CD71⁺ Red Blood Cells Mediate Increased Erythrophagocytosis and Reduce Monocyte Number in a Monocyte Suspension Assay

Adverse reactions from donor-recipient sex-mismatched transfusions have been reported in the field of transfusion medicine. Immature CD71⁺ red blood cells (CD71⁺ RBCs) may influence these reactions through an immunomodulatory mechanism potentially linked to enriched intracellular reactive oxygen species (ROS). This study was conducted to investigate the effect of CD71⁺ RBCs on erythrophagocytosis, a common in vitro test for hemolysis risk in incompatible blood transfusions. Enriched CD71⁺ RBCs and CD71^- RBCs were exposed to allogenic peripheral blood mononuclear cells (PBMCs). CD71⁺ RBCs were also incubated with anti-D sensitised CD71^- RBCs and PBMCs, in varying ratios, to assess any dose-dependent behaviour. An ROS scavenger, apocynin, was applied to understand how CD71⁺ RBCs impact erythrophagocytosis. The phagocytosis index (RBC PI or CD71⁺ RBC PI) was measured using image flow cytometry. Supernatant haemoglobin was examined spectrophotometrically. The enriched CD71⁺ RBC group showed a significantly higher RBC PI and a reduction in the monocyte number compared to the CD71^- RBC group. Compared to the non-treated group, the reduction of monocyte number and the increase of the phagocytosis index of CD71⁺ RBCs was partially reversed in the apocynin-treated group. There were positive correlations between the dose of CD71⁺ RBC and CD71⁺ RBC PI and supernatant haemoglobin. Donated CD71⁺ RBCs enhance in vitro erythrophagocytosis of CD71⁺ RBCs in a dose-dependent manner and reduce monocyte number. This work contributes to our understanding of immature CD71⁺ RBCs' role in post-transfusion immunobiology.