Innate- and adaptive-like features of human γδ T cells are associated with Vγ9+δ2+ and non-Vγ9δ2 clonotypes, respectively. Despite their presence in the blood and many organs, the role of γδ T cells in transplantation is unclear.

We performed phenotypic and clonal tracking of donor- and recipient-derived γδ T cells after human intestinal transplantation (ITx) in blood, intestinal graft and bone marrow (BM) by integrating flow cytometry and sequencing platforms.

We previously demonstrated that donor T-cell blood macrochimerism (peak level ≥4%) is associated with less rejection and slower donor T cell replacement by the recipient in the ileal graft. This is due to the migration of graft-derived graft-versus-host (GvH)-reactive donor αβ T cells into the recipient blood and BM to mediate lymphohematopoietic GvH responses (LGVHR) which counteract the host-versus-graft response. Here we found that the level of blood γδ T cell chimerism correlated with total T cell chimerism (n=19). Donor γδ T cells were detected in recipient BM between post-operative day (POD)105–357. Single-cell RNA profiling of BM-infiltrating donor γδ T cells from 3 patients with pediatric donors revealed both Vδ1- and Vδ2-dominant clonotypes with cytotoxic effector T cell (Teff) phenotypes. In one patient, the top-dominant donor clone (Vγ8Vδ1) detected during peak blood T cell chimerism (POD8–20) was also predominant in recipient BM on POD126, with high cytotoxic but low proliferation gene expression. BM-infiltrating donor δ2+ T cells were dominated by sequences with zero N-additions that likely originated during fetal life and were shared across pediatric (n=6), but not adult donors (n=5), suggesting an age-related distribution and migration pattern. With recipient γδ T cells, regardless of macrochimerism status, turnover dynamics were more rapid in patients with younger donors. Graft-repopulating recipient γδ T cells gradually acquired resident-memory (TRM) phenotypes with “private” non-Vγ9δ2 clonotypes. Single-cell profiling of recipient γδ T cells from 2 quiescent and 4 rejecting intestinal grafts late post-Tx were enriched for interchangeable Teff/TRM clusters, with higher cytotoxic Teff gene expression during rejection. In one patient, the top-dominant Vδ2 sequence (mainly Vγ5δ2) in the blood during quiescence was found at low frequencies in early quiescent graft but as the top-dominant sequence in later rejecting graft, suggesting an active exchange between the blood and graft during rejection. TCR distance analysis suggested that this top-dominant sequence unlikely recognizes MICA or CD1d, but has structural similarity with some TCRδs apparently stimulated by cytokines produced by autologous αβ T cells.

Donor γδ T cells may influence LGVHR and blood chimerism, while recipient γδ T cells may affect rejection after ITx.