Figure 1.
Defining the thrombocytopenic threshold for in utero ICH. (A) Experimental plan. E13.5 pregnant mice were injected with anti-GP1Bα antibody at a series of concentrations and analyzed 48 hours after injection for platelet numbers and hemorrhage phenotype. IgG antibody at 5 µg/g was used as a control. (B) Frequency of circulating platelets in the peripheral blood in E13.5 + 48 hours (E15.5) fetuses. No injection control: n = 7; IgG: 5 µg/g, n = 16; anti-GP1Bα: 0.5 µg/g, n = 15; 1.0 µg/g, n = 6; 1.5 µg/g, n = 14; 1.75 µg/g, n = 14; 2.0 µg/g, n = 18; 2.5 µg/g, n = 15; 5.0 µg/g, n = 15. ****P < .0001. (C) ICH frequency at 48 hours after no injection control: n = 7; IgG: 5 µg/g, n = 16; anti-GP1Bα: 0.5 µg/g, n = 15; 1.0 µg/g, n = 6; 1.5 µg/g, n = 14; 1.75 µg/g, n = 14; 2.0 µg/g, n = 18; 2.5 µg/g, n = 15; 5.0 µg/g, n = 15. **P = .002; ****P = .000007. (D) Representative images of E13.5 + 48 hours (E15.5) fetuses and dissected brains after treatment with IgG control or anti-GP1Bα antibody. Fetuses were classified according to the percentage of normal platelet count. Images shown represent the most frequent severity of hemorrhage observed in each group (scale bars represent 1 mm). (E) Frequency of ICH at platelet counts: 0% to 10%, n = 13; 11% to 20%, n = 17; 21% to 30%, n = 12; 31% to 60%, n = 8; 100%, n = 14; IgG control: n = 14. **P = .004; ****P < .00001. (F) ICH severity scores at platelet counts: 5% to 10%, n = 10; 11% to 15%, n = 6; 16% to 20%, n = 5; 21% to 25%, n = 5; 31% to 40%, n = 3; 41% to 60%, n = 8; 100% IgG: n = 8. Arbitrary values were used to define 3 as severe ICH (intraventricular involvement and/or the presence of large cortical hemorrhage), 2 as moderate ICH (large focal hemorrhage), 1 as mild ICH (small focal hemorrhage), and 0 as no ICH. *P = .05; **P < .001; ***P = .0006. (G) Lymphatic bleed severity scores: 0% to 5%, n = 8; platelet counts: 6% to 10%, n = 11; 11% to 15%, n = 6; 16% to 20%, n = 5; 21% to 25%, n = 4; 31% to 40%, n = 2; 41% to 60%, n = 8; 100% IgG: n = 8. Arbitrary values were used to define 3 as severe hemorrhage (edema, dermal hemorrhage, and extensive blood-filled lymphatics), 2 as moderate hemorrhage (extensive blood-filled lymphatics), 1 as mild hemorrhage (blood-filled lymphatics or focal dermal hemorrhage), and 0 as no hemorrhage. *P < .05; **P = .005. Data were analyzed with one-way analysis of variance (ANOVA) using the Šídák multiple comparisons test (B), or by using contingency table analysis with Fisher’s exact test (C,E-G). P values were adjusted for multiple testing using the Holm-Šídák method (C,E-G). ns, not statistically significant; Plts, platelets.

Defining the thrombocytopenic threshold for in utero ICH. (A) Experimental plan. E13.5 pregnant mice were injected with anti-GP1Bα antibody at a series of concentrations and analyzed 48 hours after injection for platelet numbers and hemorrhage phenotype. IgG antibody at 5 µg/g was used as a control. (B) Frequency of circulating platelets in the peripheral blood in E13.5 + 48 hours (E15.5) fetuses. No injection control: n = 7; IgG: 5 µg/g, n = 16; anti-GP1Bα: 0.5 µg/g, n = 15; 1.0 µg/g, n = 6; 1.5 µg/g, n = 14; 1.75 µg/g, n = 14; 2.0 µg/g, n = 18; 2.5 µg/g, n = 15; 5.0 µg/g, n = 15. ****P < .0001. (C) ICH frequency at 48 hours after no injection control: n = 7; IgG: 5 µg/g, n = 16; anti-GP1Bα: 0.5 µg/g, n = 15; 1.0 µg/g, n = 6; 1.5 µg/g, n = 14; 1.75 µg/g, n = 14; 2.0 µg/g, n = 18; 2.5 µg/g, n = 15; 5.0 µg/g, n = 15. **P = .002; ****P = .000007. (D) Representative images of E13.5 + 48 hours (E15.5) fetuses and dissected brains after treatment with IgG control or anti-GP1Bα antibody. Fetuses were classified according to the percentage of normal platelet count. Images shown represent the most frequent severity of hemorrhage observed in each group (scale bars represent 1 mm). (E) Frequency of ICH at platelet counts: 0% to 10%, n = 13; 11% to 20%, n = 17; 21% to 30%, n = 12; 31% to 60%, n = 8; 100%, n = 14; IgG control: n = 14. **P = .004; ****P < .00001. (F) ICH severity scores at platelet counts: 5% to 10%, n = 10; 11% to 15%, n = 6; 16% to 20%, n = 5; 21% to 25%, n = 5; 31% to 40%, n = 3; 41% to 60%, n = 8; 100% IgG: n = 8. Arbitrary values were used to define 3 as severe ICH (intraventricular involvement and/or the presence of large cortical hemorrhage), 2 as moderate ICH (large focal hemorrhage), 1 as mild ICH (small focal hemorrhage), and 0 as no ICH. *P = .05; **P < .001; ***P = .0006. (G) Lymphatic bleed severity scores: 0% to 5%, n = 8; platelet counts: 6% to 10%, n = 11; 11% to 15%, n = 6; 16% to 20%, n = 5; 21% to 25%, n = 4; 31% to 40%, n = 2; 41% to 60%, n = 8; 100% IgG: n = 8. Arbitrary values were used to define 3 as severe hemorrhage (edema, dermal hemorrhage, and extensive blood-filled lymphatics), 2 as moderate hemorrhage (extensive blood-filled lymphatics), 1 as mild hemorrhage (blood-filled lymphatics or focal dermal hemorrhage), and 0 as no hemorrhage. *P < .05; **P = .005. Data were analyzed with one-way analysis of variance (ANOVA) using the Šídák multiple comparisons test (B), or by using contingency table analysis with Fisher’s exact test (C,E-G). P values were adjusted for multiple testing using the Holm-Šídák method (C,E-G). ns, not statistically significant; Plts, platelets.

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