Non Cystic Fibrosis bronchiectasis is characterised by perpetual neutrophilic inflammation in the lungs. The ongoing vicious cycle of bronchiectasis leads to further damage to already damaged airways and is a culmination of repeated infection, inflammation and failure of the host response to maintain sterility of the airway, despite a sophisticated innate and adaptive immune system. Pseudomonas aeruginosa commonly colonises the lungs of patients with bronchiectasis. I hypothesised that the concept of inhibitory antibodies in the serum may be a feature of Pseudomonas aeruginosa colonisation through a specific interaction between the host adaptive immune system and strain specific features. Here I have identified a mechanism where some patients colonised with Pseudomonas aeruginosa produce IgG2 antibodies specifically against the O antigen of bacterial LPS, which rather than promote complement-mediated killing actually inhibits it.

Traumatic haemorrhagic shock (THS) is the most frequent cause of preventable death after severe injury. Shock is characterised by inadequate provision of oxygen and substrates to tissues in relation to their requirements, and it is within the microcirculation that this process is regulated. Investigation of the microcirculation is therefore key to understanding the pathological processes following THS. In Part I, some mechanisms of microcirculatory dysfunction following trauma are presented. Endotheliopathy of trauma is associated with poor microcirculatory flow, and occurs within minutes of injury. It is also associated with higher levels of circulating cell-free DNA (cfDNA), supporting the hypothesis that cfDNA is an aetiological factor in this pathological response. Both endotheliopathy and elevated cfDNA and are related to poor clinical outcomes. In Part II, clinical implications of microcirculatory monitoring are discussed for patients in the early phase following THS. It is safe and feasible to monitor the microcirculation following THS, and a novel point-of-care grading system has performed well, suggesting that targeted fluid resuscitation towards microcirculatory flow after THS may be possible. The optimal fluid strategy in this context is unknown, but physical properties (e.g. oncotic potential and viscosity) as well as endothelial restorative properties appear to be as important as oxygen-carrying capacity. Potential therapeutic interventions aimed at microcirculatory and endothelial resuscitation open intriguing possibilities for improving outcomes after THS.

B cells have been proven to have a significant role in liver fibrosis. We postulate that enrichment of B cell subsets in hepatic diseases may implicate this population in liver pathogenesis. When comparing total B cells from human immune and non-immune-mediated liver disease explants, we found an enrichment of CD20+ B cells in PBC. Furthermore, phenotypic characterization of 11 B cell subsets in matched liver and blood highlighted an enriched naïve peripheral population, and activated B cell subsets in livers. Newly identified CD19+CD24-CD38- and CD19+CD24-CD38int B cells were also augmented in livers compared to matched blood. Furthermore, CD24-CD38- B cells were elevated in PBC and formed aggregates in tissues, whereas CD24-CD38int B cells localized around bile ducts and along fibrotic tracts in PBC. CD24-CD38int B cells secreted pro-inflammatory (IL-6, IFN-γ) and immunosuppressive (IL-10) cytokines following stimulation with CpG compared to other B cell subsets, implying that CD24- B cells may play a role in liver disease pathogenesis. Our findings suggest that B cells may be influential in hepatic disease progression and pathogenesis. Elucidating their role further could provide possible therapeutic targets for prevention or treatment of chronic liver disease.

Late kidney transplant losses along with chronic kidney disease in non-renal solid organ transplantation pose major problem in this field. To address this I compared ischaemia, interstitial scarring and inflammation on the renal biopsies from failing renal transplants and chronic kidney disease outside renal transplantation using native CKD as a control group. For matched renal function, a similar degree of interstitial scarring was observed across the three study groups. Ischaemia was predominant in failing renal transplants; conversely inflammation was predominant in native CKD. The relationship between macrophages and endothelial cells indicated that, despite a lower macrophage load, there may be an allogeneic impact of macrophages on endothelial cells. In the biopsies from CKD, mRNA levels for iNOS, arginase, CX3CL1, BCL-2, MCP-1/CCL2 and FSP-1 were increased in association with an increasing macrophage load. Macrophage load correlated positively with arginase/iNOS mRNA ratio, suggesting M2 phenotypic transformation of the macrophages. Co-localization studies, demonstrated an increased number of macrophages within 5 microns from endothelial cells in failing renal transplants when compared to chronic kidney disease. Further studies directed towards manipulating macrophage-endothelial cell interaction may be potentially beneficial in improving the longevity of renal transplants.

In post-haemorrhagic communicating hydrocephalus, CSF drainage is obstructed by subarachnoid fibrosis in which the fibrogenic cytokine transforming growth factor-β1 (TGF-β1) has been aetiologically implicated. Here, the hypothesis that the TGF-β antagonist Decorin has therapeutic potential for (1) reducing fibrosis and the development of hydrocephalus and (2) degrading fibrosis and resolving hydrocephalus, was tested using a rat model of juvenile communicating hydrocephalus. In the acute study, hydrocephalus was induced by a basal cistern injection of kaolin in 3-week-old rats, immediately followed by continuous intraventricular infusion of either human recombinant Decorin or PBS. In the chronic study, hydrocephalus was allowed to develop for 7 days before starting the treatment of Decorin or PBS. Ventricular expansion was measured by magnetic resonance imaging. Inflammation, fibrosis, Decorin, TGF-β/Smad2/3 activation and hydrocephalic brain pathology were evaluated by immunohistochemistry and basic histology. In the acute study continuous Decorin infusion prevented the development of hydrocephalus by blocking TGF-β- induced subarachnoid fibrosis and protected against hydrocephalic brain damage. In the chronic study Decorin had no impact on hydrocephalus, TGF-β1 levels or subarachnoid fibrosis, however the efficiency of Decorin infusion was in disrepute. The results suggest that Decorin is a potential clinical therapeutic for the prevention of juvenile post-haemorrhagic communicating hydrocephalus.

Alpha 1 antitrypsin deficiency (AATD) is the only established genetic predisposition to chronic obstructive pulmonary disease (COPD). The development of COPD in AATD is highly variable, probably relating to complex interactions between multiple genetic and environmental factors. This thesis will describe the COPD phenotypes observed in AATD and their inter-relationships, forming the basis for examining phenotypic associations with specific candidate genes and HLA class II type. Finally it examines the potential role of ambient air pollution. Associations were seen for TNFA with chronic bronchitis, SFTPB with FEV1, TGFB with small airways disease and GC with bronchiectasis, consistent with the role of protein products in pathogenesis. Of four MMPs studied, association with gas transfer occurred in two. HLA-DQA1*0301 and HLA-DRB1*04 contributed significantly to gas transfer in regression models, and anti elastin antibodies were higher in HLA-DRB1*04 and HLA-DQA1*0301 homozygotes. Ozone levels contributed to the burden of disease in cross sectional and longitudinal models of pollution exposure, whilst PM10, NO2 and SO2 were associated only in the longitudinal model. In conclusion this thesis demonstrates the importance of genetic variation and environmental factors in determining respiratory phenotype in AATD. It also suggests a key role for adaptive immunity in pathogenesis of emphysema.

The first aim of this thesis was to systematically review the literature to determine the diagnostic accuracy of ultrasound in the first trimester to screen for twin-twin transfusion syndrome (TTTS) and predict its outcome after diagnosis, as well as the effectiveness of its two main treatments. Quantitative analysis revealed that a crown rump discordance or abnormal nuchal translucency in the first trimester was useful to screen for TTTS, but after diagnosis no single test could reliably predict outcome. In both situations a negative test was not reliable for excluding TTTS or a poor outcome. Fetoscopic laser ablation (FLA) is likely to confer a benefit both in terms of survival and morbidity in survivors. The second aim was to determine what happened to biological markers in TTTS and how FLA affected them. It appeared that TTTS may be associated with abnormal placentation as maternal serum α-fetoprotein (MSAFP) approximately doubled and free β-human chorionic gonadotrophin (f- β hCG) tripled. The balance of angiogenic factors i.e. 2-3 fold increased angiogenin 2 and 1.5 fold increased soluble vascular endothelial growth factor receptor 1 appeared to favour angiogenesis in response to hypoxia or ischaemia in TTTS. Maternal cell-free messenger RNA was reliably detected and showed similar alteration in angiogenic markers. Interestingly, TTTS was associated with minimal changes in cytokine levels. In response to FLA there was an increase in transplacental haemorrhage (MSAFP increased 445%) rather than trophoblast destruction (f-hCG unchanged) as well as a transient increase in some anti-angiogenic markers. Although, in general angiogenic factors and cytokines are altered little by this therapy. If the biomarker changes detected precede the onset of clinically apparent disease, they may be useful to improve the performance of first trimester ultrasound screening. Composite tests may be more useful to predict outcome and FLA should continue to be utilised to reduce morbidity.

Background: Diabetes-related Microvascular complications are associated with significant morbidity, mortality and economic burden. Effective treatments for microvascular complications, apart from improved metabolic and blood pressure control, are lacking. Hence, improved understanding of the pathogenesis of these complications is needed to develop new treatments. Obstructive sleep apnoea (OSA) is very common in type 2 diabetes (T2DM) and has been shown to stimulate the same harmful pathways as hyperglycaemia, particularly those that are involved in the pathogenesis of microvascular complications. Hence, it is plausible that OSA is associated with microvascular complications in patients withT2DM. Aims: To explore the interrelationships between OSA and microvascular complications in patients with T2DM and the possible mechanisms behind such relationship. Methods: A cross-sectional study of South Asians and White Europeans with T2DM were randomly recruited from the outpatients of two secondary care diabetes clinics in the UK. Patients were extensively characterised including assessments for OSA and microvascular complications. Results: Patients (n=234) were included in the analysis. OSA prevalence was 64.5%. OSA patients had worse metabolic profile than those without OSA. The prevalence of all microvascular complications (except cardiac autonomic neuropathy) was higher in patients with OSA compared to patients without. After adjustment for a wide range of confounders, OSA remained independently associated with microvascular complications. OSA and hypoxaemia severity correlated with the severity of complications. Based on blood samples and skin biopsies collected during the study, patients with OSA had increased oxidative and nitrosative stress and impaired microvascular regulation compared with patients without OSA. Furthermore, ethnic differences in OSA accounted for some of the ethnic differences in microvascular complications. Conclusion: I have identified a novel association between OSA and microvascular complications in patients with T2DM, with increased nitrosative stress and oxidative stress and impaired microvascular regulation as possible mechanisms. Further prospective observational and interventional studies are needed to assess the impact of OSA and its treatment on the development and progression of microvascular complications.

The role of the periaqueductal grey (PAG) in the central control of micturition was investigated in urethane-anaesthetised rats, with the aims of furthering understanding of the central control of micturition and identifying novel therapeutic targets for urinary incontinence. Experiments using microinjection of GABAA agonists and antagonists into the midbrain showed that transmission through a localised region of the caudal ventrolateral PAG (cvlatPAG) is critical for reflex voiding and the micturition pathway is normally subject to tonic inhibitory GABAergic control. Experiments to determine the role of dopamine in controlling micturition by selectively lesioning dopamine-containing neurons in the cvlatPAG and microinjection of dopamine agonists and antagonists were inconclusive and require further work. Micturition could however be suppressed completely by trains of electrical stimulation applied throughout the midbrain. Microinjection of an excitatory amino acid over the same area reduced the frequency of micturition without disrupting the pattern of voiding. Though further work is required to determine the mechanism by which electrical stimulation inhibits reflex micturition, collaboration with clinical colleagues has indicated the exciting translational potential of electrical stimulation of the midbrain in human patients to treat urinary disturbances that have proven refractory to pharmacotherapy.

Conservative non-pharmacological evidence-based management options for Chronic Obstructive Pulmonary Disease (COPD) primarily focus on developing physiological capacity. With co-morbidities, including those of the musculoskeletal system, contributing to the overall disease severity, further research was needed. This thesis presents a critical review of musculoskeletal management approaches used in COPD, which concluded there is insufficient evidence for using musculoskeletal interventions in COPD management. With a paucity of literature exploring chest wall flexibility and clinical guidelines advocating research into thoracic mobility exercises in COPD, a focus on thoracic spine motion analysis was taken. Soft tissue artefact (STA) threatens the validity of existing in vivo measurement techniques. Having measured and reported unacceptable levels of STA, an alternative approach was developed and tested for reliability as part of this thesis. This technique, along with other measures, was subsequently used to evaluate cervico-thoracic musculoskeletal changes and their relationship with pulmonary function in COPD. In summary, subjects with COPD had reduced spinal motion, altered posture and increased muscle sensitivity compared to controls. Reduced spinal motion and altered neck posture were associated with reduced pulmonary function and having diagnosed COPD. Results from this thesis provide evidence to support inception of a clinical trial of flexibility exercises in COPD.