Fetal and neonatal alloimmune thrombocytopenia: No evidence of systemic inflammation as a modulator of disease severity. Could placental inflammation be key?

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.

Severe intracranial haemorrhage in neonatal alloimmune thrombocytopenia due to antibodies against human platelet antigen 1b: Case report and literature review.

Fetal/neonatal alloimmune thrombocytopenia (FNAIT) is a rare life-threatening disorder, leading to severe thrombocytopenia and potentially bleeding, with intracranial haemorrhage (ICH) being the most serious complication. We report on a FNAIT case with fourth-degree ICH that arose due to antibodies against human platelet antigen (HPA)-1b. The male infant, born to an otherwise healthy mother, presented with severe signs of ICH soon after delivery. Since only moderate thrombocytopenia was noted and there were no active signs of bleeding, the infant did not receive intravenous immunoglobulins (IVIg) or platelet transfusion. Spontaneous recovery of platelets was noted on the eighth day of life, but permanent neurological impairment remained as a consequence of ICH. We report the results of HPA and human leukocyte antigen (HLA) antibodies in the mother's and the infant's sera, the family's HPA genotype and the mother's HLA genotype, and summarise previously described cases of FNAIT due to anti-HPA-1b antibodies in the literature. FNAIT with severe ICH due to anti-HPA-1b antibodies is rarely diagnosed. An association between HLA genes and sensitization to HPA-1b antibodies was not demonstrated. The severity of FNAIT and the occurrence of ICH is often difficult to predict. In this case, the infant presented with moderate thrombocytopenia and ICH, with subsequent permanent consequences.

Maternal Anti-HPA-1a Antibodies Increase Endothelial Cell Apoptosis and Permeability.

Intracranial hemorrhage (ICH) associated with fetal/neonatal alloimmune thrombocytopenia (FNAIT) is attributed mainly to endothelial damage caused by binding of maternal anti-HPA-1a antibodies to the αvß3 integrin on endothelial cells (ECs). We examined the effect of anti-HPA-1a antibodies on EC function using 2 EC lines from different vascular beds, HMVEC of dermal origin and hCMEC/D3 of cerebral origin. Anti-HPA-1a sera significantly increased apoptosis in both HMVEC and hCMEC/D3 cells and permeability in hCMEC/D3 cells only. This increase in both apoptosis and permeability was significantly inhibited by a monoclonal anti-ß3 antibody (SZ21) binding to the HPA-1a epitope. Our results indicate that (1) maternal anti-HPA-1a antibodies impair EC function by increasing apoptosis and permeability and (2) ECs from different vascular beds vary in their susceptibility to pathological effects elicited by maternal anti-HPA-1a antibodies on EC permeability. Examination of maternal anti-HPA-1a antibodies for their effect on EC permeability may predict potential ICH associated with FNAIT.

Placental Complement Activation in Fetal and Neonatal Alloimmune Thrombocytopenia: An Observational Study.

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a disease that causes thrombocytopenia and a risk of bleeding in the (unborn) child that result from maternal alloantibodies directed against fetal, paternally inherited, human platelet antigens (HPA). It is hypothesized that these alloantibodies can also bind to the placenta, causing placental damage. This study aims to explore signs of antibody-mediated placental damage in FNAIT. We performed a retrospective study that included pregnant women, their newborns, and placentas. It comprised 23 FNAIT cases, of which nine were newly diagnosed (14 samples) and 14 were antenatally treated with intravenous immune globulins (IVIg) (21 samples), and 20 controls, of which 10 had anti-HLA-class I antibodies. Clinical information was collected from medical records. Placental samples were stained for complement activation markers (C1q, C4d, SC5b-9, and mannose-binding lectin) using immunohistochemistry. Histopathology was examined according to the Amsterdam criteria. A higher degree of C4d deposition was present in the newly diagnosed FNAIT cases (10/14 samples), as compared to the IVIg-treated FNAIT cases (2/21 samples, p = 0.002) and anti-HLA-negative controls (3/20 samples, p = 0.006). A histopathological examination showed delayed maturation in four (44%) placentas in the newly diagnosed FNAIT cases, five (36%) in the IVIg-treated FNAIT cases, and one in the controls (NS). C4d deposition at the syncytiotrophoblast was present in combination with low-grade villitis of unknown etiology in three newly diagnosed FNAIT cases that were born SGA. We conclude that a higher degree of classical pathway-induced complement activation is present in placentas from pregnancies with untreated FNAIT. This may affect placental function and fetal growth.

Prospects for risk stratification of anti-HPA-1a alloimmunized pregnant women.

A diagnosis of fetal/neonatal alloimmune thrombocytopenia (FNAIT) is made if a platelet-specific antibody is detected in the mother and the fetus or newborn carries the cognate antigen. Some children will experience very low platelet counts or even intracranial hemorrhage with devastating consequences, whereas others are largely unaffected. At the moment, predictive tools to forecast the severity of FNAIT during pregnancy are not available and over- or under-treatment may put the mother or the fetus at risk. A number of potential modulators of FNAIT severity have been reported. Maternal immune responses differ in respect to the IgG subtype composition, the glycosylation pattern of the antibodies, their fine specificity, and their functional effects on platelets, the trophoblast, and endothelial cells. In addition, antibody levels are variable. The efficacy of IgG transfer and, on the fetal side, gender and inflammatory responses, were also investigated for their potential impact on FNAIT severity. These potential risk modulators are scrutinized for available experimental and clinical evidence. Antibody glycosylation and anti-endothelial activity are hot candidates which, most likely in conjunction with the antibody level, should be explored further as tools to stratify fetal risk.

Management and Neonatal Outcomes of Pregnancies with Fetal/Neonatal Alloimmune Thrombocytopenia: A Single-Center Retrospective Cohort Study.

BACKGROUND: There is no consensus regarding the optimal antenatal treatment of fetal/neonatal alloimmune thrombocytopenia (F/NAIT). We aimed to review the fetal blood sampling (FBS)-related risk, fetal response to maternal intravenous immunoglobulin (IVIG), and cesarean section (CS) rate in pregnancies with a history of F/NAIT. METHODS: Maternal demographics, alloantibodies, pregnancy management, fetal and neonatal outcomes, and index case characteristics were collected. Responders (R) and non-responders (NR) were defined as women treated with IVIG in whom fetal platelets (PLTs) were normal or low (< 50 × 109/L). RESULTS: An FBS-related risk occurred in 1.6% (2/119) of procedures. Maternal characteristics did not differ between responders (n = 21) and non-responders (n = 21). HPA-1a antibody was detected in all non-responders and in 72% of responders (p < 0.01). The index case had a significantly lower PLT count at birth in non-responders versus responders (median PLT count: R = 20 × 109/L [IQR 8-43] vs. NR = 9 × 109/L [IQR 4-18], p < 0.02). No differences were found in IVIG treatment duration or dosage. PLTs at birth were significantly lower in non-responders compared to responders. No intracranial hemorrhages occurred. CSs were performed for obstetric indications only in all but two cases. CONCLUSION: Maternal IVIG can elicit different fetal responses. The lack of prognostic factors to predict responders or non-responders suggests that there remains a role for FBS in F/NAIT in experienced hands.

Sensitivity of assays for the detection of HPA-1a antibodies: results of an international workshop demonstrating the impact of cation chelation from integrin αIIbß3 on three widely used assays.

BACKGROUND AND OBJECTIVES: HPA-1a antibodies account for 70-80% of cases of fetal-neonatal alloimmune thrombocytopenia (FNAIT) in Caucasians. However, numerous workshops have demonstrated variability in their detection. We recently showed that exposure of αIIbß3 to ethylene diamine tetraacetic acid (EDTA) affected binding of many anti-αIIbß3 monoclonal, and HPA-1a allo-, antibodies; this adversely affected sensitivity of the monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay and indirect platelet immunofluorescence test (PIFT). This study presents results from an international workshop studying the impact of cation chelation on HPA-1a antibody detection in routine diagnostic laboratories. MATERIALS AND METHODS: Serum and EDTA-anticoagulated plasma samples containing anti-HPA-1a were distributed to 39 laboratories. Participants were asked to detect and identify any HPA antibodies present. RESULTS: 2/39 (5.1%) participants were able to detect and identify anti-HPA-1a in the serum, but not in the plasma sample. EDTA plasma reduced MAIPA assay sensitivity by ≥ 20% in 17/24 (70.8%) laboratories and by ≥ 50% in 9/24 (37.5%) when using HPA-1a1a platelets (mean: 27.7%, range 0-85.1%); when using HPA-1a1b platelets 3/4 (75%), participants reported ≥ 50% loss of sensitivity (mean 65.6%, range 0-96.6%). A small but significant increase in optical densities was observed in antigen capture ELISA assays when using plasma (mean difference: 0.081, P < 0.01). Insufficient PIFT data were returned to draw firm conclusions. CONCLUSION: Use of EDTA plasma significantly affects the sensitivity of the MAIPA assay and can affect detection of even potent, FNAIT-causing examples of anti-HPA-1a. These data highlight the importance of use of αIIbß3 in an appropriate conformation for the sensitive detection of anti-HPA-1a.

[Serological Diagnosis and Clinical Data Analysis of Neonatal Alloimmune Thrombocytopenia].

OBJECTIVE: To investigate the pathogenesis and clinical diagnosis of fetal/neonatal alloimmune thrombocytopenia (FNAIT) and analyze the laboratory test results and clinical data related to the disease, so as to provide reference for clinical treatment and improvement of prognosis. METHODS: The clinical data of six neonatal patients with FNAIT in the Neonatology Department of our hospital from March 2017 to September 2020 were retrospectively analyzed, which included laboratory diagnosis, clinical symptoms, treatment, and prognosis. RESULTS: Among six patients, two cases occurred in the first pregnancy and four cases in the second pregnancy. The platelet count of six cases were decreased at admission or during hospitalization and maternal and neonatal serum autoimmune platelet antibody tests were positive. Five cases were accompanied by different degrees of skin and facial bleeding spots or petechiae and ecchymosis, intracranial hemorrhage. Four cases were treated with immunoglobulin and/or steroid hormone therapy (one of them received cross-matched platelets transfusion), while the symptoms of the other two cases improved spontaneously. Five cases recovered and were discharged from the hospital, while one case had not recovered but the family members requested to be discharged forwardly. Four cases were hospitalized within two weeks, but two cases were hospitalized for more than two weeks due to other diseases or factors (e.g., neonatal sepsis, neonatal enteritis, congenital heart disease, neonatal asphyxia, etc.). CONCLUSION: FNAIT is characterized by decreased platelet count, with or without bleeding symptoms, and may occur in the first and following pregnancy. FNAIT can recover spontaneously or have a good prognosis after treatment. However, the complication with other diseases or factors may affect the prognosis.

A single-center investigational study of CD36 antigen deficiency and platelet alloantibody distribution in different populations in Northern China as well as platelet alloantibodies effect on pregnancy.

BACKGROUND: Platelet antibodies can lead to clinical diseases such as platelet transfusion refractoriness (PTR), fetal/neonatal alloimmune thrombocytopenia (FNAIT), etc. This study is aimed at understanding CD36 expression, platelet alloantibody distribution in different populations in Northern China, and effects of platelet alloantibodies on pregnancy. STUDY DESIGN AND METHODS: Whole blood samples of 612 subjects including hematological patients, pregnant women, and blood donors were collected at a single center, then CD36 expressions were determined, followed by platelet antibody screening and characterization of platelet antibody specificity. A retrospective analysis was performed in 1552 pregnant women admitted to Department of Obstetrics, in order to investigate FNAIT occurrence. RESULTS: Rate of CD36 deficiency expression was 2.12% (13/612), all cases exhibited type II deficiency without type I deficiency being detected, and such rate is lower than that in Southern China (3.43%), Japanese (4.87%) and in the black people (4.18%), and higher than that in the White people (0.09%). Positive rates of platelet antibody screening in hematological patient group (6.86%, 14/204) and in pregnant women group (6.31%, 13/206) are higher than that in blood donor group (0.49%, 1/202), P < .01. Out of 1552 pregnant women, there were not children with FNAIT. CONCLUSION: The frequency of CD36 deficiency in northern China was low, all of them were type II deficiency, and no CD36 antibody was detected. It is speculated that the risk of immune-related thrombocytopenia caused by CD36 deficiency in this population is very low. Platelet antibodies should be monitored early in patients with hematological and multiple miscarriages pregnant.

Identification and follow-up of pregnant women with platelet-type human platelet antigen (HPA)-1bb alloimmunized with fetal HPA-1a.

INTRODUCTION: Pregnant women negative for human platelet antigen 1a (HPA-1a) are at risk of alloimmunization with fetal HPA-1a antigen inherited from the father, and their offspring may develop fetal and neonatal alloimmune thrombocytopenia (FNAIT). The aim of this study was to analyze the frequency of HPA-1a alloimmunization in pregnant Polish women, the feasibility of using maternal platelets for intrauterine transfusions in women subjected to diagnostic fetal blood sampling (FBS) and to discuss potential consequences of alloimmunization. MATERIAL AND METHODS: Fifteen thousand two hundred and four pregnant women were typed for HPA-1a; HPA-1a negative were screened for anti-HPA-1a. Alloimmunized women received specialist perinatology care; some of them were subjected to FBS, followed by transfusion of HPA-1a negative platelet concentrates (PC) prepared from maternal blood. RESULTS: Three hundred seventy-three (2.5%) women were HPA-1a negative, and 32 (8.6%) tested positively for anti-HPA-1a. Antibodies were detected in 22 women during pregnancy. Diagnostic FBS followed by PC transfusion was performed in 14 woman, who were platelet donors for their 16 unborn babies. Blood donations were tolerated well by the patients, and also intrauterine platelet transfusions were uneventful. Pharmacotherapy with intravenous immunoglobulins was implemented in 11/22 patients. CONCLUSIONS: HPA-1a negative women (ca. 2.5% of all pregnant patients) are at risk of alloimmunization with HPA-1a antigen and developing FNAIT. Alloimmunized women can be donors of platelets for their offspring providing removal of antibodies from PC. Owing to potential complications, special care should be taken if an alloimmunized woman was qualified as a blood or stem cell recipient.

Taking a wider view on fetal/neonatal alloimmune thrombocytopenia.

In fetal/neonatal alloimmune thrombocytopenia (FNAIT), platelets are destroyed by maternal antibodies directed against fetal/neonate antigens. Thrombocytopenia can be severe and lead to intracranial hemorrhage (ICH) in about 10% of cases. Although three types of antigen groups, presented on platelets [ABO blood group antigens, human leukocyte antigens (HLA) and human platelet antigens (HPA)] are known to be implicated in immune platelet destruction, antibodies against HPA are most commonly involved in FNAIT and hence are the target of extensive research. Awareness of FNAIT by physicians as well as the availability of the most sensitive diagnostic methods capable of detecting a wide range of antibodies are crucial for the diagnosis of FNAIT and the prevention of severe thrombocytopenia and its bleeding risks in subsequent pregnancies.

Fetal/Neonatal Alloimmune Thrombocytopenia: Pathogenesis, Diagnostics and Prevention.

Fetal/neonatal alloimmune thrombocytopenia (FNAIT) is a relatively rare condition (1/1000-1/2000) that was granted orphan status by the European Medicines Agency in 2011. Clinical consequences of FNAIT, however, may be severe. A thrombocytopenic fetus or new-born is at risk of intracranial hemorrhage that may result in lifelong disability or death. Preventing such bleeding is thus vital and requires a solution. Anti-HPA1a antibodies are the most frequent cause of FNAIT in Caucasians. Its pathogenesis is similar to hemolytic disease of the newborn (HDN) due to anti-RhD antibodies, but is characterized by platelet destruction and is more often observed in the first pregnancy. In 75 % of these women, alloimmunization by HPA-1a antigens, however, occurs at delivery, which enables development of antibody-mediated immune suppression to prevent maternal immunization. As for HDN, the recurrence rate of FNAIT is high. For advancing diagnostic efforts and treatment, it is thereby crucial to understand the pathogenesis of FNAIT, including cellular immunity involvement. This review presents the current knowledge on FNAIT. Also described is a program for HPA-1a screening in identifying HPA-1a negative pregnant women at risk of immunization. This program is now performed at the Institute of Hematology and Transfusion Medicine in cooperation with the Department of Obstetrics and Gynecology of the Medical Centre of Postgraduate Education in Warsaw as well as the UiT The Arctic University of Norway.

Placental histological lesions in fetal and neonatal alloimmune thrombocytopenia: A retrospective cohort study of 21 cases.

BACKGROUND: Alloimmunization against human platelet antigens (HPAs) can occur prenatally and induce fetal/neonatal alloimmune thrombocytopenia (FNAIT). The aim of this study was to identify placental histological features associated with platelet alloimmunization and their clinical significance. METHODS: This study examined 21 placentas from FNAIT-affected pregnancies and 42 age-matched control cases, all collected from pathology departments in the Rhône-Alpes region. Clinical and laboratory findings were collected for each FNAIT case. Two pathologists reviewed the placental slides of each FNAIT and control case. Histological features, with special emphasis on chronic inflammatory lesions, were evaluated. Differences between the two groups were calculated with odds ratios (ORs) and assessed with Wald's chi-square. RESULTS: FNAIT was associated with a significantly higher frequency of chronic chorioamnionitis (CC) (OR 14, 95%CI 1.7-113.8), basal chronic villitis (BCV) (OR 17, 95%CI 2-145.6) and chronic intervillositis (CIV). Chronic villitis (CV) (OR 3.7, 95%CI 0.9-15.2) and chronic deciduitis (CD) (OR 4.7, 95%CI 0.79-28.2) were also more frequent in the FNAIT than the control group, but these differences were not statistically significant. CONCLUSIONS: FNAIT is significantly associated with CC, BCV, and CIV. This chronic inflammatory reaction is preferentially localized on the maternofetal interface. Anti-HPA alloimmunization may trigger an immunological conflict similar to graft-versus-host disease.

Prenatal testing for hemolytic disease of the newborn and fetal neonatal alloimmune thrombocytopenia - current status.

Incompatibility of red cell and platelet antigens can lead to maternal alloimmunization causing hemolytic disease of the fetus & newborn and fetal neonatal alloimmune thrombocytopenia respectively. As the molecular background of these polymorphisms emerged, prenatal testing using initially fetal DNA obtained from invasively obtained amniotic fluid or chorionic villus was implemented. This evolved into testing using maternal plasma as source of fetal DNA, and this is in routine use as a safe non-invasive diagnostic that has no risk to the fetus of alloimmunization or spontaneous miscarriage. These tests were initially applied to high risk pregnancies, but has been applied on a mass scale, to screen fetuses in D-negative pregnant populations as national screening programs. Fetal neonatal alloimmune thrombocytopenia management has had comparatively small take up in non-invasive testing for causative fetal platelet alleles (e.g., HPA-1A), but mass scale genotyping of mothers to identify at risk HPA-1b1b pregnancies and their treatment with prophylactic anti-HPA-1A is being considered in at least one country (Norway).