Dissertation (Ph.D.) -- University of Texas Southwestern Medical Center at Dallas, 2006. / Vita. Bibliography: pp. 124-137

Thesis (Ph. D.)--Virginia Commonwealth University, 2009. / Prepared for: Dept. of Microbiology and Immunology. Bibliography: leaves 267 - 296. Also available online via the Internet.

The healthy immune system is a delicate and precisely orchestrated balance between activation and suppression. It is well established that regulatory T cells (Tregs) have substantial immunosuppressive properties and play a pivotal role in maintaining this balance. Many autoimmune states have been characterized by disproportionately high numbers of T effector cells, and comparatively low numbers of regulatory T cells (Hori et al., 2003; Sakaguchi et al., 1995; Choileain et al., 2006). Furthermore, mouse models in which regulatory T cells are removed or rendered ineffective show rapid development of autoimmunity. It is therefore hypothesized that regulatory T cells are essential to the acquisition and maintenance of self-tolerance. Type 1 diabetes is an increasingly common autoimmune condition, with 30,000 new diagnoses each year (JDRF Fact Sheet). Pancreatic islet transplantation holds great promise as a potential cure for this difficult disease; however human trials have had limited success. Attempts to promote self-tolerance or maintain a physical barrier to the transplanted islets have largely failed (Groot et al., 2004). Because of this, insulin dependence normally resumes fiver years post-operation. The deleterious effects of long-term immunosuppression to promote extended islet survival are considered too great to justify this treatment. Because of their important role in promoting self-tolerance, many immunologists believe regulatory T cells are the key to developing tolerance of islet allograft tissue. Rapamycin and anti-CD154 are immunoregulatory treatments that specifically inhibit the activation of T effector cells and promote the growth of regulatory T cell populations. As regulatory T cell numbers increase, self-tolerance is established and the need for immunosuppressant drugs is eliminated. Unfortunately, treatments such as anti-CD154 and rapamycin have had limited success due to the ability of toll-like receptor (TLR) pathways to bypass such activation blockades. TLR stimulation results in a potent and direct activation that acts to bolster the immune response. This TLR activation results in the release of inflammatory cytokines, which render regulatory T cells unstable. Regulatory T cells have been shown to adopt effector phenotypes in such environments and may have pathogenic potential. This study aims to elucidate aspects of Treg plasticity that result from TLR activation. In vitro models were used to demonstrate how TLR agonists change Treg phenotypic expression. Our findings indicate that the presence of lipopolysaccharides (LPS) has a relatively significant effect on regulatory T cell phenotypes. Specifically, our findings indicate that LPS causes increased GATA3 expression in Tregs, promoting differentiation to a TH2 phenotype (p= 0.0543). Regulatory T cells were also examined for the expression of RORγt and Tbet transcription factors. Neither transcription factor was significantly expressed, indicating the absence of TH17 and TH1 phenotypes, respectively. It is also worth noting that stability of the foxp3 transcript appeared to be greater in cells treated with LPS, than in those without (p= 0.0009). In addition, this study utilized an in vivo model for tracking regulatory T cell changes after pancreatic islet transplantation. Diabetic reporter mice received pancreatic islet transplants, as well as TLR agonist to induce allograft rejection. Mice were treated with rapamycin, anti-CD154 and TLR agonist. After 12 days, regulatory and ex-regulatory T cells were harvested from the transplanted area and analyzed. This experiment is still in progress and results have yet to be determined. This study establishes proof of concept of an effective system for the study of regulatory T cell plasticity. Additional investigation must be done in order to more thoroughly understand these important cells. This study is not complete, but our progress thus far is a strong foundation for further experimentation.

A presente dissertação procura investigar o incremento da racionalidade regulatória (ou otimização regulatória) a partir da teoria das falhas de regulação. Hoje, já parece existir um consenso teórico e prático de que a regulação e seu aparato institucional as agências reguladoras constituem fenômeno irreversível. Nesse contexto, as perguntas que se colocam são as seguintes: no plano dos resultados, o Estado Regulador tem alcançado as finalidades a que se propôs? Em caso negativo, que tipo de obstáculos o tem impedido? E mais: que providências devem ser adotadas para superá-los? Responder a tais indagações depende do reconhecimento de que não apenas os mercados são imperfeitos; também a intervenção estatal na economia gera riscos e efeitos negativos. Estudar seus tipos, suas fontes e a maneira como operam é o ponto de partida para se otimizar a regulação. O trabalho propõe uma sistematização das espécies de falhas regulatórias, baseada na proposta de Cass Sunstein, mas adaptada à realidade brasileira. A exposição é precedida de explicações introdutórias sobre o conceito de regulação; as razões para se regular; e características da regulação no Estado Democrático de Direito. Tais características conformam um ideal de racionalidade regulatória, o qual é comprometido pela instauração das falhas de regulação. Reconhecer a existência dos defeitos regulatórios e conhecê-los é já um primeiro passo para se melhorar a regulação. Mas há outros encaminhamentos deveras importantes para a prevenção e correção de falhas regulatórias. Dentre eles, destaca-se um conjunto de reformas institucionais em sentido amplo e estrito, as quais envolvem o sistema de controle pelos Poderes Legislativo, Executivo e Judiciário, e mecanismos procedimentais como a análise de impacto regulatório e a elaboração de agendas regulatórias. / This dissertation seeks to investigate the increase in regulatory rationality (or regulatory optimization) based on the theory of regulatory failures. Today, there already seems to be a theoretical (and practical) consensus that regulation and its institutional apparatus (regulatory agencies) constitute an irreversible phenomenon. In this context, the following questions are posed: in terms of results, has the Regulatory State achieved its established goals? If not, what obstacles prevented it from doing so? And moreover: what measures should be adopted to overcome these obstacles? Answering these questions depends on acknowledging that not only are markets imperfect; but also that state intervention in the economy creates negative risks and effects. Studying types, sources and how they operate is the starting point for optimizing regulation. This work proposes systematizing the types of regulatory failures, based on the proposal of Cass Sunstein, but adapted to Brazilian reality. The exposition is preceded by introductory explanations on the concept of regulation; the reasons for regulating; and characteristics of regulation in the Democratic State of Law. These characteristics conform an ideal of regulatory rationality, which is compromised by the introduction of regulatory failures. Acknowledging the existence of regulatory defects and knowing them is a first step in improving regulation. But there are other very important procedures for preventing and correcting regulatory failures, such as, and indeed in particular, a set of institutional reforms in a broad sense and strict, which involves the control system by the Legislative, Executive and Judiciary; and procedural mechanisms such as regulatory impact analysis and preparation of regulatory agendas.

A presente dissertação procura investigar o incremento da racionalidade regulatória (ou otimização regulatória) a partir da teoria das falhas de regulação. Hoje, já parece existir um consenso teórico e prático de que a regulação e seu aparato institucional as agências reguladoras constituem fenômeno irreversível. Nesse contexto, as perguntas que se colocam são as seguintes: no plano dos resultados, o Estado Regulador tem alcançado as finalidades a que se propôs? Em caso negativo, que tipo de obstáculos o tem impedido? E mais: que providências devem ser adotadas para superá-los? Responder a tais indagações depende do reconhecimento de que não apenas os mercados são imperfeitos; também a intervenção estatal na economia gera riscos e efeitos negativos. Estudar seus tipos, suas fontes e a maneira como operam é o ponto de partida para se otimizar a regulação. O trabalho propõe uma sistematização das espécies de falhas regulatórias, baseada na proposta de Cass Sunstein, mas adaptada à realidade brasileira. A exposição é precedida de explicações introdutórias sobre o conceito de regulação; as razões para se regular; e características da regulação no Estado Democrático de Direito. Tais características conformam um ideal de racionalidade regulatória, o qual é comprometido pela instauração das falhas de regulação. Reconhecer a existência dos defeitos regulatórios e conhecê-los é já um primeiro passo para se melhorar a regulação. Mas há outros encaminhamentos deveras importantes para a prevenção e correção de falhas regulatórias. Dentre eles, destaca-se um conjunto de reformas institucionais em sentido amplo e estrito, as quais envolvem o sistema de controle pelos Poderes Legislativo, Executivo e Judiciário, e mecanismos procedimentais como a análise de impacto regulatório e a elaboração de agendas regulatórias. / This dissertation seeks to investigate the increase in regulatory rationality (or regulatory optimization) based on the theory of regulatory failures. Today, there already seems to be a theoretical (and practical) consensus that regulation and its institutional apparatus (regulatory agencies) constitute an irreversible phenomenon. In this context, the following questions are posed: in terms of results, has the Regulatory State achieved its established goals? If not, what obstacles prevented it from doing so? And moreover: what measures should be adopted to overcome these obstacles? Answering these questions depends on acknowledging that not only are markets imperfect; but also that state intervention in the economy creates negative risks and effects. Studying types, sources and how they operate is the starting point for optimizing regulation. This work proposes systematizing the types of regulatory failures, based on the proposal of Cass Sunstein, but adapted to Brazilian reality. The exposition is preceded by introductory explanations on the concept of regulation; the reasons for regulating; and characteristics of regulation in the Democratic State of Law. These characteristics conform an ideal of regulatory rationality, which is compromised by the introduction of regulatory failures. Acknowledging the existence of regulatory defects and knowing them is a first step in improving regulation. But there are other very important procedures for preventing and correcting regulatory failures, such as, and indeed in particular, a set of institutional reforms in a broad sense and strict, which involves the control system by the Legislative, Executive and Judiciary; and procedural mechanisms such as regulatory impact analysis and preparation of regulatory agendas.

Master of Science / Department of Psychological Sciences / Clive J. A. Fullagar / Regulatory focus has emerged as an important construct in the organizational sciences. In the past ten years more than 200 papers have been published applying regulatory focus to a wide variety of contexts ranging from marketing and persuasion to feedback and performance appraisal (Johnson et al., 2015). Despite the ubiquity of RFT’s application, only a few studies have targeted the psychometric properties of measures of regulatory focus; and the findings thus far suggest that improvement is needed. Haws (2010) evaluated five measures of regulatory focus and concluded that they differed substantially with respect to their theoretical content, and that most demonstrated unacceptably low internal consistency. Summerville & Roese (2008) drew similar conclusions in their evaluation of the Regulatory Focus Questionnaire (RFQ) and the General Regulatory Focus Measure (GRFM) and added that the two scales might actually be measuring different underlying constructs. Given the inconsistencies and problems associated with existing measures of regulatory focus, the purpose of the current research is to extend the critical evaluation of existing measures of regulatory focus and then to propose the development of a new measure based on rigorous scale development practices like those set forth in Hinkin, (1995) and Crocker & Algina, (1986). A new scale of Regulatory Focus was developed designed to measure all aspects of RFT and to test whether a two-factor or a four-factor SEM model fit the data best. The final scale consisted of 14 items. CFAs were used to test whether a two-factor or a four-factor model of regulatory focus fit the data best. Results suggested that both models fit the data equally well. However, for parsimony reasons and given that one of the latent factors of the four-factor model contained only two items (making any estimates of internal consistency difficult) the two factor model of regulatory focus was retained as the preferred model.

Tariff and non-tariff barriers to trade are measures that are put in place in a country to which an exporter wants to export to. These barriers make it difficult for a new manufacturer to export their products. These measures may be considered undesirable in the context of world trade, because they restrict the flow of goods and are detrimental to the consumer because they drive prices up. The Uruguay Round of multilateral trade negotiations significantly reduced tariff barriers to trade. During the same round, the World Trade Organisation Agreement on Technical Barriers to Trade was negotiated with a view to ensure that countries use technical regulations (non- tariff barriers) for no other purpose than to protect the health and safety of the public and the environment. Many of the developing countries have not yet been able to take full advantage of this agreement. It is often difficult and costly for exporters from developing countries to meet the technical requirements of standards and technical regulations and to provide evidence of compliance. This stems from a lack of resources available to developing countries to participate and influence the work of international standards-setting bodies serving as a basis for technical regulation. The absence of internationally recognised national infrastructure for standardisation, accreditation and metrology also prevents acceptance of African products in export markets. Various regions such as Asia and Europe have initiated technical regulatory reforms to align their technical regulations with the requirements of the World Trade Organisation requirements and to establish appropriate technical institutions. These reforms are also expected to assist member countries to gain a competitive edge in global trade ensuring increased gains from trade liberalisation initiatives which provide fair market access for goods and services. Africa appears to be lagging behind the abovementioned regions. The current technical regulatory system in Africa is still too inefficient and ineffective to position African countries competitively. The technical institutions are underdeveloped and under funded and can not support market assess and thus economic development. It is in this context that this study is conducted to explore the technical regulatory framework in Africa against the background of reforms in Asia and Europe with a view of providing public policy recommendations for the establishment of an African technical regulatory system supported by appropriate institutional capacity that may expedite economic recovery for the continent. / Thesis (PhD)--University of Pretoria, 2010. / School of Public Management and Administration (SPMA) / Unrestricted

The temporal and spatial patterning that underlies morphogenetic events is controlled by gene regulatory networks (GRNs). These operate through a combinatorial code of DNA – binding transcription factor proteins, and non – coding DNA sequences (cis-regulatory elements, or CREs), that specifically bind transcription factors and regulate nearby genes. By comparatively studying the development of different species, we can illuminate lineage – specific changes in gene regulation that account for morphological evolution. The central nervous system of vertebrates is composed of diverse neural cells that undergo highly coordinated programs of specialization, migration and differentiation during development. Approximately 20% of neurons in the cerebral cortex are GABAergic inhibitory interneurons, which release the neurotransmitter gamma-aminobutyric acid (GABA). Diseases such as autism, schizophrenia and epilepsy are associated with defects in GABAergic interneuron function. Several members of the distal-less homeobox (Dlx) transcription factor family are implicated in a GRN underlying early GABAergic interneuron development in the forebrain. I examined the role played by orthologous dlx genes in the development of GABAergic interneurons in the zebrafish forebrain. I found that when ascl1a transcription factor is down-regulated through the micro-injection of translation – blocking morpholino oligonucleotides, Dlx gene transcription is decreased in the diencephalon, but not the telencephalon. Similarly, gad1a transcription is also decreased in this region for these morphants. As gad1a encodes an enzyme necessary for the production of GABA, these genes are implicated in a cascade underlying GABAergic interneuron development in the diencephalon.

Inferring regulatory relationships between genes, including the direction and the nature of influence between them, is the foremost problem in the field of genetics. One classical approach to this problem is epistasis analysis. Broadly speaking, epistasis analysis infers the regulatory relationships between a pair of genes in a genetic pathway by considering the patterns of change in an observable trait resulting from single and double deletion of genes. More specifically, a “surprising” situation occurs when the phenotype of a double mutant has a similar, aggravating or alleviating effect compared to the phenotype resulting from the single deletion of either one of the genes. As useful as this broad approach has been, there are limits to its ability to discriminate alternative pathway structures, meaning it is not always possible to infer the relationship between the genes. Here, we explore the possibility of dynamic epistasis analysis. In addition to performing genetic perturbations, we drive a genetic pathway with a dynamic, time-varying upstream signal, where the phenotypic consequence is measured at each time step. We explore the theoretical power of dynamic epistasis analysis by conducting an identifiability analysis of Boolean models of genetic pathways, comparing static and dynamic approaches. We also explore the identifiability of individual links in the pathway. Through these evaluations, we quantify how helpful the addition of dynamics is. We believe that a dynamic input in addition to epistasis analysis is a powerful tool to discriminate between different networks. Our primary findings show that the use of a dynamic input signal alone, without genetic perturbations, appears to be very weak in comparison with the more traditional genetic approaches based on the deletion of genes. However, the combination of dynamical input with genetic perturbations is far more powerful than the classical epistasis analysis approach. In all cases, we find that even relatively simple input dynamics with gene deletions greatly increases the power of epistasis analysis to discriminate alternative network structures and to confidently identify individual links in a network. Our positive results show the potential value of dynamics in epistasis analysis.

This thesis was presented by Aaron Webber on the 4th December 2013 for the degree of Doctor of Philosophy from the University of Manchester. The title of this thesis is ‘Transcriptional co-regulation of microRNAs and protein-coding genes’. The thesis relates to gene expression regulation within humans and closely related primate species. We have investigated the binding site distributions from publically available ChIP-seq data of 117 transcription regulatory factors (TRFs) within the human genome. These were mapped to cis-regulatory regions of two major classes of genes,  20,000 genes encoding proteins and  1500 genes encoding microRNAs. MicroRNAs are short 20 - 24 nt noncoding RNAs which bind complementary regions within target mRNAs to repress translation. The complete collection of ChIP-seq binding site data is related to genomic associations between protein-coding and microRNA genes, and to the expression patterns and functions of both gene types across human tissues. We show that microRNA genes are associated with highly regulated protein-coding gene regions, and show rigorously that transcriptional regulation is greater than expected, given properties of these protein-coding genes. We find enrichment in developmental proteins among protein-coding genes hosting microRNA sequences. Novel subclasses of microRNAs are identified that lie outside of protein-coding genes yet may still be expressed from a shared promoter region with their protein-coding neighbours. We show that such microRNAs are more likely to form regulatory feedback loops with the transcriptional regulators lying in the upstream protein-coding promoter region. We show that when a microRNA and a TRF regulate one another, the TRF is more likely to sometimes function as a repressor. As in many studies, the data show that microRNAs lying downstream of particular TRFs target significantly many genes in common with these TRFs. We then demonstrate that the prevalence of such TRF/microRNA regulatory partnerships relates directly to the variation in mRNA expression across human tissues, with the least variable mRNAs having the most significant enrichment in such partnerships. This result is connected to theory describing the buffering of gene expression variation by microRNAs. Taken together, our study has demonstrated significant novel linkages between the transcriptional TRF and post-transcriptional microRNA-mediated regulatory layers. We finally consider transcriptional regulators alone, by mapping these to genes clustered on the basis of their expression patterns through time, within the context of CD4+ T cells from African green monkeys and Rhesus macaques infected with Simian immunodeficiency virus (SIV). African green monkeys maintain a functioning immune system despite never clearing the virus, while in rhesus macaques, the immune system becomes chronically stimulated leading to pathogenesis. Gene expression clusters were identified characterizing the natural and pathogenic host systems. We map transcriptional regulators to these expression clusters and demonstrate significant yet unexpected co-binding by two heterodimers (STAT1:STAT2 and BATF:IRF4) over key viral response genes. From 34 structural families of TRFs, we demonstrate that bZIPs, STATs and IRFs are the most frequently perturbed upon SIV infection. Our work therefore contributes to the characterization of both natural and pathogenic SIV infections, with longer term implications for HIV therapeutics.