Pro-inflammatory profile of preeclamptic placental mesenchymal stromal cells: new insights into the etiopathogenesis of preeclampsia.

The objective of the present study was to evaluate whether placental mesenchymal stromal cells (PDMSCs) derived from normal and preeclamptic (PE) chorionic villous tissue presented differences in their cytokines expression profiles. Moreover, we investigated the effects of conditioned media from normal and PE-PDMSCs on the expression of pro-inflammatory Macrophage migration Inhibitory Factor (MIF), Vascular Endothelial Growth Factor (VEGF), soluble FMS-like tyrosine kinase-1 (sFlt-1) and free β-human Chorionic Gonadotropin (βhCG) by normal term villous explants. This information will help to understand whether anomalies in PE-PDMSCs could cause or contribute to the anomalies typical of preeclampsia. METHODS: Chorionic villous PDMSCs were isolated from severe preeclamptic (n = 12) and physiological control term (n = 12) placentae. Control and PE-PDMSCs’s cytokines expression profiles were determined by Cytokine Array. Control and PE-PDMSCs were plated for 72 h and conditioned media (CM) was collected. Physiological villous explants (n = 48) were treated with control or PE-PDMSCs CM for 72 h and processed for mRNA and protein isolation. MIF, VEGF and sFlt-1 mRNA and protein expression were analyzed by Real Time PCR and Western Blot respectively. Free βhCG was assessed by immunofluorescent. RESULTS: Cytokine array showed increased release of pro-inflammatory cytokines by PE relative to control PDMSCs. Physiological explants treated with PE-PDMSCs CM showed significantly increased MIF and sFlt-1 expression relative to untreated and control PDMSCs CM explants. Interestingly, both control and PE-PDMSCs media induced VEGF mRNA increase while only normal PDMSCs media promoted VEGF protein accumulation. PE-PDMSCs CM explants released significantly increased amounts of free βhCG relative to normal PDMSCs CM ones. CONCLUSIONS: Herein, we reported elevated production of pro-inflammatory cytokines by PE-PDMSCs. Importantly, PE PDMSCs induced a PE-like phenotype in physiological villous explants. Our data clearly depict chorionic mesenchymal stromal cells as central players in placental physiopathology, thus opening to new intriguing perspectives for the treatment of human placental-related disorders as preeclampsia

Longitudinal study of computerised cardiotocography in early fetal growth restriction.

OBJECTIVES: To explore if in early fetal growth restriction (FGR) the longitudinal pattern of short-term fetal heart rate (FHR) variation (STV) can be used for identifying imminent fetal distress and if abnormalities of FHR registration associate with two-year infant outcome. METHODS: The original TRUFFLE study assessed if in early FGR the use of ductus venosus Doppler pulsatility index (DVPI), in combination with a safety-net of very low STV and / or recurrent decelerations, could improve two-year infant survival without neurological impairment in comparison to computerised cardiotocography (cCTG) with STV calculation only. For this secondary analysis we selected women, who delivered before 32 weeks, and who had consecutive STV data for more than 3 days before delivery, and known infant two-year outcome data. Women who received corticosteroids within 3 days of delivery were excluded. Individual regression line algorithms of all STV values except the last one were calculated. Life table analysis and Cox regression analysis were used to calculate the day by day risk for a low STV or very low STV and / or FHR decelerations (DVPI group safety-net) and to assess which parameters were associated to this risk. Furthermore, it was assessed if STV pattern, lowest STV value or recurrent FHR decelerations were associated with two-year infant outcome. RESULTS: One hundred and fourty-nine women matched the inclusion criteria. Using the individual STV regression lines prediction of a last STV below the cCTG-group cut-off had a sensitivity of 0.42 and specificity of 0.91. For each day after inclusion the median risk for a low STV(cCTG criteria) was 4% (Interquartile range (IQR) 2% to 7%) and for a very low STV and / or recurrent decelerations (DVPI safety-net criteria) 5% (IQR 4 to 7%). Measures of STV pattern, fetal Doppler (arterial or venous), birthweight MoM or gestational age did not improve daily risk prediction usefully. There was no association of STV regression coefficients, a last low STV or /and recurrent decelerations with short or long term infant outcomes. CONCLUSION: The TRUFFLE study showed that a strategy of DVPI monitoring with a safety-net delivery indication of very low STV and / or recurrent decelerations could increase infant survival without neurological impairment at two years. This post-hoc analysis demonstrates that in early FGR the day by day risk of an abnormal cCTG as defined by the DVPI protocol safety-net criteria is 5%, and that prediction of this is not possible. This supports the rationale for cCTG monitoring more often than daily in these high-risk fetuses. Low STV and/or recurrent decelerations were not associated with adverse infant outcome and it appears safe to delay intervention until such abnormalities occur, as long as DVPI is in the normal range

How to monitor pregnancies complicated by fetal growth restriction and delivery below 32 weeks: a post-hoc sensitivity analysis of the TRUFFLE-study.

OBJECTIVES: In the recent TRUFFLE study it appeared that, in pregnancies complicated by fetal growth restriction (FGR) between 26 and 32 weeks, monitoring of the ductus venosus (DV) combined with computerised cardiotocography (cCTG) as a trigger for delivery, increased the chance of infant survival without neurological impairment. However, concerns in interpretation were raised as DV monitoring appeared associated with a non-significant increase in fetal death, and part of the infants were delivered after 32 weeks, after which the study protocol was no longer applied. This secondary sensitivity analysis focuses on women who delivered before 32 completed weeks, and analyses fetal death cases in detail. METHODS: We analysed the monitoring data of 317 women who delivered before 32 weeks, excluding women with absent infant outcome data or inevitable perinatal death. The association of the last monitoring data before delivery and infant outcome was assessed by multivariable analysis. RESULTS: The primary outcome (two year survival without neurological impairment) occurred more often in the two DV groups (both 83%) than in the CTG-STV group (77%), however the difference was not statistically significant (p = 0.21). Nevertheless, in surviving infants 93% was free of neurological impairment in the DV groups versus 85% in the CTG-STV group (p = 0.049). All fetal deaths (n = 7) occurred in women allocated to DV monitoring, which explains this difference. Assessment of the monitoring parameters that were obtained shortly before fetal death in these 7 cases showed an abnormal CTG in only one. Multivariable regression analysis of factors at study entry demonstrated that higher gestational age, larger estimated fetal weight 50th percentile ratio and lower U/C ratio were significantly associated with the (normal) primary outcome. Allocation to the DV groups had a smaller effect, but remained in the model (p < 0.1). Assessment of the last monitoring data before delivery showed that in the CTG-STV group abnormal fetal arterial Doppler was significantly associated with adverse outcome. In contrast, in the DV groups an abnormal DV was the only fetal monitoring parameter that was associated with adverse infant outcome, while fetal arterial Doppler, STV below CTG-group cut-off or recurrent fetal heart rate decelerations were not. CONCLUSIONS: In accordance with the results of the overall TRUFFLE study of the monitoring-intervention management of very early severe FGR we found that the difference in the proportion of infants surviving without neuroimpairment (the primary endpoint) was non-significant when comparing timing of delivery with or without changes in the DV waveform. However, the uneven distribution of fetal deaths towards the DV groups was likely by chance, and among surviving children neurological outcomes were better. Before 32 weeks, delaying delivery until abnormalities in DVPI or STV and/or recurrent decelerations occur, as defined by the study protocol, is therefore probably safe and possibly benefits long-term outcome

2 year neurodevelopmental and intermediate perinatal outcomes in infants with very preterm fetal growth restriction (TRUFFLE):a randomised trial

Background No consensus exists for the best way to monitor and when to trigger delivery in mothers of babies with fetal growth restriction. We aimed to assess whether changes in the fetal ductus venosus Doppler waveform (DV) could be used as indications for delivery instead of cardiotocography short-term variation (STV).Methods In this prospective, European multicentre, unblinded, randomised study, we included women with singleton fetuses at 26-32 weeks of gestation who had very preterm fetal growth restriction (ie, low abdominal circumference [95th percentile]). We randomly allocated women 1: 1: 1, with randomly sized blocks and stratified by participating centre and gestational age (= 29 weeks), to three timing of delivery plans, which differed according to antenatal monitoring strategies: reduced cardiotocograph fetal heart rate STV (CTG STV), early DV changes (pulsatility index &gt;95th percentile; DV p95), or late DV changes (A wave [the deflection within the venous waveform signifying atrial contraction] at or below baseline; DV no A). The primary outcome was survival without cerebral palsy or neurosensory impairment, or a Bayley III developmental score of less than 85, at 2 years of age. We assessed outcomes in surviving infants with known outcomes at 2 years. We did an intention to treat study for all participants for whom we had data. Safety outcomes were deaths in utero and neonatal deaths and were assessed in all randomly allocated women. This study is registered with ISRCTN, number 56204499.Findings Between Jan 1, 2005 and Oct 1, 2010, 503 of 542 eligible women were randomly allocated to monitoring groups (166 to CTG STV, 167 to DV p95, and 170 to DV no A). The median gestational age at delivery was 30.7 weeks (IQR 29.1-32.1) and mean birthweight was 1019 g (SD 322). The proportion of infants surviving without neuroimpairment did not differ between the CTG STV (111 [77%] of 144 infants with known outcome), DV p95 (119 [84%] of 142), and DV no A (133 [85%] of 157) groups (p(trend) = 0.09). 12 fetuses (2%) died in utero and 27 (6%) neonatal deaths occurred. Of survivors, more infants where women were randomly assigned to delivery according to late ductus changes (133 [95%] of 140, 95%, 95% CI 90-98) were free of neuroimpairment when compared with those randomly assigned to CTG (111 [85%] of 131, 95% CI 78-90; p=0.005), but this was accompanied by a non-significant increase in perinatal and infant mortality.Interpretation Although the difference in the proportion of infants surviving without neuroimpairment was non-significant at the primary endpoint, timing of delivery based on the study protocol using late changes in the DV waveform might produce an improvement in developmental outcomes at 2 years of age.</p

Phase-rectified signal averaging method to predict perinatal outcome in infants with very preterm fetal growth restriction- a secondary analysis of TRUFFLE-trial

Background Phase-rectified signal averaging, an innovative signal processing technique, can be used to investigate quasi-periodic oscillations in noisy, nonstationary signals that are obtained from fetal heart rate. Phase-rectified signal averaging is currently the best method to predict survival after myocardial infarction in adult cardiology. Application of this method to fetal medicine has established significantly better identification than with short-term variation by computerized cardiotocography of growth-restricted fetuses. Objective The aim of this study was to determine the longitudinal progression of phase-rectified signal averaging indices in severely growth-restricted human fetuses and the prognostic accuracy of the technique in relation to perinatal and neurologic outcome. Study Design Raw data from cardiotocography monitoring of 279 human fetuses were obtained from 8 centers that took part in the multicenter European “TRUFFLE” trial on optimal timing of delivery in fetal growth restriction. Average acceleration and deceleration capacities were calculated by phase-rectified signal averaging to establish progression from 5 days to 1 day before delivery and were compared with short-term variation progression. The receiver operating characteristic curves of average acceleration and deceleration capacities and short-term variation were calculated and compared between techniques for short- and intermediate-term outcome. Results Average acceleration and deceleration capacities and short-term variation showed a progressive decrease in their diagnostic indices of fetal health from the first examination 5 days before delivery to 1 day before delivery. However, this decrease was significant 3 days before delivery for average acceleration and deceleration capacities, but 2 days before delivery for short-term variation. Compared with analysis of changes in short-term variation, analysis of (delta) average acceleration and deceleration capacities better predicted values of Apgar scores <7 and antenatal death (area under the curve for prediction of antenatal death: delta average acceleration capacity, 0.62 [confidence interval, 0.19–1.0]; delta short-term variation, 0.54 [confidence interval, 0.13–0.97]; P=.006; area under the curve for prediction Apgar <7: average deceleration capacity <24 hours before delivery, 0.64 [confidence interval, 0.52–0.76]; short-term variation <24 hours before delivery, 0.53 [confidence interval, 0.40–0.65]; P=.015). Neither phase-rectified signal averaging indices nor short-term variation showed predictive power for developmental disability at 2 years of age (Bayley developmental quotient, <95 or <85). Conclusion The phase-rectified signal averaging method seems to be at least as good as short-term variation to monitor progressive deterioration of severely growth-restricted fetuses. Our findings suggest that for short-term outcomes such as Apgar score, phase-rectified signal averaging indices could be an even better test than short-term variation. Overall, our findings confirm the possible value of prospective trials based on phase-rectified signal averaging indices of autonomic nervous system of severely growth-restricted fetuses

2 Year Neurodevelopmental and Intermediate Perinatal Outcomes in Infants With Very Preterm Fetal Growth Restriction (TRUFFLE):A Randomised Trial

There is no consensus on the best methods to monitor fetal growth restriction or to trigger delivery. Previous studies have suggested that the abnormal ductus venosus (DV) pulsatility index is the best discriminating variable for neonatal outcome. This study hypothesized that changes in the fetal DV Doppler waveform, rather than cardiotocograph short-term variation, should be used as indications for delivery. The study (TRUFFLE [TRrial of Umbilical and Fetal Flow in Europe]) was conducted in 20 tertiary-care centers in 5 European countries from January 1, 2005, to October 1, 2010. Women were assigned randomly to 3 different groups. One group decided on the timing of delivery with criteria for reduced short-term variation (CTG STV). The others assessed timing of delivery based on abnormalities of the DV waveform. In the second group, delivery timing was based on early DV changes, and in the third group, delivery timing was based on late DV changes. Early DV changes were defined as pulsatility index greater than the 95th percentile (DV p95), and late DV changes were defined as no or reversed flow in the DV A wave (DV no A). Survival without neurodevelopmental impairment at 2 years of age, corrected for prematurity, was the primary outcome. Data were obtained for 503 women included in the study. Of those, 461 infants survived until 2 years of age, and 402 completed follow up at 2 years. Of the 503 infants, 354 (69%) survived without severe neonatal morbidity. One hundred forty-four infants were assigned to the CTG STV group, of whom 111 survived without impairment (77%). Of the 142 assigned to the DV-p95 group, 118 (84%) survived without impairment, and 133 (85%) of 157 survived without impairment in the DV-no-A group (P-trend = 0.09). In the CTG STV group, there were 13 deaths in 166 infants (8%); in the DV p85 group, 11 (7%) deaths of 167, and in the DV-no-A group, 17 (10%) of 170 (P-trend = 0.35). Impairment in survivors was found in 15% of the CTG STV group (20 of 131), 9% in the DV-p95 group (12 or 131), and 5% of the DV-no-A group (7 of 140) (P-trend = 0.004). No differences in short-term outcomes or baseline variables were seen between the 3 study groups. Improved outcome in survivors was shown in the DV-no-A group compared with the CTG STV group (P-trend = 0.005). At 2 years, neuroimpairment was less frequent in the DV-no-A group than in the CTG STV group, and these findings support waiting for late DV changes, contrasting with previous study suggesting a worse outcome is associated with this method