Cellular uptake and activation characteristics of naked plasmid DNA and its cationic liposome complex in human macrophages.

Plasmid DNA (pDNA) is an important macromolecular therapeutic agent suitable for DNA-based therapies, such as non-viral gene therapy and DNA vaccination. Unmethylated CpG motifs abundant in bacterial DNA, but not in vertebrate DNA, are known to trigger an inflammatory response, which inhibits transgene expression, while modulating immunological consequences following vaccination. We studied cellular uptake and activation characteristics of naked pDNA and its cationic liposome complex in human macrophage-like cells.

Macrophage activation by a DNA/cationic liposome complex requires endosomal acidification and TLR9-dependent and -independent pathways.

Previously, we showed that bacterial DNA and vertebrate DNA/cationic liposome complexes stimulate potent inflammatory responses in cultured mouse macrophages. In the present study, we examined whether endocytosis and subsequent acidification are associated with these responses. The endocytosis inhibitor, cytochalasin B, reduced tumor necrosis factor alpha (TNF-alpha) production by a plasmid DNA (pDNA)/cationic liposome complex.

Restricted cytokine production from mouse peritoneal macrophages in culture in spite of extensive uptake of plasmid DNA.

The production of inflammatory cytokines from macrophages (Mphi), upon stimulation with plasmid DNA (pDNA) containing CpG motifs, is a critical process for DNA-based therapies such as DNA vaccination and gene therapy. We compared Mphi activation, following stimulation with naked pDNA, based on the production of cytokines from cell lines (RAW264.7 and J774A1) and peritoneal Mphis in primary culture.