Testing time-, ignorance-, and danger-based models of tolerance

In this study, we present data showing that tolerance to Ags in the periphe ry is not determined by the time at which the Ag appears, or by special pro perties of tissues in newborn mice or newly developing immune systems. We p laced male grafts onto immunoincompetent female mice, allowed the grafts to heal for up to 5 mo, and then repopulated the recipients with fetal liver stem cells, We found that the newly arising T cells were neither tolerant n or ignorant of the grafts, but promptly rejected them, though they did not reject female grafts, nor show any signs of autoimmunity, We also found tha t the H-Y Ag was continuously cross-presented on host APCs, that this prese ntation was immunogenic, not tolerogenic, and that it depended on the conti nuous presence of the graft. In searching for the stimulus that might activ ate the host APCs, we analyzed mRNA expression with a highly sensitive real -time quantitative PCR assay, By using two different "housekeeping" molecul es for comparison, we analyzed the message levels for several stress and/or inflammatory molecules in the healed grafts. We found that the long-healed grafts were not equivalent to "normal" skin because the healed grafts expr essed lower levels of GAPDH. Altogether, these data suggest that acceptance vs rejection of peripheral tissues is not attributable to ignorance, timin g-based tolerance, or special circulation properties of naive T cells in ne onatal tissues. It is more likely attributable to an aspect of the context of Ag presentation that remains to be identified.