Biomedical Excellence For Safer Transfusion Research Articles

Cryopreservation of Mesenchymal Stem/Stromal Cells using a DMSO-free Solution is Comparable to DMSO-containing Cryoprotectants: Results of An International Multicenter PACT/BEST Collaborative Study

Background and AimAn essential aspect of ensuring availability and stability of mesenchymal stem/stromal cells (MSCs) products for clinical use is that these cells are cryopreserved before individual infusion into patients. Currently, cryopreservation of MSCs involves use of a cryoprotectant solution containing dimethyl sulfoxide (DMSO). However, it is recognized that DMSO may be toxic for both the patient and the MSC product. In this Production Assistance for Cellular Therapies (PACT) and Biomedical Excellence for Safer Transfusion (BEST) Collaborative study, we compared a novel DMSO-free solution with DMSO containing cryoprotectant solutions for freezing MSCs. MethodsA DMSO-free cryoprotectant solution containing sucrose, glycerol, and isoleucine (SGI) in a base of Plasmalyte A was prepared at the University of Minnesota. Cryoprotectant solutions containing 5-10% DMSO (in-house) were prepared at seven participating centers (five from USA, one each from Australia and Germany). The MSCs were isolated from bone marrow or adipose tissue and cultured ex vivo per local protocols at each center. The cells in suspension were frozen by aliquoting into vials/bags. For six out of the seven centers, the vials/bags were placed in a controlled rate freezer (one center placed them at -80°C freezer overnight) before transferring to liquid nitrogen. The cells were kept frozen for at least one week before thawing and testing. Pre- and post-thaw assessment included cell viability and recovery, immunophenotype as well as transcriptional and gene expression profiles. Linear regression, mixed effects models and two-sided t-tests were applied for statistical analysis. ResultsMSCs had an average viability of 94.3% (95% CI, 87.2-100%) before cryopreservation, decreasing by 4.5% (95% CI: 0.03-9.0%; p: 0.049) and 11.4% (95% CI: 6.9-15.8%; p<0.001), for MSCs cryopreserved in the in-house and SGI solutions, respectively. The average recovery of viable MSCs cryopreserved in the SGI was 92.9% (95% CI: 85.7-100.0%), and it was lower by 5.6% (95% CI: 1.3-9.8%, p<0.013) for the in-house solution. Additionally, MSCs cryopreserved in the two solutions had expected level of expressions for CD45, CD73, CD90 and CD105 with no significant difference in global gene expression profiles. ConclusionMSCs cryopreserved in a DMSO-free solution containing sucrose, glycerol, and isoleucine in a base of Plasmalyte A had slightly lower cell viability, better recovery, and comparable immunophenotype and global gene expression profiles compared to MSCs cryopreserved in DMSO containing solutions. The average viability of MSCs in the novel solution was above 80% and, thus, likely clinically acceptable. Future studies are suggested to test the post-thaw functions of MSCs cryopreserved in the novel DMSO-free solution.

A global analysis of the use of immunoglobulin, shortages in supply, and mitigating measures: A survey of hospital providers (a BEST Collaborative study).

Immunoglobulin (IG) therapy is widely used to treat primary and secondary immune deficiencies and as immunomodulatory agent for various disorders. There is great concern that shortages of IG may rise, potentially affecting medical treatment options. An international survey was developed to study how intravenous immunoglobulins (IVIGs) are used and managed within hospitals in case of shortages. Study data were collected and managed using REDCap electronic data capture tools hosted by the Biomedical Excellence for Safer Transfusion (BEST) Collaborative. The survey was directed to hospital pharmacists and blood bank transfusion professionals and disseminated through members of the BEST Collaborative network. Survey respondents from institutions in the USA, Canada, Europe, Japan, and Australia (n = 13) confirmed that the primary specialties utilizing IG are neurology, hematology, and immunology. More than 60% of respondents reported IG supply shortages, but mitigation strategies were not well developed. As IG is the leading driver in plasma demand, more studies are needed to understand current and future demand for IG from the clinical perspective. Necessity lies in establishing clinical guidance to address shortages.

Local manufacturing processes contribute to variability in human mesenchymal stromal cell expansion while growth media supplements contribute to variability in gene expression and cell function: a Biomedical Excellence for Safer Transfusion (BEST) collaborative study

Background aimsCulture-derived mesenchymal stromal cells (MSCs) exhibit variable characteristics when manufactured using different methods, source material and culture media. The purpose of this multicenter study was to assess the impact on MSC expansion, gene expression and other characteristics when different laboratories expanded MSCs from cultures initiated with bone marrow−MSC aliquots derived from the same donor source material yet with different growth media. MethodsEight centers expanded MSCs using four human platelet lysate (HPL) and one fetal bovine serum (FBS) products as media supplements. The expanded cells were taken through two passages then assessed for cell count, viability, doubling time, immunophenotype, cell function, immunosuppression and gene expression. Results were analyzed by growth media and by center. ResultsCenter methodologies varied by their local seeding density, feeding regimen, inoculation density, base media and other growth media features (antibiotics, glutamine, serum). Doubling times were more dependent on center than on media supplements. Two centers had appropriate immunophenotyping showing all MSC cultures were positive for CD105, CD73, CD90 and negative for CD34, CD45, CD14, HLA-DR. MSCs cultured in media supplemented with FBS compared with HPL featured greater T-cell inhibition potential. Gene expression analysis showed greater impact of the type of media supplement (HPL versus FBS) than the manufacturing center. Specifically, nine genes were decreased in expression and six increased when combining the four HPL-grown MSCs versus FBS (false discovery rate [FDR] <0.01), however, without significant difference between different sources of HPL (FDR <0.01). ConclusionsLocal manufacturing process plays a critical role in MSC expansion while growth media may influence function and gene expression. All HPL and FBS products supported cell growth.

Multicenter evaluation of the IL-3-pSTAT5 assay to assess the potency of cryopreserved stem cells from cord blood units: The BEST Collaborative study.

The IL-3-pSTAT5 assay, a new, rapid, and standardized flow-cytometry-based assay may compensate for several limitations of the colony-forming unit (CFU) assay typically used for stem cell potency assessments of cord blood units (CBU). We performed an inter-laboratory evaluation of the performance of this new assay. This Biomedical Excellence for Safer Transfusion (BEST) Collaborative multicenter, international study included 15 participants from public cord blood banks (CBBs), CBB-supporting research laboratories, and stem cell laboratories. To perform the IL-3-pSTAT5 assay, participating centers received reagents, instructions, and 10 blind CBU samples, including eight normal samples and two samples exposed to a transient warming event. We measured inter-laboratory agreement qualitatively (proportion of correctly classified samples) and quantitatively (coefficient of variation [CV], correlation coefficients, receiver operating characteristics (ROC) curve, and intraclass correlation coefficient [ICC]). The qualitative agreement was 97.3% (i.e., 107/110; Fleiss' kappa=0.835). The average CV on a per-sample basis was 11.57% among all samples, 8.99% among normal samples, and on a per-center basis was 9.42% among normal samples. In a correlation matrix that compared results across centers, the mean Pearson's correlation coefficient was 0.88 (standard deviation=0.04). The ICC was 0.83 (95% confidence interval=0.68-0.95). The area under the curve (AUC) from the ROC curve was 0.9974. Excellent qualitative and quantitative agreement was exhibited across laboratories. The IL-3-pSTAT5 assay may therefore be implemented in flow cytometry laboratories to rapidly and reliably provide standardized measures of stem cell potency in CBUs.

Whole blood donor return rates after deferral for tattooing or body piercing—Survey across blood donation services: The BEST collaborative study

To protect transfusion recipients from transfusion-transmissible infections, blood donors are deferred from donating after recent tattooing or piercing. To explore to what extent and how this deferral impacts donor availability, we performed an international study to investigate how many donors were deferred for a recent tattoo or piercing and how many of these donors returned to donate. We surveyed blood centre members of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative and the European Blood Alliance Donor Studies Working Group on their numbers of donations, tattoo and piercing deferrals, and return rates in the year 2017. Eight blood centres participated. Overall, deferral rates were lower for repeat donors compared to new donors. Repeat donors were more likely to return than new donors. Women and young donors were more often deferred than male and older donors. Men were more demotivated by tattoo or piercing deferral, resulting in lower return rates compared to women. Return rates differed greatly between blood centres. Tattoo and piercing deferrals lead to missed donations and result in lower return rates. However, the numbers vary largely internationally, probably due to cultural and policy differences. Shortening deferral periods after tattooing or piercing may reduce the impact on donor availability, which should be investigated in single-centre studies.

Blood donor eligibility criteria for medical conditions: A BEST collaborative study.

Donor eligibility questions and criteria for medical conditions vary between blood centres, suggesting that they are based more on local regulations or experience, rather than on published data, which are limited. As the donor population ages, medical conditions become more common. We assessed donor health assessment criteria at blood centre members of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative. Our aim was to compare eligibility criteria and determine their underlying basis. A REDCap survey was sent to blood centre participants, based on medical conditions of greatest interest suggested by the Donor Studies Team of the BEST Collaborative. Participants were asked about current donor health assessment questions, deferral criteria and the basis for their deferral policy (donor risk, recipient risk or both) for 20 medical conditions. Complete responses were received from 26 blood donor centres (24 separate responses) representing a combination of hospital-based centres, large regional centres and community/national blood centres in 14 different countries. Most centres specifically ask about heart and lung conditions, whereas fewer than half inquire about kidney, gastrointestinal or neurological conditions. North American blood centres tended to be less restrictive, while regulatory restrictions are more prevalent in Europe. Most participants felt that the criteria were based on regulatory requirements or experience, rather than on published data. There is considerable variability in criteria by region. Ideally, criteria would be more evidence-based rather than based on regulatory requirements or experience. Deferral criteria must balance donor and recipient safety and maintain an adequate blood supply.

Application of Standardized Residual Component Culture Criteria for Suspected Septic Transfusion Reactions Would Increase the Component Culturing Rate at a Single Academic Medical Center

The 2019 SCARED study developed the Biomedical Excellence for Safer Transfusion (BEST) criteria in an effort to standardize the decision to culture residual units in the context of suspected septic transfusion reactions (STRs). The goal of this study was to apply the BEST criteria to determine the effect on the transfusion reaction decision to culture. This retrospective, single-center, cross-sectional study assessed adult transfusion reactions identified in calendar years 2013 to 2020. Reactions following transfusion of RBCs, platelets, and plasma were included, and the decisions to culture following strict application of BEST criteria were compared with decisions to culture in actual practice. In total, 1,068 transfusion reactions were reported and 200 (19%) suspected STRs were cultured, all with negative results; 303 (28%) reactions would have been cultured per strict application of the BEST criteria. Concordance between actual culture decision and BEST criteria recommendation was 62% for cultured components and 79% for components that were not cultured. BEST criteria provide objective recommendations of when to culture residual units implicated in suspected STRs, but strict application of these criteria may result in increased culture rates. Clinical correlation to aid in the decision to culture is recommended.

Human Mesenchymal Stromal Cell (MSC) Characteristics Vary Among Laboratories When Manufactured From the Same Source Material: A Report by the Cellular Therapy Team of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative.

BackgroundCulture-derived mesenchymal stromal cells (MSCs) exhibit variable characteristics when manufactured using different methods and different source materials. The purpose of this study was to assess the impact on MSC characteristics when different laboratories propagated MSCs from cultures initiated with BM aliquots derived from the same donor source material.Methods and MethodsFive aliquots from each of three different BM donors were distributed to five independent laboratories. Three laboratories plated whole BM and two laboratories a mononuclear BM cell fraction. Four laboratories cultured in media supplemented with fetal bovine serum (FBS) and one laboratory used human platelet lysate (hPL). Initial cell seeding densities (i.e., P0) ranged from 19.7 × 103/cm2–282 × 103/cm2 and for second seeding (i.e., P1) 0.05 × 103–5.1 × 103 cells/cm2. Post-thawed MSCs from each laboratory were analyzed for cell viability, immunophenotype, tri-lineage differentiation, fibroblast colony-forming units (CFU-F), gene expression, and immunosuppressive activity.ResultsTransit times from BM collection to receipt by laboratories located in the United States ranged from 16.0–30.0 h and from 41.5–71.5 h for a laboratory in Asia. Post-thaw culture derived MSCs rom BM #1, #2, and #3 exhibited viabilities that ranged from 74–92%, 61–96%, and 23–90%, respectively. CFU activity from BM #1, #2, and #3 per 200 MSCs plated averaged 45.1 ± 21.4, 49.3 ± 26.8 and 14.9 ± 13.3, respectively. No substantial differences were observed in immunophenotype, and immunosuppressive activities. Global gene expression profiles of MSCs revealed transcriptome differences due to different inter-laboratory methods and to donor source material with the center effects showing greater molecular differences than source material.ConclusionFunctional and molecular differences exist among MSCs produced by different centers even when the same BM starting material is used to initiate cultures. These results indicated that manufacturing of MSCs by five independent centers contributed more to MSC variability than did the source material of the BM used in this study. Thus, emphasizing the importance of establishing worldwide standards to propagate MSCs for clinical use.

Variations in hemoglobin measurement and eligibility criteria across blood donation services are associated with differing low‐hemoglobin deferral rates: a BEST Collaborative study

Determination of blood donor hemoglobin (Hb) levels is a pre-requisite to ensure donor safety and blood product quality. We aimed to identify Hb measurement practices across blood donation services and to what extent differences associate with low-Hb deferral rates. An online survey was performed among Biomedical Excellence for Safer Transfusion (BEST) Collaborative members, extended with published data. Multivariable negative-binomial regression models were built to estimate adjusted associations of minimum donation intervals, Hb cut-offs (high, ≥13.5 g/dL in men or ≥ 12.5 g/dL in women, vs. lower values), iron monitoring (yes/no), providing or prescribing iron supplementation (yes/no), post-versus pre-donation Hb measurement and geographical location (Asian vs. rest), with low-Hb deferral rates. Data were included from 38 blood services. Low-Hb deferral rates varied from 0.11% to 8.81% among men and 0.84% to 31.85% among women. Services with longer minimum donation intervals had significantly lower deferral rates among both women (rate ratio, RR 0.53, 95%CI 0.33-0.84) and men (RR 0.53, 95%CI 0.31-0.90). In women, iron supplementation was associated with lower Hb deferral rates (RR 0.47, 95%CI 0.23-0.94). Finally, being located in Asia was associated with higher low-Hb deferral rates; RR 9.10 (95%CI 3.89-21.27) for women and 6.76 (95%CI 2.45-18.68) for men. Differences in Hb measurement and eligibility criteria, particularly longer donation intervals and iron supplementation in women, are associated with variations in low-Hb deferral rates. These insights could help improve both blood donation service efficiency and donor care.

The impact of age and sex on first-time donor return behavior.

This study examined the impact of age and sex of first-time donors who had not experienced an adverse event or deferral on their likelihood of and time to return. On behalf of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative, international blood collection agencies (BCAs) were invited to provide data on first-time whole blood donors in 2014, including initial presentation date, collection site type, age, sex, blood type, return to donate within 24 months (yes/no), and subsequent presentation date. Eight BCAs contributed 706,789 records. The overall odds of returning to donate were slightly lower in female versus male donors, and the overall age trend was U-shaped with younger and older donors having higher odds for returning relative to middle-aged donors. However, variations by BCA were observed. Specifically, in three BCAs, women had higher odds of returning to donate than men. Further, while across seven BCAs the smallest cohort of older first-time donors returned at a higher rate and returned more quickly than middle-aged first-time donors, the behavior of younger donors varied substantially between BCAs. While older first-time donors are more likely to return and return more quickly than middle-aged donors they make up only a small proportion of first-time donors, whereas the larger group of younger donors exhibits less clear patterns of return compared to middle-aged donors. Further research is needed to determine whether targeting the recruitment of older donors or bolstering retention of middle-aged donors would be most effective in maintaining the blood supply.

Current practices for viability testing of cryopreserved cord blood products: an international survey by the cellular therapy team of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative.

Viability testing is a common practice in laboratories. The goal of this study was to ascertain current laboratory practices internationally for performing viability testing for cryopreserved cord blood (CB) products and glean information about how to standardize the method to improve interlaboratory reproducibility. A survey to evaluate current laboratory practices for viability testing was designed and distributed internationally. The question topics included sampling and testing methods, responses to unexpected results, and the rating of the reliability of the CB quality tests, together with expectations for standardization. There were 32 respondents to the survey, of whom 28 responded to the more detailed questionnaire about viability methods. Overall, responses indicated that various stains were used among the laboratories, and when multiple sites used the same viability stain the methods differed. The majority of the respondents were in favor of standardizing the viability testing methods. A wide variety of preferences were communicated about how to standardize the method, but a majority did advocate the use of 7-aminoactinomycin D (7-AAD) with flow cytometry. The survey results revealed a variety of tests and inconsistent interlaboratory practices for performing the viability assay. Flow cytometry with a 7-AAD dye was suggested as a first step toward standardization.

Assessment of the knowledge and perceptions of Brazilian medical residents on transfusion medicine

BackgroundMedical education has a major social impact because it ultimately influences the quality and safety of the health service offered to the population. Several studies have acknowledged the relationship between medical knowledge on transfusion medicine and the proper use of this therapy. The rational use of blood has become a worldwide concern. In this context, the aim of the present study was to evaluate medical residents’ knowledge on transfusion medicine in hospitals located in Rio de Janeiro, Brazil, and their training during their medical education. MethodsOne hundred and six residents from eight medical specialties of four hospitals participated. A questionnaire developed by the Biomedical Excellence for Safer Transfusion (BEST) group was applied to evaluate the participants’ medical knowledge. Another questionnaire was also applied to evaluate participant profiles regarding frequency of transfusion prescriptions, self-perceived knowledge and relevance of the subject. ResultsThe mean number of correct answers to the questionnaire on knowledge about transfusion practices was 43.5% (range: 15–80%). A relationship between training during medical residency and the obtained result was observed (p-value=0.0007). Most residents (73%) did not receive training in transfusion medicine during their graduation or residency and 93% would like to receive additional training. ConclusionA clear deficit in the knowledge of transfusion medicine was verified, indicating the need for change in the teaching of this specialty. This result is similar to some developed countries.

A multicentred study to validate a consensus bleeding assessment tool developed by the biomedical excellence for safer transfusion collaborative for use in patients with haematological malignancy.

There continues to be uncertainty about the optimal approach to documenting bleeding data in platelet transfusion trials, with a desire to apply a common assessment tool across all trials. With this in mind, a consensus bleeding assessment tool (BAT) has been developed by the Biomedical Excellence for Safer Transfusion (BEST) collaborative, based on review of data collection forms used in published randomized trials and following content validation with a range of healthcare professionals at seven haematology centres through BEST members. This study aimed to evaluate reliability and reproducibility of the consensus BAT. Replicated clinical assessments of bleeding were undertaken by participants with haematological malignancies recruited at four haematology centres in an international, multicentred, observational study. Concordance of repeat assessments was calculated for agreement in site and grade of bleeding observed. Forty patients consented to participate, and 13 trained bleeding assessors collected these data. Bleeding assessments were carried out on 113 separate days. Of all 225 bleeding assessments, 204 were compared for grade concordance, and 160 were compared for site concordance. There was very good grade concordance (83%, 95% confidence interval 74-93%) and good bleeding site concordance (69%, 95% confidence interval 57-79%) in observations of bleeding. Discordance was primarily in relation to assessing skin bleeding. Alongside a structured training programme, levels of concordance for a consensus BAT were high. Researchers using assessment tools for bleeding need to balance comprehensive data collection against potential loss of accuracy for some types of bleeding, such as skin findings.

Comparison of donor and general population demographics over time: a BEST Collaborative group study.

We compared donor and general population demographics over time to provide insight into current donation patterns and the future adequacy of the blood supply. Seventeen blood center members of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative from 12 countries provided the number of donors and people in the general population by demographic category for 2001 and 2011, changes in age criteria, and percentage of first-time donors. We calculated the median age of donors and the general population and determined the percentage of each group in age and sex cohorts. Age criteria vary, with upper limits recently liberalized in several countries. In 2011, the percentage of first-time donors ranged from 10% to 41%. The median age of the donor and general population increased from 2001 to 2011 in most countries, as did the percentage of the general population over 60. The youngest donor cohort is overrepresented to a variable degree; this tendency increased over time. Although still underrepresented, older donors contributed more in 2011. A large middle-aged cohort is aging at a rate exceeding the progression of time, while 25- to 45-year-olds are relatively underrepresented. All participating countries are experiencing aging of their general population. Donor demographics differ substantially between countries; this can be only partly explained by population demographics and age criteria. Many countries have an aging middle-aged donor and population cohort and are increasingly relying on their youngest donors to contribute disproportionately to the blood supply.

Prothrombin complex concentrate for emergent reversal of warfarin: an international survey of hospital protocols.

For emergent warfarin reversal, four-factor prothrombin complex concentrates (4FPCCs) are recommended by many international guidelines. We surveyed international clinical sites including members of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative. Most sites have emergent warfarin reversal protocols (53% use PCC, 25% use PCC+ plasma and 2% use plasma alone); however, variation between adjusted dosing and fixed dosing was observed.

An international survey on the role of the hospital transfusion committee.

Hospital transfusion committees (HTCs) can oversee all aspects of transfusion practice at a hospital. This survey sought to identify which quality variables were being reported at HTCs around the world. A working party composed of members of the Biomedical Excellence for Safer Transfusion (BEST) collaborative developed a survey of quality variables that could be potentially presented at HTC meetings. The survey was electronically sent to all BEST members who were encouraged to complete it if they were active on an HTC and to send it to other colleagues with similar experience. An expert panel was convened to determine which quality variables are the most important for review at HTC meetings. There were 121 respondents; the majority were from Europe (52%), Asia (19%), or North America (19%). Most respondents (68%) were at university hospitals. Of the 117 (97%) respondents with an HTC, the committee most often met quarterly (42%) and reviewed transfusion reactions (79%) and risk management-reported events (52%). The HTCs most commonly included transfusion medicine physicians, anesthesiologists, and other physicians who regularly transfuse blood products. Some of the most commonly reported quality variables included number of blood products transfused, wasted, and expired and the number of improperly labeled specimens. The expert panel analysis revealed that some variables that were deemed important were not being frequently reported at HTCs. There is variability in the variables being reported at HTCs around the world with some important variables not frequently reported.

Design and Validation of a Survey Questionnaire for the Assessment of Physician Transfusion Medicine Knowledge

The clinical appropriateness of blood transfusion rests with physicians having the correct knowledge when making decisions concerning the selection and transfusion of blood and blood components. Transfusion medicine knowledge (TMK) is typically acquired during medical school, as on-the-job training during residency or fellowship, and through continuing education (e.g., online, seminars, workshops) in practice. To obtain a baseline of TMK in physicians at Augusta University Medical Center (AUMC) (formerly Georgia Regents University Medical Center), a multiple-choice survey questionnaire was designed and validated using input from individuals with expertise in the field of transfusion medicine. The survey was then deployed to physicians at AUMC. Analysis of survey response data showed that there were significant knowledge gaps among physicians that varied among specialty groups as well as with the number of formal education hours that physicians received as part of their medical training and in practice. The purpose of this research study was to establish the baseline TMK level of physicians at AUMC. ABBREVIATIONS:TMK - Transfusion Medicine Knowledge, AUMC - Augusta University Medical Center -, ARC - American Red Cross, CMS – Centers for Medicare & Medicaid Services, CFR - Code of Federal Regulations, ASCP - American Society for Clinical Pathology, BEST - Biomedical Excellence for Safer Transfusion, TJC - The Joint Commission, PBM - Patient Blood Management

BEST-TEST2: assessment of hematology trainee knowledge of transfusion medicine.

As transfusion is a common therapy and key component in every hematologist's practice, hematology training programs should dedicate significant time and effort to delivering high-quality transfusion medicine education to their trainees. The current state of hematology trainee knowledge of transfusion medicine is not known. A validated assessment tool developed by the Biomedical Excellence for Safer Transfusion (BEST) Collaborative was used to assess prior transfusion medicine education, attitudes, perceived ability, and transfusion medicine knowledge of hematology trainees. A total of 149 hematology trainees at 17 international sites were assessed. The overall mean exam score was 61.6% (standard deviation, 13.4%; range, 30%-100%) with no correlation in exam scores with postgraduate year or previous transfusion medicine education in medical school or internal medicine residency. However, better scores correlated with 3 or more hours of transfusion medicine education (p = 0.0003) and perceived higher-quality education during hematology training (p = 0.03). Hematology trainees at US sites, where hematology is often combined with oncology training, had statistically lower scores than trainees at non-US sites (56.2% vs. 67.4%; p < 0.0001). In terms of topic areas, although 93% of participants had obtained consent for transfusion, the lowest scores were on transfusion reaction-related questions. Given the overall poor performance, this study serves as an impetus for all hematology training programs to reevaluate the quality and quantity of transfusion medicine training and can assist in the development of targeted curricula.

Sqstm1 Is Required to Retain Hematopoietic Stem Cell/ Progenitors As a Negative Regulator of Macrophage-Dependent Inflammatory Signaling in the Bone Marrow Osteoblastic Niche

In the bone marrow (BM), hematopoietic stem cells and progenitors (HSC/P) reside in specific anatomical niches. Among these niches, a functional osteoblast (Ob)-macrophage (MΦ) niche has been described where Ob and MΦ (so called “osteomacs”) are in direct relationship. A connection between innate immunity surveillance and traffic of hematopoietic stem cells/progenitors (HSC/P) has been demonstrated but the regulatory signals that instruct immune regulation from MΦ and Ob on HSC/P circulation are unknown. The adaptor protein sequestosome 1 (Sqstm1), contains a Phox bemp1 (PB1) domain which regulates signal specificities through PB1-PB1 scaffolding and processes of autophagy. Using microenvironment and osteoblast-specific mice deficient in Sqstm1, we discovered that the deficiency of Sqstm1 results in macrophage contact-dependent activation of Ob IKK/NF-κB, in vitro and in vivo repression of Ccl4 (a CCR5 binding chemokine that has been shown to modulate microenvironment Cxcl12-mediated responses of HSC/P), HSC/P egress and deficient BM homing of wild-type HSC/P. Interestingly, while Ccl4 expression is practically undetectable in wild-type or Sqstm1-/- Ob, primary Ob co-cultured with wild-type BM-derived MΦ strongly upregulate Ccl4 expression, which returns to normal levels upon genetic deletion of Ob Sqstm1. We discovered that MΦ can activate an inflammatory pathway in wild-type Ob which include upregulation of activated focal adhesion kinase (p-FAK), IκB kinase (IKK), nuclear factor (NF)-κB and Ccl4 expression through direct cell-to-cell interaction. Sqstm1-/- Ob cocultured with MΦ strongly upregulated p-IKBα and NF-κB activity, downregulated Ccl4 expression and secretion and repressed osteogenesis. Forced expression of Sqstm1, but not of an oligomerization-deficient mutant, in Sqstm1-/- Ob restored normal levels of p-IKBα, NF-κB activity, Ccl4 expression and osteogenic differentiation, indicating that Sqstm1 dependent Ccl4 expression depends on localization to the autophagosome formation site. Finally, Ob Sqstm1 deficiency results in upregulation of Nbr1, a protein containing a PB1 interacting domain. Combined deficiency of Sqstm1 and Nbr1 rescues all in vivo and in vitro phenotypes of Sqstm1 deficiency related to osteogenesis and HSC/P egression in vivo. Together, this data indicated that Sqstm1 oligomerization and functional repression of its PB1 binding partner Nbr1 are required for Ob dependent Ccl4 production and HSC/P retention, resulting in a functional signaling network affecting at least three cell types. A functional 'MΦ-Ob niche’ is required for HSC/P retention where Ob Sqstm1 is a negative regulator of MΦ dependent Ob NF-κB activation, Ob differentiation and BM HSC/P traffic to circulation. DisclosuresStarczynowski:Celgene: Research Funding. Cancelas:Cerus Co: Research Funding; P2D Inc: Employment; Terumo BCT: Research Funding; Haemonetics Inc: Research Funding; MacoPharma LLC: Research Funding; Therapure Inc.: Consultancy, Research Funding; Biomedical Excellence for Safer Transfusion: Research Funding; New Health Sciences Inc: Consultancy.

Development of a validated exam to assess physician transfusion medicine knowledge

There is evidence that physicians lack adequate transfusion medicine knowledge. To design needs-based educational interventions to address this gap, a validated assessment tool is required. Previously published exams have not been created or validated using rigorous psychometric methods. A modified Delphi method was used to achieve consensus regarding the essential knowledge and skills for physicians who transfuse blood products. To ensure content validity, members of an international organization of transfusion medicine experts (Biomedical Excellence for Safer Transfusion [BEST] Collaborative) participated in the exam design process. An exam, based on the most highly rated topics, was created and administered to individuals with a priori expected basic, intermediate, and expert levels of transfusion medicine knowledge. Rasch analysis, a psychometric technique used in high-stakes medical licensure and board testing, was used to determine exam accuracy and precision. Thirty-six topics achieved ratings sufficient to be considered for inclusion in the exam (content validity index > 0.8). A 23-question exam was administered to 49 individuals. Mean scores for individuals with expected basic, intermediate, and expert knowledge were 42, 62, and 82%, respectively (p < 0.0001). The exam achieved good fit with the Rasch model. A validated exam has now been created to accurately assess transfusion medicine knowledge. This exam can be used to determine knowledge deficits and assist in the design of curricula to improve blood product utilization.