Figure 1.
LMWH prevents EV-mediated placental dysfunction. (A-C) Representative images (A; top, embryos; bottom, placenta) and bar graphs (B-C) showing reduced fetal death (B) and improved embryonic height (C) after LMWH treatment in EV-injected mice. Pregnancy outcome assessed in C57BL/6 mice at day 12.5 p.c. after IV injection of mouse endothelial cell–derived procoagulant EVs at day 10.5 p.c. and 11.5 p.c. (D-E) Hematoxylin and eosin (H&E) staining of murine placenta (representative images [D]; bar graph summarizing results [E]) showing LMWH prevents EV-induced enhanced maternal vascularization (blood lacunae; enucleated erythrocytes, arrows) and reduced fetal vascularization (nucleated erythrocytes, arrow heads) after EV injections. Vascularized areas were calculated using ImageJ. Analyses performed at day 12.5 p.c. Size bar represent 20 μm. (F-G) Ki-67 staining on murine placenta (representative images [F]; bar graph summarizing results [G]) showing LMWH restores EV-induced impaired placental proliferation. Size bar represent 20 μm. (H) Representative Immunostaining images from mice placenta showing increased CD62P (P-selectin, red; DAPI [4′,6-diamidino-2-phenylindole]–stained nucleus, blue) signals in placentas from EV-injected dams, suggesting increased activated platelets within the placenta. LMWH-treated mice (EV+LMWH) showed reduced CD62P signaling, suggesting LMWH prevents platelet activation. Size bar represent 20 μm. (I) Bar graphs summarizing results from qRT-PCR for mouse trophoblast differentiation gene (PL-II, Tpbpa, Esx1, and Gcm-1) showing that LMWH prevents EV-induced abnormal placental differentiation. Control mice, C, were injected with the supernatant obtained after the last PBS wash during EV isolation. For panels B-C,E,G,I, data shown represent mean ± standard error of the mean (SEM); n = 5 to 8 placentas from 3 dams (∗P < .05, nonsignificant [ns]; analysis of variance [ANOVA]; Šídák multiple comparisons test). C, control; EV, extracellular vesicle (EV)-injected mice; EV+LMWH, EV-injected mice with low-molecular weight heaprin (LMWH) injections; qRT-PCR, quantitative reverse transcriptase-polymerase chain reaction; PL-II, placental lactogen-II; Tpbpa, trophoblast specific protein alpha; Esx1, Extraembryonic, Spermatogenesis, Homeobox 1 and Gcm-1, Glial Cells Missing Transcription Factor 1.

LMWH prevents EV-mediated placental dysfunction. (A-C) Representative images (A; top, embryos; bottom, placenta) and bar graphs (B-C) showing reduced fetal death (B) and improved embryonic height (C) after LMWH treatment in EV-injected mice. Pregnancy outcome assessed in C57BL/6 mice at day 12.5 p.c. after IV injection of mouse endothelial cell–derived procoagulant EVs at day 10.5 p.c. and 11.5 p.c. (D-E) Hematoxylin and eosin (H&E) staining of murine placenta (representative images [D]; bar graph summarizing results [E]) showing LMWH prevents EV-induced enhanced maternal vascularization (blood lacunae; enucleated erythrocytes, arrows) and reduced fetal vascularization (nucleated erythrocytes, arrow heads) after EV injections. Vascularized areas were calculated using ImageJ. Analyses performed at day 12.5 p.c. Size bar represent 20 μm. (F-G) Ki-67 staining on murine placenta (representative images [F]; bar graph summarizing results [G]) showing LMWH restores EV-induced impaired placental proliferation. Size bar represent 20 μm. (H) Representative Immunostaining images from mice placenta showing increased CD62P (P-selectin, red; DAPI [4′,6-diamidino-2-phenylindole]–stained nucleus, blue) signals in placentas from EV-injected dams, suggesting increased activated platelets within the placenta. LMWH-treated mice (EV+LMWH) showed reduced CD62P signaling, suggesting LMWH prevents platelet activation. Size bar represent 20 μm. (I) Bar graphs summarizing results from qRT-PCR for mouse trophoblast differentiation gene (PL-II, Tpbpa, Esx1, and Gcm-1) showing that LMWH prevents EV-induced abnormal placental differentiation. Control mice, C, were injected with the supernatant obtained after the last PBS wash during EV isolation. For panels B-C,E,G,I, data shown represent mean ± standard error of the mean (SEM); n = 5 to 8 placentas from 3 dams (∗P < .05, nonsignificant [ns]; analysis of variance [ANOVA]; Šídák multiple comparisons test). C, control; EV, extracellular vesicle (EV)-injected mice; EV+LMWH, EV-injected mice with low-molecular weight heaprin (LMWH) injections; qRT-PCR, quantitative reverse transcriptase-polymerase chain reaction; PL-II, placental lactogen-II; Tpbpa, trophoblast specific protein alpha; Esx1, Extraembryonic, Spermatogenesis, Homeobox 1 and Gcm-1, Glial Cells Missing Transcription Factor 1.

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