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. 2015 Feb 17;5:8505. doi: 10.1038/srep08505

Figure 2. Inflamed cerebrovascular endothelial cells produce shedding-type E-EVs.

Figure 2

(a) Treatment of inflamed cells with PKH26, a plasma membrane-intercalating fluorescent reagent, clearly demonstrates that E-EVs are released from the inflamed endothelial cell membranes. Suspended b.End5 cells stained with PKH26 emit red fluorescence (Upper row). Azure-blue n indicates nucleus. PKH26-positive E-EVs are present in the culture medium (Bottom row). Red fluorescence is colocalized to the micron-sized vesicles as observed by differential interference contrast microscopy (DIC). The azure-blue arrowhead shows an E-EV. Scale bar, 10 μm. (b) Scanning electron microscopy (SEM) images showing that E-EVs are shed from the cell surface immediately after stimulation by CytoCombo + LPS (Upper row). The acute response implies that E-EV production is a non-genomic reaction. DIC image showing E-EVs present on the adhered cell surface (Bottom row), suggesting that E-EVs are bilaterally produced on both the luminal and abluminal surfaces of endothelial cells in the tube formed by the blood vessels in inflammatory conditions in vivo. Scale bar, 10 μm (a and b lower), and 1 μm (b upper).