DAHEAN: A Danish nationwide study ensuring quality assurance through real-world data for suspected hereditary anemia patients.

BACKGROUND: Hereditary anemias are a group of genetic diseases prevalent worldwide and pose a significant health burden on patients and societies. The clinical phenotype of hereditary anemias varies from compensated hemolysis to life-threatening anemia. They can be roughly categorized into three broad categories: hemoglobinopathies, membranopathies, and enzymopathies. Traditional therapeutic approaches like blood transfusions, iron chelation, and splenectomy are witnessing a paradigm shift with the advent of targeted treatments. However, access to these treatments remains limited due to lacking or imprecise diagnoses. The primary objective of the study is to establish accurate diagnoses for patients with hereditary anemias, enabling optimal management. As a secondary objective, the study aims to enhance our diagnostic capabilities. RESULTS: The DAHEAN study is a nationwide cohort study that collects advanced phenotypic and genotypic data from patients suspected of having hereditary anemias from all pediatric and hematological departments in Denmark. The study deliberates monthly by a multidisciplinary anemia board involving experts from across Denmark. So far, fifty-seven patients have been thoroughly evaluated, and several have been given diagnoses not before seen in Denmark. CONCLUSIONS: The DAHEAN study and infrastructure harness recent advancements in diagnostic tools to offer precise diagnoses and improved management strategies for patients with hereditary anemias.

SMIM1 absence is associated with reduced energy expenditure and excess weight.

BACKGROUND: Obesity rates have nearly tripled in the past 50 years, and by 2030 more than 1 billion individuals worldwide are projected to be obese. This creates a significant economic strain due to the associated non-communicable diseases. The root cause is an energy expenditure imbalance, owing to an interplay of lifestyle, environmental, and genetic factors. Obesity has a polygenic genetic architecture; however, single genetic variants with large effect size are etiological in a minority of cases. These variants allowed the discovery of novel genes and biology relevant to weight regulation and ultimately led to the development of novel specific treatments. METHODS: We used a case-control approach to determine metabolic differences between individuals homozygous for a loss-of-function genetic variant in the small integral membrane protein 1 (SMIM1) and the general population, leveraging data from five cohorts. Metabolic characterization of SMIM1-/- individuals was performed using plasma biochemistry, calorimetric chamber, and DXA scan. FINDINGS: We found that individuals homozygous for a loss-of-function genetic variant in SMIM1 gene, underlying the blood group Vel, display excess body weight, dyslipidemia, altered leptin to adiponectin ratio, increased liver enzymes, and lower thyroid hormone levels. This was accompanied by a reduction in resting energy expenditure. CONCLUSION: This research identified a novel genetic predisposition to being overweight or obese. It highlights the need to investigate the genetic causes of obesity to select the most appropriate treatment given the large cost disparity between them. FUNDING: This work was funded by the National Institute of Health Research, British Heart Foundation, and NHS Blood and Transplant.

Functional asymmetry of the urea transporter UT-B in human red blood cells.

We studied urea, thiourea, and methylurea transport and interaction in human red blood cells (RBCs) under conditions of self-exchange (SE), net efflux (NE), and net influx (NI) at pH 7.2. We combined four methods, a four-centrifuge technique, the Millipore-Swinnex filtering technique, the continuous flow tube method, and a continuous pump method to measure the transport of the 14C-labeled compounds. Under SE conditions, both urea and thiourea show perfect Michaelis-Menten kinetics with half-saturation constants, K½,SE (mM), of ≈300 (urea) and ≈20 (thiourea). The solutes show no concentration-dependent saturation under NE conditions. Under NI conditions, transport displays saturation or self-inhibition kinetics with a K½,NI (mM) of ≈210 (urea) and ≈20 (thiourea). Urea, thiourea, and methylurea are competitive inhibitors of the transport of analog solutes. This study supports the hypothesis that the three compounds share the same urea transport system (UT-B). UT-B functions asymmetrically as it saturates from the outside only under SE and NI conditions, whereas it functions as a high-capacity channel-like transporter under NE conditions. When the red blood cell enters the urea-rich kidney tissue, self-inhibition reduces the urea uptake in the cell. When the cell leaves the kidney, the channel-like function of UT-B implies that intracellular urea rapidly equilibrates with external urea. The net result is that the cell during the passage in the kidney capillaries carries urea to the kidney to be excreted while the urea transfer from the kidney via the bloodstream is minimized.NEW & NOTEWORTHY The kinetics of urea transport in red blood cells was determined by means of a combination of four methods that ensures a high time resolution. In the present study, we disclose that the urea transporter UT-B functions highly asymmetric being channel-like with no saturation under conditions of net efflux and saturable under conditions of net influx and self-exchange in the concentration range 1-1,000 mM (pH 7.2 and 25-38 °C).

Evaluating the inhibition of IL-17A and TNFα in a cartilage explant model cultured with Th17-derived cytokines.

Introduction: T-helper 17 (Th17) cells produce IL-17A playing a critical role in activating the pathogenic chain leading to joint tissue inflammation and destruction. Elevated levels of Th17 cells and IL-17A have been detected in skin lesions, blood, and synovial fluid from patients with psoriatic arthritis (PsA) and ankylosing spondylitis (AS). Moreover, IL-17A inhibitors suppress disease activity in psoriasis, PsA and AS, supporting the evidence of IL-17A contributing to the disease pathogenesis. Although, IL-17A inhibitors are widely approved, it remains unclear how the inhibitory effect of IL-17A alters the extracellular matrix (ECM) of the joint in a Th17-conditioned inflammatory milieu. Therefore, the aim of this study was to establish a cartilage model cultured with conditioned medium from Th17 cells and inhibitors to explore the effect of IL-17A inhibition on joint tissue remodeling. Methods: Naïve CD4+ T cells from healthy human buffy coat were differentiated into Th17 cells, followed by Th17 cell activation to secrete Th17-related cytokines and molecules into media. The activated Th17 cells were isolated from the conditioned media (CM) and analyzed using flow cytometry to verify Th17 cell differentiation. The CM were assessed with ELISA to quantify the concentrations of cytokines secreted into the media by the Th17 cells. Healthy bovine cartilage explants were cultured with the Th17-CM and treated with IL-17A and TNFα inhibitors for 21 days. In harvested supernatant from the cartilage cultures, MMP- and ADAMTS-mediated biomarker fragments of type II collagen, aggrecan, and fibronectin were measured by ELISA to investigate the ECM remodeling within the cartilage tissue. Results: Th17-CM stimulated a catabolic response in the cartilage. Markers of type II collagen and aggrecan degradation were upregulated, while anabolic marker of type II collagen formation remained on similar levels as the untreated explants. The addition of IL-17A inhibitor to Th17-CM decreased the elevated type II collagen and aggrecan degradation, however, degenerative levels were still elevated compared to untreated group. The addition of TNFα inhibitor completely reduced both type II collagen and aggrecan degradation compared to untreated explants. Moreover, the TNFα inhibitor treatment did not alter the type II collagen formation compared to untreated group. Conclusion: This study suggests that inhibition of IL-17A in Th17-conditioned cartilage tissue only partially reduced the MMP-mediated type II collagen degradation and ADAMTS-mediated aggrecan degradation, while the TNFα inhibitor treatment fully reduced both MMP- and ADAMTS-mediated ECM degradation. This exploratory study where ECM biomarkers are combined with Th17-conditioned ex vivo model may hold great potential as output for describing joint disease mechanisms and predicting structural effects of treatment on joint tissue.

Extreme Reticulocytosis After Splenectomy in a Patient With Hemoglobin Mizuho.

BACKGROUND: Patients with Hb Mizuho may be splenectomized at a young age to decrease their need for blood transfusions. OBSERVATIONS: Transfusion-dependency decreased dramatically in a 4-year-old white boy with Hb Mizuho after splenectomy. Surprisingly, he developed reticulocytosis (>1000×10 9 /L) with a peak reticulocyte percentage of 49%, and erythrocyte abnormalities, including Heinz bodies, Howell-Jolly bodies, and basophilic stippling. Manual reticulocyte counting and flow cytometric measurement with anti-CD71 antibodies supported a truly elevated reticulocyte count. CONCLUSIONS: We propose possible explanations for the extreme reticulocytosis that arose postsplenectomy and compare the reticulocyte count in the present case with previously published cases.

The postgraduate medical educational climate assessed by the Danish Residency Educational Climate Test (DK-RECT): a validation and cross-sectional observational study.

BACKGROUND: A good educational climate is essential for delivering high-quality training for medical trainees, professional development, and patient care. The aim of this study was to (1) validate the Dutch Residency Educational Climate Test (D-RECT) in a Danish setting and (2) describe and evaluate the educational climate among medical trainees. METHODS: D-RECT was adopted in a three-step process: translation of D-RECT into Danish (DK-RECT), psychometric validation, and evaluation of educational climate. Trainees from 31 medical specialties at Copenhagen University Hospital - Rigshospitalet, Denmark were asked to complete an online survey in a cross-sectional study. RESULTS: We performed a forward-backward translation from Dutch to Danish. Confirmatory factor analysis showed that DK-RECT was robust and valid. The reliability analysis showed that only seven trainees from one specialty were needed for a reliable result. With 304 trainees completing DK-RECT, the response rate was 68%. The subsequent analysis indicated a positive overall educational climate, with a median score of 4.0 (interquartile range (IQR): 3.0-5.0) on a five-point Likert scale. Analysis of the subscales showed that the subscale Feedback received the lowest ratings, while Supervision and Peer collaboration were evaluated highest. CONCLUSIONS: Psychometric validation of D-RECT in a Danish context demonstrated valid results on the educational climate in specialist training. DK-RECT can be used to evaluate the effectiveness of interventions in the future and can facilitate the conversation on the educational climate.

Classification of fetal and adult red blood cells based on hydrodynamic deformation and deep video recognition.

Flow based deformation cytometry has shown potential for cell classification. We demonstrate the principle with an injection moulded microfluidic chip from which we capture videos of adult and fetal red blood cells, as they are being deformed in a microfluidic chip. Using a deep neural network - SlowFast - that takes the temporal behavior into account, we are able to discriminate between the cells with high accuracy. The accuracy was larger for adult blood cells than for fetal blood cells. However, no significant difference was observed between donors of the two types.

Hunting for the elusive target antigen in gestational alloimmune liver disease (GALD).

The prevailing concept is that gestational alloimmune liver disease (GALD) is caused by maternal antibodies targeting a currently unknown antigen on the liver of the fetus. This leads to deposition of complement on the fetal hepatocytes and death of the fetal hepatocytes and extensive liver injury. In many cases, the newborn dies. In subsequent pregnancies early treatment of the woman with intravenous immunoglobulin can be instituted, and the prognosis for the fetus will be excellent. Without treatment the prognosis can be severe. Crucial improvements of diagnosis require identification of the target antigen. For this identification, this work was based on two hypotheses: 1. The GALD antigen is exclusively expressed in the fetal liver during normal fetal life in all pregnancies; 2. The GALD antigen is an alloantigen expressed in the fetal liver with the woman being homozygous for the minor allele and the father being, most frequently, homozygous for the major allele. We used three different experimental approaches to identify the liver target antigen of maternal antibodies from women who had given birth to a baby with the clinical GALD diagnosis: 1. Immunoprecipitation of antigens from either a human liver cell line or human fetal livers by immunoprecipitation with maternal antibodies followed by mass spectrometry analysis of captured antigens; 2. Construction of a cDNA expression library from human fetal liver mRNA and screening about 1.3 million recombinants in Escherichia coli using antibodies from mothers of babies diagnosed with GALD; 3. Exome/genome sequencing of DNA from 26 presumably unrelated women who had previously given birth to a child with GALD with husband controls and supplementary HLA typing. In conclusion, using the three experimental approaches we did not identify the GALD target antigen and the exome/genome sequencing results did not support the hypothesis that the GALD antigen is an alloantigen, but the results do not yield basis for excluding that the antigen is exclusively expressed during fetal life., which is the hypothesis we favor.

Blood donor genotyping for prediction of blood group antigens: Results from 5 years' experience (2017-2022).

BACKGROUND AND OBJECTIVES: For 5 years, routine genotyping has been performed for selected blood groups of blood donors in the Copenhagen Capital Region, Denmark. The result is summarized in the following. MATERIALS AND METHODS: Genotyping was carried out by an external service provider using the competitive allele specific PCR (KASP) technology. The genotypes were returned to the blood bank and translated into phenotypes by a proprietary IT application. RESULTS: In total, 65 alleles from 16 blood group systems (ABO, MNS, Rh, Lutheran, Kell, Duffy, Kidd, Diego, Yt, Dombrock, Colton, Landsteiner-Wiener, Cromer, Knops, Vel, secretor status) and the HPA1, HPA5 and HPA15 antigens were interrogated. After translation, phenotypes were imported into the laboratory information management system of the blood bank. The results from 31,538 genotyped blood donors were used to calculate the allele frequencies for a Danish blood donor population. ABO genotyping was done for sample ID purposes. Determination of the 1061delC single nucleotide polymorphism (SNP) (NM_020469.2), most frequently characteristic of ABO*A2, was validated against a series of 1287 samples with Dolichos biflorus lectin determination of the A1 phenotype. CONCLUSION: We report allele frequencies and phenotype frequencies for 16 blood groups from a total of 31,538 genotyped blood donors. Blood products were supplied from a total of 64,312 active blood donors, and of these active blood donors 25,396 (39.5%) were genotyped. These donors represent a valuable resource for patient treatment. This genotyping has enabled the provision of rare genotyped donor blood for patients with alloantibodies and rare reagent cells for serology.

Urea transport in human red blood cells: Donor variation compared to chloride, glucose, and water transport.

We determined the permeability (P, cm/s) of unmodified human red blood cells (HRBC) to urea (Pu), chloride (PCl), glucose (Pglu), and water diffusion (Pd) under conditions of self-exchange (SE) with the continuous flow tube method at pH 7.2, 25°C. Among 24 donors, Pu at 1 mM varied >100%. Two of the donors were also tested in 1983. Their Pu had decreased by 77 and 90%. High age in males and Kidd genotype Jk(a+,b+), but not blood types AB0, appear related to low Pu. For one of the two donors, PCl (150 mM, 38°C, pH 7.2), Pglu (1 mM, 38°C, pH 7.2), and Pd (55.5 M, 25°C, pH 7.2) were determined then and now and showed no significant changes with age. The results from six more donors show donor PCl, Pglu, and Pd in the range of ≈1%. PCl and Pglu are vital for the metabolism of cells and tissues, and we see but little donor variation, and so far, no phenotypes without glucose (GLUT1) and anion (AE1) transporters in HRBC. Phenotypes with no urea transporter (UT-B) or no water transporters (aquaporin, AQP1) are registered and are compatible with life. Our results are in line with the concept that the solutes do not share pathways in common. The great donor variation in Pu must be considered in comparative transport physiological studies.

Human blood neutrophils generate ROS through FcγR-signaling to mediate protection against febrile P. falciparum malaria.

Blood phagocytes, such as neutrophils and monocytes, generate reactive oxygen species (ROS) as a part of host defense response against infections. We investigated the mechanism of Fcγ-Receptor (FcγR) mediated ROS production in these cells to understand how they contribute to anti-malarial immunity. Plasmodium falciparum merozoites opsonized with naturally occurring IgG triggered both intracellular and extracellular ROS generation in blood phagocytes, with neutrophils being the main contributors. Using specific inhibitors, we show that both FcγRIIIB and FcγRIIA acted synergistically to induce ROS production in neutrophils, and that NADPH oxidase 2 and the PI3K intracellular signal transduction pathway were involved in this process. High levels of neutrophil ROS were also associated with protection against febrile malaria in two geographically diverse malaria endemic regions from Ghana and India, stressing the importance of the cooperation between anti-malarial IgG and neutrophils in triggering ROS-mediated parasite killing as a mechanism for naturally acquired immunity against malaria.

Reduced susceptibility to COVID-19 associated with ABO blood group and pre-existing anti-A and anti-B antibodies.

BACKGROUND: Susceptibility to severe acute respiratory syndrome coronavirus 2 shows individual variability in un-vaccinated and previously un-exposed individuals. We investigated the impact of ABO blood group, titers of anti-A and anti-B, other blood group antigens, and the extracellular deposition of ABH antigens as controlled by secretor fucosyltransferase 2 (FUT2) status. STUDY DESIGN AND METHODS: We studied incidents in three different hospitals between April to September 2020, where un-diagnosed coronavirus disease 2019 (COVID-19) patients were cared for by health care workers without use of personal protection and with close contact while delivering therapy. We recruited 108 exposed staff, of whom 34 were diagnosed with COVID-19. ABO blood type, titer of anti-A and -B, blood group specific alleles, and secretor status were determined. RESULTS: Blood group O was associated with lower risk of COVID-19 (OR 0.39, 95 %CI (0.16-0.92), p = 0.03) compared to non-O, i.e., blood groups A, B and AB. High titer anti-A immunoglobulin G (IgG) compared to low titer was associated with lower risk of COVID-19 (OR 0.24 95 %CI (0.07-0.78), p = 0.017). High titer of anti-B immunoglobulin M (IgM) compared to no anti-B (IgM) was associated with lower risk of COVID-19 (OR 0.16, 95 %CI (0.039-0.608), p = 0.006) and the same applies to low titer anti-B (IgM) compared to no titer (OR 0.23, 95 %CI (0.07-0.72), p = 0.012). The 33Pro variant in Integrin beta-3, that is part of human platelet antigen 1b (HPA-1b), was associated with lower risk of COVID-19 (OR 0.23, 95 %CI (0.034-0.86), p = 0.028). CONCLUSION: Our data showed that blood group O, anti-A (IgG) titer, anti-B (IgM) titer as well as HPA-1b are associated with lower risk for COVID-19.

Urea and water are transported through different pathways in the red blood cell membrane.

Several studies of the urea transporter UT-B expressed in Xenopus oocytes and in genetically modified red blood cells (RBC) have concluded that UT-B also transports water. In the present study, we use unmodified RBC to test that conclusion. We find that the permeability of urea, Pu (cm/s), has a 10-fold donor variation, while the diffusional water permeability, Pd (cm/s), remains unchanged. Additionally, we observe that phloretin inhibits Pu but not Pd, and that the time course of maximum p-chloromercuribenzosulfonate inhibition of Pu and Pd differs-Pu inhibition takes <2 min, whereas Pd inhibition requires ≥1 h of incubation. The findings in the present study are in line with a previous comparative study using unmodified RBC from four animals and a solvent drag study using human RBC, and they lead us to reject the conclusion that the UT-B transporter represents a common pathway for both solutes.

Secretor status of blood group O mothers is associated with development of ABO haemolytic disease in the newborn.

BACKGROUND AND OBJECTIVES: Identification of antibody characteristics and genetics underlying the development of maternal anti-A/B linked to inducing haemolytic disease of the foetus and newborn could contribute to the development of screening methods predicting pregnancies at risk with high diagnostic accuracy. MATERIALS AND METHODS: We examined 73 samples from mothers to 37 newborns with haemolysis (cases) and 36 without (controls). The secretor status was determined by genotyping a single nucleotide polymorphism in FUT2, rs601338 (c.428G>A). RESULTS: We found a significant association between secretor mothers and newborns developing haemolysis (p = 0.028). However, stratifying by the newborn's blood group, the association was found only in secretor mothers to blood group B newborns (p = 0.032). In fact, only secretor mothers were found in this group. By including antibody data from a previous study, we found higher median semi-quantitative levels of IgG1 and IgG3 among secretor mothers than non-secretor mothers to newborns with and without haemolysis. CONCLUSION: We found that the maternal secretor status is associated with the production of anti-A/B, pathogenic to ABO-incompatible newborns. We suggest that secretors experience hyper-immunizing events more frequently than non-secretors, leading to the production of pathogenic ABO antibodies, especially anti-B.

Associations of ABO and Rhesus D blood groups with phenome-wide disease incidence: A 41-year retrospective cohort study of 482,914 patients.

Background: Whether natural selection may have attributed to the observed blood group frequency differences between populations remains debatable. The ABO system has been associated with several diseases and recently also with susceptibility to COVID-19 infection. Associative studies of the RhD system and diseases are sparser. A large disease-wide risk analysis may further elucidate the relationship between the ABO/RhD blood groups and disease incidence. Methods: We performed a systematic log-linear quasi-Poisson regression analysis of the ABO/RhD blood groups across 1,312 phecode diagnoses. Unlike prior studies, we determined the incidence rate ratio for each individual ABO blood group relative to all other ABO blood groups as opposed to using blood group O as the reference. Moreover, we used up to 41 years of nationwide Danish follow-up data, and a disease categorization scheme specifically developed for diagnosis-wide analysis. Further, we determined associations between the ABO/RhD blood groups and the age at the first diagnosis. Estimates were adjusted for multiple testing. Results: The retrospective cohort included 482,914 Danish patients (60.4% females). The incidence rate ratios (IRRs) of 101 phecodes were found statistically significant between the ABO blood groups, while the IRRs of 28 phecodes were found statistically significant for the RhD blood group. The associations included cancers and musculoskeletal-, genitourinary-, endocrinal-, infectious-, cardiovascular-, and gastrointestinal diseases. Conclusions: We found associations of disease-wide susceptibility differences between the blood groups of the ABO and RhD systems, including cancer of the tongue, monocytic leukemia, cervical cancer, osteoarthrosis, asthma, and HIV- and hepatitis B infection. We found marginal evidence of associations between the blood groups and the age at first diagnosis. Funding: Novo Nordisk Foundation and the Innovation Fund Denmark.

Non-Invasive Fetal K Status Prediction: 7 Years of Experience.

Introduction: In the Kell blood group system, the K and k antigens are the clinically most important ones. Maternal anti-K IgG antibodies can lead to the demise of a K-positive fetus in early pregnancy. Intervention can save the fetus. Prenatal K status prediction of the fetus in early pregnancy is desirable and gives a good basis for pregnancy risk management. We present the results from 7 years of clinical experience in predicting fetal K status as well as some theoretical considerations relevant for design of the assay and evaluation of results. Methods: Blood was collected from 43 women, all immunized against K, at a mean gestational age of 18 weeks (range 10-38). A total of 56 consecutive samples were tested. The KEL *01.01 /KEL *02 single nucleotide variant that determines K status was amplified from maternal plasma DNA by PCR without allele specificity. The PCR product was sequenced by NGS technology, and the number of sequenced KEL *01.01 and KEL *02 reads were counted. Prediction of the fetal K status was based on this count and was compared with the serologically determined K status of the newborns. Results: All fetal K predictions were in accordance with postnatal serology where available (n = 34), using our current data analysis. Conclusion: We have developed an NGS-based method for the non-invasive prediction of fetal K status. This approach requires special considerations in terms of primer design, stringent preanalytical sample handling, and careful analytical procedures. We analyzed samples starting at GA 10 weeks and demonstrated the correct prediction of fetal K status. This assay enables timely clinical intervention in pregnancies at risk of hemolytic disease of the fetus and newborn caused by maternal anti-K IgG antibodies.

Exome-Based Trio Analysis for Diagnosis of the Cause of Congenital Severe Hemolytic Anemia in a Child.

Inborn hemolytic anemia requiring frequent blood transfusions can be a life-threatening disease. Treatment, besides blood transfusion, includes iron chelation for prevention of iron accumulation due to frequent blood transfusions. We present the results of a clinical investigation where the proband was diagnosed with severe hemolytic anemia of unknown origin soon after birth. Transfusion was required every 4-6 weeks. After whole exome sequencing of the proband and his parents as well as a healthy sibling, we established that the proband had a compound heterozygous state carrying two rare variants in the erythrocytic spectrin gene, SPTA1. The maternal allele was a stop mutation (rs755630903) and the paternal allele was a missense mutation (rs375506528). The healthy sibling had the paternal variant but not the maternal variant. These rare variants of SPTA1 most likely account for the hemolytic anemia. A severely reduced osmotic resistance in the erythrocytes from the proband was demonstrated. Splenectomy considerably improved the hemolytic anemia and obviated the need for blood transfusion despite the severe clinical presentation.

ABO haemolytic disease of the newborn: Improved prediction by novel integration of causative and protective factors in newborn and mother.

BACKGROUND AND OBJECTIVES: Prediction of haemolytic disease of the foetus and newborn (HDFN) caused by maternal anti-A/-B enables timely therapy, thereby preventing the development of kernicterus spectrum disorder. However, previous efforts to establish accurate prediction methods have been only modestly successful. MATERIALS AND METHODS: In a case-control study, we examined 76 samples from mothers and 76 samples from their newborns; 38 with and 38 without haemolysis. The IgG subclass profile of maternal anti-A and anti-B was determined by flow cytometry. Samples from newborns were genetically analysed for the A2 subgroup, secretor and FcγRIIa receptor alleles. RESULTS: Surprisingly, we found a correlation between the newborn secretor allele and haemolysis (p = 0.034). No correlation was found for FcγRIIa alleles. The A2 subgroup was found only in newborns without haemolysis. Unexpectedly, different reaction patterns were found for maternal anti-A and anti-B; consequently, the results were treated separately. For the prediction of haemolysis in A-newborns, the maternal IgG1 subclass determination resulted in an accuracy of 83% at birth. For B-newborns, an accuracy of 91% was achieved by the maternal IgG2 subclass determination. CONCLUSION: We improved the prediction of ABO-HDFN by characterizing maternal anti-A and anti-B by flow cytometry and we presented genetic traits in newborns with correlation to haemolysis. We propose a new understanding of A- and B-substances as immunogens that enhance the maternal immune response and protect the newborn, and we suggest that the development of ABO-HDFN is different when caused by maternal anti-A compared to maternal anti-B.

Managing patients on direct factor Xa inhibitors with rapid thrombelastography.

The use of direct factor Xa inhibitors rivaroxaban and apixaban (XABANs) has rapidly increased; however, there is no validated test available to monitor the effect on hemostasis. This study aims to assess how hemostatic management based on the Rapid Thromboelastography (R-TEG) variable activated clotting time (ACT) of XABAN patients with ongoing bleedings or in need for acute surgical intervention, affected patient outcome. A total of 343 XABAN patients were included in the main analysis together with 50 healthy volunteers to validate the reference value for ACT. An ACT >120 s (s) was defined as having XABAN-induced coagulopathy. Sixty-five percent of the XABAN patients presented with R-TEG ACT within the normal reference. Patients with XABAN-induced coagulopathy had a significantly increased risk of severe bleeding. Significantly more patients with extra-cerebral bleeding (ECB) and ACT above 120 s were transfused with five red blood cell (RBC) units or more compared to patients with ACT at 120 s or below (17% vs. 3%, p <.05). Significantly more XABAN-patients with ACT above 120 s received pro-hemostatic intervention with prothrombin complex concentrate (PCC) when compared to those with ACT at 120 s or below (ECB: 2% vs. 8%, p =.03, intracranial hemorrhage: 25% vs. 68%, p <.00). Patients who received PCC had a higher 30- and 90-day mortality compared to the rest of the cohort (16% vs. 6%, p = .02 and 21% vs. 7%, p =.00). Patients with XABAN-induced coagulopathy as evaluated by R-TEG ACT presented with more severe bleeding and higher transfusion requirements when compared to those with ACT in the normal range. This suggests that R-TEG ACT measurement in XABAN patients with active hemorrhage or in need for acute surgery may be of clinical value.