Antigen Density Dictates Immune Responsiveness following Red Blood Cell Transfusion

Abstract

Although RBC transfusion can result in the development of anti-RBC alloantibodies that increase the probability of life-threatening hemolytic transfusion reactions, not all patients generate anti-RBC alloantibodies. However, the factors that regulate immune responsiveness to RBC transfusion remain incompletely understood. One variable that may influence alloantibody formation is RBC alloantigen density. RBC alloantigens exist at different densities on the RBC surface and likewise exhibit distinct propensities to induce RBC alloantibody formation. However, although distinct alloantigens reside on the RBC surface at different levels, most alloantigens also represent completely different structures, making it difficult to separate the potential impact of differences in Ag density from other alloantigen features that may also influence RBC alloimmunization. To address this, we generated RBCs that stably express the same Ag at different levels. Although exposure to RBCs with higher Ag levels induces a robust Ab response, RBCs bearing low Ag levels fail to induce RBC alloantibodies. However, exposure to low Ag–density RBCs is not without consequence, because recipients subsequently develop Ag-specific tolerance. Low Ag–density RBC–induced tolerance protects higher Ag–density RBCs from immune-mediated clearance, is Ag specific, and occurs through the induction of B cell unresponsiveness. These results demonstrate that Ag density can potently impact immune outcomes following RBC transfusion and suggest that RBCs with altered Ag levels may provide a unique tool to induce Ag-specific tolerance.

Hemoglobin A clearance in children with sickle cell anemia on chronic transfusion therapy

“…[31][32][33] Further, we demonstrated that DHbA does directly impact CTT outcomes, as increased HbA clearance leads to lower pretransfusion Hb, higher HbS, and higher reticulocytes. Although the factors that dictate RBC alloimmunization continue to be an area of active investigation, [34][35][36][37][38][39] as patients with RBC antibodies have increased HbA clearance, it is possible that increased removal of transfused RBCs may reflect increased RBC uptake by immune cells that are uniquely poised to facilitate RBC alloimmunization. 40 Conversely, in a murine model of alloimmunization, donor RBCs that were subjected to pretransfusion oxidant stress to cause rapid posttransfusion clearance by the reticuloendothelial system were significantly more immunogenic than donor RBCs with longer posttransfusion survival, suggesting that clearance rate by the reticuloendothelial system influences alloimmunization respones.…”

Antibody‐mediated immunosuppression can result from RBC antigen loss independent of Fcγ receptors in mice

“…While proteolytic cleavage allowed HEL to be selectively removed in vitro , many proteins are sensitive to protease, which could impact RBC circulatory half‐life, interactions with immune cells or other RBC changes that may influence the overall immune response to HOD, precluding the use of this approach to examine the potential impact of HEL antigen levels on anti‐HOD formation. HOD founders that express different levels of HOD, as recently described for KEL, will ultimately be needed to begin defining potential antigen threshold requirements for a productive immune response against the HOD antigen.…”

“…For detection of Ter‐119, RBCs were stained with Ter‐119 (BD Biosciences), as done previously . To assess anti‐HOD immunoglobulin M (IgM) and IgG antibody development, sera were flow cross‐matched, as previously described . All antibodies were used at 1:100 in fluorescence activated cell sorting buffer (2% bovine serum albumin in phosphate‐buffered saline) unless otherwise noted, as done previously …”

Red blood cell alloimmunisation: induction of immunity and potential mitigation strategies

“…A series of transgenic mice expressing variations of the human KEL glycoprotein have been generated over the past 5 years, with some expressing very low copy number (KEL lo ) and some expressing a more moderate copy number (KEL). Recipients transfused with KEL lo RBCs fail to mount a detectable humoral immune response , whereas those transfused with RBCs expressing a moderate copy number of KEL generate anti‐KEL alloantibodies after a single transfusion . Further, animals transfused with transgenic RBCs expressing very high copy numbers fail to mount a detectable response, unless treated with an adjuvant around the time of RBC exposure.…”

“…However, these animals remain non‐alloimmunised upon subsequent exposure to hGPA RBCs in the presence of an adjuvant (which are otherwise immunogenic in naïve animals). This non‐responsiveness is antigen specific, with animals tolerised to hGPA or other RBCs capable of responding to RBCs expressing 3rd party antigens .…”