Article Figures & Data
Figures
- FIGURE 1.
Expression of human Toll receptors in CHO cells. Clonal CHO cell lines transfected with FLAG-epitope-tagged cDNA were stained by indirect immunofluorescence using anti-Flag mAb and analyzed by flow cytometry. “Control” represents the same cells stained only with FITC-anti-IgGmu.
- FIGURE 2.
TLR2, but not TLR4, expression imparts responsiveness to S. aureus synergistically with CD14. A, CHO/Neo, CHO/TLR2, CHO/CD14, and CHO/CD14/TLR2 were treated with PBS or stimulated with heat-killed S. aureus (108 or 109 CFU) for 45 min. Nuclear extracts from these cells were assessed for the presence of NF-κB using the EMSA. Shown are the NF-κB/32P-labeled probe complexes. B, CHO/CD14/TLR2 and CHO/CD14/TLR4 reporter cell lines that express surface CD25 as a result of NF-κB translocation (15 ) were exposed to either PBS or heat-killed S. aureus for 18 h. The cells were stained with FITC-labeled anti-CD25 mAb and subjected to flow cytometric analysis for transgene expression. Not shown are all cell lines responded equivalently to TNF-α (10 ng/ml) and IL-1β (5 ng/ml).
- FIGURE 3.
TLR2, but not TLR4, mediates cellular activation by heat-killed S. pneumoniae. A, CHO/CD14 and CHO/CD14/TLR2 reporter cells were treated with PBS or with a clinical strain of heat-killed S. pneumoniae for 18 h. The cells were stained with FITC-labeled anti-CD25 mAb and subjected to flow cytometric analysis for the expression of the NF-κB-dependent transgene (CD25). B, Because of the potential role of pneumolysin as a nonspecific activator of cells, a ply mutant of S. pneumoniae (P-) was used to activate CHO/CD14 reporter cells and its derivative human TLR-expressing cell lines. After 18 h, cells were again analyzed for CD25 expression by flow microfluorometry.
- FIGURE 4.
Soluble PGN from S. aureus activates TLR2 but not TLR4 transfected CHO/CD14 cells. Clonal cell lines expressing CD14 and human TLRs were stimulated with PBS or Staphylococcal sPGN (1, 10, or 100 nM) for 45 min. Nuclear extracts were prepared, and the nuclear levels of NF-κB were determined by the gel shift assay.
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