Role of intrathymic clonal deletion and peripheral anergy in transplantation tolerance induced by bone marrow transplantation in mice conditioned with a nonmyeloablative regimen.

Abstract

We have investigated the mechanism of tolerance induced by allogeneic bone marrow transplantation (BMT) in mice conditioned with a nonmyeloablative regimen. Permanent mixed chimerism and skin allograft tolerance were achieved when recipient B10 (H-2b) mice were pre-treated with anti-CD4 and anti-CD8 mAbs, thymic irradiation, and 3 Gy whole body irradiation, followed by injection of B10.A (H-2a) bone marrow cells. Vbeta11+ T cells are normally deleted in I-E+ B10.A mice, but not in I-E- B10 mice. In spleens of mixed B10.A-->B10 chimeras, Vbeta11+ B10 T cells were reduced but detectable in the first 6 wk after BMT, and later became undetectable. Splenic Vbeta11+ T cells were unresponsive to receptor cross-linking with Vbeta11-specific mAb, demonstrating anergy. B10 Vbeta11+ T cells were also detected in spleens of mice that were thymectomized before BMT. In PBL of chimeras, Vbeta11+ T cells were undetectable for at least 1.5 yr. Thymocyte chimerism and extensive clonal deletion of mature Vbeta11+ B10 thymocytes were observed as early as 10 days post-BMT and at all subsequent time points in thymi of mixed chimeras. Rare I-E+ donor cells were detected in day 10 thymi, and these increased progressively in frequency at later times. In chimeras prepared in the reciprocal strain combination, B10-->B10.A, B10 donor Vbeta11+ T cells were undetectable in PBL. Together, our results implicate both central clonal deletion and peripheral clonal anergy in tolerance induced with our regimen. A very low number of intrathymic donor cells or low density of Ag expression is sufficient to mediate extensive clonal deletion of thymocytes that recognize donor Ags.