Recognition of microbial products through TLRs triggers the expression of several cytokines that regulate innate and adaptive immunity. Signaling by various TLRs is not equivalent and leads to differential gene induction. This study analyzed the responses of human dendritic cells (DCs) and PBMCs stimulated with agonists of TLR2, TLR3, TLR4, TLR5, and TLR7, first individually and then in combination. Several cytokines were equally induced by all TLR agonists, but four genes, IFN-β, IFN-γ-inducible protein 10 (IP-10), IL-12p35, and IL-15, showed a very restricted pattern of induction. Thus, each TLR appears to possess a distinctive ability to activate DCs or PBMCs, suggesting that TLR-mediated responses cannot be simply cataloged as resembling either TLR2 (MyD88 dependent) or TLR4 (MyD88 independent) and that other signaling modalities may exist. The analysis of DC and PBMC activation by combinations of TLR agonists revealed that TLR2 agonists are able to block the induction of IP-10, IL-12p35, and IFN-γ, but not IL-15 and IFN-β, by TLR3 and TLR4. TLR2 stimulation led to rapid release of IL-10 that is responsible for inhibition of IP-10 and IL-12p35 induction. Cross-talk between different TLRs may modify the primary responses of TLR to their agonist, adding a further level of complexity to the regulation of innate immunity.