The cerebral cortex is one of the most complex and divergent of all biological structures and is composed of hundreds of different types of highly interconnected neurons. This complexity underlies its ability to perform exceedingly complex neural processes. One of the most important questions in developmental neurobiology is how such a vast degree of diversity and specificity is achieved during embryogenesis. Furthermore, understanding the cellular and genetic basis of cortical development may yield insights into the mechanisms underlying human disorders such as mental retardation, autism, epilepsies and brain tumors. <p>During this Phd-project, we set out to identify novel transcription factors involved in cortical neurogenesis. Therefore, we initially took advantage of a model of in vitro embryonic stem cell (ESC)-derived corticogenesis that was previously established in the lab (Gaspard et al. 2008) and from several previously generated ESC lines that allow overexpression of specific transcription factors potentially involved in corticogenesis (van den Ameele et al. 2012). <p>Among the genes tested, Bcl6, a B-cell lymphoma oncogene known to be expressed during cortical development but without well-characterized function in this context, displayed a strong proneurogenic activity and thus became the main focus of this thesis. <p><p>During neurogenesis, neural stem/progenitor cells (NPCs) undergo an irreversible fate transition to become neurons. The Notch pathway is well known to be important for this process, and repression of Notch-dependent Hes genes is essential for triggering differentiation. However, Notch signalling often remains active throughout neuronal differentiation, implying a change in the transcriptional responsiveness to Notch during the neurogenic transition.<p>We showed that Bcl6 starts to be expressed specifically during the transition from progenitors to postmitotic neurons and is required for proper neurogenesis of the mouse cerebral cortex. Bcl6 promotes this neurogenic conversion by switching the composition of Notch-dependent transcriptional complexes at the Hes5 promoter. Bcl6 triggers exclusion of the co-activator Mastermind-like 1 and recruitment of the NAD+-dependent deacetylase Sirt1, which we showed to be required for Bcl6-dependent neurogenesis in vitro. The resulting epigenetic silencing of Hes5 leads to neuronal differentiation despite active Notch signalling. These findings thus suggest a role for Bcl6 as a novel proneurogenic factor and uncover Notch-Bcl6-Sirt1 interactions that may affect other aspects of physiology and disease (Tiberi et al. 2012a). <p><p>A subsequent yet unpublished part of this Phd-project focused on unraveling roles for Bcl6 in regionalization of the cerebral cortex. In all mammals, the three major areas of the neocortex are the motor, somatosensory and visual areas, each subdivided in secondary domains and complemented with species-specific additional areas. All these domains comprise of neurons with different functionality, molecular profiles, electrical activity and connectivity. Spatial patterning of the cortex is mainly under the control of diffusible molecules produced by organizing centers, but is also regulated by intrinsic, cell-autonomous programs (Tiberi et al. 2012b). <p>Since Bcl6 expression is confined to frontal and parietal regions of the developing cerebral cortex and remains high in postmitotic neurons, also after completion of neurogenesis, we hypothesized it would be involved in acquisition of motor and somatosensory identity. As expected from the neurogenesis defect in these regions, we observed a trend towards a reduced size of the frontal areas in the Bcl6 mutant cortex. Preliminary data from cDNA microarray profiling after gain- and loss-of-function of Bcl6 and from in situ hybridization on mouse cortex however do not show dramatic changes in molecular markers of different cortical areas. Similarly, the coarse-grained pattern of thalamocortical and efferent projections of motor and somatosensory neurons appears to be spared. These preliminary findings thus suggest that Bcl6 is not strictly required for proper acquisition of motor and somatosensory areal identity. / Doctorat en Sciences médicales / info:eu-repo/semantics/nonPublished

The aim of this dissertation is to examine and analyse the judicial framework with regard to embryonic stem cell research and cloning in South Africa. The examination is conducted within the framework of the South African and United Kingdom's legal systems. Focus is placed on aspects of medical law, human rights law as envisaged in the Constitution of the Republic of South Africa, and the law of persons. The specific focus of this dissertation is to examine the intense debate on the moral and legal status of the embryo and fetus in South Africa. A comparative study is undertaken, with the United Kingdom as a background against which recommendations for the South African framework are made. The study firstly provides a clinical overview of stem cell research and cloning. Secondly, the concept of life, in particular human life; the protection of the embryo and fetus under the constitutional guarantee of the right to life, among other constitutionally protected rights, are examined. In this context, the most important finding is that although the fetus is not a bearer of constitutional rights the state has a constitutional duty to protect fetal life in terms of an objective value system. Thereby, the state is permitted to regulate abortion, fetal tissue research, and embryo research to protect fetal life. In particular, the aim of this dissertation is to present a critical summary of the major debates and policy responses relating to embryonic stem cell research and cloning techniques, drawing attention to some of the challenges posed by conflicting moral values in an era of global scientific endeavour, and to provide an analysis of the key ethical and regulatory implications for stem cell therapy. The most important findings are that current South African legislation remains fragmented and ineffective in the manner in which embryonic stem cell research and cloning are regulated. This finding leads to a summary of recommendations, which attempts to provide specific remedies in order to adapt the current regulatory framework in South Africa. / Dissertation (LLM (Public Law))--University of Pretoria, 2007. / Public Law / LLM / unrestricted

Au cours du développement précoce de l'embryon, une importante vague de reprogrammation épigénétique efface et rétablit les profils de méthylation d’ADN (metADN) à travers le génome. Cependant, des régions spécifiques telles que les gènes à empreinte doivent échapper à cette vague de reprogrammation et maintenir leurs profils de metADN précis par l’activité constante de l’enzyme DNMT1 (ADN méthyltransférase 1) pour assurer le bon développement embryonnaire. En utilisant un modèle de cellules souches embryonnaires (mES) de souris avec une répression inductible de Dnmt1 (Dnmt1tet/tet), nous avons précédemment montré que la perte temporaire de Dnmt1 déclenche la perte permanente des profils de metADN sur les régions à empreinte et régions similaires, ainsi que sur d'autres régions du génome. Nous ne comprenons toujours pas pourquoi certaines séquences génomiques sont incapables de rétablir leurs profils de metADN normaux après la ré-expression de Dnmt1, et comment d'autres marques épigénétiques (e.g. les modifications des histones) sont altérées. Notre hypothèse est qu’un réarrangement erroné des marques d’histones aux régions promotrices, suivant une perte temporaire du maintien de la méthylation d’ADN par DNMT1, empêchera l’expression normale dans les cellules souches embryonnaires de souris. Pour ce faire, nous avons collecté des cellules mES Dnmt1tet/tet avant l'inactivation de Dnmt1, après l'inactivation de Dnmt1, puis après la réactivation complète de l'expression de Dnmt1. Nous avons ensuite utilisé la technique ChIP-Seq pour les marques d'histones (H3K4me3, H3K27me3, H3K27ac, H3K9me3, H3K4me1), celle de RRBS pour la méthylation de l'ADN et la technique de RNA-Seq pour l'expression des gènes. En définissant une liste de 18 166 promoteurs uniques, nous les avons classés en quatre catégories (Actif, Bivalent, Déplété et Réprimé). Nous montrons que l'inactivation de Dnmt1 mène à une dérégulation drastique des marques d'histones à travers les types de promoteurs. Cependant, lors de la réactivation de Dnmt1, la plupart de ces défauts ont été corrigés. Pourtant, dans l’ensemble des catégories, nous observons des promoteurs avec des dysrégulations persistantes des marques d'histones ainsi qu'un nombre significatif de gènes avec une expression différentielle. Dans l'ensemble, nos résultats montrent qu'une absence temporaire de DNMT1 a un impact plus important sur la conservation des profils des marques d'histones et l'expression des gènes que sur le maintien des profils de metADN sur les régions promotrices, dans les cellules souches embryonnaires de souris. Cela suggère que l'absence temporaire de maintien de la méthylation d’ADN déclenche une série d'événements qui conduisent à des dérégulations permanentes de marques d'histones aux promoteurs, lesquelles ne sont pas directement associés aux altérations sous-jacentes de la méthylation d’ADN dans les régions promotrices. / During early embryo development, a major epigenetic reprogramming wave erases and re-establishes DNA methylation (DNAmet) profiles across the genome. However, specific regions such as imprinting loci must escape this reprogramming wave and maintain their precise DNAmet profiles by constant DNMT1 (DNA methyltransferase 1) activity to ensure the proper development. Using a mouse embryonic stem (mES) cell model with inducible Dnmt1 repression (Dnmt1tet/tet), we previously showed that the temporary loss of Dnmt1 triggers the permanent loss of DNAmet profiles on imprinted and imprinted-like regions, as well as on other regions across the genome. We still do not understand why particular genomic sequences are unable to re-establish their normal DNAmet profiles following Dnmt1 re-expression, and how other epigenetic marks (e.g., histone modifications) are altered. Our hypothesis is that an erroneous rearrangement of histone marks on promoter regions following a temporary lack of DNAmet maintenance by DNMT1 will prevent proper gene expression in mouse embryonic stem cells. To test this, we collected mESDnmt1tet/tet cells prior to Dnmt1 inactivation, after Dnmt1 inactivation, and following complete reactivation of Dnmt1 expression. We then performed ChIP-Seq for histone marks (H3K4me3, H3K27me3, H3K27ac, H3K9me3, H3K4me1), RRBS for DNA methylation and RNA-Seq for gene expression. By defining a list of 18 166 unique promoters we categorized them in four categories (Active, Bivalent, Depleted and Repressed). We show that inactivation of Dnmt1 lead to drastic dysregulation of histone marks across types of promoters. However, upon reactivation of Dnmt1, most of these defects were rescued. Still, across categories, we observe promoters with persistent histone mark dysregulations as well as a significant number of associated genes with differential expression. Overall, our results show that a temporary lack of DNMT1 has a greater impact on the conservation of histone mark profiles and gene expression than it has on the maintenance of DNAmet profiles on promoter regions in mouse embryonic stem cells. This suggests that the temporary lack of methylation maintenance triggers a series of events that leads to the permanent dysregulation of histone marks in promoter regions, which are not directly associated with underlying DNA methylation alterations in the promoter regions.

No description available.

No description available.

No description available.

Stem cell research offers hope to many people suffering from incurable diseases such as Alzheimer's disease, diabetes, heart disease and spinal back injuries. However this poses a moral dilemma because embryos are destroyed during embryonic stem cell research. To determine whether embryonic stem cell research is morally justifiable, two views in respect of a human being were considered: i. a human has a dualistic nature in which his body and soul are two separate entities or ii. his body and soul forms a unity which can not be separated. If a human has a dualistic nature, it means that the embryo is not a human, it does not have a soul because the soul is added later to form a human. The implication of this is that it will be morally justifiable to kill an embryo during embryonic stem cell research. However if body and soul forms a unity which can not be separated, the embryo is a human which is already developing into a full grown human with several stages of development. It will thus not be morally justifiable to kill an embryo as this will violate the sixth commandment, i.e. “Thou shalt not kill.” To determine whether a human’s body and soul is an inseparable unity or whether they are two separate entities, the Pauline letters' view on the human being was investigated. The research method employed was to do a comparative literary study to highlight the different aspects of stem cell research and then exegesis was done in respect of body (σoμα / sōma); soul (ψυχὴ / psychē) and spirit (πνεῦμα / pneuma) in the Pauline letters according to the grammatical-historical method. An electronic Bible Concordance was used to determine the texts in which the above concepts appear. A semantic word analysis was also done to analyse these concepts. Then authoritative commentaries were used to check the findings. The analysis indicated that Paul refers to a human as unity in which body and soul can not be separated. The implication of this finding is that embryonic stem cell research should be dismissed because it will result in the destruction of embryos. Humans will thus be killed in violation of the sixth commandment. On the other hand adult stem cell research should be encouraged because it has the potential to cure diseases which has up to now been incurable. / Thesis (MTh (Ethics))--North-West University, Potchefstroom Campus, 2013.

Stem cell research offers hope to many people suffering from incurable diseases such as Alzheimer's disease, diabetes, heart disease and spinal back injuries. However this poses a moral dilemma because embryos are destroyed during embryonic stem cell research. To determine whether embryonic stem cell research is morally justifiable, two views in respect of a human being were considered: i. a human has a dualistic nature in which his body and soul are two separate entities or ii. his body and soul forms a unity which can not be separated. If a human has a dualistic nature, it means that the embryo is not a human, it does not have a soul because the soul is added later to form a human. The implication of this is that it will be morally justifiable to kill an embryo during embryonic stem cell research. However if body and soul forms a unity which can not be separated, the embryo is a human which is already developing into a full grown human with several stages of development. It will thus not be morally justifiable to kill an embryo as this will violate the sixth commandment, i.e. “Thou shalt not kill.” To determine whether a human’s body and soul is an inseparable unity or whether they are two separate entities, the Pauline letters' view on the human being was investigated. The research method employed was to do a comparative literary study to highlight the different aspects of stem cell research and then exegesis was done in respect of body (σoμα / sōma); soul (ψυχὴ / psychē) and spirit (πνεῦμα / pneuma) in the Pauline letters according to the grammatical-historical method. An electronic Bible Concordance was used to determine the texts in which the above concepts appear. A semantic word analysis was also done to analyse these concepts. Then authoritative commentaries were used to check the findings. The analysis indicated that Paul refers to a human as unity in which body and soul can not be separated. The implication of this finding is that embryonic stem cell research should be dismissed because it will result in the destruction of embryos. Humans will thus be killed in violation of the sixth commandment. On the other hand adult stem cell research should be encouraged because it has the potential to cure diseases which has up to now been incurable. / Thesis (MTh (Ethics))--North-West University, Potchefstroom Campus, 2013.