Analysis of cytokine-producing Th cells from hen egg lysozyme-immunized mice reveals large numbers specific for "cryptic" peptides and different repertoires among different Th populations

Abstract

We employed an optimized ex vivo enzyme-linked immunospot assay for enumerating and defining the peptide specificity of all the hen egg lysozyme (HEL)-specific Th cells producing IL-2, IFN-γ, or IL-4, in different lymphoid organs of HEL-immunized BALB/c and CBA mice. Previous studies, employing T cell proliferation assays, demonstrated that lymph node cells from BALB/c mice immunized with HEL emulsified in complete Freund's adjuvant (CFA) are specific for HEL105-120. In contrast, we found that the spleens of BALB/c mice immunized with HEL/CFA, or with heat-aggregated HEL on aluminum hydroxide adjuvant, contain IL-4-producing T cells specific for other HEL peptides, previously characterized as "cryptic", with consistent responses to HEL11-25. The Th repertoire expressed in different lymphoid organs of the same immunized mouse can be different, as can the repertoire of Th cells producing different cytokines and present in one lymphoid organ. In addition, we found that the repertoire of Th cells generated depends upon the adjuvant employed. Lastly, the summation of responses elicited by a panel of non-overlapping HEL peptides is equal to that elicited by HEL. This high-resolution study thus illustrates that the Th repertoire generated upon HEL immunization depends upon diverse parameters, and that the natural processing of HEL gives rise to more diverse peptides then previously evident from studies employing T cell proliferation assays. © 2005 Wiley-VCH Verlag GmbH & Co. KgaA, Weinheim.