Cytokinins are plant hormones that affect the primary growth of shoots and roots. Application of the cytokinin N6-benzylaminopurine (BAP) to the shoot apical meristem of <i>Arabidopsis thaliana Landsberg erecta </i>(L.) Heynh induces aberrant flower development and a significant genetic response, and some of these phenotypes and expression patterns were carried to the next generation. Analysis of altered transcript levels with Affymetrix GeneChips® indicated significant changes in transcript levels of genes associated with shoot meristem activity, circadian rhythms, cytokinin metabolism, two-component systems, stress and defense responses, auxin regulation, ethylene and salicylic acid biosynthesis, and signal transduction. Specific genes were also mined from the data as potentially responsible for the BAP-induced aberrant floral phenotypes, increased floral organ number, buds in axils of sepals, and mosaic floral organs. Of particular note was a decrease in the transcript levels of CLAVATA1 (CLV1), a gene encoding a receptor kinase involved in organ differentiation and maintenance of shoot and floral meristems. Time course analysis by RT-PCR showed a decline and subsequent recovery of transcript levels of CLV1 and a coincident increase in WUSCHEL (WUS) transcript, consistent with the known suppression of WUS by CLV. WUS encodes a homeodomain protein associated with shoot meristem proliferation. The temporal coincidence of an increased floral organ phenotype with changes in transcript levels of CLV1 and WUS suggests that cytokinins regulate flower development by affecting the activity of genes controlling shoot meristem activity. Aberrant floral phenotypes in subsequent non-treated generations suggest epigenetic inheritance of some BAP-altered transcript patterns. Repressed expression of the majority of significant genes in the untreated T1 population suggests a mechanism of gene silencing, such as methylation, was involved in this epigenetic inheritance. Also, transcript levels of time-keeping genes, including CIRCADIAN CLOCK ASSOCIATED 1 / ELONGATED HYPOCOTYL, and associated genes with oscillating expression patterns, such as COLD-RESPONSIVE, were affected by BAP in treated plants and the subsequent generation, suggesting the capacity of cytokinins to affect the phase of the circadian clock. Hormonal regulation of heritably altered diurnal periodicity and environmental responses may provide a developmental and, therefore, evolutionary advantage to plants.

Cytokinins are plant hormones that affect the primary growth of shoots and roots. Application of the cytokinin N6-benzylaminopurine (BAP) to the shoot apical meristem of <i>Arabidopsis thaliana Landsberg erecta </i>(L.) Heynh induces aberrant flower development and a significant genetic response, and some of these phenotypes and expression patterns were carried to the next generation. Analysis of altered transcript levels with Affymetrix GeneChips® indicated significant changes in transcript levels of genes associated with shoot meristem activity, circadian rhythms, cytokinin metabolism, two-component systems, stress and defense responses, auxin regulation, ethylene and salicylic acid biosynthesis, and signal transduction. Specific genes were also mined from the data as potentially responsible for the BAP-induced aberrant floral phenotypes, increased floral organ number, buds in axils of sepals, and mosaic floral organs. Of particular note was a decrease in the transcript levels of CLAVATA1 (CLV1), a gene encoding a receptor kinase involved in organ differentiation and maintenance of shoot and floral meristems. Time course analysis by RT-PCR showed a decline and subsequent recovery of transcript levels of CLV1 and a coincident increase in WUSCHEL (WUS) transcript, consistent with the known suppression of WUS by CLV. WUS encodes a homeodomain protein associated with shoot meristem proliferation. The temporal coincidence of an increased floral organ phenotype with changes in transcript levels of CLV1 and WUS suggests that cytokinins regulate flower development by affecting the activity of genes controlling shoot meristem activity. Aberrant floral phenotypes in subsequent non-treated generations suggest epigenetic inheritance of some BAP-altered transcript patterns. Repressed expression of the majority of significant genes in the untreated T1 population suggests a mechanism of gene silencing, such as methylation, was involved in this epigenetic inheritance. Also, transcript levels of time-keeping genes, including CIRCADIAN CLOCK ASSOCIATED 1 / ELONGATED HYPOCOTYL, and associated genes with oscillating expression patterns, such as COLD-RESPONSIVE, were affected by BAP in treated plants and the subsequent generation, suggesting the capacity of cytokinins to affect the phase of the circadian clock. Hormonal regulation of heritably altered diurnal periodicity and environmental responses may provide a developmental and, therefore, evolutionary advantage to plants.

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No description available.

Bone remodelling is a complex process, which involves the coupling of bone formation to completed foci of bone resorption, the balance between these 2 processes determines if bone is lost or gained at a particular site. During bone resorption osteoclasts release growth factors sequestered in bone matrix, which are thought to initiate new bone formation. On the other hand, osteoblasts can regulate osteoclast activity through the expression of the counter-acting cytokines, RANKL and OPG. The aim of this project was to determine if factors released during bone resorption impact on the RANKL/OPG system or on osteoclasts directly to regulate bone remodelling. OPG secretion was characterized in a number of osteoblast-like cells and the osteosarcoma cell line MG-63 was chosen as a model for osteoblastic cell behaviour in vitro. EDTA bone extracts prepared from normal human cortical bone powder were used to treat MG-63 cells in vitro. The response to the extract was dependent on the purification procedure used. OPG production was inhibited by partially purified extracts prepared using hydrophobic interaction chromatography, C18 SPE. In comparison extracts prepared using size exclusion centrifugal filters stimulated OPG secretion in confluent MG-63 cells. Therefore bone matrix constituents were able to influence osteoclast activity directly and indirectly through the osteoblastic cells to produce the same response. The simplest mechanism for this co-ordinated response would be the presence of one factor in the extract that is able to influence both osteoblasts and osteoclasts. The identity of the factor responsible for the opposing effects seen in the bone matrix extracts is at the moment unknown. The work presented in this thesis clearly demonstrated that unknown growth factors present in bone matrix influence bone remodelling.

Cardiovascular disease is the leading cause of death globally. Chronic hypertension can lead to a gradual deterioration in the structure and performance of the cardiovascular system in a process described as cardiovascular remodelling. The ultimate response to this remodelling is heart failure. While cardiovascular remodelling is characterised by features such as cardiac and vascular hypertrophy, endothelial dysfunction and fibrosis; mechanisms leading to such pathologies are still unclear. However, oxidative stress, the damage caused by increased production of reactive oxygen species such as free radicals, or their reduced removal by anti-oxidants, appears to have a significant role. The major aim of this thesis is to investigate the involvement of reactive oxygen species in cardiovascular remodelling and to further investigate the potentials of three anti-oxidant approaches (a combination of alpha-lipoic acid and vitamin E, apocynin and a red-wine component, resveratrol) in preventing or reversing cardiovascular remodelling. These studies were conducted in two well-established rat models of cardiovascular remodelling including the deoxycorticosterone acetate (DOCA)-salt hypertensive rat and the ageing spontaneously hypertensive rat (SHR). Additionally, this thesis also characterises the effects of ovariectomy on the blood pressure and survival rate of female SHR. The cardiovascular structure and function in the animals have been defined using in vivo echocardiography, ex vivo isolated Langendorff heart perfusion, isolated thoracic aortic rings, histological analysis of the myocardial extracellular matrix and inflammation along with terminal organ weight measurements. Reactive oxygen species were assessed through the measurement of plasma malondialdehyde (MDA) while the anti-oxidant capacities have been assessed through measurements of plasma total anti-oxidant status (TAS) and plasma glutathione peroxidase (GPx) activity. DOCA-salt treated rats exhibited hypertension, oxidative stress and cardiovascular remodelling, evidenced by their increased left ventricular weights, excess collagen deposition in the heart and increased values for diastolic stiffness, increased plasma MDA concentrations along with impaired contraction and relaxation of the vessels. Treatment with a combination of alpha-lipoic acid and vitamin E or apocynin significantly inhibited the increases in blood pressure, left ventricular weights, cardiac stiffness, interstitial collagen deposition along with improvements in the vascular responses. The treatment of ageing male SHRs with resveratrol, although without any decrease in blood pressure, was shown to reduce left ventricular hypertrophy, fibrosis, diastolic stiffness and improve functional performance of the heart. The level of oxidative stress was also lowered in the male SHRs treated with resveratrol as evidenced by decreased plasma MDA and increased TAS. Female SHRs had a higher blood pressure and diastolic stiffness compared to their age-matched WKYs. Similar to male SHRs, treatment with resveratrol did not affect blood pressure, but attenuated the increased diastolic stiffness, in female SHRs. Furthermore, the ovariectomised SHRs were proven to be extremely hypertensive and had a significantly poorer survival rate. Overall, these studies demonstrated great potential for the alpha-lipoic acid and vitamin E combination and apocynin in the treatment of cardiovascular remodelling. However, further clinical and experimental research is essential to confirm the complete cardiovascular health benefits of resveratrol or red wine intake.

Master of Medicine / Non-melanoma skin cancer (NMSC) is the most common human cancer worldwide. Squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) make up almost all NMSC. SCC usually arises from actinic keratosis (AK) as a result of exposure to sunlight. SCC and AK provide a useful clinical model to investigate changes involved in the progression of NMSC. This project examines the expression of Brm, Brg-1, Snail 1 and Snail 2 in the progression of NMSC. Brm and Brg-1 are subunits of the SWI/SNF chromatin-remodelling complex which is involved in regulating the access of cell machinery to DNA by altering the structure of chromatin. It has been suggested that loss of this function is involved in carcinogenesis as the cell is unable to access to DNA normally in order to repair mutations or activate apoptosis. The loss of Brm or Brg-1 has been described in several human cancers. Snail 1 and Snail 2 are zinc-finger transcription factors that are known for their role in epithelial to mesenchymal transition (EMT), a process vital to embryological development. Increased expression of these factors leads to a loss of cell-cell adhesion and a migratory phenotype and has been described in some human cancers. In this project, double-label immunohistochemistry was used to determine the relative expression of these proteins in human SCC, BCC, AK and normal skin. The expression of Snail was unable to be determined due to poor specificity of the antibodies used. The expression of both Brm and Brg-1 proteins was found to be dramatically and consistently decreased in SCC and BCC when compared to normal skin and AK. This loss of Brm and Brg-1 occured as the tumour progressed from benign AK to malignant SCC. This finding suggests that the loss of either Brm or Brg-1 constitutes a key step in carcinogenesis. The results of this study identify Brm and Brg-1 as putative tumour suppressors involved in the progression of non-melanoma skin cancer from benign to malignant.

Atrial fibrillation is the most common sustained arrhythmia; however, its mechanism is not well understood. Several conditions such as valvular disease, heart failure, and hypertension predispose to atrial fibrillation. Identifying the electrophysiological substrate in these clinical conditions would yield insight into the mechanism of atrial fibrillation and aid in developing strategies to prevent or cure it. Rheumatic mitral stenosis is associated with high prevalence of atrial fibrillation. While atrial stretch itself may be adequate to explain the occurrence of atrial fibrillation in this population, it is not known if the disease process would remodel the atria so as to increase its propensity. Chapters 2 and 3 present the results of the studies evaluating the substrate for atrial fibrillation in both the left and right atria in rheumatic mitral stenosis. These studies have demonstrated extensive conduction abnormalities both regional and site specific associated with low voltage area and scar. Despite the prolonged atrial refractoriness, the propensity for atrial fibrillation was increased; lending support to the theory that structural remodelling associated with conduction abnormalities plays a greater role in the substrate predisposing to atrial fibrillation. Chapters 4 and 5 present the results of the studies evaluating the immediate effects of chronic atrial stretch reversal on the atrial electrical remodelling. These studies demonstrated that immediately after percutaneous mitral commissurotomy there was decrease in P wave duration, improvement in site specific conduction delay and conduction velocity associated with increase in the voltage. However, there was no change in atrial refractoriness. Chapter 6 studies the substrate long-term after reduction of stretch. There was further increase in conduction velocity and voltage associated with decrease in atrial refractoriness and conduction delay across the crista terminalis. These observations suggest that strategies aimed at reducing atrial stretch in different disease conditions would potentially decrease the burden or prevent atrial fibrillation. There is mounting evidence of the effect of stretch on the atria; however, the effect of stretch on the triggers of atrial fibrillation has not been evaluated before. Chapter 7 and 8 present the results of the study examining the effect of acute and chronic stretch on human pulmonary veins. Simultaneous pacing of the right ventricle and pulmonary vein induced acute stretch. The effect of chronic stretch was evaluated in patients with mitral stenosis. The atrial refractoriness was abbreviated in acute stretch while it was prolonged in the chronic form. Nevertheless, both resulted in marked pulmonary vein conduction abnormalities that were pronounced with chronic stretch and extra-stimuli. Additionally, structural remodelling was seen with chronic stretch. These abnormalities implicate stretch in the milieu for re-entry and pulmonary vein arrhythmogenesis in conditions predisposed to atrial fibrillation. In summary, this thesis has evaluated the effects of stretch on the substrate and triggers of atrial fibrillation. It provides evidence for the importance of structural changes and the associated abnormalities in conduction in predisposing to atrial fibrillation. These observations may be important in the development of tools to treat, cure and prevent atrial fibrillation. / Thesis (Ph.D.) -- University of Adelaide, School of Medicine, 2008

Abstract Cardiac hypertrophy and remodelling are mechanisms of adaptation to increased workload and acute injuries of the heart. In the long-term, these initially beneficial mechanisms become detrimental and ultimately lead to the development of heart failure. The molecular determinant of myocardial remodelling and heart failure is altered intracellular signal transduction and a modified gene expression pattern in the individual cardiomyocyte. This study was aimed at characterising the changes in mitogen-activated protein kinases (MAPKs) and their nuclear effector, GATA-4, and their functional significance and interaction in experimental models of increased cardiac workload and remodelling. To study the effects of increased cardiac workload on MAPKs and GATA-4, isolated perfused rat hearts were subjected to increased left ventricular wall stress and their activities were determined using western blot and gel mobility shift assays. Left ventricular wall stress rapidly activated the DNA binding of GATA-4, and this activation was abolished in the presence of endothelin-1 (ET-1) and angiotensin II receptor antagonists. Furthermore, the activation of GATA-4 DNA binding was significantly attenuated by p38 MAPK and extracellular signal regulated kinase (ERK) inhibition. To gain further insights into the role of p38 MAPK as a regulator of cardiac transcription factors, gene expression and remodelling, a gene transfer protocol of increased p38 MAPK activity was established. Direct adenovirus-mediated gene transfer of wild-type p38α and constitutively active upstream kinase mitogen-activated kinase kinase 3b (MKK3b) selectively increased p38 MAPK activity in the left ventricle, which was followed by up-regulation of cardiac gene expression, myocardial inflammation and fibrosis. Using a DNA microarray approach, the cardiac target genes of p38 MAPK were identified, including several cell division, inflammation and signal transduction-associated genes. Furthermore, p38 MAPK over-expression was found to increase the DNA binding activities of several transcription factors, including GATA-4. Finally, the functional role of p38 MAPK was determined using adenovirus-mediated gene transfer in an experimental model of myocardial infarction. Post-infarction remodelling was characterised by a sustained down-regulation of p38 MAPK, while rescue of p38 MAPK activity attenuated post-infarction remodelling through anti-apoptotic and angiogenic mechanisms. These results indicate that p38 MAPK is a key regulator of GATA-4 transcription factor and cardiac gene expression during left ventricular wall stress and remodelling. They demonstrate that p38 MAPK, being cardioprotective in the infarcted heart but promoting inflammation and fibrosis in the normal heart, has a unique dual role in the myocardium.

Many buildings in Pretoria’s central business district stand empty and bandoned. These buildings are being looted, stripped and vandalised, resulting in a great deal of damage that eventually leads to their destruction. The Extramural Building is an iconic modernist building that has served the city well over its lifetime. It has stood abandoned and empty for the last seven years, is in a state of disrepair, and is on the verge of being lost and forgotten. The intention with the dissertation is to connect the past and the future by investigating the potential of old buildings with the aim of retaining their existing fabric and cultural significance. Heritage practice and theories of adaptation are investigated to generate a theoretical framework for the remodelling of old buildings. A statement of significance is developed by investigating the historical context of the building. The Tshwane Inner City Regeneration Framework is used as a starting point for the development of an urban framework for the project. An appropriate program is chosen, derived from the theoretical framework, statement of significance and urban framework. Appropriate precedent studies serve as examples of the discussed theory, urban framework, program and conceptual intentions. The design of a proposed intervention is developed within the established urban framework by applying the theoretical framework. The navigation of the complexities between heritage practice and remodelling ultimately allows a new interpretation of the building to emerge. The argument and its eventual expression results in an intervention that allows the Extramural Building to reclaim its role in the city, unlocking its latent potential, and retaining its inherent cultural value. / ‘n Groot aantal geboue in Pretoria se sentrale besigheidsdistrik staan leeg en verlate. Hierdie geboue word gereeld deur vandale gestroop en geplunder; hulle rig verstommende skade aan wat uiteindelik tot die vernietiging van die geboue lei. Die Buitemuurse Gebou is ‘n ikoniese, moderne gebou wat vir lank ‘n positiewe bydrae tot die stad gelewer het. Die gebou staan al sewe jaar lank leeg en is baie vinnig besig om te verval, wat sonder drastiese ingryping ongetwyfeld tot die gebou se finale vernietiging sal lei. In die verhandeling word daar gepoog om die verlede en toekoms te versoen deur die potensiaal van ou geboue te ondersoek met die doel om die geboue en hul kulturele waarde te behou. Erfenispraktyk en aanpassingsteorieë word ondersoek om ‘n teoretiese raamwerk vir die herbenutting van ou geboue te genereer. ‘n Verklaring van betekenis word ontwikkel deur die gebou se geskiedenis te ondersoek. Die Tshwane Middestad Ontwikkeling- en Herlewingstrategie word as ‘n beginpunt vir die ontwikkeling van ‘n stedelike raamwerk vir die projek gebruik. ‘n Toepaslike gebruiksprogram word afgelei uit die teoretiese raamwerk, die verklaring van betekenis en die stedelike raamwerk. Die studie word deur toepaslike gevallestudies as voorbeelde van die tersaaklike teorieë, stedelike raamwerk, program en konseptuele doelwit van die projek ingelig. Die ontwerp van die voorgestelde ingryping word binne die bestaande stedelike raamwerk in oorleg met die teoretiese raamwerk ontwikkel. Die ondersoek van die kompleksiteite wat tussen erfenispraktyk en aanpassingsteorieë bestaan maak nuwe interpretasies van die ou gebou moontlik. Die argument, en uiteindelike uitdrukking daarvan, word gevolg deur ‘n ingryping wat die Buitemuurse Gebou toelaat om weer sy rol in die stad te vervul en so die latente potensiaal van dié gebou te ontgin, maar ook die inherente kulturele waarde daarvan te laat voortbestaan. / Mini Dissertation MArch(Prof)--University of Pretoria, 2018. / Architecture / MArch(Prof) / Unrestricted