Optimization of delivery of adeno-associated virus mediated gene transfer to a transplanted heart in a rat model.
OBJECTIVE
To optimize transgene expression levels after Adeno-associated virus (AAV)-mediated gene transfer, different delivery methods were compared in a transplant setting.


METHODS
Heterotopic abdominal heart transplants were performed in male Lewis rats (250-280 g). According to the vector application method, animals were divided into 3 groups: group 0.35 mL, containing saline solution AAV2/9-LacZ (2 x 10(11) vector genome) was injected directly into the myocardium (apex) immediately after reperfusion. Group 0.3 mL contained a cardioplegic solution AAV2/9-LacZ vectors (3 x 10(12) vector genome), which was rapidly injected into the aortic root, with the pulmonary trunk clamped. Before transplant the transfected heart was incubated for 25 minutes in iced cardioplegia. A reperfusion system was applied in group 5 mL. For 25 minutes, a cold solution of cardioplegia and AAV2/9-LacZ vectors (5 x 10(12) vector genome) was recirculated through the donor heart. Transplanted grafts were explanted after 3 weeks. To detect and to measure marker gene expression, X-gal staining was performed.


RESULTS
In groups 0.35 mL and 0.3 mL, higher transfection efficiency was observed compared to group 5 mL (P < .05). While positive-stained myocardia were detected around the injection site in group 0.35 mL, the expression pattern was much more homogenous in group 0.3 mL.


CONCLUSIONS
Results demonstrate that intracoronary injection of the vectors with the pulmonary trunk clamped leads to the highest and most homogenous distribution of transgene expression in the graft.
Xiaofeng
Jiang
J
Department of Cardiac Surgery, Ludwig-Maximilians-University of Munich, Munich, Germany. Xiaofeng.Jiang@med.uni-muenchen.de
Burdorf
Lars
L
Hinkel
Rabea
R
Gheith
Christian
C
Reichart
Bruno
B
Brandl
Ulrike
U
Schmoeckel
Michael
M
D000818:Animals; D002314:Cardioplegic Solutions / Q000008:administration & dosage; D000229:Dependovirus / Q000235:genetics*; D017930:Genes, Reporter; D005822:Genetic Vectors; D006324:Heart Arrest, Induced; D016027:Heart Transplantation; D007267:Injections; D008297:Male; D023421:Models, Animal; D009206:Myocardium / Q000378:metabolism*; D051381:Rats; D011917:Rats, Inbred Lew; D013997:Time Factors; D014161:Transduction, Genetic; D016042:Transplantation, Heterotopic; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
D002314:Cardioplegic Solutions; D001616:beta-Galactosidase