Analysis of 24-h recovery of transfused stored RBCs in recipient mice of distinct genetic backgrounds.

BACKGROUND AND OBJECTIVES: Human studies have demonstrated substantial donor-to-donor variation in refrigerated RBC storage with respect to several variables, including 24-h post-transfusion RBC recovery. However, the human studies leading to these observations are mostly performed using autologous transfusions of stored RBCs, thereby avoiding issues of infectious disease transmission and alloimmunization. Accordingly, one cannot distinguish whether variability in 24-h RBC recovery is due to alterations in RBC storage, differences in phagocytic activity of the recipient's reticuloendothelial system or both. Similar to humans, genetically distinct inbred mouse strains have substantial differences in RBC storage biology, including 24-h post-transfusion RBC recovery. MATERIALS AND METHODS: In this report, we juxtaposed 24-h recoveries in 15 distinct inbred strains of mice, holding the RBC donor constant to isolate transfusion recipient variation as an independent variable. Strains were chosen for differences in baseline reticulocyte count and haemoglobin, which may correlate to RBC life span and turnover. RESULTS: Unlike large differences observed in storage of RBCs obtained from different strains of mice, only subtle strain-to-strain differences were observed regarding 24-h post-transfusion RBC recoveries. CONCLUSIONS: These findings indicate that the murine strains examined are not likely to be useful in sorting out mechanisms of clearance of stored RBCs, and suggest that such mechanisms may be generally conserved in the strains of mice analysed.

Glucose-6-phosphate dehydrogenase deficiency in transfusion medicine: the unknown risks.

The hallmark of glucose-6-phosphate dehydrogenase (G6PD) deficiency is red blood cell (RBC) destruction in response to oxidative stress. Patients requiring RBC transfusions may simultaneously receive oxidative medications or have concurrent infections, both of which can induce haemolysis in G6PD-deficient RBCs. Although it is not routine practice to screen healthy blood donors for G6PD deficiency, case reports identified transfusion of G6PD-deficient RBCs as causing haemolysis and other adverse events. In addition, some patient populations may be more at risk for complications associated with transfusions of G6PD-deficient RBCs because they receive RBCs from donors who are more likely to have G6PD deficiency. This review discusses G6PD deficiency, its importance in transfusion medicine, changes in the RBC antioxidant system (of which G6PD is essential) during refrigerated storage and mechanisms of haemolysis. In addition, as yet unanswered questions that could be addressed by translational and clinical studies are identified and discussed.

Effects of genetic, epigenetic, and environmental factors on alloimmunization to transfused antigens: Current paradigms and future considerations.

Transfused red blood cells, platelets, or coagulation factors have the capacity to induce alloantibodies, which once formed, can be a clinical barrier to future transfusion therapy and/or transplantation. Large observational studies over the last 50 years have characterized some of the general properties of transfusion induced alloimmunization, which appear to vary to a considerable extent from what is generally observed for human responses to other immunogens, such as microbial pathogens and vaccines. Transfused cells and factor only induce immune responses in the minority of recipients. There are data to suggest that differences in the unit may play a role. However, there are clearly differences in recipient biology, as once a recipient makes one antibody they are much more likely to make additional antibodies; indeed, recipients have been categorized as "responder" and "non-responder" by the field. Recent mechanistic studies have begun to define potential causes for such differences in alloimmunization from patient to patient, but much progress needs to be made to understand how, why, and in whom alloimmunization occurs. This review gives a general background on immunology in the context of transfusion, summarizes recent progress in the field, and discusses future directions for exploration. Particular attention is paid to the general concept that the human immune system is melded by the wide range of antigens encountered in our environment, and that the effects of such on the immune system may have a profound effect upon response to transfused cells.

Mechanisms of alloimmunization and subsequent bone marrow transplantation rejection induced by platelet transfusion in a murine model.

For many nonmalignant hematological disorders, HLA-matched bone marrow transplantation (BMT) is curative. However, due to lack of neoplasia, the toxicity of stringent conditioning regimens is difficult to justify, and reduced intensity conditioning is used. Unfortunately, current reduced intensity regimens have high rates of BMT rejection. We have recently reported in a murine model that mHAs on transfused platelet products induce subsequent BMT rejection. Most nonmalignant hematological disorders require transfusion support prior to BMT and the rate of BMT rejection in humans correlates with the number of transfusions given. Herein, we perform a mechanistic analysis of platelet transfusion-induced BMT rejection and report that unlike exposure to alloantigens during transplantation, platelet transfusion primes alloimmunity but does not stimulate full effector function. Subsequent BMT is itself an additional and distinct immunizing event, which does not induce rejection without antecedent priming from transfusion. Both CD4(+) and CD8(+) T cells are required for priming during platelet transfusion, but only CD8(+) T cells are required for BMT rejection. In neither case are antibodies required for rejection to occur.

Minor antigens on transfused RBCs crossprime CD8 T cells but do not induce full effector function.

HLA-matched bone marrow transplantation (BMT) is a cure for nonmalignant hematological disorders; however, rejection rates are high and correlate with the number of antecedent transfusions. Recently, using murine models, we reported that minor antigens (mHAs) in transfused leukoreduced red blood cell (RBC) or platelet units induce rejection of subsequent BMT. To study RBCs as an immunogen, we utilized transgenic donors that express a model mHA selectively on RBCs (HOD mouse). Transfusion of HOD blood did not induce BMT rejection of marrow that shared mHAs with the HOD RBCs. Similarly, no endogenous anti-HOD CD8(+) T-cell response was detected with antigen-specific tetramer reagents. Adoptively transferred OT-I T cells rapidly expanded after HOD blood transfusion; however, only a semi-effector phenotype was observed (tumor necrosis factor-α and interferon-γ secretion, but essentially no Granzyme B). After initial expansion, OT-I T cells contracted rapidly to very low levels. A similar trend was observed by in vivo CTL assay, with only transient lytic activity. Together, these data indicate that RBCs may not be the component of RBC units that induces BMT rejection, and suggest that contaminating platelets or leukocytes may be responsible.

MHC II on transfused murine blood is not required for alloimmunization against MHC I.

BACKGROUND AND OBJECTIVES: Transfusion of allogeneic platelet products can result in antibodies against donor major histocompatibility complex (MHC) I antigens, leading to a refractory state to subsequent platelet transfusions. However, there is disagreement in the field regarding the molecular mechanisms of humoral alloimmunization. One hypothesis states that donor MHC II is a requirement for alloimmunization. However, other studies have suggested that donor MHC I is alone sufficient and MHC II is not required. MATERIALS AND METHODS: We utilized a mouse model of anti-MHC I alloimmunization to transfused blood, which employed donors with a complete deletion of all MHC II genes. BALB/c (H-2(d)) recipients were transfused with blood from either C57BL/6 (H-2(b)) or MHC II null donors on a C57BL/6 background. Anti-MHC I alloimmunization was monitored by indirect immunofluorescence. RESULTS: Recipients of either wild type or MHC II null blood produced equivalent humoral responses against donor MHC I antigens. However, there was variation in the relative amounts of IgG subclasses. CONCLUSION: These data reject the hypothesis that donor MHC II expression is required for alloimmunization to MHC I antigens.

Use of mouse models to study the mechanisms and consequences of RBC clearance.

Mice provide tractable animal models for studying the pathophysiology of various human disorders. This review discusses the use of mouse models for understanding red-blood-cell (RBC) clearance. These models provide important insights into the pathophysiology of various clinically relevant entities, such as autoimmune haemolytic anaemia, haemolytic transfusion reactions, other complications of RBC transfusions and immunomodulation by Rh immune globulin therapy. Mouse models of both antibody- and non-antibody-mediated RBC clearance are reviewed. Approaches for exploring unanswered questions in transfusion medicine using these models are also discussed.

Cross-linking induces non-haemolytic antigen-loss from transfused red blood cells: a potential role for rheumatoid factor.

Transfusion of cross-match incompatible blood can lead to haemolysis. However, in some cases, incompatible transfused red blood cells are bound by antibody and then converted to being negative for both the incompatible antigen and the direct antiglobulin test. Using a murine model of this phenomenon, we have recently reported that antibodies binding to multiple epitopes are required. Herein, we report that antibodies against one epitope can induce antigen-loss if an anti-immunoglobulin G (IgG) antibody is also present. These findings support a model of cross-linking being required, and raise the possibility that naturally occurring anti-IgG, such as rheumatoid factor, may contribute to antigen-loss.

CD40 ligation induced phenotypic and functional expression of CD80 by human cardiac microvascular endothelial cells.

BACKGROUND: CD40/CD40L (gp39) interactions are known to play a central role in the function of the immune system (1). CD40 is constitutively expressed on professional antigen presenting cells, such as macrophages and dendritic cells, as well as at low levels on other cell lineages, including human umbilical vein endothelial cells (HUVECs). On antigen-presenting cells, ligation of CD40 causes expression of the costimulatory molecule CD80 (B7-1). Similar ligation of CD40 on HUVECs, however, leads to up-regulation of the adhesion molecules VCAM-1, ICAM-1, and E-selectin, but not CD80. METHODS: In efforts to provide evidence that microvascular endothelial cells (MECs) are distinct from HUVECs and that the distinguishing features play a role in allograft rejection, MEC cultures were prepared from the explanted hearts of human heart transplant recipients and primary cell lines were established. These MECs were induced to express higher levels of CD40 with interferon-gamma pretreatment, co-cultured with CD40L-transfected HeLa cells, and fluorescence-activated cell sorter-assisted phenotypic studies, in addition to functional allogeneic mixed lymphocyte reaction and accessory-cell dependent mitogen induced proliferation assays were performed. RESULTS: CD40-CD40L interactions induced the expression of the adhesion molecules VCAM-1 and E-selectin and the costimulatory molecule CD80 but not CD86 (B7-2) on the MECs. The expressed CD80 proved functional in both allo-MLR assays and accessory-cell dependent mitogen proliferation assays. CONCLUSIONS: MECs are distinct from HUVECs by their potential to express VCAM-1 after interferon-gamma pretreatment and CD80 after CD40 ligation, properties which enable this cell lineage to play a central role in initiating and maintaining allograft rejection in human cardiac transplants.

Human herpesvirus 8 encodes an interferon regulatory factor (IRF) homolog that represses IRF-1-mediated transcription.

Human herpesvirus 8 (HHV-8) is the probable viral etiologic agent for Kaposi's sarcoma. The HHV-8 genome encodes viral interferon regulatory factor (vIRF), a gene product that has homology to the IRF family of transcription factors. We demonstrate that vIRF inhibits responses to type I and type II interferons and blocks IRF-1-mediated transcription. vIRF does not compete with IRF-1 for binding to DNA or complex directly with IRF-1. The ability of vIRF to block IRF-1-mediated transcription is independent of the DNA binding domains of both vIRF and IRF-1. These data suggest that vIRF may contribute to viral pathogenesis and cellular transformation by interfering with interferon- and IRF-1-mediated gene expression through a novel mechanism.