Donor-Specific Unresponsiveness to Murine Cardiac Allografts Induced by Intrathymic-Soluble Alloantigens Is Dependent on Alternate Pathway of Antigen Presentation

Abstract

This study extends the finding that intrathymic (IT) inoculation of uv-B irradiated donor spleen cells (SC) or soluble alloantigens (Ag) induces peripheral tolerance to organ allografts in the rat to the murine cardiac allograft model. In our initial experiment, we showed that IT inoculation of uv-B irradiated SC combined with transient immunosuppression of the recipient with either sublethal TBI or ALS on Day-7 led to donor-specific, long-term cardiac allograft survival (>300 days) in the completely mismatched A/J-to-C57BL/6 mice combination. To test our hypothesis that peripheral tolerance induced by IT injection of donor Ag is dependent on presentation of the foreign MHC molecule by thymic APC to T cell precursors, we examined the effect of IT injection of donor APC-free-soluble Ag inoculum obtained from 3 M KCl extracts of purified MHC class I resting T cells on cardiac allograft survival in the A/J-to-C3H mice combination. The results showed that IT injection of the optimal dose of 500 μg soluble Ag combined with 0.5 ml ALS on Day-7 led to donor-specific permanent graft survival (>200 days). This finding could not be reproduced by intravenous administration of soluble Ag, thus confirming the privileged position of the thymus in tolerance induction. To further define the role of host APC in allorecognition, we studied the presentation of soluble Ag by responder APC in MLR. The results showed that primed T cells obtained from A/J skin graft-sensitized C3H T cells specifically developed alloreactivity to A/J-soluble Ag in MLR. Addition of anti-Ia mAb blocked the proliferative response, thus confirming that primed T cells can respond to soluble Ag presented by responder-type APC. This study suggests that induction of peripheral tolerance by IT inoculation of soluble Ag is reproducible in the murine cardiac allograft model, where tolerance is dependent on the Ag dose and the indirect pathway of Ag presentation. This approach of immunologic manipulation of the adult thymus may have clinical therapeutic applications.