Safety and feasibility pilot study of continuous low-dose maternal supplemental oxygen in fetal single ventricle heart disease.

OBJECTIVES: Fetuses with single ventricle physiology (SVP) exhibit reductions in fetal cerebral oxygenation with associated delays in fetal brain growth and neurodevelopmental outcomes. Maternal supplemental oxygen (MSO) has been proposed to improve fetal brain growth but current evidence on dosing, candidacy, and outcomes are limited. In this pilot study, we evaluated the safety and feasibility of continuous low-dose MSO in the setting of SVP. METHODS: This single-centre, open-label, pilot phase 1 safety and feasibility clinical trial included 25 pregnant individuals with a fetal diagnosis of SVP. Participants self-administered continuous supplemental oxygen using medical-grade oxygen concentrators for up to 24 hours per day from the second half of gestation until delivery. The primary aim was the evaluation of the safety profile and feasibility of MSO. A secondary preliminary analysis was performed to assess the impact of MSO on the fetal circulation by echocardiography and late-gestational cardiovascular magnetic resonance, early outcomes including brain growth and pre-operative brain injury, and 18-month neurodevelopmental outcomes by the Bayley Scales of Infant and Toddler Development 3rd Edition compared to a contemporary fetal SVP cohort that received standard of care (SOC). RESULTS: Among 25 participants, the average maternal age at conception was 35 years, and fetal SVP diagnoses included 16 right ventricle dominant, 8 left ventricle dominant, and 1 indeterminant ventricular morphology. Participants started the trial at approximately 29.3 gestational weeks and took MSO for a median 16.1 hours per day for 63 days, cumulating a median 1029 hours of oxygen intake from enrollment until delivery. The only treatment-associated adverse events were nasal complications that were typically resolved by attaching a humidifier unit to the oxygen concentrator. No premature closure of the ductus arteriosus or unexpected fetal demise was observed. In the secondary analysis, MSO was not associated with any changes in fetal growth, middle cerebral artery pulsatility index, cerebroplacental ratio, nor head circumference to abdominal circumference ratio Z-scores over gestation compared to SOC. Although MSO was associated with changes in umbilical artery pulsatility index Z-score over gestation compared to SOC (p=0.02), this was likely due to initial baseline differences in placental resistance. At late-gestational cardiovascular magnetic resonance, MSO was not associated with any significant increase in umbilical vein oxygen saturation, fetal oxygen delivery, or fetal cerebral oxygen delivery. Similarly, we observed no differences in newborn outcomes including brain volume and pre-operative brain injury, nor mortality by 18 months of age, nor neurodevelopmental outcomes at 18 months of age. CONCLUSIONS: This pilot phase 1 clinical trial indicates low-dose maternal supplemental oxygen therapy is safe and well tolerated in pregnancies diagnosed with fetal SVP. However, our protocol was not associated with any significant changes in fetal circulatory physiology or improvements in early neurologic or neurodevelopmental outcomes. This article is protected by copyright. All rights reserved.

Peripheral cord insertion is associated with adverse pregnancy outcome only when accompanied by clinically significant placental pathology.

OBJECTIVE: To examine the relationship between umbilical cord insertion site, placental pathology and adverse pregnancy outcome in a cohort of normal and complicated pregnancies. METHODS: Sonographic measurement of the cord insertion and detailed placental pathology were performed in 309 participants. Associations between cord insertion site, placental pathology and adverse pregnancy outcome (pre-eclampsia, preterm birth, small-for-gestational age) were examined. RESULTS: A total of 93 (30%) participants were identified by pathological examination to have a peripheral cord insertion site. Only 41 of the 93 (44%) peripheral cords were detected by prenatal ultrasound. Peripherally inserted cords were associated significantly (P < 0.0001) with diagnostic placental pathology (most commonly with maternal vascular malperfusion (MVM)); of which 85% had an adverse pregnancy outcome. In cases of isolated peripheral cords, without placental pathology, the incidence of adverse outcome was not statistically different when compared to those with central cord insertion and no placental pathology (31% vs 18%; P = 0.3). A peripheral cord with an abnormal umbilical artery (UA) pulsatility index (PI) corresponded to an adverse outcome in 96% of cases compared to 29% when the UA-PI was normal. CONCLUSIONS: This study demonstrates that peripheral cord insertion is often part of the spectrum of findings of MVM disease and is associated with adverse pregnancy outcome. However, adverse outcome was uncommon when there was an isolated peripheral cord insertion and no placental pathology. Therefore, additional sonographic and biochemical features of MVM should be sought when a peripheral cord is observed. © 2023 International Society of Ultrasound in Obstetrics and Gynecology.

MRI characterization of hemodynamic patterns of human fetuses with cyanotic congenital heart disease.

OBJECTIVES: To characterize, using magnetic resonance imaging (MRI), the distribution of blood flow and oxygen transport in human fetuses with subtypes of congenital heart disease (CHD) that present with neonatal cyanosis. METHODS: Blood flow was measured in the major vessels of 152 late-gestation human fetuses with CHD and 40 gestational-age-matched normal fetuses, using cine phase-contrast MRI. Oxygen saturation (SaO2 ) was measured in the major vessels of 57 fetuses with CHD and 40 controls. RESULTS: Compared with controls, we found lower combined ventricular output in fetuses with single-ventricle physiology, with the lowest being observed in fetuses with severe forms of Ebstein's anomaly. Obstructive lesions of the left or right heart were associated with increased flow across the contralateral side. Pulmonary blood flow was reduced in fetuses with Ebstein's anomaly, while those with Ebstein's anomaly and tricuspid atresia had reduced umbilical flow. Flow in the superior vena cava was elevated in fetuses with transposition of the great arteries, normal in fetuses with hypoplastic left heart, tetralogy of Fallot or tricuspid atresia and reduced in fetuses with Ebstein's anomaly. Umbilical vein SaO2 was reduced in fetuses with hypoplastic left heart or tetralogy of Fallot. Ascending aorta and superior vena cava SaO2 were reduced in nearly all CHD subtypes. CONCLUSIONS: Fetuses with cyanotic CHD exhibit profound changes in the distribution of blood flow and oxygen transport, which result in changes in cerebral, pulmonary and placental blood flow and oxygenation. These alterations of fetal circulatory physiology may influence the neonatal course and help account for abnormalities of prenatal growth and development that have been described in newborns with cyanotic CHD. © 2021 International Society of Ultrasound in Obstetrics and Gynecology.

Customized birth-weight centiles and placenta-related fetal growth restriction.

OBJECTIVE: The value of using customized birth-weight centiles to improve the diagnostic accuracy for fetal growth restriction (FGR), in comparison with using population-based charts, remains a matter of debate. One potential explanation for the conflicting data is that most studies used measures of perinatal mortality and morbidity as proxies for placenta-mediated FGR, many of which are not specific and may be confounded by other factors such as prematurity. The aim of this study was to compare the diagnostic accuracy of small-for-gestational age (SGA) at birth, defined according to customized vs population-based charts, for associated abnormal placental pathology. METHODS: This was a secondary analysis of data from a prospective cohort study on risk factors for placenta-mediated complications and abnormal placental pathology in low-risk nulliparous women. All placentae were sent for detailed histopathological examination by two perinatal pathologists. The primary exposure was SGA, defined as birth weight < 10th centile for gestational age using either a customized (SGAcust ) or a population-based (SGApop ) birth-weight reference. The outcomes of interest were one of three types of abnormal placental pathology associated with FGR: maternal vascular malperfusion (MVM), chronic villitis and fetal vascular malperfusion (FVM). Adjusted relative risks (aRR) with 95% CIs were estimated using modified Poisson regression analysis, with adjustment for smoking, body mass index and aspirin treatment. RESULTS: A total of 857 nulliparous women met the study criteria. The proportions of infants identified as SGA based on the customized and population-based charts were 12.6% (108/857) and 11.4% (98/857), respectively. A diagnosis of SGA using either customized or population-based charts was associated with an increased risk of any placental pathology (aRR, 3.04 (95% CI, 2.29-4.04) and 1.60 (95% CI, 1.10-2.31), respectively) and MVM pathology (aRR, 12.33 (95% CI, 6.60-23.03) and 5.29 (95% CI, 2.87-9.76), respectively). SGAcust , but not SGApop , was also associated with an increased risk for chronic villitis (aRR, 1.85 (95% CI, 1.07-3.18)) and FVM pathology (aRR, 2.48 (95% CI, 1.25-4.93)). SGAcust had a higher detection rate for any placental pathology (30.3% vs 17.1%; P < 0.001), MVM pathology (63.2% vs 39.5%; P = 0.003) and chronic villitis (20.8% vs 8.3%; P = 0.007) than did SGApop , for a similar false-positive rate. This was mainly the result of a higher detection rate for abnormal pathology in the white and East-Asian subgroups and a lower false-positive rate for abnormal pathology in the South-Asian subgroup by SGAcust than by SGApop . In addition, pregnancies in the SGAcust group, but not those in the SGApop group, were more likely to be complicated by preterm birth and a low 5-min Apgar score than were the corresponding non-SGA group. CONCLUSION: These findings suggest that customized birth-weight centiles may be superior to population-based birth-weight centiles in detecting FGR that is due to underlying placental disease. © 2020 International Society of Ultrasound in Obstetrics and Gynecology.

Determination of fetal heart rate short-term variation from umbilical artery Doppler waveforms.

OBJECTIVE: To evaluate the feasibility of using umbilical artery (UA) Doppler waveforms to measure fetal heart rate (FHR) short-term variation (STV) across gestation. METHODS: This was a prospective longitudinal study, conducted at two study sites, of 195 pregnancies considered low risk. Pulsed-wave Doppler of the UAs was performed at 4-weekly intervals, between 14 and 40 weeks of gestation, using a standardized imaging protocol. Up to 12 consecutive UA Doppler waveforms were analyzed using offline processing software. FHR STV was calculated using average R-R intervals extracted from the waveforms and baseline corrected for FHR. RESULTS: Baseline-corrected FHR STV increased significantly with gestational age (conditional R2 = 0.37; P < 0.0001) and was correlated inversely with FHR (conditional R2 = 0.54; P < 0.0001). The STV ranged (median (interquartile range)) from 3.5 (2.9-4.1) ms at 14-20 weeks' gestation to 6.3 (4.8-7.7) ms at 34-40 weeks' gestation. The change in heart rate STV did not differ between study sites or individual sonographers. CONCLUSIONS: UA Doppler waveforms offer a robust and feasible method to derive STV of the FHR. It should be emphasized that the UA Doppler-derived STV is not interchangeable with measurements derived with computerized cardiotocography. Accordingly, further investigations are needed to validate associations with outcome, in order to determine the value of concurrent fetal cardiovascular and heart rate evaluations that are possible with the technique described here. © 2020 International Society of Ultrasound in Obstetrics and Gynecology.

Essential variables for reporting research studies on fetal growth restriction: a Delphi consensus.

OBJECTIVE: To determine, by expert consensus using a Delphi procedure, a minimum reporting set of study variables for fetal growth restriction (FGR) research studies. METHODS: A panel of experts, identified based on their publication record as lead or senior author of studies on FGR, was asked to select a set of essential reporting study parameters from a literature-based list of variables, utilizing the Delphi consensus methodology. Responses were collected in four consecutive rounds by online questionnaires presented to the panelists through a unique token-secured link for each round. The experts were asked to rate the importance of each parameter on a five-point Likert scale. Variables were selected in the three first rounds based on a 70% threshold for agreement on the Likert-scale scoring. In the final round, retained parameters were categorized as essential (to be reported in all FGR studies) or recommended (important but not mandatory). RESULTS: Of the 100 invited experts, 87 agreed to participate and of these 62 (71%) completed all four rounds. Agreement was reached for 16 essential and 30 recommended parameters including maternal characteristics, prenatal investigations, prenatal management and pregnancy/neonatal outcomes. Essential parameters included hypertensive complication in the current pregnancy, smoking, parity, maternal age, fetal abdominal circumference, estimated fetal weight, umbilical artery Doppler (pulsatility index and end-diastolic flow), fetal middle cerebral artery Doppler, indications for intervention, pregnancy outcome (live birth, stillbirth or neonatal death), gestational age at delivery, birth weight, birth-weight centile, mode of delivery and 5-min Apgar score. CONCLUSIONS: We present a list of essential and recommended parameters that characterize FGR independent of study hypotheses. Uniform reporting of these variables in prospective clinical research is expected to improve data quality, study consistency and ultimately our understanding of FGR. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

Molecular actions of heparin and their implications in preventing pre-eclampsia.

Pre-eclampsia, a hypertensive disorder of pregnancy, continues to be a significant cause of global maternal morbidity. Low-dose aspirin remains the only standard-of-care prophylactic therapy for preventing pre-eclampsia, but is limited in efficacy. Heparin and its derivatives may further enhance the efficacy of aspirin therapy to prevent pre-eclampsia, but the mechanisms mediating this augmentative effect are not known. Although heparin is an anticoagulant agent, it also possesses many anticoagulant-independent properties that may be relevant in the prevention of pre-eclampsia, including effects on placental, vascular and inflammatory function. This review summarizes the non-anticoagulant properties of heparin, and extrapolates how these actions may influence the trajectory of pre-eclampsia pathogenesis as a means of pathway-specific therapy.

Unfractionated heparin and placental pathology in high-risk pregnancies: secondary analysis of a pilot randomized controlled trial.

INTRODUCTION: Heparin is often prescribed during pregnancy with the intention of improving perinatal outcomes on the basis that it exerts an anticoagulant action in the inter-villous space. Accumulating in-vitro and in-vivo evidence indicates that heparin's beneficial effects in pregnancy may result from 'non-anticoagulant' effects including the promotion of angiogenesis. METHODS: To study the effect of heparin within the placenta, we performed secondary analyses on a pilot trial where 32 women with negative thrombophilia screens and second-trimester evidence of placental insufficiency were randomized to standard care or antenatal self-administration of unfractionated heparin (UFH) 7500 IU twice-daily. Serial placental ultrasound images were reviewed and compared with histo-pathologic findings following delivery. RESULTS: There were no differences between the two arms in either the evolution of abnormal placental lesions on ultrasound (p = 0.75) or evidence of maternal vascular under-perfusion on histopathology (p = 0.89). In pregnancies considered at increased risk for adverse pregnancy outcomes based on previous history or abnormal serum marker screen, early (second-trimester) placental ultrasound, reflecting developmental pathology had better test characteristics (sensitivity 77.8%; positive predictive value 80.8%) for predicting adverse pregnancy outcomes than third-trimester ultrasound that is reflective of placental thrombotic injury. CONCLUSIONS: Administration of UFH did not prevent the development or evolution of abnormal placental lesions on placental ultrasound or evidence of maternal vascular underperfusion on placental histo-pathology. Second-trimester placental ultrasound may be of value in predicting those at greatest risk of adverse outcomes.

Leptin is differentially expressed and epigenetically regulated across monochorionic twin placenta with discordant fetal growth.

Severely growth-discordant monochorionic (MC) twins offer a unique opportunity to study fetal and placental growth based on a similar genetic background and maternal host environment where the healthy twin serves as an ideal control. Differences in development of MC twins may therefore be due to differential epigenetic regulation of genes involved in placental development and function. Growth-discordant twins are known for abnormal angio-architecture in the placenta of the smaller twin. Since the reasons for this phenotype are mostly unknown this study was aimed to investigate the expression and regulation of genes known to be involved in angiogenesis. We studied 10 severely growth-discordant MC twin placentas (birthweight difference ≥20%) without twin-twin-transfusion syndrome and 5 growth-concordant MC twin placentas. Growth-discordant twin placentas were phenotyped by histology. Placental mRNA expression of 88 angiogenesis-related genes was measured by PCR array. ELISA assay and immunohistochemistry were used to confirm PCR results. EpiTYPTER for DNA methylation was used to determine if methylation ratios were responsible for differential gene expression. The PCR array analysis showed significant mRNA up-regulation in the placental share of the smaller twin for several genes. These included leptin (24.6-fold, P = 0.017), fms-like tyrosine kinase 1 (Flt1, 2.4-fold, P = 0.016) and Endoglin (Eng, 1.86-fold, P = 0.078). None of the other 84 angiogenesis-related genes showed significant differences. ELISA confirmed significantly increased leptin protein expression (49.22 versus 11.03 pg/ml, P = 0.049) in the smaller twin of the discordant growth cohort. Leptin expression in smaller twins' placentas was associated with elevated DNA methylation of the leptin promotor region suggesting the inhibition of binding of a transcriptional activator/inhibitor in that region. We attempted to overcome the limitation of sample size by careful patient selection. We minimized any bias in placental sampling by random sampling from two different sites and by avoiding sampling from areas with grossly visible abnormalities using a standardized sampling protocol. In conclusion, the smaller twin's placenta is characterized by differentially increased gene expressions for Flt1 and Eng mRNA that may be causally associated with the villous pathology driven by abnormal feto-placental angiogenesis. The substantial up-regulation of leptin mRNA may be epigenetically conferred and relevant to the post-natal risk of metabolic syndrome in intrauterine growth restriction offspring with placental pathology. Growth-discordant MC twins offer unique insights into the epigenetic basis of perinatal programming.

Emerging role of SUMOylation in placental pathology.

INTRODUCTION: Small ubiquitin-like modifiers (SUMO) conjugate to target proteins in a dynamic, reversible manner to function as post-translational modifiers. SUMOylation of target proteins can impinge on their localization, in addition to their activity or stability. Differential expression of deSUMOylating enzymes (SENP 1 and 2) contributes to altered mammalian placental development and function in mice. Severe preeclampsia (sPE) is associated with abnormal placental development and chronic ischemic injury. Extra- and intracellular stimuli/stressors that include hypoxic-activated pathways are known modulators of SUMOylation. In this current study we hypothesized that placentas from sPE patients will display up regulation in the SUMO regulatory pathway. METHODS: Utilizing qRT-PCR, immuno-blotting and Western techniques, we determined the expression levels of SUMO pathway genes in healthy and diseased placentas. We also exposed placental explants to hypoxia to study the effect on the SUMOylation pathway. RESULTS: We observed steady-state expression of SUMO1-3, SUMO-conjugated enzyme-UBC9 and deSUMOylating enzymes - SENPs, throughout normal gestation. An elevated level of free SUMO1-3 and SUMO-protein conjugates was observed in sPE placentas. Furthermore, placental UBC9 levels were strikingly increased in the same sPE patients. Hypoxia-induced SUMOylation in first trimester placental explants. DISCUSSION: Our data demonstrate an elevated steady-state of SUMOylation in sPE placentas compared with gestational aged-matched controls. The observed hyper-SUMOylation in sPE placentas correlates with elevated expression of UBC9 rather than with reduced expression of SENPs Hypoxia may contribute to alterations in placental SUMOylation pathway. CONCLUSION: Increased placental SUMOylation may contribute to the pathogenesis of serious placental pathology that causes extreme preterm birth.

Heme oxygenase-1 in placental development and pathology.

Pregnancy is accompanied by several adaptations in the mother, such as increased blood volume, higher cardiac output and reduced peripheral vascular resistance. Inability to accomplish these changes places both her and her pregnancy at risk of major placental complications such severe pre-eclampsia (sPE) or severe intra-uterine growth restriction (sIUGR). sPE is characterized by wide-spread maternal vascular dysfunction expressed as increased systemic vascular resistance; this state is accompanied by elevated levels of anti-angiogenic factors and lower production of vasodilatory gases. One of the key molecules implicated in sPE pathogenesis is heme oxygenase-1 (HO-1), a rate-limiting enzyme that breaks down heme into carbon monoxide (CO), biliverdin and free iron. CO and bilirubin (a downstream product of biliverdin processing) account for the angiogenic, vasodilatory and anti-oxidant properties of HO-1. These collective actions of the heme breakdown metabolites generated by HO-1 offer protection against cytotoxicity, inflammation, hypoxia and other forms of cellular stress that are central to the pathogenesis of sPE. Placental HO-1 expression and exhaled CO levels are both lower in women with sPE, consistent with a pathogenic role of HO-1. In vitro experiments demonstrate that induction of HO-1 downregulates secretion of the anti-angiogenic factor soluble fms-like tyrosine kinase-1 (sFLT-1) and increases CO production. Advancing our understanding of regulatory pathways promoting placental HO-1 expression may offer new pharmacological tools to reduce maternal and perinatal morbidity in severe placental insufficiency syndromes, especially in women at greatest risk of developing sPE.

Placental hyperinflation and the risk of adverse perinatal outcome.

OBJECTIVES: To determine the pathological basis and clinical associations of excessively thick placentae observed at second-trimester ultrasound examination. METHODS: In a retrospective cohort of 19 singleton high-risk second-trimester pregnancies noted to have a placental length-to-maximum thickness ratio ≤ 2.0, maximum sonographic placental thickness was correlated with clinical outcome, maximum placental thickness after delivery and placental pathological findings. Results were compared with those of an intermediate group of 21 high-risk pregnancies with normal placental dimensions and a control group of 18 low-risk pregnancies also with normal placental dimensions. Increased maximum placental thickness (> 28 mm) and abnormal placental deflation following delivery (pathology - sonography difference in maximum placental thickness < -2 mm) were defined by the upper and lower quartile values, respectively, in the control group. RESULTS: The study group exhibited significantly more adverse outcomes and gross pathological placental features compared with both intermediate and control groups. Despite increased sonographic maximum placental thickness in the study group (median, 55 (range, 40-75) mm compared with both the intermediate group (median, 27 (range, 22-41) mm, P < 0.0001) and the control group (median 26 (range, 23-36) mm, P < 0.0001)), all three groups had similar maximal placental thickness following delivery (study group: median, 24 (range, 10-50) mm vs intermediate group: median, 27 (range, 15-40) mm, P = 0.82 and vs control group: median, 28.5 (range, 18-44), P = 0.42). Pathology-sonography difference in maximum placental thickness in the study group (median, -30 (range, -42 to 0) mm) was significantly greater than that in either the intermediate (median, -2 (range, -11 to 9) mm, P < 0.0001) or the control (median, 1.5 (range, -10 to 18) mm, P < 0.0001) group and was significantly associated with abnormal development of the gas-exchanging placental villi (distal villous hypoplasia) (P = 0.0001). CONCLUSIONS: Increased second-trimester sonographic maximum placental thickness represents a pathological finding associated with severe adverse perinatal outcome. This observation is due to overinflation of the intervillous space by maternal blood rather than to adaptive formation of functional placental tissue.

Dual specificity phosphatase 9 (DUSP9) expression is down-regulated in the severe pre-eclamptic placenta.

OBJECTIVES: Intrauterine growth restriction (IUGR) and pre-eclampsia are severe and clinically important manifestations of placental insufficiency. In the mouse, dual specificity phosphatase 9 (DUSP9) is critical to the normal development of the placenta, where knock-outs are growth restricted and have a placental phenotype similar to that seen in syndromes of human placental insufficiency. Our purpose was to characterize DUSP9 expression in normal human pregnancy and in cases of placental insufficiency. STUDY DESIGN: We used RT-PCR, immuno-histochemistry and Western blotting to characterize DUSP9 gene expression and protein levels across human gestation and in pregnancies complicated by severe IUGR and/or severe pre-eclampsia. DUSP9 promoter methylation was studied in pathologic and pre-term control placentas to investigate potential epigenetic regulation. First trimester villous explants and BeWo cells were treated with DUSP9 silencing RNA to determine the effect on downstream pathways. Placental hypoxia is a hallmark of pre-eclampsia; therefore explants were subjected to hypoxic culture conditions to determine the effect of oxygen on DUSP9 expression in vitro. RESULTS: DUSP9 expression was evident in villous trophoblast and declined during development. DUSP9 protein was significantly lower in severe pre-eclamptic placentas compared to severe growth restriction. This was not epigenetically mediated by promoter hyper-methylation, and the downstream pathway ERK1/2 was not significantly affected. DUSP9 expression in first trimester explants was significantly decreased by 74 ± 20% in hypoxic (3% oxygen) culture conditions. In BeWo cells and explanted placental villi treated with DUSP9 silencing RNA, expression of DUSP9 was down-regulated by 61% and 62% respectively. There was a trend to increased phosphorylation of the downstream target ERK1/2 in DUSP9 down-regulated BeWo cells and explanted placental villi. CONCLUSION: DUSP9 protein levels were markedly suppressed in severe pre-eclampsia, but not in severe IUGR. This suppression might be attributable to the prolonged hypoxic conditions found in pre-eclampsia.