Hemolytic disease of the fetus and newborn: systematic literature review of the antenatal landscape.

BACKGROUND: Prevention of pregnancy-related alloimmunization and the management of hemolytic disease of the fetus and newborn (HDFN) has significantly improved over the past decades. Considering improvements in HDFN care, the objectives of this systematic literature review were to assess the prenatal treatment landscape and outcomes of Rh(D)- and K-mediated HDFN in mothers and fetuses, to identify the burden of disease, to identify evidence gaps in the literature, and to provide recommendations for future research. METHODS: We performed a systematic search on MEDLINE, EMBASE and clinicaltrials.gov. Observational studies, trials, modelling studies, systematic reviews of cohort studies, and case reports and series of women and/or their fetus with HDFN caused by Rhesus (Rh)D or Kell alloimmunization. Extracted data included prevalence; treatment patterns; clinical outcomes; treatment efficacy; and mortality. RESULTS: We identified 2,541 articles. After excluding 2,482 articles and adding 1 article from screening systematic reviews, 60 articles were selected. Most abstracted data were from case reports and case series. Prevalence was 0.047% and 0.006% for Rh(D)- and K-mediated HDFN, respectively. Most commonly reported antenatal treatment was intrauterine transfusion (IUT; median frequency [interquartile range]: 13.0% [7.2-66.0]). Average gestational age at first IUT ranged between 25 and 27 weeks. weeks. This timing is early and carries risks, which were observed in outcomes associated with IUTs. The rate of hydrops fetalis among pregnancies with Rh(D)-mediated HDFN treated with IUT was 14.8% (range, 0-50%) and 39.2% in K-mediated HDFN. Overall mean ± SD fetal mortality rate that was found to be 19.8%±29.4% across 19 studies. Mean gestational age at birth ranged between 34 and 36 weeks. CONCLUSION: These findings corroborate the rareness of HDFN and frequently needed intrauterine transfusion with inherent risks, and most births occur at a late preterm gestational age. We identified several evidence gaps providing opportunities for future studies.

Hemolytic disease of the fetus and newborn: rapid review of postnatal care and outcomes.

BACKGROUND: Advances in postnatal care for hemolytic disease of the fetus and newborn (HDFN) have occurred over the past decades, but little is known regarding the frequency of postnatal treatment and the clinical outcomes of affected neonates. Most studies reporting on HDFN originate from high-income countries or relatively large centers, but important differences between centers and countries may exist due to differences in prevalence and available treatment options. We therefore aimed to evaluate the postnatal treatment landscape and clinical outcomes in neonates with Rhesus factor D (Rh(D))- and/or K-mediated HDFN and to provide recommendations for future research. METHODS: We conducted a rapid literature review of case reports and series, observational retrospective and prospective cohort studies, and trials describing pregnancies or children affected by Rh(D)- or K-mediated HDFN published between 2005 and 2021. Information relevant to the treatment of HDFN and clinical outcomes was extracted. Medline, ClinicalTrials.gov and EMBASE were searched for relevant studies by two independent reviewers through title/abstract and full-text screening. Two independent reviewers extracted data and assessed methodological quality of included studies. RESULTS: Forty-three studies reporting postnatal data were included. The median frequency of exchange transfusions was 6.0% [interquartile range (IQR): 0.0-20.0] in K-mediated HDFN and 26.5% [IQR: 18.0-42.9] in Rh(D)-mediated HDFN. The median use of simple red blood cell transfusions in K-mediated HDFN was 50.0% [IQR: 25.0-56.0] and 60.0% [IQR: 20.0-72.0] in Rh(D)-mediated HDFN. Large differences in transfusion rates were found between centers. Neonatal mortality amongst cases treated with intrauterine transfusion(s) was 1.2% [IQR: 0-4.4]. Guidelines and thresholds for exchange transfusions and simple RBC transfusions were reported in 50% of studies. CONCLUSION: Most included studies were from middle- to high-income countries. No studies with a higher level of evidence from centers in low-income countries were available. We noted a shortage and inconsistency in the reporting of relevant data and provide recommendations for future reports. Although large variations between studies was found and information was often missing, analysis showed that the postnatal burden of HDFN, including need for neonatal interventions, remains high. SYSTEMATIC REVIEW REGISTRATION: PROSPERO 2021 CRD42021234940. Available from:  https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42021234940 .

Live birth prevalence of hemolytic disease of the fetus and newborn in the United States from 1996 to 2010.

BACKGROUND: Hemolytic disease of the fetus and newborn (HDFN) is mediated by maternal alloantibodies, a consequence of immune sensitization during pregnancy with maternal-fetal incompatibility with ABO, Rhesus factor (Rh), and/or other red blood cell antigens. RhD, Kell, and other non-ABO alloantibodies are the primary cause of moderate to severe HDFN, whereas ABO HDFN is typically mild. HDFN live birth prevalence owing to Rh alloimmunization among newborns in the United States was last estimated to be 106 per 100,000 births in 1986. HDFN live birth prevalence owing to all alloantibodies was estimated to be 817 to 840 per 100,000 in Europe. There is a need for updated prevalence estimates in the United States and a better understanding of disease demographics, severity, and treatments. OBJECTIVE: This study aimed to estimate the live birth prevalence of HDFN and the proportion of severe cases of HDFN in the United States, to describe the associated risk factors, and to compare the clinical outcomes and treatments among healthy newborns, newborns with HDFN, and newborns who are sick without HDFN using a nationally representative hospital discharge database. STUDY DESIGN: In this retrospective, observational cohort study, we used data from the 1996 to 2010 National Hospital Discharge Survey to identify live births, defined by inpatient visits with the newborn flag, with and without a diagnosis of HDFN across 200 to 500 sampled hospitals (≥6 beds) per year. Patient and hospital characteristics, alloimmunization status, disease severity, treatment, and clinical outcomes were evaluated. Frequencies and weighted percentages were calculated for all variables. Logistic regression was used to compare the characteristics between newborns with HDFN and other newborns using odds ratios. RESULTS: Of 480,245 live births identified, 9810 HDFN cases were recorded. When weighted to the United States population, this corresponded to a live birth prevalence of 1695 per 100,000 live births. Compared with other newborns, newborns with HDFN were more likely to be female, Black, living in the South (vs the Midwest or West), and treated at larger (>100 beds) and government-owned hospitals. ABO and Rh alloimmunization accounted for 78.1% and 4.3% of newborns with HDFN, respectively, whereas HDFN caused by other antigens, such as Kell and Duffy, accounted for 17.6% of the cases. Among newborns with HDFN, 22% received phototherapy, 1% received simple transfusions, and 0.5% received exchange transfusions or intravenous immunoglobulin. Newborns affected by HDFN caused by Rh alloimmunization were more likely to require medical interventions, including simple or exchange transfusions, and more likely to be delivered by cesarean delivery. Overall, HDFN was associated with a longer hospital length of stay in the neonatal intensive care unit when compared with healthy and other sick newborns, a higher rate of cesarean delivery, and a higher rate of nonroutine discharge than healthy newborns. CONCLUSION: Overall, the live birth prevalence of HDFN was higher than those previously reported, whereas Rh-induced HDFN live birth prevalence was similar to those previously reported. HDFN live birth prevalence owing to Rh alloimmunization decreased over time, likely because of continued Rh immune globulin prophylaxis. Treatment patterns for newborns with HDFN and the comparative clinical outcomes when compared with healthy newborns confirm the continued clinical needs of this population.

Angiogenic factors and preeclampsia.

Preeclampsia is a major cause of maternal and neonatal morbidity and mortality worldwide. Although the etiology of preeclampsia is still unclear, recent studies suggest that its major phenotypes, hypertension and proteinuria, may be due to an excess of circulating anti-angiogenic growth factors, most notably soluble fms-like tyrosine kinase 1 (sFlt1) and soluble endoglin (sEng). sFlt1 is an endogenous protein that is produced by the placenta. sFlt1 is able to bind to the angiogenic growth factors vascular endothelial growth factor and placental growth factor, thereby neutralizing their functions. High serum concentrations of sFlt1 and low concentrations of free vascular endothelial growth factor and free placental growth factor have been observed during and prior to clinical manifestation of preeclampsia. More recently, serum levels of sEng were also shown to be significantly elevated in preeclamptic women and levels of sEng correlated strongly with disease severity. Therefore, measurement of sFlt1 and sEng in the maternal circulation may be a useful diagnostic and screening tool for preeclampsia. The availability of such a test to predict preeclampsia would have significant impact on current obstetrical care and may help reduce preeclampsia-induced morbidity and mortality. This review will focus on the role of angiogenic factors in normal and abnormal placental development and indicate how measurement of circulating angiogenic factors may help identify women at risk of preeclampsia.

Fetal nucleic acids in maternal body fluids: an update.

Our laboratory continues to be actively involved in the development of new biomarkers for prenatal diagnosis using maternal blood and amniotic fluid. We have also developed a mouse model that demonstrates that cell-free fetal (cff) DNA is detectable in the pregnant maternal mouse. In human maternal plasma and serum we have analyzed factors that are important in the clinical interpretation of cff DNA levels. Maternal race, parity, and type of conception (natural or assisted) do not affect cff DNA levels, but maternal weight does. We have also analyzed the relationship between placental volume, using a three-dimensionsal ultrasound examination, and cff DNA levels. Surprisingly, there is no association between these values. Finally, we are using specific disease models (such as congenital diaphragmatic hernia and twin-to-twin transfusion) to understand the effects of gestational age and specific pathology on fetal gene expression by analyzing cell-free mRNA levels in maternal plasma. In the amniotic fluid we have focused on improvements in recovery of cff DNA and mRNA. By optimizing recovery we have made some interesting observations about differences in fetal DNA between blood and amniotic fluid. In addition, we have successfully hybridized cff DNA in amniotic fluid to DNA microarrays, permitting assessment of fetal molecular karyotype. We also have preliminary data on fetal gene expression in amniotic fluid. Finally, we remain actively involved in promoting noninvasive prenatal testing in the United States, such as encouraging the use of fetal DNA for fetal rhesus D assessment. On the other hand, we are cautious and concerned about the accuracy of "at-home" kits for fetal gender detection.

Circulating cell-free fetal messenger RNA levels after fetoscopic interventions of complicated pregnancies.

OBJECTIVE: The aim of this study was to examine fetal gene expression in maternal plasma after fetoscopic intervention for twin-twin transfusion syndrome or congenital diaphragmatic hernia. STUDY DESIGN: Twelve women with pregnancies that were complicated by twin-twin transfusion syndrome and 10 women carrying fetuses with congenital diaphragmatic hernia were sampled before and sequentially after treatment. Levels of glyceraldehyde-3-phosphate dehydrogenase, human placental lactogen, and gamma globin messenger RNA were measured by real-time reverse transcriptase polymerase chain reaction amplification. RESULTS: At all time points, glyceraldehyde-3-phosphate dehydrogenase messenger RNA levels were higher in the congenital diaphragmatic hernia cases than in the twin-twin transfusion syndrome cases (P < .05), but during the immediate postoperative observation period, there were no significant changes in glyceraldehyde-3-phosphate dehydrogenase, human placental lactogen, or gamma globin messenger RNA levels in individual patients or patients who were grouped by procedure. CONCLUSION: Fetoscopic intervention of complicated pregnancies does not affect circulating fetal messenger RNA levels, which is in contrast to earlier observations that circulating fetal DNA levels increase after laser ablation for twin-twin transfusion syndrome. Plasma glyceraldehyde-3-phosphate dehydrogenase messenger RNA levels could be a potential novel biomarker for fetal trauma.

Markers for presymptomatic prediction of preeclampsia and intrauterine growth restriction.

Preeclampsia and intrauterine growth restriction are both characterized by placental malfunction. The pathological processes of abnormal trophoblast invasion, partial absence of maternal spiral artery modification, increased apoptosis of trophoblast cells, and placental ischemia are all associated with the release of specific molecules. These proteins, as well as cell-free fetal DNA and RNA might be detected in the maternal peripheral circulation, quantified, and used for early identification and prediction of preeclampsia and intrauterine growth restriction, prior to the appearance of the clinical symptoms. As preeclampsia and intrauterine growth restriction are associated with increased maternal, perinatal, and neonatal morbidity and mortality, early identification of these pregnancy associated complications will permit the design of appropriate preventive measures. In this review a variety of factors reported to be useful as potential markers for early detection of pregnancies at increased risk will be discussed. Molecules associated with the establishment of the placenta and essential in fetal-maternal interactions, like interleukin 2-receptor, insulinlike growth factor-1, and insulinlike growth factor binding protein-1, placenta growth factor, hepatocyte growth factor, inhibin A, activin A, and human chorionic gonadotrophin seem to be the most likely candidates for presymptomatic markers for preeclampsia and/or intrauterine growth restriction. Detection and discrimination of these molecules through the placental RNA in maternal plasma based strategy has become a realistic option.

Plasma hepatocyte growth factor as a marker for small-for-gestational age fetuses.

OBJECTIVE: To study the association between hepatocyte growth factor (HGF) levels and pregnancy outcome. STUDY DESIGN: Hepatocyte growth factor levels were measured in 42 plasma samples between weeks 14 and 21 of gestation using an enzyme-linked immunosorbent assay (ELISA). Results were correlated to pregnancy outcome and Mann-Whitney U-test applied to study the differences. RESULTS: Hepatocyte growth factor values in pregnancies that develop preeclampsia (n=12) were not significantly different from unaffected pregnancies (n=21, multiples of the median (MoM)=1.38, P=0.47). However, hepatocyte growth factor values were significantly elevated in pregnancies of small-for-gestational age (SGA) fetuses (n=9) compared to uncomplicated pregnancies (MoM=2.66, P<0.001). CONCLUSION: Measurement of hepatocyte growth factor in peripheral blood between 14 and 21 weeks gestation may offer new possibilities in the early diagnosis and prediction of fetal birth weight but not of preeclampsia.

First-trimester follistatin-like-3 levels in pregnancies complicated by subsequent gestational diabetes mellitus.

OBJECTIVE To determine whether maternal levels of follistatin-like-3 (FSTL3), an inhibitor of activin and myostatin involved in glucose homeostasis, are altered in the first trimester of pregnancies complicated by subsequent gestational diabetes mellitus (GDM). RESEARCH DESIGN AND METHODS This was a nested case-control study of subjects enrolled in a prospective cohort of pregnant women with and without GDM (> or =2 abnormal values on a 100-g glucose tolerance test at approximately 28 weeks of gestation). We measured FSTL3 levels in serum collected during the first trimester of pregnancy. Logistic regression analyses were used to determine the risk of GDM. RESULTS Women who developed GDM (n = 37) had lower first-trimester serum levels of FSTL3 compared with women who did not (n = 127) (median 10,789 [interquartile range 7,013-18,939] vs. 30,670 [18,370-55,484] pg/ml, P < 0.001). When subjects were divided into tertiles based on FSTL3 levels, women with the lowest levels demonstrated a marked increase in risk for developing GDM in univariate (odds ratio 11.2 [95% CI 3.6-35.3]) and multivariate (14.0 [4.1-47.9]) analyses. There was a significant negative correlation between first-trimester FSTL3 levels and approximately 28-week nonfasting glucose levels (r = -0.30, P < 0.001). CONCLUSIONS First-trimester FSTL3 levels are associated with glucose intolerance and GDM later in pregnancy.

Trophoblastic oxidative stress and the release of cell-free feto-placental DNA.

Considerable quantities of cell-free fetal DNA circulate in the maternal blood during human pregnancy, but the origin of the DNA remains uncertain. Circumstantial evidence suggests the placenta is the principal source, so we tested the hypothesis that release occurs from the syncytiotrophoblast after the induction of apoptotic changes. Villous explants from normal placentas delivered by elective caesarean section were cultured under normoxic conditions (10% oxygen) for up to 20 hours or exposed to hypoxia (0.5% oxygen) for 1 hour followed by reoxygenation. The concentration of beta-globin cell-free DNA in the supernatant, measured using real-time polymerase chain reaction methodology, was significantly increased at 20 hours after hypoxia-reoxygenation. Release was associated with increased apoptosis, confirmed by increased activation of caspase-3 on Western blotting, and immunolocalized to the syncytiotrophoblast; necrosis was also evidenced by release of lactate dehydrogenase. Both release of cell-free DNA and apoptosis could be significantly reduced by the addition of antioxidant vitamins C and E to the culture medium. This study provides the first evidence of a mechanistic and quantitative link between placental apoptosis/necrosis and release of cell-free DNA, hence confirming that maternal serum/plasma concen-trations of cell-free DNA may act as a biomarker of trophoblast well-being during pregnancy.

Neurokinin B levels in maternal circulation during early pregnancy.

Neurokinin B levels were measured between the 10th-20th weeks of pregnancy, i.e., prior to the development of clinical symptoms, in women who developed preeclampsia or delivered a growth-restricted baby. No difference was found in plasma neurokinin B levels, although neurokinin B levels increased slightly towards term.

Circulating trophoblast in maternal blood.

This review describes the status of circulating trophoblast, but is considered in the perspective that only a specific subset of trophoblast cells circulates in the maternal blood. The consequences for isolation, identification and clinical potential are described.