Fig. 2: Midlife APC-derived EVs exhibit a reduced capacity to maintain adipose immune homeostasis.
a A t-SNE plot displaying single-nucleus RNA sequencing data of cells from WAT samples from young and middle-aged humans. b Cell identity annotation in human WAT samples. c Number of DEGs for each identified cell subgroup. d, e Western blot results of p19ARF and p53 in mouse APCs (d), P16Ink4a and P21 in human APCs (e) from the young (mice, 3-month-old; humans, aged 18-29), middle-aged (mice, 12-month-old; humans, aged 30-49) and aged (mice, 24-month-old; humans, aged 50-69) groups (n = 3 independent experiments). f, g Transcriptional levels of senescence-associated genes in the APCs of mice (p16Ink4a, p21, Il-6, and Igfbp3) (f) and humans (P16Ink4a, P21, IL-6, and IGFBP3) (g) from the indicated groups (n = 6 per group). h Number of DEGs annotated through gene ontology (GO) enrichment-based clustering based on bulk RNA-seq analysis of young and middle-aged APCs from mice or humans. i The engulfment of PKH26-labeled EVs (red) by APCs, adipocytes and macrophages was detected by confocal microscopy. The experiments were repeated independently three times. Scale bar, 20 µm. j The engulfment of yEVs and mEVs by macrophages was detected by flow cytometry. k Western blot analysis of iNOS expression from BMDMs of blank, yEVs and mEVs groups in the presence of PBS or LPS (n = 3 per group). l RT‒qPCR results of inflammation-associated genes (Tnf-α, Il-1β, and Il-6) in BMDMs of blank, yEVs and mEVs groups in the presence of PBS or LPS (n = 3 biological replicates per group). The data were presented as mean ± SEM. Statistical significance was assessed by one-way ANOVA (f, g, bottom row in k, l). See also Supplementary Fig. 2 and Supplementary Fig. 3. Source data are provided as a Source Data file.
