Thesis (Ph. D.)--University of Queensland, 2002. / Includes bibliographical references.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1963. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Background Superparamagnetic particles of iron oxide (SPIO) are part of a novel and exciting class of ‘smart’ magnetic resonance imaging (MRI) contrast agents that are taken up by inflammatory cells. Ultrasmall SPIO (USPIO; ~30 nm diameter) can be used to assess cellular tissue inflammation and SPIO (80-150 nm) have the potential to be used to label cells ex vivo for in vivo cell tracking studies. Objectives The aims of the thesis were therefore (i) to develop and validate quantitative MRI methodology for assessing SPIO uptake within tissues, (ii) to demonstrate USPIO accumulation within the aortic wall and its implications in patients with abdominal aortic aneurysms (AAA), and (iii) to develop and apply a Good Manufacturing Practice (GMP) compliant method of SPIO cell labelling in healthy volunteers. Methods Patients with asymptomatic AAA >4.0 cm in diameter were recruited. Imaging sequences were optimised in eight patients using a 3 tesla MRI scanner. Data were analysed using the decay constant for multi echo T2* weighted (T2*W) sequences (T2*) or its inverse (R2*) and the repeatability of these measurements was established. A further twenty-nine patients underwent MRI scanning before and 24- 36 hours after administration of USPIO. T2 and multi echo T2*W sequences were performed and ultrasound-based growth rate data were collected. Operative aortic wall tissue samples were obtained from patients undergoing open surgical aneurysm repair. A GMP compliant protocol was developed for labelling cells with SPIO for clinical cell tracking studies. The effects of SPIO-labelling on cell viability and function were assessed in vitro. A phased-dosing protocol was used to establish the safety of intravenous administration of SPIO-labelled cells in healthy volunteers. The feasibility of imaging cells at a target site in vivo following local or systemic administration was assessed. Tracking of SPIO-labelled cells to a target site was investigated by inducing an iatrogenic inflammatory focus in the skin of the anterior thigh of healthy volunteers, following which autologous SPIO-labelled cells were administered and their accumulation was assessed using MRI scanning and histology of skin biopsies. Results Robust and semi-quantitative data acquisition and image analysis methodology was developed for the assessment of SPIO accumulation in tissues. In patients with AAA, histological analysis of aortic wall tissue samples confirmed USPIO accumulation in areas of cellular inflammation. USPIO-enhanced MRI detected aortic wall inflammation and mural USPIO uptake was associated with a 3-fold higher aneurysm expansion rate. Human mononuclear cells were labelled with SPIO under GMP compliant conditions without affecting cell viability or function. Both local and intravenous administration of SPIO-labelled cells was safe and cells were detectable in vitro and in vivo using a clinical MRI scanner. SPIO-labelled cells tracked to a focal iatrogenic inflammatory focus following intravenous administration in humans and were detectable on MRI scanning and histological examination of skin biopsies. Conclusions SPIO contrast agents have an extensive range of potential clinical applications. USPIO uptake in the wall of AAA appears to identify cellular inflammation and predict accelerated aneurysm expansion. This is therefore a promising investigative tool for stratifying the risk of disease progression in patients with AAA, and may also be considered as a biomarker for response to novel pharmacological agents. The ability to label cells for non-invasive cell tracking studies would facilitate the further development of novel cell-based therapies and would enable assessment of dynamic inflammatory processes through inflammatory cell tracking.

M.Cur. / The natural progress of an abdominal aortic aneurysm is enlargement and rupture. The incidence of abdominal aortic aneurysms has increased in the past 30 years and up to 50% of the patients with untreated aneurysms will die due to rupture within 5 years. Open surgery is effective in the prevention of rupture and can be performed with a mortality of 2 -5% in most cases. However, patients with aneurysms are generally older and have associated medical co-morbidities, which increase the risk in surgical intervention. In view of these associated risks with open surgery for abdominal aorta aneurysm repair, a less invasive option such as endoluminal stent-grafts, are often preferred. This new, less invasive technique with Parodi as pioneer has several advantages for patients, the greatest being the reduction in peri-operative risks of aneurysm repair. As in all new procedures, this new intervention sets specific requirements for quality peri-operative nursing. Within the legal-ethical framework of nursing there is no room for random nursing, and we as nurses must turn to protocols and standards applicable to quality nursing, and in effect the quality assurance process. Quality nursing care delivery to the patient remains the ideal of each nurse. The endovascular repair of abdominal aortic aneurysms, although less invasive, is still associated with major morbidity and mortality. The potential for complications is a reality. Complications are mainly systemic and/or procedure related. The reality of these complications affects the quality of nursing. Finally, the need to accommodate this problem requires that protocol/standards are established for the nursing of the patient with an endoluminal repair of an abdominal aortic aneurysm by means of an endovascular stent-graft. The following question can be asked in view of the above arguments and problem statement: How must these patients be nursed peri-operatively to ensure quality nursing care? The aim of this study is to compile protocol/standards for quality nursing of patients with an erldovascular stent-graft repair of an abdominal aortic aneurysm in a Coronary Intensive Care Unit in a private hospital in Cape Town.

TEVAR has been used extensively to treat TBAD. The principle of treatment involve placement of a stent graft in the true lumen to cover the primary tear, thereby excluding the false lumen. Success depends on a combination of factors: reduction of false lumen pressure and perfusion, thrombosis of the false lumen, and remodeling of the aorta leading to eventual healing. The long-term goals are to prevent continuous growth of the false lumen, reinterventions, and aneurysm rupture. The success of TEVAR depends on a combination of factors, including the blood flow and pressure in the two aortic lumens, and remodeling is a dynamic process. Much controversy exists regarding the ideal timing of TEVAR, its efficacy in effecting complete false lumen exclusion, the long-term durability of the repair, and the fate of the aortic size. The objective of this thesis is to examine the morphological and hemodynamic changes within the aortic lumens after TEVAR, using a combination of ex-vivo animal models and computational tomography analysis. The residual pressure of the true and false lumens in TBAD models was studied. Volumetric analyses of CT scan of patients were compared. The ultimate goals are to determine if it is beneficial to treat type B dissections early, and to determine long-term morphological results. In ex-vivo hemodynamic study, 28 fresh porcine aortas models were created to simulate three different pathological scenarios of TBAD: model A represented pre-treated TBAD; model B represented post-treated TBAD with patent false lumen; and model C represented chronic stage of post-treated TBAD with false lumen thrombosis. True lumen and false lumen pressure differences were compared between the three models. Pressure effect was successfully reduced by 30% in model C in comparison with the other two models. No hemodynamic parameters were significantly different between model A and model B. Aortic remodeling parameters were volumetrically analyzed and compared between two groups of patients who underwent endo-grafting for uncomplicated TBAD (group A) and dissecting aneurysms (group B). Modern DIOCM processing workstations and software were used to reconstruct thoracic aorta with serial CT scans. The true lumen, false lumen, thrombus and aortic size were measured volumetrically. Stent graft migration and area of inlet and outlet were also quantified. There were progressive migration and continuous expansion of the stent graft on patients in both groups. Favorable aortic remodeling was observed in most. One fourth of all patients demonstrated aortic volume increase at 36 months. However, there was no difference between group A and group B in terms of stent graft re-shaping and aortic remodeling. In conclusion, Aortic remodeling after TEVAR in treating TBAD is a continuous process. There were no significant differences between chronic dissections and dissecting aneurysms in all morphological parameters. Treating chronic dissections before aneurysm formation does not seem to have a morphologic advantage. Sealing of primary entry tear with introducing thrombosis could significantly reduce false lumen pressure. However, the remaining pressure accumulations should be considered to reduce by further excluding distal reentry tears in those patients who undergo progressive false lumen expansion after TEVER. / published_or_final_version / Surgery / Doctoral / Doctor of Philosophy

Abdominal Aortic Aneurysms (AAA) are amongst the top ten most common cause of death in those over 55 years of age. The disease is usually asymptomatic, often being diagnosed incidentally. Once diagnosed, elective repair of an AAA results in excellent long-term survival with a 3-5% operative mortality. However, up to one half of patients present with ruptured aneurysms, a complication that carries an 80% mortality in the community, and of those reaching hospital, a 50% mortality. Clearly early diagnosis and treatment results in improved survival. Screening for AAA, with ultrasound, would detect aneurysms early, prior to rupture. However, debate continues over the cost effectiveness of population based screening programmes. The identification of a sub-population at a higher risk of developing AAA would increase the yield of a screening prograrmne. A number of populations have been examined, none of which have received international acceptance. About 20% of patients with an AAA have a family history of an aneurysm. The disease is also considered to be a disease of Caucasians, both facts suggesting a strong genetic component to the disease. Perhaps a genetically identified sub-population at a high risk of developing an AAA would prove to be an ideal population for screening. This thesis examines the incidence of aneurysms and the family histories of patients with AAA in the Otago region of New Zealand. Almost twenty percent of the population has a family history of AAA. DNA was collected from each of these patients for genetic analysis. The population was divided into familial AAA and non-familial AAA for the purpose of genetic analysis and compared to a control population. AAA is believed to be a disease of Caucasians; a non-Caucasian population with a low incidence of AAA may prove to be a good control population for genetic studies. A literature review demonstrated a higher incidence of AAA in Caucasians than other ethnic groups and within Caucasians a higher incidence in patients of Northern European origin. The incidence was low in Asian communities, even in studies involving of migrant Asian populations. The New Zealand Maori are believed to have originated from South East Asia, therefore could be expected to have a low incidence of AAA and would make an ideal control population for genetic studies. A pilot study was undertaken to examine the incidence of AAA in the New Zealand Maori. The age standardised incidence of AAA proved to be at least equal in Maori to non-Maori, with a more aggressive form of the disease in Maori, manifesting with a younger age at presentation and a higher incidence of ruptured aneurysms at diagnosis. It is well known that at the time of surgery, an AAA

No description available.

Actin isoform specific mutations have been identified as causes for various human diseases. These include twelve missense mutations in γ-nonmuscle actin leading to early onset autosomal dominate nonsyndromic hearing loss and twenty two missense mutations in α-smooth muscle actin leading to thoracic aortic aneurysms and dissections (TAAD). The molecular mechanisms leading to these human pathologies are mainly unknown, principally due to the inability to isolate pure mutant γ-nonmuscle actin and α-smooth muscle actin in quantities required for biochemical analysis. To begin to address these problems, I have individually expressed the human nonmuscle actin isoforms, β– and γ– nonmuscle actin, in a baculovirus expression system and characterized their biochemical properties. Surprisingly, despite a conserved amino acid difference of only 4 residues at or near the N-terminus, Ca-γ-actin exhibits slower monomeric and filamentous biochemical properties than β-actin. In the Mg-form, the difference between the two is smaller. Mixing experiments with Ca-actins reveal the two will readily co-polymerize. Calcium bound in the high affinity binding site of γ-actin may cause a selective energy barrier relative to β-actin that retards the equilibration between G– and F-monomer conformations resulting in a slower polymerizing actin with greater filament stability. This difference may be particularly important in sites such as the γ-actin-rich cochlear hair cell stereocilium where local mM calcium concentrations may exist. In hair cells γ-nonmuscle actin seems to play a central role in stereocilia maintenance. To determine how the deafness causing D51N-γ-mutant actin mutation leads to deafness, I expressed and characterized it in the γ-actin background. The D51N mutation, lethal when cloned into yeast, displayed decreased filament stability and polymerization kinetics of an actin more dynamic than γ-actin. This result suggests that the hearing effects of the γ-actin mutations on the hearing apparatus are not simply caused by an inability to polymerize. The observed increased polymerization rates and decreased filament stability may have major implications for the human disease, as the mutation may alter the ability of the γ-actin to fulfill its maintenance functions. To address the basis by which TAAD mutations cause vascular dysfunction I introduced two of the know human mutations, N115T and R116Q, into yeast actin, 86% identical to human α-smooth muscle actin. I then generated yeast strains expressing each of these mutations as the sole actin in the cell to assess their effect on actin function in vivo and in vitro. Both mutant strains exhibited reduced ability to grow under a variety of stress conditions, although the N115T cells were more severely affected. In vitro the mutations caused exhibited altered thermostability and nucleotide exchange rates indicating effects on monomer conformation with R116Q the most severely affected. The N115T actin demonstrated a biphasic elongation phase during polymerization, while R116Q actin demonstrated a markedly extended nucleation phase. Allele-specific effects were also seen on critical concentration, rate of depolymerization and filament treadmilling. R116Q filaments were hypersensitive to severing by the actin-binding protein cofilin. In contrast, N115T filaments were hyposensitive to cofilin, despite near normal binding affinities of actin for cofilin. The mutant specific effects on actin behavior suggest that individual mechanisms may contribute to TAAD. Understanding the mechanisms of actin dependent human diseases requires elucidation of the effects of the mutations on the behavior of actin per se, its regulation, and the impact on actin mediated processes within the cell. The work provided in this thesis and future studies will provide the information required to understand the pathways involved in these diseases and form innovative treatments for deafness and TAAD.

The success of endoluminal stent-graft treatment for abdominal aortic aneurysm relies on maintenance of an effective seal when the stent expands into the healthy artery. Clinical observation of aortic neck dilation following endoluminal grafting has led to the hypothesis that excessive stent expansion forces may cause remodelling and dilation of the artery to accommodate the strong forces. This may lead to failure of the seal, hence so-called endoleak. In this research, we analysed the force field generated by aortic stent-grafts and investigated in vitro approaches for studying the effects of these forces on cells within the vascular wall. The pressure-deformation behaviour of ovine arteries was examined experimentally and was found to vary with artery type. A finite element model of abdominal aorta (AA) characterised by Mooney-Rivlin hyperelastic material properties was validated. The property inputs were derived from the polynomial form of the strain energy density function proposed by Patel and Vaishnav. Stent-artery contact simulations revealed stresses 1.2-19 times higher than within a normal vessel at 120 mmHg when contacted by a zig-zag, square cross-section stent that expanded the AA by 3-16%. Streses 1.3-23 times normal were predicted for circular cross-section stents at the same range of expansions. The stress distribution was determined to be concentrated at the contacting surface and within the inner region of the aortic wall. These results confirmed that the forces within the vessel wall are likely to place unnatural physiological demands on the cells within. We then developed an in vitro system for studying the impact of this mechanical stress on cells within a three dimensional (3D) structure. A 20 wt% poly(vinyl alcohol) (PVA) - 5 wt% collagen tubular construct was developed to support cells, and was shown to sustain physiological blood pressures. Two cell-seeding techniques were examined, direct cell encapsulation and surface cell-seeding. Both demonstrated the capability of entrapping viable cells within the construct that remained viable for up to 4 days. In conclusion, stent contact does create abnormal stress concentrations within the vessel wall with a magnitude severely higher than physiological levels. A feasible tubular construct and an in vitro system were developed, enabling further assessments on the effects of these abnormality on the cells.

Abdominal Aortic Aneurysms (AAA) are amongst the top ten most common cause of death in those over 55 years of age. The disease is usually asymptomatic, often being diagnosed incidentally. Once diagnosed, elective repair of an AAA results in excellent long-term survival with a 3-5% operative mortality. However, up to one half of patients present with ruptured aneurysms, a complication that carries an 80% mortality in the community, and of those reaching hospital, a 50% mortality. Clearly early diagnosis and treatment results in improved survival. Screening for AAA, with ultrasound, would detect aneurysms early, prior to rupture. However, debate continues over the cost effectiveness of population based screening programmes. The identification of a sub-population at a higher risk of developing AAA would increase the yield of a screening prograrmne. A number of populations have been examined, none of which have received international acceptance. About 20% of patients with an AAA have a family history of an aneurysm. The disease is also considered to be a disease of Caucasians, both facts suggesting a strong genetic component to the disease. Perhaps a genetically identified sub-population at a high risk of developing an AAA would prove to be an ideal population for screening. This thesis examines the incidence of aneurysms and the family histories of patients with AAA in the Otago region of New Zealand. Almost twenty percent of the population has a family history of AAA. DNA was collected from each of these patients for genetic analysis. The population was divided into familial AAA and non-familial AAA for the purpose of genetic analysis and compared to a control population. AAA is believed to be a disease of Caucasians; a non-Caucasian population with a low incidence of AAA may prove to be a good control population for genetic studies. A literature review demonstrated a higher incidence of AAA in Caucasians than other ethnic groups and within Caucasians a higher incidence in patients of Northern European origin. The incidence was low in Asian communities, even in studies involving of migrant Asian populations. The New Zealand Maori are believed to have originated from South East Asia, therefore could be expected to have a low incidence of AAA and would make an ideal control population for genetic studies. A pilot study was undertaken to examine the incidence of AAA in the New Zealand Maori. The age standardised incidence of AAA proved to be at least equal in Maori to non-Maori, with a more aggressive form of the disease in Maori, manifesting with a younger age at presentation and a higher incidence of ruptured aneurysms at diagnosis. It is well known that at the time of surgery, an AAA is at the end stage in its life. At this time, inflammation and matrix metalloproteinases (MMP) enzymes are prevalent within the aneurysm wall and have destroyed the wall of the aorta. One of the most important genetic pathways regulating these enzymes is the plasminogen activator inhibiter 1-Tissue plasminogen activator-plasmin pathway. Genetic analysis of this pathway demonstrated an association of the 4G5G polymorphism in the promoter of the PAl-1 gene with familial AAA. In this insertion:deletion polymorphism, the 5G variant binds an activator and repressor, resulting in reduced PAI-1 expression and ultimately increased MMP activation. This allele was associated with familial aneurysms, 47% versus 62% non-familial AAA and 61% controls (p=0.024). A polymorphism within the tissue plasminogen activator gene was also examined and no association was found with AAA. Another way the MMPs expression could be increased is from mutations or polymorphisms in their own genetic structure. Stromelysin 3 is itself a MMP capable of destroying the aortic wall and it has a role in activating other MMPs. A 5A6A insertion:deletion polymorphism exists in the promoter of this gene. The 5A allele variant results in increased stromelysin expression and is associated with AAA 46% versus 33% in controls p=0. 0006. The actions of the MMPs are themselves inhibited by the tissue inhibitors of matrix metalloproteinases. The TIMP genes have been sequenced; two polymorphisms have been identified in the non-coding promoter area of the TIMP 1 gene. Further studies are necessary to examine the effect of these polymorphisms. Inflammation has been implicated in aneurysm progression. One of the roles of the inflammatory cells found in an aneurysm is to deliver the MMP�s to the AAA. The HLA system is integral in controlling this inflammation and was therefore examined. From this series of studies it is concluded that there is a genetic component to AAA. This thesis presents the first genetic polymorphism associated with familial AAA and explores the role of a genetic pathway in the formation of AAA.