Optimisation of Polyethylenimine-Based Gene Delivery to Mouse Brain

Abstract

Polyethylenimine (PEI) is proving to be an efficient and versatile vector for gene delivery in vivo. However, a limiting factor is the relatively short duration of gene expression in some sites. Given the particularly high levels of expression seen in the short term we postulated that loss of expression could result from overloading the nucleus with foreign DNA. To address this problem we first followed DNA delivery and localisation with digoxin-labelled plasmid DNA complexed with 22 kD linear PEI and used these complexes for intraventricular injection into brains of anaesthetised newborn mice. At 24 h post-injection, labelled DNA was found exclusively in the nuclear and perinuclear regions. We next carried out a dose response curve using decreasing amounts of DNA, either in a constant volume (2 μl) or at a constant concentration (500 ng/μl). In both conditions, transgene expression yield was maximum at 100 ng DNA per injection. Using this optimal amount of DNA increased yield of transgene expression significantly at 24 h and one week post-injection as compared to 1 μg DNA. A final point addressed was whether co-expressing an anti-apoptotic gene could enhance gene expression in the longer term. Co-expressing bcl-X_L with luciferase or LacZ significantly increased expression of both these genes at one week post-injection.

Key Words:

• Apoptosis
• Cationic polymers
• CNS
• In vivo gene transfer
• Synthetic vectors