Recipient intramuscular administration of naked plasmid TGF-beta1 attenuates lung graft reperfusion injury.
BACKGROUND
Gene therapy may be an effective strategy for modulating lung graft ischemia-reperfusion injury. We investigated whether recipient intramuscular (IM) naked plasmid gene transfer of transforming growth factor beta1-active (TGF-beta1-active) ameliorates lung graft ischemia-reperfusion injury.


METHODS
Preliminary studies in F344 rats demonstrated that gastrocnemius muscle transfection of TGF-beta1-active produced muscle and plasma protein expression at 24 and 48 hours after transfection. Recipients (n = 8) received IM injection of naked plasmid-encoding chloramphenicol acetyl transferase (CAT), TGF-beta1-latent or TGF-beta1-active, respectively, at 24 or at 48 hours before left lung transplantation. We did not treat the control group before transplantation (18-hour cold ischemia). Donor lungs were flushed with low-potassium dextran-1% glucose and stored for 18 hours at 4 degrees C. All groups were killed at 24 hours after transplantation. Immediately before killing the animals, we clamped the contralateral right hilum and assessed graft function. We measured wet-to-dry ratio (W/D), myeloperoxidase, pro-inflammatory cytokines (interleukin 1 [IL-1], tumor necrosis factor alpha [TNF-alpha], interferon-gamma [INF-gamma], and IL-2) and performed immunohistochemistry.


RESULTS
Arterial oxygenation was greatest in the recipient group transfected with TGF-beta1-active at 24 hours before transplantation compared with CAT, TGF-beta1-latent, and 18-hour cold ischemia groups (p < 0.01). The W/D ratio and myeloperoxidase decreased in both 24- and 48-hour groups, with TGF-beta1-active compared with CAT, and 18-hour cold ischemia groups (W/D, p < 0.02 and p < 0.004, respectively; myeloperoxidase, p < 0.05 and p < 0.01, respectively). All pro-inflammatory cytokines decreased in the 24-hour TGF-beta1-active group compared with CAT, TGF-beta1-latent, 18-hour and 1-hour cold ischemia, and non-treated lung groups (IL-1beta, p < 0.03; TNF-alpha, p < 0.02; IFN-gamma, p < 0.001; IL-2, p < 0.0001). In 24- and 48-hour groups with TGF-beta1-active, immunohistochemistry showed marked staining of Type I and Type II alveolar cells and of macrophages from the apical to the caudal sections of the lung grafts.


CONCLUSIONS
Recipient IM administration of naked plasmid encoding TGF-beta1-active before transplantation ameliorates lung isograft reperfusion injury after prolonged ischemia.
Daddi
Niccolò
N
Division of Cardiothoracic Surgery, Washington University School of Medicine, St. Louis, MO 63110, USA.
Kanaan
Samer A
SA
Suda
Takashi
T
Tagawa
Tsutomu
T
D'Ovidio
Franco
F
Grapperhaus
Kathleen
K
Kozower
Benjamin D
BD
Ritter
Jon H
JH
Mohanakumar
T
T
Patterson
G Alexander
GA
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D005240:Feasibility Studies; D018014:Gene Transfer Techniques; D007273:Injections, Intramuscular; D016040:Lung Transplantation / Q000009:adverse effects*; D008297:Male; D023421:Models, Animal; D010957:Plasmids / Q000008:administration & dosage* / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000209:etiology / Q000378:metabolism* / Q000517:prevention & control*; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D053773:Transforming Growth Factor beta1; D019076:Transgenes / Q000235:genetics
C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D015500:Chloramphenicol O-Acetyltransferase