Ethnicity and Language Proficiency Differences in the Provision of and Intention to Use Prenatal Screening for Down's Syndrome and Congenital Anomalies. A Prospective, Non-selected, Register-Based Study in the Netherlands.

Objective We aimed to conduct an analysis of the associations between the information provision procedure of prenatal screening for Down's syndrome and congenital anomalies and the intention to participate in prenatal screening (PS) of ethnicity groups and Dutch language proficiency groups. Design Using a prospective web-based registration form, we asked counselors (midwives, general practitioners, nurses and gynecologists) to report whether and how they offered information about PS to pregnant women. Duration The study was conducted from 2008 to 2010. Participants We collected data on the characteristics of the women who received an information offer about PS from counselors. Measurements Measures included socio-demographic and language proficiency level (LPL) characteristics, key elements of the provision procedure of PS, and intentional participation in PS. Findings The dataset represents 37% of the total population in the study area. Women with a non-native Dutch background and/or insufficient Dutch LPL received fewer information offers about PS, faced a reduced chance of receiving counseling, and showed lower intentional participation rates for PS. Key Conclusions Women with a non-native Dutch background and/or with an insufficient LPL are underserved in the Dutch PS program. These findings present evidence indicating that the fundamental principle of the Dutch Population Screening Act, namely, equal access to PS for all pregnant women, is not being realized. Implications for Practice Therefore, the study findings are important for national and international healthcare, policy makers and governmental professionals to allow ethnic and LPL-related differences in the provision and intentional uptake of PS.

An Audit of Second-Trimester Fetal Anomaly Scans Based on a Novel Image-Scoring Method in the Southwest Region of the Netherlands.

OBJECTIVES: Since 2007 the second-trimester fetal anomaly scan is offered to all pregnant women as part of the national prenatal screening program in the Netherlands. Dutch population-based screening programs generally have a well-described system to achieve quality assurance. Because of the absence of a uniform system to monitor the actual performance of the fetal anomaly scan in 2012, we developed a standardized image-scoring method. The aim of this study was to evaluate the scanning performance of all sonographers in the southwestern region of the Netherlands using this image-scoring method. METHODS: Each sonographer was requested to set up a digital portfolio. A portfolio consists of five logbooks from five different pregnant women, each containing 25 fetal anatomical structures and six biometric measures of randomly selected fetal anomaly scans. RESULTS: During the study period, 425 logbooks of 85 sonographers were assessed as part of the audit process. Seventy-three out of 85 sonographers (86%) met the criteria in the primary audit, and 12 sonographers required individual hands-on training. A successful assessment was achieved for 11 sonographers in the re-audit and one sonographer ceased her contract. Moreover, 2.1% of the required images were not digitally stored and therefore could not be reviewed. CONCLUSIONS: Quality assessment using the image-scoring method demonstrated that most of the sonographers met the expectations of the audit process, but those who had subpar performance met the expectations after retraining.

The relation between viscero-abdominal disproportion and type of omphalocele closure.

OBJECTIVE: To investigate the relation between prenatal ultrasound measurements of viscero-abdominal disproportion and the expected type of postnatal surgical closure of an omphalocele. STUDY DESIGN: Retrospectively, 24 fetuses diagnosed with an isolated omphalocele in the 2nd trimester of pregnancy were selected (period 2003-2013). An image of the axial plane of the abdomen at the level of the defect was retrieved. The ratio of omphalocele circumference to abdominal circumference (OC/AC), and the ratio of defect diameter to abdominal diameter (DD/DA) were calculated. Prognostic outcome was primary closure. Sensitivity and specificity and the corresponding area under the ROC curve of these ratios were calculated as measurements of prognostic accuracy. RESULTS: Primary closure was achieved in 15/24 cases. For the OC/AC-ratio a cut-off value of 0.82 successfully predicted outcome in 23/24 cases with an area under the ROC curve of 0.99. A cut-off value of 0.61 for the DD/DA-ratio successfully predicted type of closure in 20/24 cases with an area under the ROC curve of 0.88. In all cases without eviscerated liver tissue, the defect was primarily closed. CONCLUSION: In prenatal isolated omphalocele cases, the OC/AC-ratio is better at predicting postnatal surgical closure than the DD/DA-ratio and can be used as a prognostic tool for expected type of closure in the 2nd trimester of pregnancy.

Homocysteine exposure affects early hemodynamic parameters of embryonic chicken heart function.

Maternal hyperhomocysteinemia has been associated with an increased risk of newborns with a congenital heart defect. This has been substantiated in the chicken embryo, as congenital heart defects have been induced after homocysteine treatment. Comparable heart defects are observed in venous clipping studies, a model of altered embryonic blood flow. Because of this overlap in heart defects, our aim was to test the hypothesis that homocysteine would cause alterations in embryonic heart function that precede the structural malformations previously described. Therefore, Doppler flow velocity waveforms were recorded in both primitive ventricles and the outflow tract of the embryonic heart of homocysteine treated and control chicken embryos at embryonic day 3.5. Homocysteine treatment consisted of 50 µL 0.05 M L-homocysteine thiolactone at 24, 48, and 72 hr. Homocysteine-treated embryos displayed significantly lower mean heart rates of 134 (SD 22) bpm, compared to 150 (14) bpm in control embryos. Homocysteine treatment caused an inhibiting effect on hemodynamic parameters, and altered heart function was presented by a shift in the proportions of the different wave times in percentage of total cycle time. Homocysteine induces changes in hemodynamic parameters of early embryonic chicken heart function. These changes may precede morphological changes and contribute to the development of CHD defects through alterations in shear stress and shear stress related genes, as seen before in venous clipping studies.

The effects of homocysteine and folic acid on angiogenesis and VEGF expression during chicken vascular development.

Homocysteine (Hcy) has been implicated in the development of cardiovascular developmental defects. Additionally, in experimental studies, vasculotoxic properties of Hcy have been described. Although Hcy has been identified as a vascular pathogen, little is known about the direct effects Hcy exerts during early embryonic vascular development. Angiogenesis is a critical process involved in embryo survival and development. There are limited studies on the effects of Hcy on early embryonic vasculogenesis and angiogenesis. Folic acid (FA) is a B vitamin essential in embryo development, and FA supplementation may lead to reduced Hcy levels. Therefore, the purpose of our study was to explore the effects of Hcy and FA on early embryonic vascular development. Embryonic day (E) 3.5 chicken embryos were treated with a sham, Hcy or FA solution. We developed a computational program for systematic analysis of microscopic images obtained from the extra embryonic vascular beds. These results were combined with real-time PCR data on the expression of VEGF-A and its receptor in these vascular beds. Our data show that Hcy exposure inhibits early vascular development, displayed by a significant reduction of vessel area and altered composition of the vascular beds. Vascular beds of Hcy embryos for the greater part consisted of vessels of the smallest diameters, compared to middle size vessels in control and FA embryos. Hcy also reduced expression of VEGF-A and VEGFR-2. No significant effects of FA were found. We conclude that Hcy exposure causes impaired early extra embryonic vascular development, shown by altered composition of the vascular beds as well as reduced expression of VEGF-A and VEGFR-2. These effects of Hcy, and the consecutive cascade of events, may be involved in the development of cardiovascular developmental defects.

Multiple reaction monitoring assay for pre-eclampsia related calcyclin peptides in formalin fixed paraffin embedded placenta.

Although the cause of pre-eclampsia during pregnancy has not been elucidated yet, it is evident that placental and maternal endothelial dysfunction is involved. We previously demonstrated that in early onset pre-eclampsia placental calcyclin (S100A6) expression is significantly higher compared to controls ( De Groot , C. J. ; Clin. Proteomics 2007 , 1 , 325 ). In the current study, the results were confirmed and relatively quantified by using multiple reaction monitoring (MRM) on two peptide fragments of calcyclin. Cells were obtained from control (n = 5) and pre-eclamptic placental (n = 5) tissue collected by laser capture microdissection from formalin-fixed paraffin-embedded (FFPE) material treated with a solution to reverse formalin fixation. Two calcyclin peptides with an extra glycine inserted in the middle of the amino acid sequence were synthesized and used as an internal reference. Data presented show that MRM on laser microdissected material from FFPE tissue material is possible. The developed MRM assay to study quantitative levels of proteins in FFPE laser microdissected cells using nonisotopic-labeled chemical analogs of mass tagged internal references showed that in pre-eclamptic patients elevated levels of calcyclin is observed in placental trophoblast cells compared to normal trophoblast cells. By immunohistochemistry, we were able to confirm this observation in a qualitative manner.

The endothelin-1 pathway and the development of cardiovascular defects in the haemodynamically challenged chicken embryo.

BACKGROUND/AIMS: Ligating the right lateral vitelline vein of chicken embryos (venous clip) results in cardiovascular malformations. These abnormalities are similar to malformations observed in knockout mice studies of components of the endothelin-1 (ET-1)/endothelin-converting enzyme-1/endothelin-A receptor pathway. In previous studies we demonstrated that cardiac ET-1 expression is decreased 3 h after clipping, and ventricular diastolic filling is disturbed after 2 days. Therefore, we hypothesise that ET-1-related processes are involved in the development of functional and morphological cardiovascular defects after venous clip. METHODS: In this study, ET-1 and endothelin receptor antagonists (BQ-123, BQ-788 and PD145065) were infused into the HH18 embryonic circulation. Immediate haemodynamic effects on the embryonic heart and extra-embryonic vitelline veins were examined by Doppler and micro-particle image velocimetry. Ventricular diastolic filling characteristics were studied at HH24, followed by cardiovascular morphologic investigation (HH35). RESULTS: ET-1 and its receptor antagonists induced haemodynamic effects at HH18. At HH24, a reduced diastolic ventricular passive filling component was demonstrated, which was compensated by an increased active filling component. Thinner ventricular myocardium was shown in 42% of experimental embryos. CONCLUSION: We conclude that cardiovascular malformations after venous clipping arise from a combination of haemodynamic changes and altered gene expression patterns and levels, including those of the endothelin pathway.

Complex congenital malformations and the impact of the plasminogen activator system and beta-hCG in amniotic fluid.

OBJECTIVE: The plasminogen activator system and beta-hCG may affect neural crest cells and angiogenesis, and thereby embryogenesis. Therefore, we investigated these parameters in amniotic fluids of pregnancies with a complex congenital malformation. STUDY DESIGN: In a case-control study amniotic fluid samples were collected from 62 pregnancies with a complex congenital malformation and from 110 healthy control pregnancies at an obstetric department of a large university hospital in the Netherlands. We determined concentrations of tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA), plasminogen activator inhibitors (PAI-1, PAI-2), tPA approximately PAI-1 and uPA approximately PAI-1 complexes, and beta-hCG with enzyme-linked immunosorbent assays. Mann-Whitney U-tests and analysis of covariance, adjusting for gestational and maternal age, were performed for data comparisons. RESULTS: Compared with controls, cases demonstrated significantly lower adjusted geometric mean levels of uPA (24%), tPA (> or =19%) and tPA approximately PAI-1 (35%). Cases showed significantly higher adjusted mean levels of beta-hCG (> or =48%) and PAI-2 (10 ng/mL) than controls. Mean PAI-1 and uPA approximately PAI-1 levels were comparable between both groups. CONCLUSIONS: Disturbances in the plasminogen activator system and beta-hCG levels are suggested to be involved in the pathogenesis of complex congenital malformations by affecting neural crest cell migration and angiogenesis.

Large variations in absolute wall shear stress levels within one species and between species.

Wall shear stress (WSS), the frictional force between blood and endothelium, is an important determinant of vascular function. It is generally assumed that WSS remains constant at a reference value of 15 dyn/cm(2). In a study of small rodents, we realized that this assumption could not be valid. This review presents an overview of recent studies in large and small animals where shear stress was measured, derived from velocity measurements or otherwise, in large vessels. The data show that large variations exist within a single species (human: variation of 2-16 N/m(2)). Moreover, when we compared different species at the same location within the arterial tree, an inverse relationship between animal size and wall shear stress was noted. When we related WSS to diameter, a unique relationship was derived for all species studied. This relationship could not be described by the well-known r(3) law of Murray, but by the r(2) law introduced by Zamir et al. in 1972. In summary, by comparing data from the literature, we have shown that: (i) the assumption of a physiological WSS level of approximately 15 dyn/cm(2) for all straight vessels in the arterial tree is incorrect; (ii) WSS is not constant throughout the vascular tree; (iii) WSS varies between species; (iv) WSS is inversely related to the vessel diameter. These data support an "r(2) law" rather than Murray's r(3) law for the larger vessels in the arterial tree.

Dietary intake of B-vitamins in mothers born a child with a congenital heart defect.

BACKGROUND: Periconceptional use of multivitamins reduces the risk of a child with a congenital heart defect (CHD). Data on the impact of maternal diet, however, are lacking. AIM OF THE STUDY: We investigated the association between the maternal dietary intake of B-vitamins and having a child with a CHD. METHODS: A case-control study was performed in 192 mothers of a child with a CHD and 216 mothers of a healthy child. Mothers filled out food frequency questionnaires covering the current dietary intake, and general questionnaires at 17 months after the index-pregnancy. Maternal blood samples were taken to determine B-vitamin and plasma total homocysteine (tHcy) concentrations as nutritional biomarkers. Pregnant and lactating mothers and those with another diet compared with the preconceptional period were excluded for analysis. Case-mothers and controls were compared using the Mann-Whitney U test and logistic regression. RESULTS: The dietary intake of macronutrients and B-vitamins was comparable between both groups, but all mothers had a substantially lower median folate intake (cases 161 microg, controls 175 microg) than the Dutch recommended dietary allowance of 300 microg. Within the case-group, the intake of proteins and vitamin B(6) and the concentrations of serum vitamin B(12) and folate were significantly lower in hyperhomocysteinemics (tHcy > or = 14.5 micromol/l) than in normohomocysteinemics. The maternal educational level was positively associated with B-vitamin intake, except for vitamin B(12) in controls. Low educated case-mothers showed a significantly lower median vitamin B(12) intake than controls (2.8 microg and 3.8 microg, P = 0.01). The CHD risk doubled if vitamin B(12) intake in these mothers reduced by 50% (OR 2.0; 95% CI: 1.1-3.5). CONCLUSIONS: A diet low in vitamin B(12) is associated with an increased risk of a child with a CHD, especially in low educated women. A disbalance in the maternal intake of proteins and low folate intake may play a role as well, but needs further investigation. As hyperhomocysteinemia is a strong risk factor for adult cardiovascular disease, these data may imply that the hyperhomocysteinemic mothers and their children should be targeted for nutritional interventions.

Incidence of parvovirus B19 infection among an unselected population of pregnant women in the Netherlands: A prospective study.

OBJECTIVE: To evaluate seroprevalence of anti-parvovirus B19 IgG immunoglobulins and the rate of seroconversion in seronegative pregnant women. DESIGN: Prospective assessment of anti-parvovirus B19 IgG immunoglobulins in an unselected population of pregnant women booked for antenatal care from 1998 to 2000. SETTING: Maternity departments of an academic hospital and four affiliated teaching hospitals in the Netherlands. SUBJECTS: Two thousand five hundred and sixty seven pregnant women. MAIN OUTCOME MEASURES: Seroprevalence of anti-parvovirus B19 IgG immunoglobulin in the first trimester of pregnancy and subsequent seroconversion in those women who were tested negative for parvovirus B19 antibodies in the first trimester of pregnancy. RESULTS: The estimated seroprevalence of anti-parvovirus B19 IgG immunoglobulins among the study population is 70% (95% CI: 68-71) in the first trimester of pregnancy. Seven hundred and seventy nine women tested negative for parvovirus B19 antibodies in the first trimester of pregnancy. Paired testing in these women confirmed 18 seroconversions. Based on these findings the estimated incidence of maternal parvovirus B19 infection in this population among seronegative Dutch women is 2.4% (95% CI: 1.4-3.7). CONCLUSION: Maternal infection with parvovirus B19 is relatively common. However, it is argued that in the Netherlands routine assessment of parvovirus antibodies in pregnant women is not warranted as there is a low risk of adverse fetal outcome and measures to prevent the parvovirus B19 infection and its consequences are very limited.

In vivo micro particle image velocimetry measurements of blood-plasma in the embryonic avian heart.

The measurement of blood-plasma velocity distributions with spatial and temporal resolution in vivo is inevitable for the determination of shear stress distributions in complex geometries at unsteady flow conditions like in the beating heart. A non-intrusive, whole-field velocity measurement technique is required that is capable of measuring instantaneous flow fields at sub-millimeter scales in highly unsteady flows. Micro particle image velocimetry (muPIV) meets these demands, but requires special consideration and methodologies in order to be utilized for in vivo studies in medical and biological research. We adapt muPIV to measure the blood-plasma velocity in the beating heart of a chicken embryo. In the current work, bio-inert, fluorescent liposomes with a nominal diameter of 400 nm are added to the flow as a tracer. Because of their small dimension and neutral buoyancy the liposomes closely follow the movement of the blood-plasma and allow the determination of the velocity gradient close to the wall. The measurements quantitatively resolve the velocity distribution in the developing ventricle and atrium of the embryo at nine different stages within the cardiac cycle. Up to 400 velocity vectors per measurement give detailed insight into the fluid dynamics of the primitive beating heart. A rapid peristaltic contraction accelerates the flow to peak velocities of 26 mm/s, with the velocity distribution showing a distinct asymmetrical profile in the highly curved section of the outflow tract. In relation to earlier published gene-expression experiments, the results underline the significance of fluid forces for embryonic cardiogenesis. In general, the measurements demonstrate that muPIV has the potential to develop into a general tool for instationary flow conditions in complex flow geometries encountered in cardiovascular research.

Systolic and diastolic ventricular function assessed by pressure-volume loops in the stage 21 venous clipped chick embryo.

Cardiac pressure-volume relations enable quantification of intrinsic ventricular diastolic and systolic properties independent of loading conditions. The use of pressure-volume loop analysis in early stages of development could contribute to a better understanding of the relationship between hemodynamics and cardiac morphogenesis. The venous clip model is an intervention model for the chick embryo in which permanent obstruction of the right lateral vitelline vein temporarily reduces the mechanical load on the embryonic myocardium and induces a spectrum of outflow tract anomalies. We used pressure-volume loop analysis of the embryonic chick heart at stage 21 (3.5 d of incubation) to investigate whether the development of ventricular function is affected by venous clipping at stage 17, compared with normal control embryos. Steady state hemodynamic parameters demonstrated no significant differences between the venous clipped and control embryos. However, analysis of pressure-volume relations showed a significantly lower end-systolic elastance in the clipped embryos (slope of the end-systolic pressure-volume relation: 5.68 +/- 0.85 versus 11.76 +/- 2.70 mm Hg/microL, p < 0.05), indicating reduced contractility. Diastolic stiffness tended to be increased in the clipped embryos (slope of end-diastolic pressure-volume relation: 2.74 +/- 0.56 versus 1.67 +/- 0.21, p = 0.103), but the difference did not reach statistical significance. The results of the pressure-volume loop analysis show that 1 d after venous obstruction, development of ventricular function is affected, with reduced contractility. Pressure-volume analysis may be applied in the chick embryo and is a sensitive technique to detect subtle alterations in ventricular function.

Ventricular diastolic filling characteristics in stage-24 chick embryos after extra-embryonic venous obstruction.

Alteration of extra-embryonic venous blood flow in stage-17 chick embryos results in well-defined cardiovascular malformations. We hypothesize that the decreased dorsal aortic blood volume flow observed after venous obstruction results in altered ventricular diastolic function in stage-24 chick embryos. A microclip was placed at the right lateral vitelline vein in a stage-17 (52-64 h of incubation) chick embryo. At stage 24 (4.5 days of incubation), we measured simultaneously dorsal aortic and atrioventricular blood flow velocities with a 20-MHz pulsed-Doppler velocity meter. The fraction of passive and active filling was integrated and multiplied by dorsal aortic blood flow to obtain the relative passive and active ventricular filling volumes. Data were summarized as means +/- S.E.M. and analyzed by t-test. At similar cycle lengths ranging from 557 ms to 635 ms (P>0.60), dorsal aortic blood flow and stroke volume measured in the dorsal aorta were similar in stage-24 clipped and normal embryos. Passive filling volume (0.07+/-0.01 mm(3)) was decreased, and active filling volume (0.40+/-0.02 mm(3)) was increased in the clipped embryo when compared with the normal embryo (0.15+/-0.01 mm(3), 0.30+/-0.01 mm(3), respectively) (P<0.003). In the clipped embryos, the passive/active ratio was decreased compared with that in normal embryos (P<0.001). Ventricular filling components changed after partially obstructing the extra-embryonic venous circulation. These results suggest that material properties of the embryonic ventricle are modified after temporarily reduced hemodynamic load.

Assessment of fetal liver volume and umbilical venous volume flow in pregnancies complicated by insulin-dependent diabetes mellitus.

OBJECTIVES: To determine fetal liver volume and its relation with umbilical venous volume flow and maternal glycosylated haemoglobin (HbA1c) in pregnancies complicated by diabetes mellitus type I. DESIGN: A cross sectional matched control study. Obstetric out patient clinic, Erasmus MC-University Medical Centre, Rotterdam. POPULATION: Data from fetuses of diabetic women (n = 32; 18-36 weeks) were compared with data from normal controls (n = 32) matched for gestational age. METHODS: Umbilical venous cross sectional area (mm(2)) and time-averaged velocity (mm/s Doppler) were determined for calculation of volume flow (mL/min) and flow per kilogram fetal weight (mL/min/kg). Umbilical artery pulsatility index was determined. Fetal liver volume measurements were obtained using a Voluson 530-D. MAIN OUTCOME MEASURES: Fetal liver volume, umbilical venous volume flow and downstream impedance. RESULTS: A statistically significant difference between fetuses of diabetic women and normal controls was found for liver volume (mean [SD]: 45.9 [34.0] vs 38.3 [28.7] mL), abdominal circumference (22.2 [6.6] vs 21.3 [5.6] cm), estimated fetal weight (1162 [898] vs 1049 [765] g) and fetoplacental weight ratio (0.22 vs 0.19) and liver volume/estimated fetal weight ratio (4.13% [0.007] vs 3.62% [0.009]). Umbilical venous volume flow (mL/min) and umbilical artery pulsatility index were not essentially different between the two study groups, but umbilical venous volume flow per kilogram fetal weight was lower (P < 0.05) in the diabetes group (94.3 [26.1] mL/min kg) compared with normal controls (109.5 [28.0] mL/min/kg). A positive correlation existed between fetal liver volume and maternal HbA1c (P = 0.002). CONCLUSIONS: Measurement of fetal liver volume by three-dimensional ultrasound may play a role in identifying fetal growth acceleration in diabetic pregnancies. Fetal liver volume increase is positively related to maternal HbA1c levels reflecting degree of maternal glycemic control. Fetal liver volume normalised for estimated fetal weight is significantly higher in the fetuses of diabetic women. In the present study, umbilical venous volume flow and fetoplacental downstream impedance are not different between diabetic and normal pregnancies.

Acutely altered hemodynamics following venous obstruction in the early chick embryo.

In the venous clip model specific cardiac malformations are induced in the chick embryo by obstructing the right lateral vitelline vein with a microclip. Clipping alters venous return and intracardiac laminar blood flow patterns, with secondary effects on the mechanical load of the embryonic myocardium. We investigated the instantaneous effects of clipping the right lateral vitelline vein on hemodynamics in the stage-17 chick embryo. 32 chick embryos HH 17 were subdivided into venous clipped (N=16) and matched control embryos (N=16). Dorsal aortic blood flow velocity was measured with a 20 MHz pulsed Doppler meter. A time series of eight successive measurements per embryo was made starting just before clipping and ending 5h after clipping. Heart rate, peak systolic velocity, time-averaged velocity, peak blood flow, mean blood flow, peak acceleration and stroke volume were determined. All hemodynamic parameters decreased acutely after venous clipping and only three out of seven parameters (heart rate, time-averaged velocity and mean blood flow) showed a recovery to baseline values during the 5h study period. We conclude that the experimental alteration of venous return has major acute effects on hemodynamics in the chick embryo. These effects may be responsible for the observed cardiac malformations after clipping.