Cell Mediated Immunity in Pigeon Breeders' Disease

This thesis explored the role of cell mediated immunity in the pathogenesis of extrinsic allergic alveolitis (EAA), using pigeon breeders disease (PBD) as a model. The first part of the thesis looked at the total and differential white blood cell count (WBCC) and lymphocyte subsets in the peripheral blood of pigeon breeders, grouped into well defined categories according to symptoms, presence of serum antibody to pigeon antigen and whether or not they were acutely symptomatic at the time of blood sampling. The aim was to determine whether there were any changes in peripheral blood leucocyte phenotypes and if these changes reflected the symptomatic status of the subjects. The initial analysis revealed that symptomatic, antibody positive, pigeon breeders, who were acutely symptomatic at the time of blood sampling, had a leucocytosis, comprising neutrophils and T lymphocytes of the CD4+ and CDS+ subsets. The number of CD56+ cells was also increased, but B cell numbers were unchanged. Pigeon breeders who were normally symptomatic and antibody positive, but who were not acutely symptomatic at the time of blood sampling, had normal peripheral blood cell profiles. These results suggest that acute exacerbations of PBD lead to a transient leucocytosis. An increased proportion of asymptomatic, antibody positive subjects had high CD4:CD8 ratios compared with controls, due to increased numbers of CD4+ cells. The increase in the numbers of CD4+ cells may reflect the presence of an active humoral immune response, in these subjects. Asymptomatic, antibody positive and asymptomatic, antibody negative pigeon breeders had normal peripheral blood cell profiles. A proportion of pigeon breeders (41%) with severe longstanding PBD, had reversed CD4:CD8 ratios in their peripheral blood compared with controls. This was due predominantly to increased numbers of CDS cells, which were both CD3+ T cells and CD3- cells, possibly CD56+ NK cells. Further phenotypic analysis of these cells revealed increased percentages of CD3-CD16+ NK cells, which were predominantly CD8-, but this was not associated with increased cytotoxicity of these cells, as measured (against K562 target cells) in a chromium release assay. It is possible that these cells may be involved in an antibody dependent cellular cytotoxicity reaction in the lung. They might also act as inflammatory mediators. These pigeon breeders also had significantly reduced percentages of gammadelta T cells, predominantly due to a decrease in BB3+ cells, the predominant subpopulation of gammadelta T cells in the lung and the peripheral blood. Both CDS+ and CD8-gammadelta T cells were reduced. It is possible that these cells are being recruited to the lung. Although there was no change in the levels of "naive" (CD45RA+) CD4+ and "memory" (CDW29+) CD4+ cells in the peripheral blood of these pigeon breeders, total levels of CD45RA'*' cells were increased, predominantly due to CD4-CD45RA+ cells. These cells were not identified. The changes recorded, in the levels of different lymphocyte subpopulations in the peripheral blood of these pigeon breeders, with severe longstanding PBD, did not correlate with each other. This suggests that the changes occurring, in each lymphocyte subpopulation, are independent and may be associated with different pigeon breeders, who are perhaps at a different stage of disease. The remaining part of the thesis attempted to determine the stimulus for the changes in the lymphocyte subpopulations, within the group of pigeon breeders with severe longstanding PBD. This was done by assessing whether antigens extracted from pigeon material could provoke cell mediated immune responses by lymphocytes in vitro. The initial studies used pigeon serum as an antigen source. This was found to stimulate lymphocytes from both pigeon breeders and controls. To determine whether this mitogenicity was masking an underlying antigenic response, pigeon serum was fractionated into a "globulin" and "albumin rich" fraction. The "globulin" fraction failed to induce a proliferative response in either pigeon breeders or controls. In contrast, the "albumin rich" fraction was found to possess significant mitogenicity, although with reduced activity, compared with whole pigeon serum. This mitogenicity was not masking a specific antigenic response to pigeon albumin, since a commercial source of pure pigeon albumin failed to induce lymphocyte proliferation. Therefore these studies did not identify any specific response of lymphocytes from pigeon breeders to pigeon serum antigens. The nature of the mitogenic agent in pigeon serum was explored by testing the hypothesis that it was endotoxin. which could be detected in pigeon serum at a concentration of 5pg/ml. Equivalent doses of LPS, similar to those that would be present in a mitogenic dose of pigeon serum, were used to try to mimic the effect of pigeon serum. Although no proliferative response of lymphocytes from either pigeon breeders or controls was induced, the results were inconclusive, since recommended doses of LPS also failed to stimulate lymphocytes to proliferate