1074. Interaction of LPD Nanoparticles with Dendritic Cells
Authors: Lisa M. Shollenberger, Leaf Huang
Journal: Molecular Therapy (2006)

Liposome-polycation-DNA (LPD) nanoparticles made with cationic lipids are self-assembling transfection reagents with the ability to act as an adjuvant for the immune system through unmethylated CpG motifs in the plasmid DNA. These properties, as well as the ease of large-scale preparation and lack of a specific immune response to the particle itself, make these particles potentially important nonviral vaccine vectors. This study focuses on the interaction of LPD particles with dendritic cells (DCs), which are mediators of immunity in vivo. Understanding the internalization and intracellular trafficking of these particles in dendritic cells is important for exploitation of this knowledge during the generation of new vaccines. DC2.4 cells, a transformed dendritic cell line, were exposed to LPDs overnight and were shown by flow cytometry to be fully activated as compared to lipopolysaccharide (LPS) activation, presumably through the DNA component. Activated or not activated dendritic cells were incubated in the presence LPD particles made with fluorescent labeled plasmid DNA for 1 hour and cells were analyzed for internalization of the particles. Both flow cytometry and fluorescence microscopy results indicate DCs can internalize LPDs but do not internalize free plasmid DNA and also suggest that dendritic cells internalize more particles after activation. To examine the intracellular trafficking and localization after particle internalization, preliminary confocal microscopy studies confirm the dendritic cells internalize LPD nanoparticles and, when activated, target those particles to the endosomal/lysosomal compartment for degradation. Supported by NIH grant AI48851. Liposome-polycation-DNA (LPD) nanoparticles made with cationic lipids are self-assembling transfection reagents with the ability to act as an adjuvant for the immune system through unmethylated CpG motifs in the plasmid DNA. These properties, as well as the ease of large-scale preparation and lack of a specific immune response to the particle itself, make these particles potentially important nonviral vaccine vectors. This study focuses on the interaction of LPD particles with dendritic cells (DCs), which are mediators of immunity in vivo. Understanding the internalization and intracellular trafficking of these particles in dendritic cells is important for exploitation of this knowledge during the generation of new vaccines. DC2.4 cells, a transformed dendritic cell line, were exposed to LPDs overnight and were shown by flow cytometry to be fully activated as compared to lipopolysaccharide (LPS) activation, presumably through the DNA component. Activated or not activated dendritic cells were incubated in the presence LPD particles made with fluorescent labeled plasmid DNA for 1 hour and cells were analyzed for internalization of the particles. Both flow cytometry and fluorescence microscopy results indicate DCs can internalize LPDs but do not internalize free plasmid DNA and also suggest that dendritic cells internalize more particles after activation. To examine the intracellular trafficking and localization after particle internalization, preliminary confocal microscopy studies confirm the dendritic cells internalize LPD nanoparticles and, when activated, target those particles to the endosomal/lysosomal compartment for degradation. Supported by NIH grant AI48851.