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Functional role of Islet1 in pancreatic development
Malfatti, Jessica ; Pavlínková, Gabriela (advisor) ; Krausová, Michaela (referee)
1 Abstract Diabetes mellitus is characterized by the dysfunction and reduction of insulin-producing cells, resulting in hyperglycemia, which in long term harms the organism. For future therapy, it is crucial to understand the function of various factors participating in the differentiation and maturation of endocrine pancreatic cells. The aim of this study was to unravel the functional role of ISL1 during the development of the pancreas. ISL1 is expressed in all endocrine cells of the islets of Langerhansbut its function remains unclear, especially during early pancreatogenesis. As the global deletion of this gene is embryonically lethal, we used the tissue specific deletion of Isl1 in Neurod1 possitive cells using the Cre-loxP system. In this work we studied the effect of this deletion on the structure of islets of Langerhans, the formation of endocrine cell types and relative expression of genes during early pancreatic development. A defective achitecture of islets together with postnatal absence of α-cells was found in the Isl1 deletion mutant. Also, the expression of genes important for the specification of α-cell lineage and their subsequent function was decreased. The secondary outcome was the optimalization of a protocol for effective sorting of endocrine cells using fluorescent flow cytometry, which...
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Functional role of SOX2 in inner ear neurosensory development
Dvořáková, Martina ; Pavlínková, Gabriela (advisor) ; Rohlena, Jakub (referee) ; Machoň, Ondřej (referee)
The main functional cells of the inner ear are neurons and sensory cells that are formed from a common embryonic epithelial neurosensory domain. Discovering genes important for specification and differentiation of sensory cells and neurons in the inner ear is a crucial basis for understanding the pathophysiology of hearing loss. Some of these factors are necessary not only for the inner ear but also for the development of other neurosensory systems such as the visual and olfactory system. The aim of this work was to reveal functions of transcription factor SOX2 in inner ear development by using mouse models with different conditional deletions of Sox2 gene. Sox2 gene was deleted by cre-loxP recombination. In Isl1-cre, Sox2 CKO mutant, reduced number of hair cells differentiated only in some inner ear organs (utricle, saccule and cochlear base) and not in others (cristae and cochlear apex). Early forming inner ear neurons in the vestibular ganglion and neurons innervating the cochlear base developed in these mutants but died by apoptosis due to the lack of neurotrophic support from sensory cells. Late forming neurons in the cochlear apex never formed. In Foxg1-cre, Sox2 CKO mutant, only rudimental ear with no sensory cells was formed. The initial formation of vestibular ganglion with peripheral and...
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Molecular mechanisms in transcriptional regulation of neurosensory development
Filová, Iva ; Pavlínková, Gabriela (advisor) ; Tureček, Rostislav (referee) ; Valihrach, Lukáš (referee)
The development, maturation, and viability of inner ear neurosensory cells depend on the spatiotemporal expression of multiple transcriptional factors. Based on three mouse models [Tg(Pax2-Isl1)], Sox2CKO, and Neurod1CKO, this thesis investigates the function of three transcriptional factors ISL1, SOX2 and NEUROD1 in the neurosensory development of the inner ear. The mouse mutant [Tg(Pax2-Isl1)] carries transgenic sequence containing Isl1 gene under Pax2 regulatory sequence in its genome. ISL1 ectopic expression driven by Pax2 regulatory sequence resulted in the enlarged cochleovestibular ganglion and accelerated neurite extension in [Tg(Pax2- Isl1)] embryos. In adult mutants, we detected an early onset of age-related hearing loss correlating with the worsening function of outer hair cells. These changes were associated with the loss of medial olivocochlear efferent neuron fibers innervating outer hair cells. For the first time, we showed that the age-related hearing loss (presbyacusis) might be caused by efferent innervation defects besides hair cell loss and spiral ganglion degeneration. In addition to presbyacusis, [Tg(Pax2-Isl1)] mice suffered from hyperactivity that was diminished by the administration of picrotoxin - channel blocker for GABA receptor chloride channels. This indicates that...
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Development and function of endocrine cells of the pancreas
Hamplová, Adéla ; Pavlínková, Gabriela (advisor) ; Berková, Zuzana (referee)
Diabetes mellitus affects nearly 300 million people in the world. The development of diabetes is caused by dysfunction or by reduction of insulin-producing β-cells that are part of the endocrine pancreas. Therefore, the most critical step for understanding the pathophysiology of diabetes and for restoring lost β cells is the identification of molecular cues that specify the cellular phenotype in the pancreas. This work is based on the hypothesis that the transcription factor NEUROD1 is a key factor for the development of the pancreas and for the maintenance of endocrine tissue function. Neurod1 conditional KO mutants (Neurod1CKO) were generated using the Cre-loxP system by crossing floxed Neurod1 mice with Isl1-Cre line. Immunohistochemical analyses of the pancreas at embryonic day 17.5 and postnatal day 0 showed that the deletion of Neurod1 negatively affected the development, organization of endocrine tissue, and total mass of pancreatic endocrine cells. To better understand molecular changes, quantitative PCR was used to analyse mRNA expression in the developing pancreas at the age of embryonic day 14.5 and postnatal day 1. Genes important for the development and function of the pancreas have been selected for the study of expression changes. These analyses showed changes in expression of genes...
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Transcriptional regulation in the development of neurosensory cells in the inner ear
Vochyánová, Simona ; Pavlínková, Gabriela (advisor) ; Schierová, Michaela (referee)
To understand the pathophysiology of hearing loss, it is necessary to identify genes responsible for specification and differentiation of sensory cells and neurons from a common neurosensory progenitor. These factors include LIM-homeodomain transcription factor ISLET1, high-mobility group protein SOX2, and basic helix-loop-helix transcription factors ATOH1, NEUROG1 and NEUROD1. This study aims to map important factors in inner ear development and their interactions with specific focus on transcription factor NEUROD1 and its role in mouse neurosensory inner ear development and function. Key words: inner ear, transcriptional regulation, mouse model, targeted deletion, embryonal development, Neurod1
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Development and function of beta-cells
Hamplová, Adéla ; Pavlínková, Gabriela (advisor) ; Tlapáková, Tereza (referee)
Insulin producing β-cells are located in the endocrine pancreas. They are a part of pancreatic islets of Langerhans along with α-, β-, δ-, ε- a PP-cells producing glucagon, somatostatin, ghrelin and pancreatic polypeptide. Insulin regulates glucose uptake into cells and thus contributes to the regulation of energy metabolism. The development of β-cells as well as the development of the pancreas is a complex process. Developmental processes of proliferation, differentiation and total pancreatic organogenesis are best described in the mouse model. The developmental processes and pancreatic functions are regulated by a network of transcription factors. Pancreatic duodenal homeobox gene 1 is a transcription factor that is expressed in the precursors of endocrine, exocrine and ductal cells. Neurogenin 3 is expressed in precursors of the islets of Langerhans cells. Islet 1 regulates the formation of the islets of Langerhans as well as the pair domains of transcription factors 4 and 6, whose expression is later limited only to β-cells. Transcription factors Islet 1 and Neurod 1 regulate insulin production in β-cells. Mutations in transcription factors lead to the abnormal development and altered function of pancreatic cells, including β-cells. Diabetes mellitus is a disease resulting from defects in...
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