FcγRIIA activates mouse and human mast cells in vitro. (A) Representative expression of FcγRIIA on PCMCs, FSMCs, and BMMCs from 3KOIIA (open histogram) and 3KO mice (filled histogram). (B) PCMCs from indicated mice were incubated with indicated preformed mouse IgG-IC (αOVA: anti-OVA mAb; αGPI: polyclonal anti-GPI antiserum; open histograms) or not (filled histograms). Binding of ICs was detected by staining with F(ab′)2 GAM-PE. (C) Calcium fluxes in PCMCs from 3KO or 3KOIIA mice incubated with indicated IC (black curves) or Ag alone (gray curves). Ionomycin was used as control. (D) Western blot analysis of PCMC lysates after stimulation with indicated reagents for different periods of time. PCMCs sensitized overnight with IgE anti-DNP and challenged with DNP-HSA for 3 minutes served as positive controls. Actin was used as a loading control. FcγRIIA was used as a genotype control (reprobe after pERK1/2 staining). (E-F) Mediator release by PCMCs from 3KO (open bars) and 3KOIIA (black bars) mice challenged with indicated reagents. PCMCs sensitized overnight with IgE anti-DNP and challenged with DNP-HSA served as positive controls. NB: GPI+αGPI correspond to ICs made of GPI and polyclonal anti-GPI antiserum in panel E, and to ICs made of GPI and IgG purified from anti-GPI antiserum in panel F. (G) Representative histogram plots of human FcR expression on human SMCs. (H) Percentage of β-hexosaminidase release and quantification of histamine release by human SMCs incubated with anti-FcϵRI mAb or with preformed complexes of mAb IV.3 and GAM. (E-F,H) Data are represented as mean ± SEM. (A-H) Data are representative from at least 2 independent experiments (*P < .05; **P < .01; ***P < .001).
