Background. Administration of cyclosporine after syngeneic bone marrow transplantation elicits a systemic autoaggression syndrome termed syngeneic graft-versus-host disease (SGVHD). The effector T cells recognize a peptide from the invariant chain termed CLIP (MHC class II invariant chain peptide) presented on MHC class II molecules. Moreover, the N-terminal flanking region of CLIP interacts with the T-cell receptor (TcR) β chain. Methods. The current study uses a novel approach to isolate and examine the responding T cells ex vivo. A soluble MHC class II molecule using immunoglobulin (Ig)G (MHC class II-Ig) as a scaffold was constructed. The MHC class II-Ig molecule loaded with different peptide variants of CLIP (lacking the N-terminal or C-terminal flanking regions of CLIP) was used to isolate antigen-specific T cells from animals with SGVHD by panning or by flow cytometric sorting. Results. Two subsets of antigen specific T cells restricted by the N- and C-terminal flanking regions of CLIP can be isolated during acute SGVHD that express the Vβ8.5 TcR determinant but secrete different cytokines (interferon [IFN]-γ interleukin [IL]-10) as detected by real-time quantitative polymerase chain reaction (PCR). Spectratyping of the complementaritydetermining region 3 regions reveals that the N-terminal CLIP reactive population has greater diversity in both the CDR3 and J regions than the C-terminal CLIP reactive subset. Interestingly, the N-terminal CLIP reactive cells can be preferentially detected in the target tissues during acute SGVHD. However, only IFN-γ messenger (m)RNA was detected in the tissues. Conclusion. The ability to isolate and examine antigen specific T cells ex vivo has revealed unexpected differences in TcR diversity and cytokine production in SGVHD.
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