Fetal gene defects precipitate platelet-mediated pregnancy failure in factor V Leiden mothers

Abstract

We describe a mouse model of fetal loss in factor V Leiden (FvL) mothers in which fetal loss is triggered when the maternal prothrombotic state coincides with fetal gene defects that reduce activation of the protein C anticoagulant pathway within the placenta. Fetal loss is caused by disruption of placental morphogenesis at the stage of labyrinth layer formation and occurs in the absence of overt placental thrombosis, infarction, or perfusion defects. Platelet depletion or elimination of protease-activated receptor 4 (Par4) from the mother allows normal placentation and prevents fetal loss. These findings establish a cause-effect relationship for the observed epidemiologic association between maternal FvL status and fetal loss and identify fetal gene defects as risk modifiers of pregnancy failure in prothrombotic mothers. Pregnancy failure is mediated by Par4-dependent activation of maternal platelets at the fetomaternal interface and likely involves a pathogenic pathway independent of occlusive thrombosis. Our results further demonstrate that the interaction of two given thrombosis risk factors produces markedly disparate consequences on disease manifestation (i.e., thrombosis or pregnancy loss), depending on the vascular bed in which this interaction occurs.

Figure 1.

Phenotype of Thbd^ProPro embryos in FV^QQThbd^Pro+ mothers. (a, c, and e) Thbd^Pro+ littermates at E9.5, E12.5, and E18.5, respectively, of Thbd^ProPro embryos (b, d, and f). At E9.5, some Thbd^ProPro embryos are smaller (b). By E11.5, most Thbd^ProPro embryos are in an advanced stage of decay (d). Platelet depletion of the mother with anti-GP1bα antibody treatment (e and f) results in normal-appearing Thbd^ProPro embryos. Pictures were taken at different magnifications. Bars, 1 mm.

Figure 2.

Placental size of Thbd^ProPro embryos relative to controls. Each symbol represents a measurement from a single fetoplacental unit. All litters were analyzed at E9.5. Mean ratios and SDs are shown.

Figure 3.

Placental phenotype of Thbd^ProPro embryos and littermate controls in Fv^QQThbd^Pro+ mothers. Histological sections through placentas of Thbd^Pro+ (a, c, and e) and a phenotypically normal Thbd^ProPro (b, d, and f) embryo at E9.5. a and b show cytokeratin-expressing trophoblast cells (brown staining). The placenta of Thbd^ProPro embryos (stained area) is smaller than that of Thbd^Pro+ embryos. c and d are higher magnification images of hematoxylin and eosin–stained sections adjacent to a and b, respectively. e and f correspond to boxes in c and d, respectively. Dashed lines in c and d mark the border of the placenta and decidua, as determined by cytokeratin staining. Thbd^Pro+ placentas (c and e) have formed a labyrinth characterized by maternal (bright red enucleated maternal red blood cells; dashed arrows) and fetal (purple hematoxylin-stained nucleated fetal red blood cells; continuous arrow) blood spaces. The labyrinth is distinctly absent in the placentas of littermate Thbd^ProPro embryos (d and f). Bars, 0.1 mm.

Figure 4.

Placental thrombosis in FvL mice. The number of placental thrombi was similar in placentas of wild-type and Thbd^ProPro embryos (see Results). Thrombi were identified on serial sections by hematoxylin and eosin staining (a, c, and e), and immunostaining with antifibrinogen antibodies (b) and anti–p-selectin antibodies (d and f). e and f are enlarged views of the boxed areas in c and d. Arrows indicate clots. Bars, 0.1 mm.

Figure 5.

Assessment of decidual and placental vascular patency by FITC-dextran infusion. Whole-mount (a and b) and sections (c–f) of decidual-placental units corresponding to a wild-type embryo in a wild-type mother (a, c, and e) and a Fv^Q+Thbd^ProPro embryo in a Fv^QQThbd^Pro+ mother (b, d, and f) are shown. From the fetal aspect of the placenta, a characteristic ring of maternal placental vasculature, marked by FITC-dextran, is observed in wild-type pregnancies but not in placenta of a Fv^Q+Thbd^ProPro embryo in a Fv^QQThbd^Pro+ mother (a and b). Decidual and placental regions in both types of pregnancies are perfused with FITC-dextran (c and d). Hematoxylin and eosin–stained sections adjacent to c and d are shown in e and f. Arrows show maternal blood spaces in the placenta. Bars, 1 mm.