A randomized trial of blood donor iron repletion on red cell quality for transfusion and donor cognition and well-being.

Although altruistic regular blood donors are vital for the blood supply, many become iron deficient from donation-induced iron loss. The effects of blood donation-induced iron deficiency on red cell transfusion quality or donor cognition are unknown. In this double-blind, randomized trial, adult iron-deficient blood donors (n = 79; ferritin < 15 µg/L and zinc protoporphyrin >60 µMol/mol heme) who met donation qualifications were enrolled. A first standard blood donation was followed by the gold-standard measure for red cell storage quality: a 51-chromium posttransfusion red cell recovery study. Donors were then randomized to intravenous iron repletion (1 g low-molecular-weight iron dextran) or placebo. A second donation ∼5 months later was followed by another recovery study. Primary outcome was the within-subject change in posttransfusion recovery. The primary outcome measure of an ancillary study reported here was the National Institutes of Health Toolbox-derived uncorrected standard Cognition Fluid Composite Score. Overall, 983 donors were screened; 110 were iron-deficient, and of these, 39 were randomized to iron repletion and 40 to placebo. Red cell storage quality was unchanged by iron repletion: mean change in posttransfusion recovery was 1.6% (95% confidence interval -0.5 to 3.8) and -0.4% (-2.0 to 1.2) with and without iron, respectively. Iron repletion did not affect any cognition or well-being measures. These data provide evidence that current criteria for blood donation preserve red cell transfusion quality for the recipient and protect adult donors from measurable effects of blood donation-induced iron deficiency on cognition. This trial was registered at www.clinicaltrials.gov as NCT02889133 and NCT02990559.

Admission Viscoelastic Hemostatic Assay Parameters Predict Poor Long-Term Intracerebral Hemorrhage Outcomes.

BACKGROUND: Viscoelastic hemostatic assays (VHAs) provide more comprehensive assessments of coagulation compared with conventional coagulation assays. Although VHAs have enabled guided hemorrhage control therapies, improving clinical outcomes in life-threatening hemorrhage, the role of VHAs in intracerebral hemorrhage (ICH) is unclear. If VHAs can identify coagulation abnormalities relevant for ICH outcomes, this would support the need to investigate the role of VHAs in ICH treatment paradigms. Thus, we investigated whether VHA assessments of coagulation relate to long-term ICH outcomes. METHODS: Patients with spontaneous ICH enrolled into a single-center cohort study receiving admission Rotational Thromboelastometry (ROTEM) VHA testing between 2013 and 2020 were assessed. Patients with previous anticoagulant use or coagulopathy on conventional coagulation assays were excluded. Primary ROTEM exposure variables were coagulation kinetics and clot strength assessments. Poor long-term outcome was defined as modified Rankin Scale ≥ 4 at 6 months. Logistic regression analyses assessed associations of ROTEM parameters with clinical outcomes after adjusting for ICH severity and hemoglobin concentration. RESULTS: Of 44 patients analyzed, the mean age was 64 years, 57% were female, and the median ICH volume was 23 mL. Poor 6-month outcome was seen in 64% of patients. In our multivariable regression models, slower, prolonged coagulation kinetics (adjusted odds ratio for every second increase in clot formation time 1.04, 95% confidence interval 1.00-1.09, p = 0.04) and weaker clot strength (adjusted odds ratio for every millimeter increase of maximum clot firmness 0.84, 95% confidence interval 0.71-0.99, p = 0.03) were separately associated with poor long-term outcomes. CONCLUSIONS: Slower, prolonged coagulation kinetics and weaker clot strength on admission VHA ROTEM testing, not attributable to anticoagulant use, were associated with poor long-term outcomes after ICH. Further work is needed to clarify the generalizability and the underlying mechanisms of these VHA findings to assess whether VHA-guided treatments should be incorporated into ICH care.

Apixaban to Prevent Recurrence After Cryptogenic Stroke in Patients With Atrial Cardiopathy: The ARCADIA Randomized Clinical Trial.

Importance: Atrial cardiopathy is associated with stroke in the absence of clinically apparent atrial fibrillation. It is unknown whether anticoagulation, which has proven benefit in atrial fibrillation, prevents stroke in patients with atrial cardiopathy and no atrial fibrillation. Objective: To compare anticoagulation vs antiplatelet therapy for secondary stroke prevention in patients with cryptogenic stroke and evidence of atrial cardiopathy. Design, Setting, and Participants: Multicenter, double-blind, phase 3 randomized clinical trial of 1015 participants with cryptogenic stroke and evidence of atrial cardiopathy, defined as P-wave terminal force greater than 5000 µV × ms in electrocardiogram lead V1, serum N-terminal pro-B-type natriuretic peptide level greater than 250 pg/mL, or left atrial diameter index of 3 cm/m2 or greater on echocardiogram. Participants had no evidence of atrial fibrillation at the time of randomization. Enrollment and follow-up occurred from February 1, 2018, through February 28, 2023, at 185 sites in the National Institutes of Health StrokeNet and the Canadian Stroke Consortium. Interventions: Apixaban, 5 mg or 2.5 mg, twice daily (n = 507) vs aspirin, 81 mg, once daily (n = 508). Main Outcomes and Measures: The primary efficacy outcome in a time-to-event analysis was recurrent stroke. All participants, including those diagnosed with atrial fibrillation after randomization, were analyzed according to the groups to which they were randomized. The primary safety outcomes were symptomatic intracranial hemorrhage and other major hemorrhage. Results: With 1015 of the target 1100 participants enrolled and mean follow-up of 1.8 years, the trial was stopped for futility after a planned interim analysis. The mean (SD) age of participants was 68.0 (11.0) years, 54.3% were female, and 87.5% completed the full duration of follow-up. Recurrent stroke occurred in 40 patients in the apixaban group (annualized rate, 4.4%) and 40 patients in the aspirin group (annualized rate, 4.4%) (hazard ratio, 1.00 [95% CI, 0.64-1.55]). Symptomatic intracranial hemorrhage occurred in 0 patients taking apixaban and 7 patients taking aspirin (annualized rate, 1.1%). Other major hemorrhages occurred in 5 patients taking apixaban (annualized rate, 0.7%) and 5 patients taking aspirin (annualized rate, 0.8%) (hazard ratio, 1.02 [95% CI, 0.29-3.52]). Conclusions and Relevance: In patients with cryptogenic stroke and evidence of atrial cardiopathy without atrial fibrillation, apixaban did not significantly reduce recurrent stroke risk compared with aspirin. Trial Registration: ClinicalTrials.gov Identifier: NCT03192215.

ABO blood type and thromboembolic complications after intracerebral hemorrhage: An exploratory analysis.

BACKGROUND AND PURPOSE: Non-O blood types are known to be associated with thromboembolic complications (TECs) in population-based studies. TECs are known drivers of morbidity and mortality in intracerebral hemorrhage (ICH) patients, yet the relationships of blood type on TECs in this patient population are unknown. We sought to explore the relationships between ABO blood type and TECs in ICH patients. METHODS: Consecutive adult ICH patients enrolled into a prospective observational cohort study with available ABO blood type data were analyzed. Patients with cancer history, prior thromboembolism, and baseline laboratory evidence of coagulopathy were excluded. The primary exposure variable was blood type (non-O versus O). The primary outcome was composite TEC, defined as pulmonary embolism, deep venous thrombosis, ischemic stroke or myocardial infarction, during the hospital stay. Relationships between blood type, TECs and clinical outcomes were separately assessed using logistic regression models after adjusting for sex, ethnicity and ICH score. RESULTS: Of 301 ICH patients included for analysis, 44% were non-O blood type. Non-O blood type was associated with higher admission GCS and lower ICH score on baseline comparisons. We identified TECs in 11.6% of our overall patient cohort. . Although TECs were identified in 9.9% of non-O blood type patients compared to 13.0% in O blood type patients, we did not identify a significant relationship of non-O blood type with TECs (adjusted OR=0.776, 95%CI: 0.348-1.733, p=0.537). The prevalence of specific TECs were also comparable in unadjusted and adjusted analyses between the two cohorts. In additional analyses, we identified that TECs were associated with poor 90-day mRS (adjusted OR=3.452, 95% CI: 1.001-11.903, p=0.050). We did not identify relationships between ABO blood type and poor 90-day mRS (adjusted OR=0.994, 95% CI:0.465-2.128, p=0.988). CONCLUSIONS: We identified that TECs were associated with worse ICH outcomes. However, we did not identify relationships in ABO blood type and TECs. Further work is required to assess best diagnostic and prophylactic and treatment strategies for TECs to improve ICH outcomes.

Genome-wide metabolite quantitative trait loci analysis (mQTL) in red blood cells from volunteer blood donors.

The red blood cell (RBC)-Omics study, part of the larger NHLBI-funded Recipient Epidemiology and Donor Evaluation Study (REDS-III), aims to understand the genetic contribution to blood donor RBC characteristics. Previous work identified donor demographic, behavioral, genetic, and metabolic underpinnings to blood donation, storage, and (to a lesser extent) transfusion outcomes, but none have yet linked the genetic and metabolic bodies of work. We performed a genome-wide association (GWA) analysis using RBC-Omics study participants with generated untargeted metabolomics data to identify metabolite quantitative trait loci in RBCs. We performed GWA analyses of 382 metabolites in 243 individuals imputed using the 1000 Genomes Project phase 3 all-ancestry reference panel. Analyses were conducted using ProbABEL and adjusted for sex, age, donation center, number of whole blood donations in the past 2 years, and first 10 principal components of ancestry. Our results identified 423 independent genetic loci associated with 132 metabolites (p < 5×10-8). Potentially novel locus-metabolite associations were identified for the region encoding heme transporter FLVCR1 and choline and for lysophosphatidylcholine acetyltransferase LPCAT3 and lysophosphatidylserine 16.0, 18.0, 18.1, and 18.2; these associations are supported by published rare disease and mouse studies. We also confirmed previous metabolite GWA results for associations, including N(6)-methyl-L-lysine and protein PYROXD2 and various carnitines and transporter SLC22A16. Association between pyruvate levels and G6PD polymorphisms was validated in an independent cohort and novel murine models of G6PD deficiency (African and Mediterranean variants). We demonstrate that it is possible to perform metabolomics-scale GWA analyses with a modest, trans-ancestry sample size.

Relationships of Hemoglobin Concentration, Ischemic Lesions, and Clinical Outcomes in Patients With Intracerebral Hemorrhage.

BACKGROUND: Hemoglobin concentration and diffusion-weighted imaging (DWI) ischemic lesions are separately known to be associated with poor intracerebral hemorrhage (ICH) outcomes. While hemoglobin concentrations have known relationships with ischemic stroke, it is unclear whether hemoglobin concentration is associated with DWI ischemic lesions after ICH. We sought to investigate the hypothesis that hemoglobin concentrations would associate with DWI lesions after ICH and further investigated their relationships with clinical outcomes. METHODS: Supratentorial ICH patients enrolled between 2010 and 2016 to a prospective, multicenter, observational cohort study (ERICH study [Ethnic/Racial Variations of Intracerebral Hemorrhage]) were assessed. Patients from this study with baseline, admission hemoglobin, and hospitalization magnetic resonance imaging were analyzed. Hemoglobin was examined as the primary exposure variable defined as a continuous variable (g/dL). Magnetic resonance imaging DWI ischemic lesion presence was assessed as the primary radiographic outcome. Primary analyses assessed relationships of hemoglobin with DWI lesions. Secondary analyses assessed relationships of DWI lesions with poor 3-month outcomes (modified Rankin Scale score, 4-6). These analyses were performed using separate multivariable logistic regression models adjusting for relevant covariates. RESULTS: Of 917 patients with ICH analyzed, mean baseline hemoglobin was 13.8 g/dL (±1.9), 60% were deep ICH, and DWI lesions were identified in 27% of the cohort. In our primary analyses, increased hemoglobin, defined as a continuous variable, was associated with DWI lesions (adjusted odds ratio, 1.21 per 1 g/dL change in hemoglobin [95% CI, 1.07-1.37]) after adjusting for sex, race, ICH severity, time to magnetic resonance imaging, and acute blood pressure change. In secondary analyses, DWI lesions were associated with poor 3-month outcomes (adjusted odds ratio, 1.83 [95% CI, 1.24-2.69]) after adjusting for similar covariates. CONCLUSIONS: We identified novel relationships between higher baseline hemoglobin concentrations and DWI ischemic lesions in patients with ICH. Further studies are required to clarify the role of hemoglobin concentration on both cerebral small vessel disease pathophysiology and ICH outcomes.

Impacts of ABO-incompatible platelet transfusions on platelet recovery and outcomes after intracerebral hemorrhage.

Acute platelet transfusion after intracerebral hemorrhage (ICH) given in efforts to reverse antiplatelet medication effects and prevent ongoing bleeding does not appear to improve outcome and may be associated with harm. Although the underlying mechanisms are unclear, the influence of ABO-incompatible platelet transfusions on ICH outcomes has not been investigated. We hypothesized that patients with ICH who receive ABO-incompatible platelet transfusions would have worse platelet recovery (using absolute count increment [ACI]) and neurological outcomes (mortality and poor modified Rankin Scale [mRS 4-6]) than those receiving ABO-compatible transfusions. In a single-center cohort of consecutively admitted patients with ICH, we identified 125 patients receiving acute platelet transfusions, of whom 47 (38%) received an ABO-incompatible transfusion. Using quantile regression, we identified an association of ABO-incompatible platelet transfusion with lower platelet recovery (ACI, 2 × 103cells per µL vs 15 × 103cells per µL; adjusted coefficient ß, -19; 95% confidence interval [CI], -35.55 to -4.44; P = .01). ABO-incompatible platelet transfusion was also associated with increased odds of mortality (adjusted odds ratio [OR], 2.59; 95% CI, 1.00-6.73; P = .05) and poor mRS (adjusted OR, 3.61; 95% CI, 0.97-13.42; P = .06); however, these estimates were imprecise. Together, these findings suggest the importance of ABO compatibility for platelet transfusions for ICH, but further investigation into the mechanism(s) underlying these observations is required.

Rapid clearance of storage-induced microerythrocytes alters transfusion recovery.

Permanent availability of red blood cells (RBCs) for transfusion depends on refrigerated storage, during which morphologically altered RBCs accumulate. Among these, a subpopulation of small RBCs, comprising type III echinocytes, spheroechinocytes, and spherocytes and defined as storage-induced microerythrocytes (SMEs), could be rapidly cleared from circulation posttransfusion. We quantified the proportion of SMEs in RBC concentrates from healthy human volunteers and assessed correlation with transfusion recovery, investigated the fate of SMEs upon perfusion through human spleen ex vivo, and explored where and how SMEs are cleared in a mouse model of blood storage and transfusion. In healthy human volunteers, high proportion of SMEs in long-stored RBC concentrates correlated with poor transfusion recovery. When perfused through human spleen, 15% and 61% of long-stored RBCs and SMEs were cleared in 70 minutes, respectively. High initial proportion of SMEs also correlated with high retention of RBCs by perfused human spleen. In the mouse model, SMEs accumulated during storage. Transfusion of long-stored RBCs resulted in reduced posttransfusion recovery, mostly due to SME clearance. After transfusion in mice, long-stored RBCs accumulated predominantly in spleen and were ingested mainly by splenic and hepatic macrophages. In macrophage-depleted mice, splenic accumulation and SME clearance were delayed, and transfusion recovery was improved. In healthy hosts, SMEs were cleared predominantly by macrophages in spleen and liver. When this well-demarcated subpopulation of altered RBCs was abundant in RBC concentrates, transfusion recovery was diminished. SME quantification has the potential to improve blood product quality assessment. This trial was registered at www.clinicaltrials.gov as #NCT02889133.

Reticulocytes in donor blood units enhance red blood cell alloimmunization.

Although red blood cell (RBC) transfusions save lives, some patients develop clinically-significant alloantibodies against donor blood group antigens, which then have adverse effects in multiple clinical settings. Few effective measures exist to prevent RBC alloimmunization and/or eliminate alloantibodies in sensitized patients. Donor-related factors may influence alloimmunization; thus, there is an unmet clinical need to identify which RBC units are immunogenic. Repeat volunteer blood donors and donors on iron supplements have elevated reticulocyte counts compared to healthy non-donors. Early reticulocytes retain mitochondria and other components, which may act as danger signals in immune responses. Herein, we tested whether reticulocytes in donor RBC units could enhance RBC alloimmunization. Using a murine model, we demonstrate that transfusing donor RBC units with increased reticulocyte frequencies dose-dependently increased RBC alloimmunization rates and alloantibody levels. Transfusing reticulocyte-rich RBC units was associated with increased RBC clearance from the circulation and a robust proinflammatory cytokine response. As compared to previously reported post-transfusion RBC consumption patterns, erythrophagocytosis from reticulocyte-rich units was increasingly performed by splenic B cells. These data suggest that reticulocytes in a donated RBC unit impact the quality of blood transfused, are targeted to a distinct compartment, and may be an underappreciated risk factor for RBC alloimmunization.

Blood donor, component, and recipient-specific factors associated with venous thromboembolism in transfused hospitalized adult patients: Data from the recipient epidemiology and donor evaluation Study-III (REDS-III).

OBJECTIVE: Growing evidence suggests multiple pathophysiological mechanisms linking red blood cells (RBC) transfusions to thrombosis. This study examined blood donor, component, and recipient factors which may be associated with thromboembolic outcomes following RBC transfusion. METHODS: We utilized the Recipient Epidemiology Donor Evaluation Study-III (REDS-III) database on patients transfused in 12 hospitals between 2013-2016. Stratified Cox proportional hazards regression models with time-dependent exposures were used to examine associations of donor and component modification characteristics on venous thromboembolism (VTE) in patients transfused RBC units. RESULTS: 59,603 patients were transfused 229,500 RBC units during 79,298 hospitalizations with post-transfusion VTE occurring in 1869 (2.4%) of patients. In adjusted regression analyses, a per RBC-unit risk of VTE was present for gamma irradiation (HR = 1.03; 95% CI: 1.02-1.03), female donor sex (HR = 1.01; 95% CI: 1.00-1.01), storage duration greater than 5 weeks (HR = 1.01; 95% CI: 1.01-1.02), AS-1 storage solution (HR = 1.01; 95% CI: 1.00-1.01), and apheresis-derived collections (HR = 1.01; 95% CI: 1.01-1.02). Among recipient factors, male sex (HR = 1.03; 95% CI: 1.02-1.04), pre-transfusion hemoglobin level (HR = 0.94; 95% CI: 0.94-0.94), body mass index strata (HR = 1.11; 95% CI: 1.08-1.14), and principal diagnoses including malignancy (HR = 1.13; 95% CI: 1.10-1.16), cardiac arrest (HR = 1.38; 95% CI:1.07-1.77) and hip fracture (HR = 1.59; 95% CI:1.53-1.66) were associated with VTE in adjusted analyses. DISCUSSION: We identified several donor, component, and recipient-specific factors associated with VTE in transfused hospitalized adult patients. In adjusted models, the dose-dependent associations of donor and component-specific factors with VTE were modest and unlikely to be clinically significant in the majority of transfused patients. Additional mechanistic and clinical studies linking blood donor and component factors with thrombotic outcomes are needed.

Associations between ABO non-identical platelet transfusions and patient outcomes-A multicenter retrospective analysis.

BACKGROUND: Due to platelet availability limitations, platelet units ABO mismatched to recipients are often transfused. However, since platelets express ABO antigens and are collected in plasma which may contain ABO isohemagglutinins, it remains controversial as to whether ABO non-identical platelet transfusions could potentially pose harm and/or have reduced efficacy. STUDY DESIGN AND METHODS: The large 4-year publicly available Recipient Epidemiology and Donor Evaluation Study-III (REDS-III) database was used to investigate patient outcomes associated with ABO non-identical platelet transfusions. Outcomes included mortality, sepsis, and subsequent platelet transfusion requirements. RESULTS: Following adjustment for possible confounding factors, no statistically significant association between ABO non-identical platelet transfusion and increased risk of mortality was observed in the overall cohort of 21,176 recipients. However, when analyzed by diagnostic category and recipient ABO group, associations with increased mortality for major mismatched transfusions were noted in two of eight subpopulations. Hematology/Oncology blood group A and B recipients (but not group O) showed a Hazard Ratio (HR) of 1.29 (95%CI: 1.03-1.62) and intracerebral hemorrhage group O recipients (but not groups A and B) showed a HR of 1.75 (95%CI: 1.10-2.80). Major mismatched transfusions were associated with increased odds of receiving additional platelet transfusion each post-transfusion day (through day 5) regardless of the recipient blood group. DISCUSSION: We suggest that prospective studies are needed to determine if specific patient populations would benefit from receiving ABO identical platelet units. Our findings indicate that ABO-identical platelet products minimize patient exposure to additional platelet doses.

Proceedings of the 2022 NHLBI and OASH state of the science in transfusion medicine symposium.

BACKGROUND: State of the Science (SoS) meetings are used to define and highlight important unanswered scientific questions. The National Heart, Lung, and Blood Institute (NHLBI), National Institutes of Health, and the Office of the Assistant Secretary for Health (OASH), Department of Health and Human Services held a virtual SoS in transfusion medicine (TM) symposium. STUDY DESIGN AND METHODS: In advance of the symposium, six multidisciplinary working groups (WG) convened to define research priorities in the areas of: blood donors and the supply, optimizing transfusion outcomes for recipients, emerging infections, mechanistic aspects of components and transfusion, new computational methods in transfusion science, and impact of health disparities on donors and recipients. The overall objective was to identify key basic, translational, and clinical research questions that will help to increase and diversify the volunteer donor pool, ensure safe and effective transfusion strategies for recipients, and identify which blood products from which donors best meet the clinical needs of specific recipient populations. RESULTS: On August 29-30, 2022, over 400 researchers, clinicians, industry experts, government officials, community members, and patient advocates discussed the research priorities presented by each WG. Dialogue focused on the five highest priority research areas identified by each WG and included the rationale, proposed methodological approaches, feasibility, and barriers for success. DISCUSSION: This report summarizes the key ideas and research priorities identified during the NHLBI/OASH SoS in TM symposium. The report highlights major gaps in our current knowledge and provides a road map for TM research.

Irradiation Causes Alterations of Polyamine, Purine, and Sulfur Metabolism in Red Blood Cells and Multiple Organs.

Investigating the metabolic effects of radiation is critical to understand the impact of radiotherapy, space travel, and exposure to environmental radiation. In patients undergoing hemopoietic stem cell transplantation, iron overload is a common risk factor for poor outcomes. However, no studies have interrogated the multiorgan effects of these treatments concurrently. Herein, we use a model that recapitulates transfusional iron overload, a condition often observed in chronically transfused patients. We applied an omics approach to investigate the impact of both the iron load and irradiation on the host metabolome. The results revealed dose-dependent effects of irradiation in the red blood cells, plasma, spleen, and liver energy and redox metabolism. Increases in polyamines and purine salvage metabolites were observed in organs with high oxygen consumption including the heart, kidneys, and brain. Irradiation also impacted the metabolism of the duodenum, colon, and stool, suggesting a potential effect on the microbiome. Iron infusion affected the response to radiation in the organs and blood, especially in erythrocyte polyamines and spleen antioxidant metabolism, and affected glucose, methionine, and glutathione systems and tryptophan metabolism in the liver, stool, and the brain. Together, the results suggest that radiation impacts metabolism on a multiorgan level with a significant interaction of the host iron status.

Red blood cell transfusion-induced non-transferrin-bound iron promotes Pseudomonas aeruginosa biofilms in human sera and mortality in catheterized mice.

Transfusion of storage-damaged red blood cells (RBCs) increases non-transferrin-bound iron (NTBI) levels in humans. This can potentially enhance virulence of microorganisms. In this study, Pseudomonas aeruginosa replication and biofilm production in vitro correlated with NTBI levels of transfused subjects (R2 = 0·80; P < 0·0001). Transfusion of stored RBCs into catheterized mice enhanced P. aeruginosa virulence and mortality in vivo, while pre-administration of apotransferrin reduced NTBI levels improving survival (69% vs 27% mortality; P < 0·05). These results suggest that longer RBC storage, by modulating the bioavailability of iron, may increase the risk of P. aeruginosa biofilm-related infections in transfused patients.

Retention of functional mitochondria in mature red blood cells from patients with sickle cell disease.

Sickle cell disease (SCD) is an inherited blood disorder characterized by sickled red blood cells (RBCs), which are more sensitive to haemolysis and can contribute to disease pathophysiology. Although treatment of SCD can include RBC transfusion, patients with SCD have high rates of alloimmunization. We hypothesized that RBCs from patients with SCD have functionally active mitochondria and can elicit a type 1 interferon response. We evaluated blood samples from more than 100 patients with SCD and found elevated frequencies of mitochondria in reticulocytes and mature RBCs, as compared to healthy blood donors. The presence of mitochondria in mature RBCs was confirmed by flow cytometry, electron microscopy, and proteomic analysis. The mitochondria in mature RBCs were metabolically competent, as determined by enzymatic activities and elevated levels of mitochondria-derived metabolites. Metabolically-active mitochondria in RBCs may increase oxidative stress, which could facilitate and/or exacerbate SCD complications. Coculture of mitochondria-positive RBCs with neutrophils induced production of type 1 interferons, which are known to increase RBC alloimmunization rates. These data demonstrate that mitochondria retained in mature RBCs are functional and can elicit immune responses, suggesting that inappropriate retention of mitochondria in RBCs may play an underappreciated role in SCD complications and be an RBC alloimmunization risk factor.

The Recipient Epidemiology and Donor Evaluation Study-IV-Pediatric (REDS-IV-P): A research program striving to improve blood donor safety and optimize transfusion outcomes across the lifespan.

BACKGROUND: The Recipient Epidemiology and Donor Evaluation Study-IV-Pediatric (REDS-IV-P) is a new iteration of prior National Heart, Lung, and Blood Institute (NHLBI) REDS programs that focus on improving transfusion recipient outcomes across the lifespan as well as the safety and availability of the blood supply. STUDY DESIGN AND METHODS: The US program includes blood centers and hospitals (22 including 6 free-standing Children's hospitals) in four geographic regions. The Brazilian program has 5 participating hemocenters. A Center for Transfusion Laboratory Studies (CTLS) and a Data Coordinating Center (DCC) support synergistic studies and activities over the 7-year REDS-IV-P program. RESULTS: The US is building a centralized, vein-to-vein (V2V) database, linking information collected from blood donors, their donations, the resulting manufactured components, and data extracts from hospital electronic medical records of transfused and non-transfused patients. Simultaneously, the Brazilian program is building a donor, donation, and component database. The databases will serve as the backbone for retrospective and prospective observational studies in transfusion epidemiology, transfusion recipient outcomes, blood component quality, and emerging blood safety issues. Special focus will be on preterm infants, patients with sickle cell disease, thalassemia or cancer, and the effect of donor biologic variability and component manufacturing on recipient outcomes. A rapid response capability to emerging safety threats has resulted in timely studies related to Severe Acute Respiratory Syndrome Corona Virus-2 (SARS-CoV-2). CONCLUSIONS: The REDS-IV-P program endeavors to improve donor-recipient-linked research with a focus on children and special populations while also maintaining the flexibility to address emerging blood safety issues.

Prevalence and predictors of SARS-CoV-2 antibodies among solid organ transplant recipients with confirmed infection.

It remains uncertain whether immunocompromised patients including solid organ transplant (SOT) recipients will have a robust antibody response to SARS-CoV-2 infection. We enrolled all adult SOT recipients at our center with confirmed SARS-CoV-2 infection who underwent antibody testing with a single commercially available anti-nucleocapsid antibody test at least 7 days after diagnosis in a retrospective cohort. Seventy SOT recipients were studied (56% kidney, 19% lung, 14% liver ± kidney, and 11% heart ± kidney recipients). Thirty-six (51%) had positive anti-nucleocapsid antibody testing, and 34 (49%) were negative. Recipients of a kidney allograft were less likely to have positive antibody testing compared to those who did not receive a kidney (p = .04). In the final multivariable model, the years from transplant to diagnosis (OR 1.26, p = .002) and baseline immunosuppression with more than two agents (OR 0.26, p = .03) were significantly associated with the antibody test result, controlling for kidney transplantation. In conclusion, among SOT recipients with confirmed infection, only 51% of patients had detectable anti-nucleocapsid antibodies, and transplant-related variables including the level and nature of immunosuppression were important predictors. These findings raise the concern that SOT recipients with COVID-19 may be less likely to form SARS-CoV-2 antibodies.

Hemolytic anemia blunts the cytokine response to transfusion of older red blood cells in mice and dogs.

BACKGROUND: Transfusion of red blood cells (RBCs) stored for longer durations induces hemolysis and inflammatory cytokine production in murine and canine models. Despite immune system activation by stored RBCs, human randomized trials suggest that fresher RBC transfusions do not improve clinical outcomes. We hypothesized that underlying recipient hemolysis may affect cytokine responses to older RBC transfusions. STUDY DESIGN AND METHODS: C57BL/6 mouse cohorts were infused with anti-TER119 antibody to induce hemolysis, rabbit anti-platelet antiserum to induce immune thrombocytopenia (ITP), or appropriate control antibodies. Two days later, mice were transfused with fresh or stored RBCs. Furthermore, in a prospective, randomized, blinded trial, 38 client-owned dogs with primary autoimmune hemolytic anemia (AIHA) and two dogs with ITP, requiring RBC transfusion, were enrolled and randomized to receive fresh (≤7 days) or old (≥21 days) stored RBC transfusions. Monocyte chemoattractant protein (MCP)-1 levels were assessed at defined times after transfusion. RESULTS: Prior immune-mediated hemolysis blunted the MCP-1 response to stored RBC transfusion in mice (361 ± 111 pg/ml vs. 6836 ± 1528 pg/ml in mice with immune hemolysis vs. ITP, respectively; mean ± SD; p < .0001). Although hemolysis markers increased after transfusion of older RBCs, the cytokine response was also muted in dogs with AIHA. No differences in morbidity or mortality were evident comparing dogs randomized to fresh or old RBCs. CONCLUSION: These data suggest that underlying hemolysis blunts inflammatory responses to old RBC transfusions. The canine data support randomized trial results suggesting a lack of clinical benefit with fresh RBC transfusions in subjects with underlying, baseline hemolysis.

Serum Proteomics in COVID-19 Patients: Altered Coagulation and Complement Status as a Function of IL-6 Level.

Over 5 million people around the world have tested positive for the beta coronavirus SARS-CoV-2 as of May 29, 2020, a third of which are in the United States alone. These infections are associated with the development of a disease known as COVID-19, which is characterized by several symptoms, including persistent dry cough, shortness of breath, chills, muscle pain, headache, loss of taste or smell, and gastrointestinal distress. COVID-19 has been characterized by elevated mortality (over 100 thousand people have already died in the US alone), mostly due to thromboinflammatory complications that impair lung perfusion and systemic oxygenation in the most severe cases. While the levels of pro-inflammatory cytokines such as interleukin-6 (IL-6) have been associated with the severity of the disease, little is known about the impact of IL-6 levels on the proteome of COVID-19 patients. The present study provides the first proteomics analysis of sera from COVID-19 patients, stratified by circulating levels of IL-6, and correlated to markers of inflammation and renal function. As a function of IL-6 levels, we identified significant dysregulation in serum levels of various coagulation factors, accompanied by increased levels of antifibrinolytic components, including several serine protease inhibitors (SERPINs). These were accompanied by up-regulation of the complement cascade and antimicrobial enzymes, especially in subjects with the highest levels of IL-6, which is consistent with an exacerbation of the acute phase response in these subjects. Although our results are observational, they highlight a clear increase in the levels of inhibitory components of the fibrinolytic cascade in severe COVID-19 disease, providing potential clues related to the etiology of coagulopathic complications in COVID-19 and paving the way for potential therapeutic interventions, such as the use of pro-fibrinolytic agents. Raw data for this study are available through ProteomeXchange with identifier PXD020601.

Evidence of Structural Protein Damage and Membrane Lipid Remodeling in Red Blood Cells from COVID-19 Patients.

The SARS-CoV-2 beta coronavirus is the etiological driver of COVID-19 disease, which is primarily characterized by shortness of breath, persistent dry cough, and fever. Because they transport oxygen, red blood cells (RBCs) may play a role in the severity of hypoxemia in COVID-19 patients. The present study combines state-of-the-art metabolomics, proteomics, and lipidomics approaches to investigate the impact of COVID-19 on RBCs from 23 healthy subjects and 29 molecularly diagnosed COVID-19 patients. RBCs from COVID-19 patients had increased levels of glycolytic intermediates, accompanied by oxidation and fragmentation of ankyrin, spectrin beta, and the N-terminal cytosolic domain of band 3 (AE1). Significantly altered lipid metabolism was also observed, in particular, short- and medium-chain saturated fatty acids, acyl-carnitines, and sphingolipids. Nonetheless, there were no alterations of clinical hematological parameters, such as RBC count, hematocrit, or mean corpuscular hemoglobin concentration, with only minor increases in mean corpuscular volume. Taken together, these results suggest a significant impact of SARS-CoV-2 infection on RBC structural membrane homeostasis at the protein and lipid levels. Increases in RBC glycolytic metabolites are consistent with a theoretically improved capacity of hemoglobin to off-load oxygen as a function of allosteric modulation by high-energy phosphate compounds, perhaps to counteract COVID-19-induced hypoxia. Conversely, because the N-terminus of AE1 stabilizes deoxyhemoglobin and finely tunes oxygen off-loading and metabolic rewiring toward the hexose monophosphate shunt, RBCs from COVID-19 patients may be less capable of responding to environmental variations in hemoglobin oxygen saturation/oxidant stress when traveling from the lungs to peripheral capillaries and vice versa.