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• W2016030595 abstract "Objective: To optimize transgene expression levels after AAV-mediated gene transfer different delivery methods were compared in a rat (A) and porcine (B) heterotopic heart transplantation model. Methods: (A) Heterotopic abdominal heart transplantations were performed in male Lewis rats. After harvesting the donor hearts, the viral vectors were delivered to the graft by the following methods: (1) 0.35 ml saline solution containing AAV2/9-LacZ (2 × 1011 vector genome, vg) was injected directly into the myocardium (apex) immediately after reperfusion. (2) cardioplegic solution (0.3 ml) containing AAV-2(HBSD), 2/9-LacZ vectors was rapidly injected into the aortic root with the pulmonary trunk clamped. Before transplantation the transfected heart was incubated for 20 min in iced cardioplegia. (3) A reperfusion system was applied: For 20 min a cold solution of cardioplegia (5 ml) and AAV-2(HBSD) or AAV2/9-LacZ vectors were recirculated through the donor heart. Transplanted grafts were explanted after 3 weeks. To detect and measure marker gene expression X-gal staining or a luciferase assay was performed. In a second series we compared the effects of the transduction of PD-L1 to LacZ using the optimum method (intraaortic root injection) in the same heart transplantation model. (B) Heterotopic abdominal HTX was performed in pigs (Landrace, 10–18 kg) following vector application to the donor heart in an in situ-Langendorff perfusion system (AAV2/GFP and AAV2/Luciferase). Recipients were given tacrolimus (0.3 mg/kg BW), after 21 days the transplanted hearts were explanted for transgene expression analysis. In a second series we compared AAV2/9-mediated transduction of PD-L1 and LacZ in the in situ-Langendorff model and consequent allogeneic heterotopic abdominal heart transplantation. Results: (A) Highest transfection efficiency was observed in the grafts treated with intracoronary infusion of AAV2/9 at the higher dosage, and the expression pattern was global and homogenous in the grafts. hPD-L1 transduction resulted in no significant difference of survival time and signs of rejection after allogeneic rat heart transplantation. (B) AAV2-mediated gene delivery was unable to yield sufficient transgene expression after in situ-Langendorff perfusion of porcine hearts. AAV2/9 based gene transfer of LacZ and hPD-L1 led to excellent myocardial gene expression lasting up to 2 months. Due to species incompatibility no protective effects were observed in our allogeneic porcine transplantation model. Conclusions: (A) We demonstrated that infusion of AAV vectors into the aortic root with the pulmonary trunk clamped is a simple and efficient method for gene delivery to the donor heart in an allogeneic rat heart transplantation setting. Gene transfer of hPD-L1 was ineffective to protect against allorejection, on the contrary there was a trend to aggravated rejection. Therefore the exact mechanism of hPD-L1 in allograft rejection needs further investigation. (B) The in situ-Langendorff model was developed to allow isolated target organ perfusion with high vector concentrations under physiological conditions. Using this method AAV2/9 mediated gene delivery into porcine hearts proved effective to induce excellent marker gene expression. Expression of hPD-L1 could also be achieved but was ineffective in pig allotransplantation due to species incompatibility. Double transgenic pig hearts (e.g. Gal-KO+CD46) can now efficiently be transduced with hPD-L1 or hCTLA4-Ig to further optimize long-term survival after pig-to-primate cardiac xenotransplantation." @default.
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• W2016030595 date "2010-03-01" @default.
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• W2016030595 title "AAV based gene delivery to myocardium in rodents and pigs" @default.
• W2016030595 doi "https://doi.org/10.1111/j.1399-3089.2010.00573_13.x" @default.
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