title: Adenovirus-mediated augmentation of cell transfection with unmodified plasmid vectors.
abstract: The present study demonstrates that the human adenovirus (Ad) can augment transfer and expression of a gene within plasmid DNA unmodified by nonspecific linkers or by linker-ligand complexes. Following the transfection of COS-7 cells with pRSVL, a luciferase expression plasmid vector directed by the Rous sarcoma virus-long terminal repeat promoter, luciferase activity in the target cells was 10(3)- to 10(4)-fold higher when the cells were also infected with Ad-CFTR, a replication-deficient recombinant Ad containing human cystic fibrosis transmembrane conductance regulator (CFTR) cDNA. The enhancement of luciferase gene expression in COS-7 cells was also observed with Ad-dl312 (a replication-deficient E1a deletion mutant Ad with no exogenous gene) and wild type Ad5. The efficiency of cell transfection with pRSVL in the presence of an Ad was achieved in a dose-dependent fashion with progressively higher luciferase activity in cells infected by increasing amounts of Ad-CFTR, Ad-dl312, or Ad5. The augmentation by Ad-CFTR of the transfer and expression of the luciferase gene in cells was similar to that of another transfection reagent, cationic liposomes. Further, when Ad-CFTR and liposomes were used in combination, 4- to 100-fold more efficient expression of the luciferase gene was achieved than with Ad-CFTR or liposomes alone. When COS-7, HeLa, and CV-1 cells were evaluated in parallel in the presence or absence of liposomes, Ad-mediated enhancement of luciferase activity was observed in all cell lines. Thus, exposure of target cells to replication-deficient or competent human Ad will markedly augment transfer and expression of the genes within plasmid DNA in mammalian cells in vitro without modifying the plasmid with linkers or linker-ligand complexes, a strategy that should be useful for in vitro and in vivo gene transfer applications.
authors.affiliation: Pulmonary Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: K
authors.initials: K
authors.lastname: Yoshimura
authors.forename: M A
authors.initials: MA
authors.lastname: Rosenfeld
authors.forename: P
authors.initials: P
authors.lastname: Seth
authors.forename: R G
authors.initials: RG
authors.lastname: Crystal
mesh_terms: D000256:Adenoviridae / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004705:Endocytosis; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D066298:In Vitro Techniques; D008156:Luciferases / Q000378:metabolism; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection; D014779:Virus Replication
chemical_list: D012333:RNA, Messenger; D008156:Luciferases