Figure 6
Figure 6. P-Rex and Vav control the pulmonary recruitment of eosinophils, monocytes/macrophages, and lymphocytes during Ova-induced lung allergy in a platelet- and leukocyte-adhesion–dependent manner. (A-B) Absence of pulmonary leukocyte recruitment in P1V1 and P1V3 mice during Ova-induced lung allergy. Mice were sensitized to Ova, challenged with aerosolized Ova to induce allergy, or were mock treated, and pulmonary recruitment of eosinophils, monocytes/macrophages, lymphocytes, or total leukocytes, as indicated, was assessed by lung lavage 24 hours after Ova challenge (A), as were total leukocyte numbers in peripheral blood (B). Data are mean ± SEM of 3 experiments with 4 to 14 mice/group; statistics in (A) unpaired Student t test between sham and Ova treatment, and 1-way ANOVA with Tukey’s multiple comparisons or Kruskal-Wallis with Dunn’s multiple comparisons for genotypes; statistics in (B) are 2-way ANOVA with Bonferroni multiple comparisons. (C) Normal IL-5–dependent eosinophil mobilization. Mice were IV injected with IL-5 to induce eosinophil mobilization from the bone marrow into the blood stream or were sham treated, and eosinophil numbers in the peripheral blood were assessed after 24 hours by enzyme-linked immunosorbent assay. Data are mean ± SEM of 3 mice/group; statistics as in (B). (D-E) Reduced Ova-induced leukocyte transmigration and airway infiltration in lungs of P1V1 and P1V3 mice. Quantitative histologic analysis of leukocyte transmigration into lung tissue (D) and airway infiltration (E) during Ova-dependent lung inflammation, quantitated as in Figure 2B-D. Data are mean ± SEM of 4 to 9 mice/group; statistics 2-way ANOVA with Bonferroni multiple comparisons. (F) Reconstitution of Ova-induced pulmonary eosinophil recruitment in platelet-depleted PVWT mice with PVWT but not P1V1 or P1V3 platelets. Platelets were transfused into thrombocytopenic PVWT mice as in Figure 5 and eosinophil recruitment into the Ova-inflamed lung was assessed as in (A). Data are mean ± SEM of n = 4 to 8 mice/group; statistics 1-way ANOVA with Bonferroni multiple comparisons. (G) Altered eosinophil rolling and impaired adhesion in postcapillary venules of the Ova-inflamed allergic lung. Lung allergy was induced as in (A) and intravital microscopy of the tracheal postcapillary venules was performed and analyzed as in Figure 3 for numbers of rolling eosinophils (i), rolling velocity (ii; both mean velocity and categories), and firm adhesion (iii). Data are mean ± SEM of 2 to 4 sham and 4 to 6 Ova-treated mice/group for numbers of rolling and adhering cells. Rolling velocity is from ≥120 cells; statistics as in (D).

P-Rex and Vav control the pulmonary recruitment of eosinophils, monocytes/macrophages, and lymphocytes during Ova-induced lung allergy in a platelet- and leukocyte-adhesion–dependent manner. (A-B) Absence of pulmonary leukocyte recruitment in P1V1 and P1V3 mice during Ova-induced lung allergy. Mice were sensitized to Ova, challenged with aerosolized Ova to induce allergy, or were mock treated, and pulmonary recruitment of eosinophils, monocytes/macrophages, lymphocytes, or total leukocytes, as indicated, was assessed by lung lavage 24 hours after Ova challenge (A), as were total leukocyte numbers in peripheral blood (B). Data are mean ± SEM of 3 experiments with 4 to 14 mice/group; statistics in (A) unpaired Student t test between sham and Ova treatment, and 1-way ANOVA with Tukey’s multiple comparisons or Kruskal-Wallis with Dunn’s multiple comparisons for genotypes; statistics in (B) are 2-way ANOVA with Bonferroni multiple comparisons. (C) Normal IL-5–dependent eosinophil mobilization. Mice were IV injected with IL-5 to induce eosinophil mobilization from the bone marrow into the blood stream or were sham treated, and eosinophil numbers in the peripheral blood were assessed after 24 hours by enzyme-linked immunosorbent assay. Data are mean ± SEM of 3 mice/group; statistics as in (B). (D-E) Reduced Ova-induced leukocyte transmigration and airway infiltration in lungs of P1V1 and P1V3 mice. Quantitative histologic analysis of leukocyte transmigration into lung tissue (D) and airway infiltration (E) during Ova-dependent lung inflammation, quantitated as in Figure 2B-D. Data are mean ± SEM of 4 to 9 mice/group; statistics 2-way ANOVA with Bonferroni multiple comparisons. (F) Reconstitution of Ova-induced pulmonary eosinophil recruitment in platelet-depleted PVWT mice with PVWT but not P1V1 or P1V3 platelets. Platelets were transfused into thrombocytopenic PVWT mice as in Figure 5 and eosinophil recruitment into the Ova-inflamed lung was assessed as in (A). Data are mean ± SEM of n = 4 to 8 mice/group; statistics 1-way ANOVA with Bonferroni multiple comparisons. (G) Altered eosinophil rolling and impaired adhesion in postcapillary venules of the Ova-inflamed allergic lung. Lung allergy was induced as in (A) and intravital microscopy of the tracheal postcapillary venules was performed and analyzed as in Figure 3 for numbers of rolling eosinophils (i), rolling velocity (ii; both mean velocity and categories), and firm adhesion (iii). Data are mean ± SEM of 2 to 4 sham and 4 to 6 Ova-treated mice/group for numbers of rolling and adhering cells. Rolling velocity is from ≥120 cells; statistics as in (D).

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