Ex vivo transfection of pulmonary artery segments in lung isografts

Background. Gene transfer to lung grafts may be useful in ameliorating isch emia-reperfusion injury and rejection. Proximal pulmonary artery endothelia l transfection may provide beneficial downstream effects on the whole lung graft. We have already demonstrated the feasibility of in vivo and ex vivo transfection in proximal pulmonary artery segments of rat lung grafts. The aim of this study was to determine the optimal conditions for and duration of transfection. Methods. Orthotopic left lung transplantation was performed in F344 rats af ter donor lung proximal pulmonary artery segments were isolated and injecte d with lipid 67/DOPE-chloramphenicol acetyl transferase (CAT) complementary deoxyribonucleic acid construct. Effect of exposure time was studied by ex posing donor pulmonary artery segments to the construct for 0, 30, and 60 m inutes prior to transplantation. In another series of experiments, pulmonar y artery segments were exposed to the construct for 60 minutes prior to tra nsplantation. Onset and duration of gene expression were determined after s acrificing animals at 3, 6, 12, and 24 hours and 3 days as well as 1 week, 2, 4, and 8 weeks after transplantation. Effect of exposure temperature was studied by exposing pulmonary artery segments to the construct for 60 minu tes at 4 degrees, 10 degrees, and 23 degrees C. These recipients were sacri ficed on postoperative day 3. Effect of exposure pressure was studied by us ing two volumes of the construct (0.01 and 0.03 mt). These recipients were sacrificed on postoperative day 3. Transgene expression was assessed by chl oramphenicol acetyl transferase activity assay. Results. Transgene expression was similar after 30- and 60-minute exposure. Transgene expression was evident within 3 to 6 hours after operation and p ersisted at 8 weeks after operation. Expression was detected at all tempera tures and was equivalent at both exposure pressures. Conclusions. Gene transfection into graft pulmonary artery segments is poss ible under a range of conditions applicable to clinical lung transplantatio n. (C) 1999 by The Society of Thoracic Surgeons.