Improved lipid-mediated gene transfer in C6 glioma cells and primary glial cells using FuGene.
Gene therapy is a potent method to counteract neurodegeneration by introducing genetic information encoding neuroprotective factors. In this study cationic lipids were used to transfer DNA into C6 glioma cells and primary glial cells. When comparing the novel compound FuGene with other commercially-available lipids, it was found that FuGene markedly enhanced gene transfer of a beta-galactosidase reporter plasmid into C6 glioma cells. FuGene had several advantages compared to other lipids, such as a very low toxicity and the capability of transfection under serum conditions. When optimizing, a DNA-lipid ratio of 150 ng DNA/1 microl FuGene and a concentration of 3 microl FuGene/1 ml medium was found to be optimal. The incubation time peaked after 8 h and the expression time reached an optimum between 2 and 6 days. When cells were transfected on 3 consecutive days for 6 h each ('boosting'), the transfection efficiency was markedly enhanced in primary glial cells. When using endotoxin-free DNA the transfection efficiency could be enhanced up to 3 times. The optimal transfection efficiency in C6 glioma cells and in primary glial cells was found to be 16.3 +/- 0.3% and 5.1 +/- 0.37% of total cells, respectively. In conclusion this study shows that the novel compound FuGene has a very high potential to transfer DNA into cells of glial origin, and it might be an interesting canditate for ex vivo and in vivo gene therapeutic approaches.
Wiesenhofer
B
B
Department of Psychiatry, University Hospital Innsbruck, Austria.
Kaufmann
W A
WA
Humpel
C
C
D000818:Animals; D000831:Animals, Newborn; D004247:DNA / Q000008:administration & dosage / Q000235:genetics*; D004731:Endotoxins; D018014:Gene Transfer Techniques; D005910:Glioma / Q000235:genetics*; D008081:Liposomes / Q000008:administration & dosage / Q000235:genetics*; D009457:Neuroglia; D051381:Rats; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000235:genetics*
D004731:Endotoxins; D008081:Liposomes; D004247:DNA; D001616:beta-Galactosidase