Decidual mesenchymal stem/stromal cells from preeclamptic patients secrete endoglin, which at high levels inhibits endothelial cell attachment invitro.

INTRODUCTION: In preeclampsia (PE), inadequate remodelling of spiral arterioles in the decidua basalis causes oxidative stress and subsequent increased release of antiangiogenic soluble endoglin (sENG) into the maternal circulation. Decidual mesenchymal stem/stromal cells (DMSCs) reside adjacent to endothelial cells in this vascular niche. Surprisingly, DMSCs express membrane-bound ENG (CD105). PE-affected DMSCs (PE-DMSCs) are abnormal and due to reduced extravillous invasion, more of them are present, but the significance of this is not known. METHODS: DMSCs were isolated and characterised from normotensive control and severe-PE placentae. Extracellular vesicle (EV) types, shed microvesicles (sMV) and exosomes, were isolated from DMSC conditioned media (DMSCCM), respectively. Secretion of ENG by DMSCs was assessed by ELISA of DMSCCM, with and without EV depletion. The effects of reducing ENG concentration, by blocking antibody, on human umbilical vein endothelial cell (HUVEC) attachment were assessed by xCELLigence real-time functional assays. RESULTS: ENG was detected in DMSCCM and these levels significantly decreased when depleted of exosomes and sMV. There was no significant difference in the amount of ENG secreted by control DMSCs and PE-DMSCs. Blocking ENG in concentrated DMSCCM, used to treat HUVECs, improved endothelial cell attachment. DISCUSSION: In normotensive pregnancies, DMSC secretion of ENG likely has a beneficial effect on endothelial cells. However, in PE pregnancies, shallow invasion of the spiral arterioles exposes more PE-DMSC derived sources of ENG (soluble and EV). The presence of these PE-DMSCs in the vascular niche contributes to endothelial cell dysfunction.

Decreased placental glypican expression is associated with human fetal growth restriction.

Placental mediated fetal growth restriction (FGR) is a leading cause of perinatal morbidity and mortality. Heparan sulphate proteoglycans (HSPG) are highly expressed in placentae and regulate haemostasis. We hypothesise that altered expression of HSPGs, glypicans (GPC) may contribute to the development of FGR and small-for-gestational-age (SGA). GPC expression was determined in first-trimester chorionic villous samples collected from women with later SGA pregnancies and in placentae from third-trimester FGR and gestation-matched uncomplicated pregnancies. The expression of both GPC1 and GPC3 were significantly reduced in first-trimester SGA as well as in the third-trimester FGR placentae compared to controls. This is the first study to report a relationship between altered placental GPC expression and subsequent development of SGA/FGR.

Soluble fms-like tyrosine kinase-1 to placental growth factor ratio: ruling out pre-eclampsia for up to 4 weeks and value of retesting.

OBJECTIVES: The soluble fms-like tyrosine kinase-1 (sFlt-1) to placental growth factor (PlGF) ratio is generally elevated some time before and at the clinical onset of pre-eclampsia. The PROGNOSIS study validated a sFlt-1/PlGF ratio cut-off of ≤ 38 to rule out the onset of pre-eclampsia within 1 week of testing in women with suspected disease. The aim of this study was to assess the predictive value of the sFlt-1/PlGF ratio to rule out the onset of pre-eclampsia for up to 4 weeks, and to assess the value of repeat measurements. METHODS: This was an exploratory post-hoc analysis of data from the PROGNOSIS study performed in pregnant women aged ≥ 18 years with suspected pre-eclampsia, who were at 24 + 0 to 36 + 6 weeks' gestation at their first clinic visit. Serum samples were collected at the first visit and weekly thereafter. sFlt-1 and PlGF levels were measured using Elecsys® sFlt-1 and PlGF immunoassays. Whether the sFlt-1/PlGF ratio cut-off of ≤ 38 used to rule out the onset of pre-eclampsia within 1 week could predict the absence of pre-eclampsia 2, 3, and 4 weeks post-baseline was assessed. The value of repeat sFlt-1/PlGF testing was assessed by examining the difference in sFlt-1/PlGF ratio 2 and 3 weeks after the first measurement in women with, and those without, pre-eclampsia or adverse fetal outcome. RESULTS: On analysis of 550 women, sFlt-1/PlGF ratio ≤ 38 ruled out the onset of pre-eclampsia 2 and 3 weeks post-baseline with high negative predictive values (NPV) of 97.9% and 95.7%, respectively. The onset of pre-eclampsia within 4 weeks was ruled out with a high NPV (94.3%) and high sensitivity and specificity (66.2% and 83.1%, respectively). Compared with women who did not develop pre-eclampsia, those who developed pre-eclampsia had significantly larger median increases in sFlt-1/PlGF ratio at 2 weeks (∆, 31.22 vs 1.45; P < 0.001) and at 3 weeks (∆, 48.97 vs 2.39; P < 0.001) after their initial visit. Women who developed pre-eclampsia and/or adverse fetal outcome compared with those who did not had a significantly greater median increase in sFlt-1/PlGF ratio over the same period (∆, 21.22 vs 1.40; P < 0.001 at 2 weeks; ∆, 34.95 vs 2.30; P < 0.001 at 3 weeks). CONCLUSION: The Elecsys® immunoassay sFlt-1/PlGF ratio can help to rule out the onset of pre-eclampsia for 4 weeks in women with suspected pre-eclampsia. © 2018 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.

Decorin expression is decreased in first trimester placental tissue from pregnancies with small for gestation age infants at birth.

Fetal growth restriction (FGR) is a leading cause of perinatal morbidity and mortality. FGR pregnancies are often associated with histological evidence of placental vascular thrombosis. The proteoglycans are important components and regulators of vascular homeostasis. Previous studies from our laboratory highlighted mRNA and protein expression differences in placental proteoglycan decorin (DCN), within a clinically well-characterised cohort of third-trimester idiopathic FGR compared with gestation-matched uncomplicated control pregnancies. We also showed that decorin contributes to abnormal angiogenesis and increased thrombin generation in vitro. These observations suggest that DCN gene expression may contribute to the etiology of FGR. Small for gestational age (SGA) is frequently used as a proxy for FGR and is defined as a birth weight below the 10th percentile of a birth weight curve. We therefore made use of a unique resource of first trimester tissues obtained via chorionic villus sampling during the first trimester to investigate the temporal relationship between altered DCN expression and any subsequent development of SGA. We hypothesized that placental DCN expression is decreased early in gestation in SGA pregnancies. Surplus chorionic villus specimens from 15 women subsequently diagnosed with FGR and 50 from women with uncomplicated pregnancies were collected. DCN mRNA and DCN protein were determined using real-time PCR and immunoblotting, respectively. Both DCN mRNA and protein were significantly decreased in placentae from first-trimester SGA-pregnancies compared with controls (p < 0.05). This is the first study to report a temporal relationship between altered placental DCN expression and subsequent development of SGA.

Placental vitamin D receptor expression is decreased in human idiopathic fetal growth restriction.

UNLABELLED: Fetal growth restriction (FGR) affects up to 5 % of pregnancies worldwide, and trophoblast function plays a significant role on the outcome. An epidemiological study has linked vitamin D deficiency to adverse perinatal outcomes, which include decreased birth weight. The placenta as an important source of vitamin D regulates its metabolism through the vitamin D receptor (VDR), but the mechanism by which VDR regulates trophoblast function is poorly understood. Our study aimed at determining placental VDR expression in FGR and gestation-matched control (GMC) pregnancies and identifying the actions of VDR in trophoblast differentiation and apoptosis. Placentae were collected from a well-defined cohort of idiopathic FGR and GMC pregnancies. VDR mRNA and protein expressions were determined by PCR, immunohistochemistry and immunoblotting, while functional consequences of VDR inactivation in vitro were determined on BeWo cells by determining changes in differentiation, attachment and apoptosis. Significant decreases in VDR mRNA expression (p = 0.0005) and protein expression (p = 0.0003) were observed in the FGR samples, while VDR inactivation, which showed markers for differentiation, cell attachment and apoptosis, was significantly increased. Thus, decreased placental VDR may contribute to uncontrolled premature differentiation and apoptosis of trophoblasts that are characteristics of idiopathic FGR pregnancies. KEY MESSAGE: Fetal growth restriction (FGR) affects up to 5 % of all pregnancies worldwide. FGR is the second highest cause of perinatal mortality and morbidity. The placenta plays a pivotal role in vitamin D metabolism during pregnancy. Vitamin D deficiency is associated with adverse pregnancy outcomes. Placental vitamin D receptor expression is decreased in FGR.

Mesenchymal stem cells reside in a vascular niche in the decidua basalis and are absent in remodelled spiral arterioles.

INTRODUCTION: Maternal decidua basalis tissue attached to the placenta following delivery is a source of decidual mesenchymal stem cells (DMSCs). The in vitro characteristics of DMSCs have been partly defined but their in vivo function(s) are poorly understood. The anatomic location, or niche, provides clues regarding potential in vivo function(s) of DMSCs, but the niche has not been described. METHODS: Cells were isolated from the decidua basalis and flow cytometric analyses showed the expected phenotypic profile for MSC cell surface markers. In vitro, the cells differentiated into adipocytes, osteocytes, and chondrocytes. DMSCs were then stained with antibodies by immunofluorescence detection. RESULTS: Immunocytochemistry revealed that DMSCs were positive for FZD-9, STRO-1, 3G5, and α-SMA as expected and lacked expression of vWF and Ck7. Fluorescence in situ hybridization analysis showed the cultured cells were of maternal origin. Immunofluorescence was carried out on placental bed biopsies using the FZD-9, STRO-1, 3G5, and α-SMA antibodies. DMSCs were located in the vascular niche in decidua basalis. Immunofluorescence with antibodies to FZD-9, Ck7 and vWF revealed DMSCs in the vascular niche surrounding intact non-transformed spiral arterioles but DMSCs were absent in fully transformed spiral arterioles. DISCUSSION: Spiral arteriole remodelling is a critical feature of human pregnancy. The DMSC niche was investigated in fully transformed and non-transformed spiral arterioles. DMSCs have not been previously implicated in spiral arteriole remodelling. The absence of DMSCs around fully transformed spiral arterioles suggests they are a target for replacement or destruction by invading placental extravillous trophoblast cells, which carry out spiral arteriole remodelling.

Altered decorin leads to disrupted endothelial cell function: a possible mechanism in the pathogenesis of fetal growth restriction?

OBJECTIVE: Fetal growth restriction (FGR) is a key cause of adverse pregnancy outcome where maternal and fetal factors are identified as contributing to this condition. Idiopathic FGR is associated with altered vascular endothelial cell functions. Decorin (DCN) has important roles in the regulation of endothelial cell functions in vascular environments. DCN expression is reduced in FGR. The objectives were to determine the functional consequences of reduced DCN in a human microvascular endothelial cell line model (HMVEC), and to determine downstream targets of DCN and their expression in primary placental microvascular endothelial cells (PLECs) from control and FGR-affected placentae. APPROACH: Short-interference RNA was used to reduce DCN expression in HMVECs and the effect on proliferation, angiogenesis and thrombin generation was determined. A Growth Factor PCR Array was used to identify downstream targets of DCN. The expression of target genes in control and FGR PLECs was performed. RESULTS: DCN reduction decreased proliferation and angiogenesis but increased thrombin generation with no effect on apoptosis. The array identified three targets of DCN: FGF17, IL18 and MSTN. Validation of target genes confirmed decreased expression of VEGFA, MMP9, EGFR1, IGFR1 and PLGF in HMVECs and PLECs from control and FGR pregnancies. CONCLUSIONS: Reduction of DCN in vascular endothelial cells leads to disrupted cell functions. The targets of DCN include genes that play important roles in angiogenesis and cellular growth. Therefore, differential expression of these may contribute to the pathogenesis of FGR and disease states in other microvascular circulations.

Increased decidual mRNA expression levels of candidate maternal pre-eclampsia susceptibility genes are associated with clinical severity.

INTRODUCTION: Pre-eclampsia (PE) has a familial association, with daughters of women who had PE during pregnancy having more than twice the risk of developing PE themselves. Through genome-wide linkage and genetic association studies in PE-affected families and large population samples, we previously identified the following as positional candidate maternal susceptibility genes for PE; ACVR1, INHA, INHBB, ERAP1, ERAP2, LNPEP, COL4A1 and COL4A2. The aims of this study were to determine mRNA expression levels of previously identified candidate maternal pre-eclampsia susceptibility genes from normotensive and severe PE (SPE) pregnancies and correlate mRNA expression levels with the clinical severity of SPE. METHODS: Third trimester decidual tissues were collected from both normotensive (n = 21) and SPE pregnancies (n = 24) and mRNA expression levels were determined by real-time PCR. Gene expression was then correlated with several parameters of clinical severity in SPE. Statistical significance was determined by Mann-Whitney U test and Spearman's Correlation. RESULTS: The data demonstrate significantly increased decidual mRNA expression levels of ACVR1, INHBB, ERAP1, ERAP2, LNPEP, COL4A1 and COL4A2 in SPE (p < 0.05). Increased mRNA expression levels of several genes - INHA, INHBB, COL4A1 and COL4A2 were correlated with earlier onset of PE and earlier delivery of the fetus (p < 0.05). CONCLUSION: These results suggest altered expression of maternal susceptibility genes may play roles in PE development and the course of disease severity.

Decidua parietalis-derived mesenchymal stromal cells reside in a vascular niche within the choriodecidua.

Mesenchymal stromal cells (MSCs) from gestational tissues represent promising cell populations with stem cell-like properties for use in regenerative medicine. Previously, we reported that MSCs in the chorionic villi of the human placenta reside in a vascular niche. However, the niche(s) in which MSCs reside in the fetal membranes, another rich source of MSCs, remains to be determined. The cell surface markers STRO-1 and 3G5 were previously employed to identify niches in a variety of tissues and here we use these markers to report the location of the MSC niche in the human decidua parietalis. The cultured decidua parietalis MSCs (DPMSCs) isolated from the choriodecidua component of the fetal membranes possessed stem cell-like properties such as adherence to plastic, colony forming ability, and multipotent differentiation potential. Fluorescence in situ hybridization analysis showed cultured DPMSCs were of maternal origin. Immunocytochemistry demonstrated that cultured DPMSCs stained positively with stem cell surface markers 3G5, CD105, CD106, STRO-1, CD146, CD49a, and α-SMA but were negative for hematopoietic markers (CD117, CD34) and vascular markers (CD34, von Willebrand factor [vWF]). Immunohistochemistry with antibodies to stem cell surface markers and the endothelial markers on term fetal membranes revealed a vascular niche for DPMSCs, which was confirmed by immunofluorescence analysis. Both STRO-1 and vWF fluorescence signals showed substantial overlap, while CD146 and vWF signals showed partial overlap. These observations were consistent with a vascular niche.

Placental CLIC3 is increased in fetal growth restriction and pre-eclampsia affected human pregnancies.

Chloride intracellular channel (CLIC) proteins constitute a subgroup of the glutathione-S-transferase (GSTs) superfamily. In humans, the CLIC family of proteins consists of six members, designated CLIC 1-6, which have a conserved C-terminal 240 residue module and one major transmembrane domain. CLIC proteins regulate fundamental cellular processes including regulation of chloride ion concentration, stabilization of cell membrane potential, trans-epithelial transport, regulation of cell volume and stimulation of apoptotic processes in response to cellular stress. Previously, we described the expression profile of a member of the CLIC family of proteins, CLIC3, in human placentae and fetal membranes. In the current study, we determined CLIC3 expression in placentae from pregnancies complicated with either fetal growth restriction (FGR, n=19), pre-eclampsia (PE, n=16) or both FGR and PE combined (n=12) compared to gestation-matched controls (n=13) using real-time PCR and a CLIC3 specific immunoassay. Significantly increased CLIC3 mRNA and protein were detected in placental extracts from pregnancies with FGR, PE and PE with FGR compared to controls. Our results suggest that increased expression of CLIC3 may play a role in abnormal placental function associated with the human pregnancy disorders FGR and PE.

The expression of placental proteoglycans in pre-eclampsia.

BACKGROUND/AIMS: Pre-eclampsia (PE) is one of the leading causes of maternal and perinatal morbidity and mortality. PE is defined clinically as the onset of maternal hypertension and proteinuria following 20 weeks of gestation. It is associated with altered maternal uterine decidual spiral artery remodelling, which may lead to reduced blood flow and increased thrombosis within the uteroplacental vasculature. Proteoglycans (PGs) are macromolecules which have (in combination with glycosaminoglycans) important anticoagulant roles in vascular endothelial environments, including the uteroplacental circulation. The hypothesis under consideration in this study was that differential expression of placental PGs may be associated with PE. METHODS: PE and control placental samples were collected with ethics approval and patient consent. RNA and protein were extracted and real-time PCR and Western immunoblotting were performed to determine the expression of the PGs in the samples. RESULTS: Of the nine PGs investigated, none showed increased expression, whereas the mRNA and protein expression of five of them was significantly decreased in the placentae of pre-eclamptic women compared to gestation-matched controls. CONCLUSION: Therefore, the results of this study support the hypothesis that a placental PG deficiency may contribute to the placental thrombotic lesions characteristic of PE.

The PAI-1 4G/5G polymorphism is not associated with an increased risk of adverse pregnancy outcome in asymptomatic nulliparous women.

BACKGROUND: Plasminogen activator inhibitor type 1 (PAI-1) is an important regulator of fibrinolysis. A common deletion polymorphism that results in a sequence of 4G instead of 5G in the promoter region of the gene is associated with a small increase in the risk of venous thromboembolism. Its potential association with adverse pregnancy events remains controversial. OBJECTIVE: We aimed to assess the impact of the 4G PAI-1 polymorphism on pregnancy outcomes in women who had no prior history of adverse pregnancy outcomes or personal or family history of venous thromboembolism. PATIENTS/METHODS: This study represents a secondary investigation of a prior prospective cohort study investigating the association between inherited thrombophilias and adverse pregnancy events in Australian women. Healthy nulliparous women were recruited to this study prior to 22 weeks gestation. Genotyping for the 4G/5G PAI-1 gene was performed using Taqman assays in an ABI prism 7700 Sequencer several years after the pregnancy was completed. Pregnancy outcome data were extracted from the medical record. The primary outcome was a composite comprising development of severe pre-eclampsia, fetal growth restriction, major placental abruption, stillbirth or neonatal death. RESULTS: Pregnancy outcome data were available in 1733 women who were successfully genotyped for this polymorphism. The primary composite outcome was experienced by 139 women (8% of the cohort). Four hundred and fifty-nine women (26.5%) were homozygous for the 4G deletion polymorphism, while 890 (51.4%) were heterozygous. Neither homozygosity nor heterozygosity for the PAI-1 4G polymorphism was associated with the primary composite outcome (homozygous OR = 1.30, 95% CI = 0.81-2.09, P = 0.28, heterozygous OR = 0.84, 95% CI = 0.53-1.31, P = 0.44) or with the individual pregnancy complications. CONCLUSION: The PAI-1 4G polymorphism is not associated with an increase in the risk of serious adverse pregnancy events in asymptomatic nulliparous women.

Calreticulin has opposing effects on the migration of human trophoblast and myometrial endothelial cells.

Calreticulin is a calcium binding, endoplasmic reticulum resident protein best known for its roles in intracellular calcium homeostasis and the quality control processes of the endoplasmic reticulum. There is evidence for a range of activities for calreticulin outside the endoplasmic reticulum, including in the cytosol, on the surface of different cells types and in the extracellular matrix. Recent evidence indicates that human pregnancy is a condition of elevated circulating calreticulin. Calreticulin was increased in the plasma of women throughout pregnancy compared to the non-pregnant state. Calreticulin was also further increased during the hypertensive disorder of human pregnancy, pre-eclampsia. To clarify the roles of circulating calreticulin in pregnancy and pre-eclampsia, the aim of this study was to determine the effects of exogenous calreticulin on two cell types that are relevant to normal human pregnancy and to pre-eclampsia. Human primary myometrial microvascular endothelial cells (UtMVEC-Myo) and the human trophoblast cell line, HTR8/SVneo, were cultured with exogenous calreticulin at concentrations (2 µg/ml and 5 µg/ml) comparable to that measured in maternal blood. The higher concentration of calreticulin significantly increased the migration of the UtMVEC-Myo cells, but significantly reduced the migration of the HTR8/SVneo cells. In the presence of only FGF, FBS and antibiotics calreticulin at 5 µg/ml significantly reduced the number of UtMVEC-Myo cells during in vitro culture for 120 h. These results demonstrate that exogenous calreticulin can alter both HTR8/SVneo and UtMVEC-Myo cell functions in vitro at a (patho-) physiologically relevant concentration. Increased calreticulin may also contribute to altered functions of both cell types during pre-eclampsia.

OS070. Shared genetic risk factors for preeclampsia and cardiovascular disease.

INTRODUCTION: There is compelling evidence to support the hypothesis that a maternal constitutional predisposition to cardiovascular disease (CVD) is a key component in development of preeclampsia. In particular, CVD and preeclampsia share pathological features such as endothelial dysfunction and inflammation, and have several metabolic abnormalities in common. In support of this hypothesis, our recent genetic dissection of the Australian preeclampsia susceptibility locus on chromosome 2q22 revealed shared novel genetic risk factors for preeclampsia and CVD-related traits. OBJECTIVES: To replicate association between our recently reported 2q22 preeclampsia risk variants and CVD-related traits in an independent Australian population based cohort. METHODS: Four independent SNPs from four genes, rs35821928 (LRP1B), rs17783344 (GCA), rs115015150 (RND3) and rs2322659 (LCT), were recently found to be significantly associated with preeclampsia susceptibility and CVD-related traits. These SNPs were genotyped in a large independent Australian cohort rich in quantitative CVD risk traits; The Western Australian Pregnancy Cohort (Raine) Study. This cohort comprises of blood samples from 1246 mothers and 1461 adolescents and clinical measures such as, but not limited to, anthropometric measures of adiposity and lipid-related measures. Genetic association analyses of these four potential preeclampsia susceptibility SNPs against the CVD-related risk traits were performed using the software package R. All statistical analyses assumed an additive model of gene action. RESULTS: Several significant associations (p<0.05) for all four SNPs with a variety of CVD-related risk traits were detected, both for the mothers and the adolescents. The LRP1B SNP was associated with HDL/cholesterol ratio, LDL cholesterol, triglycerides, skinfold measures and weight. The GCA SNP was associated with total cholesterol, HDL cholesterol, serum insulin, hemoglobin, blood glucose, BMI and skinfold measures. The RND3 SNP was associated with triglycerides and waist-hip ratio. The LCT SNP was associated with hemoglobin, blood glucose and abdominal skinfold. CONCLUSION: We have recently identified genetic variants within the LRP1B, GCA, RND3 and LCT genes to be significantly associated with preeclampsia susceptibility and CVD-related risk traits. We have now demonstrated thatthese specific genetic variants are associated with CVD-related risk traits in an independent population. Our collective findings provide substantial empirical data to support the hypothesis that genetic risk factors for preeclampsia and CVD are, at least in part, shared.

OS074. Brachial artery flow-mediated dilatation as a predictor of pre-eclampsia in the first trimester of pregnancy.

INTRODUCTION: Increased vascular reactivity secondary to endothelial injury has been reported to be involved in the pathogenesis of pre-eclampsia (PE). However, whether these observations of vascular activity are the result or the cause of PE is still not fully understood. Some studies have shown that pregnant women who subsequently suffer from PE have an impaired flow-mediated dilatation in the brachial artery at the beginning of the second half of pregnancy. However this parameter has not been tested as a first trimester predictor of PE. OBJECTIVES: To investigate the role of maternal brachial artery reactivity during the first trimester of pregnancy (11-14 weeks) for the prediction of PE. METHODS: Prospective study with singleton pregnancies examined at 11-14 weeks of gestation, presenting consecutively for antenatal care in a tertiary Brazilian hospital. Maternal endothelial dysfunction was assessed by flow-mediated dilatation (FMD) of the brachial artery. FMD was assessed in 550 pregnant women by the same examiner. Obstetric and perinatal outcomes were available in 506 patients (92%). A total of 19 cases were excluded, because 6 miscarried, 12 presented isolated fetal growth restriction (FGR) and 1 was considered an "outlier" because of a significant inflammatory medical co-morbidity. The main outcome was PE based on the ISSHP classification, and we considered early-onset PE when delivery occurred before 34 weeks and late-onset PE when delivery occurred after 34 weeks. RESULTS: In a total of 487 patients, 31 cases developed PE (6.3%), with 9 cases (1.8%) of early-PE and 22 cases (4.5%) of late-PE. The mean gestational age of the study was 12 weeks (range 11-14 weeks).The diameter of the brachial artery at rest was similar between control and PE groups (2.83mm and 2.93mm; p = 0.60) and the resistance and pulsatility index measurements also showed no significant differences among these groups. The mean FMD was 7.4 ± 8.2% in the control group and 7.3 ± 8.2% in the PE group. Logistic regression analysis determined that FDM was not a predictor of PE (OR=0.99, CI 95% 0.94-1.04; p=0.90). Dividing the cases of PE in early and late-onset, it was also observed that the mean diameter of the brachial artery at rest was similar among these groups, and that after the shear stress test the diameter was not significantly different in patients who developed early-PE. CONCLUSION: Maternal brachial artery reactivity assessed by FMD in the first trimester of pregnancy is not a predictor of PE.

OS088. First trimester triple vascular test for pre-eclampsia prediction.

INTRODUCTION: Although PE represents a major cause of maternal and fetal morbidity and mortality, the vascular mechanisms underlying this disorder have not been clearly identified. During the past three decades, while numerous clinical, biophysical, and biochemical screening tests have been proposed for the early detection of preeclampsia, maternal circulation changes during early pregnancy have yet to be fully evaluated for their contribution to PE prediction. OBJECTIVES: The aim of this study was to examine a combination of maternal risk factors, mean arterial blood pressure, uterine artery Doppler, brachial artery flow-mediated dilatation (FMD), and ophthalmic artery Doppler for pre-eclampsia prediction during the first trimester of pregnancy. METHODS: Prospective study with singleton pregnancies examined at 11-14 weeks of gestation, presenting consecutively for antenatal care in a tertiary Brazilian hospital. The base-cohort population constituted of 487 singleton pregnancies, including 9 case subjects who developed pre-eclampsia (PE) requiring delivery before 34 weeks (early PE) and 22 with late PE, 47 with gestational hypertension, and 409 cases subjects (84%) who were unaffected by PE or gestational hypertension. Maternal history (nulliparity, previous and family history of PE), body mass index (BMI), mean arterial pressure (MAP), uterine artery pulsatility index, brachial artery FMD and ophthalmic artery Doppler were recorded in all of the cases. Univariate and logistic regression analysis was used to derive algorithms for the prediction of hypertensive disorders. RESULTS: Uterine artery percentile of mean PI was higher in the PE than in the control group (p<0.01). The mean brachial artery FMD was 7.4%±8.2% in the control group and 7.3%±8.2% in the PE group. Logistic regression analysis determined that FDM was not a predictor of PE (OR=0.99, CI 95% 0.94-1.04; p=0.90) and this test was withdrawn from the predictive model. The average of the first diastolic peak velocity in the ophthalmic artery was higher in the PE group compared with controls (24.56cm/s×21.13cm/s; p<0.01).It was estimated that, with the prediction algorithm for PE, a combination of maternal factors + MAP + uterine artery Doppler or ophthalmic artery Doppler can detect 78% of early-onset PE with 10% false-positive rate. CONCLUSION: Maternal ophthalmic artery Doppler in the first trimester of pregnancy is a novel predictive parameter for PE (especially early-onset PE), it has the same detection rate contribution in a multi-parameter predictive model as would be the case uterine artery Doppler was used instead.

PP114. First trimester multi-parameter prediction of pre-eclampsia.

INTRODUCTION: Pre-eclampsia (PE), which affects about 3-5% of pregnant women, is the most frequent serious medical complication in pregnancy and a major cause of maternal and perinatal morbidity and mortality. During the past three decades, numerous clinical, biophysical, and biochemical screening tests have been proposed for the early detection of PE. Literature shows large variations in the sensitivity and predictive value of these tests. No single screening test used for PE prediction has gained widespread acceptance into clinical practice. Instead, panels of tests, which combine several clinical measurements, seem to be of more value for increasing the predictive value for PE. OBJECTIVES: The aim of this study was to examine a combination of maternal risk factors, mean arterial blood pressure, and uterine artery Doppler for pre-eclampsia prediction during the first trimester of pregnancy. METHODS: Prospective study with singleton pregnancies examined at 11-14 weeks of gestation, presenting consecutively for antenatal care in a tertiary Brazilian hospital. The base-cohort population was 487 singleton pregnancies, including nine case subjects who developed PE requiring delivery before 34 weeks (early PE) and 22 with late PE, 47 with gestational hypertension, and 409 cases subjects (84%) who were unaffected by PE or gestational hypertension. Maternal history, body mass index (BMI), mean arterial pressure (MAP), and uterine artery pulsatility index were recorded in all of the cases. Univariate and logistic regression analysis was used to derive algorithms for the prediction of hypertensive disorders. RESULTS: The maternal characteristics selected by regression analysis to be part of the final predictive model were nulliparity, previous personal and family history of PE. MAP was higher (86 versus 78 mmHg) in patients who developed PE (p<0.01). The uterine artery percentile of mean PI was higher in the PE than in the control group (50.3%±31.7% versus 37.4%±30.0%; p<0.01). It was estimated that, with the algorithm for PE, 78%, 45%, and 26% of early PE, late PE, and gestational hypertension, respectively, could be detected with a 10% false-positive rate. CONCLUSION: The traditional approach to screening for PE, which is based on maternal demographic characteristics and medical history, identifies ∼60% of cases destined to develop early PE for a false-positive rate of 10%. This study proposes that a combination of maternal risk factors, mean arterial blood pressure, and uterine artery Doppler, for the same false-positive rate of 10%, could identify 78% of cases of early PE.

Homeobox gene Distal-less 3 (DLX3) is a regulator of villous cytotrophoblast differentiation.

Dlx3, a member of the large homeobox gene family of transcription factors, is important for murine placental development. Targeted deletion of Dlx3 in the mouse results in embryonic death due to placental failure. This study investigated the role of human DLX3 in villous cytotrophoblast (VCT) differentiation in the placenta. Primary VCT from human term placentae, which spontaneously differentiate when maintained in culture over 72 h, showed a significant increase in mRNA and protein expression of DLX3 and 3ßHSD. The functional role of DLX3 was determined using trophoblast derived-cell line, BeWo. Forskolin treated BeWo cells showed significantly increased DLX3 mRNA and protein expression. Forskolin stimulation also showed a significant increase in syncytin and 3ßHSD mRNA expression, and increased release of ßhCG into the cell culture supernatant. To determine whether DLX3 had a direct or indirect effect on VCT differentiation, mRNA and protein expression of DLX3 was increased using a plasmid DLX3 over-expression construct. Over-expression of DLX3 resulted in increased mRNA expression of 3ßHSD and syncytin, as well as increased secretion of ß-hCG protein in the cell culture medium. In conclusion, we provide evidence that DLX3 acts upstream of syncytin, 3ßHSD and ßhCG and that DLX3 has a regulatory role in VCT differentiation.