Table 1.
TLR agonists used to stimulate primary murine AECsa
| Control treatmentb | Treatment designation | Value |
|---|---|---|
| TLR1/2 agonist Pam3CSK4 | A | 0.1 μg/ml |
| B | 0.5 μg/ml | |
| C | 1 μg/ml | |
| TLR2 agonist HKLM | A | 106 cells/ml |
| B | 107 cells/ml | |
| C | 108 cells/ml | |
| TLR2/6 agonist FSL1 | A | 10 ng/ml |
| B | 100 ng/ml | |
| C | 1 μg/ml | |
| TLR3 agonist poly(I·C) | A | 1 μg/ml |
| B | 10 μg/ml | |
| C | 100 μg/ml | |
| TLR4 agonist LPS | A | 1 ng/ml |
| B | 10 ng/ml | |
| C | 100 ng/ml | |
| TLR5 agonist ST-FLA | A | 10 ng/ml |
| B | 100 ng/ml | |
| C | 1 μg/ml | |
| TLR7 agonist ssRNA40 | A | 0.25 μg/ml |
| B | 2.5 μg/ml | |
| C | 10 μg/ml | |
| TLR9 agonist ODN1826 | A | 0.5 μM |
| B | 1 μM | |
| C | 5 μM | |
| TNF-α | 10 ng/ml | |
| Pneumocystis-to-AEC ratios of: | ||
| 0.5:1 | 0.75 × 5 Pneumocystis cysts/well | |
| Pc 1:1 | 1.5× 105 Pneumocystis cysts/well | |
| Pc 3:1 | 4.5 × 105 Pneumocystis cysts/well | |
| Pc depleted |
a
Primary murine AECs were isolated from C57BL/6 (WT) mice. Confluent monolayers were stimulated for 6 h with a panel of TLR ligands used at the indicated concentrations. CCL2 and CXCL2 levels were measured in supernatants (Fig. 5).
b
Pam3CSK4, Pam3CysSerLys4; HKLM, heat-killed preparation of L. monocytogenes; FSL1, Pam2CGDPKHPKSF; ST-FLA, flagellin from S. Typhimurium; ODN1826, CpG synthetic oligonucleotide; Pc depleted, supernatant of Pneumocystis-depleted preparation.