Homeostatic Expansion and Repertoire Regeneration of Donor T Cells during Graft Versus Host Disease Is Constrained by the Host Environment.

Graft versus host disease (GVHD) typically results in impaired T cell reconstitution characterized by lymphopenia and repertoire skewing. One of the major causes of inadequate T cell reconstitution is that T cell survival and expansion in the periphery is impaired. In this report, we performed adoptive transfer studies to determine whether the quantitative reduction in T cell numbers is due to an intrinsic T cell defect or whether the environmental milieu deleteriously affects T cell expansion. Irradiated (750 cGy) AKR/J (H-2^k) recipients were initially transplanted with C57BL/6(H-2^b) BM plus B6 spleen cells to induce GVHD. Lethally irradiated (1100 cGy) secondary AKR mice were then transplanted with B6 Rag-1 BM alone or together with equivalent numbers of T cells from either normal B6 or day 19 GVHD (B6→AKR) chimeras. Whereas the majority of animals transplanted with B6 naïve T cells died from GVHD (31% survival by day 90), mice transplanted with splenic T cells from GVHD (B6→AKR) mice had superior survival (p<0.0001) that was not significantly different from mice reconstituted with Rag-1 BM alone (p=0.35), indicating that re-exposure of splenic T cells from GVHD animals to host APCs/host antigens did not effectively transfer the disease. The ability of these T cells to undergo homeostatic expansion in the absence of a GVHD environment was then assessed in secondary animals. We observed that GVHD T cells were capable of five-fold expansion in secondary recipients, in contrast to animals transplanted with naive T cells that had severe T cell hypoplasia and no T cell expansion. The CD4:CD8 ratio that was inverted in primary GVHD animals (0.3) prior to adoptive transfer also normalized in secondary recipients (2.1 ± 0.2). This resulted in a 7-fold expansion of CD4⁺ T cells and 3.5 fold expansion of CD8⁺ T cells in animals transplanted with T cells from GVHD mice compared to lethally irradiated animals reconstituted with naïve T cells. The ability of GVHD T cells to regenerate a complex T cell repertoire was then assessed using Vβ TCR spectratyping. T cells from primary GVHD animals demonstrated a high degree of repertoire skewing with many Vβ families [i.e ~90% (96/106)] showing only a few intense bands. In contrast, after transfer of these T cells into secondary AKR recipients, there was substantial improvement in the T cell repertoire. Whereas a complex repertoire was observed in only 10% of Vβ families in primary GVHD animals, approximately 61% of Vβ families analyzed in splenic T cells from secondary recipients had evidence of reconstitution that was either partial or complete after 90 days. To determine the rapidity with which animals were able to generate complex repertoires, peripheral blood samples that had been serially obtained 38 and 66 days post-BMT were analyzed by TCR spectratyping. Forty percent of the Vβ families had normal complex distributions by day 38, while 65% were complex by day 66, indicating that T cells from GVHD mice had the capacity to develop complex repertoires early post-BMT. These data indicate that T cells from mice with GVHD can respond to homeostatic signals in the periphery and are not intrinsically compromised once they are removed from the GVHD environment. We thereby conclude that the host environment and not an intrinsic T cell defect is primarily responsible for the lack of effective T cell expansion and diversification of complex T cell repertoires that occurs during GVHD.