799 The transfer of genes encoding immunosuppressive agents holds promise as an inductive therapy in transplantation. It has been suggested that gene therapy would provide local, transient immunosuppression thereby avoiding the side affects of systemic immunosuppression. However, fundamental questions regarding the immunobiology of immunosuppressive gene therapy, the suitability of viral and non-viral vectors for transfecting allografts, and the control of transgene expression must be addressed before gene transfer based therapies may be applied to clinical transplantation. In the current study, mouse vascularized cardiac allografts were perfused with DNA-liposome (γ-DLRIE/DOPE) complexes or adenoviral vectors encoding the active form of TGFβ1 as a model in which to study these aspects of suppressive gene therapy. DNA-liposome mediated transfection with TGFβ1 resulted in prolonged allograft survival in approximately 2/3 of transplant recipients and was associated with markedly reduced Th1 and alloantibody responses. However, TGFβ1 transfected grafts were eventually rejected, and loss of graft function was associated with a unique pathology characterized by an aberrant deposition of collagen and fibrous material within the graft. Despite an intense mononuclear cell infiltration, overt myocyte necrosis was not apparent, further suggesting that loss of graft function was not associated with "classical" immune effector mechanisms. In contrast, adenoviral mediated transfection with TGFβ1 did not delay graft rejection, nor did it inhibit Th1 development and alloantibody responses. Preliminary data indicate that cytokines induced by adenoviral vectors (but not DNA-liposome complexes) over-ride the suppressive activities of TGFβ1. Interestingly, in vivo depletion of CD8+ T cells markedly prolonged survival (>60 days) of allografts transfected with either DNA-liposome complexes or adenoviral vectors, indicating that CD8+ T cells are less susceptible to TGFβ1 gene transfer than are CD4+ cells. Prolonged graft survival in CD8 depleted mice was associated with persistent transgene expression (>60 days) and continued inhibition of Th1 activation. This study documents the utility of DNA-liposome complexes for gene transfer into vascularized allografts and demonstrates that transgene expression is persistent rather than transient. Further, these data indicate that CD4+ and CD8+ T cells exhibit distinct sensitivities to TGFβ1 gene transfer and suggest that transient depletion of CD8+ cells may augment immunosuppressive gene therapies.