A Novel And Unexpected Role Of Donor T Cells In The Efficacy Of Tolerogenic Therapies For Transplant Rejection
Marcos Iglesias Lozano, DVM, PhD, Darrel Bibicheff, BS, Maria Chicco, MD, Gerald Brandacher, MD, Giorgio Raimondi, PhD.
Johns Hopkins University, Baltimore, MD, USA.
Costimulation blockade-based regimens are a promising immunomodulatory strategy to prevent transplant rejection and promote lasting tolerance. However, their efficacy is affected by multiple factors, many of which remain unknown. Recently, some reports have highlighted the unexpected capacity of passenger donor lymphocytes to directly fuel the recipient's anti-graft response. Further studies are then necessary to understand their possible role in settings of regulation of alloreactivity. In this study we aimed to assess if T lymphocytes contained in the donor specific transfusion (DST) inoculum, used as part of a very effective CoB regimen, contribute to the observed limited regulation of skin transplant rejection.
Full mismatch skin transplants were performed using dorsal skin from Balb/c into C57BL/6 mice and animals received a peri-transplant regimen based on DST, (107 splenocytes; on day 0) and anti-CD154 mAb (MR-1; on day 0, 7, 14). To study the role of donor T cells, DST inocula were depleted in either total T cells, CD8, or CD4 T cell subpopulations by negative-selection. IgG Donor Specific Antibodies (DSA) in serum of transplanted animals was determined by flow cytometry, and T cell-IFNg production to determine the strength of recipients alloresponse, measured by ELISpot.
A peri-transplant regimen based on DST+anti-CD154 (MR-1) has a profound protective effect on mouse skin allotransplantation. However, it does not induce long term survival (MST=58 days). When donor T cells were depleted from the splenocytes used as DST, the beneficial effect was statistically increased (MST=105 days). We studied then the specific role of donor CD4 and CD8 T cell by performing two additional transplant groups receiving DST depleted of CD4 or CD8 T cells respectively. The absence of CD4 abrogated the prolongation of survival observed with the total-T cell depletion in the DST, bringing survival back to a value, MST=64, comparable to that of unmanipulated DST. This suggested a deleterious role for donor CD8 T cells. Unexpectedly, the absence of donor CD8 T lymphocytes induced a remarkable improvement in transplant survival, with 65% of the grafts surviving beyond 180 days. In ongoing experiments, we are aiming to determine the role of donor memory CD4/CD8 T cells in the effect observed and what correlation exists between the presence/absence/type of donor T lymphocytes and the levels of donor specific antibodies, or variations in the strength of the direct and indirect alloresponses.
Overall, these data reveal the existence of a novel and very important opposing role for donor passenger lymphocytes in the modulation of recipient's alloimmunity by DST+MR1 based regimens: a deleterious role for donor CD8, and a beneficial one for CD4 T cells. Identification of the specific mechanisms through which these divergent modulations of the anti-donor alloresponse are exerted will be pivotal for the optimization of clinically effective tolerogenic therapies.
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