Fetal and neonatal alloimmune thrombocytopenia (FNAIT) occurs when a pregnant individual becomes alloimmunized against paternally-inherited fetal platelet antigens. Maternal-fetal incompatibility for HPA-1a is the most common. The pathophysiology involved in fetal platelet destruction is not fully understood. The objective of this study was to assess the ability of human anti-HPA-1a antibodies to mediate Fc gamma receptor-dependent platelet phagocytosis, complement fixation, platelet activation and desialylation to determine if the combination of these potential mechanisms could explain thrombocytopenia in FNAIT.

Ten sera (anti-HPA-1a positive and anti-HLA-antibody negative) were studied. An in vitro platelet phagocytosis assay was conducted. Briefly, donor platelets were fluorescently labelled, incubated with human serum, and added to macrophages for phagocytosis. Non-phagocytosed, surface-bound platelets were identified by immunofluorescence. Macrophages were mounted on to glass slides and photomicrographs were taken by spinning-disc confocal microscopy. Z-stacked images were 3D reconstructed for analysis using Imaris, and the phagocytic index was calculated. To determine the sera"s ability to fix complement, a complement fixation assay based on sheep red cell hemolysis was performed. Sera were first incubated with platelets, and rabbit-anti-sheep sensitized red cells were used and indicator cells in a second step. In vitro platelet activation, P-selectin expression, and desialylation, Ricinus Communis Agglutinin I (RCA I) binding, was determined by flow cytometry.

Nine of the 10 patients had neonates with moderate/severe thrombocytopenia with all of these sera inducing either platelet phagocytosis (8/9) and/or complement fixation (2/9) while the only patient with mild disease (platelet count of 64) induced neither. In addition, 2/8 of sera with severe FNAIT induced platelet desialylation (RCA-I binding) as well as platelet activation (CD62P) of healthy donor platelets suggesting a more complex pathophysiology in some FNAIT patients. Phagocytosis was mediated by FcγRI and FcγRIII and was dependent on the presence of IgG.

This study suggests phagocytosis and complement fixation as potentially contributing to thrombocytopenic severity in FNAIT with the presence of platelet activation and desialylation as potential additional contributors.

No conflicts of interest.