Adeno-associated virus mediated gene transfer into primary rat brain neuronal and glial cultures: enhancement with the pH-sensitive surfactant dodecyl 2-(1 '-imidazolyl) propionate

This study evaluated the effects of a novel, pH-sensitive surfactant, dodec yl 2-(1'-imidazolyl) propionate (DIP), on cationic lipid mediated transfect ion in primary rat brain neuronal and glial cultures. The cationic lipid co mplex DOTAP/DOPE (1,2-dioleoyl-3-trimethylammonium propionate and dioleoyl phosphatidylethanolamine, respectively) was added over a range of concentra tions (0-120 mu g/ml) with DNA concentration kept constant (1.6 mu g/ml). T he neuron-specific enolase (NSE) and cytomegalovirus (CMV) promoters were f ound to drive green fluorescent protein (GFP) expression in neuron-enriched and glial cultures, respectively, using adeno-associated virus (AAV) deriv ed constructs. NSE-driven GFP expression was not observed in glial cultures . Addition of DOTAP/DOPE increased transfection efficiency over a wide rang e of lipid concentrations (5-50 mu g/ml) keeping DNA concentration constant (1.6 mu g/ml). Addition of DIP to the lipid/DNA complex increased maximum transfection efficiencies in glial and neuronal cultures 2-3-fold. Transfec tion efficiencies were at their maximum with a similar total lipid concentr ation (50 mu g/ml) in both cell-types in the presence of DIP. Neuronal cult ures were more sensitive than glia to the toxic actions of DOTAP/DOPE, with or without DIP. These results indicate that AAV-mediated gene-transfer to neurons and glia can be facilitated by addition of a pH-sensitive surfactan t to cationic liposome/DNA complexes and that endosomal escape could be a l imiting factor in transgene expression. (C) 2000 Elsevier Science Ltd. All rights reserved.