Natural and Synthetic TLR7 Ligands Inhibit CpG-A- and CpG-C-Oligodeoxynucleotide-Induced IFN-α Production

Abstract

Plasmacytoid dendritic cells (pDCs) are unique with respect to their capacity to produce unsurpassed amounts of IFN-α and coexpress TLR7 and TLR9, mediating IFN-α production. Although TLRs are critical receptors of innate immunity, little is known about the immunological effects of TLR7/TLR9 costimulation. We have analyzed the effects of TLR7/TLR9 costimulation on IFN-α production by leukocytes and pDCs. Our experiments revealed that both synthetic (resiquimod and loxoribine) and natural (ssRNA40) TLR7 ligands abrogate CpG-A- and CpG-C-oligodeoxynucleotide (ODN)-induced IFN-α production by human leukocytes. Because TLR7 ligands themselves represent important IFN-α inducers, we demonstrated that substimulatory TLR7 ligand concentrations significantly inhibited CpG-A-induced IFN-α. Delayed addition of TLR7 ligands still resulted in complete suppression of CpG-A-ODN-induced IFN-α production, suggesting that the inhibition is unlikely to be caused by a kinetic uptake advantage. Unlike for CpG-A and CpG-C, TLR7 ligands did not inhibit CpG-B-ODN-induced IFN-α production. Experiments with purified human pDCs demonstrated that the inhibitory effects of TLR7/TLR9 costimulation were mediated directly by pDCs. Suppression of IFN-α production was not related to increased cell death and was also detectable in enriched mouse pDCs. Analyses of pDCs suggested that the TLR7 signal regulates the outcome of TLR7 ligand/CpG-A-ODN costimulation and can either inhibit (IFN-α) or promote (IL-8/CD40) cytokine and surface marker expression. Our data reveal for the first time a strong inhibitory effect of TLR7 stimulation on IFN-α production induced by CpG-A- and CpG-C-ODNs. These findings provide novel insight into the effects of TLR7/TLR9 costimulation and may support the development of novel TLR9 inhibitors.