B. Kruse

Preoperative low-dose irradiation promotes long- term allograft acceptance and induces regulatory T cells in a porcine model of pulmonary transplantation

Background. A simplified conditioning protocol including single-dose preoperative thymic and low-dose whole body irradiation with or without subsequent donor bone marrow transplantation (BMTx) can be applied in porcine lung transplantation. We hypothesized that this protocol would prolong allograft survival. Methods. Left-sided single lung transplantation from major histocompatibility complex (MHC)-mismatched donors was performed in 27 minipigs. Recipients received whole body (1.5 Gy) and thymic irradiation (7 Gy) before transplantation (IRR group, n=6), intravenous immunosuppression with methylprednisolone, cyclosporine, and azathioprine for 27 postoperative days (IS group, n=5) or both (IRR+IS group, n=10). BMTx group recipients were treated with irradiation, immunosuppression and a donor bone marrow infusion on postoperative day 1. Peripheral blood leukocyte phenotype and donor cell chimerism were monitored by flow cytometry. Purified CD25+ T cells from long-term survivors or rejecting animals were used for in vitro MLR suppression assays. Results. Median graft survival was: IRR 12 days, IS 55 days, IRR+IS 239 days, and BMTx 80 days (P<0.0001). Early peripheral blood macrochimerism was substantial in both the IRR+IS and the BMTX group but was lost in all groups after day 80. The frequency and suppressive function of CD4+CD25+ T cells were enhanced in IRR+IS group long-term survivors. Conclusion. Although donor bone marrow infusion was not beneficial in our model, a substantial proportion of the animals treated with irradiation and a 28-day course of immunosuppression accepted their lung allografts long term. The mechanism involved in maintaining allograft tolerance may be based on peripheral T-cell regulation.

Postoperative intravenous infusion of donor-derived transplant acceptance-inducing cells as an adjunct immunosuppressive therapy in a porcine pulmonary allograft model

There is very limited published information testifying to the safety and possible complications of cell‐based therapies. Accurately assessing the potential risks of translating novel, cell‐based immunosuppressive protocols into clinical trials is therefore extremely difficult. This report describes the use of a pulmonary allograft model in outbred miniature pigs as a preliminary step in the development of a safe, clinically feasible, cell‐based immunosuppressive protocol. Single lung transplants were performed in 22 MHC Class I‐mismatched donor–recipient pairs, which were randomized between four treatment groups. For the first 28 days postoperatively, all animals were immunosuppressed with methylprednisilone and tacrolimus, with or without preoperative irradiation; subsequently, pharmacological immunosuppression was stopped in all treatment groups. Animals in two groups also received a central venous infusion of donor‐derived ‘transplant acceptance‐inducing cells’ (TAICs) on the seventh and 14th days postoperatively. Allograft survival was monitored by sequential chest X‐rays, bronchoscopies and transbronchial biopsy histologies. No acute adverse events were associated with the administration of TAICs and there was no evidence of accelerated graft rejection. The observations presented in this report represent an important first step towards the development of a clinically applicable protocol for the use of TAIC therapy in lung transplantation.

Correlation of donor leukocyte chimerism with pulmonary allograft survival after immunosuppressive drug withdrawal in a porcine model.

Background. This study was designed to analyze the role of postoperative donor cell chimerism for the induction and maintenance of transplantation tolerance in a porcine lung transplantation model. Methods. Left-sided single lung transplantation from major histocompatibility mismatched male donors was performed in 27 female minipigs. All received a 28-day course of pharmacologic immunosuppression using various agents, some in combination with preoperative irradiation. Groups for eventual analysis were strictly defined by outcome, that is, pigs with acute rejection before postoperative day 178 (n=16) were allocated into one group, long-term surviving animals (n=11) into the other. Peripheral blood chimerism was monitored by flow cytometry and real-time polymerase chain reaction. Intragraft chimerism was detected from bronchoalveolar lavage fluid (BALF) by fluorescent in situ hybridization. Results. Blood chimerism peaked 1 hour after transplantation and was significantly higher in the group of long-term survivors at that time. Thereafter chimerism rapidly decreased, but tended to remain higher in long-term survivors. In case of acute rejection donor cells were lost, but remained detectable for up to 36 postoperative months in tolerant animals. In BALF, the percentage of male nuclei was equally high under immunosuppression in both groups. Rejecting animals showed a rapid decrease of Y-bearing cells in BALF after drug withdrawal and an almost complete loss when acute rejection occurred. In tolerant pigs, intragraft chimerism remained detectable throughout the follow-up. Conclusions. This study demonstrates a clear correlation of donor leukocyte chimerism with long-term allograft survival in a porcine allogeneic lung transplantation model.

ALLOANTIGEN REQUIREMENTS FOR THE INDUCTION AND THE MAINTENANCE OF PERIPHERAL TOLERANCE FOLLOWING PORCINE LUNG TRANSPLANTATION: 1938

G. Warnecke1, K. Dreckmann2, M. Avsar2, A. Knöfel3, N. Madrahimov1, W. Sommer1, B. Kruse1, S. Thissen1, J. Karstens1, A.R. Simon1, A. Haverich4, M. Strüber1 1, Hannover Medical School, Hannover/GERMANY, 2Department Of Cardiothoracic, Transplantation And Vascular Surgery, Hannover Medical School, Hannover/GERMANY, 3Httg, Hannover Medical School, Hannover/GERMANY, 4Heart Transplant Surgery, Medical School Hannover, Hannover/GERMANY