Background-Allograft deterioration is the major obstacle to organ transplantation as a long-term treatment of end-stage heart failure. In this study, we transduced the antioxidant gene, heme oxygenase-1 (HO-1), to heart grafts using a recombinant adeno-associated viral vector (rAAV) in a rat heart transplantation model and investigated its potentiality in prevention of chronic graft deterioration. Methods and Results-rAAV/HO-1 was administered to heart grafts through the coronary arteries during cold preservation. We investigated the expression patterns and activities of transgene, graft survival, graft histomorphology, and relevance of HO-1 expression on graft survival and chronic graft deterioration by itself. Long-term allograft survival can be achieved by rAAV/HO-1-mediated stable transgene expression. The development of graft arteriosclerosis and interstitial fibrosis was prevented in rAAV/HO-1-transduced allografts on day 100. rAAV/HO-1-mediated long-term graft protection was accompanied by remarkable downregulation of the intragraft mRNA level of macrophage migration inhibitory factor, tumor necrosis factor-␣, and transforming growth factor-␤ 1 . Blockage of HO activities by zinc protoporphyrin IX at different posttransplant phases showed that the stable expression of HO-1 is a prerequisite for both survival of grafts and prevention of graft arteriosclerosis.

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Ischemia/reperfusion (I/R) injury is an unavoidable barrier that significantly affects outcome of solid organ transplantation. Here, we establish a protein transduction system to extend graft preservation time and to prevent I/R injury in heart transplantation. We generated a recombinant heme oxygenase-1 (HO-1) protein containing a modified protein transduction domain (PTD). PTD could cross cover cell membrane and carry target molecule to parenchymal cells of cold-preserved heart grafts. The newly generated PTD-HO-1 protein localized mainly in subcellular membrane organelle and nucleus after delivery that significantly prolonged cold preservation of heart grafts. This effect was associated with significantly less endothelial cell activation, less neutrophil and macrophage infiltration in PTD-HO-1-transduced heart grafts after reperfusion as compared with controls. In addition, transduction of PTD-HO-1 protein to heart graft significantly suppressed the I/R injury-associated myocardiocyte apoptosis. The infarct areas of heart graft after I/R injury were significantly reduced after PTD-HO-1 protein treatment. We show here for the first time that PTD can maintain its biological activities during cold preservation. Transduction of cell penetrating HO-1 protein significantly prolongs the cold preservation time and protects the graft from the I/R injury. This approach represents a novel method for the improvement of the overall outcome of organ transplantation.