Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts: method and efficacy of adenovirus-mediated transduction.
OBJECTIVE
Allograft-targeted immunosuppressive gene therapy may inhibit recipient immune activation and provide an alternative to systemic immunosuppression. We studied the optimal technique and efficacy of intracoronary gene transfer of viral interleukin-10 and human transforming growth factor-beta 1 in a rabbit model of heterotopic heart transplantation.


METHODS
Replication-defective adenoviral vectors were constructed, expressing viral interleukin-10 (AdSvIL10) or transforming growth factor-beta 1 (AdCMVTGF-beta 1). Intracoronary delivery of vectors was accomplished ex vivo by either bolus injection or slow infusion. The allografts were implanted heterotopically in recipient rabbits and collected 4 days after the operation. Vector dose was 4 x 10(9) to 6 x 10(10) pfu/gm of donor heart. Transfer was confirmed by DNA amplification for both genes. Gene product expression in tissue was quantified by immunoassay and visualized by immunohistochemical staining.


RESULTS
Allograft viral uptake was only 9.9% +/- 2.4% with bolus injection, but increased to 80.5% +/- 6.8% at 1 ml/min infusion rate (p = 5 x 10(-14)). Uptake ratio was not affected by vector quantity or slower infusion rates. Transforming growth factor-beta 1 was consistently detected in allografts infected with AdCMVTGF-beta 1, but not with control adenovirus or AdSvIL10. Expression was proportional to infused vector quantity and reached 10 ng/gm of allograft at infused 10(10) pfu/gm. Transforming growth factor-beta 1 was also detected in recipient's serum at less than 1 ng/ml. Viral interleukin-10 was detected in minor amounts only (< 1 ng/gm) in allografts infected with AdvIL10 up to 5 x 10(10) pfu/gm. Nevertheless, it was detected in recipient serum at concentrations up to 0.4 ng/ml.


CONCLUSIONS
Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts during cold preservation is feasible. Slow infusion is superior to bolus injection. In vivo effects on allograft rejection remain to be determined.
Brauner
R
R
Division of Cardiothoracic Surgery, University of California, Los Angeles School of Medicine, USA.
Wu
L
L
Laks
H
H
Nonoyama
M
M
Scholl
F
F
Shvarts
O
O
Berk
A
A
Drinkwater
D C
DC
Wang
J L
JL
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D015925:Cryopreservation; D004195:Disease Models, Animal; D005240:Feasibility Studies; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D016027:Heart Transplantation / Q000276:immunology* / Q000502:physiology*; D007150:Immunohistochemistry; D007261:Infusions, Intra-Arterial; D016753:Interleukin-10 / Q000008:administration & dosage / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011817:Rabbits; D014161:Transduction, Genetic; D016212:Transforming Growth Factor beta / Q000008:administration & dosage / Q000235:genetics*; D016042:Transplantation, Heterotopic
D016212:Transforming Growth Factor beta; D016753:Interleukin-10