test this, BALB/c (H-2 d ) were transplanted with allogenic grafts from FVB transgenic mice (b-CM Tg) (H-2 q ) whose hearts are expressing b-CM in 85% in adult life. No immunosuppression was used. Graft function was monitored by palpation; rejection was confirmed histologically (H&E, EVG). Allo- and CM-specific T cell response was determined by ELISPOT; CM-specific ELISA measured anti-CM autoantibodies in recipient’s sera. Control littermates donor hearts were all rejected within 7 days. Conversely, b-CM Tg grafts enjoyed long-term graft survival (100 days). Long-term survival of b-CM Tg grafts was associated with decrease in fre- quency of inflammatory IFNg-producing allo- and a-CM-specific host T cells, and with expansion of IL-5-producing T cells. We also detected significant increase in titres of anti-CM antibodies in recipients of b-CM Tg hearts when compared to their controls (p=0.009). Our data show, that overexpression of b-CM in donor heart results in significant prolongation of cardiac allograft sur- vival, even in fully mismatched combination. Long-term survival of b-CM allografts is accompanied by alterations in host anti-graft immunity, and suggests involvement of regulatory immune mech- anisms associated with neonatal forms of cardiac myosin. 264 TOLERANCE TO CARDIAC ALLOGRAFTS IN MINIATURE SWINE USING DONOR-SPECIFIC TRANSFUSIONS COMBINED WITH CYCLOSPORINE BUT NOT WITH RAPAMYCIN R. Hoerbelt, 1 T. Shoji, 1 A. Muniappan, 1 D.A. Guenther, 1 S.L. Houser, 1 J.S. Allan, 1 M. Bravard, 1 D.H. Sachs, 1 J.C. Madsen, 1 1 Surgery, MGH, TBRC, Charlestown, MA Purpose: To evaluate the effect of pretransplant donor-specific transfusion (DST)combined with CyA and rapamycin on tolerance induction to cardiac allografts in a large animal model. Methods: Heterotopic cardiac transplants were performed across a major MHC class I barrier in MHC-inbred miniature swine. Experi- mental animals were treated with two DSTs, each containing 1.4x10 8 viable PBMC, on POD -14 and -7. All animals received CyA or rapamycin (RPM) from POD 0 until POD 11. Control groups received CyA or RPM alone, DST alone or no treatment. CML assays were performed at regular intervals. T cell priming and activation induced cell death were assessed by CFSE proliferation assays and Annexin V staining. Results: Untreated (n=2) and DST-only (n=2) treated control animals rejected between POD 6 and 8. Animals treated with CyA (n=3) or RPM (n=3) alone exhibited graft survival to 53, 52 and 59 days or 41, 50 and 53 days, respectively. In contrast, DST- pretreatment combined with CyA (n=3) led to stable graft func- tion for 200 days in all animals. Grafts in recipients treated with pretransplant DST and RPM (n=4) survived 52, 45, 100 and 200 days. Long-term acceptors in the DST+CyA group and DST+RPM group mostly maintained peripheral CML anti- donor and -3rd party response. Following DSTs, the donor-specific proliferative re- sponse of CD8+ recipient T cells was significantly increased on CFSE assay (p=0.011), and a significant number of recipient CD8+ T cells underwent apoptosis (10.1% on POD 0 versus 5.2% on POD –14; p=0.04). Conclusion: The combination of DST and a short course of CyA consistently prolonged survival of cardiac allografts in miniature swine. The sustained anti-donor CML response and activation of alloreactive T cells by the DST suggest an active regulatory mecha- nism. In contrast, RPM did not demonstrate with the tolerogenic effect of pretransplant DST potentially because it failed to achieve appropriate regulatory cells. 265 HEART AND EN-BLOC THYMUS TRANSPLANTATION IN CYNOMOLGUS MONKEYS A. Muniappan, 1 D.R. Johnston, 1 D.A. Guenther, 1 T. Shoji, 1 S. Boskovic, 1 R. Hoerbelt, 1 S.L. Houser, 1 J.S. Allan, 1 K. Yamada, 1 T. Kawai, 1 J.C. Wain, 1 D.H. Sachs, 1 J.C. Madsen, 11 Transplantation Biology Research Center, Massachusetts General Hospital, Boston, MA Purpose: To determine whether cotransplantation of a whole vascu- larized donor thymic lobe can prolong cardiac allograft survival in cynomolgus monkeys. Background: We have previously shown that transplantation of vascularized donor thymic tissue can prolong cardiac allograft sur- vival in miniature swine. To test this modality in primates and, at the same time, test a procurement technique applicable to human cadaveric heart transplantation, we have developed a heart and en-bloc thymus transplant procedure for cynomolgus monkeys. Methods: The right mediastinal thymus was harvested en-bloc with the heart, preserving the entire arterial supply and venous drainage to the thymus. The heart and en-bloc thymus grafts were transplanted heterotopically into fully MHC mismatched cynomolgus monkeys. All recipients were treated with rabbit anti-thymocyte globulin at the time of transplant, and daily FK506 for 28 days. Some recipients were thymectomized on the day of transplant. Results: Isolated cardiac allograft survival was 40 and 61 days in euthymic recipients, and 62 days in a thymectomized recipient. Heart and en-bloc thymus allograft survival was 158 and 48 days in euthymic recipients, and 127 days in a thymectomized recipient. Acute rejec- tion was eventually seen in all grafts, and all recipients maintained donor specific responsiveness by MLR. Conclusions: Heart and en-bloc thymus transplantation is a clinically feasible technique which transfers large volumes of donor thymus to the recipient at the time of heart transplantation. Our obligate use of older monkeys, with limited thymic mass, may have prevented tolerance in this set of experiments. We hypothesize that the use of younger animals, along with more effective T cell depletion, may facilitate establishment of cardiac allograft tolerance. This is the first report of heart and en-bloc vascularized thymus transplantation in a primate model. 266 CYTOTOXIC HYPORESPONSIVENESS IS ASSOCIATED WITH HIGH IL-10 mRNA EXPRESSION IN IMMUNOSUPPRESSED CARDIAC ALLOGRAFT PATIENTS E. Dijke, 1 J. Velthuis, 1 A. Balk, 2 S. Korevaar, 1 L. Maat, 2 W. Weimar, 1 C. Baan, 11 Internal Medicine, Erasmus MC; 2 Thorax Center, Erasmus MC, Rotterdam, Netherlands At present, transplant patients are prescribed immunosuppressive drugs to prevent rejection. However, this is accompanied by severe side-effects. Identification of allograft recipients who do not need full dosage of immunosuppressive medication is therefore required. In the present study, we analysed the donor-specific proliferative and cytotoxic T cell responses to asses whether cardiac allograft patients (n=18) have developed donor-antigen specific hyporesponsive- ness  1 year after transplantation (median 4 y; range 1 1 /2 - 7 y). In addition, we analysed the mechanisms controlling immune responses e.g. Foxp3+/GITR+ cells and IL-10 producing cells. All patients were treated with CNI, anti-proliferative agents and corticosteroids. In the presence of an intact proliferative response against donor- antigens, no IL-2 or IL-15 driven cytotoxic responsiveness (CTLpf  10/10 6 PBMC) could be measured in a number of allograft patients (IL-2: 6/18 and IL-15: 16/18, respectively). Overall, the cytotoxic response against donor cells was significantly lower than against S128 Abstracts The Journal of Heart and Lung Transplantation February 2005