In vitro induction of naive cytotoxic T lymphocytes with complexes of peptide and recombinant MHC class I molecules coated onto beads: role of TCR/ligand density.

Abstract

We previously reported that complexes of peptide with soluble single-chain recombinant MHC (SC-MHC) class I molecules are able to induce cytotoxic T lymphocytes (CTL) in vitro in a murine system with an efficiency comparable to that observed with peptide-pulsed dendritic cells as antigen-presenting cells. In this report, we have assessed the capacity of preformed peptide/SC-Kd complexes in monomeric or dimeric form as well as of peptide/SC-Kd-loaded beads to generate in vitro specific CTL responses from naive DBA/2 spleen cells. Peptide/SC-Kd-coated beads were consistently more efficient. We evaluated the role of costimulatory molecules, using monoclonal antibodies anti-CD80 or anti-CD86. In addition, the capacity of peptide/SC-Kd-coated beads to generate a CTL response from purified naive CD8+ T cells was ascertained. Taken together, the results indicate that, under our conditions, CTL priming does not require the participation of co-stimulatory molecules and is the consequence of a direct interaction between the cognate TCR on peptide-specific CTL precursors and the peptide/SC-Kd-loaded beads. Titration of the amount of preformed complexes of SC-Kd and peptide 170-179 of HLA-CW3 that need to be coated onto the beads to prime CTL precursors shows an activation threshold which can be calculated to be between 25000 and 50000 complexes. In effect, in cultures stimulated with specific peptide CW3/SC-Kd complexes representing less than 50% occupancy of the total (10(5)) complexes on the beads, no peptide-specific cytolytic activity was observed. These results suggest that the efficiency of the primary CTL induction depends on the density of specific peptide/SC-Kd complexes present on the beads.