Gene delivery strategies for targeting microglia cells (101.17)

Microglia (MG) are the resident immune cells of the CNS and are implicated as both important sensors for biological defense as well as mediators of various pathological conditions through the various inflammatory and oxidative molecules they produce. In the current study we compared different techniques for gene targeting of microglia using non-viral and viral methods in vitro for eventual application to the in vivo setting. For this investigation we employed the EOC2 MG cell line and three methods of gene introduction for pmax plasmid-GFP: electroporation,cationic lipid co-delivery andlentiviral vector infection. Electroporation when employed with various buffers and at different voltages led to low transfection efficacies (>1%). Transfection with cationic lipids (LipofectAmine, NovaFector, GeneFector) showed higher transfection rates (%), but these were still relatively low. In contrast, lentiviral vector delivery produced robust infection (>95%) and GFP expression. In addition, the activation of the cells, so that they underwent morphological (rounded) and immunophenotypical (ED1 immunoreactivity) changes, prior to or during transfection with the lower efficiency methods, did dramatically enhance efficiency (>95%). We conclude, that in CNS pathologies, where MG cells are activated, both non-viral and viral methods of gene therapy will likely be highly successful for the introduction of specific anti-inflammatory or immune suppressive genes. FUNDING: The Miami Project to Cure Paralysis, The Buoniconti Fund, The US Army Medical Research and Material Command