The Development and Consequences of Red Blood Cell Alloimmunization.

While red blood cell (RBC) transfusion is the most common medical intervention in hospitalized patients, as with any therapeutic, it is not without risk. Allogeneic RBC exposure can result in recipient alloimmunization, which can limit the availability of compatible RBCs for future transfusions and increase the risk of transfusion complications. Despite these challenges and the discovery of RBC alloantigens more than a century ago, relatively little has historically been known regarding the immune factors that regulate RBC alloantibody formation. Through recent epidemiological approaches, in vitro-based translational studies, and newly developed preclinical models, the processes that govern RBC alloimmunization have emerged as more complex and intriguing than previously appreciated. Although common alloimmunization mechanisms exist, distinct immune pathways can be engaged, depending on the target alloantigen involved. Despite this complexity, key themes are beginning to emerge that may provide promising approaches to not only actively prevent but also possibly alleviate the most severe complications of RBC alloimmunization.

Polymorphonuclear neutrophil activation by Src phosphorylation contributes to HLA-A2 antibody-induced transfusion-related acute lung injury.

Human leukocyte antigen (HLA)-A2 antibody contributes to the pathogenesis of transfusion-related acute lung injury (TRALI) via polymorphonuclear neutrophil (PMN) activation, but the signaling pathways involved this process remain largely undefined. In this study, we sought to study the signaling pathways involved in the pathogenesis of HLA-A2-induced TRALI. Lipopolysaccharide (LPS), and the plasma from the HLA-A2 antibody-positive donors were utilized to establish a rat model of TRALI. Human pulmonary endothelial cells (HPMECs) were in vitro co-cultured with HLA-A2 antibody-treated PMNs and then treated with LPS to induce a cytotoxicity model. The effects of HLA-A2 antibody on HPMEC injury were evaluated in this model. Besides, dasatinib was used to block the Src phosphorylation to explore whether Src involved in the TRALI or HPMEC injury induced by HLA-A2 antibody. The HLA-A2 antibody plus LPS induced TRALI and stimulated PMN activation in rats. HLA-A2 antibody-induced TRALI could be attenuated via depletion of PMN. HLA-A2 antibody activated NF-κB and NLRP3 inflammasome. In addition, HLA-A2 antibody aggravated the HPMEC injuries and the release of PMN surfaces makers, but dasatinib treatment reversed this effect, indicating that HLA-A2 antibody activated PMNs and exacerbated TRALI by stimulating phosphorylation of Src followed by activation of NF-κB and NLRP3 inflammasome, which was validated in vivo. In summary, HLA-A2 induced PMNs by activating NF-κB/NLRP3 inflammasome via phosphorylated-Src elevation, thereby exacerbating TRALI. This study highlights promising target for the treatment of antibody-mediated TRALI.

Do We Store Packed Red Blood Cells under "Quasi-Diabetic" Conditions?

Red blood cell (RBC) transfusion is one of the most common therapeutic procedures in modern medicine. Although frequently lifesaving, it often has deleterious side effects. RBC quality is one of the critical factors for transfusion efficacy and safety. The role of various factors in the cells' ability to maintain their functionality during storage is widely discussed in professional literature. Thus, the extra- and intracellular factors inducing an accelerated RBC aging need to be identified and therapeutically modified. Despite the extensively studied in vivo effect of chronic hyperglycemia on RBC hemodynamic and metabolic properties, as well as on their lifespan, only limited attention has been directed at the high sugar concentration in RBCs storage media, a possible cause of damage to red blood cells. This mini-review aims to compare the biophysical and biochemical changes observed in the red blood cells during cold storage and in patients with non-insulin-dependent diabetes mellitus (NIDDM). Given the well-described corresponding RBC alterations in NIDDM and during cold storage, we may regard the stored (especially long-stored) RBCs as "quasi-diabetic". Keeping in mind that these RBC modifications may be crucial for the initial steps of microvascular pathogenesis, suitable preventive care for the transfused patients should be considered. We hope that our hypothesis will stimulate targeted experimental research to establish a relationship between a high sugar concentration in a storage medium and a deterioration in cells' functional properties during storage.

NLRP3 inflammasome of renal tubular epithelial cells induces kidney injury in acute hemolytic transfusion reactions.

BACKGROUND: Blood transfusion, a common basic supporting therapy, can lead to acute hemolytic transfusion reaction (AHTR). AHTR poses a great risk to patients through kidney function damage in a short time. Previous reports found that heme from destroyed red blood cells impaired kidney function, and NLR family pyrin domain containing 3 (NLRP3) inflammasome was augmented in case of kidney injury. However, the detailed mechanism regarding whether NLRP3 inflammasome is involved in kidney function injury in AHTR is not fully understood yet. METHODS: Hemolysis models were established by vein injection with human blood plasma or mouse heme from destroyed red blood cells. The injured renal tubular epithelial cells (RTECs) were evaluated by tubular damage markers staining in hemolysis models and in primary RTECs in vitro. The activation of NLRP3 inflammasome in RTECs by hemes was investigated by Western blot, ELISA, scanning electron microscopy, immunofluorescent staining, flow cytometry, and hemolysis models. NLRP3 gene knockout mice were employed to confirm these observations in vitro and in vivo. The binding between a novel inhibitor (66PR) and NLRP3 was affirmed by molecule docking and co-immunoprecipitation. The rescue of 66PR on kidney function impairment was explored in murine hemolysis models. RESULTS: We found that heme could activate NLRP3 inflammasome in RTECs to induce kidney function injury. NLRP3 gene knockout could prevent the damage of RTECs caused by hemes and recover kidney function in AHTR. Moreover, NLRP3 inflammasome chemical inhibitor, 66PR, could bind to NLRP3 protein and inhibit inflammasome activation in RTECs, which consequently relieved the injury of RTECs caused by hemes, and alleviated kidney function damage in the AHTR model. CONCLUSIONS: Hemes could activate NLRP3 inflammasome in RTECs, and a novel NLRP3 inflammasome inhibitor named 66PR relieved kidney function damage in AHTR. Our findings provided a new possible strategy to treat kidney function failure in AHTR.

Characterization of glycoprotein IIb/IIIa-specific alloantibodies induced by cross-strain platelet immunization in mice.

BACKGROUND: We previously described a mouse model in which platelet immunization between selected strains leads to production of alloantibodies and severe autoimmune thrombocytopenia and mimics the human condition posttransfusion purpura (PTP). This report describes studies defining epitopes recognized by these alloantibodies. STUDY DESIGN: Hybridomas were produced from spleen cells of immunized mice. Glycoprotein (GP) targets of resulting monoclonal antibodies were characterized by immunoprecipitation using platelets from the immunizing strains. Antigens defined by single amino acid (AA) polymorphisms recognized by monoclonal antibodies were identified by mutagenizing target glycoproteins expressed in Chinese hamster ovary cells and observing the effects on antibody binding. RESULTS: Three monoclonal antibodies (417.1, 417.3, 425.1) were produced that recognized GPIIb on immunizing platelets. Monoclonal antibodies 417.1 and 417.3 both required G111 and 425.1 required V37, located on the beta propeller domain of GPIIb, for binding to platelets from the immunizing strains C57 and PWK, respectively. Injection of 417.3 and 425.1 into mice caused platelet destruction only in mice with GPIIb containing the targeted AAs. CONCLUSIONS: Findings made provide evidence that alloantibodies produced by mice experiencing thrombocytopenia in a mouse model of PTP are specific for single AA polymorphisms that differ in GPIIb/IIIa integrin of the immunizing and immunized strains and therefore closely resemble the potent alloantibodies found in patients with PTP. The observations show that naturally occurring single AA differences in GPIIb/IIIa integrin of various mouse strains are highly immunogenic in the mouse strains studied and readily induce antibodies comparable to human platelet antigen-specific antibodies found in transfused and pregnant humans.

The Role of Circulating Cell-Free Hemoglobin in Sepsis-Associated Acute Kidney Injury.

Sepsis is a heterogeneous clinical syndrome that is complicated commonly by acute kidney injury (sepsis-AKI). Currently, no approved pharmacologic therapies exist to either prevent sepsis-AKI or to treat sepsis-AKI once it occurs. A growing body of evidence supports a connection between red blood cell biology and sepsis-AKI. Increased levels of circulating cell-free hemoglobin (CFH) released from red blood cells during hemolysis are common during sepsis and can contribute to sepsis-AKI through several mechanisms including tubular obstruction, nitric oxide depletion, oxidative injury, and proinflammatory signaling. A number of potential pharmacologic therapies targeting CFH in sepsis have been identified including haptoglobin, hemopexin, and acetaminophen, and early phase clinical trials have suggested that acetaminophen may have beneficial effects on lipid peroxidation and kidney function in patients with sepsis. Bedside measurement of CFH levels may facilitate predictive enrichment for future clinical trials of CFH-targeted therapeutics. However, rapid and reliable bedside tests for plasma CFH will be required for such trials to move forward.

ABO-incompatible platelets are associated with increased transfusion reaction rates.

BACKGROUND: ABO compatibility can affect platelet transfusion safety and efficacy, and ABO-incompatible (ABOi) platelets likely increases the risks of transfusion reactions though the magnitude of this risk is unclear. STUDY DESIGN AND METHODS: Data collected on all platelet transfusions administered over 36+ months were classified based on patient and product ABO blood group type and merged with a data set that included all transfusion reactions reported during that period. The transfusion reaction rates among various subsets was calculated. RESULTS: In patients greater than 1 year of age, the transfusion reaction rate in the ABO-compatible (ABO-identical) platelet group was 1.0%, while the ABOi platelet group had an elevated reaction rate of 1.7%. The increased reaction rate for ABOi platelets held true even if the analysis were limited to Centers for Disease Control and Prevention/National Healthcare Safety Network qualifying reactions or just allergic or febrile nonhemolytic reactions. The increased reaction rate with ABOi platelets was independent of unit age. Surprisingly, major-incompatible transfusions (A/B antigen incompatible) had the highest rate of reactions, at 2.0%. During the study period, three acute hemolytic reactions were reported out of 2522 plasma-incompatible platelet transfusions (0.12%). CONCLUSIONS: Our results find that compatible platelet transfusions have the lowest rate of transfusion reactions. While hemolytic reactions were observed with plasma-incompatible transfusions, the rate was low. Transfusion of ABO antigen-incompatible platelets had the highest rate of transfusion reactions and resulted in a transfusion reaction rate 1.5 to 2 times that of ABO compatible transfusions.

Platelet-derived extracellular vesicles convey mitochondrial DAMPs in platelet concentrates and their levels are associated with adverse reactions.

BACKGROUND: Whereas platelet transfusion is a common medical procedure, inflammation still occurs in a fraction of transfused individuals despite the absence of any apparent infectious agents. Platelets can shed membrane vesicles, called extracellular vesicles (EVs), some of which contain mitochondria (mito+EV). With its content of damage-associated molecular pattern (DAMP), the mitochondrion can stimulate the innate immune system. Mitochondrial DNA (mtDNA) is a recognized DAMP detected in the extracellular milieu in numerous inflammatory conditions and in platelet concentrates. We hypothesized that platelet-derived mitochondria encapsulated in EVs may represent a reservoir of mtDNA. STUDY DESIGN AND METHODS: Herein, we explored the implication of mito+EVs in the occurrence of mtDNA quantified in platelet concentrate supernatants that induced or did not induce transfusion adverse reactions. RESULTS: We observed that EVs were abundant in platelet concentrates, and platelet-derived mito+EVs were more abundant in platelet concentrates that induced adverse reactions. A significant correlation (rs = 0.73; p < 0.0001) between platelet-derived mito+EV levels and mtDNA concentrations was found. However, there was a nonsignificant correlation between the levels of EVs without mitochondria and mtDNA concentrations (rs = -0.11; p = 0.5112). The majority of the mtDNA was encapsulated into EVs. CONCLUSION: This study suggests that platelet-derived EVs, such as those that convey mitochondrial DAMPs, may be a useful biomarker for the prediction of potential risk of adverse transfusion reactions. Moreover, our work implies that investigations are necessary to determine whether there is a causal pathogenic role of mitochondrial DAMP encapsulated in EVs as opposed to mtDNA in solution.

Consequences of hemolysis: Pro-inflammatory cytokine response to erythrophagocytosis.

Red blood cells (RBCs) can be cleared from the circulation either intravascularly or extravascularly. In the setting of an IgG-mediated delayed hemolytic transfusion reaction (HTR), most RBC clearance is typically extravascular, presumably by phagocytosis by liver and splenic macrophages. Animal models of HTRs suggest that this RBC clearance is associated with a pro-inflammatory cytokine response. Although IgG-mediated HTRs are typically benign, they can lead to vaso-occlusive crises and further complications, such as hyperhemolysis, in patients with sickle cell disease (SCD). Why the manifestations of HTRs are more severe in the setting of SCD has yet to be determined; however, in this symposium paper, we provide a review of the evidence that robust RBC phagocytosis results in a pro-inflammatory cytokine response, which may induce vaso-occlusive crises and further complications in the setting of SCD.

Strategies for managing transfusional iron overload: conventional treatments and novel strategies.

PURPOSE OF REVIEW: For individuals who have transfusion-dependent anemia, iron overload is the long-term complication, which results in significant morbidity. Ameliorating this is now the biggest unmet need. This review specifically addresses this issue. RECENT FINDINGS: Over the last decade or so, major advances in the treatment of these individuals, has resulted from novel strategies aimed at reducing transfusion requirement as well as optimizing chelation therapy. This review will summarize these advances and provide insights into some of the therapies in the pipeline. Strategies aimed at reducing transfusion requirement include modulation of erythropoietic regulation by reducing ineffective red cell production through activin trapping, as well as stem cell gene modification approaches, which aim for a cure, and transfusion independence. Refined means of assessing tissue iron and the introduction of oral chelators have facilitated tailoring chelation regimens with closer monitoring and improved compliance. Newer approaches to ameliorate iron toxicity have focused on the hepcidin pathway, all of which would result in increased hepcidin levels and reduction of iron absorption from the intestine, sequestration of iron in normal storage sites and reduced exposure of more susceptible organs, such as the heart and endocrine organs, to the toxic effects of increased iron. SUMMARY: These advances offer the promise of improved management of transfusion-dependent individuals.

Genetically-engineered pigs as sources for clinical red blood cell transfusion: What pathobiological barriers need to be overcome?

An alternative to human red blood cells (RBCs) for clinical transfusion would be advantageous, particularly in situations of massive acute blood loss (where availability and compatibility are limited) or chronic hematologic diseases requiring frequent transfusions (resulting in alloimmunization). Ideally, any alternative must be neither immunogenic nor pathogenic, but readily available, inexpensive, and physiologically effective. Pig RBCs (pRBCs) provide a promising alternative due to their several similarities with human RBCs, and our increasing ability to genetically-modify pigs to reduce cellular immunogenicity. We briefly summarize the history of xenotransfusion, the progress that has been made in recent years, and the remaining barriers. These barriers include prevention of (i) human natural antibody binding to pRBCs, (ii) their phagocytosis by macrophages, and (iii) the T cell adaptive immune response (in the absence of exogenous immunosuppressive therapy). Although techniques of genetic engineering have advanced in recent years, novel methods to introduce human transgenes into pRBCs (which do not have nuclei) will need to be developed before clinical trials can be initiated.

Metabolomics Approach Based on Multivariate Techniques for Blood Transfusion Reactions.

Blood transfusions temporarily improve the physical state of the patient but exert widespread effects on immune and non-immune systems. Perioperative allogeneic blood transfusions (ABT) are associated with various risks, including coagulopathy, incompatibility, transmission of infectious agents, and allergic reactions. Nevertheless, little is known about the global metabolic alterations that reflect the possible reactions of blood transfusions. In this study, we investigated metabolite changes generated by ABT in a rat model using metabolomics technology. To further profile the "metabolome" after blood transfusions, we used both liquid chromatography-quadrupole time-of-flight high-definition mass spectrometry and gas chromatography-mass spectrometry. ABT promoted a stimulatory microenvironment associated with a relative increase in glucose transporter 1/4 (GLUT1/GLUT4) expression. Supporting this result, glucose metabolism-related enzyme IRS1 and interleukin-6 (IL-6) were abnormally expressed, and levels of lysophosphatidylcholine (LysoPC) and its related enzyme phospholipase A2 (PLA2) were significantly altered in allogeneic groups compared to those in autologous groups. Finally, amino acid metabolism was also altered following ABT. Taken together, our results show a difference between autologous and allogeneic blood transfusions and demonstrate correlations with cancer-associated metabolic changes. Our data provide endogenous information for a better understanding of blood transfusion reactions.

Diagnosis and treatment of cardiac iron overload in transfusion-dependent thalassemia patients.

INTRODUCTION: Thalassemia is among the most common genetic diseases. Patients with severe forms of the disease are transfusion-dependent, leading to iron overload. A condition which can eventually develop in the iron-loaded heart is iron overload cardiomyopathy, a debilitating disease that accounts for the majority of deaths in thalassemia patients. Areas covered: This review article provides a comprehensive summary of the diagnosis and treatment of cardiac iron overload in transfusion-dependent thalassemia patients, with discussion covering current weak points and potential improvements of the relevant diagnostic and therapeutic strategies. Expert commentary: Current limitations of various diagnostic techniques for iron overload cardiomyopathy include suboptimal accuracy, untimely detection, or inadequate accessibility, and novel modalities are required to overcome these shortcomings. Treatment should address key pathophysiologic mechanisms of iron overload cardiomyopathy, which include cardiac iron mishandling and iron-induced oxidative injury. Apart from the promotion of iron removal by chelators, prevention of cardiac iron deposition and attenuation of oxidative damage should also be rigorously investigated on a cell-to-bedside basis.

Role of heme in lung bacterial infection after trauma hemorrhage and stored red blood cell transfusion: A preclinical experimental study.

BACKGROUND: Trauma is the leading cause of death and disability in patients aged 1-46 y. Severely injured patients experience considerable blood loss and hemorrhagic shock requiring treatment with massive transfusion of red blood cells (RBCs). Preclinical and retrospective human studies in trauma patients have suggested that poorer therapeutic efficacy, increased severity of organ injury, and increased bacterial infection are associated with transfusion of large volumes of stored RBCs, although the mechanisms are not fully understood. METHODS AND FINDINGS: We developed a murine model of trauma hemorrhage (TH) followed by resuscitation with plasma and leukoreduced RBCs (in a 1:1 ratio) that were banked for 0 (fresh) or 14 (stored) days. Two days later, lungs were infected with Pseudomonas aeruginosa K-strain (PAK). Resuscitation with stored RBCs significantly increased the severity of lung injury caused by P. aeruginosa, as demonstrated by higher mortality (median survival 35 h for fresh RBC group and 8 h for stored RBC group; p < 0.001), increased pulmonary edema (mean [95% CI] 106.4 µl [88.5-124.3] for fresh RBCs and 192.5 µl [140.9-244.0] for stored RBCs; p = 0.003), and higher bacterial numbers in the lung (mean [95% CI] 1.2 × 10(7) [-1.0 × 10(7) to 2.5 × 10(7)] for fresh RBCs and 3.6 × 10(7) [2.5 × 10(7) to 4.7 × 10(7)] for stored RBCs; p = 0.014). The mechanism underlying this increased infection susceptibility and severity was free-heme-dependent, as recombinant hemopexin or pharmacological inhibition or genetic deletion of toll-like receptor 4 (TLR4) during TH and resuscitation completely prevented P. aeruginosa-induced mortality after stored RBC transfusion (p < 0.001 for all groups relative to stored RBC group). Evidence from studies transfusing fresh and stored RBCs mixed with stored and fresh RBC supernatants, respectively, indicated that heme arising both during storage and from RBC hemolysis post-resuscitation plays a role in increased mortality after PAK (p < 0.001). Heme also increased endothelial permeability and inhibited macrophage-dependent phagocytosis in cultured cells. Stored RBCs also increased circulating high mobility group box 1 (HMGB1; mean [95% CI] 15.4 ng/ml [6.7-24.0] for fresh RBCs and 50.3 ng/ml [12.3-88.2] for stored RBCs), and anti-HMGB1 blocking antibody protected against PAK-induced mortality in vivo (p = 0.001) and restored macrophage-dependent phagocytosis of P. aeruginosa in vitro. Finally, we showed that TH patients, admitted to the University of Alabama at Birmingham ER between 1 January 2015 and 30 April 2016 (n = 50), received high micromolar-millimolar levels of heme proportional to the number of units transfused, sufficient to overwhelm endogenous hemopexin levels early after TH and resuscitation. Limitations of the study include lack of assessment of temporal changes in different products of hemolysis after resuscitation and the small sample size precluding testing of associations between heme levels and adverse outcomes in resuscitated TH patients. CONCLUSIONS: We provide evidence that large volume resuscitation with stored blood, compared to fresh blood, in mice increases mortality from subsequent pneumonia, which occurs via mechanisms sensitive to hemopexin and TLR4 and HMGB1 inhibition.

Iron overload in myelodysplastic syndromes (MDS).

Iron overload (IOL) starts to develop in MDS patients before they become transfusion-dependent because ineffective erythropoiesis suppresses hepcidin production in the liver and thus leads to unrestrained intestinal iron uptake. However, the most important cause of iron overload in MDS is chronic transfusion therapy. While transfusion dependency by itself is a negative prognostic factor reflecting poor bone marrow function, the ensuing transfusional iron overload has an additional dose-dependent negative impact on the survival of patients with lower risk MDS. Cardiac dysfunction appears to be important in this context, as a consequence of chronic anemia, age-related cardiac comorbidity, and iron overload. Another potential problem is iron-related endothelial dysfunction. There is some evidence that with increasing age, high circulating iron levels worsen the atherosclerotic phenotype. Transfusional IOL also appears to aggravate bone marrow failure in MDS, through unfavorable effects on mesenchymal stromal cells as well a hematopoietic cells, particularly erythroid precursors. Patient series and clinical trials have shown that the iron chelators deferoxamine and deferasirox can improve hematopoiesis in a minority of transfusion-dependent patients. Analyses of registry data suggest that iron chelation provides a survival benefit for patients with MDS, but data from a prospective randomized clinical trial are still lacking.

Acute Effects of Blood Transfusion on Insulin Sensitivity and Pancreatic ß-Cell Function in Children with ß-Thalassemia/Hemoglobin E Disease.

OBJECTIVE: To assess the acute effects of blood transfusion on insulin sensitivity and pancreatic ß-cell function in thalassemia patients. METHODS: Fifty children and adolescents with ß-thalassemia/HbE disease were enrolled in a prospective cohort study. Hemoglobin, serum ferritin and oral glucose tolerance test (OGTT) were performed prior to, and one week after blood transfusion. Insulin sensitivity indices [homeostatic model assessment (HOMA) of insulin resistance (HOMA-IR), whole body insulin sensitivity index (WBISI)] and ß-cell function indices [HOMA of ß-cell function (HOMA-ß), insulinogenic index (IGI), and disposition index (DI)] were calculated from glucose and insulin levels obtained during the OGTT. RESULTS: Following blood transfusion, hemoglobin and serum ferritin increased significantly; 8.5 to 10.1 g/dL (p<0.001) and 1764 to 2160 ng/mL (p<0.001), respectively. ß-Cell function indices also increased significantly [median HOMA-ß: 74.3 vs. 82.7 (p=0.033); median IGI: 59.6 vs. 79.3 (p=0.003); median DI: 658 vs. 794 (p=0.01)]. However, the insulin sensitivity index (WBISI) tended to decrease and the insulin resistance index (HOMA-IR) tended to increase although this did not reach significance. Multivariate analysis showed that pre-transfusion serum ferritin was the major factor negatively associated with WBISI and positively associated with HOMA-IR, but pre-transfusion hemoglobin had no significant association with insulin sensitivity indices post-transfusion. CONCLUSION: This study demonstrated that acute increases in serum ferritin and hemoglobin following blood transfusion in patients with thalassemia might contribute to an increase in insulin secretion and to a trend towards increased insulin resistance.

Discriminating the hemolytic risk of blood type A plasmas using the complement hemolysis using human erythrocytes (CHUHE) assay.

BACKGROUND: The agglutination-based cross-matching method is sensitive for antibody binding to red blood cells but is only partially predictive of complement-mediated hemolysis, which is important in many acute hemolytic transfusion reactions. Here, we describe complement hemolysis using human erythrocytes (CHUHE) assays that directly evaluate complement-mediated hemolysis between individual serum-plasma and red blood cell combinations. The CHUHE assay is used to evaluate correlations between agglutination titers and complement-mediated hemolysis as well as the hemolytic potential of plasma from type A blood donors. STUDY DESIGN AND METHODS: Plasma or serum from each type A blood donor was incubated with AB or B red blood cells in the CHUHE assay and measured for free hemoglobin release. RESULTS: CHUHE assays for serum or plasma demonstrate a wide, dynamic range and high sensitivity for complement-mediated hemolysis for individual serum/plasma and red blood cell combinations. CHUHE results suggest that agglutination assays alone are only moderately predictive of complement-mediated hemolysis. CHUHE results also suggest that plasma from particular type A blood donors produce minimal complement-mediated hemolysis, whereas plasma from other type A blood donors produce moderate to high-level complement-mediated hemolysis, depending on the red blood cell donor. CONCLUSION: The current results indicate that the CHUHE assay can be used to assess complement-mediated hemolysis for plasma or serum from a type A blood donor, providing additional risk discrimination over agglutination titers alone.

Clinical response and transfusion reactions of sheep subjected to single homologous blood transfusion.

Studies in relation to blood conservation and responses to transfusion are scarce for ruminants. We evaluated the clinical manifestations of sheep that received a single homologous transfusion of whole blood, focusing on transfusion reactions. Eighteen adult sheep were subjected to a single phlebotomy to withdraw 40% of the total blood volume, which was placed into CPDA-1 bags and then divided into G0, animals that received fresh blood, and G15 and G35, animals that received blood stored for 15 or 35 days, respectively. Clinical observations were recorded throughout the transfusion, whereas heart rate, respiratory rate, and rectal temperature were assessed at the following times: 24 hours after phlebotomy and before transfusion; 30 minutes, six, twelve, 24, 48, 72, and 96 hours and eight and 16 days after transfusion. All groups presented transfusion reactions, among which hyperthermia was the most frequent (50% of animals). Tachycardia occurred most frequently in the G35 animals (50% of them). During transfusion G35 animals presented more clinical manifestation (P < 0.05). Transfusion of fresh or stored total blood improved the blood volume, but transfusion reactions occurred, demonstrating that a single transfusion of fresh or stored blood can cause inflammatory and febrile nonhemolytic transfusion reactions in sheep.