Annika K. Hult

RIFINs are adhesins implicated in severe Plasmodium falciparum malaria

Rosetting is a virulent Plasmodium falciparum phenomenon associated with severe malaria. Here we demonstrate that P. falciparum–encoded repetitive interspersed families of polypeptides (RIFINs) are expressed on the surface of infected red blood cells (iRBCs), where they bind to RBCs—preferentially of blood group A—to form large rosettes and mediate microvascular binding of iRBCs. We suggest that RIFINs have a fundamental role in the development of severe malaria and thereby contribute to the varying global distribution of ABO blood groups in the human population.

Forssman expression on human erythrocytes: biochemical and genetic evidence of a new histo-blood group system

In analogy with histo-blood group A antigen, Forssman (Fs) antigen terminates with α3-N-acetylgalactosamine and can be used by pathogens as a host receptor in many mammals. However, primates including humans lack Fs synthase activity and have naturally occurring Fs antibodies in plasma. We investigated individuals with the enigmatic ABO subgroup A(pae) and found them to be homozygous for common O alleles. Their erythrocytes had no A antigens but instead expressed Fs glycolipids. The unexpected Fs antigen was confirmed in structural, serologic, and flow-cytometric studies. The Fs synthase gene, GBGT1, in A(pae) individuals encoded an arginine to glutamine change at residue 296. Gln296 is present in lower mammals, whereas Arg296 was found in 6 other primates, > 250 blood donors and A(pae) family relatives without the A(pae) phenotype. Transfection experiments and molecular modeling showed that Agr296Gln reactivates the human Fs synthase. Uropathogenic E coli containing prsG-adhesin-encoding plasmids agglutinated A(pae) but not group O cells, suggesting biologic implications. Predictive tests for intravascular hemolysis with crossmatch-incompatible sera indicated complement-mediated destruction of Fs-positive erythrocytes. Taken together, we provide the first conclusive description of Fs expression in normal human hematopoietic tissue and the basis of a new histo-blood group system in man, FORS.

ABO Blood Groups Influence Macrophage-mediated Phagocytosis of Plasmodium falciparum-infected Erythrocytes

Erythrocyte polymorphisms associated with a survival advantage to Plasmodium falciparum infection have undergone positive selection. There is a predominance of blood group O in malaria-endemic regions, and several lines of evidence suggest that ABO blood groups may influence the outcome of P. falciparum infection. Based on the hypothesis that enhanced innate clearance of infected polymorphic erythrocytes is associated with protection from severe malaria, we investigated whether P. falciparum-infected O erythrocytes are more efficiently cleared by macrophages than infected A and B erythrocytes. We show that human macrophages in vitro and mouse monocytes in vivo phagocytose P. falciparum-infected O erythrocytes more avidly than infected A and B erythrocytes and that uptake is associated with increased hemichrome deposition and high molecular weight band 3 aggregates in infected O erythrocytes. Using infected A1, A2, and O erythrocytes, we demonstrate an inverse association of phagocytic capacity with the amount of A antigen on the surface of infected erythrocytes. Finally, we report that enzymatic conversion of B erythrocytes to type as O before infection significantly enhances their uptake by macrophages to observed level comparable to that with infected O wild-type erythrocytes. These data provide the first evidence that ABO blood group antigens influence macrophage clearance of P. falciparum-infected erythrocytes and suggest an additional mechanism by which blood group O may confer resistance to severe malaria.

Many genetically defined ABO subgroups exhibit characteristic flow cytometric patterns.

BACKGROUND: A flow cytometric method for detection of low levels of A/B antigen had been developed previously in our laboratory. The aim of this study was to investigate if this approach could be utilized to characterize different ABO subgroups and constitute a useful tool in a reference laboratory.

Do ABO Blood Group Antigens Hamper the Therapeutic Efficacy of Mesenchymal Stromal Cells

Investigation into predictors for treatment outcome is essential to improve the clinical efficacy of therapeutic multipotent mesenchymal stromal cells (MSCs). We therefore studied the possible harmful impact of immunogenic ABO blood groups antigens – genetically governed antigenic determinants – at all given steps of MSC-therapy, from cell isolation and preparation for clinical use, to final recipient outcome. We found that clinical MSCs do not inherently express or upregulate ABO blood group antigens after inflammatory challenge or in vitro differentiation. Although antigen adsorption from standard culture supplements was minimal, MSCs adsorbed small quantities of ABO antigen from fresh human AB plasma (ABP), dependent on antigen concentration and adsorption time. Compared to cells washed in non-immunogenic human serum albumin (HSA), MSCs washed with ABP elicited stronger blood responses after exposure to blood from healthy O donors in vitro, containing high titers of ABO antibodies. Clinical evaluation of hematopoietic stem cell transplant (HSCT) recipients found only very low titers of anti-A/B agglutination in these strongly immunocompromised patients at the time of MSC treatment. Patient analysis revealed a trend for lower clinical response in blood group O recipients treated with ABP-exposed MSC products, but not with HSA-exposed products. We conclude, that clinical grade MSCs are ABO-neutral, but the ABP used for washing and infusion of MSCs can contaminate the cells with immunogenic ABO substance and should therefore be substituted by non-immunogenic HSA, particularly when cells are given to immunocompentent individuals.

Blood group genotype analysis for the quality improvement of reagent test red blood cells.

Background and Objectives  Reagent red blood cells (RBCs) for antibody detection should express certain important antigens as a double dose, that is, the donors must be homozygous for the corresponding alleles. Traditionally, dose is determined by serological typing and known allele frequencies. However, RHD zygosity cannot be predicted serologically owing to the absence of an antithetical antigen, and FY zygosity is confounded by two variant haplotypes, FY*0 and FY*X. Furthermore, lack of reagents hampers our ability to type for some clinically important antigen pairs such as Doa/Dob.

Weak A phenotypes associated with novel ABO alleles carrying the A2‐related 1061C deletion and various missense substitutions

BACKGROUND: The 1061delC single‐nucleotide polymorphism (SNP) has been reported mostly in the context of the common A2[A201] allele and typically produces an A2 phenotype. This study evaluated new Aweak alleles, each containing 1061delC.

Flow cytometry evaluation of red blood cells mimicking naturally occurring ABO subgroups after modification with variable amounts of function-spacer-lipid A and B constructs.

BACKGROUND: Kodecytes bearing synthetic blood group A and B antigens are increasingly being used in transfusion laboratories as serologic mimics of red blood cell (RBC) Aweak and Bweak subtypes. The aim of this study was to compare the flow cytometry profile of kodecytes with native ABO subgroups.

Identification of the molecular and genetic basis of PX2, a glycosphingolipid blood group antigen lacking on globoside-deficient erythrocytes

Background: Expression of x2 glycosphingolipid (PX2) is elevated on erythrocytes from individuals with the rare P/P1/Pk-negative p phenotype. Results: Globoside-deficient individuals with mutated P synthase (β1,3GalNAc-T1) lack PX2 and have anti-PX2 in plasma. Transfection of B3GALNT1 induces P and PX2 expression. Conclusion: PX2 synthesized by β1,3GalNAc-T1 fulfills blood group criteria. Significance: β1,3GalNAc-T1 uses different acceptors to form immunologically distinct glycosphingolipids. The x2 glycosphingolipid is expressed on erythrocytes from individuals of all common blood group phenotypes and elevated on cells of the rare P/P1/Pk-negative p blood group phenotype. Globoside or P antigen is synthesized by UDP-N-acetylgalactosamine:globotriaosyl-ceramide 3-β-N-acetylgalactosaminyltransferase encoded by B3GALNT1. It is the most abundant non-acid glycosphingolipid on erythrocytes and displays the same terminal disaccharide, GalNAcβ3Gal, as x2. We encountered a patient with mutations in B3GALNT1 causing the rare P-deficient P1k phenotype and whose pretransfusion plasma was unexpectedly incompatible with p erythrocytes. The same phenomenon was also noted in seven other unrelated P-deficient individuals. Thin-layer chromatography, mass spectrometry, and flow cytometry were used to show that the naturally occurring antibodies made by p individuals recognize x2 and sialylated forms of x2, whereas x2 is lacking on P-deficient erythrocytes. Overexpression of B3GALNT1 resulted in synthesis of both P and x2. Knockdown experiments with siRNA against B3GALNT1 diminished x2 levels. We conclude that x2 fulfills blood group criteria and is synthesized by UDP-N-acetylgalactosamine: globotriaosylceramide 3-β-N-acetylgalactosaminyltransferase. Based on this linkage, we proposed that x2 joins P in the GLOB blood group system (ISBT 028) and is renamed PX2 (GLOB2). Thus, in the absence of a functional P synthase, neither P nor PX2 are formed. As a consequence, naturally occurring anti-P and anti-PX2 can be made. Until the clinical significance of anti-PX2 is known, we also recommend that rare P1k or P2k erythrocyte units are preferentially selected for transfusion to Pk patients because p erythrocytes may pose a risk for hemolytic transfusion reactions due to their elevated PX2 levels.

A and B antigen levels acquired by group O donor‐derived erythrocytes following ABO‐non‐identical transfusion or minor ABO‐incompatible haematopoietic stem cell transplantation

ABO‐incompatible haematopoietic stem cell transplantation (HSCT) presents a challenge to blood component transfusion. The aim of this study was to investigate the weak blood group A or B antigen expression by donor‐derived group O red blood cells (RBC) observed following transfusion or minor ABO‐incompatible HSCT. In addition, in vitro experiments were performed to elucidate possible mechanisms underlying this phenomenon.