Fig. 2

ASPH guides breast cancer cells to secrete pro-oncogenic/pro-metastatic extracellular vesicles. (a) Exosomes secreted by MDA-MB-231 cells stably overexpressing ASPH were actively taken up by parental breast cancer cells. (b) ECM degradation/remodeling in parental cells incubated with exosomes released from MDA-MB-231 expressing vector or ASPH. (c) Tube formation of parental cells incubated with exosomes secreted by MDA-MB-231 cells vs. with PBS. (d) Transendothelial migration and extravasation; (e) Invasion through basement membrane and subsequent mammosphere formation of parental MDA-MB-231 cells incubated with exosomes secreted by MDA-MB-231 expressing vector or ASPH. (f) Compared to empty vector, the exosomes (expressing classic biomarkers CD63 and CD9) released by MDA-MB-231 stably expressing WT-ASPH exhibited enrichment of pro-metastatic components activated Notch1, JAG1/2, CD44, EpCAM, MMPs, and ADAMs. (g) Representative protein cargoes of extracellular vesicles released by MDA-MB-231 cells expressing WT-ASPH was deciphered with proteomics using Mass Spectrometry. (h) GW4869 blocked ECM degradation of parental cells, which was rescued by addition of exosomes secreted by MDA-MB-231 stably expressing ASPH (vector to a much less extent). (i) 3-D invasion in response to GW4869. (j) Mammosphere formation in response to GW4869. (k) Transendothelial migration and extravasation; (l) Invasion through basement membrane and subsequent mammosphere formation in response to GW4869. ^*p < 0.05; ^**p < 0.01; ^***p < 0.001