Single-cell analyses reveal two defects in peptide-specific activation of naive T cells from aged mice

Confocal fluorescent microscopy was used to study redistribution of membran e-associated proteins in naive T cells from young and old mice from a trans genic stock whose T cells express a TCR specific for a peptide derived from pigeon cytochrome C, About 50% of the T cells from young mice that formed conjugates,vith peptide-pulsed APC were found to form complexes, at the sit e of binding to the APC, containing CD3 epsilon, linker for activation of T cells (LAT), and Zap-70 in a central area and c-Cbl, p95(vav), Grb-2, PLC gamma, Fyn. and Lck distributed more uniformly across the interface area. T wo-color staining show ed that those cells that were able to relocalize c-C bl, LAT, CD3 epsilon, or PLC gamma typically relocalized all four of these components of the activation complex. About 75% of conjugates that rearrang ed LAT, c-Cbl, or PLC gamma also exhibited cytoplasmic NF-AT migration to t he T cell nucleus. Aging had two effects, First, it led to a diminution of similar to2-fold in the proportion of T cell/APC conjugates that could relo calize any of the nine tested proteins to the immune synapse. Second, aging diminished by similar to2-fold the frequency of cytoplasmic NF-AT migratio n among cells that could generate immune synapses containing LAT, c-Cbl, or PLC gamma, Thus naive CD4 T cells from old mice exhibit at least two separ able defects in the earliest stages of activation induced by peptide/MHC co mplexes.