FKBPL and SIRT-1 Are Downregulated by Diabetes in Pregnancy Impacting on Angiogenesis and Endothelial Function.

Diabetes in pregnancy is associated with adverse pregnancy outcomes including preterm birth. Although the mechanisms leading to these pregnancy complications are still poorly understood, aberrant angiogenesis and endothelial dysfunction play a key role. FKBPL and SIRT-1 are critical regulators of angiogenesis, however, their roles in pregnancies affected by diabetes have not been examined before in detail. Hence, this study aimed to investigate the role of FKBPL and SIRT-1 in pre-gestational (type 1 diabetes mellitus, T1D) and gestational diabetes mellitus (GDM). Placental protein expression of important angiogenesis proteins, FKBPL, SIRT-1, PlGF and VEGF-R1, was determined from pregnant women with GDM or T1D, and in the first trimester trophoblast cells exposed to high glucose (25 mM) and varying oxygen concentrations [21%, 6.5%, 2.5% (ACH-3Ps)]. Endothelial cell function was assessed in high glucose conditions (30 mM) and following FKBPL overexpression. Placental FKBPL protein expression was downregulated in T1D (FKBPL; p<0.05) whereas PlGF/VEGF-R1 were upregulated (p<0.05); correlations adjusted for gestational age were also significant. In the presence of GDM, only SIRT-1 was significantly downregulated (p<0.05) even when adjusted for gestational age (r=-0.92, p=0.001). Both FKBPL and SIRT-1 protein expression was reduced in ACH-3P cells in high glucose conditions associated with 6.5%/2.5% oxygen concentrations compared to experimental normoxia (21%; p<0.05). FKBPL overexpression in endothelial cells (HUVECs) exacerbated reduction in tubule formation compared to empty vector control, in high glucose conditions (junctions; p<0.01, branches; p<0.05). In conclusion, FKBPL and/or SIRT-1 downregulation in response to diabetic pregnancies may have a key role in the development of vascular dysfunction and associated complications affected by impaired placental angiogenesis.

Role of A Novel Angiogenesis FKBPL-CD44 Pathway in Preeclampsia Risk Stratification and Mesenchymal Stem Cell Treatment.

CONTEXT: Preeclampsia is a leading cardiovascular complication in pregnancy lacking effective diagnostic and treatment strategies. OBJECTIVE: To investigate the diagnostic and therapeutic target potential of the angiogenesis proteins, FK506-binding protein like (FKBPL) and CD44. DESIGN AND INTERVENTION: FKBPL and CD44 plasma concentration or placental expression were determined in women pre- or postdiagnosis of preeclampsia. Trophoblast and endothelial cell function was assessed following mesenchymal stem cell (MSC) treatment and in the context of FKBPL signaling. SETTINGS AND PARTICIPANTS: Human samples prediagnosis (15 and 20 weeks of gestation; n ≥ 57), or postdiagnosis (n = 18 for plasma; n = 4 for placenta) of preeclampsia were used to determine FKBPL and CD44 levels, compared to healthy controls. Trophoblast or endothelial cells were exposed to low/high oxygen, and treated with MSC-conditioned media (MSC-CM) or a FKBPL overexpression plasmid. MAIN OUTCOME MEASURES: Preeclampsia risk stratification and diagnostic potential of FKBPL and CD44 were investigated. MSC treatment effects and FKBPL-CD44 signaling in trophoblast and endothelial cells were assessed. RESULTS: The CD44/FKBPL ratio was reduced in placenta and plasma following clinical diagnosis of preeclampsia. At 20 weeks of gestation, a high plasma CD44/FKBPL ratio was independently associated with the 2.3-fold increased risk of preeclampsia (odds ratio = 2.3, 95% confidence interval [CI] 1.03-5.23, P = 0.04). In combination with high mean arterial blood pressure (>82.5 mmHg), the risk further increased to 3.9-fold (95% CI 1.30-11.84, P = 0.016). Both hypoxia and MSC-based therapy inhibited FKBPL-CD44 signaling, enhancing cell angiogenesis. CONCLUSIONS: The FKBPL-CD44 pathway appears to have a central role in the pathogenesis of preeclampsia, showing promising utilities for early diagnostic and therapeutic purposes.

Preeclampsia Prevention Using Routine Versus Screening Test-Indicated Aspirin in Low-Risk Women.

The objective was to evaluate whether routine aspirin 75 mg is more cost-effective than the Fetal Medicine Foundation screen-and-treat approach for preeclampsia prevention in low-risk nulliparous women. A health economic decision analytical model was devised to estimate the discounted net health and cost outcomes of routine aspirin versus Fetal Medicine Foundation screening test-indicated aspirin for a cohort of 100 000 low-risk nulliparous women. Both strategies were compared with no intervention. A subanalysis also compared disaggregated components of the algorithm. The analysis used data from hospital administration, literature, and a randomized controlled trial. Sensitivity analyses assessed the impact of aspirin adherence, test cost, and accuracy on study results. Presumed rates of preeclampsia were 3.75% with no intervention versus 0.45% with aspirin use. Results found that routine aspirin was the preferred strategy, in terms of greater health gains and larger cost savings. It provided 163 quality-adjusted life-years relative to no intervention, whereas the screen-and-treat policy achieved 108 quality-adjusted life-years. Routine aspirin would result in an estimated cost saving of €14.9 million annually relative to no intervention, whereas screen-and-treat approach would result in a smaller cost saving of €3.1 million. When the analysis was extended to consider alternative screen-and-treat strategies, routine aspirin remained the optimally cost-effective approach. In conclusion, routine aspirin use in low-risk nulliparous women has a greater health gain and cost saving compared with both the Fetal Medicine Foundation and other screen-and-treat approaches.

Trial of feasibility and acceptability of routine low-dose aspirin versus Early Screening Test indicated aspirin for pre-eclampsia prevention (TEST study): a multicentre randomised controlled trial.

OBJECTIVE: Evaluate the feasibility and acceptability of routine aspirin in low-risk women, compared with screening-test indicated aspirin for the prevention of pre-eclampsia and fetal growth restriction. DESIGN: Multicentre open-label feasibility randomised controlled trial. SETTING: Two tertiary maternity hospitals in Dublin, Ireland. PARTICIPANTS: 546 low-risk nulliparous women completed the study. INTERVENTIONS: Women underwent computerised randomisation to: Group 1-routine aspirin 75 mg from 11 until 36 weeks; Group 2-no aspirin and; Group 3-aspirin based on the Fetal Medicine Foundation screening test. PRIMARY AND SECONDARY OUTCOME MEASURES: (1) Proportion agreeing to participate; (2) compliance with protocol; (3) proportion where first trimester uterine artery Doppler was obtainable and; (4) time taken to issue a screening result. Secondary outcomes included rates of pre-eclampsia and small-for-gestational-age fetuses. RESULTS: 546 were included in the routine aspirin (n=179), no aspirin (n=183) and screen and treat (n=184) groups. 546 of 1054 were approached (51.8%) and enrolled. Average aspirin adherence was 90%. The uterine artery Doppler was obtained in 98.4% (181/184) and the average time to obtain a screening result was 7.6 (0-26) days. Of those taking aspirin, vaginal spotting was greater; n=29 (15.1%), non-aspirin n=28 (7.9%), OR 2.1 (95% CI 1.2 to 3.6). Postpartum haemorrhage >500 mL was also greater; aspirin n=26 (13.5%), no aspirin n=20 (5.6%), OR 2.6 (95% CI 1.4 to 4.8). CONCLUSION: Low-risk nulliparous women are open to taking aspirin in pregnancy and had high levels of adherence. Aspirin use was associated with greater rates of vaginal bleeding. An appropriately powered randomised controlled trial is now required to address the efficacy and safety of universal low-dose aspirin in low-risk pregnancy compared with a screening approach. TRIAL REGISTRATION NUMBER: ISRCTN (15191778); Post-results.

Prenatal surgery for spina bifida: a therapeutic dilemma. Proceedings of the SHINE conference, Belfast.

This is a transcript of a scientific conference on the subject of prenatal surgery for spina bifida. It represents the views of three patients, an obstetrician, a postnatal neurosurgeon, a neonatologist, a paediatric neurologist, two surgeons who practice open spina bifida foetal surgery, a fetoscopic surgeon and an obstetrician experienced in randomised trials and systematic reviews. Implications for current practice and recommendations for future research are also discussed in detail.

The impact of maternal obesity on completion of fetal anomaly screening.

OBJECTIVE: To examine the impact of maternal obesity on completion of fetal anomaly screening. METHODS: A retrospective analysis of 500 anomaly scans (19+0-21+6 weeks) was included. Women were categorised according to the World Health Organisation (WHO) body mass index (BMI) classification: normal weight (18.50-24.99 kg/m2), overweight (25.00-29.99 kg/m2), obese class I (30-34.99 kg/m2), obese class II (35.00-39.99 kg/m2) and obese class III (≥40.00 kg/m2). A fetal anomaly imaging scoring system was developed from the National Health Service (NHS) Fetal Anomaly Screening Programme standard to evaluate scans. RESULTS: Image quality deteriorated as BMI increased and was significantly different across the BMI categories (P<0.001). Performance was poorest in imaging of the fetal chest and was significantly different across BMI categories (P<0.001). In obese class III, 33% of four-chamber cardiac views and 38% of outflow tract views were not obtained. In total, 119 women (23.6%) had an incomplete scan. In obese class III, 44.1% of scans were incomplete compared with 10.2% in the normal BMI category (P<0.001). Of 117 women attending for repeat scans, 78.6% were complete, 11.1% were incomplete, 6.8% were advised to re-attend and 3.4% were referred to Fetal Medicine. CONCLUSION: Maternal obesity has a significant impact on completion of fetal anomaly screening.

Defining the residual risk of adverse perinatal outcome in growth-restricted fetuses with normal umbilical artery blood flow.

OBJECTIVE: We sought to determine the cause of adverse perinatal outcome in fetal growth restriction (FGR) where umbilical artery (UA) Doppler was normal, as identified from the Prospective Observational Trial to Optimize Pediatric Health (PORTO). We compared cases of adverse outcome where UA Doppler was normal and abnormal. STUDY DESIGN: The PORTO study was a national multicenter study of >1100 ultrasound-dated singleton pregnancies with an estimated fetal weight <10th centile. Each pregnancy underwent intensive ultrasound, including multivessel Doppler. UA Doppler was considered abnormal when the pulsatility index was >95th centile or end-diastolic flow was absent/reversed. Adverse perinatal outcome was defined as a composite of intraventricular hemorrhage, periventricular leukomalacia, hypoxic ischemic encephalopathy, necrotizing enterocolitis, bronchopulmonary dysplasia, sepsis, or death. RESULTS: In all, 57 (5.0%) of the 1116 fetuses had an adverse perinatal outcome. Nine (1.3%) of 698 fetuses with normal UA Doppler had an adverse outcome, compared with 48 (11.5%) of 418 with abnormal UA Doppler (P < .0001). There were 2 perinatal deaths in the normal group and 6 in the abnormal group (P = .01). The perinatal deaths in the normal group were 1 case of pulmonary hypoplasia after prolonged preterm rupture of the membranes from 12 weeks' gestation and a case of placental abruption. Gestation at delivery was 33 ± 3 vs 31 ± 4 weeks (P = .05) and mean birthweight was 1830 ± 737 vs 1146 ± 508 g (P = .001) in the respective groups. Neonatal sepsis was the commonest adverse outcome in both groups: 0.1% and 0.4%, respectively (P = .01). CONCLUSION: Adverse perinatal outcome is uncommon in FGR with normal UA Doppler. The cases we identified were associated with heterogenous pathologies. FGR with normal UA blood flow is a largely benign condition.

The role of brain sparing in the prediction of adverse outcomes in intrauterine growth restriction: results of the multicenter PORTO Study.

OBJECTIVE: The aim of the Prospective Observational Trial to Optimize Pediatric Health in IUGR Study was to evaluate the optimal management of fetuses with an estimated fetal weight less than the 10th centile. The objective of this secondary analysis was to describe the role of the cerebroplacental ratio (CPR) in the prediction of adverse perinatal outcome. STUDY DESIGN: More than 1100 consecutive singleton pregnancies with intrauterine growth restriction (IUGR) were recruited over 2 years at 7 centers, undergoing serial sonographic evaluation including multivessel Doppler measurement. CPR was calculated using the pulsatility and resistance indices of the middle cerebral and umbilical artery. Adverse perinatal outcome was defined as a composite of intraventricular hemorrhage, periventricular leukomalacia, hypoxic ischemic encephalopathy, necrotizing enterocolitis, bronchopulmonary dysplasia, sepsis, and death. RESULTS: Data for CPR calculation was available in 881 cases, which was performed at a mean gestational age of 33 weeks (interquarile range, 28.7-35.9). Of the 146 cases with CPR less than 1, 18% (n = 27) had an adverse perinatal outcome. This conferred an 11-fold increased risk (odds ratio, 11.7; P < .0001) when compared with cases with normal CPR (2%; 14 of 735). An abnormal CPR was present in all 3 cases of mortality. Prediction of adverse outcomes was comparable when using all definitions of abnormal CPR. CONCLUSION: Irrespective of the CPR calculation used, brain sparing is significantly associated with an adverse perinatal outcome in IUGR. This adds further weight to integrating CPR evaluation into the clinical assessment of IUGR pregnancies. The impact of this finding on long-term neurodevelopmental outcomes in this patient cohort is underway.

Optimizing the definition of intrauterine growth restriction: the multicenter prospective PORTO Study.

OBJECTIVE: The objective of the Prospective Observational Trial to Optimize Pediatric Health in Intrauterine Growth Restriction (IUGR) (PORTO Study), a national prospective observational multicenter study, was to evaluate which sonographic findings were associated with perinatal morbidity and mortality in pregnancies affected by growth restriction, originally defined as estimated fetal weight (EFW) <10th centile. STUDY DESIGN: Over 1100 consecutive ultrasound-dated singleton pregnancies with EFW <10th centile were recruited from January 2010 through June 2012. A range of IUGR definitions were used, including EFW or abdominal circumference <10th, <5th, or <3rd centiles, with or without oligohydramnios and with or without abnormal umbilical arterial Doppler (pulsatility index >95th centile, absent or reversed end-diastolic flow). Adverse perinatal outcome, defined as a composite outcome of intraventricular hemorrhage, periventricular leukomalacia, hypoxic ischemic encephalopathy, necrotizing enterocolitis, bronchopulmonary dysplasia, sepsis, and death was documented for all cases. RESULTS: Of 1116 fetuses, 312 (28%) were admitted to neonatal intensive care unit and 58 (5.2%) were affected by adverse perinatal outcome including 8 mortalities (0.7%). The presence of abnormal umbilical Doppler was significantly associated with adverse outcome, irrespective of EFW or abdominal circumference measurement. The only sonographic weight-related definition consistently associated with adverse outcome was EFW <3rd centile (P = .0131); all mortalities had EFW <3rd centile. Presence of oligohydramnios was clinically important when combined with EFW <3rd centile (P = .0066). CONCLUSION: Abnormal umbilical artery Doppler and EFW <3rd centile were strongly and most consistently associated with adverse perinatal outcome. Our data call into question the current definitions of IUGR used. Future studies may address whether using stricter IUGR cutoffs comparing various definitions and management strategies has implications on resource allocation and pregnancy outcome.

The endogenous anti-angiogenic family of splice variants of VEGF, VEGFxxxb, are down-regulated in pre-eclamptic placentae at term.

PET (pre-eclamptic toxaemia) has recently been linked with alterations in production of a VEGFR1 [VEGF (vascular endothelial growth factor) receptor 1] splice variant that acts as a circulating inhibitor. We have recently described a family of naturally occurring splice variants of VEGF, termed VEGFxxxb, that also appear to act as inhibitors of conventional VEGFxxx-mediated angiogenesis. To determine whether alteration in splicing of VEGF-VEGFR family members extended beyond VEGFR1, we investigated the effect of pre-eclampsia on placental VEGFxxxb mRNA and protein expression. VEGFxxx and VEGFxxxb mRNA and protein were both found in normal human term placentae. VEGFxxx protein formed the majority of the total VEGF protein (980+/-195 pg/mg), whereas VEGFxxxb (11.5 pg/mg) was found to form a small part of the total VEGF protein expression (1.5+/-0.24%). Evidence for VEGF165b, VEGF121b and VEGF145b expression was found. In pre-eclamptic placentae, there was a significant down-regulation of VEGFxxxb isoforms, but a small up-regulation of VEGFxxx isoforms. In normal placenta VEGFxxxb and VEGFxxx concentrations were positively correlated (r=0.69, P<0.02), whereas in pre-eclamptic placentae, there was a significant negative correlation between VEGFxxxb and VEGFxxx protein expression (r=-0.8, P<0.02), indicating that there was a significant uncoupling of the splicing regulation of the VEGF isoforms. Combined with previous studies showing increased soluble VEGFR1 isoforms in human pre-eclampsia, these data suggest that there may be a common mechanism in pre-eclampsia that involves dysregulation of mRNA splicing of members of the VEGF-VEGFR axis.

Soluble vascular endothelial growth factor receptor-1 (sFlt-1) is increased throughout gestation in patients who have preeclampsia develop.

OBJECTIVE: This study was undertaken to analyze prospectively circulating vascular endothelial growth factor (VEGF) and its soluble receptor, (s) Flt-1, throughout normotensive and preeclamptic pregnancies and to assess the importance of these proteins in the development of preeclampsia. STUDY DESIGN: In this longitudinal cohort study, serum samples were collected from recruited subjects throughout pregnancy at 12, 20, 30, and 37 weeks and in the 24 hours before and after delivery. Subjects were divided retrospectively into normotensive and preeclamptic groups. Circulating VEGF and sFlt-1 concentrations were analyzed by radioimmunoassay and enzyme-linked immunosorbent assay, respectively. RESULTS: Circulating sFlt-1 and VEGF significantly increased as gestation progressed and both were further elevated in preeclampsia compared with normotensive pregnancy. Soluble Flt-1 concentrations were elevated early in gestation and were significantly increased at 30 weeks' gestation in those who subsequently developed preeclampsia. CONCLUSION: These results indicate a definite association between elevated sFlt-1 concentrations and the onset of preeclampsia suggesting that sFlt-1 is linked with disease pathogenesis.