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Title: Investigation of the abberent immune resonse in Behçet Syndrome
Author: Ambrose, Nicola
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2013
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Behçet Syndrome (BS) is a multisystem disorder characterized by mucocutaneous lesions (oral ulcers, genital ulcers, and skin pustules), arthritis and intraocular inflammation. The pathogenesis of BS is poorly understood but it is considered a complex polygenetic syndrome, with environmental triggers. A Th1 skew has been reported. Recent genome wide association studies have confirmed the association with HLA B51, as well as identifying novel single nucleotide polymorphisms including IL-10, IL23/12R, STAT4, ERAP1 and CPLX1. Aims: This project aimed to dissect abnormal cutaneous inflammatory response in BS to gain an understanding as to which pathways are responsible for the pathergy response. Secondary aims were to dissect any observations in more detail in vitro. In Chapter 3, the hypothesis that the abnormal cutaneous inflammatory response to "non-specific" stimulation in BS is associated with a failure of deactivation of macrophages during the resolution phase of inflammation was explored using human in vivo models of inflammation (MSU injections and skin blisters). In Chapter 4, the hypothesis that the abnormal cutaneous inflammatory response may involve neuropeptides was explored. We applied capsaicin topically to BS and HC, and measured the resultant change in dermal blood flow using laser doppler imaging to evaluate non-invasively if neuropeptides such as CGRP may play a role in the abnormal cutaneous inflammatory response in BS. Chapter 5 summarizes key in vitro experiments undertaken alongside in vivo experiments. The IFN-γ / chemokine pathways were explored in detail in CD14+ monocyte populations in BS, HC and disease control subjects. Results: CXCL10 was identified as a chemokine of interest using the Cantharidin skin blister model of inflammation. We did not find statistically significant differences in neuropeptide responses between BS and HC, although there was a trend towards less CGRP in the BS cohort. In vitro work focused on peripheral blood derived monocytes stimulated with IFN-γ. BS monocytes produced significantly more CXCL10 protein versus healthy control (HC) monocytes from two hours onwards, despite equivalent quantities of mRNA, revealing a significantly increased protein:mRNA ratio. This increased protein:mRNA ratio was found to be a specific response. Other IFN-γ induced chemokines examined did not show this response profile (CXCL9,CXCL11, CCL2); monocytes stimulated with other cytokines (e.g. TNF-γ) did not show this specific response; and the increased CXCL10 protein mRNA ratio was not observed in either Rheumatoid Arthritis (RA) or Systemic Lupus Erythematosus (SLE) patients. Sucrose density gradients, to segregate cell lysate mRNA into free RNA or polysome-associated RNA, did not further differentiate HCs from BS, with equal quantities of mRNA in BS and HC samples being found in association with polysomes, possibly implicating microRNAs as translation control factors of interest. Conclusions: We have discovered that BS monocytes have dysfunctional post-transcriptional regulation of CXCL-10 mRNA, resulting in overexpression of CXCL-10 protein upon IFN-γ stimulation. This may contribute to the exaggerated inflammatory responses that characterises BS and could also be diagnostically useful.
Supervisor: Haskard, Dorian ; Botto, Marina Sponsor: Arthritis Research UK
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available