Cell culturing and cytokine analysis. Bone marrowderived immature dendritic cells (DCs) were prepared by culturing bone marrow cells isolated from the femurs and tibia of C57BL/6j and TLR4 2/2 mice in complete RPMI 1640 (cRPMI; 5% [v/v] heat inactivated Fetal Calf Serum [FCS] [30 mins at 60uC], 100 U/ml penicillin, 100 mg/ml streptomycin, 2 mM L-glutamine and 50 mM 2-mercaptoethanol) with recombinant mouse GM-CSF (20 ng/ml; R&D Systems), at 37uC. On days 3 and 6 of culture, fresh medium with GM-CSF (20 ng/ml) was added to the cells. On day 8, cells were harvested, counted and stained with CD11c (Caltag Laboratories) for analysis by flow cytometry to determine purity (.90%). J774 cells and RAW264.7, murine macrophage cell lines were cultured in cRPMI 1640 medium containing 10% (v/v) FCS. All cells were used to conduct experiments when they reached ,90% confluence.