In-depth phenotypic characterization of reticulocyte maturation using mass cytometry

Abstract

Progress towards an in-depth understanding of the final steps of the erythroid lineage development is paramount for many hematological diseases. We have characterized the final stages of reticulocyte maturation from bone marrow to peripheral blood using for the first time single-cell Mass Cytometry (CyTOF). We were able to measure the expression of 31 surface markers within a single red blood cell (RBC). We demonstrate the validity of CyTOF for RBC phenotyping by confirming the progressive reduction of transferrin receptor 1 (CD71) during reticulocyte maturation to mature RBC. We highlight the high-dimensional nature of mass cytometry data by correlating the expression of multiple proteins on individual RBCs. We further describe a more drastic reduction pattern for a component of the alpha4/beta1 integrin CD49d at the very early steps of reticulocyte maturation in bone marrow and directly linked with the mitochondria remnants clearance pattern. The enhanced and accurate RBC phenotyping potential of CyTOF described herein could be beneficial to decipher RBC preferences, as well as still not well understood receptor-ligand interaction of some hemotropic parasites such as the malaria causing agent Plasmodium vivax.

CD71 improves delineation of PNH type III, PNH type II, and normal immature RBCS in patients with paroxysmal nocturnal hemoglobinuria

“…In order to improve both quantitative and qualitative assessments of the erythroid component in PNH we have introduced CD71 (transferrin receptor) as a marker of immature RBCs (iRBC) and examined the utility of assessing the clone size within this fraction and whether a clearer delineation of Type II and Type III cells can be made. iRBCs are unaffected by complement‐mediated lysis (Morgan et al, ; Sadallah et al, ) or by transfusions of mature RBCs (Thomson‐Luque et al, ).…”

“…Earlier assays relied on nucleic acid stains to detect reticulocytes (Chin‐Yee, Keeney, & Lohmann, ; Ware, Rosse, & Hall, ), whereas more recently, antibodies targeting the transferrin receptor (CD71) have been employed (Naumova, Plekhanova, Lugovskaia, Pochtar, & Bugrov, ; Sutherland et al, ). The latter detects immature RBCs (nucleated RBCs and reticulocytes) (Thomson‐Luque et al, ) and does not require the use of nucleic acid stains.…”

Scald Injury-Induced T Cell Dysfunction Can Be Mitigated by Gr1+ Cell Depletion and Blockage of CD47/CD172a Signaling

“…Immature reticulocytes express the erythroid lineage marker CD235 (human)/Ter119 (murine), transferrin receptor (CD71), VLA-4 (CD49d) and have residual RNA. The expression/presence of the last three decreases with increasing maturation (44)(45)(46). Immature reticulocytes were first attributed immunosuppressive properties by hampering immune cell activation and cytokine production through arginase 2 in a study using newborn mice, expanding the old concept that neonatal infection susceptibility is solely based on an underdeveloped immune system (44).…”

Naturally acquired blocking human monoclonal antibodies to Plasmodium vivax reticulocyte binding protein 2b

“…TfR1 mediates the uptake of iron into cells in complex with its human ligand, iron-loaded Tf 6 . On the surface of reticulocytes, TfR1 is abundantly expressed and is shed during reticulocyte maturation into normocytes 7 . TfR1 is critical for P. vivax invasion, as cells expressing a mutant form of TfR1 are completely refractory to P. vivax invasion 2 .…”