EFFICIENT GENE-TRANSFER METHOD INTO THE WHOLE HEART THROUGH THE CORONARY-ARTERY WITH HEMAGGLUTINATING VIRUS OF JAPAN LIPOSOME

Objective: To confirm gene transfer techniques especially into the who le heart, we tried out a gene transfer method involving liposome with the viral envelope hemagglutinating virus of Japan liposome as an alte rnative to existing techniques such as cationic lipofection or other v iral vectors, Method: For this study, hemagglutinating virus of Japan liposome (H group) or cationic liposome(L group) was used to compare t he efficacy of gene transfection of oligonucleotide labeled with fluor escein isothiocyanate and cDNA of beta-galactosidase and human mangane se-superoxide dismutase, Fluorescein-labeled oligonucleotide, cDNA of beta-galactosidase, or manganese-superoxide dismutase was complexed wi th liposomes, DNA-binding nuclear protein, and the viral protein coat of hemagglutinating virus of Japan, After donor rat hearts arrested by cardioplegia had been harvested, the coronary artery during cardiople gic arrest was infused via an aortic cannula with the liposome-gene co mplex, Next, the hearts were transplanted into the abdomen of recipien t rats of the same strain, and all recipients were put to death after 3 days of transfection. Results: Fluorescein isothiocyanate was detect ed in the nuclei of more than 70% of the myocytes (75% +/- 14%, n = 5) in the Fl group compared with fewer than 10% in the L group (7% +/- 5 %, n = 5), The intensity of fluorescein isothiocyanate was significant ly higher in the H group (979 +/- 112 FI) than in the L group (116 +/- 68 Fl), beta-Galactosidase was expressed in the cytosol of more than 50% of the myocytes in the H group (61% +/- 7%, n = 5) compared with n one in the L group (0%, n = 5), After 3 days of gene transfection, and when exposed to ischemia (30 minutes, 37 degrees C) and reperfusion ( 30 minutes, 37 degrees C) with Langendorff apparatus, the hearts trans fected with manganese-superoxide dismutase (S group, n = 5) showed a s ignificantly higher percentage of recovery of left ventricular end-dia stolic pressure (S vs C, 86% +/- 3% vs 54% +/- 12%) and coronary flow (98% +/- 2% vs 66% +/- 12%) than did the control hearts (C group, n = 5), Western blotting analysis showed an apparent increased expression of manganese-superoxide dismutase in the hearts transfected with manga nese-superoxide dismutase compared with the control hearts, These resu lts clearly demonstrated that the donor hearts were transfected with f luorescein-labeled oligonucleotide and the beta-galactosidase gene as a result of coronary infusion of the hemagglutinating virus of Japan l iposome during cardioplegic arrest at the time of harvest, Furthermore , the hearts transfected with manganese-superoxide dismutase showed si gnificant improvement in tolerance against ischemiareperfusion injury. Conclusion: We believe that this method represents a novel in vivo ge ne transfer technique for the heart and thus may provide a new tool fo r research and therapy of heart transplantation.