title: Receptor-mediated transfer of pSV2CAT DNA to a human hepatoblastoma cell line HepG2 using asialofetuin-labeled cationic liposomes.
abstract: Asialofetuin-labeled liposomes (AF-lps) were developed as a vector for gene transfer to hepatocytes. Plasmid pSV2CAT DNA which encodes bacterial chloramphenicol acetyltransferase (CAT) was associated with (meaning, in this report, the sum of 'to be adsorbed on the surface of' and 'to be encapsulated into the internal phase of') AF-lps (AF-lps-pSV2CAT) prepared by a tandem combination of the detergent removal and freeze-thaw methods. Ninety-six percent of input pSV2CAT was associated with AF-lps containing N-(alpha-trimethylammonioacetyl)-didodecyl-D-glutamate chloride, and approx. two-thirds of the associated DNA was encapsulated into the internal phase. The uptake of AF-lps by the cultured human hepatoblastoma cell line HepG2, having asialoglycoprotein receptors (AGPR) on their plasma membrane, was decreased by the addition of free AF and cytochalasin B. AF-lps bound to HepG2 cells through specific interaction with AGPR, and were internalized into the cells by the receptor-mediated endocytotic pathway. HepG2 cells transfected by AF-lps-pSV2CAT showed a significantly higher CAT activity than those transfected by pSV2CAT associated with non-labeled control lps (N-lps-pSV2CAT) or a mixture of pSV2CAT and empty AF-lps. Pretreatment with EDTA-encapsulated AF-lps increased the transfection efficiency of AF-lps-pSV2CAT. The CAT activity in A431 and Swiss/3T3 cells transfected with AF-lps-pSV2CAT was low and almost the same as those transfected with N-lps-pSV2CAT. Since DNA encapsulated in lps is likely to be protected against digestion by nucleases in the blood circulation, AF-lps could be used as a gene transfer vector targeting the hepatocytes in vivo.
authors.affiliation: Tokyo College of Pharmacy, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Hara
authors.forename: Y
authors.initials: Y
authors.lastname: Aramaki
authors.forename: S
authors.initials: S
authors.lastname: Takada
authors.forename: K
authors.initials: K
authors.lastname: Koike
authors.forename: S
authors.initials: S
authors.lastname: Tsuchiya
mesh_terms: D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism*; D001692:Biological Transport; D004339:Drug Compounding; D060748:Fetuins; D005822:Genetic Vectors; D018197:Hepatoblastoma; D006801:Humans; D008081:Liposomes; D008113:Liver Neoplasms; D010957:Plasmids; D011956:Receptors, Cell Surface / Q000378:metabolism*; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured; D000509:alpha-Fetoproteins / Q000378:metabolism*
chemical_list: D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D060748:Fetuins; D008081:Liposomes; D011956:Receptors, Cell Surface; D000509:alpha-Fetoproteins; C017029:asialofetuin