NEONATAL TOLERANCE IS DEPENDENT UPON THYMIC MECHANISMS

Abstract

490 Introduction: Neonatal tolerance promotes specific long-term acceptance to skin allografts. It is believed that this tolerant state is induced and maintained by the establishment of semi-allogeneic (F1) lymphoid chimerism, which effectively silences potentially allo-reactive T-cells. However, the role of the thymus in this model has not been clearly defined, and it has been suggested that the observed unresponsiveness may be the result of non-professional antigen presentation to peripheral naive T-cells, leaving them functionally tolerant. To determine if neonatal tolerance is thymus dependent, we employed a technique of vascularized thymus transplantation which has been shown to successfully reconstitute T-cell function in thymectomized hosts. Methods: Neonatal chimeras were created by injecting newborn Lewis rats with a mixture of Lew/ACI F1 splenocytes and bone marrow cells. Chimerism was documented by FACS. Successful chimeras were used as thymus donors, and splenocytes from each thymus donor were tested for allo-response by MLR. Thymectomized Lewis recipients underwent lethal irradiation followed by reconstitution with T-cell depleted syngeneic bone marrow prior to vascularized thymus transplantation. Thirty days after thymus transplantation in vitro tolerance was measured by MLR, whilein vivo tolerance was assessed by challenge with an ACI heterotopic cardiac allograft. Simultaneously, recipient T-cell reconstitution was determined by FACS. All animals with long-term surviving ACI allografts received third party BN cardiac grafts. Results: Chimeric thymus recipients demonstrated normal syngeneic T-cell reconstitution, MLR unresponsiveness to both parent stains with strong proliferation to an unrelated third party (BN), were tolerant to ACI allografts (mean survival> 100 days, n=6), and acutely rejected BN grafts (mean survival = 14.5 days). Tolerance and rejection were confirmed histologically. ACI cells were not detected by FACS or PCR in extrathymic tissue of the thymus recipient.Conclusion: Tolerance to cardiac allografts can be successfully induced by vascularized chimeric thymus transplantation. This tolerance is characterized by T-cell unresponsiveness to alloantigen using both in vivo and in vitro testing, and is highly specific as shown by normal response to unrelated third party antigens. The absence of detectable ACI T-cells in recipient peripheral tissues argues against explanations based upon peripheral tolerance or suppressor mechanisms. These results suggests that neonatal tolerance is primarily dependent upon intrathymic processes which direct T-cell maturation, rather than a state of functional peripheral tolerance or anergy.