title: Adenovirus-mediated gene transfer in the transplant setting. II. Successful expression of transferred cDNA in syngeneic liver grafts.
abstract: These experiments establish a model for gene transfer to transplanted liver grafts ex vivo using adenoviral vectors. Rat liver grafts (n = 8) were harvested, and preserved in UW or lactated Ringer's. The grafts were infected ex-vivo via portal vein perfusion with replication-defective Ad vectors encoding the beta-galactosidase (beta-gal) gene diluted in UW solution. The infected grafts were stored at 4 degrees C for 1 hr, then transplanted into syngeneic hosts. Liver biopsies were taken at 1, 7, and 15 days after transplantation. Infection rate was assessed by histochemical staining for beta-gal. Liver DNA and RNA were assayed for the beta-gal sequences, and recombinant protein production measured at 24 hr and 7 days after transplantation. Under conditions mimicking liver graft cold storage, efficient gene transfer was achieved with an infection rate of 10-15%, as assessed by X-gal staining. Viral DNA and RNA presence in the graft was confirmed; gene expression with protein production were verified by western blots and a functional protein assay. All studies were negative in control livers. Gene expression persisted for at least 2 weeks after transplantation. We conclude that efficient adenovirus-mediated gene insertion and expression of gene products can be accomplished in whole-liver grafts under hypothermic preservation conditions currently used in clinical transplantation.
authors.affiliation: Department of Surgery, University of California at Los Angeles, CA 90024-6904.
authors.forename: A
authors.initials: A
authors.lastname: Shaked
authors.forename: M E
authors.initials: ME
authors.lastname: Csete
authors.forename: K E
authors.initials: KE
authors.lastname: Drazan
authors.forename: D
authors.initials: D
authors.lastname: Bullington
authors.forename: L
authors.initials: L
authors.lastname: Wu
authors.forename: R W
authors.initials: RW
authors.lastname: Busuttil
authors.forename: A J
authors.initials: AJ
authors.lastname: Berk
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D016031:Liver Transplantation / Q000379:methods*; D008297:Male; D008969:Molecular Sequence Data; D051381:Rats; D011917:Rats, Inbred Lew
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary