Isolation and culture of murine peptone-elicited peritoneal macrophages (PEM) PEM were isolated from 7 to 10 week-old C57BL/6J and genetically-matched tlr2 2/2 , tlr3 2/2 , tlr4 2/2 , mal/tirap 2/2 , trif 2/2 , irf3 2/2 and irf7 2/2 mice as previously described [44] . The percentage of macrophages was determined by flow cytometry using CD11b (1:100, eBiosciences) and F4/80 (1:100, eBiosciences) antibodies. More than 90% of the cells were macrophages. PEM were seeded onto 6-well plates at a concentration of 2610 6 cells per well in DMEM (PAA) supplemented with 10% FCS, 10% L929 conditioned medium (LCM) and 1% penicillin-streptomycin or RPMI-1640 (Gibco) supplemented with 10% FBS and 1% penicillin-streptomycin.