Fig. 4: Intracellular Ca²⁺ influx-mediated EV biosynthesis.

a Intracellular Ca²⁺ in M2 BV2 macrophages under electrical stimulation of ePOWER at different time points. Ca²⁺ was probed with fluo-4 dye (scale bars, 20 μm). b Quantification of the varied calcium ions in (a). c, d Flow cytometry analysis of the intracellular Ca²⁺ level after stimulating cells with either ePOWER or CaCl₂ incubation (c) and corresponding quantification (d) (n = 3). e Quantitative comparison of EV generation between the treatments of CaCl₂, ePOWER, and the addition of EGTA plus ePOWER stimulation (n = 3). f Representative TEM images of cells with and without ePOWER stimulation. ePOWER stimulation leads to MVBs with more intraluminal vesicles (scale bars, 0.5 μm). Statistical differences were analyzed using one-way ANOVA with Turkey’s multiple comparisons test. The data is represented by an average of ± SD.