Upon stimulation with a wide range of concentrations of CpG oligodeoxynucleotide 2216 (CpG 2216), plasmacytoid DC are induced to produce type I IFN (IFN-alpha/beta). In contrast, CpG 1668 shows a bell- shaped dose-response correlation, i.e. only intermediate but not high doses of CpG 1668 induce IFN-alpha/beta. Interestingly, high-dose CpG 1668 completely inhibited IFN-alpha responses induced by CpG 2216. Experiments using supernatant of high-dose CpG-1668-treated cells indicated that secreted inhibitor(s) mediated the IFN-alpha shut-off. Among modulating cytokines, IL-10 turned out to be one important negative regulator. In line with this, supernatants of IL-10-deficient DC cultures stimulated with high-dose CpG 1668 did not inhibit IFN-alpha production. Interestingly, high-dose CpG 1668 also inhibited IFN-alpha responses induced by the DNA-encoded mouse cytomegalovirus, whereas IFN-alpha responses induced by negative-strand RNA-encoded vesicular stomatitis virus were only marginally affected. Experiments with DC cultures devoid of TLR9 indicated that TLR9 was critically required to mediate stimulatory and modulatory signals by low and high concentrations of CpG 1668, respectively. Analysis of purified DC subsets showed that conventional DC were the main IL-10 producers, whereas plasmacytoid DC hardly produced any IL-10.