Differential behaviour of lipid based and polycation based gene transfer systems in transfecting primary human fibroblasts: a potential role of polylysine in nuclear transport.
DNA delivery systems for gene therapy applications have to be able to trigger the uptake of plasmid DNA into the nucleus. We have tested two types of non-viral vector systems, lipofection (cationic lipid-based, using Lipofectamine) and polyfection (cationic polymer-based, using glycerol enhanced transferrinfection), for their ability to transfect confluent, contact inhibited primary human fibroblasts. While both systems worked well with growing fibroblasts, polyfection was superior with confluent cells. A slight reduction in cell associated plasmid DNA was observed with resting cells, but it was similar for both types of complexes. Lipofectamine showed a prevalence for transfecting cycling cells as judged by costaining transfected cells with cell cycle markers. No such bias was observed when glycerol enhanced transferrinfection was used. Microinjection of plasmid DNA/polylysine complexes into the cytoplasm of fibroblasts resulted in a higher percentage of expressing cells than injection of plasmid DNA, offering an explanation for the higher transfection levels obtained with transferrinfection in non-growing cells.
Zauner
W
W
Institute of Biochemistry, University of Vienna, Dr. Bohrgasse 9/3, A-1030, Vienna, Austria. wolfgang.zauner@cobrat.com
Brunner
S
S
Buschle
M
M
Ogris
M
M
Wagner
E
E
D001692:Biological Transport; D002411:Cation Exchange Resins; D002467:Cell Nucleus / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism*; D015316:Genetic Therapy; D006801:Humans; D008055:Lipids; D008156:Luciferases; D008845:Microinjections; D010957:Plasmids; D011107:Polylysine / Q000031:analogs & derivatives* / Q000378:metabolism; D014162:Transfection / Q000379:methods*; D014168:Transferrin / Q000031:analogs & derivatives*
D002411:Cation Exchange Resins; D008055:Lipids; C086724:Lipofectamine; D014168:Transferrin; C072481:transferrin-polylysine conjugate; D011107:Polylysine; D008156:Luciferases