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DNA/protein complexes delivered in conjunction with adenovirus generate high‐efficiency in vitro transfection: A powerful transfection reagent with potential broad applications
Author(s) -
Askari Fred
Publication year - 1993
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1002/hep.1840180639
Subject(s) - transfection , gene delivery , microbiology and biotechnology , reporter gene , biology , dna , genetic enhancement , hek 293 cells , gene , gene expression , biochemistry
We have combined a receptor‐mediated DNA delivery system with the endosomal lysis ability of adenovirus and shown that DNA can be delivered into primary hepatocytes, resulting in a high level of gene expression. When asialoorosomucoid conjugated with poly(L‐lysine) was used to deliver the Escherichia coli β‐galactosidase gene into primary hepatocytes through binding with the hepatic asialoglycoprotein receptor, only a low level of β‐galactosidase was detectable, with less than 0.1% of the hepatocytes being transfected. This level of activity can be greatly enhanced by the cointernalization of the DNA protein complex with a replication‐defective adenovirus, resulting in 100% of the hepatocytes staining blue with 5‐bromo‐4‐chloro‐3‐indolyI β‐D‐galactoside. Quantitative analysis of β‐galactosdase expression also showed a 1000‐fold enhancement of activity. To test the applicability of this DNA delivery system for the correction of phenylketonuria, a metabolic disorder that causes severe mental retardation in children, we have delivered the human phenylalanine hydroxylase (PAH) gene to hepatocytes derived from a PAH‐deficient mouse strain and dem‐onstrated complete reconstitution of enzymatic activity. This method shows great promise for efficient gene delivery to the liver for correction of hepatic disorders.