|
|
Evolution of the gene regulatory network underlying the formation of the gastrula organizer
Macháčová, Simona ; Kozmikova, Iryna (advisor) ; Krylov, Vladimír (referee) ; Buchtová, Marcela (referee)
During gastrulation, the vertebrate embryo is organized from the clump of cells into a bilaterally symmetric body. This organization process is driven by the gastrula organizer. Its establishment is induced by maternal Wnt/β-catenin signaling and Nodal/Activin signaling localized in the presumptive dorsal region of the embryo. The regulative environment then triggers the expression of the organizer-specific genes which create morphogen gradients in the embryonic body and therefore give each cell positional information. However, the evolution of vertebrate organizer establishment remains vague. Here we aim to test the compatibility of the invertebrate cis-regulatory modules with the vertebrate gene regulatory network (GRN). We introduced fluorescent reporter genes under the control of the invertebrate regulatory sequence of organizer-specific genes into a vertebrate model to observe their behavior in the context of the vertebrate GRN. We found and functionally verified a 500 bp-long amphioxus sequence (an enhancer) that is necessary and sufficient to drive a correct Chordin gene expression in the gastrula organizer in zebrafish. Chordin is a prominent organizer-specific gene antagonizing Bone Morphogenetic Protein (BMP) signaling. We tested also other invertebrate genes for their compatibility with...
|
|
|
Huntington's disease modeling and stem cell therapy in spinal cord disorders and injury
Hruška-Plocháň, Marián ; Motlík, Jan (advisor) ; Bjarkam, Carsten (referee) ; Roth, Jan (referee)
Neurological disorders affect more than 14% of the population worldwide and together with traumatic brain and spinal cord injuries represent major health, public and economic burden of the society. Incidence of inherited and idiopathic neurodegenerative disorders and acute CNS injuries is growing globally while neuroscience society is being challenged by numerous unanswered questions. Therefore, research of the CNS disorders is essential. Since animal models of the CNS diseases and injuries represent the key step in the conversion of the basic research to the clinics, we focused our work on generation of new animal models and on their use in pre-clinical research. We generated and characterized transgenic minipig model of Huntington's disease (HD) which represents the only successful establishment of a transgenic model of HD in minipig which should be valuable for testing of long term safety of HD therapeutics. Next, we crossed the well characterized R6/2 mouse HD model with the gad mouse model which lacks the expression of UCHL1 which led to results that support the theory of "protective" role of mutant huntingtin aggregates and suggest that UCHL1 function(s) may be affected in HD disturbing certain branches of Ubiquitin Proteasome System. Traumatic spinal cord injury and Amyotrophic Lateral...
|
|
|
Modelovanie ochorenia a štúdium regeneračných procesov v Huntingtonovej chorobe a ALS in vivo
Hruška-Plocháň, Marián
Neurological disorders affect more than 14% of the population worldwide and together with traumatic brain and spinal cord injuries represent major health, public and economic burden of the society. Incidence of inherited and idiopathic neurodegenerative disorders and acute CNS injuries is growing globally while neuroscience society is being challenged by numerous unanswered questions. Therefore, research of the CNS disorders is essential. Since animal models of the CNS diseases and injuries represent the key step in the conversion of the basic research to the clinics, we focused our work on generation of new animal models and on their use in pre-clinical research. We generated and characterized transgenic minipig model of Huntington's disease (HD) which represents the only successful establishment of a transgenic model of HD in minipig which should be valuable for testing of long term safety of HD therapeutics. Next, we crossed the well characterized R6/2 mouse HD model with the gad mouse model which lacks the expression of UCHL1 which led to results that support the theory of "protective" role of mutant huntingtin aggregates and suggest that UCHL1 function(s) may be affected in HD disturbing certain branches of Ubiquitin Proteasome System. Traumatic spinal cord injury and Amyotrophic Lateral...
|
|
|
Development of advanced in-vivo imaging methods for non-invasive study of tumour growth dynamic
Michalčíková, Tereza ; Procházka, Jan (advisor) ; Rohlena, Jakub (referee)
Non-invasive imaging techniques, such as micro-CT or optical imaging, provide valuable information about tumour microstructure, size, volume and growth dynamics. Although histology is an approach capable of describing several of these characteristics, the invasiveness of this analysis remains a disadvantage. The main aim of this work is the methodological development of non-invasive imaging of the dynamics of tumour growth and progression. The preparation of a dual-reporter lentiviral vector enables non-invasive study of tumour growth and dissemination of metastasis. The same dual reporter will also be a part of a second vector designed as a construct for targeting mouse embryonic stem cells with aim to produce corresponding transgenic reporter mouse line. This reporter mouse line can be beneficial for future projects by providing a novel approach for studying the dynamics of tumour growth under various genetic conditions. In addition to optical imaging, this project will also include the use of micro-CT technology which, as a non-invasive approach, has the potential to provide information about the microstructure of tumour tissue in 3D that histology is not able to report.
|
|
|
Generation of large animal models using genome editing
Dvořáková, Nikola ; Ellederová, Zdeňka (advisor) ; Kašpárek, Petr (referee)
The principle of gene engineering is the intervention to the DNA of the studied organism. After the discovery of the programmed endonucleases, there has been a great expansion of this technique and it also accelerated the possibilities to create large animal models. Until recently, large animal models were very difficult to be generated. These endonucleases include zinc finger nuclease (ZFN), transcription activator like effector nuclease (TALEN) and CRISPR/Cas9. All endonucleases produce locally specific splicing in the targeted segment of the genome. This splicing is most easily corrected by the non-homologous ends joining (NHEJ), so then it is possible to create a so -called knock-out (KO) model. The second type of repair is homologous recombination (HR) using a DNA template with homologous arms. This makes it possible to create a knock-in (KI) model that cannot be created without specific endonucleases in large animal models due to the low natural HR. This work summarizes the history, technique and the use of programmed endonucleases for the creation of large animal models. These models have a great use in biomedicine, mostly in preclinical research, they are also significant in agriculture and even in the environment protection. Key words: large animal model, transgenesis, genome editing,...
|
|
|
Transgenic technologies based on transposons
Dobiášovská, Ivana ; Kozmik, Zbyněk (advisor) ; Čáp, Michal (referee)
Genenetic engineering is one of the leading technologies in biological research. Transgenesis, one of the most important genetic engineering technologies, enables to study genetic aspects of organismal systems and thus helps us to better understand to the functional characteristics of genomes. Transposons are naturally occuring mobile genetic elements, which can be used to artificially integrate transgenes into host cell genomes. Catalysis of this essential step during transgenesis makes from transposons an useful genetic tool.The aim of this work is to present eukaryotic DNA transposons that transpose in a cut-and-paste-fashion, together with particular mechanisms affecting their function, that can be used as gene delivery system.
|
|
|
Modelovanie ochorenia a štúdium regeneračných procesov v Huntingtonovej chorobe a ALS in vivo
Hruška-Plocháň, Marián
Neurological disorders affect more than 14% of the population worldwide and together with traumatic brain and spinal cord injuries represent major health, public and economic burden of the society. Incidence of inherited and idiopathic neurodegenerative disorders and acute CNS injuries is growing globally while neuroscience society is being challenged by numerous unanswered questions. Therefore, research of the CNS disorders is essential. Since animal models of the CNS diseases and injuries represent the key step in the conversion of the basic research to the clinics, we focused our work on generation of new animal models and on their use in pre-clinical research. We generated and characterized transgenic minipig model of Huntington's disease (HD) which represents the only successful establishment of a transgenic model of HD in minipig which should be valuable for testing of long term safety of HD therapeutics. Next, we crossed the well characterized R6/2 mouse HD model with the gad mouse model which lacks the expression of UCHL1 which led to results that support the theory of "protective" role of mutant huntingtin aggregates and suggest that UCHL1 function(s) may be affected in HD disturbing certain branches of Ubiquitin Proteasome System. Traumatic spinal cord injury and Amyotrophic Lateral...
|
|
|
Huntington's disease modeling and stem cell therapy in spinal cord disorders and injury
Hruška-Plocháň, Marián ; Motlík, Jan (advisor) ; Bjarkam, Carsten (referee) ; Roth, Jan (referee)
Neurological disorders affect more than 14% of the population worldwide and together with traumatic brain and spinal cord injuries represent major health, public and economic burden of the society. Incidence of inherited and idiopathic neurodegenerative disorders and acute CNS injuries is growing globally while neuroscience society is being challenged by numerous unanswered questions. Therefore, research of the CNS disorders is essential. Since animal models of the CNS diseases and injuries represent the key step in the conversion of the basic research to the clinics, we focused our work on generation of new animal models and on their use in pre-clinical research. We generated and characterized transgenic minipig model of Huntington's disease (HD) which represents the only successful establishment of a transgenic model of HD in minipig which should be valuable for testing of long term safety of HD therapeutics. Next, we crossed the well characterized R6/2 mouse HD model with the gad mouse model which lacks the expression of UCHL1 which led to results that support the theory of "protective" role of mutant huntingtin aggregates and suggest that UCHL1 function(s) may be affected in HD disturbing certain branches of Ubiquitin Proteasome System. Traumatic spinal cord injury and Amyotrophic Lateral...
|
|
|
Biomedical models of Huntington disease
Žižková, Martina ; Motlík, Jan (advisor) ; Moravec, Jan (referee)
Huntington's diease is a dominant inherited neurodegenerative disorder that is caused by an expansion of a CAG repeats within a huntingtin gene. Mutant protein causes a neuron degeneration in a brain of HD pacients which leads to a motor abnormalities and personality decay. This disease is very malign because of its late onset. An equal therapy does not exist yet, but a lot of research teams focus on designig a suitable medical treatment. It is necessary to create animal models of Huntington disease which can be used for testing the therapies. In my work I aim to summarize the animal models of HD which are used in research. A rodent model is the most common due to its low price and easy breeding. However, more important are human related large animals like sheep, pigs or non-human primates. The principal criterion of animal model is its method of creation. We can divide the models into two categories, genetic and non-genetic. The memebers of the first one are able to reproduce better expression of human Huntington disease. Generation of animal models of HD leads to better comprehension the principles of HD, and developing an equal therapy for HD pacients.
|
guest ::
