dose of triiodothyronine (200 μg/kg rat body weight) increased mRNA expression of uncoupling protein 2 in liver tissue almost 2-fold (P < 0.01 vs. control group) in rats 12 hours after T3 administration. Concentrations of total triiodothyronine and free triiodothyronine in serum were increased 122-fold and 77-fold (p < 0.001), respectively....
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Mitochondrial production of reactive oxygen species and its role in physiological regulations
Holzerová, Eliška ; Mráček, Tomáš (advisor) ; Kalous, Martin (referee)
The production of mitochondrial reactive oxygen species and the resulting oxidative stress is an important phenomenon driving long-lasting research and intense discussions. Knowledge of exact mechanisms of reactive oxygen species production and pathways leading to their formation could help us to directly affect their production, a task with potential terapeutic implications. The molecular nature of the production of reactive oxygen species by some enzymes has already been well documented, but others still remain controversial and current theories are obviously far from the truth. Much more interesting is the question of physiological importace of this production. The reactive oxygen species were considered harmful factors clearly distorting the integrity of the organism for a long time. However, recent research suggest that their existence can also be beneficial and effective. Evidently they can serve as a signaling molecules in several metabolic and regulatory pathways occurring in the organism. This bachelor thesis offers insight into the current state of knowledge. It focuses on the most detailed description of the reactive oxygen species production by mitochondrial respiratory chain enzymes. Furthermore, it deals with some signaling cascades, where involvement of mitochondrially generated...
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Mitochondrial dynamics in myocardium.
Weissová, Romana ; Nováková, Olga (advisor) ; Kalous, Martin (referee)
The heart is an absolutely vital body organ, which requires sufficient amount of active mitochondria for its energy demanding activity. The functionality of a mitochondrial population is maintained through mitochondrial turnover, encompassing mitophagy removing damaged mitochondria and mitochondrial biogenesis responsible for the emergence of new organelles. Dynamic processes of mitochondrial fusion and fission can also contribute to the maintenance of a healthy mitochondrial population. Mitochondrial fusion and fission have not yet been proven in cardiomyocytes, although these cells possess all the proteins required for these events. These processes, however, take on the importance during pathological conditions, when changes in the amount of protein applied in the mitochondrial dynamics occur. The modification in mitochondrial phenotype leads to the cell damage. Understanding the role of mitochondrial dynamics in myocardium may contribute to the development of new heart diseases treatments.
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Affecting the growth of a tumor on an experimental animal model
Kučera, Alexandr ; Škába, Richard (advisor) ; Eckschlager, Tomáš (referee) ; Havránek, Petr (referee) ; Kalous, Martin (referee)
In this study we present the models of preventive and therapeutic vaccination of sarcoma-bearing rats with dendritic cells that present tumor antigens from killed tumor cells. We present the characteristics of DCs-based vaccine and its capacity to induce antitumor immune response both in v vitro and in vivo. We show that preventive vaccination efficiently prevents the growth of tumors. On the other hand vaccination of rats with established tumors did not led to the eradication of tumors. Despite the induction of a vigorous immune response after administration of DCs-based vaccine and transient decrease in tumor progression, tumors eventually resumed their growth and animals vaccinated with DCs succumbed to cancer. In both settings, preventive and therapeutic, DCs-based vaccination induced a vigorous tumor specific T cells response. These results argue for the timing of cancer immunotherapy to the stages of low tumor load. Immunotherapy initiated at the stage of minimal residual disease, after reduction of tumor load by other modalities, will have much better chance to offer a clinical benefit to cancer patients than the immunotherapy at the stage of metastatic disease. Powered by TCPDF (www.tcpdf.org)
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Mitochondrial uncoupling and mitochondrial morphology in relation to type-2 diabetes
Špaček, Tomáš ; Ježek, Petr (advisor) ; Kalous, Martin (referee) ; Vařecha, Miroslav (referee)
- 2 - 1 SUMMARY Increased ATP/ADP ratio resulting from enhanced glycolysis and oxidative phosphorylation represents a plausible mechanism controlling the glucose-stimulated insulin secretion (GSIS) in pancreatic -cells. Using high resolution respirometry and parallel mitochondrial membrane potential ( m) monitoring we have quantified bioenergetics in rat insulinoma INS-1E cells representing a suitable model for studies of insulin secretion in vitro. Upon glucose addition to glucose-depleted cells, we have demonstrated a simultaneous increase in respiration and m during GSIS and shown that the endogenous state 3/state 4 respiratory ratio hyperbolically increases with glucose, approaching the maximum oxidative phosphorylation rate at maximum GSIS. Attempting to assess the basis of the "toxic" effect of fatty acids on insulin secretion, GSIS has been studied after linoleic acid addition. The linoleic acid addition diminished the observed respiration increase, m jump, and magnitude of insulin release, and reduced state 3/state 4 dependencies on glucose, which was caused by mitochondrial uncoupling. Energetic status of mitochondria, the rate of oxidative phosphorylation, is expressed not only by bioenergetic parameters measurable by classical biophysical methods (e.g. above mentioned respiration and...
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