ZusammenfassungDie fetale und neonatale Alloimmunthrombozytopenie (FNAIT) wird durch mütterliche Antikörper hervorgerufen, die gegen ein vom Vater ererbtes Blutgruppenmerkmal an fetalen Thrombozyten gerichtet sind. Während Teil 1 des Beitrags Ausgabe die Ätiologie, die Pathogenese und die Diagnostik der FNAIT thematisiert hatte, widmet sich dieser 2. Teil der Risikostratifizierung und Behandlung 1.

The transport of maternal IgG antibodies into the fetal circulation provides the offspring with passive humoral immunity. The transplacental transport of IgG antibodies takes place in a complex process in which the neonatal receptor for the crystallizable fragment of IgG molecules (fragment cristallizable receptor neonatal, FcRn) is essentially involved. FcRn is ubiquitously expressed in the adult organism, regulates IgG and albumin homeostasis as well as innate and adaptive immunity against IgG immune complexes and is thus involved in the defense against infectious diseases and anti-tumor immunity. Therapeutic FcRn antagonists inhibit the recycling of IgG molecules and lead to a reduction in IgG serum levels. In the context of pregnancy, therapeutic FcRn antagonists inhibit transplacental IgG transport. This review is intended to present the status of the potential use of FcRn antagonists in immunohematological diseases caused by autoantibodies and in diseases of the fetus and newborn caused by maternal alloantibodies.

Zusammenfassung Durch den Transport von mütterlichen IgG-Antikörpern in die Zirkulation des Fetus erhalten die Nachkommen den humoralen „Nestschutz“ bzw. die mütterliche „Leihimmunität“. Der transplazentare Transport von IgG-Antikörpern erfolgt in einem komplexen Prozess, an dem der neonatale Rezeptor für das kristallisierbare Fragment von IgG-Molekülen (Fragment cristallizable receptor neonatal, FcRn) essenziell beteiligt ist. FcRn ist im adulten Organismus ubiquitär exprimiert, reguliert die IgG- und Albumin-Homöostase, sowie die angeborene und adaptive Immunität gegen IgG-Immunkomplexe und ist damit an der Abwehr infektiöser Erkrankungen und der Anti-Tumor-Immunität beteiligt. Therapeutische FcRn-Antagonisten blockieren das Recycling von IgG-Molekülen und führen zu einer Absenkung der IgG-Serumspiegel. Im Rahmen einer Schwangerschaft blockieren therapeutische FcRn-Antagonisten den transplazentaren IgG-Transport. Die vorliegende Übersichtsarbeit soll den aktuellen Stand der potenziellen Anwendung von FcRn-Antagonisten bei immunhämatologischen Erkrankungen durch Autoantikörper sowie im Rahmen von Erkrankungen des Fetus und Neugeborenen durch mütterliche Alloantikörper darstellen.

Zusammenfassung In der vorliegenden Übersichtsarbeit diskutieren wir, wie iatrogene Blutverluste durch immunhämatologische Untersuchungen bei Früh- und Reifgeborenen minimiert werden können. Die Hauptursache für die Transfusion von Erythrozytenkonzentraten (EK) sind dabei iatrogene Blutverluste durch diagnostische Blutentnahmen. In einer Beobachtungsstudie betrug der iatrogene Blutverlust bei Frühgeborenen in den ersten 28 Lebenstagen im Median 24,2 mL/kg im Vergleich zum transfundierten EK-Volumen von 30 mL/kg im selben Zeitraum 1. Die Reduktion von diagnostischen Blutentnahmen stellt somit eine effiziente Maßnahme zur Reduzierung von EK-Transfusionen bei Frühgeborenen dar. Rationale und bedarfsadaptierte immunhämatologische Untersuchungen können den Blutverlust reduzieren, Transfusionen vermeiden und die entstehenden Kosten senken. Ferner stellen wir dar, wie durch differenzierte Auswahl von Blutkomponenten das Auftreten unerwünschter Ereignisse bei der Transfusion von Neugeborenen verhindert werden kann und regen an, restriktivere Indikationsstellungen zur Transfusion insbesondere in der Frühgeborenenmedizin zu diskutieren.

Pemphigus vulgaris (PV) is an autoimmune blistering disorder of the skin and/or mucous membranes caused by IgG autoantibodies that predominantly target two transmembrane desmosomal cadherins: desmoglein (DSG)1 and DSG3. DSG-specific T cells play a central role in PV pathogenesis because they provide help to autoreactive B cells for autoantibody production. In this study, we characterized DSG3-specific peripheral T cells in a cohort of 52 patients with PV and 41 healthy controls with regard to cytokine profile and epitope specificity. By ELISpot analysis, type 2 T cells reactive with the DSG3 ectodomain were significantly increased in patients with PV compared with those in healthy controls. By dextramer analysis, CD4 thorn T cells specific for an epitope within the extracellular domain of DSG3, DSG3(206-220), were found at significantly higher frequencies in patients with PV than in HLA-matched healthy controls. T-cell recognition of two distinct DSG3 epitopes, that is, DSG3(206-220) and DSG3(378-392), correlated significantly, suggesting a synergistic effect in B-cell help. Immunization of HLADRB1*04:02-transgenic mice with PV with the same set of DSG3 peptides induced pathogenic DSG3-specific IgG antibodies, which induced loss of keratinocyte adhesion in vitro. Thus, DSG3 peptide-specific T cells are of particular interest as surrogate markers of disease activity and potential therapeutic targets in PV.

In the present review, we discuss how iatrogenic blood loss through immunohematologic testing can be minimized in preterm and mature infants. In this context, the main cause of red blood cell (RBC) concentrate transfusion is iatrogenic blood loss due to diagnostic blood sampling. In an observational study, the median iatrogenic blood loss in preterm infants during the first 28 days of life was 24.2 mL/kg compared with the transfused RBC volume of 30 mL/kg during the same period 1 . Thus, reduction of diagnostic blood sampling is an efficient measure to reduce RBC concentrate transfusions in preterm infants. Evidence-based immunohematologic testing can reduce blood loss, avoid transfusions, and reduce the costs incurred. Furthermore, we present how the occurrence of adverse events in neonatal transfusion can be prevented by evidence-based selection of blood components and suggest discussing more restrictive indications for transfusion, especially in premature infants.

In fetal/neonatal alloimmune thrombocytopenia (FNAIT), maternal alloantibodies against paternal human platelet antigens (HPA) cross the placenta and lead to platelet destruction. The extent of thrombocytopenia varies among neonates, and inflammation may constitute an important trigger. A set of stable inflammatory markers was measured in serum samples from neonates with low platelet counts, of which n = 50 were diagnosed with FNAIT due to anti-HPA-1a antibodies and n = 50 were thrombocytopenic without detectable maternal HPA antibodies. Concentrations of C-reactive protein, soluble CD14, procalcitonin, and sFlt-1 did not differ between the two cohorts. There was no correlation between C-reactive protein or soluble CD14 and the platelet count, but a negative correlation between procalcitonin concentrations and the neonatal platelet count in both cohorts. sFlt-1 concentration and the platelet count were correlated in FNAIT cases exclusively. None of the inflammatory markers was statistically different between cases with and without intracranial haemorrhage. We were unable to identify systemic inflammation as a relevant factor for thrombocytopenia in FNAIT. The antiangiogenic enzyme sFlt-1, released by the placenta, did correlate with the platelet count in FNAIT cases. Our findings may give rise to the hypothesis that placental inflammation rather than systemic inflammation modulates disease severity in FNAIT.

Most cases of fetal and neonatal thrombocytopenia (FNAIT) are caused by maternal anti-human platelet antigen-1a antibodies (anti-HPA-1a). Anti-HPA-5b antibodies are the second most common antibodies in suspected FNAIT cases. Given the high prevalence of anti-HPA-5b antibodies in pregnant women delivering healthy newborns, the association with FNAIT may be coincidental. This review of the literature related to FNAIT using the MEDLINE database was conducted according to PRISMA guidelines. A retrospective analysis of a single-centre cohort of 817 suspected FNAIT cases was conducted. The pooled prevalence of anti-HPA-5b antibodies in unselected pregnant women of European descent was 1.96% (n = 3113), compared with 3.4% (n = 5003) in women with suspected FNAIT. We found weak evidence that a small proportion of pregnant women presenting with anti-HPA-5b antibodies will give birth to a newborn with mild thrombocytopenia. The neonatal platelet counts were not different between suspected FNAIT cases (n = 817) with and without maternal anti-HPA-5b antibodies. The prevalence of maternal anti-HPA-5b antibodies was not different between neonates with intracranial haemorrhage and healthy controls. The current experimental and epidemiological evidence does not support the hypothesis that anti-HPA-5b antibodies cause severe thrombocytopenia or bleeding complications in the fetus or newborn.

(1) Background: ABO blood group incompatibility between the mother and fetus protects against anti-D immunization by pregnancy. The possible role of ABO incompatibility in protecting against anti-human platelet antigen-1a immunization is unclear. (2) Methods: This study retrospectively screened 817 families (mother-father-neonate trios) of suspected fetal and neonatal alloimmune thrombocytopenia for inclusion. ABO genotypes were determined in 118 mother-child pairs with confirmed alloimmune thrombocytopenia due to anti-HPA-1a antibodies, and 522 mother-child pairs served as the control group. The expression of blood group antigen A on platelets was determined in 199 consecutive newborns by flow cytometry and compared with adult controls. (3) Results: ABO incompatibility between mother and fetus did not protect against anti-human platelet antigen-1a immunization by pregnancy. ABO blood groups of mothers and/or fetuses were not associated with the severity of fetal and neonatal alloimmune thrombocytopenia. The expression pattern of blood group A antigens on the platelets of newborns mirrored that of adults, albeit on a lower level. Blood group A antigen was detected on a subpopulation of neonatal platelets, and some newborns revealed high platelet expression of A determinants on all platelets (type II high-expressers). (4) Conclusion: The lack of a protective effect of ABO incompatibility between mother and fetus against anti-human platelet antigen-1a immunization by pregnancy may indicate that fetal platelets are not the cellular source by which the mother is immunized.

Summary The causative role of maternal, anti‐human leukocyte antigen (anti‐HLA) class I antibodies in foetal and neonatal alloimmune thrombocytopenia (FNAIT) remains controversial. Furthermore, in FNAIT cases caused by anti‐human platelet antigen‐1a (anti‐HPA‐1a) antibodies, the possible additive effect of maternal anti‐HLA class I antibodies on outcomes is unclear. Among 817 mother–father–neonate trios of suspected FNAIT, we assessed the possible association of maternal anti‐HLA class I antibodies with neonatal platelet count, and the incidence of FNAIT caused by anti‐HPA‐1a antibodies. In 144 FNAIT cases caused by anti‐HPA‐1a antibodies, we investigated the possible association of maternal anti‐HLA class I antibodies with neonatal platelet count, birth weight, and the incidence of intracranial haemorrhage ( n = 16). Maternal anti‐HLA class I antibodies were not associated with neonatal platelet count in suspected cases of FNAIT. There was no significant interaction between the presence of anti‐HLA class I antibodies and anti‐HPA‐1a antibodies. In FNAIT cases caused by anti‐HPA‐1a antibodies, there was no association between the presence of anti‐HLA class I antibodies and neonatal platelet count, birth weight, or occurrence of intracranial haemorrhage. This study’s findings do not support the concept that maternal anti‐HLA class I antibodies represent a risk factor of FNAIT or disease severity.

Individuals with the Bombay phenotype (Oh) in the ABO blood group system do not express the H, A, and B antigens but have no clinical symptoms. Bombay phenotype with clinical symptoms has been described in leukocyte adhesion deficiency type II (LAD II), a fucosylation disorder caused by mutations in SLC35C1. Only few LAD II patients have been described so far. Here we describe an additional patient, a 22-year old male, born to unrelated parents, presenting with inflammatory skin disease, periodontitis, growth, and mental retardation, admitted to the department of dentistry for treatment under general anesthesia. Pre-operative routine investigations revealed the presence of the Bombay phenotype (Oh). Genomic sequencing identified two novel triplet deletions of the SLC35C1 gene. Functional investigations confirmed the diagnosis of LAD II. Therapy with oral fucose led to the disappearance of the chronic skin infections and improvements in behavior and attention span.

Introduction In pregnant women with a history of fetal and neonatal alloimmune thrombocytopenia (FNAIT), prenatal intervention in subsequent pregnancies may be required to prevent fetal bleeding. Several invasive and non-invasive protocols have been published: amniocentesis for fetal genotyping, fetal blood sampling for the determination of fetal platelet count, intrauterine platelet transfusions, and weekly maternal i.v. immunoglobulin (IVIG) infusion with or without additional corticosteroid therapy. This is the first retrospective study that report the experience with a non-invasive protocol focused on side effects of maternal IVIG treatment and neonatal outcome. Methods Pregnant women with proven FNAIT in history and an antigen positive fetus were treated with IVIG (1 g/kg/bw) every week. To identify potential IVIG-related hemolytic reactions isoagglutinin titer of each IVIG lot and maternal blood count were controlled. IVIG-related side effects were prospectively documented and evaluated. Furthermore, ultrasound examination of the fetus was performed before starting IVIG administration and continued regularly during treatment. Outcome of the index and subsequent pregnancy was compared. Corresponding data of the newborns were analyzed simultaneously. Results IVIG was started at 20 weeks of gestation (median). Compared to the index pregnancy, platelet counts of the newborns were higher in all cases. No intracranial hemorrhage occurred (Index pregnancies: 1 case). Platelet counts were 187 × 10 9 /l (median, range 22–239, 95% CI) and one newborn had mild bleeding. No severe hemolytic reaction was observed and side effects were moderate. Conclusion Among pregnant women with FNAIT history, the use of non-invasive fetal risk determination and maternal IVIG resulted in favorite outcome of all newborns. Invasive diagnostic or therapeutic procedures in women with a history of FNAIT should be abandoned.

Hemolytic disease of the fetus and newborn and fetal and neonatal alloimmune thrombocytopenia are caused by maternal antibodies against fetal alloantigens on red blood cells or platelets that are inherited from the father. After transplacental transport to the fetal circulation, antibodies of the IgG class may cause severe fetal anemia or bleeding complications. The indication for noninvasive fetal blood group genotyping is given if a clinically relevant antibody is detected in a pregnant woman and if the father is heterozygous (or unknown) for the implicated blood group allele. This mini-review will focus on the advantages and current limitations of next-generation sequencing (NGS) for noninvasive diagnosis of fetal blood groups which is, in contrast to fetal aneuploidy screening, proposed only by some research groups. Targeted massively parallel sequencing of short DNA fragments from maternal cell-free plasma samples enables counting of fetal alleles for many single nucleotide polymorphisms in parallel. This information can be utilized for estimation of the fetal fraction of cell-free DNA (cfDNA) as well as detection of the paternal blood group allele in question. Adherence to a cut-off of ≥4% fetal fraction for reporting conclusive results is recommended to avoid false-negative results due to low fetal fraction. For screening purposes of fetal RHD in RhD-negative pregnant women, real-time PCR methods are very well established. However, for diagnostic purposes, the targeted amplicon-based NGS approach has the inherent capability to estimate the fetal fraction of cfDNA. In the future, improving the accuracy of NGS by consensus sequencing of single cfDNA molecules may enable reliable fetal blood group genotyping already in the first trimester of pregnancy.

The degree and type of T cell infiltration influence rectal cancer prognosis regardless of classical tumor staging. We asked whether clonal expansion and tumor infiltration are restricted to selected-phenotype T cells; which clones are accessible in peripheral blood; and what the spatial distribution of their target antigens is. From five rectal cancer patients, we isolated paired tumor-infiltrating T cells (TILs) and T cells from unaffected rectum mucosa (TUM) using 13-parameter FACS single cell index sorting. TCRαβ sequences, cytokine, and transcription factor expression were determined with single cell sequencing. TILs and TUM occupied distinct phenotype compartments and clonal expansion predominantly occurred within CD8+ T cells. Expanded TIL clones identified by paired TCRαβ sequencing and exclusively detectable in the tumor showed characteristic PD-1 and TIM-3 expression. TCRβ repertoire sequencing identified 49 out of 149 expanded TIL clones circulating in peripheral blood and 41 (84%) of these were PD-1− TIM-3−. To determine whether clonal expansion of predominantly tumor-infiltrating T cell clones was driven by antigens uniquely presented in tumor tissue, selected TCRs were reconstructed and incubated with cells isolated from corresponding tumor or unaffected mucosa. The majority of clones exclusively detected in the tumor recognized antigen at both sites. In summary, rectal cancer is infiltrated with expanded distinct-phenotype T cell clones that either i) predominantly infiltrate the tumor, ii) predominantly infiltrate the unaffected mucosa, or iii) overlap between tumor, unaffected mucosa, and peripheral blood. However, the target antigens of predominantly tumor-infiltrating TIL clones do not appear to be restricted to tumor tissue.

BACKGROUNDMost cases of fetal and neonatal alloimmune thrombocytopenia (FNAIT) are caused by maternal alloantibodies against human platelet antigen‐1a (HPA‐1a). Alloimmunization mainly occurs in HPA‐1a–negative mothers who are carriers of the HLA‐DRB3*01:01 allele. Recently, it has been reported that the combined presence of HLA‐DRB3*01:01 and HLA‐DRB4*01:01P was associated with severity of FNAIT. We tested this hypothesis by analyzing a large cohort of cases and controls.STUDY DESIGN AND METHODSIn total, 101 mothers with a history of FNAIT caused by anti‐HPA‐1a were investigated. HLA‐DRB1, ‐DRB3, ‐DRB4, and ‐DRB5 genotypes were determined by Luminex technology. Haplotype frequencies were compared between cases and 100 controls. The platelet (PLT) counts of neonates and the incidence of intracranial hemorrhage (ICH) were compared between subgroups defined by genotype.RESULTSOf the HPA‐1a–immunized mothers, 98% (99/101) carried at least one copy of HLA‐DRB3*01:01. Carriage of HLA‐DRB3*01:01 was significantly associated with immune response to HPA‐1a (odds ratio, 92.3; 95% confidence interval, 26.9‐317.1; p = 1.34 × 10−12). No association between HLA‐DRB3*01:01 and HLA‐DRB4*01:01P alone or in combination with the PLT count of the newborns or the incidence of ICH was detected.CONCLUSIONIn contrast to HLA‐DRB4*01:01P, the inheritance of HLA‐DRB3*01:01 is strongly associated with the propensity for mounting a humoral immune response against fetal HPA‐1a antigen. Neither a homozygous nor a compound heterozygous gene dose predicts the severity of the disease. Testing for the presence of HLA‐DRB3*01:01 might be very useful in counseling women at risk of FNAIT due to anti‐HPA‐1a.