Human FcγRIIIA mediates both inhibition and activation through the FcRγ ITAM depending on the type of ligand. (A) FcγRIIIA chimeric constructs are schematically presented and their surface expression levels after transfection into RBL-2H3 cells are shown compared with nontransfected cells (gray vs blue histograms). (B) IgE-sensitized RBL-2H3 transfectants were incubated with either 3G8 F(ab′)2 or 320 F(ab′)2 at 10 μg/mL for 30 minutes at 37°C as indicated and degranulation was triggered with DNP-HSA (0.1 μg/mL) for 45 minutes. Net β-hexosaminidase release was determined. (*P < .05; n = 3; nonparametric Mann-Whitney test). (C) RBL-2H3 transfectants were incubated with 10 μg/mL of 3G8 F(ab′)2 at 4°C before stimulation with rabbit anti–mouse F(ab′)2 (RAM at 40 μg/mL) for 45 minutes at 37°C. Net β-hexosaminidase release was determined. (**P < .01; n = 3; nonparametric Mann-Whitney test). (D) IgE-sensitized huFcγRIII+ RBL-2H3 transfectants were preincubated at 37°C with either BSA at 10 mg/mL for 240 minutes or 3G8 F(ab′)2 or with IVIg, at the indicated concentrations and for the indicated time. Degranulation was measured as in panel B (*P < .05; n = 3, nonparametric Mann-Whitney test).
