Inflammasome-induced exosome secretion in HeLa cells. (A) Secretion of IL-18 in SeV-infected HeLa cells. Cont., control. (B) SeV infection activates the inflammasome as noted by the increase in cleaved caspase-1. (C) The caspase-1 inhibitor VI (Z-YVAD-FMK, 100 µM) inhibits (Inh.) RILP cleavage in SeV-infected HeLa cells. (D) Expression of the NLRP3 component of the inflammasome increases basal RILP cleavage. (E) SeV infection increases exosome secretion in HeLa cells by approximately eightfold. (F) EVs isolated from SeV-infected HeLa cells can be characterized by the exosome markers CD63, heat shock protein 70, Flotillin, and Tsg101. They do not contain Annexin A1 (microvesicles), histone H2B (apoptotic bodies), GM10 (Golgi), or Tom20 (mitochondria). All markers were undetected in exosome-depleted medium. WCL, whole-cell lysate. Exo, exosome. (G) Overexpression of empty vector (MG) or specific miRNAs does not alter exosome secretion. Data are shown as mean ± SD; *, P ≤ 0.05 compared with untreated for n = 4–6. (H) Intracellular expression of overexpressed miRNAs.