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Title: Cellular and humoral mechanisms of allergic disease
Author: Smith, Karen
Awarding Body: University of Brighton
Current Institution: University of Brighton
Date of Award: 2012
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CD 154 is a T cell activation marker, transiently expressed following ligation of the T cell receptor, therefore providing direct access to an antigen-specific T cell population. Using peripheral blood samples taken from allergic individuals and healthy non-atopic controls, this project identified and phenotyped CD 154 + T helper cells following ex vivo stimulation with native allergen extracts (birch pollen, cat dander, grass pollen). Peripheral blood mononuclear cells were stimulated with allergen extract for 16 hours in the presence of Brefeldin A. Responding CD 154 + T cells were identified and phenotyped using multiparametric flow cytometry. Activated CD154+ TH1, TH2 and TRl-Iike cells, that co-expressed IFNy, IL4 and ILIO respectively, were identified in allergic and non-allergic participants. A close correlation was observed between TH1, TH2 and TR1-like cell frequency in non- allergic participants, such that the three parameters increased together to maintain a low TH2:TH1 ratio. The relationship between TH1, TH2 and TR1-like responses was dysregulated in allergic individuals, with abrogation of the IL10 response and a higher TH2:THI ratio. A close correlation was observed between Th2 cell frequency and the absolute concentration of birch-specific IgE. This work confirms previous reports of a more differentiated T cell phenotype in allergic subjects with regard to seasonal allergens. The detection of CD154+ T cells after short-term antigen stimulation may be a useful method for the detection of T cell responses to allergens when cost, speed and convenience are priorities. The CD 154 assay was also used to investigate allergen-specific T cells in two situations: [1] during allergoid immunotherapy and [2] at peak pollen season compared to out of season. This preliminary data suggests an increased frequency of TH2 cells in allergic individuals during the birch pollen season compared to healthy non-allergic controls, and a decrease in the TH2:THI ratio following successful immunotherapy. A proliferation assay utilising the cell surface PKH dye was also optimised to investigate proliferative responses to native allergen extracts in allergic and non- allergic subjects. Colonisation with superantigen-producing staphylococci is common in atopic diseases and may contribute to the initiation and maintenance of allergic sensitisation. However, little is known regarding T cell responses to superantigens in atopic individuals. This project sought to investigate T helper cell responses to the superantigen Staphylococcal Enterotoxin B (SEB) from Staphylococcus aureus in non-atopic individuals and highly atopic polysensitised individuals. Peripheral blood mononuclear cells were stimulated with SEB for 16 hours in the presence of Brefeldin A. Responding TH1, TH2, TH17, TR1-like and naturally occurring regulatory T cells were identified using multiparametric flow cytometry. This preliminary study identified a downregulated T H 17 response to the superantigen SEB in highly atopic individuals that does not relate to T Reg cell frequency or TCRVP3 expression. The Pollen-Food Syndrome (PFS) is caused by sensitisation to homologous panallergens within aeroallergens and food proteins. Information regarding the sensitisation profiles of individuals with PFS in the UK is limited and investigations into causative panallergens are not routinely performed. In a small study, patients with symptoms suggestive of PFS were recruited. from the Allergy Clinic at Brighton and Sussex University Hospital NHS trust. A standardised food allergy questionnaire was completed and serum analysed by component-resolved diagnosis using ImmunoCAP ISAC technology. This study cohort conformed to the Northern European pattern of birch-pollen associated PFS with cross-reactivity between the major birch pollen allergen Bet v 1 and homologues in food proteins. Sensitisation to LTPs and profilins was also noted, but the clinical relevance of these remains to be elucidated. Profilin sensitisation was associated with reactions to a significantly larger number of foods compared to PR-10 mono sensitised individuals.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: B130 Pathology