Fig. 4.

Inactivation of PI3K p110δ with IC87114 inhibits eosinophil migration and prevents eotaxin-1-induced changes in cell morphology. A: migration of fluorescently labeled BM-Eos treated with IC87114 or vehicle toward 100 nM murine eotaxin-1 in Transwell plates after 2 h at 37°C. Combined data from 5 independent experiments in duplicate are shown. B: expression of CCR3, the receptor for eotaxin-1, by vehicle- and IC87114-treated eosinophils by flow cytometry using FITC-conjugated anti-mouse CCR3 (5 μg/ml). Rat IgG2a was used as the isotype control. Histogram shown is representative of 3 experiments with eosinophils from 3 different mice. C: confocal microscopy of FITC-phalloidin-stained BM-Eos adhering to VCAM-1 or ICAM-1 in the presence of vehicle or 10 μM IC87114 for 20 min followed by eotaxin-1 (100 nM, 5 min). Magnification ×600. D: quantitation of adherent vehicle- and IC87114-treated BM-Eos exhibiting changes in morphology after eotaxin-1 treatment. Adhered cells in 5 different fields of each coverslip exhibiting cell spreading with distinct leading edges, lamellipodia, or filopodia after eotaxin-1 treatment were counted and expressed as a percentage of the total number of cells in the field. Combined data of 3 experiments for adhesion on VCAM-1 and 2 experiments for adhesion on ICAM-1 are shown. Data represent means ± SE in A and D. *P < 0.05 in A and < 0.01 in D for comparison of IC87114- vs. vehicle-treated eosinophils.