High maternal body-mass-index (BMI) is linked to adverse pregnancy outcomes, partly through impaired placental development. An inverse association between BMI and placental biomarkers has been described, however it is unclear whether this relationship is restricted to obesity or occurs across the full-BMI range. We assessed the relationship between maternal BMI and placental biomarkers across multiple cohorts. METHODS:We analysed three UK cohorts: ALSPAC (n = 994), CBGS (n = 1202) and POPS (n = 3869). Concentrations of growth-differentiation-factor15 (GDF15), beta-human chorionic gonadotropin (βhCG), pregnancy-associated plasma protein-A (PAPP-A), and alpha-fetoprotein (AFP) were measured at 8-16 weeks of gestation in maternal serum and expressed as gestational age-adjusted z-scores. Cohort-specific regression and individual-participant-data-meta-analyses (IPD-MA) were used. RESULTS:Across cohorts, each 1SD increase in maternal BMI was associated with significant reduction in: GDF15 (β: -0.16, 95%CI: -0.23 to -0.10; p<0.001), βhCG (β: -0.26, 95%CI: -0.28 to -0.23; p<0.001), PAPP-A (β: -0.38 95%CI: -0.41 to -0.36; p<0.001), and AFP (β: -0.21, 95%CI: -0.24 to -0.18; p<0.001). Associations were linear across the full BMI range, not driven solely by obesity. Biomarker levels correlated more strongly with early-pregnancy BMI than with estimated blood volume and associated directionally with the maternal polygenic-risk-scores (PRS) for BMI. CONCLUSIONS:Maternal BMI across its full range was inversely associated with biomarkers arising from, or transferred through, the placenta. We propose that the maternal caloric environment may regulate the development of the materno-fetal interface and its capacity for nutrient transfer, supporting similar rates of early fetal-growth in the face of substantial inter-individual variation in maternal body mass.

Preeclampsia and fetal growth restriction (FGR) are major causes of global morbidity and mortality. Both conditions are associated with impaired invasion of the uterus by extravillous trophoblast (EVT). We performed proteomics in maternal serum obtained at ~12 weeks of gestational age in a prospective pregnancy cohort (Pregnancy Outcome Prediction Study). Here we show that low maternal serum isthmin-2 (ISM2) was the strongest protein signal (out of 2,904) in the first trimester of pregnancy for preeclampsia or FGR. We validated the association in two independent cohorts (Pregnancy Outcome Prediction Study 2 and Improving Maternal Pregnancy And Child ouTcomes study). ISM2 protein and mRNA are almost exclusively produced in the placenta, and, within the placenta, ISM2 mRNA is highly enriched in EVT. Knocking down ISM2 in cultured human trophoblast stem cells profoundly inhibited EVT invasion. Conversely, expressing ISM2 in a cell line lacking endogenous ISM2 (HEK293 cells) promoted migration. We conclude that ISM2 may be causally involved in the early pathophysiology of failed trophoblast invasion and that the protein and its associated pathways are potential targets for the prediction and prevention of preeclampsia and FGR.

Introduction Current UK guidelines recommend measurement of symphyseal fundal height and measurement of maternal blood pressure and urinalysis, with the aim of detecting women at increased risk of fetal growth restriction (FGR) and pre-eclampsia. Between 2008 and 2013, we conducted a prospective cohort study recruiting 4512 nulliparous women at the Rosie Hospital, Cambridge, where we performed serial ultrasonic imaging and serial blood sampling and generated a novel screening test for pre-eclampsia and FGR. The method involved measuring the ratio of two placental biomarkers (soluble fms-like tyrosine kinase receptor-1 and placenta growth factor) at ~36 weeks of gestational age (wkGA) and combining the result with maternal characteristics and ultrasonic imaging. Women who screened positive had a ~50% risk of a composite outcome, consisting of pre-eclampsia±delivery of a baby with a birth weight <3rd percentile for sex and gestational age±perinatal morbidity or death. It is plausible that screening and intervention using this method might improve pregnancy outcome. Methods and analysis Nulliparous women with an apparently normal singleton pregnancy will be recruited at their dating ultrasound scan. Blood will be obtained at this visit, at their anomaly scan (20 wkGA), and at two research appointments (28 wkGA and 36 wkGA) when research ultrasound scans will be performed. Blood for DNA will be obtained from the father of the baby where possible and the placenta will be sampled following birth. At 36 wkGA, women will be consented for participation in the randomised controlled trial (RCT) element of the study and their risk of term pre-eclampsia and FGR will be assessed using the novel approach. Women who screen high-risk will then be randomly allocated to either having the result revealed or masked. Women randomised to having the result revealed will be offered early delivery and/or enhanced monitoring. Where the result is masked, there will be no communication between the research team and the participant, and she will continue to receive routine care at the Rosie Hospital. The primary outcome is a composite of pre-eclampsia, FGR and perinatal morbidity and mortality. The study will also generate data and biological samples to support future research in novel screening methods and disease mechanisms. Ethics and dissemination The study received ethical approval from the East of England Research Ethics Committee. All women provide written informed consent to participate in the cohort. Women provide a second written informed consent to participate in the RCT. The study results will be disseminated by presentation at international conferences and publication in peer reviewed journals. Trial registration number ISRCTN1218142 .

BACKGROUND AND AIM:Mendelian randomization (MR) is a widely used tool to estimate causal effects using genetic variants as instrumental variables. MR is limited to cross-sectional summary statistics of different samples and time points to analyze time-varying effects. We aimed at using longitudinal summary statistics for an exposure in a multivariable MR setting and validating the effect estimates for the mean, slope, and within-individual variability. SIMULATION STUDY:We tested our approach in 12 scenarios for power and type I error, depending on shared instruments between the mean, slope, and variability, and regression model specifications. We observed high power to detect causal effects of the mean and slope throughout the simulation, but the variability effect was low powered in the case of shared SNPs between the mean and variability. Mis-specified regression models led to lower power and increased the type I error. REAL DATA APPLICATION:We applied our approach to two real data sets (POPS, UK Biobank). We detected significant causal estimates for both the mean and the slope in both cases, but no independent effect of the variability. However, we only had weak instruments in both data sets. CONCLUSION:We used a new approach to test a time-varying exposure for causal effects of the exposure's mean, slope and variability. The simulation with strong instruments seems promising but also highlights three crucial points: (1) The difficulty to define the correct exposure regression model, (2) the dependency on the genetic correlation, and (3) the lack of strong instruments in real data. Taken together, this demands a cautious evaluation of the results, accounting for known biology and the trajectory of the exposure.

Preeclampsia (PE) and fetal growth restriction (FGR) complicate 5-10% of pregnancies and are major causes of maternal and fetal morbidity and mortality. Here we demonstrate that measuring circulating cell-free RNAs (cfRNAs) from maternal plasma can accurately predict pregnancies complicated by the combination of PE and FGR. We investigated 751 maternal plasma samples from 195 pregnant women (39 cases; 156 non-cases). We developed machine learning models from our discovery cohort (15 cases; 60 non-cases) and evaluated their predictive performances internally (24 cases; 96 controls) and externally (40 cases; 73 non-cases). We found circulating leptin (LEP) and pappalysin2 (PAPPA2) cfRNAs are the strongest cfRNA predictors of complicated pregnancies, each with an area under the receiver operating characteristic curve (AUC) of ~0.82. Using an external validation dataset of women with established PE, the combination of LEP and PAPPA2 had an AUC ~0.951. Our findings show that cfRNAs can predict complications of human pregnancy.

Genetic factors related to pregnancy-related traits are understudied, especially in ancestrally diverse cohorts. To assess maternal contributions to hypertensive disorders of pregnancy (HDP), we performed a multi-ancestry genome-wide association study (GWAS) of HDP in data from the North Carolina-based Personalized Environment and Genes Study (PEGS) cohort with validation in the UK Biobank (UKBB). The GWAS revealed two maternal loci associated with HDP at the genome-wide significance level. The lead independent variants were rs114954125 on chromosome 2 (near LRP1B; odds ratio [OR] [95% confidence interval {CI}]): 2.96 [2.02-4.34]; p = 2.82 3 10-8) and rs61176331 on chromosome 3 (on RARB; OR (95% CI): 3.08 (2.12-4.48); p = 3.52 3 10-9). We validated the associations near RARB with a meta-analysis of PEGS and the UKBB. We also identified cis-expression quantitative trait loci in the candidate region associated with decreased RARB expression in macrophage cells exposed to Salmonella. Chromatin mapping in FUMA identified a significant interaction within chromosome 3's enhancer and open chromatin regions, with strong effects observed for RARB and H3P10 gene regulation in mesendoderm cells, mesenchymal stem cells, and trophoblast-like stem cells. We applied existing polygenic scores (PGS) for preeclampsia and gestational hypertension and found that the scores were significantly associated with HDP in PEGS. The findings demonstrate the power of multi-ancestry studies for genetic discovery and highlight the relationship between immune response, regulation, and HDP and the utility of PGS for risk prediction. (PEGS is registered at ClinicalTrials.gov: NCT00341237.)

BACKGROUND:Preeclampsia is a major cause of maternal and fetal mortality and morbidity. Early risk stratification enables timely preventative therapy in high-risk women. Polygenic risk scores (PGS) improve prediction in complex diseases, but their added value for preeclampsia remains unclear, particularly in comparison to gold-standard first-trimester prediction models and across non-European ancestries. METHODS:We evaluated the performance of both a preeclampsia and systolic blood pressure PGS in 2 prospective pregnancy cohorts with detailed phenotyping: the Fetal Medicine Foundation study (n=5207; 2127 cases) and the Pregnancy Outcome Prediction study (n=3659; 228 cases). Risk models included (1) clinical factors; (2) clinical factors plus PGS; (3) advanced model including first-trimester mean arterial pressure, PAPP-A (pregnancy-associated plasma protein-A), and uterine artery pulsatility index; and (4) advanced model plus PGS. Discriminative performance, measured by the area under the receiver operating characteristic curve, was assessed overall and by ancestry. RESULTS:The preeclampsia PGS was independently associated with preeclampsia (odds ratio per SD, 1.24 [95% CI, 1.17-1.31]; P<0.001). It modestly improved prediction over clinical models (area under the receiver operating characteristic curve 0.746 versus 0.750; P=0.017) but not over the advanced model (area under the receiver operating characteristic curve 0.817 versus 0.818; P=0.326). The systolic blood pressure PGS showed stronger performance, improving prediction over both models in women of European ancestry. No improvement was observed with either score in women of African ancestry. CONCLUSIONS:PGSs for preeclampsia and SBP provide modest added predictive value beyond clinical risk factors in European ancestry women. Limited utility in African ancestry women reflects underrepresentation in the genome-wide association studies used to develop current scores. As cohort sizes grow and models are refined, PGSs may become important tools for equitable risk stratification in maternal health.

OBJECTIVE:To determine the inter-relationships between five first-trimester biomarkers (pregnancy associated plasma protein A [PAPP-A], alpha-fetoprotein [AFP], beta human chorionic gonadotrophin [beta-hCG], placenta growth factor [PlGF] and soluble fms-like tyrosine kinase receptor-1 [sFlt-1]) and a range of adverse pregnancy outcomes (APOs).DESIGN:Prospective cohort study of nulliparous singleton pregnancy.SETTING:Cambridge, UK.POPULATION OR SAMPLE:4056 pregnancy outcome prediction study participants.METHODS:The biomarker concentrations were measured in maternal serum at ~12 weeks of gestation. Univariable analysis of APOs was performed using logistic regression. Multivariable analysis used best subsets logistic regression with cross-validation.MAIN OUTCOME MEASURES:Pre-eclampsia (PE), small for gestational age (SGA), including severe SGA (birthweight <3rd), fetal growth restriction (FGR), preterm birth (PTB, both induced and spontaneous [iPTB and sPTB, respectively]), pre-viable loss and stillbirth, plus combinations of outcomes.RESULTS:Lower values of PAPP-A, PlGF and sFlt-1 and higher values of AFP were associated with FGR (OR for 1 SD higher value 0.59 [95% CI 0.48-0.74], OR 0.56 [95% CI 0.44-0.70], OR 0.68 [95% CI 0.54-0.87] and OR 1.53 [95% CI 1.25-1.88]), severe SGA (OR 0.59 [95% CI 0.49-0.72], OR 0.71 [95% CI 0.57-0.87], OR 0.74 [95% CI 0.60-0.91] and OR 1.41 [95% CI 1.17-1.71]), sPTB (OR  0.61 [95% CI 0.50-0.73], OR 0.79 [95% CI 0.66-0.96], OR 0.57 [95% CI 0.47-0.70] and OR 1.41 [95% CI 1.18-1.67]) and iPTB (OR  0.72 [95% CI 0.57-0.91], OR 0.62 [95% CI 0.49-0.78], OR 0.71 [95% CI 0.56-0.90] and OR 1.44 [95% CI 1.16-1.78]), respectively. When combinations of biomarkers were assessed, PAPP-A and AFP were independently associated with severe SGA; PAPP-A alone with PE + PTB; PlGF alone with severe PE; PlGF, beta-hCG, AFP and PAPP-A with the combination of PE and SGA; AFP and sFlt-1 with sPTB; and AFP and PlGF with iPTB.CONCLUSIONS:Combinations of first-trimester placental biomarkers are associated with APOs. However, the patterns vary for different types of APO, indicating heterogeneity in the underlying pathophysiological pathways.

BACKGROUND: Elevated maternal serum sFLT1 (soluble fms-like tyrosine kinase 1) has a key role in the pathophysiology of preeclampsia. We sought to determine the relationship between the maternal and fetal genome and maternal levels of sFLT1 at 12, 20, 28, and 36 weeks of gestational age (wkGA). METHODS: We studied a prospective cohort of nulliparous women (3968 mother-child pairs). We related maternal and fetal genotype to the adjusted sFLT1 Z score and sFLT1:placental growth factor (PlGF) ratio Z score at each wkGA and the change in the Z score between 28 and 36 wkGA (Δ36-28). We studied genetic variants from a previous fetal genome-wide association study of preeclampsia and an externally defined polygenic score from a maternal genome-wide association study of preeclampsia. RESULTS: Four variants from the fetal preeclampsia genome-wide association study were positively associated with sFLT1 and sFLT1:PlGF Z score at 36 wkGA, and FLT1 enhancer single-nucleotide polymorphisms were associated with increased Δ36-28 of sFLT1. The associations were specific for the fetal genome or stronger for the fetal than the maternal genome. An increased risk of preeclampsia based on the maternal polygenic score for preeclampsia was associated with lower levels of sFLT1 and sFLT1:PlGF ratio in the first trimester and a greater Δ36-28 for sFLT1. CONCLUSIONS: The current data are consistent with a causal association between sFLT1 released by the placenta in late pregnancy and the pathophysiology of preeclampsia. The data are also consistent with maternal components to the protective effect of high sFLT1 in the first trimester and the rise in third-trimester sFLT1 levels and preeclampsia.

BACKGROUND:Although maternal hemoglobin levels during pregnancy are commonly associated with perinatal outcomes, their link to childhood neurodevelopment remains uncertain. OBJECTIVE:This study aimed to examine the associations between maternal hemoglobin in early and late pregnancy and the educational attainment of offspring mid-childhood in a high-resource obstetric setting. STUDY DESIGN:Pregnancy data from a prospective birth cohort (Pregnancy Outcome Prediction Study, Cambridge, United Kingdom, 2008-2012, N=3285) were linked to mid-childhood educational outcomes (Department for Education, United Kingdom). Regression models adjusted for maternal, child, and socioeconomic factors were used to determine associations between maternal hemoglobin, pregnancy complications, and offspring educational outcomes (aged 5-7 years). RESULTS:No association was observed between maternal hemoglobin at 12 weeks and the likelihood of either adverse pregnancy outcomes or children meeting expected educational standards between ages 5-7 years. Higher maternal hemoglobin at 28 weeks was associated with an increased risk of small-for-gestational-age infants (adjusted odds ratio, 1.26 [95% confidence interval, 1.11-1.59]; P=.002) and preterm birth (adjusted odds ratio, 1.38 [95% confidence interval, 1.11-1.81]; P=.005). There were no adverse birth outcomes associated with anemia. However, children of mothers who were anemic at 28 weeks had ∼40% increased risk of not attaining expected educational standards at age 5 (adjusted odds ratio, 1.42 [95% confidence interval, 1.03-1.95]; P=.03). There was no association between maternal anemia at 28 weeks and educational performance at ages 6-7. No associations were found between high maternal hemoglobin concentrations (top decile) or change in hemoglobin concentrations between 12 and 28 weeks and childhood educational attainment. CONCLUSION:Maternal anemia at 28 weeks of pregnancy is associated with reduced educational attainment at 5 years old but not at older ages (6-7 years old). A proactive approach to increasing maternal hemoglobin in high-resource settings is unlikely to impact long-term childhood educational attainment.

To identify and internally validate metabolites predictive of spontaneous preterm birth (sPTB) using multiple machine learning methods and sequential maternal serum samples, and to predict spontaneous early term birth (sETB) using these metabolites.

BACKGROUND: Previous studies suggest that gestational diabetes mellitus is associated with poorer cognitive outcomes in children. However, confounding factors, especially maternal body mass index, have been poorly accounted for. OBJECTIVE: This study aimed to examine the independent associations between maternal body mass index, gestational diabetes mellitus status, and educational outcomes. STUDY DESIGN: Antenatal data from a prospective birth cohort (Pregnancy Outcome Prediction Study, 2008-2012, Cambridge, United Kingdom) were linked to mid-childhood educational outcomes (Department for Education, United Kingdom). A total of 3249 children born at term were stratified by maternal gestational diabetes mellitus status and body mass index at booking (<25 vs >25 kg/m2). 2 ). Regression models adjusted for relevant maternal, child, and socioeconomic factors were used to determine associations with academic outcomes at ages of 5 to 7 years. RESULTS: No differences in educational attainment were found between children exposed to gestational diabetes mellitus and nonexposed children. Neither maternal glucose levels measured at 11 to 14 or 24 to 28 weeks, nor acceleration of the fetal abdominal circumference growth velocity were related to educational attainment at ages of 5 to 7 years. Children of mothers with booking body mass index >25 kg/m2 2 (vs <25 kg/m(2)) were w50% more likely to not meet expected educational standards regardless of gestational diabetes mellitus status (age 5: adjusted odds ratio, 1.44; 95% confidence interval, 1.19-1.74; P<.001; age 6: adjusted odds ratio, 1.61; 95% confidence interval, 1.28-2.02; P<.001). The association between maternal body mass index and offspring educational attainment is dose-dependent and robust to stratification by gestational diabetes mellitus status and adjustment for socioeconomic factors. CONCLUSION: Mid-childhood educational attainment is not associated with maternal glucose status. This may provide important reassurance for pregnant women and clinicians. However, maternal body mass index is associated with lower childhood educational attainment and may be modifiable with intervention before or during pregnancy.

It is hypothesized that increased placental release of soluble fms-like tyrosine kinase-1 (sFLT1) is causally related to the pathophysiology of preeclampsia (PE). Genome-wide association studies identified four PE-associated fetal variants near the FLT1 gene: rs4769612, rs4769613, rs7318880, and rs12050029, but previous studies have failed to show associations between these variants in the mother and sFLT1 levels in the first and second trimester of pregnancy. We sought to determine the association between maternal or fetal carriage of these variants and maternal circulating levels of sFLT1 across all three trimesters of pregnancy. We analyzed data from the Pregnancy Outcome Prediction study, a prospective cohort of nulliparous women, where 3,968 mother-child pairs passed genotyping quality control. Maternal serum sFLT1 levels were measured at 12-, 20-, 28-, and 36 weeks of gestational age (wkGA) using a Roche Cobas e411 analyzer. In cross-sectional and longitudinal regression models, the dependent variable was the z-score of log-transformed sFLT1 at each wkGA or the change in z-score between 28- and 36wkGA, adjusted for fetal sex, age, BMI, maternal race/ethnicity, alcohol use, smoking, deprivation index, and the top 10 genetic principal components. There was no association between the maternal variants and sFLT1 levels at any individual wkGA (P > 0.05). There was no association between fetal variants and sFLT1 levels at 12, 20 or 28wkGA (P > 0.05). However, all four fetal variants were positively associated with the sFLT1 z-score at 36wkGA (Figure 1). All maternal and fetal variants except rs12050029 were associated with increased change in the sFLT1 z-score from 28 to 36wkGA. Fetal associations with third trimester sFLT1 z-score change were consistently stronger than maternal associations (P < 1 × 10-5). Fetal variants near FLT1 associated with an increased risk of PE are associated with higher maternal serum sFLT1 levels in the third trimester, supporting a causal role for placentally derived sFLT1 in the pathophysiology of PE.

Streptococcus agalactiae (Group B Streptococcus ; GBS) is a common cause of sepsis in neonates. Previous work detected GBS DNA in the placenta in ~5% of women before the onset of labour, but the clinical significance of this finding is unknown. Here we re-analysed this dataset as a case control study of neonatal unit (NNU) admission. Of 436 infants born at term (≥37 weeks of gestation), 7/30 with placental GBS and 34/406 without placental GBS were admitted to the NNU (odds ratio (OR) 3.3, 95% confidence interval (CI) 1.3–7.8). We then performed a validation study using non-overlapping subjects from the same cohort. This included a further 239 cases of term NNU admission and 686 term controls: 16/36 with placental GBS and 223/889 without GBS were admitted to the NNU (OR 2.4, 95% CI 1.2–4.6). Of the 36 infants with placental GBS, 10 were admitted to the NNU with evidence of probable but culture-negative sepsis (OR 4.8, 95% CI 2.2–10.3), 2 were admitted with proven GBS sepsis (OR 66.6, 95% CI 7.3–963.7), 6 were admitted and had chorioamnionitis (inflammation of the foetal membranes) (OR 5.3, 95% CI 2.0–13.4), and 5 were admitted and had funisitis (inflammation of the umbilical cord) (OR 6.7, 95% CI 12.5–17.7). Foetal cytokine storm (two or more pro-inflammatory cytokines >10 times median control levels in umbilical cord blood) was present in 36% of infants with placental GBS DNA and 4% of cases where the placenta was negative (OR 14.2, 95% CI 3.6–60.8). Overall, ~1 in 200 term births had GBS detected in the placenta, which was associated with infant NNU admission and morbidity.

Pubertal timing varies considerably and has been associated with a range of health outcomes in later life. To elucidate the underlying biological mechanisms, we performed multi-ancestry genetic analyses in ~800,000 women, identifying 1,080 independent signals associated with age at menarche. Collectively these loci explained 11% of the trait variance in an independent sample, with women at the top and bottom 1% of polygenic risk exhibiting a ~11 and ~14-fold higher risk of delayed and precocious pubertal development, respectively. These common variant analyses were supported by exome sequence analysis of ~220,000 women, identifying several genes, including rare loss of function variants in ZNF483 which abolished the impact of polygenic risk. Next, we implicated 660 genes in pubertal development using a combination of in silico variant-to-gene mapping approaches and integration with dynamic gene expression data from mouse embryonic GnRH neurons. This included an uncharacterized G-protein coupled receptor GPR83, which we demonstrate amplifies signaling of MC3R, a key sensor of nutritional status. Finally, we identified several genes, including ovary-expressed genes involved in DNA damage response that co-localize with signals associated with menopause timing, leading us to hypothesize that the ovarian reserve might signal centrally to trigger puberty. Collectively these findings extend our understanding of the biological complexity of puberty timing and highlight body size dependent and independent mechanisms that potentially link reproductive timing to later life disease.