Phenotypic and proteomic characterization of the human erythroid progenitor continuum reveal dynamic changes in cell cycle and in metabolic pathways.

Human erythropoiesis is a complex process leading to the production of 2.5 million red blood cells per second. Following commitment of hematopoietic stem cells to the erythroid lineage, this process can be divided into three distinct stages: erythroid progenitor differentiation, terminal erythropoiesis, and reticulocyte maturation. We recently resolved the heterogeneity of erythroid progenitors into four different subpopulations termed EP1-EP4. Here, we characterized the growth factor(s) responsiveness of these four progenitor populations in terms of proliferation and differentiation. Using mass spectrometry-based proteomics on sorted erythroid progenitors, we quantified the absolute expression of ~5500 proteins from EP1 to EP4. Further functional analyses highlighted dynamic changes in cell cycle in these populations with an acceleration of the cell cycle during erythroid progenitor differentiation. The finding that E2F4 expression was increased from EP1 to EP4 is consistent with the noted changes in cell cycle. Finally, our proteomic data suggest that the protein machinery necessary for both oxidative phosphorylation and glycolysis is present in these progenitor cells. Together, our data provide comprehensive insights into growth factor-dependence of erythroid progenitor proliferation and the proteome of four distinct populations of human erythroid progenitors which will be a useful framework for the study of erythroid disorders.

Impairment of human terminal erythroid differentiation by histone deacetylase 5 deficiency.

Histone deacetylases (HDACs) are a group of enzymes that catalyze the removal of acetyl groups from histone and nonhistone proteins. HDACs have been shown to have diverse functions in a wide range of biological processes. However, their roles in mammalian erythropoiesis remain to be fully defined. This study showed that, of the 11 classic HDAC family members, 6 (HDAC1, -2, -3, and HDAC5, -6, -7) are expressed in human erythroid cells, with HDAC5 most significantly upregulated during terminal erythroid differentiation. Knockdown of HDAC5 by either short hairpin RNA or small interfering RNA in human CD34+ cells followed by erythroid cell culture led to increased apoptosis, decreased chromatin condensation, and impaired enucleation of erythroblasts. Biochemical analyses revealed that HDAC5 deficiency resulted in activation of p53 in association with increased acetylation of p53. Furthermore, although acetylation of histone 4 (H4) is decreased during normal terminal erythroid differentiation, HDAC5 deficiency led to increased acetylation of H4 (K12) in late-stage erythroblasts. This increased acetylation was accompanied by decreased chromatin condensation, implying a role for H4 (K12) deacetylation in chromatin condensation. ATAC-seq and RNA sequencing analyses revealed that HDAC5 knockdown leads to increased chromatin accessibility genome-wide and global changes in gene expression. Moreover, pharmacological inhibition of HDAC5 by the inhibitor LMK235 also led to increased H4 acetylation, impaired chromatin condensation, and enucleation. Taken together, our findings have uncovered previously unrecognized roles and molecular mechanisms of action for HDAC5 in human erythropoiesis. These results may provide insights into understanding the anemia associated with HDAC inhibitor treatment.

αI-spectrin represents evolutionary optimization of spectrin for red blood cell deformability.

Spectrin tetramers of the membranes of enucleated mammalian erythrocytes play a critical role in red blood cell survival in circulation. One of the spectrins, αI, emerged in mammals with enucleated red cells after duplication of the ancestral α-spectrin gene common to all animals. The neofunctionalized αI-spectrin has moderate affinity for ßI-spectrin, whereas αII-spectrin, expressed in nonerythroid cells, retains ancestral characteristics and has a 10-fold higher affinity for ßI-spectrin. It has been hypothesized that this adaptation allows for rapid make and break of tetramers to accommodate membrane deformation. We have tested this hypothesis by generating mice with high-affinity spectrin tetramers formed by exchanging the site of tetramer formation in αI-spectrin (segments R0 and R1) for that of αII-spectrin. Erythrocytes with αIIßI presented normal hematologic parameters yet showed increased thermostability, and their membranes were significantly less deformable; under low shear forces, they displayed tumbling behavior rather than tank treading. The membrane skeleton is more stable with αIIßI and shows significantly less remodeling under deformation than red cell membranes of wild-type mice. These data demonstrate that spectrin tetramers undergo remodeling in intact erythrocytes and that this is required for the normal deformability of the erythrocyte membrane. We conclude that αI-spectrin represents evolutionary optimization of tetramer formation: neither higher-affinity tetramers (as shown here) nor lower affinity (as seen in hemolytic disease) can support the membrane properties required for effective tissue oxygenation in circulation.

Comprehensive phenotyping of erythropoiesis in human bone marrow: Evaluation of normal and ineffective erythropoiesis.

Identification of stage-specific erythroid cells is critical for studies of normal and disordered human erythropoiesis. While immunophenotypic strategies have previously been developed to identify cells at each stage of terminal erythroid differentiation, erythroid progenitors are currently defined very broadly. Refined strategies to identify and characterize BFU-E and CFU-E subsets are critically needed. To address this unmet need, a flow cytometry-based technique was developed that combines the established surface markers CD34 and CD36 with CD117, CD71, and CD105. This combination allowed for the separation of erythroid progenitor cells into four discrete populations along a continuum of progressive maturation, with increasing cell size and decreasing nuclear/cytoplasmic ratio, proliferative capacity and stem cell factor responsiveness. This strategy was validated in uncultured, primary erythroid cells isolated from bone marrow of healthy individuals. Functional colony assays of these progenitor populations revealed enrichment of BFU-E only in the earliest population, transitioning to cells yielding BFU-E and CFU-E, then CFU-E only. Utilizing CD34/CD105 and GPA/CD105 profiles, all four progenitor stages and all five stages of terminal erythroid differentiation could be identified. Applying this immunophenotyping strategy to primary bone marrow cells from patients with myelodysplastic syndrome, identified defects in erythroid progenitors and in terminal erythroid differentiation. This novel immunophenotyping technique will be a valuable tool for studies of normal and perturbed human erythropoiesis. It will allow for the discovery of stage-specific molecular and functional insights into normal erythropoiesis as well as for identification and characterization of stage-specific defects in inherited and acquired disorders of erythropoiesis.

Low Seroprevalence of SARS-CoV-2 in Rhode Island blood donors during may 2020 as determined using multiple serological assay formats.

BACKGROUND: Epidemic projections and public health policies addressing Coronavirus disease (COVID)-19 have been implemented without data reporting on the seroconversion of the population since scalable antibody testing has only recently become available. METHODS: We measured the percentage of severe acute respiratory syndrome- Coronavirus-2 (SARS-CoV-2) seropositive individuals from 2008 blood donors drawn in the state of Rhode Island (RI). We utilized multiple antibody testing platforms, including lateral flow immunoassays (LFAs), enzyme-linked immunosorbent assays (ELISAs) and high throughput serological assays (HTSAs). To estimate seroprevalence, we utilized the Bayesian statistical method to adjust for sensitivity and specificity of the commercial tests used. RESULTS: We report than an estimated seropositive rate of RI blood donors of approximately 0.6% existed in April-May of 2020. Daily new case rates peaked in RI in late April 2020. We found HTSAs and LFAs were positively correlated with ELISA assays to detect antibodies specific to SARS-CoV-2 in blood donors. CONCLUSIONS: These data imply that seroconversion, and thus infection, is likely not widespread within this population. We conclude that IgG LFAs and HTSAs are suitable to conduct seroprevalence assays in random populations. More studies will be needed using validated serological tests to improve the precision and report the kinetic progression of seroprevalence estimates.

Serological Assays Estimate Highly Variable SARS-CoV-2 Neutralizing Antibody Activity in Recovered COVID-19 Patients.

The development of neutralizing antibodies (NAbs) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) following infection or vaccination is likely to be critical for the development of sufficient population immunity to drive cessation of the coronavirus disease of 2019 (COVID-19) pandemic. A large number of serologic tests, platforms, and methodologies are being employed to determine seroprevalence in populations to select convalescent plasma samples for therapeutic trials and to guide policies about reopening. However, the tests have substantial variations in sensitivity and specificity, and their ability to quantitatively predict levels of NAbs is unknown. We collected 370 unique donors enrolled in the New York Blood Center Convalescent Plasma Program between April and May of 2020. We measured levels of antibodies in convalescent plasma samples using commercially available SARS-CoV-2 detection tests and in-house enzyme-linked immunosorbent assays (ELISAs) and correlated serological measurements with NAb activity measured using pseudotyped virus particles, which offer the most informative assessment of antiviral activity of patient sera against viral infection. Our data show that a large proportion of convalescent plasma samples have modest antibody levels and that commercially available tests have various degrees of accuracy in predicting NAb activity. We found that the Ortho anti-SARS-CoV-2 total Ig and IgG high-throughput serological assays (HTSAs) and the Abbott SARS-CoV-2 IgG assay quantify levels of antibodies that strongly correlate with the results of NAb assays and are consistent with gold standard ELISA results. These findings provide immediate clinical relevance to serology results that can be equated to NAb activity and could serve as a valuable roadmap to guide the choice and interpretation of serological tests for SARS-CoV-2.

TET2 deficiency leads to stem cell factor-dependent clonal expansion of dysfunctional erythroid progenitors.

Myelodysplastic syndromes (MDSs) are clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis. Anemia is the defining cytopenia of MDS patients, yet the molecular mechanisms for dyserythropoiesis in MDSs remain to be fully defined. Recent studies have revealed that heterozygous loss-of-function mutation of DNA dioxygenase TET2 is 1 of the most common mutations in MDSs and that TET2 deficiency disturbs erythroid differentiation. However, mechanistic insights into the role of TET2 on disordered erythropoiesis are not fully defined. Here, we show that TET2 deficiency leads initially to stem cell factor (SCF)-dependent hyperproliferation and impaired differentiation of human colony-forming unit-erythroid (CFU-E) cells, which were reversed by a c-Kit inhibitor. We further show that this was due to increased phosphorylation of c-Kit accompanied by decreased expression of phosphatase SHP-1, a negative regulator of c-Kit. At later stages, TET2 deficiency led to an accumulation of a progenitor population, which expressed surface markers characteristic of normal CFU-E cells but were functionally different. In contrast to normal CFU-E cells that require only erythropoietin (EPO) for proliferation, these abnormal progenitors required SCF and EPO and exhibited impaired differentiation. We termed this population of progenitors "marker CFU-E" cells. We further show that AXL expression was increased in marker CFU-E cells and that the increased AXL expression led to increased activation of AKT and ERK. Moreover, the altered proliferation and differentiation of marker CFU-E cells were partially rescued by an AXL inhibitor. Our findings document an important role for TET2 in erythropoiesis and have uncovered previously unknown mechanisms by which deficiency of TET2 contributes to ineffective erythropoiesis.

How did we rapidly implement a convalescent plasma program?

Since the beginning of the COVID-19 pandemic, the use of convalescent plasma as a possible treatment has been explored. Here we describe our experience as the first U.S. organization creating a COVID-19 convalescent plasma program to support its use through the single-patient emergency investigational new drug, the National Expanded Access Program, and multiple randomized controlled trials. Within weeks, we were able to distribute more than 8000 products, scale up collections to more than 4000 units per week, meet hospital demand, and support randomized controlled trials to evaluate the efficacy of convalescent plasma treatment. This was through strategic planning; redeployment of staff; and active engagement of hospital, community, and public health partners. Our partners helped with donor recruitment, testing, patient advocacy, and patient availability. The program will continue to evolve as we learn more about optimizing the product. Remaining issues to be resolved are antibody titers, dose, and at what stage of disease to transfuse.

Distinct roles for TET family proteins in regulating human erythropoiesis.

The ten-eleven translocation (TET) family of proteins plays important roles in a wide range of biological processes by oxidizing 5-methylcytosine (5mC) to 5-hydroxy-methylcytosine. However, their function in erythropoiesis has remained unclear. We show here that TET2 and TET3 but not TET1 are expressed in human erythroid cells, and we explore the role of these proteins in erythropoiesis. Knockdown experiments revealed that TET2 and TET3 have different functions. Suppression of TET3 expression in human CD34+ cells markedly impaired terminal erythroid differentiation, as reflected by increased apoptosis, the generation of bi/multinucleated polychromatic/orthochromatic erythroblasts, and impaired enucleation, although without effect on erythroid progenitors. In marked contrast, TET2 knockdown led to hyper-proliferation and impaired differentiation of erythroid progenitors. Surprisingly, knockdown of neither TET2 nor TET3 affected global levels of 5mC. Thus, our findings have identified distinct roles for TET2 and TET3 in human erythropoiesis, and provide new insights into their role in regulating human erythroid differentiation at distinct stages of development. Moreover, because knockdown of TET2 recapitulates certain features of erythroid development defects characteristic of myelodysplastic syndromes (MDSs), and the TET2 gene mutation is one of the most common mutations in MDS, our findings may be relevant for improved understanding of dyserythropoiesis of MDS.

Unexpected role for p19INK4d in posttranscriptional regulation of GATA1 and modulation of human terminal erythropoiesis.

Terminal erythroid differentiation is tightly coordinated with cell-cycle exit, which is regulated by cyclins, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors (CDKI), yet their roles in erythropoiesis remain to be fully defined. We show here that p19INK4d, a member of CDKI family, is abundantly expressed in erythroblasts and that p19INK4d knockdown delayed erythroid differentiation, inhibited cell growth, and led to increased apoptosis and generation of abnormally nucleated late-stage erythroblasts. Unexpectedly, p19INK4d knockdown did not affect cell cycle. Rather, it led to decreased expression of GATA1 protein. Importantly, the differentiation and nuclear defects were rescued by ectopic expression of GATA1. Because the GATA1 protein is protected by nuclear heat shock protein family (HSP) member HSP70, we examined the effects of p19INK4d knockdown on HSP70 and found that p19INK4d knockdown led to decreased expression of HSP70 and its nuclear localization. The reduced levels of HSP70 are the result of reduced extracellular signal-regulated kinase (ERK) activation. Further biochemical analysis revealed that p19INK4d directly binds to Raf kinase inhibitor PEBP1 and that p19INK4d knockdown increased the expression of PEBP1, which in turn led to reduced ERK activation. Thus we have identified an unexpected role for p19INK4d via a novel PEBP1-p-ERK-HSP70-GATA1 pathway. These findings are likely to have implications for improved understanding of disordered erythropoiesis.

Deubiquitylase USP7 regulates human terminal erythroid differentiation by stabilizing GATA1.

Ubiquitination is an enzymatic post-translational modification that affects protein fate. The ubiquitin-proteasome system (UPS) was first discovered in reticulocytes where it plays important roles in reticulocyte maturation. Recent studies have revealed that ubiquitination is a dynamic and reversible process and that deubiquitylases are capable of removing ubiquitin from their protein substrates. Given the fact that the UPS is highly active in reticulocytes, it is speculated that deubiquitylases may play important roles in erythropoiesis. Yet, the role of deubiquitylases in erythropoiesis remains largely unexplored. In the present study, we found that the expression of deubiquitylase USP7 is significantly increased during human terminal erythroid differentiation. We further showed that interfering with USP7 function, either by short hairpin RNA-mediated knockdown or USP7-specific inhibitors, impaired human terminal erythroid differentiation due to decreased GATA1 level and that restoration of GATA1 levels rescued the differentiation defect. Mechanistically, USP7 deficiency led to a decreased GATA1 protein level that could be reversed by proteasome inhibitors. Furthermore, USP7 interacts directly with GATA1 and catalyzes the removal of K48-linked poly ubiquitylation chains conjugated onto GATA1, thereby stabilizing GATA1 protein. Collectively, our findings have identified an important role of a deubiquitylase in human terminal erythroid differentiation by stabilizing GATA1, the master regulator of erythropoiesis.

Donor incentives improve cardiovascular disease risk profile and donation rates.

BACKGROUND: Blood centers may offer point-based reward systems or cardiovascular disease (CVD) screening to incentivize donors. However, combining these incentives to improve CVD risk and blood donation rates has not been studied. STUDY DESIGN AND METHODS: Study was a three-arm prospective controlled trial: Group 1, control (routine points, no CVD screening); Group 2, CVD screening with routine points; and Group 3, CVD screening plus incentive double points. The primary objective was to determine if double versus routine incentive points led to improvement or maintenance of CVD risk profile assessed using self-reported changes in 1) reading food labels for calorie and fat content, 2) exercising daily, 3) reduced fat intake, and 4) increase in eating fruits and vegetables. Outcomes were compared at first and final (2-year) follow-up visits. As secondary outcome, median blood donation rates before enrollment and during study were compared. RESULTS: A total of 570 donors (290 in Group 1, 134 in Group 2, 146 Group 3) were selected. At first follow-up visit, 71.4% in Group 3 versus 62.0% in Group 2 subjects reported at least one of four positive behavioral changes (p < 0.001). Increase in reading food labels for calorie and fat content was the most common change and higher in Group 3 (Group 3 from 60.9% to 79.1%; Group 2 from 67.6% to 77.5%; p < 0.001). Final evaluation showed significant increase in self-reported exercise in Group 3 only (from baseline 52.9% to 68.3%; p < 0.05). Group 3 reported higher increase in median number of donations/year during study enrollment (6.8 [IQR, 4.3-12] vs. baseline 4.6 [IQR, 3.2-7.1] donations/year) than Group 2 (5.6 [IQR, 4.2-10.5] vs. baseline 4.9 [IQR, 3.5-10.2]) and Group 1 (4.4 [IQR, 2.7-8.0] vs. baseline 4.4 [IQR, 2.5-6.0] donations/year; p < 0.001). CONCLUSION: Positive donor reinforcement (double vs. routine points) resulted in better self-reported health maintenance behavior and increased donation rates.

Ten years of TRALI mitigation: measuring our progress.

BACKGROUND: Transfusion-related acute lung injury (TRALI) is a leading cause of transfusion-associated mortality for which multiple mitigation strategies have been implemented over the past decade. However, product-specific TRALI rates have not been reported longitudinally and may help refine additional mitigation strategies. STUDY DESIGN AND METHODS: This retrospective multicenter study included analysis of TRALI rates from 2007 through 2017. Numerators included definite or probable TRALI reports from five blood centers serving nine states in the United States. Denominators were components distributed from participating centers. Rates were calculated as per 100,000 components distributed (p < 0.05 significant). RESULTS: One hundred four TRALI cases were reported from 10,012,707 components distributed (TRALI rate of 1.04 per 100,000 components). The TRALI rate was 2.25 for female versus 1.08 for male donated components (p < .001). The TRALI rate declined from 2.88 in 2007 to 0.60 in 2017. From 2007 to 2013, there was a significantly higher TRALI rate associated with female versus male plasma (33.85 vs. 1.59; p < 0.001) and RBCs (1.97 vs. 1.15; p = 0.03). From 2014 through 2017, after implementation of mitigation strategies, a significantly higher TRALI rate only from female-donated plateletpheresis continued to be observed (2.98 vs. 0.75; p = 0.04). CONCLUSION: Although the TRALI rates have substantially decreased secondary to multiple strategies over the past decade, a residual risk remains, particularly with female-donated plateletpheresis products. Additional tools that may further mitigate TRALI incidence include the use of buffy coat pooled platelets suspended in male donor plasma or platelet additive solution due to the lower amounts of residual plasma.

Impact of predictive scoring model and e-mail messages on African American blood donors.

BACKGROUND: Expanding the African American (AA) donor pool is critical to sustain transfusion support for sickle cell disease patients. STUDY DESIGN AND METHODS: The aims were to: 1) apply cognitive computing on donation related metrics to develop a predictive model that effectively identifies repeat AA donors, 2) determine whether a single e-mail communication could improve AA donor retention and compare retention results on higher versus lower predictive score donors, and 3) evaluate the effect of e-mail marketing on AA donor retention with culturally versus nonculturally tailored message. RESULTS: Between 2011 and 2012, 30,786 AA donors donated blood at least once on whom predictive repeat donor scores (PRDSs) was generated from donor-related metrics (frequency of donations, duration between donations, age, blood type, and sex). In 2013, 28% (8657/30,786) of 2011 to 2012 donors returned to donate on whom PRDS was validated. Returning blood donors had a higher mean PRDS compared to nonreturning donors (0.649 vs. 0.268; p < 0.001). In the e-mail pilot, high PRDS (≥0.6) compared to low PRDS (<0.6) was associated with 89% higher donor presentation rate (p < 0.001), 20% higher e-mail opening rate (p < 0.001), and, specifically among those who opened the e-mail, 159% higher presentation rate (p < 0.001). Finally, blood donation rate did not differ (p = 0.79) as a function of generic (n = 9312, 1.4%) versus culturally tailored (n = 9326, 1.3%) message. CONCLUSION: Computational algorithms utilizing readily available donor metrics can identify highly committed AA donors and in conjunction with targeted e-mail communication has the potential to increase the efficiency of donor marketing.

Spotlight on pathogenesis of TRALI: HNA-3a (CTL2) antibodies.

Human neutrophil antigen-3a (HNA-3a) antibodies contained in donor plasma can result in severe, sometimes fatal transfusion-related acute lung injury (TRALI). Recent developments in TRALI secondary to antibodies to HNA-3a antigen span diagnosis, pathophysiology, treatment, and prevention resulting in improved understanding, potential treatments, and mitigation strategies. First, on the molecular level, characterization of HNA-3 antigen has allowed for genotyping methods that clarify population prevalence. Related work has led to generation of multiple antibody detection assays. These assays aid in determining potential populations at risk and potential mitigation strategies. Second, the development of TRALI requires a hit from the patient and from the product. Anti- HNA-3a is one of the product-derived factors and appears to result in TRALI by binding directly to pulmonary endothelium as well as to neutrophils expressing the corresponding antigen. Finally, potential mitigation strategies include red blood cell product filtration to remove anti-HNA-3a as well as other antibodies.

Storage characteristics of multiple-donor pooled red blood cells compared to single-donor red blood cell units.

BACKGROUND: Each unit of blood donated is processed and stored individually resulting in variability in the amount of red blood cells (RBCs) collected, RBC properties, and the 24-hour posttransfusion RBC survivability. As a result, each unit differs in its ability to deliver oxygen and potentially its effects on the recipient. The goal of this study was to investigate the storage of pooled RBCs from multiple donors in comparison to control standard RBC units. STUDY DESIGN AND METHODS: Two units of irradiated, leukoreduced RBCs of same ABO, D, E, C, and K antigen phenotype were collected from each of five donors using apheresis. One unit from each donor was pooled in a 2-L bag and remaining units were used as controls. After being pooled, RBCs were separated in five bags and stored at 4°C along with the controls. Quality indexes were measured on Days 2, 14, and 28 for all the units. RESULTS: Adenosine triphosphate assays for both pooled and controls showed a slight decrease from Day 2 to Day 28 (pooled/control from 5.22/5.24 to 4.35/4.33 µmol/g hemoglobin [Hb]). 2,3-Diphosphoglycerate was successfully rejuvenated for all RBC units on Day 28 (pooled 11.46 µmol/g Hb; control 11.86 µmol/g Hb). The results showed a nonsignificant difference between pooled and control units, with a general trend of lower standard deviation for pooled units when compared to controls. CONCLUSION: Pooled units have reduced unit-to-unit variability. Future exploration of their immunogenicity is required before using pooled units for transfusion.

Riboflavin and ultraviolet light for pathogen reduction of murine cytomegalovirus in blood products.

BACKGROUND: Two studies were performed to test the effectiveness of riboflavin and ultraviolet (UV) light treatment (Mirasol PRT, Terumo BCT) against murine cytomegalovirus (MCMV). The first study utilized immune-compromised mice to measure the reduction of cell-free MCMV. A second study used a murine model to evaluate the ability of Mirasol PRT to prevent transfusion-transmitted (TT)-MCMV infection. STUDY DESIGN AND METHODS: Human plasma was inoculated with MCMV and then treated with Mirasol PRT. The viral titer was measured using an infectious dose 50% assay in nude mice. Mice were euthanized on Day 10 posttransfusion, and their spleens were tested for the presence of MCMV DNA using polymerase chain reaction (PCR). Mirasol PRT was also evaluated to determine its effectiveness in preventing TT-MCMV in platelets (PLTs) stored in PLT additive solution. PLTs were inoculated with either cell-associated MCMV or cell-free MCMV and then treated with Mirasol PRT. Mice were transfused with treated or untreated product and were euthanized 14 days posttransfusion. Blood and spleens were assayed for MCMV DNA by real-time-PCR. RESULTS: Using nude mice to titer MCMV, a modest 2.1-log reduction was observed in plasma products after Mirasol PRT treatment. TT-MCMV was not observed in the mouse transfusion model when either cell-free or cell-associated MCMV was treated with Mirasol PRT; MCMV transmission was uniformly observed in mice transfused with untreated PLTs. CONCLUSIONS: These results suggest that using riboflavin and UV light treatment may be able to reduce the occurrence of transmission of human CMV from infectious PLTs and plasma units.

Evaluating a program to increase blood donation among racial and ethnic minority communities in New York City.

BACKGROUND: Individuals with sickle cell disease (SCD), thalassemia, and leukemia often require frequent transfusion and run the risk of red blood cell (RBC) alloimmunization. To prevent alloimmunization or when alloimmunization is present, phenotype-matched and antigen-negative RBCs are transfused. To increase the probability of a phenotypic match, donors and recipients should share the same racial and/or ethnic background. Because the majority of patients with SCD are of African and Hispanic or Latino descent, a donor base of racial and ethnic minority donors providing an adequate supply of antigen-negative RBC units that can be phenotypically matched is required to meet the needs of frequently transfused patients. STUDY DESIGN AND METHODS: The New York Blood Center began the PreciseMatch program in 2005 to increase donations among African American and Hispanic/Latino donors by 150 incremental units per month. To evaluate the program, we conducted a systematic analysis of program documentation, focus group results, and collections data by race and ethnicity over time. RESULTS: The program achieved 75% of the operationalized goal of a 150-unit-per-month increase; 75% of donors were first-time donors, with deferral rates at new drives as high as 50%. Significant time and effort was involved in cultivating the community connections that facilitated new drives. CONCLUSIONS: Although PreciseMatch fell short of targets, it served as a foundation for relationships with diverse communities. Further research is needed to understand better how to increase minority donation using existing infrastructure and in the face of market pressures to collect blood as efficiently as possible.

The value of area-based analyses of donation patterns for recruitment strategies.

BACKGROUND: Lack of ready access to a donation site may be a potential barrier to or influence the frequency of blood donations. In this study, we applied geographic analysis to blood donor behavior and use of different donation sites. STUDY DESIGN AND METHODS: The study population consisted of blood donors who gave whole blood in Georgia between 2004 and 2008. Zip code, city, and county of donor's residence were matched with the addresses of their donation sites. Donors were dichotomized as either nonmetro Atlanta or metro Atlanta residents. Six donation site categories were defined: donation within the same or a different zip code, within the same or a different city, and within the same or a different county. Logistic regression was used to compare donations by zip code, city, and county. RESULTS: The study population consisted of 402,692 blood donors who donated 1,147,442 whole blood units between 2004 and 2008, more than half of whom (56.4%) resided in the metro Atlanta area. The majority of donors were white (75.0%) and female (55.7%). In nonmetro Atlanta, repeat donors were more likely to have donated at fixed sites (p < 0.001). In metro Atlanta, repeat donors were more likely to have donated at a mobile site than at a fixed site (p < 0.001). CONCLUSION: Geographic and demographic differences in blood donation patterns exist. The locations of donor residences and blood donation sites influence donor behaviors. Understanding the geographic influence on donation patterns provides an important tool for optimizing donor recruitment strategies.

Dose of prophylactic platelet transfusions and prevention of hemorrhage.

BACKGROUND: We conducted a trial of prophylactic platelet transfusions to evaluate the effect of platelet dose on bleeding in patients with hypoproliferative thrombocytopenia. METHODS: We randomly assigned hospitalized patients undergoing hematopoietic stem-cell transplantation or chemotherapy for hematologic cancers or solid tumors to receive prophylactic platelet transfusions at a low dose, a medium dose, or a high dose (1.1x10(11), 2.2x10(11), or 4.4x10(11) platelets per square meter of body-surface area, respectively), when morning platelet counts were 10,000 per cubic millimeter or lower. Clinical signs of bleeding were assessed daily. The primary end point was bleeding of grade 2 or higher (as defined on the basis of World Health Organization criteria). RESULTS: In the 1272 patients who received at least one platelet transfusion, the primary end point was observed in 71%, 69%, and 70% of the patients in the low-dose group, the medium-dose group, and the high-dose group, respectively (differences were not significant). The incidences of higher grades of bleeding, and other adverse events, were similar among the three groups. The median number of platelets transfused was significantly lower in the low-dose group (9.25x10(11)) than in the medium-dose group (11.25x10(11)) or the high-dose group (19.63x10(11)) (P=0.002 for low vs. medium, P<0.001 for high vs. low and high vs. medium), but the median number of platelet transfusions given was significantly higher in the low-dose group (five, vs. three in the medium-dose and three in the high-dose group; P<0.001 for low vs. medium and low vs. high). Bleeding occurred on 25% of the study days on which morning platelet counts were 5000 per cubic millimeter or lower, as compared with 17% of study days on which platelet counts were 6000 to 80,000 per cubic millimeter (P<0.001). CONCLUSIONS: Low doses of platelets administered as a prophylactic transfusion led to a decreased number of platelets transfused per patient but an increased number of transfusions given. At doses between 1.1x10(11) and 4.4x10(11) platelets per square meter, the number of platelets in the prophylactic transfusion had no effect on the incidence of bleeding. (ClinicalTrials.gov number, NCT00128713.)