LMWH prevents platelet and EV-induced inflammasome activation via HBEGF-AKT signaling. (A-B) LMWH activates HBEGF-AKT signaling in placenta. Immunoblots of placenta from EV-injected pregnant mice (representative immunoblots [A]; bar graphs summarizing results [B]) showing restored HBEGF, pEGFR, pAKT, and pERK1/2 levels upon LMWH treatment. Placentas were assessed from C57BL/6 mice at day 12.5 p.c. after IV injection of mouse endothelial cell–derived procoagulant EVs at days 10.5 p.c. and 11.5 p.c. Control mice (C) were injected with the supernatant obtained after the last PBS wash during EV isolation. (C-D) Inhibition of HBEGF (HBEGFi, CRM197) or AKT (AKTi, LY294002) in vitro inhibits (representative immunoblots [C]; bar graphs summarizing results [D]) LMWH-mediated protective effects on platelet and EV-induced expression of NLRP3, cleaved IL-1β, and cleaved caspase-1. Cells were pretreated respective inhibitors before LMWH treatment. Control cells, C, were exposed to supernatant obtained after the last PBS wash during EV isolation. EV represents cells exposed to EV and platelets (PRP); EV+LMWH, cells pretreated with LMWH and exposed to EVs and platelets; EV+LMWH+HBEGFi, cells pretreated with HBEGFi and LMWH and exposed to EVs and platelets; EV+LMWH+AKTi, cells pretreated with AKTi and LMWH and exposed to EVs and platelets. Inactive (proform) and active (cleaved form) of IL-1β or caspase-1 are indicated in panels A,C. Only the active form (cleaved form) was quantified in panels B,D. Data shown represent mean ± SEM from 6 to 8 placentas from 3 dams for panels A-B or 3 independent repeat experiments for panels C-D. ∗P < .05, ns; ANOVA; Šídák multiple comparisons test. EV, EV-injected mice; EV+LMWH, EV-injected mice with LMWH injections.