FIG 4.

TLR4- and MyD88-dependent TNF-α induction. (A) Peritoneal macrophages derived from TLR2 KO mice were treated with 10 μM OD2088 or 50 μM VIPER for 30 min and then infected with M. pneumoniae. After 6 h of incubation, TNF-α concentrations in the culture medium were measured using ELISA. All values are presented as the means and SD of the results of 3 assays. *, P < 0.01 compared with PBS treatment by multiple comparison. (B) Peritoneal macrophages derived from TLR2 KO, TLR4 KO, and TLR2/4 double-KO mice were infected with M. pneumoniae. After 6 h of incubation, TNF-α concentrations in the culture medium were measured using ELISA. All values are presented as the means and SD of the results of 3 assays. *, P < 0.01 compared with WT macrophages by multiple comparison. (C) Peritoneal macrophages derived from TLR2 KO, TLR4 KO, and MyD88 KO mice were infected with M. pneumoniae (OD₅₉₅ = 0.1). After 6 h of incubation, TNF-α concentrations in the culture medium were measured using ELISA. All values are presented as the means and SD of the results of 3 assays. *, P < 0.01 compared with WT macrophages by multiple comparison. (D) WT, TLR2 KO, and TLR2/4 double-KO mice were intranasally infected with M. pneumoniae. After 24 h, the mice were infected with same amounts of M. pneumoniae again for an additional 24 h. TNF-α concentrations in the BALF were measured using ELISA. All values are presented as the means and SD of the results of 3 assays. **, P < 0.05 compared with WT mice by multiple comparison. (E) Peritoneal macrophages derived from TLR2 KO mice were treated with 10 μM SB203580, 10 μM U0126, or 50 μM SP600123 for 30 min. The treated cells were infected with M. pneumoniae. After 6 h of incubation, TNF-α concentration in the culture medium were measured using ELISA. All values are presented as the means and SD of the results of 3 assays. *, P < 0.01 compared with DMSO treatment by multiple comparison. **, P < 0.05 compared with DMSO treatment by multiple comparison.