Background Bos d 2, a major bovine allergen of the lipocalin family, stimulates very weakly cow dust-asthmatic subjects' peripheral blood mononuclear cells and the spleen cells of several inbred mouse strains immunized with the allergen.
Objective To identify the immune mechanisms accounting for the weak stimulatory capacity of Bos d 2.
Methods The spleen cell responses of BALB/c mice immunized with the allergen and with hen egg lysozyme and tetanus toxoid as control antigens were examined using several in vitro methods.
Results Analysis of the numbers of spleen cells in the antigen-stimulated in vitro cultures with the vital dye 7-amino-actinomycin D showed that Bos d 2 induced a smaller expansion of cells in comparison with the control antigens. Increased cell death in vitro did not account for the weak response against Bos d 2. The number of spleen cells reacting against Bos d 2 also proved to be the lowest when they were analysed by labelling the stimulated cells with 5-6-carboxyfluorescein diacetate succinimidyl ester or by enumerating cytokine-secreting cells by ELISPOT. Eliminating CD8+ cells in the in vitro culture did not enhance the response against Bos d 2. Bos d 2 was also the weakest of the antigens to stimulate the production of soluble cytokines. Adding IL-2, IL-4 or antibody against TGF-β in the antigen-stimulated spleen cell cultures enhanced the proliferative responses against all the antigens, whereas adding IL-12 or antibody against IL-4 or IL-10 did not enhance the responses.
Conclusion The results exclude several mechanisms of peripheral tolerance as an explanation for the poor immune response against Bos d 2, and suggest that the allergen is recognized by a low number of specific T cells. The weak immunogenicity of Bos d 2 may be related to its allergenicity.