The immunogenic properties of DNA were first discovered in 1984 when DNA extracts from Mycobacter tuberculosis were reported to activate NK cells. Subsequent studies showed that synthetic oligodeoxynucleotides (ODNs) containing CpG motifs (CpG-ODN) display similar properties to bacterial DNA, showing a broad activation of the immune system: B-cells, NK cells, macrophages, dendritic cells and Th1-biased T-cells. The biological activity of these CpG-ODNs is further increased if they are rendered nuclease resistant by a phosphorothioate backbone modification.
Toll-like receptor 9 (TLR9) was recently identified as a major component in CpG-ODN recognition, thus defining CpG-ODNs as TLR9 agonists. Specific TLR9 expression has been demonstrated in CpG-ODN sensitive cells, such as B-lymphocytes and plasmacytoid dendritic cells.
On B-cells, TLR9 agonists are directly mitogenic, induce secretion of a number of cytokines such as Interleukin (IL)-6 or IL-10, prevent apoptosis triggered by surface antigen-receptor cross linking or other apoptotic agents and promote Ig secretion.
Dendritic cells are directly activated by TLR9 agonists to express co-stimulatory molecules (CD40, CD80, CD86) and secrete a wide variety of cytokines such as TNFa, interferons, IL-6 or IL-12, allowing activation of T-lymphocytes. Most interestingly, TLR9 agonists are able to "license" dendritic cells to directly prime CD8 T cells by a T-helper cell-independent mechanism.
Direct activation of T-cells by TLR9 agonists is unlikely. However, cytokine secretion by dendritic cells activates T-cells. Secretion of IL-12 and IFN? drives the T-cell differentiation towards the Th1 phenotype and can even redirect established Th2-biased responses to Th1 immune responses.
NK cells are strongly activated by TLR9 agonists but they are not primary targets of TLR9 agonists. Actually, NK cell activation depends upon cytokine secretion by dendritic cells, such as IL-12, TNF? and IFN.
Immune stimulation of dendritic and B-cells by TLR9 agonists requires entry of the molecule into the cell. There is no evidence for a specific CpG receptor on the cell surface. Bacterial DNA or CpG-ODNs are probably taken up within cells by endocytosis and then acidified and degraded within endosomes. Internalized TLR9 agonists rapidly activate multiple signalling transduction pathways, including the activation of reactive oxygen species and the mitogen-activated protein kinase (MAP) pathways.
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