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The standard cryoprotectant for human cellular products is dimethyl sulfoxide (DMSO), which is associated with hematopoietic cell infusion-related adverse events (HCI-AEs) in hematopoietic stem cell transplantation including peripheral blood stem cell (PBSC) transplantation (PBSCT). DMSO is often used with hydroxyethyl starch (HES), which reduces DMSO concentration while maintaining the postthaw cell recovery. The cryoprotectant medium CP-1 (Kyokuto Pharmaceutical Industrial) is widely used in Japan. After mixture of a product with CP-1, DMSO and HES concentrations are 5% and 6%, respectively. However, the safety profile of CP-1 in association with HCI-AEs has not been investigated.To compare CP-1 with other cryoprotectants, we conducted a subgroup analysis of PBSCT recipients in a prospective surveillance study for HCI-AEs. Moreover, we validated the toxicity of CP-1 in 90 rats following various dose administration.The PBSC products cryopreserved with CP-1 (CP-1 group) and those with other cryoprotectants, mainly 10% DMSO (non-CP-1 group), were infused into 418 and 58 recipients, respectively. The rate of ≥grade 2 HCI-AEs was higher in the CP-1 group, but that of overall or ≥grade 3 HCI-AEs was not significantly different, compared to the non-CP-1 group. Similarly, after propensity score matching, ≥grade 2 HCI-AEs were more frequent in the CP-1 group, but the ≥grade 3 HCI-AE rate did not differ significantly between the groups. No significant toxicity was detected regardless of the CP-1 dose in the 90 rats.Infusion of a CP-1-containing PBSC product is feasible with the respect of HCI-AEs.

The objectives of this study were to determine the longer-term trends in childhood obesity and glucose metabolism abnormalities among residents of Fukushima Prefecture 5 years after the Great East Japan Earthquake.We evaluated the changes for height, weight, body mass index (BMI), BMI SD score, fasting plasma glucose (FPG) concentration, and hemoglobin A1c (HbA1c) in elementary and junior high school residents who had lived in the evacuation zone between 2011 and 2015.1) Of the Residents, 11,112 received health checks, while in 2012, 2013, 2014 and 2015, 5,737, 4,522, 4,297 and 3,405 received health checks, respectively. 2) The mean BMI SD score for all participants in 2011 was 0.149, and this score gradually decreased from 2011 to 2015. 3) FPG levels and HbA1c levels for all participants with a BMI value≧+2SD in 2011 were higher than those in residents with a BMI value+2SD. 4) The frequency of participants with a FPG level ≧126 mg/dl and the frequency of participants with a HbA1c level ≧6.5 % in 2011 were higher than those in 2012, 2013, and 2015.These results suggest that a number of pediatric residents suffered from obesity and glucose metabolism abnormalities. Furthermore, the longer-term observations indicated an improvement in obesity and glucose metabolism abnormalities. There was a strong association observed between obesity and glucose metabolism, thus, it is considered important to continue with health checks for children with obesity and strive to improve their health.

Serologic RhD-negative red cells can cause anti-D alloimmunization if they carry the Asian-type DEL or other DEL variants.

Background Haploidentical hematopoietic stem cell transplantation (HSCT) is a useful therapy for relapsed/refractory acute leukemia or lymphoma because of the strong graft-versus-leukemia (GvL) effect. However, it is often accompanied by severe acute graft-versus-host disease (GvHD), which is the most serious complication after haploidentical HSCT. Thus, it is important to control the severity of acute GvHD while maintaining the GvL effect. In our experiences of pediatric haploidentical HSCT, it takes several days for acute GvHD to become severe after the appearance of initial symptoms, mostly skin rashes. In this study, we aimed to identify useful biomarkers at the onset of acute GvHD that predict subsequent development of severe acute GvHD. Methods Forty-five consecutive children with relapsed/refractory acute leukemia or lymphoma who developed acute GvHD after haploidentical HSCT were enrolled. We analyzed possible biomarkers from samples collected at the onset of acute GvHD. Results Nineteen patients developed grade 1-2 acute GvHD, and 26 patients developed grade 3-4 acute GvHD. There was no significant difference in patient characteristics between the two groups. Transplant-related mortality occurred only in the grade 3-4 acute GvHD group (34.5%). Multivariate analysis revealed that serum albumin was an independent biomarker for predicting the severity of acute GvHD (p=0.009). The area under the receiver operating characteristic curve of serum albumin was 0.864. Conclusions The serum albumin level at the onset of acute GvHD could be a useful biomarker for the development of subsequent severe acute GvHD in pediatric patients after haploidentical HSCT.

Background and objectives Anti-CD38 monoclonal antibodies, including daratumumab and isatuximab, often interfere with pretransfusion testing. Dithiothreitol (DTT) treatment of red blood cells (RBCs) negates this interference. However, the optimum DTT concentration and treatment time have not been well defined. Here, we quantified CD38 on RBCs before and after DTT treatment using a flow cytometric antibody binding assay (FABA) to specify the optimum conditions for CD38 inactivation. Materials and methods For FABA, untreated or DTT-treated RBCs were incubated with fluorescein isothiocyanate-labelled anti-CD38 antibody, in the presence or absence of 100-fold or more excess of unlabelled anti-CD38 antibody, and then analysed by flow cytometry (FCM). Dissociation of CD38-positive and control histograms was determined from the D-value using the Kolmogorov-Smirnov test. The results from FABA were compared with those from conventional FCM, indirect antiglobulin test (IAT) and Western blotting. Results The results from FABA were more consistent than those from conventional FCM. The D-value was found to be reliable in the analysis of difference between CD38 before and after DTT treatment. Our data showed that 0·0075 mol/l DTT for 30 min is sufficient to inactivate CD38 on RBCs. These results were stable and consistent with the findings from IAT. Conclusion Flow cytometric antibody binding assay is an objective way of evaluating the efficacy of DTT treatment for CD38 on RBCs. This approach allows the detection of a small number of cell surface antigens and will be useful for assessing the various chemical treatments to denature RBC antigens.

Background During storage, the potassium level of red blood cell (RBC) components increases, especially after irradiation. Neonates are prone to hyperkalemia, for example, non-oliguric hyperkalemia, so using potassium adsorption filters during transfusion may be helpful. To overcome dilution of RBC components caused by saline priming of existing potassium adsorption filters, a downsized potassium adsorption filter for neonates (PAF-n, Kawasumi Laboratories Inc., Tokyo, Japan) was developed. Study design and methods To assess the performance of PAF-n, its adsorption efficiency and RBC recovery rate were evaluated by testing pre-filtration and serial post-filtration (0-30 mL, 30-60 mL, 60-90 mL, and 90-120 mL) samples from 8 RBC components. Results The average potassium adsorption rate of the PAF-n was 90.5% ± 0.78%, and never less than 89.0% in any of 8 RBC components. RBC recovery rates were 99.3% ± 1.12%. Conclusion The PAF-n showed an effective potassium ability with negligible RBC dilution.

Background Hematopoietic cell infusion-related adverse events (HCI-AEs) in hematopoietic stem cell transplantations (HSCTs) have been largely attributed to toxicity of dimethyl sulfoxide (DMSO) for cryopreservation, but HSC products also contain various cells and plasma components. Our recent prospective study of 1125 HSCT recipients revealed the highest overall HCI-AE rate in bone marrow transplantation (BMT) using fresh/noncryopreserved products, although products of peripheral blood stem cell transplantation and cord blood transplantation (CBT) are generally cryopreserved with DMSO containing smaller plasma volumes. We aimed to clarify if product volume and component effects are more substantial in small recipients including children. Study design and methods We performed subgroup analysis on 219 recipients of 45 kg or less body weight (whole small recipients), including 90 children (pediatric recipients), from the original cohort (general recipients). Results Whereas overall HCI-AE rates did not differ among hematopoietic stem cell sources in the general recipients, bradycardia most often occurred after CBT in whole small recipients. Conversely, whole small and general recipients shared the same trend of having the highest rate of hypertension in BMT. The overall HCI-AE rate was higher in allogeneic HSCT compared with autologous HSCT. Notably, pediatric recipients showed a 10-fold higher incidence of nausea and vomiting in allogeneic HSCT compared with autologous HSCT, suggesting a possible role of allogeneic antigens. Multivariate analysis identified a relatively large infusion volume per body weight as a significant factor correlating with HCI-AE in whole small recipients. Conclusions We should be aware of product volume and specific HCI-AEs such as nausea and vomiting in small patients including children.

Background Despite intensive multimodal therapies, the prognosis of relapsed/ refractory Ewing sarcoma family tumors (RR-ESFTs) is dismal. Some case reports using allogeneic stem cell transplantation (allo SCT) for RR-ESFTs have been reported, however, the efficacy of allo SCT is yet to be established. Aim The purpose of this study was to evaluate the response and toxicity of T-cell replete haploidentical SCT (TCR-haplo-SCT) in RR-ESFTs. Methods and results In this study, we retrospectively analyzed six patients with RR-ESFTs who received TCR-haplo-SCT. Four patients had relapsed and two patients had refractory Ewing sarcoma. Before the TCR-haplo-SCT, all patients received a reduced intensity-conditioning regimen containing fludarabine, melphalan, and low-dose rabbit anti-thymocyte globulin (2.5 mg/kg), as well as graft-versus-host disease (GVHD) prophylaxis, which consisted of tacrolimus, methotrexate, and prednisolone. Primary neutrophil engraftment was achieved in all the patients. Four patients developed acute GVHD (aGVHD) (grade I, 1; grade II, 1; grade III, 2), and two patients developed chronic GVHD (cGVHD). Among the four that developed aGVHD, three survived for 14, 116, and 129 months without relapse, while one died due to a transplant-related complication. In contrast, the two patients who did not develop aGVHD experienced relapse early after TCR-haplo-SCT. Conclusions In this study, three of the six patients with RR-ESFTs survived for more than one year without relapse, and the treatment toxicity was considered acceptable even for patients who underwent high-intensity pretreatment. TCR-haplo-SCT could be a potential therapeutic option for patients with RR-ESFTs.

Human T-cell lymphotropic virus type 1 (HTLV-1), the causative agent of adult T-cell leukaemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), is endemic in sub-Saharan Africa (SSA) and poses a high morbidity and mortality risk. Its prevalence in the general population is poorly understood. The potential for prevention motivated us to do a systematic review and meta-analysis of population-based studies to estimate the prevalence of HTLV-1 in SSA.A comprehensive, no-limit search was conducted in EMBASE, PubMed, Web of Science and the Cochrane Library from their inception dates to March 2019. Population-based studies presenting data on HTLV-1 in sub-Saharan Africa were included. Pooled prevalence was estimated using a random-effects meta-analysis.A total of 21 studies were included, representing 42 297 participants. The pooled HTLV-1 seroprevalence was 3.19% (95% CI 2.36-4.12%) with variations across year of study. Prevalence of HTLV-1 positively correlated with year of study (β = 0.0036, P = 0.007). Participants from Central, Western and Southern Africa had a seroprevalence of 4.16% (95% CI 2.43-6.31%), 2.66% (95% CI 1.80-3.68%) and 1.56% (95% CI 0.48-3.15%), respectively.Our findings suggest that HTLV-1 infection is a public health concern in SSA and highlight the need to implement effective preventive programmes and interventions aimed at reducing the burden of this common yet neglected infection.PRÉVALENCE DANS LA POPULATION DU VIRUS T-LYMPHOTROPIQUE HUMAIN DE TYPE 1 (HTLV-1) EN AFRIQUE SUBSAHARIENNE: OBJECTIF: Le virus lymphotropique T humain 1 (HTLV-1), l'agent causal de la leucémie T de l'adulte/lymphome (ATL) et la myélopathie associée à HTLV-1/paraparésie spastique tropicale (HAM/TSP), est endémique en Afrique subsaharienne (ASS) et présente un risque élevé de morbidité et de mortalité. Sa prévalence dans la population générale est mal comprise. Le potentiel de prévention nous a incité à procéder à une revue systématique et à une méta-analyse des études basées sur la population afin d'estimer la prévalence du HTLV- 1 en ASS. MÉTHODES: Une recherche approfondie et sans limite a été effectuée dans EMBASE, PUBMED, Web of Science et dans la Cochrane Library, depuis leur création jusqu’à mars 2019. Des études basées sur la population présentant des données sur HTLV-1 en ASS ont été incluses. La prévalence poolée a été estimée à l'aide d'une méta-analyse à effet aléatoire. RÉSULTATS: Un total de 21 études ont été incluses, représentant 42.297 participants. La séroprévalence poolée du HTLV-1 était de 3,19% (IC95%: 2,36% à 4,12%), avec des variations au cours de l'année de l’étude. La prévalence du HTLV-1 était corrélée positivement avec l'année d’étude (bêta = 0,0037, p = 0,007). Les participants d'Afrique centrale, de l'Ouest et Australe présentaient une séroprévalence de 4,16% (IC95%: 2,43% à 6,31%), de 2,66% (IC95%: 1,80% à 3,68%) et 1,56% (IC95%: 0,48% à 3,15%), respectivement. CONCLUSIONS: Nos résultats suggèrent que l'infection au HTLV-1 est une préoccupation de santé publique en ASS et soulignent la nécessité de mettre en œuvre des programmes et des interventions préventives efficaces visant à réduire la charge de cette infection commune mais négligée.

Background Anti-KANNO, a broadly reactive RBC alloantibody, is found among some Japanese pregnant women, but the genetic basis of the corresponding antigen remains unclear. Study design and methods We integrated a statistical approach to identify the coding gene for KANNO antigen by conducting a genome-wide association study (GWAS) on four KANNO-negative individuals and 415 healthy Japanese. We also applied whole-exome sequencing to them and performed a replication study to confirm the identified genome variation using independent 14 KANNO-negative individuals. A monoclonal antibody-specific immobilization of erythrocyte antigens (MAIEA) assay was used to locate KANNO antigen on RBC-specific membrane protein. In vivo and in vitro binding assays of anti-KANNO were further applied to the cells expressing a candidate protein. Results The GWAS revealed a genome-wide significant association of chromosome 20p13 locus (p = 2.76E-08; odds ratio > 1000 [95% confidence interval = 48-23,674]). The identified single-nucleotide polymorphism located in an intronic region of the prion protein (PRNP) gene. Whole-exome sequencing revealed a missense variant in the PRNP gene (rs1800014, E219K), which is in linkage disequilibrium with the single-nucleotide polymorphism identified in the GWAS. All 18 KANNO-negative individuals possessed the homozygous genotype of the missense variant. The MAIEA assay using anti-KANNO and mouse antihuman prion protein showed a clear difference between KANNO-positive and KANNO-negative RBCs. Anti-KANNO showed direct binding to CHO-K1 cells expressing wild-type PRNP but not to those expressing E219K PRNP. Conclusion We first identified the coding gene of the high-frequency antigen KANNO located in PRNP and the missense variation (E219K) that affects the seropositivity of the KANNO antigen, which were confirmed by PRNP overexpressed cells.

Background Weak D-type RBCs have fewer D epitopes, but it remains unclear whether individuals with certain types of weak D produce alloanti-D directed at D epitopes absent from the RBCs, and whether it is an alloantibody or an autoantibody. We report the first case of a patient with a weak D Type 15 who produced autoantibodies mimicking alloanti-D. Case report A 52-year-old Japanese male with weak D developed anti-D 3 months after transfusion of D-negative and -positive RBCs, and the antibody persisted for 24 months with a consistently negative direct antiglobulin test. Eluates from the patient's RBCs demonstrated anti-D specificity. The recipient did not exhibit any signs of delayed hemolytic transfusion reaction. As his anti-D was removed by the different adsorbing cells of weak D Type 15 and autologous as well as D positive, D negative, weak D Type 24, and partial DVa, it was thought to be an autoantibody mimicking anti-D rather than an alloantibody. The patient's RBCs reacted weakly with the 13 anti-D reagents used in the study. Polymerase chain reaction and nucleotide sequencing revealed that the patient had an RHD genotype of RHD*01N.01/RHD*15. Conclusion Anti-D, produced in a patient with weak D Type 15 after transfusion, was found to be mimicking autoanti-D. Alloanti-D was excluded by an adsorption study with different RBC types.

BACKGROUND Improving apheresis technology may lead to an efficient and safe peripheral blood stem cell (PBSC) collection. Recently, the Spectra Optia (Optia, Terumo BCT) was introduced as an automated apheresis instrument, but comparisons with other instruments have been few. This is the first randomized multicenter and crossover comparison of the Optia with the automated program of the established apheresis instrument, the Spectra (Spectra-Auto, Terumo BCT). STUDY DESIGN AND METHODS A total of 233 apheresis procedures performed in 46 autologous patients and 108 allogeneic donors were investigated. Apheresis performed in the first day for all subjects using the Spectra-Auto (n = 79) and the Optia (n = 75) were evaluated as first-day analysis. Seventy-nine subjects, who required another session on the second day, underwent apheresis using the other instrument than the first-day instrument and were compared with each other in a paired crossover analysis. RESULTS The two instruments processed similar volumes with comparable run times and volumes of acid-citrate-dextrose used. The volumes of collected products were greater in the Optia. Yields of mononuclear cells and CD34+ cells were not different, but collection efficiencies were higher in the Optia (p = 0.008 in CE1 of crossover analysis). Spectra-Auto–collected products contained more contaminating red blood cells (RBCs), whereas there was a trend of more contaminating platelets (PLTs) in the Optia-collected products. Slight reductions were noted in the RBC or PLT counts of subjects who underwent apheresis with the Spectra-Auto or the Optia, respectively. CONCLUSION The Optia is safe and more efficient in the PBSC collection compared with the Spectra-Auto.

Background and objectives The presence of red blood cell (RBC) alloantibodies in pregnant women has been linked to the significant perinatal morbidity and mortality. A limited number of African studies have assessed alloimmunization to RBC antigens in pregnant women, but this literature has not been systematically reviewed. Thus, the aim of this study was to synthetize, by systematic review, the current evidence on RBC alloantibodies among pregnant women in Africa. Methods We systematically searched MEDLINE, EMBASE and the Africa-Wide Information database to identify relevant studies in any language. Case reports, comments, letters, conference abstracts, editorials and review articles were excluded. Results Of 269 potentially relevant articles, 12 studies representing 93 871 pregnant women fulfilled our selection criteria. Overall proportions of RBC alloimmunization exhibited a wide variation ranging from 1·1 (95% CI: 1·0, 1·2) to 12·1 (95% CI: 9·8, 14·7) per 100 pregnant women. Among clinically relevant antibodies, anti-D ranked as the most common, followed by anti-K and anti-E. Conclusion The review of the available literature characterized the clinical challenge of RBC alloimmunization among pregnant women in Africa and revealed the insufficient volume and quality of research conducted in this topic. Improvement of quality of research should be a priority to gather stronger evidence that should drive policy decisions and strengthen practice in favour of RBC alloantibody screening in pregnant women as a standard of care throughout Africa.

Background and Objectives Red blood cell (RBC) transfusion prior to allogeneic hematopoietic stem cell transplantation (allo-HSCT) is linked to poor outcomes, but outcomes related to post-transplant, especially postengraftment transfusions, have not been fully investigated. We investigated allo-HSCT outcome in association with early postengraftment transfusion. Materials and Methods We reviewed RBC and platelet transfusions in 50 recipients of bone marrow transplantation, specifically analysing postengraftment transfusions from 31 to 60 days post-transplant with no relapse in our institution. Results Postengraftment RBC and/or platelet transfusions were needed in 22 (44·0%) patients. Pretransplant haemoglobin concentrations were negatively correlated with RBC transfusions performed from days 0 to 30, but not postengraftment. Regardless of product type, transfused patients were more anaemic and thrombocytopenic even at day 100, when most of them no longer required transfusion. Patients with postengraftment transfusions, especially those who were transfused both RBC and platelet, showed inferior overall survival (P = 0·016) with increases of both chronic graft-versus-host disease and relapse (P = 0·002 and 0·019, respectively). Postengraftment RBC and platelet transfusion were significant predictors for poor survival in multivariable analysis. Conclusions Postengraftment transfusion may be associated with adverse outcomes of allo-HSCT.

SUMMARY Background: Despite improvements in first-line therapies, the outcomes of relapsed or refractory childhood acute leukaemia that has not achieved complete remission after relapse, has relapsed after stem cell transplantation (SCT), has primary induction failure and has relapsed with a very unfavourable cytogenetic risk profile, are dismal. Objectives and Methods: We evaluated the feasibility and efficacy of T-cell-replete haploidentical peripheral blood stem cell transplantation (haplo-SCT) with low-dose anti-human thymocyte immunoglobulin (ATG), tacrolimus, methotrexate and prednisolone (PSL) in 14 paediatric patients with high-risk childhood acute leukaemia. Results: All patients achieved complete engraftment. The median time to reaching an absolute neutrophil count of more than 0.5 × 10 9 L −1 was 14 days. Acute graft-vs-host disease (aGVHD) of grades II–IV and III–IV developed in 10 (71%) and 2 (14%) patients, respectively. Treatment-related mortality and relapse occurred in one (7%) patient and six (43%) patients, respectively. Eleven patients were alive and seven of them were disease-free with a median follow-up of 36 months (range: 30–159 months). The probability of event-free survival after 2 years was 50%. Conclusion: These findings indicate that T-cell-replete haplo-SCT, with low-dose ATG and PSL, provides sustained remission with an acceptable risk of GVHD in paediatric patients with advanced haematologic malignancies.

Background Leukofiltration of blood components is currently implemented worldwide as a precautionary measure against white blood cell–associated adverse effects and the potential transmission of variant Creutzfeldt-Jakob disease (vCJD). A newly developed bifunctional filter (Sepacell Prima, Asahi Kasei Medical) was assessed for prion removal, leukoreduction (LR), and whether the filter significantly affected red blood cells (RBCs). Study Design and Methods Sepacell Prima's postfiltration effects on RBCs, including hemolysis, complement activation, and RBC chemistry, were compared with those of a conventional LR filter (Sepacell Pure RC). Prion removal was measured by Western blot after spiking RBCs with microsomal fractions derived from scrapie-infected hamster brain homogenate. Serially diluted exogenous prion solutions (0.05 mL), with or without filtration, were injected intracerebrally into Golden Syrian hamsters. Results LR efficiency of 4.44 log with the Sepacell Prima was comparable to 4.11 log with the conventional LR filter. There were no significant differences between the two filters in hemoglobin loss, hemolysis, complement activation, and RBC biomarkers. In vitro reduction of exogenously spiked prions by the filter exceeded 3 log. The titer, 6.63 (log ID50/mL), of prefiltration infectivity of healthy hamsters was reduced to 2.52 (log ID50/mL) after filtration. The reduction factor was calculated as 4.20 (log ID50). Conclusion With confirmed removal efficacy for exogenous prion protein, this new bifunctional prion and LR filter should reduce the residual risk of vCJD transmission through blood transfusion without adding complexity to component processing.

Background In the Spectra apheresis instrument (Terumo BCT), both manual (Spectra-MNC) and automated (Spectra-Auto) programs have been widely used to collect peripheral blood progenitor cells (PBPCs). However, direct comparison of these programs remains extremely limited. Study Design and Methods We investigated 188 collections and products from autologous (patient) and allogeneic (donor) subjects and analyzed a subset of 89 allogeneic collections and products. Twenty-nine subjects who received apheresis for 2 consecutive days using both programs were also evaluated with a paired crossover comparison. Results The two programs processed similar volumes, but run time was longer with Spectra-Auto. Yield and efficiency of CD34+ cell collection were similar between these programs in the whole cohort, although white blood cell (WBC) and mononuclear cell (MNC) yields were higher with Spectra-MNC. In the allogeneic cohort, yield and efficiency of WBC collection were greater in Spectra-MNC. However, collected WBCs, MNCs, and CD34+ cells were similar between these programs in paired comparison. Regardless of program, preapheresis peripheral WBC, MNC, and CD34+ cell counts correlated with the number of cells collected. In contrast, preapheresis WBC counts in the whole cohort were negatively correlated with collection efficiencies of CD34+ cells in Spectra-MNC but not Spectra-Auto. The products collected using Spectra-MNC contained more contaminating platelets (PLTs) than Spectra-Auto, with a corresponding reduction in postdonation circulating PLTs. Conclusion Spectra-MNC and Spectra-Auto showed distinct features that should be considered on a case-by-case basis. Similar investigations should be undertaken as new collection platforms are introduced.

S. Panzer, S. Engelbrecht, M. F. Cole-Sinclair, E. M. Wood, S. Wendel, S. Biagini, Z. Zhu, J.-J. Lefrere, G. Andreu, T. Zunino, J.-J. Cabaud, P. Rouger, O. Garraud, K. Janetzko, M. Muller-Steinhardt, P. van der Burg, A. Brand, P. Agarwal, T. Triyono, A. Gharehbaghian, N. Manny, O. Zelig, A. Takeshita, Y. Yonemura, H. Fujihara, K. E. Nollet, H. Ohto, K.-S. Han, V. S. Nadarajan, G. Berlin, S. G. Sandler, R. G. Strauss & H. W. Reesink

Myeloid/natural killer cell precursor acute leukemia (MNKL) is an aggressive disease with a high relapse rate even after allogeneic hematopoietic stem cell transplantation (SCT). We report a patient with MNKL who had a donor lymphocyte infusion (DLI) for relapse after T cell-replete human leukocyte antigen (HLA)-haploidentical SCT, but relapsed again 20 months later with loss of mismatched HLA. This case suggests that a strong graft-versus-leukemia effect of haploidentical SCT can be expected in MNKL patients. In the haploidentical setting, DLI should be considered for patients with relapsed leukemia whose leukemic cells have not lost HLA cell surface expression.

In this study, we evaluated the feasibility of our graft-versus-host disease (GVHD) prophylaxis with tacrolimus, methotrexate, and prednisolone in non-T-cell-depleted haploidentical hematopoietic stem cell transplantation (HSCT) for children. Twenty-one consecutive patients including those with hematological malignancies (n = 11), solid tumors (n = 7), and non-malignancies (n = 3) were analyzed. Myeloablative and reduced intensity conditionings were carried out in 5 and 16 patients, respectively, and both of the regimens contained anti-human T-lymphocyte immunoglobulin. Twenty (95%) of the 21 patients achieved primary engraftment. Acute GVHD of grades II-IV and III-IV were observed in nine (47%) and one (5%) patient, respectively, all of which were controllable by steroids. Chronic GVHD was observed in eight (51%) of the 17 evaluable patients, and one of them developed steroid refractory chronic GVHD. Treatment-related mortality occurred in three patients (15%), as a result of acute pancreatitis, chronic GVHD, and EB virus associated lymphoproliferative disease. The median follow-up of the 13 survivors was 24 months, and the two-yr probability of overall survival was 68%. The Karnofsky performance scale score of the 13 survivors was 100%. These results indicated the feasibility of our GVHD prophylaxis in non-T-cell-depleted haploidentical HSCT for children.

Background Apheresis platelets (APs) have gained favour over whole blood-derived platelets on the presumption that they are less likely to provoke alloimmunization to red-blood-cell antigens. Case Reports Non-D Rh antibodies appeared in three patients after apheresis platelet transfusion. Anti-C and anti-E arose in two female patients with previous antigen exposure. Both anti-c and anti-E arose in a male recipient with no prior transfusion history. Materials and Methods Fifty APs were analysed for residual RBCs and RBC-derived microparticles, using samples obtained from a local blood centre. Cells and microparticles were quantified with a flow cytometry gating scheme, using PE-labelled anti-CD235a (glycophorin A) and FITC-labelled anti-CD41a (platelet gp IIb/IIIa) to distinguish lineage. Results Apheresis platelets were found to contain a mean of 7·5 × 106 (95% C.I. [6·3–8·5 × 106]) RBCs on one manufacturer’s device and 5·2 × 106 (95% C.I. [4·0–6·3 × 106]) RBCs on another’s. RBC-derived microparticles averaged 210·7 × 106 (95% C.I. [166·2–254·2 × 106]) on one manufacturer’s device and 232·3 × 106 (95% C.I. [194·3–272·9 × 106]) on another’s. These counts all correspond to volumes of

BACKGROUND: Platelet (PLT) transfusion–associated bacterial sepsis has remained a substantial patient risk, primarily due to lacking effective and point-of-issue measures to detect bacterial contamination. This study describes near infrared (NIR) spectroscopy to examine inoculated PLTs without sampling within a few seconds. STUDY DESIGN AND METHODS: This study evaluated apheresis PLTs inoculated with low concentrations of Bacillus cereus and Staphylococcus epidermidis, comparing with sterile bags. Short-wavelength NIR spectra over the range from 700 to 1100 nm in the transmittance mode were obtained using research (NIRS6500, Foss NIRSystems) and portable (NIRGun, Shizuoka Shibuya Seiki) instruments at 6-hour intervals from 0 to 72 hours after inoculation (HAI). RESULTS: The sensitivity of the NIRS6500 was 100% (43/43) and 98% (50/51) after incubating PLTs inoculated with B. cereus for 42 HAI or more and with S. epidermidis for 54 HAI or more, respectively. Difference spectra calculated by subtracting the NIR spectra of stored PLTs with that of the same PLTs measured at 0 HAI improved the discrimination results compared with conventional second derivative spectra. CONCLUSION: The NIRS6500 system can provide a PLT monitoring system based on difference spectra. The chemical components of PLTs that were influenced by bacterial metabolism seemed to play an important role in the calibration structure. Further studies should examine samples spiked with various species of prevalent bacteria.

BACKGROUND: Screening blood donors for hepatitis C virus (HCV) antibody has effectively mitigated transfusion transmission of HCV. We conducted a post hoc analysis to clarify the impact of donor screening on a general population of reproductive-age females and their offspring. STUDY DESIGN AND METHODS: Anti-HCV screening in Japan started in late-1989. In a cohort studied between May 1990 and November 2004, a total of 22,664 consecutive serum samples from pregnant women were screened for anti-HCV. Reactive samples were further tested for HCV RNA. Linear structural regression was applied to identify causal relationships. RESULTS: Anti-HCV–reactive rates declined significantly by two measures. First, among women known to have been transfused, rates fell from 14.8% to 3.1% with the implementation of anti-HCV screening (p

Although the natural life span of platelets is 7–10 days,platelet transfusion to stop or prevent bleeding is currentlyrestricted by storage limits of 4–7 days, depending on theregulatory authority and whether bacterial risk mitigationis performed. Not only the risk of bacterial growth, but alsoplatelet function and storage-dependent deterioration,referred to as platelet storage lesion (PSL) should be consid-ered. The extent of PSL correlates with reduced platelet invivorecovery and survival as well as haemostatic activityafter transfusion [1].

Background and Objectives To reveal the associations between cytokines in blood and transfusion-related adverse events, we studied whether pre-storage leucoreduction of autologous blood could reduce the degree of inflammatory responses, infection rates, or the duration of hospitalizations. Materials and Methods Patients scheduled to donate autologous blood for elective orthopaedic surgery were assigned to receive either leucoreduced (LR) or non-leucoreduced (N-LR) autologous blood based on their date of birth. Levels of cytokines in the autologous blood, values for C-reactive protein, complete blood count and body temperature of the patients, as well as adverse clinical events, were evaluated periodically. Results Four hundred patients entered this study (LR group: 196, N-LR group: 204). The production of cytokines, excluding interleukin 1β (IL-1β), was suppressed for the LR group. However, for unknown reasons, IL-1β actually increased during storage for the LR group. There were no differences between the two groups in the length of hospital stay, postoperative C-reactive protein changes, leucocyte count, or body temperature, and no clinical problems associated with blood transfusion were observed in either group. Conclusion Pre-storage leucoreduction for autologous blood may be effective to suppress cytokine accumulation. However, clinical benefits such as prevention of febrile non-haemolytic reactions could not be demonstrated.

Title of published version: Culture-based bacterial detection systems for platelets: the effect of time prior to sampling and duration of incubation required for detection with aerobic culture, BACKGROUND: Bacterial contamination of platelet products (PLTs) occurs at low concentrations requiring a period of incubation for growth to minimize sampling error. Culture-based detection methods also need sufficient incubation time; together these periods may limit the useful life of PLTs. This study characterizes the impact of sampling and detection times using two commercially-available bacteria detection products. STUDY DESIGN AND METHODS: Apheresis PLTs inoculated with nine bacterial species at low concentrations were sampled immediately, and after 24 hours following inoculation. Test results were analyzed following incubation at 16, 20 and 24 hours after sampling using two bacterial detection systems. RESULTS: When sampled immediately after inoculation, BacT/ALERT (BioMerieux) and Pall eBDS (Pall Corporation) failed to detect some PLTs inoculated with S. epidermidis, S. liquefaciens or P. aeruginosa and S. epidermidis, S. liquefaciens, B. cereus or P. aeruginosa, respectively. The BacT/ALERT was better at 20 hours (p

The aim of this study was to examine the extended storage of granulocyte concentrates mobilized by granulocyte-colony-stimulating factor (G-CSF) with/without dexamethasone (DEX) and collected by a bag separation method. Ten healthy adult volunteers donated blood three times: twice after granulocyte mobilization by (1) injecting G-CSF at 3 microg kg(-1) subcutaneously (s.c.) and (2) injecting G-CSF at 3 microg kg(-1) s.c. + DEX at 8 mg per oral and once (3) for a baseline control without any forms of mobilization. Granulocytes were collected by a bag separation method. The functions (phagocytosis and oxidative killing levels), viability and levels of interleukin (IL)-1beta, IL-8, IL-6 and tumour necrosis factor-alpha of granulocytes were measured. The average numbers of granulocytes collected from 200-mL samples of whole blood from the G-CSF and G-CSF + DEX groups were 35.1 x 10(8) and 49.4 x 10(8), respectively. Phagocytosis level, oxidative killing level and the viability of the granulocytes mobilized by G-CSF with/without DEX were well maintained for up to 72 h of storage after collection. The levels of the cytokines increased in a time-dependent manner. The in vitro phagocytosis level, oxidative killing level and the viability of granulocytes mobilized by G-CSF with/without DEX and collected by bag separation method can be maintained for as long as 72 h after collection.

BACKGROUND: Under the rationale that children undergoing elective surgery are the best candidates for autologous blood donors because of their long life expectancy, aggressive donations of autologous blood, even from infants, have been reported. A number of problems are associated with the procedure, however, whereas the risks of homologous blood are very low. STUDY DESIGN AND METHODS: From 1987 through 2005, of 5792 patients referred to blood transfusion services at two Japanese university hospitals for autologous blood donations, 314 children younger than 16 years old served as subjects for assessment. RESULTS: Of 314 children, 7 were not suitable as autologous donors. In most cases this was due to uncooperative behavior. Over a follow‐up period of 19 years, the authors encountered 53 cases (17.3%) of donation‐related problems, and this rate was higher than the 6 percent rate recorded for adult cases (316/5305). Nine children suffered crucial complications such as vasovagal reactions, and one 14‐year‐old boy required a vasopressor drug. Important findings were that 6 of these were first‐time donors, and the amount of blood drawn was under 10 percent of their estimated blood volume. CONCLUSION: Of 53 donation‐related problems, 9 (17.0%) were accompanied by marked hypotension. Drawing autologous blood from children has become easier with advanced devices; however, lessening of anxiety and tension are essential for the safety of children's autologous blood donation programs. Aggressive donation should be avoided.

We report a 65-year-old woman with chronic graft-versus-host disease (GVHD) who developed severely sclerotic skin on the fingers, hand and trunk following autologous peripheral blood stem cell transplantation (APBSCT). The patient had suffered from breast cancer and been treated with surgery and chemotherapy. She showed pancytopenia and was treated with APBCST. Four years after APBSCT, she developed the severe sclerotic changes on the fingers, hands, extremities and trunk. The skin biopsy showed a flattened epidermis and a proliferation of collagen bundles in the dermis. No anti-nucleolar DNA titers were detected in the serum. She was diagnosed with chronic GVHD. Despite treatment with oral prednisolone, the skin sclerotic change developed and the breast cancer recurred. She died due to pericarditis. This is a rare case of sclerodermatous GVHD following APBSCT. The serum interleukin (IL)-12 levels were examined during the treatment.

BACKGROUND: It has been controversial whether HLA antibodies cause hemolytic transfusion reactions (HTR) or shortened red blood cell (RBC) survival. A patient is reported who had two episodes of HTR, the latter of which was likely due to RBC-reactive HLA antibodies. CASE REPORT: A 77-year-old woman, admitted for gastric varix rupture, had no RBC-irregular antibodies detected before transfusion. On Hospital Day 12, after transfusion of 2 units of RBCs and 2 units of fresh-frozen plasma, the first delayed hemolytic episode occurred and anti-E, anti-c, anti-Jka, and unidentified RBC-reactive antibodies were detected in a serum sample from Day 14. Two additional units of matched RBCs were transfused with a leukoreduction filter on Days 19 and 22. After 4 hours of starting a transfusion on Day 22, the patient had fever, and a second hemolytic episode was recorded. Multireactive HLA antibodies (reactive against 20 of 20 donor panel lymphocytes) were detected in serum samples from Day 15 to Day 21. These HLA antibodies reacted strongly with HLA-A2 and HLA-B7 antigens, corresponding to Bgc and Bga antigens on RBCs, respectively. RBCs transfused on Day 22 were found to be HLA-A2 by genotyping. CONCLUSION: Strong HLA alloantibodies in this recipient appear to have caused a HTR. It is suggested that HLA antibodies be considered in patients with unexplained HTRs.

BACKGROUND: Recent molecular studies of the RHD gene have revealed that Del individuals retain a grossly intact RHD gene or have a portion of RHD in their genomes. No Del phenotype has yet been shown to induce a primary or secondary alloanti-D immunization, however. CASE REPORT: A 67-year-old D– Japanese woman with a history of allosensitization from transfusion of D+ red blood cells (RBCs) was negative for anti-D at admission. After she received RBCs from 19 apparently D– donors, she developed anti-D with an 8-fold titer. The titer of anti-D increased further to 128-fold after transfusions of cross-match–compatible D− negative RBCs from 40 donors over the next 2 years. Two of 59 donors were found to be RHD gene–positive and antigen D– with a Del phenotype, that is, RHD(K409K). CONCLUSION: This is the first case in which RBCs having the Del phenotype induced a secondary alloanti-D immunization. A D– donor with the RHD(K409K) allele was associated with the development of anti-D. Adverse episodes or evidence of hemolysis was not observed after the transfusion of RHD(K409K) RBCs. Further clinical evidence is needed to reveal whether the Del phenotype has a clinically relevant potential for anti-D immunization.

Background: There is little data on the evolution of hepatitis C virus (HCV) quasispecies in infants infected by mother-to-infant transmission during long-term follow up. The hypervariable region 1 (HVR1) of the HCV genome was investigated in two mother–infant pairs from birth to 7.6 and 10.2 years, respectively. Methods: Ten cDNA clones of HVR1 generated from HCV-RNA and extracted from serum samples of both pairs were analyzed. The sequences were compared with regard to variability, identity, and hydrophobia profile, and analyzed by phylogenetic studies. Results: The alanine aminotransferase (ALT) level was high with fluctuation in infant A and almost within the normal range in infant B. Sequence diversity was higher in infant A at 7.6 years than in infant B at 9.3 years (sequence identity with the mothers’; 69.3–70.7% vs 85.3–90.7% for nucleotides, and 48% vs 68–72% for amino acids, respectively). Compared to the first samples, amino acid changes greatly increased in infant A (35.2% at 4.9 years and 52% at 7.6 years), but not in infant B (4% at 5.6 years and 27.5% at 9.3 years). Phylogenetic studies revealed that quasispecies in infant A evolved to a greater extent than that in infant B. Hydrophobia profile analyses revealed that dynamic shifts between hydrophilia and hydrophobia occurred in both infants. Conclusions: As in adults, the evolution of HVR1 and variability of quasispecies increased in infants infected through mother-to-infant transmission for 10 years after birth. A large episode of ALT elevation suggested the emergence of escape mutants and the evolution of new quasispecies.

Summary This study evaluated a newly developed polyolefin bag, which has a 50%/m 2 higher oxygen permeability to extend the shelf-life of platelets. Single-donor aphaeresis platelets were pooled, separated equally into two bags, PO-80 (0AE8 l capacity) and KBP-PO (1 l capacity) for control, and stored in plasma for up to 7 d. Platelet biochemical and functional parameters were monitored in bags containing high (4AE2 · 10 11 /250 ml/bag, n ¼ 9) and low (2AE0 · 10 11 / 200 ml/bag, n ¼ 3) concentrations of platelets over the storage period. After 7 d of storage, the PO-80 bags containing high concentrations of platelets had a better pH (mean pH 6AE74) than those stored in KBP-PO (pH 6AE32, P

BACKGROUND : ACD-A solution containing sodium citrate and citric acid is used as an anticoagulant agent during peripheral blood progenitor cell (PBPC) harvesting, and in rare cases can cause fatal citrate intoxication. The aim of this study was to establish effective methods for stabilizing ionized calcium (ICa) levels during PBPC harvesting. STUDY DESIGN AND METHODS : ICa was measured during 46 apheresis procedures conducted in 26 patients. Four patients in four procedures were infused with calcium gluconate solution before PBPC harvesting; three patients in six procedures were infused with calcium gluconate when symptoms of citrate intoxication appeared; and four patients in five procedures received a continuous infusion. Five patients in five procedures took an isotonic sports drink containing calcium when hypocalcemic symptoms appeared. The ICa level, blood pressure, and pulse rate were measured. RESULTS : ICa declined rapidly from the preapheresis level of 1.081(±0.092) m M to 0.937(±0.081) m M (13.3%, p < 0.0001) 10 minutes after the start of apheresis and continued to decline until the completion of the procedure. When patients received a continuous infusion of calcium during apheresis, ICa was relatively stabilized. ICa significantly rose (6.1 ± 3.6%, p < 0.02) within 2 to 5 minutes after oral intake of an isotonic sports drink containing calcium and was maintained within normal range for 31 to 55 minutes. CONCLUSION : An isotonic sports drink containing calcium has a quick stabilizing and a longer maintenance effect on ICa. Thus, we recommend the intake of an isotonic sports drink containing calcium as the easiest and best method for preventing hypocalcemia during apheresis.

BACKGROUND: TT virus (TTV) is widespread in the general population, however, the mode of its transmission and the mechanism of maintaining it in the general population are unclear. STUDY DESIGN AND METHODS: To determine the possible mother-to-infant route of transmission, 54 infants born to 50 anti-HCV-positive mothers were assessed longitudinally. Nucleotide sequences amplified by seminested PCR with primers targeting the N22 variable coding region of genotypes 1 through 6 were compared in mothers and their infants. RESULTS: The prevalence of TTV DNA was 30 percent (15/50; 95% CI, 18-45) in mothers and 44 percent (24/54; 95% CI, 31-59) in their infants. TTV DNA was detected during a follow-up period in 13 (87%; 95% CI, 60–98) of 15 infants born to infected mothers and in 11 (28%; 95% CI, 15-45) of 39 infants born to DNA-negative mothers. None of 38 cord blood samples, but one of 14 blood samples, obtained at 1 month of age had detectable TTV DNA. The lowest infection rate at the earliest ages and the subsequent increasing prevalence of infection (22% at 6 months and 33%[43% cumulative rate] at 2 years) is consistent with an age-dependent acquisition of TTV by nonparenteral routes. In 13–mother-infant pairs positive for TTV DNA, six showed a high degree of nucleotide sequence similarity (99.1-100%), whereas the remaining seven pairs differed more than 10 percent from each other (46.8-89.2%). The viral load of maternal blood was not a plausible risk factor for transmission. Genotype 1, of which pathogenicity failed to be shown by measurement of hepatic enzymes, was more rapidly cleared (88 vs. 8% other genotypes, p < 0.001) among infants. CONCLUSIONS: These observations strongly suggest that the main factor for TTV acquisition in children involves their age-associated increase in environmental interactions with infectious materials. Genotype 1 might be involved in a weak or a limited pathologic role, which can possibly be diluted by other harmless genotypes.

The Japanese Red Cross analysed the results of questionnaires sent in 1993 regarding post-transfusion graft-vs.-host disease (PT-GVHD) from hospitals; the majority of patients with PT-GVHD in 1993 were transfused for cardiovascular or cancer surgery, and about 10 patients had died yearly from PT-GVHD in the following few years. The Japan Society of Blood Transfusion (JSBT) organized a subcommittee for the prevention of PT-GVHD, and issued a fourth version of guidelines for the irradiation of blood to prevent PT-GVHD. These guidelines recommended transfusion of irradiated blood for cardiosurgery, cancer surgery, elderly recipients and severe trauma, as well as congenital immunodeficient recipients, newborn infants and other immunocompromised patients. Also recommended was irradiation not only of blood within 72 h after collection but also of blood stored for 14 days. Reported PT-GVHD has diminished to a few cases in recent years.

Summary. The proliferative and differentiative response of neutrophils to granulocyte‐macrophage colony-stimulating factor (GM-CSF) is known to be impaired in patients with myelodysplastic syndromes (MDS). To investigate the mechanisms of the defective response in MDS, we examined expression levels of GM-CSF receptor a (GMRa) and common b (bc) subunits on CD16 1 neutrophils, CD14 1 monocytes and CD3 1 T cells from 26 MDS patients and 10 healthy controls using flow cytometry. Expression of GMRa was significantly decreased on the neutrophils of five out of 26 patients and was not specific for any FAB subtype. In contrast, bc expression on neutrophils was significantly reduced in 14 out of 26 patients with a higher proportion occurring in the advanced stages of MDS including refractory anaemia with excess of blasts (RAEB), RAEB in transformation (RAEBt) and overt leukaemia compared with refractory anaemia (RA)/RA with ringed sideroblasts (RARS) or healthy controls. Decreased bc also correlated with the degree of hypogranular neutrophil morphology and increased infection. Expression of both subunits on T cells and monocytes in MDS was similar to normal controls. Polymerase chain reaction amplification of reversetranscribed mRNA isolated from the affected neutrophils suggests that the reduction of bc may result from decreased message levels. The observed reduction in GM-CSF receptor expression could account for the impaired proliferative and maturational responses in MDS.

Anti-HPA-5b is the most commonly found platelet-specific antibody among pregnant women, but it does not cause severe fetal-neonatal alloimmune thrombocytopenia in the majority of affected infants. However, as the sequelae of the affected children may become severe, it is necessary to identify the risk factors for neonatal alloimmune thrombocytopenia. Of 21 354 consecutive pregnant women, 138 [0.65%; 95% confidence interval (CI) 0.54-0.75%], corresponding to 13.2% of the 1049 HPA-5b- women calculated by the gene frequency, were positive for anti-HPA-5b at the first trimester. Anti-HPA-5b was titrated in specimens obtained at the third trimester and antibody-positive women and their neonates were HPA-5 genotyped. Platelet counts in cord blood and 3 d after birth were assessed in the infants born to these mothers. Chi-square analysis showed no significant relationship between the titres of maternal antibody to HPA-5b and the number of pregnancies. There was a significant difference in platelet counts at d 3 between neonates who were compatible (267 x 109/l) or incompatible (220 x 109/l, P /= 64 were related to the development of thrombocytopenia ( /= 64) had a positive predictive value of 50% for thrombocytopenia 3 d after birth when the infant was HPA-5b+ and a negative predictive value of 100%. These results indicate that a high titre (>/= 64) of anti-HPA-5b is associated with a higher risk of neonatal thrombocytopenia, even if anti-HPA-5b-induced severe thrombocytopenia rarely develops.

BACKGROUND: The potentially hepatotropic flavivirus-like virus, GB virus type C (GBV-C)/HGV, has been detected in a few patients with acute and chronic hepatitis and in a certain proportion of blood donors and recipients of blood or blood components. STUDY DESIGN AND METHODS: Sera from 2979 pregnant Japanese women were examined for the presence of GBV-C/HGV RNA by nested RT-PCR. Mothers who were positive for viral RNA and their 34 infants were followed and tested for infection. RESULTS: Of the 2979 women, 32 (1.1%) were positive for GBV-C/HGV RNA. Twenty-six (76.5%) of 34 babies born to these women were positive for the virus when first tested. A significantly higher titer of viral RNA was observed in mothers whose infants were infected than in those whose infants were uninfected (mean ± SD, 106.3 ± 0.9 vs. 104.6 ± 0.9/mL; p

Background and Objectives: The genetic background of the DVa category phenotype has been described in two Caucasian individuals. We were interested in the RHD sequence of 7 Japanese DVa individuals and their families. Materials and Methods: With SSP–PCR we tested exons 4, 5 and 7 of the RHD gene and used restriction enzymes for testing nucelotide associated with the DVa phenotype. Results: A single RHD G667 C697 allele was present in 5 individuals with DVa category phenotype, and in 2 individuals we found a D–CE–D hybrid gene (exon 5 had been replaced). The DVaCe gene complex was found in all families. Conclusion: The changes of the RHD gene described in European DVa individuals were also observed in Japanese families.

BACKGROUND: Weak D phenotypes involve a quantitative variation of D. The genomic basis in weak D has been disputed, however. STUDY DESIGN AND METHODS: Five sequence-specific polymerase chain reactions (SSP- PCRs) on exons 2, 5, and 7 of the RHD gene were evaluated in 248 white and 98 Japanese blood donors and compared with the results obtained by amplification of intron 4 and serology. All methods and SSP-PCR testing on the 3′ non-coding region of the RHD gene were applied to the genotyping of 94 DNA samples derived from individuals expressing weak D phenotypes. RESULTS: Concordant results were obtained with all genotyping and phenotyping methods in testing 201 D-positive and 145 D- negative donors. Four of 94 weak D samples were typed as D-negative by amplification of intron 4 and SSP-PCR on exon 5. Phenotyping with monoclonal antibodies revealed a DVI category in one of these cases and DFR phenotype in three of these cases. One weak D sample, which reacted like normal D-positive cells with all applied monoclonal antibodies, was typed falsely negative by SSP-PCR on exon 5 because of a point mutation at nucleotide 667 (T–>G) that resulted in a Phe223Val amino acid substitution. In this individual, heterozygosity was found at two other amino acid positions (Glu233Gln and Val238Met) by restriction fragment length polymorphism analysis. CONCLUSION: Genetic diversity in weak D phenotypes is rare. Only 1 of 90 true weak D phenotypes (1.1%) had a genetic variation in testing on seven gene regions of the RHD gene.

Background Microparticles in blood components might contribute to transfusion-related immunomodulation or other side effects. To elucidate the role of leukofiltration, we compared three commercially available filters for their effect on platelet (PLT)-derived (PDMP), leukocyte-derived (LDMP), and red blood cell–derived (RDMP) microparticle formation in apheresis PLTs. Study Design and Methods Apheresis PLTs from pairs of ABO-identical male donors were pooled and divided into four volumes. One volume was stored without filtration, whereas the other three were filtered with different devices. PDMPs, LDMPs, and RDMPs were measured by flow cytometry during 2 weeks of controlled-temperature (22°C) agitated storage. Results On average, PDMPs doubled over 5 days of storage, followed by a much steeper increase by which PDMPs on Day 14 were nearly 20 times higher than on Day 0. LDMP and RDMP counts were relatively stable over 14 days. Significant differences among filtered and nonfiltered products did not emerge. Conclusion Although the conditions of this study showed no favorable or unfavorable effects of three different filters on microparticle formation, surveillance and investigation of unanticipated outcomes in other experimental and clinical circumstances should continue.

BACKGROUND: Platelet-specific antibodies may be involved in refractoriness to platelet transfusions, disorders such as neonatal alloimmune thrombocytopenia, and post-transfusion purpura. Genotyping for the major human platelet antigen (HPA) systems HPA-1 through HPA-5 is of considerable help in establishing the diagnoses of these diseases. STUDY DESIGN AND METHODS: A new genotyping method is described. Alleles of all five systems are amplified in a multiplex polymerase chain reaction. Subsequently, aliquots of the amplification products are thermocycled in the presence of a pair of allele-specific oligonucleotide probes and a heat-stable ligase. After heat denaturation, the probes hybridize adjacent to complementary sequences of the amplification product. In a perfect match, the two probes become covalently joined. Detection of the ligation product is performed with an enzyme-linked immunosorbent assay. RESULTS: Complete concordance of genotypes between the ligation-based typing and established genotyping methods was determined in 54 Austrian (HPA-1, −2, −3, and −5) and 56 Japanese (HPA-4) individuals. Ligation-based genotyping of HPA-1 polymorphism using platelet-derived RNA as starting material gave concordant results in all 15 cases tested. CONCLUSION: Multiplex polymerase chain reaction in combination with ligation-based typing allows fast typing of large numbers of platelet donors and screening for critical antigens in pregnant women.

Background: The importance of autologous blood donation for elective surgery is recognized, and the method is being used at many hospitals. Not all patients are able to deposit a sufficient amount of blood before surgery because they cannot recover rapidly enough from phlebotomy-induced anemia. The ability to donate sufficient blood for autologous use was studied in patients who are particularly susceptible to phlebotomy-induced anemia. Study Design and Methods: Of 840 patients who donated blood for autologous use in elective surgery from November 1987 through May 1993, 20 with rheumatoid arthritis, 24 with iron deficiency anemia, and 37 aged 65 years and above with normocytic anemia were compared with 24 nonanemic elderly patients who donated a total of 1000 mL of blood for autologous use. Patients received iron sulfate orally and donated blood once a week until operation. Results: The amount of blood collected before surgery per control patient was more than that in others. Consequently, there was a tendency to allogeneic blood transfusion in patients with rheumatoid arthritis or elderly patients. The ferritin levels in controls and in patients with iron deficiency anemia during the donation period were almost within the normal range in spite of iron supplementation, which implied a good utilization of iron sulfate for erythropoiesis. On the other hand, the rise in ferritin levels in the elderly and in patients with rheumatoid arthritis suggested inappropriate iron availability for erythropoiesis and resulted in an increase in iron storage. Since an adequate endogenous erythropoietin response to phlebotomy-induced anemia was not observed in these patients, impaired erythropoietin production was considered one of the reasons for anemia. Conclusion: Patients with iron deficiency anemia are able to continue donating blood for autologous use so long as they have sufficient iron supplementation. However, the elderly or those with rheumatoid arthritis occasionally fail to donate a sufficient volume of blood before surgery as a result of phlebotomy-induced anemia, which is caused in turn by impaired erythropoietin production.