Intrathymic inoculation of donor antigen: an ineffective strategy for prolonging xenograft survival

The aim of this study was to determine whether intrathymic inoculation of the recipient with donor antigen and short-term depletion of peripheral lymphocytes would lead to donor-specific unresponsiveness to rat pancreatic islet xenografts. The results were compared directly with an allograft model to determine whether there were substantial differences in the mechanisms of graft prolongation between allografts and xenografts using an identical conditioning regimen. Streptozotocin-induced diabetic C57B6 mice were injected with up to 0.3 ml of rat anti-mouse lymphocyte serum (ALS) 1 day before intrathymic injection of donor splenocytes. DA rat islet xenografts or Balb/c mouse islet allografts were transplanted 3 days later. ALS depleted CD3(+) and CD4(+) peripheral blood T lymphocytes to less than 5% of values in control mice by 24 h. Median islet xenograft survival (MGS) was 9 days in untreated mice, 28 days in mice receiving 0.2 ml of ALS and 32 days in mice receiving 0.3 mi of ALS alone. Intrathymic injection of 5 x 10(6) DA splenocytes plus 0.2 mi of ALS did not improve islet xenograft survival beyond 28 days. Increasing the intrathymic inoculum to 10(7) DA splenocytes with or without a higher dose of AES (0.3 mi) did not increase MGS beyond 26 days, although 2 out of 18 animals survived beyond 100 days. These long-term surviving mice rejected a second DA rat islet graft in less than 22 days, indicating that tolerance was not achieved. To confirm the efficacy of this treatment regimen in allotransplantation, diabetic C57B6 mice received 10(7) Balb/c splenocytes intrathymically and 0.3 mi of ALS. A Balb/c islet allograft was performed 3 days later. Allograft survival was similar to that of rat islet xenografts with 40% (4 out of 10) of grafts surviving beyond 100 days. In contrast to the xenograft recipients, a second Balb/c islet allograft survived indefinitely, indicating that tolerance was achieved. Histology of the long-surviving allografts showed intact islets with a sparse cellular infiltrate, whereas the long-surviving xenografts (> 100 days) showed a large cellular infiltrate and significant islet destruction. To investigate further the role of the thymus, adult thymectomized C57B6 mice were treated with 0.3 mi of ALS and received a DA rat islet xenograft. The median graft survival was 52 days and no graft survived beyond 80 days, suggesting that peripheral xenoreactive T cells remained after ALS treatment and greater T-cell depletion was necessary to obtain permanent engraftment. These results show that peripheral xenoreactive T cells which remain after profound T-cell depletion are capable of rejecting an islet xenograft despite intrathymic inoculation of donor antigen. The T-cell-mediated xenograft response appears to be stronger and more difficult to suppress than the allograft response using this strategy.