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No description available.

Objectives: The aims of the study were to identify associations between ACE I/D and MTHFR C677T and AAA. Methods: A retrospective case-control study in which polymerase chain reaction (PCR) methodology was employed to identify associations between ACE I/D and MTHFR C677T polymorphisms and AAA. DNA was extracted from reasonably matched cases and controls after suitable screening for group assignment. There were a total of 1352 subjects genotyped for the MTHFR C677T polymorphism comprising 674 controls and 678 cases. Comparative figures for ACE I/D polymorphism genotyping were 812 and 1107, respectively. All statistical analyses were conducted using R programming software with user-written codes. Results: The ACE II, ID and DD genotype distributions in controls (177, 410 and 225) and cases (218, 529 and 270) were in Hardy-Weinberg Equilibrium (HWE), P=0.21.There was no difference in allele (“I” and “D”) distributions between cases and controls (odds ratio(OR),1.001; 95% CI, 0.88-1.14; P =0.98). There was no difference between cases and controls in terms of the II, ID and DD distributions irrespective of the genetic model adopted. Similarly, the MTHFR CC, CT and TT genotype distributions for controls (358, 257 and 59) and cases (321, 292, and 65) were in HWE (P = 0.39) but the allele (“C” and “T”) distributions were not significantly different between groups (OR, 1.172; 95%CI, 0.99 -1.38; P=0.057). However, MTHFR C677T polymorphism was significantly associated with AAA under a heterozygote co-dominant (OR, 1.27; 95% CI, 1.01-1.59) and dominant (CT+TT vs. CC) (OR, 1.26; 95% CI, 1.02-1.56; P= 0.034) genetic inheritance models, respectively. However, there was no association under the over-dominant (CT vs. CC +TT) model (OR, 1.23; 95% CI, 0.99-1.53; P=0.06). Similarly, the trend test was not significant (OR, 1.14; P=0.06) and when corrected for confounders. Conclusion: The ACE I/D and MTHFR C677T genetic polymorphisms were not independently associated with AAA in this study.

Abdominal aortic aneurysm (AAA), age-related macular degeneration (AMO) and invasive meningococcal disease (IMO) are multifactorial diseases which have genetic and environmental risk factors. The complement system is an ancient part of the innate immune system that is activated in all three of these diseases. AAA is a pathological dilatation of the abdominal aorta that can rupture, often causing death. AMD is a common sight-impairing inflammatory retinal disease. IMD is due to infection with Neisseria meningilidis, which can cause potentially fatal illness in some people. The aim of this work is to better understand the effect of human genetic polymorph isms on susceptibility to these diseases. Genome-wide association study data from collaborators (for AAA) and a public data repository (for AM D) were reanalysed to find novel genetic risk factors. Replication studies were conducted for both of these investigations. Complement system candidate gene studies were conducted for AAA, AMD and IMD using primer extension and sequencing methods. Genetic risk factors for AAA were identified in the genome-wide study, one of which, rs6511720 T in low density lipoprotein receptor (LDLR) , was associated at genome-wide significance (odds ratio 0.78; 95% confidence interval 0.69-0.86; P= 1.03x10-s). Complement gene polymorphisms were not found to be important for AAA risk. A coding polymorphism in CFB, rs12614, was identified as a novel independent risk factor for AMD. The AMD genome-wide study reanalysis identified several bio logically plausible loci that did not reach significance in a small follow-up study. Investigation of complement genes in IMO revealed that MBL2 structural polymorphisms, previously reported to be of major importance, were not associated. The AMO- and IMO-associated CFH locus was characterised in detail. The results of these investigations improve understanding of the pathogenesis of all three diseases. This knowledge may play a part in facilitating the development of new preventative and therapeutic strategies.

Abdominal aortic aneurysms (AAA) are characterised by a chronic inflammatory infiltrate within the abdominal aortic wall and aortic smooth muscle cell (AoSMC) apoptosis. It is postulated that the inflammatory infiltrate causes AoSMC apoptosis, with resultant aortic wall weakening and aneurysmal degeneration. This putative immune-mediated reaction against aortic wall component suggests that AAA may have features of an auto immune disease. It has been previously demonstrated that natural killer (NK) cells are elevated in the peripheral blood (PB) of AAA patients and display increased cytotoxicity against AoSMC. This study aimed to identify the molecular basis of the increased NK cell cytotoxicity and why an immune-mediated reaction occurs against AoSMC. Using multi-parametric flow cytometry (FC), expression of the activatory receptors NKp30, NKp44, NKp46 and NKG2D were analysed on PB NK cells from AAA patients and age-sex-matched healthy controls. No difference in activatory receptor expression or cell surface density (ΔMFI) existed between the two groups. Region specific (intra-luminal blood and AAA tissue) activatory receptor phenotypes were also investigated in AAA patients. The significant finding was a reduction in the ΔMFI of NKG2D on tissue NK cells, suggesting an interaction between this receptor and potential cognate ligands within the aortic wall. Characterised AoSMC explanted from AAA tissue were subjected to analysis using qRT-PCR and FC to identify the expression of death receptors (Fas, TRAIL-RI and TRAIL R2) and NKG2D ligands (MICA, MICB, ULBPI-3). AoSMC expressed mRNA for all NKG2D ligands. FC confirmed the cell-surface expression of NKG2D ligands and the death receptors. A significantly greater percentage of NK cells from AAA patients were CD107a+ when co-cultured with AAA AoSMC, thus accounting for the increased cytotoxicity in this group. Despite using anti-NKG2D it was not possible to inhibit NK cell degranulation in response to the NKG2D ligands on AoSMC. This work has demonstrated that AoSMC from AAA express death receptors and NKG2D ligands, potentially accounting for the NK cell molecular mechanism that leads to AoSMC apoptosis. The expression of NKG2D ligands, which have been demonstrated in other auto immune diseases, favours the hypothesis that AAA are an immune-mediated process directed against the abdominal aortic wall.

Patient specific wall stress analysis demonstrated that wall stress was higher in AAA's at risk of rupture. Furthermore, there is a correlation between sites of rupture and high wall stress regions. We set out to investigate the gene expression in regions of high and low wall stress to identify the role of wall stress in the pathogenesis of AAA Methods Finite element analysis (PEA, ADINA R&D Inc., Watertown, USA) was performed on patients with AAA scheduled for open repair. Regions of high and low wall stress were identified from the obtained patient specific wall stress map. Coordinates of regions of high and low stress were mapped on a three-dimensional reconstruction of each aneurysm which included major visceral branches as reference points to aid in intra-operative localization. High and low stress regions were marked intra-operatively on the aneurysm surface according to their distance from the reference points and full-thickness biopsies were obtained. RNA was extracted (RNeasy Fibrous Tissue RNA Extraction Kit, Qiagen, UK) and whole genome profiling was performed (Illurnina HumanRef-8 v3.0 Expression BeadChips). Protein expression was determined by Western blotting. Results PEA was performed on 11 patients. Paired samples were obtained from high and low wall stress regions. AAA wall was found to be thinner in regions exposed to high wall stress. There was over-expression of LMNA (Larnin AlC) in high wall stress regions. Over-expression of lamin AlC was also demonstrated on Western blotting. Conclusion Our results identify novel pre-rupture changes in AAA's in regions exposed to high stress. This is the first study to identify a role for lamin AlC in AAA pathogenesis. Over expression of lamin A/C in high wall stress regions highlights the role of cytoskeletal and nuclear mechanics, mechanotransduction and apoptotic transcriptional pathways in AAA development and rupture.

No description available.

AAA disease remains a common and life threatening condition, predominantly affecting men of retirement age. Whilst clinical studies have done much to predict the course of this disease, understanding the pathogenesis has been complicated by both a multi-factorial aetiology as well as several poorly defined stages to the disease (formation, growth and rupture). Evidence points to a considerable inheritable component to this condition, but as yet, associations with identified genetic variants are few and weak. This thesis describes the current understanding of the molecular mechanisms behind AAA pathogenesis, concentrating on aneurysm formation and growth. A meta-analysis of published candidate gene studies identified three genes with small but significant effects on risk of developing AAA (ACE, MTHFR and MMP9) and none with a significant effect on AAA growth. Further examination of five genes connected the Renin-Angiotensin System, using three distinct case control series, demonstrated the strongest reported association to date with AAA disease risk, with AGTR1+1166A>C. (OR 1.55 [1.30-1.83, p=5x10-7]). An interest in the role of the TGF-β pathway in AAA formation and growth has developed from the recent illumination of the mechanism behind aneurysm aetiology in Marfan syndrome. Haplotype analysis was used to investigate five genes coding for TGF-β and its binding protein (LTBP). Variants in TGFB3 and LTBP4 were significantly associated with altered AAA growth. The importance of inflammatory process was also supported by observations made in a very large longitudinal data set of AAA growth. Anti-inflammatory drugs, together with anti-platelet drugs and drugs used in diabetes, were significantly associated with decreased AAA growth independent of confounding factors. In conclusion, this thesis demonstrates; a role for the RAS in AAA formation; TGF-β in AAA growth; and anti-inflammatory drugs as potentially disease modifying. In addition, observations have also been made concerning a two tier effect illuding to the nature AAA progression.

An estimated 85% of haemorrhagic strokes are secondary to the rupture of an intracranial aneurysm (IA), a localised, blood-filled dilation of the artery wall. The clinically observed rupture of occluded IAs has led to hypothesise that the presence of thrombus may restrict the transport of nutrients, most notably oxygen, to the aneurysmal wall, thus heightening the risk of rupture through the deleterious effects of hypoxia on cellular functionality. The limited research into O2 transport within IAs demonstrate the need for further exploration into the possible detrimental hypoxic conditions as a result of intrasaccular haemodynamics and thrombusformation in untreated, treated and evolving IAs, with the ultimate goal of further understanding disease evolution and developing prognostic decision support models for clinical intervention. Preliminary computational fluid dynamic simulations conducted on a 2Daxisymmetric model of a thrombosed artery highlighted the relative importance of wall-side versus the fluid-side mass transport of oxygen. A sensitivity analysis demonstrated that variations in thrombus thickness, and arterial wall cellular respiration rates have the greatest influence on the oxygen distribution to the portion of wall in direct contact with the thrombus. The results of the coupled flow-mass transport computational fluid dynamic simulations within patient-specific IA show that a reduction in intrasaccular flow as a consequence of stent deployment affects the rate at which oxygenated blood reaches the entirety of the dome. Nonetheless, the distribution ofO2 to the aneurysmal wall itself does not differ from the observed oxygen distribution across the wall when the same IA is left untreated. Conversely, the low velocity recirculations observed in an IA presenting with a very high aspect ratio (i.e a narrow neck and high sack height) limited the transport of oxygen to such an extent as to completely deprive the delivery of oxygen to the fundus. The presence of thrombus within the IA dome results in a dramatic reduction in oxygen delivery to the wall, the extent of which is dependent on the local thrombus thickness. Finally, a novel fluid-solid-growth-mass transport (FSGT) mathematical model is conceived to explore the biochemical role of thrombus on the evolution of an IA. The shear-regulate propagation of a thrombus layer during membrane expansion leads to the gradual decrease in oxygen tension within the wall. Moreover, as a consequence of coupling this oxygen deficiency to fibroblast functionality, the collagen fibre mass density was shown to increase at an insufficient rate to compensate for the transfer in load from the degrading elastinous consitituents to the collagenous constituents, thus resulting in the increased stretch of collagen fibres in order to maintain mechanical equilibrium. Moreover this over-expansion results in the gradual unstable evolution of the IA. The observed obstruction to oxygen delivery as a result of intrasaccular haemodynamics and thrombosis compounds the need for further development of more comprehensive chemo-mechano-biological models of IAs so as to better ascertain the level of rupture risk posed by a hypoxic environment. Refinement to the models proposed within this work would prove invaluable to creating a fully integrated multi-physics, multi-scale in silico framework in aid to patient diagnostics and individual treatment planning of IAs.