title: Gene delivery via the hybrid vector of recombinant adeno-associated virus and polyethylenimine.
abstract: The aim of this study was to investigate the cellular delivery mechanism of the hybrid vector comprising the recombinant adeno-associated virus (rAAV) and polyethylenimine (PEI). The rAAV vector, rAAV-rIns1-hInsM2-ΔEGFP, was fluorescently labeled with Cy3, a cyanine dye, and complexed with PEI. The interaction of the hybrid vector with the Huh7 hepatoma cells was monitored by confocal microscopy. Complexation of rAAV with PEI enhanced the transduction efficiency, which was decreased by pretreatment of the cells with sodium chlorate, an inhibitor of glycosaminoglycan sulfation, suggesting the roles of heparan sulfate proteoglycans (HSPG) in the uptake of the hybrid vector by the cells. Examination by flow cytometry and confocal microscopy demonstrated an enhanced interaction between the cells and the virus when complexed with PEI. Pretreatment with wortmannin or cytochalasin B significantly reduced the virus uptake by the cells, suggesting the involvement of phosphatidylinositol 3-kinase (PI3K) signaling and phagocytosis in the interaction between the cells and the hybrid vectors. Treatment of cells with the antioxidants, including l-ascorbic acid, δ-tocotrienol, or N-acetylcysteine (NAC), impaired the rAAV-PEI-mediated transduction. Results obtained in this study illustrated the involvement of PI3K/Akt signaling and the ROS production in gene delivery via the rAAV-PEI hybrid vector.
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, 1, Jen-Ai Road, Section 1, Taipei 10051, Taiwan.
authors.forename: Paul Y-J
authors.initials: PY
authors.lastname: Hsu
authors.affiliation: School of Pharmacy, College of Medicine, National Taiwan University, 1, Jen-Ai Road, Section 1, Taipei 10051, Taiwan. Electronic address: ywyang@ntu.edu.tw.
authors.forename: Ya-Wun
authors.initials: YW
authors.lastname: Yang
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology; D045744:Cell Line, Tumor; D003571:Cytochalasin B / Q000494:pharmacology; D000229:Dependovirus / Q000737:chemistry* / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011094:Polyethyleneimine / Q000737:chemistry*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D014810:Vitamin E / Q000031:analogs & derivatives / Q000494:pharmacology; D000077191:Wortmannin
chemical_list: D000111:Acetylcysteine; D000730:Androstadienes; D000975:Antioxidants; D001205:Ascorbic Acid; D003571:Cytochalasin B; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011094:Polyethyleneimine; D051057:Proto-Oncogene Proteins c-akt; D017382:Reactive Oxygen Species; D014810:Vitamin E; D000077191:Wortmannin; C082097:tocotrienol, delta
keywords: 2′,7′-dichlorodihydrofluorescein diacetate; DHE; Gene delivery; H(2)DCF-DA; HSPG; Hybrid vector; N-acetyl-L-cysteine; NAC; PEI; PI3K; ROS; dihydroethidium; heparan sulfate proteoglycans; phosphatidylinositol 3-kinase; polyethylenimine; rAAV; reactive oxygen species; recombinant adeno-associated virus