Anti-αvβ3 antibodies in FNAIT are associated with disturbed angiogenesis and intracranial hemorrhage in men and mice

Abstract

In Caucasians, maternal allo-immunization against human platelet antigen (HPA)-1a is the most common cause for fetal and neonatal alloimmune thrombocytopenia (FNAIT). Intracranial hemorrhage (ICH) is a serious complication. The lack of a consistent correlation between antibody concentration, severity of thrombocytopenia, and ICH makes it difficult to predict thrombocytopenia in subsequent pregnancies. HPA-1a is located on integrin β3, which forms heterodimers with different alpha chains. The αvβ3 heterodimer is present on endothelial cells (ECs), and a specific antibody interaction with fetal ECs may explain the sporadic nature of ICH. Analysis of human sera was consistent with the presence of anti-HPA-1a of the anti-αvβ3 type in ICH positive cases. We show evidence that these sera induce VEGFR2 fixation to the cell surface, lead to VEGFR2 phosphorylation, enhance ROS production, and interfere with endothelial sprouting. Analysis of blinded human FNAIT samples demonstrates that combined biochemical and functional testing correctly identifies 6/6 ICH positive and 4/4 ICH negative cases. In a murine model, female b3 knockout mice develop anti-platelet antibodies after immunization with wild-type platelets. After mating with b3 wildtype males, only pups born to mice with anti-αvβ3 develop ICH. Matching our findings obtained with human material, only murine sera containing anti-αvβ3 induce VEGFR2 fixation on the cell surface, lead to VEGFR2 phosphorylation, induce ROS production, and hamper endothelial loop formation. In conclusion, we find evidence that ICH in FNAIT is based on vascular impairment and substantiate previous reports that impaired angiogenesis could represent a mechanism of ICH in FNAIT.