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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., Competing Interests: Declaration of interests J.S. is the deputy CEO and 50% owner of BLUsang AB. He holds patents on Vel genotyping (inventors: Jill Storry, Magnus Jöud, Björn Nilsson, and Martin L. Olsson). J.S. has received speaker fees, royalties, and honoraria from the following companies: Grifols Diagnostic Solutions, QuidelOrtho Inc., and Biorad Laboratories. J.S. receives an honorarium for Section Editor work, Vox Sanguinis from John Wiley & Sons Ltd. J.S. is Vice President of the International Society of Blood Transfusion and married to Professor M.L.O. M.L.O. is CEO and 50% owner of BLUsang AB. M.L.O. holds patents on Vel genotyping (inventors: Jill Storry, Magnus Jöud, Björn Nilsson, and Martin L. Olsson). M.L.O. received speaker fees, royalties, and honoraria from the following companies: Grifols Diagnostic Solutions, QuidelOrtho Inc., and Biorad Laboratories. M.L.O. is married to Adjunct Professor J.S. W.N.E. is chair of the International Council for Standardization in Haematology. W.N.E. works as advisor for Scorpio Labs and is on the editorial board of the Journal of Clinical Pathology. W.H.O. is chair of the Blood Transfusion Genomics Consortium. W.H.O. is in receipt of an educational/research grant from Thermo Fisher Scientific. N.G. offers scientific consulting services to Thermo Fisher Scientific., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

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., (© 2024. The Author(s).)

A central issue in immunology is the immunological response against nonself. The prerequisite for a specific immunological response is the exposure to the immune system of a nonself antigen. Mathematical equations are presented, which define the fraction of all outcomes with a nonself allele in biallelic systems at the population level in pregnancy and transfusion/transplantation medicine. When designing assays, the mathematical descriptions can be used for estimating the number of genetic markers necessary to obtain a predetermined probability level in detecting nonself alleles of a given frequency. For instance, the equations can be helpful in the design of assays, where the nonself allele can be detected by analysis of cfDNA in plasma from pregnant women, to estimate fetal fraction or to monitor changes in cfDNA in plasma of transplantation patients. The equations give exact, quantitative descriptions of all nonself situations in pregnancy and transfusion/transplantation., (© 2024 The Author(s). APMIS published by John Wiley & Sons Ltd on behalf of Scandinavian Societies for Pathology, Medical Microbiology and Immunology.)

Deep sequencing by NGS of targeted amplicons can identify rare genetic variants in a pool of DNA where the vast majority of genomic DNA does not contain the variant. This approach can be used to detect a previously described paternally inherited, fetal variant in cell-free DNA (cfDNA) in maternal plasma. This is useful in cases where risk for the fetus is contingent upon inheritance of a paternal variant that the woman does not have. Both pathogenic and non-pathogenic variants that the woman does not have can be detected. In cases of compound heterozygosity, presence of the paternal pathogenic variant also requires detection of the maternal variant for risk assessment, which requires a chorion villus biopsy.We have used this approach to focus on detection of fetal blood groups in cases of presence of maternal alloantibodies against blood group antigens in pregnancy, to predict whether the fetus has inherited a blood group antigen that is targeted by the alloantibodies. In cases of maternal alloantibodies against blood group antigens, the fetus is at risk of hemolytic disease of the fetus and newborn (HDFN). With a known specificity of the maternal antibodies and if the fetal blood group can be determined in the pregnancy, then it can be ascertained if the fetus is at risk of HDFN and rational pregnancy care can be instituted. A noninvasive procedure avoids risks for the fetus. We have reported a procedure based on NGS analysis of PCR amplified cfDNA from maternal plasma. Some fetuses may die as early as week 18. We use this approach to predict fetal K, k, RhC, Rhc, RhE, and ABO blood groups in cases with a risk of HDFN due to the corresponding maternally produced antibodies.The NGS-based analysis can predict the presence or absence of incompatible antigens on the fetal RBCs.In this chapter, a noninvasive method for predicting some fetal blood groups early in pregnancy is described. There is a clinical need for such assays, and they may be a useful tool for management of pregnancies complicated by these alloantibodies within the field of precision medicine., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

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., (© 2023 International Society of Blood Transfusion.)

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., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Rieneck et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Aim: To study the effect of sulfatide on gene expression and proliferation of human primary fibroblasts induced by insulin, insulin-like growth factor-1 and human growth hormone., Materials and Methods: Human primary fibroblasts were exposed to 1, 3 and 30 μM of sulfatide or its precursor galactosylceramide (GalCer). Proliferation was determined by 3 H-thymidine incorporation and gene expression via microarray analysis., Results: Sulfatide and GalCer reduced the growth rate of fibroblasts by 32%-82% when exposed to 0.5 nM insulin. After challenge with 120 μM of H 2 O 2 , sulfatide reduced membrane leakage. Fibroblast gene expression was altered by sulfatide in gene pathways associated with cell cycle/growth, transforming growth factor-β function, and encoding of proteins involved in intracellular signalling. NFKBIA, a key control element in NF-кB regulation, was decreased 2-fold by sulfatide., Conclusions: Sulfatide strongly inhibits fibroblast growth. We therefore suggest the addition of sulfatide to injectable commercial insulin formulations, which would reduce adverse fibroblast growth and improve well-being in patients with diabetes., (© 2023 The Authors. Diabetes, Obesity and Metabolism published by John Wiley & Sons Ltd.)

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., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier GmbH. All rights reserved.)

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., Competing Interests: The authors have no conflicts of interest to declare., (Copyright © 2022 by The Author(s). Published by S. Karger AG, Basel.)

Quantitative methods for assessing differentiative potency of adipose-derived stem/stromal cells may lead to improved clinical application of this multipotent stem cell, by advancing our understanding of specific processes such as adipogenic differentiation. Conventional cell staining methods are used to determine the formation of adipose areas during adipogenesis as a qualitative representation of adipogenic potency. Staining methods such as oil-red-O are quantifiable using absorbance measurements, but these assays are time and material consuming. Detection methods for cell characteristics using advanced image analysis by machine learning are emerging. Here, live-cell imaging was combined with a deep learning-based detection tool to quantify the presence of adipose areas and lipid droplet formation during adipogenic differentiation of adipose-derived stem/stromal cells. Different detection masks quantified adipose area and lipid droplet formation at different time points indicating kinetics of adipogenesis and showed differences between individual donors. Whereas CEBPA and PPARG expression seems to precede the increase in adipose area and lipid droplets, it might be able to predict expression of ADIPOQ . The applied method is a proof of concept, demonstrating that deep learning methods can be used to investigate adipogenic differentiation and kinetics in vitro using specific detection masks based on algorithm produced from annotation of image data.

Background: The Rh blood group system has considerable clinical importance. The C, c, and E antigens are targets of alloantibodies. Anti-C, anti-c or anti-E alloreactive antibodies produced in pregnant women can cause anemia of a fetus carrying the corresponding antigens., Aims: Based on NGS technology, we have developed a noninvasive diagnostic assay to predict the fetal blood group of C, c or E antigens by sequencing cell-free DNA (cfDNA) during pregnancy., Materials and Methods: The SNVs underlying either the C, c or E antigens were PCR amplified and sequenced using NGS on a MiSeq instrument. The DNA sequences encoding the C, c or E antigen were counted, as were the number of total sequences. Based on the percentage of fetally derived target SNVs inherited from the father, the fetal blood group could be predicted., Results: The results of 55 consecutive RHCE prenatal analyses with postnatal serological blood group determination of 30 newborns showed no discordant results. A threshold discerning positive from negative samples was set at 0.05% specific reads., Discussion: Noninvasive, prenatal prediction of fetal blood groups by sequencing cfDNA for the detection of low-level RHCE*C, RHCE*c and RHCE*E sequences was established as an accurate and robust assay applicable for use in clinical settings., (© 2021 John Wiley & Sons Ltd.)

Background: Laboratory monitoring of mother, fetus, and newborn in hemolytic disease of fetus and newborn (HDFN) aims to guide clinicians and the immunized women to focus on the most serious problems of alloimmunization and thus minimize the consequences of HDFN in general and of anti-D in particular. Here, we present the current approach of laboratory screening and testing for prevention and monitoring of HDFN at the Copenhagen University Hospital in Denmark., Summary: All pregnant women are typed and screened in the 1st trimester. This serves to identify the RhD-negative pregnant women who at gestational age (GA) of 25 weeks are offered a second screen test and a non-invasive fetal RhD prediction. At GA 29 weeks, and again after delivery, non-immunized RhD-negative women carrying an RhD-positive fetus are offered Rh immunoglobulin. If the 1st trimester screen reveals an alloantibody, antenatal investigation is initiated. This also includes RhD-positive women with alloantibodies. Specificity and titer are determined, the fetal phenotype is predicted by non-invasive genotyping based on cell-free DNA (RhD, K, Rhc, RhC, RhE, ABO), and serial monitoring of titer commences. Based on titers and specificity, monitoring with serial peak systolic velocity measurements in the fetal middle cerebral artery to detect anemia will take place. Intrauterine transfusion is given when fetal anemia is suspected. Monitoring of the newborn by titer and survival of fetal red blood cells by flow cytometry will help predict the length of the recovery of the newborn., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 by S. Karger AG, Basel.)