FIG. 3.

Impaired recruitment of neutrophils and CD4⁺ T cells to the lung in the absence of IL-1R1. Groups of mice were infected intranasally with 100 PFU influenza virus PR8. Lung-infiltrating cells were isolated by BAL. (A) Total BAL cell number (T) was measured using a Coulter Counter (Instrumenten Gesellschaft IG, Basel, Switzerland), and eosinophils (E), neutrophils (N), monocytes/macrophages (M), and lymphocytes (L) were determined by differential counts according to morphological criteria as described in Materials and Methods. Numbers represent average values for four mice ± standard deviations. (B) On days 7 and 10, BAL cells were incubated with anti-CD32/CD16 MAb to block unspecific binding and subsequently stained with allophycocyanin-labeled anti-CD4 and PE-labeled anti-CD8 MAbs. Cells were acquired using a FACSCalibur (Becton Dickinson) and data analyzed with Cell Quest software. CD4⁺ and CD8⁺ T-cell numbers were calculated from percentages of BAL total cell numbers excluding dead cells. Values show averages for four mice per group. *, significant differences (Student test). (C) On days 7 (upper panels) and 10 (lower panels), aliquots of BAL cells were stained with PE-conjugated class I tetramers loaded with influenza virus peptide NP68 and allophycocyanin-conjugated anti-CD8 MAb before analysis by flow cytometry. Values show percentages of virus-specific CD8⁺ T cells gated on lymphocytes of an individual mouse representative of the group. Average percentages from groups of mice (n = 4) are shown in parentheses.