Pluripotency is defined as the ability of a cell to give rise to all the cell types of the adult organism. In vivo this property is possessed transiently by the cells of the epiblast in the developing embryo but it can be maintained indefinitely by deriving embryonic stem (ES) cells. How the pluripotent state is established in the cells of the early embryo and how it is ‘captured’ and maintained in the form of ES cells is a fascinating question for biology with practical implications. It is hoped that ES cells will be of use in biomedical research and cell replacement therapy. Our understanding of their biology and our ability to manipulate the cells in vitro will be of great importance if these hopes are to be realised. The starting point for the work presented in this thesis was the development of a novel culture condition for the derivation and maintenance of mouse ES cells (Q-L. Ying and J. Nichols). The media is formed by the addition of three small molecule inhibitors to a previously described serum-free media, N2B27, and is termed 3i (three inhibitors).The inhibitors are SU5402, PD184352 and CHIRON99021, and they inhibit the FGF receptor, mitogen activated protein/extracellular signalregulated kinase (ERK) kinase (MEK), and glycogen synthase kinase 3 (GSK3) respectively. I attempt to further our understanding of pluripotency and self-renewal in ES cells by genetic and biochemical examination of ES cells cultured in 3i. Analysis of intracellular signalling pathways together with descriptions of genetic mutants for the targets of the inhibitors validates the mode of action and the specificity of the three inhibitors. Self-renewal of mouse ES cells is considered dependent on activation of STAT3 through provision of the cytokine leukaemia inhibitory factor (LIF). I demonstrate unequivocally that this pathway is not required for self-renewal in 3i by characterising Stat3-null ES cells. Further experiments reveal that preventing activation of ERK downstream of the growth factor FGF4, produced by the ES cells themselves, is key to preventing differentiation. Pleiotropic effects of GSK3 inhibition are observed and candidate GSK3 targets with known or predicted effects on self-renewal are investigated as potential downstream effectors. I propose that activation of canonical Wnt signalling, together with a global derepression of biosynthetic capacity, mediate the pro-self-renewal effects of GSK3 inhibition. The description of culture conditions that function independently of signalling pathways previously thought essential for self-renewal provides fresh insight into the nature of ES cell self-renewal and the relationship of ES cells to the pluripotent cells of the developing embryo. There are practical implications for ES cell biology as there is reason to hope that the new conditions will translate more readily to other mammalian species to facilitate the derivation of ES cells and will provide an optimal platform for differentiation of ES cells into somatic cell types of interest.

This dissertation presents three separate studies designed to provide structure and evidence-based insight into the impact of religious beliefs on public perceptions and U.S. government policies regarding embryonic stem cell research. First, a systematic literature review of nine (n=9) empirical studies that examined individuals' religious beliefs and perceptions/utilization of genetic technologies/services will be presented. Based on the finding from the review, there was an equal balance between studies that found that religion was a factor positively affecting intention to submit to genetic testing and those that illustrated a negative association. Secondly, a qualitative examination of college students' from various racial/ethnic and religious backgrounds exploring the definition, interpretation, and conceptualization of the influence of religious beliefs on perceptions regarding embryonic stem cell research will be offered. Employing an emergent design, the data collection process encompassed thirty-seven in-depth interviews. The majority of participants in this study believed that ESCR should be conducted and federally funding in the United States, regardless of their religious beliefs. Lastly, the findings from the analysis of congressional records from the U.S. Congress for areas of convergence and divergence between discussions, voting, and legislation regarding stem cell research with the official stances of the major religious groups in the United States accessing the influence of religious rhetoric on political discourse regarding embryonic stem cell research will be discussed. Findings from this study suggest that religious rhetoric has a substantial influence on political rhetoric regarding ESCR.

Human haematopoiesis resembles a complex hierarchy, however most intermediate stages are only poorly defined. Efforts to characterise human progenitors have been inconsistent and failed to integrate previous knowledge. Furthermore, characterisation of normal progenitors has important implications in acute myeloid leukaemia (AML) biology. We previously established that leukaemic stem cells (LSCs) resemble the immunophenotypic progenitor compartments more closely than the stem cell fraction. Therefore, I set out to characterise human stem and progenitor cells (HSCPs) on phenotypic, molecular and functional level to complete the picture of human haematopoiesis. I purified HSPCs based on their immunophenotype from adult bone marrow (BM) and umbilical cord blood (CB) to investigate steady state and neonatal haematopoiesis. To define differentiation potentials, HSPCs were subjected to functional in vitro assays on bulk and clonal level. Limit dilution assays were used to determine the frequency of cells with multiple differentiation potentials. RNA sequencing revealed underlying lineage priming and specific gene expression signatures. I successfully characterized the incompletely defined Lin^-CD34⁺CD38^-CD45RA⁺ fraction in BM and CB, containing a CD10^lo lymphoid-primed multipotent progenitor (LMPP) with T cell, B cell, NK cell, granulocytic and monocytic differentiation potential, and succeeded in placing it in the haematopoietic hierarchy with relation to similar lympho-myeloid progenitors defined by other groups. This research lays the foundation to characterise early human progenitors with a comprehensive toolkit on a phenotypic, molecular and functional level. Findings from this thesis might provide knowledge about potential targets in LSCs.

This thesis analyses the nature, rationale, and implementation of United Kingdom and Australian regulation of research involving human embryos and cloning using legal materials, other documents and qualitative interviews with researchers, practitioners and regulators. It considers how law-makers have decided upon what to regulate and where to draw the line between permissible and prohibited activities, and the type of regulatory design strategies and enforcement approaches adopted in each jurisdiction (the ‘how to regulate’ question). It is argued that both jurisdictions have effectively decided upon permissible and prohibited activities as a result of thorough public consultation, research, reviews and the parliamentary process, and have appropriately balanced competing rationales for regulation. However, the type of regulation used in relation to those who are licensed to research in this area is unsuitable due to an over-emphasis on deterrence and the authoritarian approach taken by the regulatory bureaucracies. The central thesis is that a responsive regulatory system for licence-holders should be adopted. It is proposed that such a system would maintain the top level ‘command and control’ design strategies and deterrence approaches present in the current regulatory systems for breaches of legislation by non-licence holders and serious breaches by licence holders. However, greater use of co-regulatory design strategies and cooperative, educative and persuasive enforcement approaches should be used for regulating licensed research activities.

The focus of this thesis is the analysis of the relation of the patent law legislation and the regulatory law covering all types of stem cells. In the first part all types of stem cells are identified and a variety of issues connected to several types of stem cell is addressed in subsequent parts. Nevertheless, due to the controversy of the ES cells and in particular the human ones, the major part of the thesis discuses and analyses a regulation and case law relating to human ES stem cells. In order to analyse the relationship, the thesis focuses on the prerequisites in the morality clause of the patent law and identifies basic types of regulatory systems. Other conditions of the patentability are therefore not discussed by the thesis and the third part covering patent law focuses exclusively on the role of morality clause and the question of a scope of subject matter to be evaluated in respect to inventions relating to stem cells. In order to cover and analyse the relationship, the author finds crucial to analyse the morality clause in Article 53 (a) EPC, Rule 28 EPC and Article 6 Biotech Directive to get a more comprehensive understanding of the topic and therefore the harmonized legislation of patent law morality exclusions in Europe is the key part of the thesis. Conversely, as the regulatory legislation is harmonized at neither international level nor European one, a brief overview of national regulatory systems is presented in the part 4 of the thesis. However, due to wide range of approaches differing significantly among the countries only common characteristics of basic approaches are presented without the necessity to cover national legislations in details. An exception is made in respect to the regulatory legislation of the Czech Republic which is presented as a model example of one the approaches in order to provide a practical example of the relation of the patent law and the regulatory law.

Influenza is an important respiratory pathogen that infects several million people each year. Currently available flu vaccines have to be updated regularly in order to be effective as the virus changes its composition by antigenic drift and shift. Most of the antibody response generated by these vaccines is strain specific as it is directed against the head domain (HA1) of HA. The HA2 subunit of hemagglutinin is highly conserved and immunogens designed from this subunit are likely to provide protection against multiple strains of the virus. However, expression of HA2 alone in the absence of HA1 resulted in a protein that took up the low pH conformation of HA. Our goal was to design immunogens from HA2 that would fold into the neutral pH form. Sequence analysis of a large number of HA protein sequences was carried out to identify conserved and exposed regions on HA. Several peptide and protein constructs were designed from the stem region of HA. These proteins were expressed in bacteria and purified proteins were used to immunize mice. Immunized mice were challenged with a lethal dose of virus to test for efficacy of the immunogen. Using this approach, stem domain constructs of HA were successfully designed and shown to take up the neutral pH form. These immunogens were also shown to be capable of providing broad range protection. Residues involved in the low pH induced conformational change of HA were identified from studies on HA2 derived peptides.

This thesis addresses the puzzle of variation in first-generation regulatory policies for controversial science and technology, as demonstrated in the cases of agricultural genetically modified organisms (GMOs) and human embryonic stem cell research in the United Kingdom and the United States. Why did policy outcomes vary in each technology case? This study answers this question by placing greater emphasis on institutional factors. Although works within institutional analysis, bureaucracy and regulation literatures make significant progress in revealing how existing institutions can shape outcomes, how far one can characterize bureaucratic behavior and whether interest groups capture regulation, they nevertheless create an opening for research that: describes a mechanism for path dependence to explain variation in policies; shows the degree to which bureaucratic behaviors can influence outcomes; and, highlights instances in which regulatory officials hold power. This thesis makes an original contribution by providing new historical details relating to these cases, and by providing an extensive elaboration of Pierson’s criteria for increasing returns and a so-called secondary test of path dependence to explain outcomes. The study recounts the biography of key policy documents in each case by tracing the process of decision-making through government and archival sources, secondary literature and more than 40 elite interviews. In doing so, it details the activities of key governmental bodies within the European Union, UK and US. Moreover, it shows how the Coordinated Framework (1986) and Human Fertilisation and Embryology Act 1990 framework represented decision-making structures which triggered changes in actors and interests and shaped permissive outcomes for GMOs and stem cell research in the US and UK, respectively. Furthermore, lack of comparable structures may help account for restrictive policies for GMOs in Europe and the UK, and for stem cell research in the US.

Thesis (MA (Journalism))--University of Stellenbosch, 2006. / Moral questions arising from advances in science and technology are proliferating exponentially. Much controversy surrounds the ways in which biotechnology is used to eradicate a vast range of diseases and injuries. Stem cell research is one such way. Throughout the world stem cell research has been met with varying responses that range from opposition and criticism to approval and advocacy. As a result, it has attracted significant attention from the news media. The media have been accused of bias by focusing only on the controversial aspects of the research as opposed to reporting fully and fairly on the remarkable scientific advances. In this study I look at the patterns of media attention paid to stem cell research in the international weekly magazine Time between November 1998 and September 2005 inclusive. Contrary to the results expected on the basis of my literature study which pointed out the notion that the media tend to focus on sensational news more than non-controversial issues, I found that Time did a fair job in reporting on the scientific aspects of stem cell research. The percentage content of articles by year, focusing on scientific information of stem cells, dominated other news frames. The two years following the 2000 and 2004 American presidential elections, are however marked by the dominance of policy frames. This study found that Time covered controversial issues like embryonic stem cell research, public funding debates and political policy development in direct relation to their rise and fall on the political agenda in the United States.

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.