Transfection of pulmonary artery segments in lung isografts during storage.
BACKGROUND
Proximal pulmonary artery segment (PPAS) endothelial transfection of lung grafts may be useful in ameliorating ischemia-reperfusion injury and rejection and may provide beneficial downstream effects on the whole lung graft. Transfection immediately after lung transplantation may be efficacious in ameliorating allograft dysfunction after transplantation.


METHODS
In F344 rats, the PPAS was isolated and injected with 0.03 mL of GL-67/DOPE-chloramphenicol acetyl transferase (CAT) plasmid DNA. The PPASs were exposed for 60 minutes at several temperatures. The lung grafts were stored in saline solution (group 1, n = 24) or LPDG solution (group 2, n = 27) for 12 or 24 hours at 4 degrees to 37 degrees C. In group 3 (n = 42), PPASs were stored in endothelial cell culture medium and incubated at 10 degrees or 37 degrees C in a carbon dioxide incubator for 3 to 72 hours. Group 4 (n = 18) served as transplanted controls; after 3 to 24 hours' preservation at 4 degrees C in LPDG solution, lung grafts were transplanted. Transgene expression of PPASs was assessed with two CAT activity assays, thin-layer chromatography enzyme-linked immunosorbent assay and immediately after the preservation period (groups 1 to 3) or 24 hours after transplantation (group 4).


RESULTS
In group 1, transgene expression did not appear. In groups 2 and 3, transgene expression was apparent after any storage duration at 37 degrees C. Transgene expression increased successively with longer storage periods. In group 4, transgene expression was detected after any storage duration. The enzyme-linked immunosorbent assay is able to quantify the expression of CAT activity, but thin-layer chromatography is more sensitive.


CONCLUSIONS
Transgene expression did not occur during conventional cold storage. Transgene expression in rat PPASs during storage is possible with warm storage (37 degrees C) and appropriate storage solution.
Yano
M
M
Division of Cardiothoracic Surgery, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Hiratsuka
M
M
Mora
B N
BN
Scheule
R K
RK
Patterson
G A
GA
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression / Q000502:physiology; D018014:Gene Transfer Techniques; D016040:Lung Transplantation / Q000473:pathology*; D008297:Male; D009926:Organ Preservation; D011651:Pulmonary Artery / Q000473:pathology / Q000637:transplantation*; D051381:Rats; D011916:Rats, Inbred F344; D014162:Transfection / Q000235:genetics*; D019076:Transgenes / Q000235:genetics; D014185:Transplantation, Isogeneic
D015500:Chloramphenicol O-Acetyltransferase