Factors Influencing RBC Alloimmunization: Lessons Learned from Murine Models

Abstract

Red blood cell (RBC) alloimmunization may occur following transfusion or pregnancy/delivery. Although observational human studies have described the immunogenicity of RBC antigens and the clinical significance of RBC alloantibodies, studies of factors influencing RBC alloimmunization in humans are inherently limited by the large number of independent variables involved. This manuscript reviews data generated in murine models that utilize transgenic donor mice, which express RBC-specific model or authentic human blood group antigens. Transfusion of RBCs from such donors into nontransgenic but otherwise genetically identical recipient mice allows for the investigation of individual donor or recipient-specific variables that may impact RBC alloimmunization. Potential donor-related variables include methods of blood product collection, processing and storage, donor-specific characteristics, RBC antigen-specific factors, and others. Potential recipient-related variables include genetic factors (MHC/HLA type and polymorphisms of immunoregulatory genes), immune activation status, phenotype of regulatory immune cell subsets, immune cell functional characteristics, prior antigen exposures, and others. Although murine models are not perfect surrogates for human biology, these models generate phenomenological and mechanistic hypotheses of RBC alloimmunization and lay the groundwork for follow-up human studies. Long-term goals include improving transfusion safety and minimizing the morbidity/mortality associated with RBC alloimmunization.

Keywords: Alloimmunization; Murine models; RBC; Transfusion.

Fig. 1

Transgenic HOD RBCs on an FVB background were leukoreduced using a Pall neonatal leukoreduction filter, with the equivalent of 1 human ‘unit’ of RBCs transfused into C57BL/6 recipients. A Anti-HEL responses were measured in sera 2 weeks post-transfusion. B Nucleated cells were evaluated pre and post-filtration, using propridium iodide staining. C Platelets were evaluated pre and post-filtration, using CD41 staining (and trucount beads).

Fig. 2

Blood from transgenic HOD.FVB or HOD.B6 animals was leukoreduced and stored for 28–35 days. A, B The equivalent of 1 human ‘unit’ was transfused into C57BL/6 mice, with recipient anti-HOD Ig immune responses measured by flow cytometric cross-match 14 days post-transfusion. C, D Post-transfusion RBC survival and recovery studies were completed, using monoclonal antibodies against Fy3 to track the transfused HOD RBCs.

Fig. 3

Transgenic RBCs expressing the KEL2B antigen were transfused every 4 weeks (for a total of 3 transfusions) into Townes mice homozygous for Hgb SS, heterozygous for Hgb S (AS), or homozygous for Hgb A (AA). A Anti-KEL glycoprotein Igs were measured by flow cytometric crossmatch 28 days after the first transfusion, and B measured again 28 days after the 3rd and final transfusion.

Fig. 4

The equivalent of 1 human ‘unit’ of leukoreduced mHEL or HOD RBCs, or KEL2B or hGPA RBCs were transfused into wild-type recipients, in the presence or absence of recipient poly (I:C) pre-treatment. Alloantibodies were measured 2–4 weeks post-transfusion by HEL specific ELISA or by flow cytometric cross-match using transfused and wild-type RBCs as targets.