Studies on the molecular mechanism of T cell triggering and detection and characterization of antigen-specific T cells

Abstract

(cont.) Identification and characterization of T cell epitopes derived from infectious agents or vaccines can greatly enhance the ability to study and eventually direct the immune response. This work contains a description of a novel technique for identifying and characterizing specific T cell responses in parallel. The system involves incubating heterogeneous T cell mixtures with artificial antigen-presenting microarrays, which include immobilized cytokine capture antibodies, co-stimulatory molecules, and MHC complexes presenting many different potential T cell epitope peptides. These microarrays can rapidly, conveniently, and sensitively detect antigen-specific T cells and characterize the functional responses to many different epitopes in parallel in a location-dependent manner.

T cells recognize cognate peptide antigen in complex with major histocompatibility complex (MHC) proteins; however, the molecular events which trigger T cells upon binding of MHC-peptide to the T cell receptor (TCR) are unclear. To gain a better understanding of this mechanism, CD4+ T cells were treated with soluble class II MHC-peptide monomers and oligomers instead of antigen-presenting cells, and the activation response was monitored. These experiments showed a requirement for multivalent TCR engagement to induce activation. Mathematical modeling of oligomeric equilibrium binding states indicates that the level of the T cell response correlates with the predicted number of receptor cross-links formed by soluble MHC oligomers. Treatment of CD8+ T cells with class I MHC monomers and oligomers revealed a confusing process whereby peptide derived from soluble MHC reagents was loaded on to endogenous MHC complexes on the T cell surface and re-presented to other cells. When this method of stimulation was circumvented, multivalent TCR engagement was found to be required for CD8+ T cell activation, similar to CD4+ T cells. In both types of cells, monomeric MHC-peptide binding can compete off activation responses induced by MHC-peptide oligomers in the same mixture, further emphasizing the non-productive nature of monovalent TCR engagement. However, exquisite antigen sensitivity might be achieved on the surface of an APC due to the contribution of non-activating MHC-peptide complexes. Even though solubme monomeric MHC does not cause activation, soluble heterodimeric MHC-peptide ligands with only one activating peptide induce T cell activation responses.