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The motor and cognitive consequences of inborn cerebellar degeneration and some possibilities how to influence it
Křížková, Alena ; Vožeh, František (advisor) ; Pokorný, Jaroslav (referee) ; Mareš, Jan (referee)
The motor and cognitive consequences of inborn cerebellar degeneration and some possibilities how to influence it The cerebellum is traditionally considered to be a coordinator of movements and equilibrium and a regulator of muscular tone. Participation of the cerebellum on the motor learning has been discovered later. Besides that, there is increasing experimental and clinical evidence supporting the idea of an important role of the cerebellum in common cognitive functions. Increasing number of experimental works - especially those on animals - demonstrate an effect of the the cerebellum on the memory and different types of learning. A suitable model to study the cerebellar degeneration are Lurcher mutant mice. During their lives, there is a progressive loss of Purkinje cells followed by a loss of granular cells and inferior oliva neurons. We concentrated on the following problems in this work: - development of motor learning and topical motor skills during the first month of life - positive and negative changes in NO synthesis and subsequent effect on discriminative learning - effect of administration of MK-801(non-competitive antagonist of NMDA receptors) on discriminative learning of Lurcher mutants and wild mice derived from two strains (C3H and C57B1/7). Lurcher mutants and wild mice were put in motor...
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Abilities and importance long-term cultivation embryonic cells from different species of animals
Houdek, Zbyněk ; Vožeh, František (advisor) ; Bártová, Eva (referee) ; Pokorný, Jaroslav (referee)
Embryonic cells inside the blastocyst, which divide and differentiate into cells of all tissues of organism, are called embryonic stem (ES) cells. Using the ES cells for transplantations is one the potential ways in regenerative medicine. The present study deals with two different lines of mouse pluripotent cells: embryonic carcinoma (EC) cells line P19 and ES cells line D3. The in vitro cultivation was performed in basal cultivation medium with fetal bovine serum (FBS) and in addition of leukemia inhibitory factor (LIF) in case of ES cells. The neurodifferentiation was induced by medium without FBS and without LIF in case of ES cells and supported by retinoic acid in case of P19 cells and embryoid bodies cultivation in case of ES cells. Pluripotency and neurodifferentiation of the cells were confirmed by presence of molecular markers of pluripotency and neurodifferentiation in both lines in nondifferentiated and differentiated states. Mouse EC P19 cells were transfected by gene for green fluorescent protein (GFP) and used for transplantation in cerebellum of wild type (WT) and Lurcher (Lc) mutant mice. Nondifferentiated cells (P19) and neuroprogenitors (NPG) derived from these cells were used. The study followed the survival, morphology and localization of the grafts. The survival of both types of GFP...
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Cendelín, Jan ; Vožeh, František (advisor) ; Mareš, Jan (referee) ; Pokorný, Jaroslav (referee)
Lurcher mutant mice represent a model of olivocerebellar degeneration. They are heterozygots carrying a mutation in the glutamate receptor 52-subunit gene. The receptor malfunction causes excitotoxic apoptosis of Purkinje cells, extinction of which leads to secondary degeneration of cerebellar granule cells and inferior olive neurons. Lurcher mutants suffer from ataxia and deterioration of spatial learning and orientation ability. The aim of the work was to assess the effect of embryonic cerebellar tissue transplantation and enforced physical activity as well as the effect of their combination on spatial orientation ability in Lurcher mutant mice and to observe cerebellar graft survival in adult and young Lurcher and adult wild type mice. For the graft survival analysis adult wild type mice and young and adult Lurcher B6CBA mice were used. To study the effect of the transplantation and enforced activity on spatial orientation adult and young Lurchers of the same strain were used. The graft obtained from mouse embryos without the Lurcher mutation was applied into the cerebellar area of the host as solid pieces. Control mice were treated only with vehicle. Enforced activity was represented with repeated rotarod training. Spatial orientation was tested in the Morris maze 9 weeks after the surgery. Histological...
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Cognitive and emotional abnormalities in cerebellar mutant mice
Tůma, Jan ; Vožeh, František (advisor) ; Pokorný, Jaroslav (referee) ; Delgado García, José María (referee)
3 SUMMARY Hereditary cerebellar ataxias represent a heterogeneous group of neurodegenerative disorders. Variability of human hereditary ataxias is also reflected in animal models of cerebellar disorders. The most frequently used animal models are Lurcher and Purkinje cell degeneration (pcd) mutant mice. The main aim of this thesis was to analyze and compare the spatial and emotional behavior of these cerebellar mutants. Additional aims were to study the impact of abnormal behavior on breeding capacity in Lurcher mice and to assess the applicability of cerebellar mutants as models for experimental therapy of cerebellar degeneration. We have confirmed several behavioral impairments in both Lurcher and pcd mutant mice. Nevertheless, we have found that the manifestation of spatial behavior deficit is different in these two cerebellar mutants. Based on our findings, we propose that the deficit of spatial performance in cerebellar mutants may potentially arise from a combination of 1) cognitive disturbances, 2) sensory deficits, 3) motor impairments, and finally, 4) affective disorder. Moreover, resulting spatial behavior could also be modified by the specific effect of mutation, genetic background, and sex. We have also shown that abnormal behavior, e.g. maternal infanticide leads to decreased breeding...
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Relation of tumor genotype and phenotype to diagnosis, prognosis and prediction of colorectal cancer
Pitule, Pavel ; Králíčková, Milena (advisor) ; Vožeh, František (referee) ; Slabý, Ondřej (referee)
Colorectal cancer is one of the most common type of malignity. Despite of the existence of numerous studies focused on this carcinoma, there are still many unknown features regarding its diagnosis, treatment or prognosis. In the thesis we focused on the identification of novel prognostics markers that could be useful for the stratification of patients based on the disease outcomes. In the first study we immunohistochemically assessed expression of two proteins associated with cancer stem cells in the samples of primary colorectal cancer and matched liver metastasis. Goal of the study was to evaluate relation among expression of CD44 and CD133 and overall survival and disease free interval in our set of patients. We observed that increased ratio of CD133 positive compared to CD133 negative tumor glands resulted in longer disease free interval, finding which is opposite to the general view on the CD133 role in the cancer development. Our hypothesis is that we analyzed confined group of patients and followed a bit different goal, where we measured ratio between positive and negative glands in the view-field and not the intensity of staining as the previous studies did. Our second study was focused on the transcriptional analysis of the selected set of twelve genes using frozen samples from colorectal...
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Modulation of consequences of ischemia and hypoxia
Nohejlová, Kateryna ; Mareš, Jan (advisor) ; Šonka, Karel (referee) ; Vožeh, František (referee)
Modulation of consequences of ischemia and hypoxia Kateryna Nohejlová (maiden Deykun), MD Abstract It is known from literature that that many types of insults are accompanied by the increase of free radicals level. The models of three pathological states, which are known to be related to the changes in free radicals production, were used for the purposes of the work. These models were also chosen because the intensity of action of pathogenic factor borderlines the threshold of mainly functional CNS disturbances. It was need for the animals to survive the interventions in order use behavioral methods of assessment. For the estimation of the degree of disturbances were used tests of postural motor function and motor learning and spatial orientation. The action of free radicals was evaluated indirectly by application of their scavengers and antioxidants: tempol and melatonin, which act extracellularly as well as intracellularly. Scavengers were administered as prevention, before an insult, and as therapy, after an insult. The results were divided into three sets of experiments: 1. Focal photothrombotic ischemia of sensorimotor cortex: The performance of animals that received tempol treatment did not significantly differ from the animals subjected to plain ischemia in either of tests. Melatonin application...
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The role of whiskers in compensation of visual deficit and the influence of a neurodegenerative disorder on cross-modal compensation in a mousse model of retinal and olivocerebellar degeneration
Voller, Jaroslav ; Vožeh, František (advisor) ; Jagla, Fedor (referee) ; Mysliveček, Jaromír (referee)
Sensory deprivation in one modality can enhance the development of the remaining modalities via mechanisms of synaptic plasticity. Mice of C3H strain suffers from RD1 retinal degeneration that leads to visual impairment at weaning age. Independently on the retinal degeneration there is also present olivocerebellar degeneration caused by Lurcher mutation. This neurodegenerative disorder causes motor deficits, increased CNS excitability as well as changes in synaptic plasticity. The aim of this study was to evaluate a role of whiskers in compensation of the visual deficit and to assess the influence of the olivocerebellar degeneration on this process. To differentiate contribution of the whiskers from other mechanisms that can take part in the compensation, we investigated the effect of both chronic and acute tactile deprivation. We focused on motor skills (rotarod, beam walking test), gait control (CatWalk system), spontaneous motor activity (open field) and the CNS excitability (audiogenic epilepsy). In the seeing mice without olivocerebellar degeneration, the removal of the whiskers had no effect. In the blind animals without olivocerebellar degeneration, chronic tactile deprivation caused changes in gait and impaired the performance in motor tests. Some other compensatory mechanisms were involved but the...
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Experimental Study of Mechanisms of Neurodegeneration in Different Conditions
Purkartová, Zdeňka ; Vožeh, František (advisor) ; Pokorný, Jaroslav (referee) ; Mareš, Jan (referee)
EXPERIMENTAL STUDY OF MECHANISMS OF NEURODEGENERATION IN DIFFERENT CONDITIONS - SUMMARY MUDr. Zdeňka Purkartová The cerebellum is an important structure involved in motor coordination and many other functions including cognitive and emotional processes. Severe cerebellar diseases include a wide group of cerebellar hereditary degenerations with different types of heredity, different pathogenesis and diverse phenotypic manifestation. Many types of mouse models can be used to investigate these diseases and their therapy. These mice carry spontaneous mutations or are genetically modified. The aim of this work was to analyse the dynamics of morphological changes of cerebellar degeneration in Lurcher mice using fluorescent double staining. We also performed transplantation of embryonic cerebellar tissue suspension in adult Lurcher B6CBA mice, B6.BR mice and wild-type mice of both strains with two-month survival after surgery and transplantation of identical suspension in adult Lurcher B6CBA mice and C3H mice and wild- type mice of both these strains with six-month survival after surgery. We have confirmed these main characteristics of the degenerating Purkinje cells in Lurcher mutants: the disrupted continuity of the Purkinje cell layer, the presence of dark spots in the cell nuclei, the fragmentation of nuclei...
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Cognitive and emotional abnormalities in cerebellar mutant mice
Tůma, Jan ; Vožeh, František (advisor) ; Pokorný, Jaroslav (referee) ; Delgado García, José María (referee)
3 SUMMARY Hereditary cerebellar ataxias represent a heterogeneous group of neurodegenerative disorders. Variability of human hereditary ataxias is also reflected in animal models of cerebellar disorders. The most frequently used animal models are Lurcher and Purkinje cell degeneration (pcd) mutant mice. The main aim of this thesis was to analyze and compare the spatial and emotional behavior of these cerebellar mutants. Additional aims were to study the impact of abnormal behavior on breeding capacity in Lurcher mice and to assess the applicability of cerebellar mutants as models for experimental therapy of cerebellar degeneration. We have confirmed several behavioral impairments in both Lurcher and pcd mutant mice. Nevertheless, we have found that the manifestation of spatial behavior deficit is different in these two cerebellar mutants. Based on our findings, we propose that the deficit of spatial performance in cerebellar mutants may potentially arise from a combination of 1) cognitive disturbances, 2) sensory deficits, 3) motor impairments, and finally, 4) affective disorder. Moreover, resulting spatial behavior could also be modified by the specific effect of mutation, genetic background, and sex. We have also shown that abnormal behavior, e.g. maternal infanticide leads to decreased breeding...
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Relation of tumor genotype and phenotype to diagnosis, prognosis and prediction of colorectal cancer
Pitule, Pavel ; Králíčková, Milena (advisor) ; Vožeh, František (referee) ; Slabý, Ondřej (referee)
Colorectal cancer is one of the most common type of malignity. Despite of the existence of numerous studies focused on this carcinoma, there are still many unknown features regarding its diagnosis, treatment or prognosis. In the thesis we focused on the identification of novel prognostics markers that could be useful for the stratification of patients based on the disease outcomes. In the first study we immunohistochemically assessed expression of two proteins associated with cancer stem cells in the samples of primary colorectal cancer and matched liver metastasis. Goal of the study was to evaluate relation among expression of CD44 and CD133 and overall survival and disease free interval in our set of patients. We observed that increased ratio of CD133 positive compared to CD133 negative tumor glands resulted in longer disease free interval, finding which is opposite to the general view on the CD133 role in the cancer development. Our hypothesis is that we analyzed confined group of patients and followed a bit different goal, where we measured ratio between positive and negative glands in the view-field and not the intensity of staining as the previous studies did. Our second study was focused on the transcriptional analysis of the selected set of twelve genes using frozen samples from colorectal...
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