Loss of Rap1b leads to increased neutrophil infiltration to inflamed lungs. (A) Immunoblot analysis of Rap1b in the DRMs of WT and Cdc42^−/− neutrophils. (B) Immunoblotting of neutrophil lysates from mice reconstituted with WT and Rap1b^−/− BM using anti-Rap1b, anti-total Rap1, and β-actin antibodies. Blots are representative of three independent experiments. (C) BAL analysis of mice reconstituted with WT or Rap1b^−/− hematopoietic cells that were challenged with interstitial saline or LPS (1.25 mg/kg). Numbers of neutrophils, RBCs, and macrophages recovered in BAL at 4 and 24 h after LPS challenge; n = 4 mice per group; two independent experiments. (D) Total protein analysis of BAL fluid (BALF) at 4 and 24 h after LPS challenge. (E) Hematoxylin and eosin staining of lung sections 24 h after saline or LPS challenge. Bar, 20 µm. Quantification of neutrophil infiltration in interstitial tissue and alveolar spaces using lung sections from n = 6 mice per group from three independent experiments. (F) Relative recruitment of CSFE and SNARF1 labeled and adoptively transferred WT and Rap1b^−/− neutrophils in BAL of LPS challenged mice. Data are representative of three independent experiments with 3 mice each group (C–F; mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001; NS, not significant using unpaired Student’s t test). (G) Survival of mice (n = 8 per genotype) injected i.p. with 20 mg/kg LPS. **, P = 0.00259, log-rank test. Representative of two independent experiments.