Extended Data Fig. 7: THY1 signalling facilitate neutrophil unhealthy mitochondria extrusion via migrasome.
a, Schematic representation of the protocol used for MEP sorting. Mitochondrial proteins from each indicated layer were analysed by immunoblotting (n = 5). b, Schematic overview of the isolation of different THY1-CM fractions. Marker proteins for migrasomes, exosomes, and apoptotic bodies in the purified THY1-CM fractions obtained from the same centrifugation process were analysed by immunoblotting. Samples were normalized to total protein concentration determined by the bicinchoninic acid (BCA) assay (n = 5). c, Time-lapse imaging of untreated neutrophils. The white dashed line outlined the cells. (n = 3; scale bar, 5 µm). The cell membrane was labelled with DIL. d, Membrane potential in excreted and intracellular mitochondria from untreated or THY1-Fc -treated neutrophils was analysed using Mitotracker Deep Red staining. (n = 3; scale bar, 5 µm). Representative images of THY1-Fc-treated neutrophils were shown. CFSE staining was employed to outline the cells. e, Lysosome-, endoplasmic reticulum-, and Golgi-related proteins in migrasomes from THY1-Fc-treated neutrophils were analysed (n = 5). f, Dot plot of ITGAM expression in each cell type, with dot size representing the fraction of expressing cells and colour indicating mean expression intensity, based on a publicly available scRNA dataset (skrx2fz79n). g, Monocytes were left untreated or treated with THY1-Fc. Migrasomal and mitochondrial proteins in MEP derived from CM were determined (n = 5). Data are from five (a,b,e,g) independent experiments. Data are presented as mean ± s.e.m. of over 100 cells (c,d) across three independent experiments. Statistical significance was determined using unpaired two-tailed Student’s t-test (d) or the two-sided Wilcoxon rank-sum test, and false discovery rate correction was applied for multiple comparisons across cell types (f). Violin plots show data distribution with median and interquartile range (d). Panel b created with BioRender.com.
