Sewell, G.W. (2011) An investigation of molecular defects underlying impaired acute inflammation in Crohn’s disease. Doctoral thesis, UCL (University College London).
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Mounting evidence suggests the pathogenesis of Crohn’s disease (CD) involves an impaired acute inflammatory response. Monocyte-derived macrophages from CD patients release deficient levels of pro-inflammatory cytokines in response to Escherichia coli, arising from post-translational trafficking abnormalities. In this thesis, the macrophage responses to microbial stimuli were further characterised, and the underlying molecular lesions investigated. Macrophage TNF release was attenuated in response to Escherichia coli, Candida albicans and Toll-like receptor (TLR) stimulation in CD. This deficit was unrelated to use of medication, age, gender and smoking status. Patients with stricturing and colonic disease demonstrated the most profound defects in response to TLR2 and TLR4 stimulation respectively. Genotyping for 34 known CD susceptibility polymorphisms revealed no detectable association between any individual variant and impaired TNF release. However in CD, TNF secretion in response to Escherichia coli was weakly correlated with overall genetic risk score. Macrophage sphingolipid and phospholipid compositions were subsequently investigated, given the prominent role of these molecules in vesicle trafficking. Mass spectrometric analysis revealed no gross abnormalities in CD macrophages, although a reduced percentage of phosphatidylinositol 16:0/18:1 was synthesised over 3 hours; the same species was also present at reduced levels in ileal biopsies. Given the heterogeneity of CD, a microarray outlier analysis strategy was developed to identify specific gene expression abnormalities in individual patients. A subset of patients had deficient expression of optineurin, a molecule implicated in vesicle trafficking and autophagy. These individuals shared a number of allelic variants, which were associated with optineurin expression levels. Macrophages from these patients had attenuated TNF secretion downstream of TLR2. Concordantly, depletion of optineurin in THP-1 cells impaired TNF release after TLR2 activation. This study supports a role for immune deficiency in the pathogenesis of CD and identifies abnormal optineurin expression as a relevant molecular abnormality in a subset of patients.
|Title:||An investigation of molecular defects underlying impaired acute inflammation in Crohn’s disease|
|Additional information:||Third party copyright material (articles) has been removed from this thesis|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Medical Sciences > Medicine (Division of)|
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