Fig. 1.

SLC15A4 is required for the proper regulation of mast-cell effector functions. (A) Histamine levels in the culture supernatant or cell lysates of BMMCs from WT or Slc15a4^−/− mice, determined by competitive EIA. Means and SD of three technical replicates are shown, and data are representative of three independent experiments. **P < 0.01. (B) Histidine decarboxylase (Hdc) levels in WT and Slc15a4^−/− BMMCs were determined by quantitative RT–PCR. Means and SD of three technical replicates are shown, and data are representative of three independent experiments. *P < 0.05. (C) Serotonin in the culture supernatant or cell lysates of WT and Slc15a4^−/− BMMCs upon IgE-mediated FcεRI ligation was quantified by competitive EIA. Means and SD of three technical replicates are shown, and data are representative of two independent experiments. **P < 0.01. (D) Cell-surface LAMP1 was detected on WT and Slc15a4^−/− BMMCs by flow cytometry. BMMCs were sensitized with anti-TNP IgE and stimulated by TNP₄-BSA. Histograms represent LAMP1 levels without stimulation or 15 min after stimulation, respectively (left). The proportion of LAMP1⁺ cells was tracked during IgE-mediated degranulation (~120 min.) (right); LPS was used as a negative control. Results are representative of three independent experiments. (E) Degranulation of WT or Slc15a4^−/− BMMCs was measured by β-Hex activity in the culture supernatant at 15 min (left), or over 30 min (right). ***P < 0.001. (F, G) IgE-binding capacity on the surface of WT or Slc15a4^−/− BMMCs. The amount of bound IgE was measured by flow cytometry. Results are representative of three independent experiments. (H) Morphology of histamine-containing granules in BMMCs. The intracellular histamine in WT or Slc15a4^−/− BMMCs was detected by anti-Histamine antibody and visualized on confocal microscopy. Results are representative of three independent experiments.