Women with gestational diabetes mellitus (GDM) have increased risk of adverse pregnancy outcomes such as delivering Large-for-gestational-age babies, preeclampsia or birth trauma, as well as increased risk of developing type 2 diabetes (T2DM) later in life. Lifestyle management through a healthy diet and physical activity both during and after a GDM pregnancy is the first line treatment option in GDM management and for delaying the onset of T2DM. The research for this Master thesis had two main aims: firstly, to investigate the dietary intake and beliefs related to dietary intake of pregnant women with GDM in Cape Town, and whether they adhere to established dietary recommendations and secondly, to investigate the change in dietary intake, physical activity and associated factors as well as beliefs related to these lifestyle behaviours in women with GDM from pregnancy to a postpartum follow-up assessment. Methods: For the first aim a cross-sectional study was conducted on 239 pregnant women with GDM in Cape Town and for the second aim, 98 women were followed-up 3 to 15 months postpartum. Assessments included: a quantified Food Frequency Questionnaire (qFFQ), General Practice Physical Activity Questionnaire (GPPAQ) and beliefs relating to specific dietary components were assessed using the Theory of Planned Behaviour (TPB). Results: At baseline, the majority of the sample had inadequate intakes of vitamin D (87.4%), folate (96.5%) and iron (91.3%), and the dietary intake of these women was not optimal and fell short in meeting several nutritional guidelines for pregnant women with hyperglycaemia. At follow-up, the dietary changes made during pregnancy were not maintained postpartum. Fruit and vegetable intake (F&V) fell short of the recommended 400g intake at both baseline and follow-up. The intake of carbohydrates, added sugar, table sugar, sugar sweetened beverages (SSBs), pulses and energy-dense foods increased significantly from pregnancy to postpartum. In conclusion, women with prior GDM fail to maintain the dietary changes made during pregnancy. These women being at risk for the development of T2DM would benefit from interventions supporting behaviour change towards a healthier lifestyle in pregnancy and continued in the postpartum period.

Bovine leukaemia virus (BLV) and lumpy skin disease virus (LSDV) are endemic to Africa and cause significant economic losses to the beef and dairy industries. Vaccines are the most cost-effective and efficient way to prevent infection and outbreaks. Currently, there is no commercially available vaccine against BLV. In contrast, there are several live attenuated vaccines against LSDV. A recombinant LSDV which could protect cattle against both LSDV and BLV would be of great benefit to the African continent. This Master’s degree project involved three objectives. Firstly, the genetic variabilities and phylogenetic relationships of eight South African BLV isolates with other BLV strains from different geographical regions worldwide with known genotypes were determined. The BLV full-length envelope (env) and gag genes were successfully sequenced from total DNA extracted from the blood of BLV-infected cattle from a single herd. The analyses indicated that the seven of the South African isolates characterised in this study belonged to genotype 4 and the eighth to genotype 1. Furthermore, amino acid substitutions in the BLV Env and Gag sequences unique to the South African isolates were identified. Secondly, the activity of five selected poxvirus promoters in cells infected with LSDV was assessed by the detection of transient expression of an enhanced green fluorescent protein (eGFP) reporter gene driven by the poxvirus promoters. The promoters tested were a modified early fowlpox virus promoter (pmFP), an early-late promoter of a 7.5 kilodalton polypeptide gene of vaccinia virus (VACV) (p7.5), a synthetic early-late promoter of VACV (pS), a modified early-late promoter of the H5 gene of VACV (pmH5) and a synthetic early-late optimised promoter of VACV (pLEO). The results showed that all the poxvirus promoters were functional in the LSDV-infected cells and the eGFP expression was stable over the 72-hour study period. Lastly, two LSDV-vectored dual vaccines containing BLV immunogen(s) were developed and characterised. The first recombinant LSDV-vectored vaccine contained the BLV Env and Gag immunogens and the second recombinant LSDV-vectored vaccine contained the BLV Env immunogen alone. The presence of the BLV env gene in the recombinant LSDV vaccine was confirmed by polymerase chain reaction (PCR) and the BLV env sequence was confirmed by Sanger sequencing. Furthermore, BLV Env and Gag protein expression were confirmed by immunofluorescent staining and Western blotting, respectively. Future work will involve further purification of the recombinant viruses, confirmation of the production of BLV Gag virus-like particles and the preparation of high titre stocks of the vaccines to test in cattle.

Includes bibliographical references (leaves 83-89). / The corneal endothelium (CE), a mesenchyme-derived tissue, is a monolayer of squamous cells on the inner corneal surface. In Foxc1-1• mice, the CE fails to form. The understanding of the cause of this defect has implications for the study of human eye disorders that are related to FOXC1 mutations. To understand the basis of CE defects in Foxc1-1- mice, an analysis of normal CE development was performed using scanning electron microscopy. Results showed that in normal mice the transformation from mesenchyme to endothelium was initiated at embryonic day (E) 12.5 and was characterised by a change from stellate to cobblestone shape and the formation of junctions. In FoxcN- mice, the process was initiated but a cobblestone shape not attained. The expression of adherens (N-cadherin) and tight junction (Z0-1) proteins was investigated by immunoflouresence microscopy. In the normal embryo, the expression of N-cadherin was initially in cytoplasmic vesicles and later at the cell membranes. ZO-l was first detected at the cell peripheries at E13.5. In Foxct-I- mice, N-cadherin peripheral bands failed to form. ZO-l was not expressed. These results suggest that the failure to form a monolayered CE in Foxc1 mice is due to incomplete mesenchyme-endothelial conversion. Junction formation was further investigated in vitro. N-cadherin was cytoplasmic in pre-confluent cells and at cell edges in confluent cells. ZO-l was not detected. These results suggest that in vitro, these cells are either unable to form tight junctions or the culture medium does not contain the appropriate signalling molecules.

Includes bibliographical references (leaves 130-173). / Metabolic remodeling is thought to be an important contributor towards the development of various cardiac pathophysiologic conditions. Therefore, studies attempting to delineate undenying mechanisms driving cardiac metabolic remodeling represent an important initiative toward the development of novel therapeutic interventions. To further investigate the role of metabolic substrate switches in the heart, we focused on a pivotal, rate-limiting step of cardiac fatty acid metabolism i.e. an upstream modulator of long-chain fatty acid importation into the mitochondrion. In the heart, long-chain fatty acids are transported into the mitochondrion by the rate-limiting enzyme, carnitine palmitoyl transferase 1 (CPT1). CPT1 is potently inhibited by malonyl-CoA, the product of the acetylCoA carboxylation reaction that is catalyzed by acetyl-CoA carboxylase (ACC). Recent studies have demonstrated that metabolic fuels such as fatty acids and glucose can function as signaling ligands, directing transcriptional regulation of numerous metabolic genes. However, transcriptional mechanisms directing the gene expression of the cardiac isoform of acetyl-CoA carboxylase (ACCβ) are less well understood. Previously, four E-box (CANNTG) sequence motifs were identified on the human ACCβ promoter. Since E-boxes act as binding sites for upstream stimulatory factors (US F), putative glucose-responsive transcriptional modulators, we hypothesized that ACCβ is induced by USF1 in a glucosedependent manner. To investigate this, we began by acutely fasting and subsequently refeeding Balb/C mice with a carbohydrate-enriched diet. Here, high carbohydrate feeding resulted in elevated systemic glucose levels associated with increased cardiac ACCβ gene and protein expression. To further explore these interesting findings, we tranSiently cotransfected neonatal card iom yocytes , H9C2 myoblasts, CV-1 fibroblasts and HepG2 hepatocytes with the full-length and deletion constructs of the human ACC[3 gene promoter together with a putative activator and repressor expression vector, respectively: a) USF1 (glucose-responsive transcription factor) - the rationale that it should elevate ACCβ gene promoter activity in accordance with the glucose-fatty acid cycle, and b) nuclear respiratory factor 1(NRF1) - the hypothesis being that this mitochondrial biogenesis and β-oxidation enhancing modulator would be expected to attenuate ACCβ promoter activity in order to increase fatty acid oxidation capacity.

Includes abstract. / Includes bibliographical references (leaves 226-256). / Obesity is a worldwide problem and is a factor in the pathogenesis of the metabolic syndrome and kidney disease through the development of obesity-related hypertension and neurohormonal mechanisms that include the action of leptin. As there appear to be no focussed studies that have looked at the association of the LEP gene with kidney disease phenotypes or cardiovascular disease markers like hypertension, the metabolic syndrome and obesity, and especially so in native black Africans, this study sought to establish an association between the obesity gene (LEP) and kidney disease phenotypes (independent of diabetes and hypertension) in a homogenous black African population.

With the advent of effective antiretroviral therapy (ART), human immunodeficiency virus (HIV) is now a chronic disease. With increasing survival, cardiovascular disease (CVD) in people living with HIV is increasing in the ART era, with a changing epidemiology that is now largely characterised by diastolic dysfunction. Our hypothesis was that ART would be associated with regression of myocardial fibrosis in HIV. Myocardial fibrosis is associated with an unfavourable outcome in many different clinical settings. In this study, we used cardiovascular magnetic resonance (CMR) measurements of extracellular volume fraction (ECV) and tissue characterisation to assess diffuse myocardial fibrosis and determine the effect of ART use on diffuse myocardial fibrosis in HIV infected individuals on ART compared to untreated HIV infected persons. Forty-four asymptomatic individuals with no known CVD who were HIV infected were included and classified into two groups: 25 on ART for >1 year (60% male, mean age 40 ± 9 years) and 19 ART-naïve (37% male; mean age 36 ± 8 years). All patients underwent CMR on a 3T Siemens Skyra scanner. Imaging included cine, T2 weighted (STIR), native T1 and T2 mapping, late gadolinium imaging (LGE) and ECV imaging. HIV infected patients not on ART had a median time from diagnosis to entry in the study of 9 months. Treated patients had been stable on ART for over 12 months. There was no difference in left ventricular volumes, mass and function between treated and untreated HIV-infected patients. We found that, while elevated in both groups, the native T1 values were lower in the treated group compared to untreated patients. Again, while elevated in both groups, the ECV was slightly lower in the treated group, this did not reach statistical significance. There was no correlation between native T1 value or ECV with either disease duration or nadir CD4 count. There was no difference in left ventricular volumes, mass and function between treated and untreated HIV-infected patients. We found that, while elevated in both groups, the native T1 values were lower in the treated group compared to untreated patients. Again, while elevated in both groups, the ECV was slightly lower in the treated group, this did not reach statistical significance. There was no correlation between native or ECV with either disease duration or nadir CD4 count. We conclude that in patients with HIV, diffuse myocardial fibrosis provides valuable insights into the pathophysiology of HIV associated CVD and mechanism of diastolic dysfunction. Importantly, in this study, with a short lead period on ART, ART was associated with regression of diffuse myocardial fibrosis, as assessed by native T1, but not by ECV. Larger prospective studies are needed with longer follow-up to assess the role of CMR in both risk stratification and in tracking disease progression in HIV.

Includes bibliographical references. / Two plants, Dicoma anomala [Sond.]; a member of the Asteraceae and Sclerocarya birrea [(A. Rich) Hoechst. subspecies caffra (Sond.) Kokwaro], a member of the Anarcadiaceae families, are used widely in Africa for a variety of traditional treatments. In this thesis it has been shown that these plants possess in vitro pharmacological activities against the malaria parasite. The extracts of D. anomala are active in vitro against Plasmodium falciparum and those of S. birrea selectively enhance the in vitro accumulation of chloroquine in resistant strains of Plasmodium falciparum.

Includes bibliographical references. / Recent studies have investigated the underlying regulatory mechanisms that may explain the cardioprotective role of PKCε. Sub-proteome analysis has identified interactions between activated PKCε and various mitochondrial proteins, which orchestrate mitochondrial homeostasis, including proteins governing mitochondrial oxidative phosphorylation, electron transfer, ion transport and control of mitochondrial permeability transition (MPT). MPT disruption is regarded as a key step in the initiation of an apoptotic cascade. However, brief pore opening may be beneficial in triggering the generation of small amounts of protective reactive oxygen species (ROS) and restoring calcium homeostasis. PKCε also interacts with adenine nucleotide translocases (ANTs), inner mitochondrial membrane proteins essential for ATP production and an integral component of the permeability transition pore. An augmented capacity to generate ATP would fundamentally enhance resilience to ischemia.

Bibliography: leaves 168-190.

Includes abstract. Includes bibliographical references (leaves 131-147).