Background—Two populations of T cells contribute to allograft rejection. T cells with direct allospecificity are activated after recognition of intact MHC alloantigens displayed at the surface of donor passenger leukocytes carried within the graft. In contrast, T cells with indirect allospecificity recognize donor alloantigens as processed peptides associated with self (recipient)–MHC class II molecules. In small animal models of transplantation, direct pathway T cells dominate the acute rejection process and are rendered tolerant to the graft after the loss of donor passenger leukocytes. It has been argued that indirect pathway T cells contribute substantially to continual graft damage after passenger cell loss. The purpose of this study was to determine whether donor-specific tolerance could be detected in T cells with direct anti-donor allospecificity in human heart transplant recipients after prolonged graft residence.

Methods and Results—Alloreactive helper (HTLf) and cytotoxic (CTLf) T cells were enumerated by use of limiting dilution analysis. These assay systems were refined to make them specific for the direct pathway of allorecognition and more sensitive in the case of the HTLf assay. Recipient:anti-donor frequencies were generated in 10 long-term recipients of heart grafts with progressive chronic rejection and compared with those against equivalently HLA mismatched recipient:third-party controls. For HTLf, direct pathway donor-specific hyporesponsiveness was detected in 5 of the 10 recipients (HTLf <1:100 000). Of these 5 recipients, 4 also had low anti-donor CTLf (<1:100 000). In the 5th recipient, although the CTLf was >1:100 000, it was significantly lower than that estimated against the third-party control.

Conclusions—Donor-specific hyporesponsiveness is demonstrated in 50% of recipients in both the HTLf and CTLf compartments of the direct alloresponse. Direct allorecognition therefore appears unlikely to be responsible for the progression of chronic rejection, implicating indirect allorecognition as the predominant immunological driving force. Furthermore, these data have potential implications for graft outcome, adjustment of immunosuppression, and recipient monitoring.