Fig. 2: Spermine is enriched in MCEGs.
From: Mast cell extracellular granules are bioactive condensates assembled by heparin and polyamine
a, BMMCs loaded with BODIPY–spermine (green) were stimulated with anti-TNP IgE plus TNP–BSA (IgE/Ag) for 30 min. Released granules were captured by avidin beads (top, schematic workflow) and subjected to avidin–sulforhodamine 101 (magenta) staining and fluorescence imaging (bottom left). Bottom right, quantification of fluorescence intensity of BODIPY–spermine and avidin along the indicated yellow dashed line in the merged image. Scale bar, 5 μm. b, Top 25 proteins identified in extracellular granules by mass spectrometry analysis were plotted against their molecular weight (MW). Gzmb, granzyme B; CMA1, chymase; Mcpt4, mast cell protease 4; Tpsb2, tryptase β2; Anpep, aminopeptidase N. c, TNF, IL-6, CCL3 and GM-CSF concentrations in the extracellular granules released by 1 million resting and IgE/Ag-stimulated BMMCs were measured by LEGENDplex (n = 2 biological replicates). IFN, interferon. d, MCEGs released from BMMCs expressing mCherry–tryptase (red) or mCherry–TNF (magenta) preloaded with 20 μM BODIPY–spermine (green) were collected by avidin beads and subjected to confocal imaging. White arrows denote MCEGs. Scale bar, 5 μm. e, Heparin–spermine condensate formation with TNF. Condensates were formed with 50 μM heparin and 5 mM spermine in the presence of indicated TNF concentrations. Bottom, quantification of the clustering levels and partition coefficients of spermine and heparin. Data represent the analysis of n = 20 individual images per group. Scale bar, 10 μm. Data are presented as the mean ± s.d. One representative of three independent experiments is shown.
