title: Cationic liposome-mediated expression of HIV-regulated luciferase and diphtheria toxin a genes in HeLa cells infected with or expressing HIV.
abstract: HIV-regulated expression of the diphtheria toxin A fragment gene (HIV-DT-A) is a potential gene therapy approach to AIDS. Since cationic liposomes are safe and non-immunogenic for in vivo gene delivery, we examined whether LipofectAMINE or DMRIE reagent could mediate the transfection of HIV-DT-A (pTHA43) or the HIV-regulated luciferase gene (pLUCA43) into HIV-infected or uninfected HeLa cells. pLUCA43 was expressed at a 10(3)-fold higher level in HeLa/LAV cells than in uninfected HeLa cells, while the extent of expression of RSV-regulated luciferase was the same in both cell lines. Co-transfection of HeLa cells with pTHA43 and the proviral HIV clone, HXB deltaBgl, resulted in complete inhibition of virus production. In contrast, the delivery of HIV-DT-A to chronically infected HeLa/LAV or HeLa/IIIB cells, or to HeLa CD4+ cells before infection, did not have a specific effect on virus production, since treatment of cells with control plasmids also reduced virus production. This reduction could be ascribed to cytotoxicity of the reagents. The efficiency of transfection, as measured by the percentage of cells expressing beta-gal, was approximately 5%. Thus, cationic liposome-mediated transfection was too inefficient to inhibit virus production when the DT-A was delivered by cationic liposomes to chronically- or de novo- infected cells. However, when both the virus and DT-A genes were delivered into the same cells by cationic liposomes, DT-A was very effective at inhibiting virus production. Our results indicate that the successful use of cationic liposomes for gene therapy will require the improvement of their transfection efficiency.
authors.affiliation: Department of Microbiology, School of Dentistry, University of the Pacific, San Francisco, CA 94115, USA.
authors.forename: K
authors.initials: K
authors.lastname: Konopka
authors.forename: G S
authors.initials: GS
authors.lastname: Harrison
authors.forename: P L
authors.initials: PL
authors.lastname: Felgner
authors.forename: N
authors.initials: N
authors.lastname: Düzgüneş
mesh_terms: D004167:Diphtheria Toxin / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D006678:HIV / Q000235:genetics*; D016655:HIV Core Protein p24 / Q000096:biosynthesis; D015658:HIV Infections / Q000517:prevention & control; D006367:HeLa Cells; D006801:Humans; D008081:Liposomes; D008156:Luciferases / Q000235:genetics*; D010446:Peptide Fragments / Q000235:genetics*; D010957:Plasmids; D015533:Transcriptional Activation; D014162:Transfection; D014779:Virus Replication
chemical_list: D004167:Diphtheria Toxin; D016655:HIV Core Protein p24; D008081:Liposomes; D010446:Peptide Fragments; C029795:diphtheria toxin fragment A; D008156:Luciferases