Fig. 3: Depletion of spermine impedes MCEG assembly.
From: Mast cell extracellular granules are bioactive condensates assembled by heparin and polyamine
a, Schematic illustration of the chemical inhibitors used in this study and their targets. b, MCEGs released from chemical inhibitor-treated BMMCs before and after anti-TNP IgE plus TNP–BSA (IgE/Ag) stimulation were collected with avidin beads. Granules were visualized by avidin–sulforhodamine 101 staining and the number of granules on each bead was enumerated (n = 50 images per group). P values were determined using a Kruskal–Wallis with Dunn’s multiple-comparison test. c, Protein lysates prepared from MCEGs released by 5 million inhibitor-treated BMMCs were analyzed by SDS–PAGE and Coomassie blue staining. d,e, Western blot analysis of mast cell mediators in MCEGs (d) and in the soluble fraction (e). Band intensities for each mediator released in MCEGs (d) and soluble fractions (e) after IgE/Ag stimulation were quantified and normalized to the corresponding mediator in whole-cell lysates (Extended Data Fig. 3c). Note that TNF and IL-1β appeared as multiple bands on the blot because of different isoforms and post-translational modifications113,114,115,116. These bands were included in quantification. Right, all normalized values are expressed as the fold change relative to the vehicle control under IgE/Ag stimulation (set as 1) and plotted (each data point represents one independent experiment; n = 3). P values were determined using an ordinary one-way ANOVA with Dunn’s multiple-comparison test. f, Degranulation capacity of inhibitors-treated BMMCs was analyzed by the β-hexosaminidase assay (n = 3 technical replicates). Data are presented as the mean ± s.d. One representative of two (b) or three (c–f) independent experiments is shown.
