Long-term expression of foreign genes in normal human epidermal keratinocytes after transfection with lipid/DNA complexes

Abstract

Normal human epidermal keratinocytes were isolated and cultivated in serum-free medium. The expression of the integrin subunits α₆ and β₁ indicated that a high number of keratinocytes from the stem cell system was present. These cells were transfected with complexes made of different cationic lipids and marker genes. Effectene showed a 20-fold higher transfection efficiency, compared to Lipofectin and Lipofectamine, and a similar low toxicity. The transfection protocol was optimised. A DNA/lipid ratio of 0.133 showed the highest transfection efficiency. Keratinocytes expressed the marker gene luciferase for 20 days. The maximum expression occurred after 3–4 days, where individual patches of fluorescent keratinocytes were detected. Transfected keratinocytes, cultivated at the air–liquid interface, expressed the marker gene β-galactosidase for at least 7 weeks.