Innate immune response is believed to be among the earliest provisional cellular responses, and mediates the interactions between microbes and cells. Toll-like receptors (TLRs) are critical to these interactions. We hypothesize that TLRs also play an important role in interactions between nanoparticles (NPs) and cells, although little information has been reported concerning such an interaction. In this study, we investigated the role of TLR3, TLR4 and TLR7 in cellular uptake of titanium dioxide NP (TiO NP) agglomerates and the resulting inflammatory responses to these NPs. Our data indicate that TLR4 is involved in the uptake of TiO NPs and promotes the associated inflammatory responses. The data also suggest that TLR3, which has a subcellular location distinct from that of TLR4, inhibits the denaturation of cellular protein caused by TiO NPs. In contrast, the unique cellular localization of TLR7 has middle-ground functional roles in cellular response after TiO NP exposure. These findings are important for understanding the molecular interaction mechanisms between NPs and cells.