Asunto(s)
Trasplante de Células , Endotelio Vascular/citología , Técnicas de Transferencia de Gen , Hígado , beta-Galactosidasa/genética , Actinas/genética , Adenoviridae , Animales , Aorta , Células Cultivadas , Pollos , Citomegalovirus/genética , Elementos de Facilitación Genéticos , Escherichia coli/genética , Femenino , Vectores Genéticos , Hígado/citología , Porcinos , beta-Galactosidasa/biosíntesisRESUMEN
In vivo gene transfer to the porcine liver was tested with adenoviral vector to achieve molecular biological graft modulation. In adult female pigs immunosuppressed with cyclophosphamide, cyclosporine, and prednisolone, the liver was surgically isolated and flushed out with cold lactate Ringer solution (4 degrees C), by means of a pump-controlled bypass of the portal vein and the inferior vena cava in Groups A and D. In Group A (n = 4), 2 x 1011 pfu of the adenoviral vectors (pAdexCALacZ) were injected through the left hepatic artery during cold ischemia. In Group (n = 4), 2 x 1011 pfu of adenovirus vectors were injected through the auricular vein in a one-shot manner without a laparotomy. In Group C (n = 4), 2 x 1011 pfu/ml of adenoviral vectors were injected through the hepatic artery in a one-shot manner, without a surgical isolation of the liver. Group D (n = 4) animals received the same protocol as Group A except for the fact that they did not receive the immunosuppressive regimen. In a polymerase chain reaction, a transfected LacZ sequence was detected until POD 28 in Group A, but not in the other groups. In 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) staining, only the Group A animals revealed apparent staining predominantly in the portal area at POD 2, which then continued to be recognized until POD 28. The in situ perfusion of the liver combined with immunosuppression is thought to provide an ideal environment for the liver-directed adenovirus-mediated gene transfer to the porcine liver, by enabling a long contact with a high titer of the adenoviral vector.