Figure 2

TLR4-MyD88-NF-κB signaling pathway mediates S. pneumoniae -induced TLR2 up-regulation. (A) Cells were transiently transfected with TLR2 promoter reporter gene with TLR4 DN mutant plasmid or empty vector, and S.p.-induced luciferase activity was measured 5 hours after S.p. treatment. (B) Cells were transiently transfected either with TLR4 DN mutant plasmid or empty vector, and S.p.-induced TLR2 mRNA expression was measured by Q-PCR analysis. (C) WT and TLR4 KO mice were intratracheally inoculated with S.p., and TLR2 mRNA expression was measured from the lungs of inoculated mice by Q-PCR analysis. (D) WT and MyD88 KO mice were intratracheally inoculated with S.p., and TLR2 mRNA expression was measured from the lungs of inoculated mice by Q-PCR analysis. (E) Bacterial number in the lungs of WT and TLR4 KO mice following S.p. inoculation. WT and TLR4 KO mice were intratracheally inoculated with 1 × 10⁷ CFU of S.p. D39, and CFU of the lungs were measured 24 hours after inoculation. (F) Inflammatory cell migration into airway in WT and TLR4 KO mice following S.p. inoculation. WT and TLR4 KO mice were intratracheally inoculated with 1 × 10⁷ CFU of S.p., and total cells were measured from the BAL fluid of the lungs of saline- or S.p.-inoculated mice 6 hours after inoculation. (G & H) Cells were transfected with NF-κB reporter gene, and S.p.- or PLY-induced NF-κB promoter activity was measured as relative luciferase activity. (I & J) Cells were transiently transfected with IκBα DN, IKKα DN, IKKαsiRNA, empty vector, or control siRNA, and S.p.-induced TLR2 mRNA expression was measured by Q-PCR analysis. Data in A-D & G-J are means ± S.D. (n = 3). *, p < 0.05 compared with CON, **, p < 0.05 compared with S.p. in MOCK or WT mice, CON, control inoculated with vehicle.