Background: Perforin is a cytolitic mediator produced by cytotoxic T cells (CD8+ cells) and natural killer cells. We previously reported that ex vivo interleukin (IL)-10 gene therapy induced apoptosis of allogenic infiltrative CD8+ cells, and significantly prolonged the cardiac allograft survival. Methods: To further test the hypothesis that localized IL-10 overexpression in cardiac allografts may also effect the alloreactive CD8+ T cell function by downregulating its Perforin production, a rabbit functional heterotopic allograft heart transplant model was used. Human recombinant IL-10 gene complexed with liposome was intracoronary delivered into the cardiac allografts ex vivo. Results: The percentage of apoptotic infiltrative CD8+ cells in cardiac allografts was increased 6 fold in the gene therapy group (GG) versus that in the control group (CG), while the percentage of Perforin-positive CD8+ cells was decreased 2.9 fold (p<0.01). Perforin expression level in the allograft myocardium of GC was deceased 3.2 fold (p<0.01). The amount of infiltrative Perforin+ CD8+ cells and Perforin expression level were inversely correlated with IL-10 transgene and protein expression level in the myocardium of the cardiac allografs (p<0.01), the percentage of apoptotic cardiac myocytes (p<0.01) and the peak left ventricular systolic pressure of the cardiac allografts (p<0.01), but significantly correlated with the infiltrative T cell cytotoxicity (p<0.01) and the allograft rejection score (p<0.01). Conclusion: These results suggest that localized IL-10 gene therapy prolongs cardiac allograft survival at least in part, through downregulating Perforin production by activated allogenic CD8+ T cells. Reduction of cytolitic function of cytotoxic effector cells prevents the apoptosis of cardiac myocytes.