Doppler flow velocity waveforms in the embryonic chicken heart at developmental stages corresponding to 5-8 weeks of human gestation.

OBJECTIVES: To obtain Doppler velocity waveforms from the early embryonic chicken heart by means of ultrasound biomicroscopy and to compare these waveforms at different stages of embryonic development. METHODS: We collected cardiac waveforms using high-frequency Doppler ultrasound with a 55-MHz transducer at Hamburger-Hamilton (HH) stages 18, 21 and 23, which are comparable to humans at 5 to 8 weeks of gestation. Waveforms were obtained at the inflow tract, the primitive left ventricle, the primitive right ventricle and at the outflow tract in 10 different embryos per stage. M-mode recordings were collected to study opening and closure of the cushions. By exploring the temporal relationship between the waveforms, using a secondary Doppler device, cardiac cycle events were outlined. RESULTS: Our results demonstrate that stage- and location-dependent intracardiac blood flow velocity waveforms can be obtained in the chicken embryo. The blood flow profiles assessed at the four locations in the embryonic heart demonstrated an increase in peak velocity with advancing developmental stage. In the primitive ventricle the 'passive' (P) filling peak decreased whereas the 'active' (A) filling peak increased, resulting in a decrease in P to A ratio with advancing developmental stage. M-mode recordings demonstrated that the fractional closure time of the atrioventricular cushions increased from 20% at stage HH 18 to 60% at stage HH 23. CONCLUSION: High-frequency ultrasound biomicroscopy can be used to define flow velocity waveforms in the embryonic chicken heart. This may contribute to an understanding of Doppler signals derived from valveless embryonic human hearts at 5 to 8 weeks of gestation, prior to septation.

Systolic and diastolic ventricular function in the normal and extra-embryonic venous clipped chicken embryo of stage 24: a pressure-volume loop assessment.

OBJECTIVES: Fluid mechanical forces affect cardiac development. In the chicken embryo, permanent obstruction of the right lateral vitelline vein by clipping reduces the mechanical load on the embryonic myocardium, which has been shown to induce a spectrum of outflow tract anomalies. Insight into the effects of this intervention on the mechanical function of the developing myocardium could contribute to a better understanding of the relationship between hemodynamics and cardiac morphogenesis. We aimed to explore the effects of clipping on intrinsic systolic and diastolic ventricular function at stage 24 in the chicken embryo METHODS: Cardiac pressure-volume relationships enable load-independent quantification of intrinsic ventricular systolic and diastolic properties. We determined ventricular function by pressure-volume loop analysis of in-ovo stage-24 chicken embryos (n = 15) 2 days after venous obstruction at 2.5 days of incubation (stage 17, venous clipped embryos). Control embryos (n = 15) were used for comparison. RESULTS: End-systolic volume was significantly higher in clipped embryos (0.36 +/- 0.02 microL vs. 0.29 +/- 0.02 microL, P = 0.002). End-systolic and end-diastolic pressure were also increased compared with control animals (2.93 +/- 0.07 mmHg vs. 2.70 +/- 0.08 mmHg, P = 0.036 and 1.15 +/- 0.06 mmHg vs. 0.82 +/- 0.05 mmHg, P < 0.001, respectively). No significant differences were demonstrated for other baseline hemodynamic parameters. Analysis of pressure-volume relationships showed a significantly lower end-systolic elastance in the clipped embryos (slope of end-systolic pressure-volume relationship: 2.91 +/- 0.24 mmHg/microL vs. 7.53 +/- 0.66 mmHg/microL, P < 0.005) indicating reduced contractility. Diastolic stiffness was significantly increased in the clipped embryos (slope of end-diastolic pressure-volume relationship: 1.54 +/- 0.21 vs. 0.60 +/- 0.08, P < 0.005), indicating reduced compliance. CONCLUSION: Venous obstruction apparently interferes with normal myocardial development, resulting in impaired intrinsic systolic and diastolic ventricular function. These changes in ventricular function may precede morphological derangements observed in later developmental stages.

Validation of the assessment of folate and vitamin B12 intake in women of reproductive age: the method of triads.

OBJECTIVE: To validate the folate and vitamin B12 intakes estimated by a food-frequency questionnaire (FFQ) designed to be used in a case-control study on the association between maternal dietary intake and the risk of having a child with a congenital heart defect. DESIGN AND SUBJECTS: The FFQ was filled out by 53 women of reproductive age. Immediately thereafter, blood samples were taken to determine serum folate, red blood cell (RBC) folate and serum vitamin B12 concentrations. Subsequently, three dietary 24-h recalls (24HR) were completed during a period of three successive weeks and used as a reference method. The recalls comprised two weekdays and one weekend day. Using the method of triads, validity coefficients were calculated by comparing nutrient intakes derived from the FFQ and 24HR with the corresponding nutritional biomarkers in blood. The validity coefficient is the correlation between the dietary intake reported by the FFQ and the unknown 'true' dietary intake. RESULTS: The comparison of B-vitamin intakes reported by the FFQ and the mean of the 24HR revealed deattenuated correlation coefficients of 0.98 for folate and 0.66 for vitamin B12. The correlation coefficients between the B-vitamin intakes estimated by the FFQ and concentrations of serum folate, RBC folate and serum vitamin B12 were 0.20, 0.28 and 0.21, respectively. The validity coefficients for serum folate, RBC folate and serum vitamin B12 were 0.94, 0.75 and 1.00, respectively. The estimated folate and vitamin B12 intakes were comparable with the results of the most recent Dutch food consumption survey. CONCLUSIONS: The adapted FFQ is a reliable tool to estimate the dietary intake of energy, macronutrients, folate and vitamin B12 in women of reproductive age. Therefore, this FFQ is suitable for the investigation of nutrient-disease associations in future. SPONSORSHIP: Funding was provided by the Netherlands Heart Foundation (Grant 2002.B027).

Maternal hyperhomocysteinaemia is a risk factor for congenital heart disease.

OBJECTIVE: To investigate the inter-relation between mother and infant homocysteine, folate and vitamin B12 status and the risk of a child with congenital heart disease (CHD). DESIGN: Case-control study. SETTING: Erasmus MC, University Medical Centre, Rotterdam, the Netherlands. POPULATION: Participants were 149 case-mothers and their children with CHD (n = 151) and 183 control-mothers with their children (n = 175). METHODS: Approximately 17 months after the index-pregnancy maternal fasting, children's random venous blood samples were drawn to measure plasma total homocysteine, serum and red blood cell (RBC) folate, and serum vitamin B12 concentrations. Data were compared between cases and controls using the Mann-Whitney U test. The biochemical parameters were dichotomised according to the cutoff value of the 10th percentile of vitamin concentrations and the 90th percentile of homocysteine concentrations based on control data. Risk estimates for the association between CHD and the biochemical parameters were estimated in a logistic regression model. MAIN OUTCOME MEASURES: Medians (minimum-maximum) and odds ratios (OR) (95% confidence intervals [CI]). RESULTS: The OR (95% CI) of having a child with CHD was 2.9 (1.4-6.0) for maternal hyperhomocysteinaemia (>14.3 micromol/l). This finding is substantiated by a significant concentration-dependent risk (Ptrend = 0.004). Hyperhomocysteinaemic case-mothers showed significantly lower serum folate and vitamin B12 concentrations than normohomocysteinaemic case-mothers. Serum and RBC folate concentrations were significantly higher in case-children than that in control-children. CONCLUSIONS: Maternal hyperhomocysteinaemia is associated with an increased risk of CHD, partially due to low folate and vitamin B12 status. The folate status of children warrants further investigation.

Mild renal pyelectasis in the second trimester: determination of cut-off levels for postnatal referral.

OBJECTIVE: To establish guidelines for postnatal referral of fetuses presenting with mild pyelectasis in the second trimester of pregnancy. METHODS: In a retrospective study, 87 fetuses with a renal pelvis anteroposterior (RPAP) diameter of > or = 4 mm and < or = 10 mm before 28 weeks of gestation were included. All patients had a third-trimester scan and fetuses with an RPAP diameter of > or = 10 mm at that stage were referred for postnatal assessment. The family practitioner of all infants with an RPAP of < 10 mm in the third trimester was contacted for follow-up information. The RPAP diameter most predictive of renal pathology was determined with receiver-operating characteristics (ROC) curve analysis for both the first and second scans. RESULTS: In 36 of 87 infants, 49 abnormal kidneys were diagnosed. Seven infants required surgery on eight renal tracts. The ROC curves of the first scan, second scan and differences between scans resulted in an area under the curve of 0.60, 0.87 and 0.85, respectively. The sensitivities and specificities for a cut-off level of 8, 9 and 10 mm at the second scan were 80%, 71% and 61% and 79%, 90% and 93%, respectively. At a cut-off level of 10 mm, only cases of insignificant minimal dilatation and a case of vesicoureteric reflux (VUR) requiring surgery were not detected. CONCLUSION: After establishing a diagnosis of mild pyelectasis before 28 weeks, a second scan is mandatory to determine which infants need postnatal evaluation. A cut-off level of 8 mm has a low specificity but includes most cases of pathology. A cut-off level of 10 mm detects most significant pathology; however, VUR may not be detected.

Fetal heart rate and umbilical artery flow velocity variability in intrauterine growth restriction: a matched controlled study.

OBJECTIVES: To study heart rate and umbilical artery blood flow velocity variability in growth-restricted fetuses and investigate the influence of the autonomic nervous system on these parameters. METHODS: Doppler velocity waveforms were collected from long-lasting umbilical artery recordings in 15 fetuses with growth restriction and 15 normal age-matched controls at 23-35 weeks of gestation. Absolute heart rate and umbilical artery blood flow velocity as well as the coefficient of variation were determined. Using power spectral analysis the low- and high-frequency bands of heart rate variability and blood flow velocity variability were calculated. The low-to-high (LH) ratio of heart rate variability and blood flow velocity variability were examined as a measure of sympathovagal balance. RESULTS: In growth-restricted fetuses umbilical artery velocities were significantly reduced. Heart rate variability was significantly reduced in the presence of growth restriction, but no significant difference was demonstrated for blood flow velocity variability. The LH ratio for heart rate variability was significantly decreased in growth restriction, but no difference in LH ratio was demonstrated for blood flow velocity variability. CONCLUSION: Flow velocity variability in growth restriction seems not to be predominantly influenced by the autonomic nervous system, whereas the decreased heart rate variability seems to be influenced by altered sympathetic-parasympathetic balance.

Fetal heart rate and blood flow velocity variability in the presence of increased nuchal translucency: a preliminary study.

OBJECTIVE: To study the power spectrum distribution of heart rate and umbilical artery flow velocity variability in fetuses with increased nuchal translucency thickness (NT). METHODS: Doppler velocity waveforms were collected from long-lasting (>20 s) umbilical artery recordings in 18 fetuses with increased NT (>3 mm) and 18 normal controls matched for gestational age at 11-14 (median, 12) weeks. The NT group included 11 abnormal karyotypes: trisomy 18 (n = 3), 45,X (n = 4), trisomy 21 (n = 3) and a balanced translocation. Absolute heart rate as well as the coefficient of variation for both beat-to-beat heart rate variability and umbilical artery blood flow velocity variability were determined. The ratios of the integrated low-frequency components (0.05-0.2 Hz) and the integrated high-frequency ones (0.25-1.6 Hz; LH ratio) from normalized power spectrum distributions were established to reflect sympathovagal balance. RESULTS: The mean heart rate was not significantly different between the two groups. However, mean heart rate variability and time-averaged flow velocity variability were significantly increased in the NT group, while there was no significant difference in the LH ratios between the two groups. The mean umbilical artery pulsatility index was significantly higher in the NT group. CONCLUSION: The autonomic nervous system does not seem to play a role in the altered cardiovascular homeostasis in the presence of increased fetal NT.

Umbilical venous volume flow and fetal behavioral states in the normally developing fetus.

OBJECTIVE: To determine the relationship between umbilical venous (UV) volume flow and fetal behavioral states 1F (quiet sleep) and 2F (active sleep) in normal pregnancies at 36-40 weeks of gestation. METHODS: Fetal behavioral states were established in 17 normal pregnancies by means of combined assessment of fetal heart rate patterns (FHRP), and fetal eye and body movements. UV vessel area (mm(2)) as obtained by tracing the inner vessel area using Labview and Imaq Vision software and UV time-averaged flow velocity (mm/s Doppler) were multiplied to calculate UV volume flow (mL/min) including flow/kg fetus. The pulsatility index (PI) in the umbilical artery was also determined. In each woman, all parameters were measured between three and five times in each behavioral state. Data are reported as mean +/- 1 SD and analyzed by paired t-test. RESULTS: No statistically significant behavioral-state-related changes were observed for UV time-averaged velocity and UV volume flow, resulting in UV volume flow/kg fetus of 69.1 +/- 14.9 mL/min*kg at 1F and 71.6 +/- 12.1 mL/min*kg at 2F (not significant). A statistically significant increase (P = 0.02) was established for UV cross-sectional area (46.4 +/- 8.6 mm(2) vs. 49.0 +/- 10.1 mm(2)) and for fetal heart rate (FHR) from 134.2 +/- 10.3 bpm in 1F to 144.2 +/- 7 bpm in 2F. Umbilical artery PI was not significantly different between the two behavioral states. CONCLUSIONS: On the basis of high venous vessel wall compliance, the significant increase in UV cross-sectional area during fetal behavioral state 2F may be determined by a rise in mean venous pressure. The significant rise in FHR may reflect increased fetal cardiac output during state 2F while the resistance at the hepato-ductal pathway remains relatively constant with the purpose of meeting raised energy demands during the active sleep state. This is further supported by the observed trend towards an increase in UV volume flow.

Fetal brain/liver volume ratio and umbilical volume flow parameters relative to normal and abnormal human development.

OBJECTIVES: To estimate fetal brain volume from head circumference and published postmortem data; to determine normal values for the fetal brain/liver volume ratio relative to gestational age; to establish the relationship between the brain/liver volume ratio and fetal circulatory parameters during normal and restricted (SGA) fetal growth. PATIENTS AND METHOD: This was a cross-sectional study involving a total of 47 uncomplicated pregnancies appropriate-for-gestational age (AGA) and 23 pregnancies resulting in the delivery of a SGA fetus. At enrollment gestational age ranged between 20 and 36 weeks in both groups. Umbilical venous cross-sectional area and time-averaged velocity for calculation of volume flow as well as velocity waveforms from the umbilical artery, middle cerebral artery and ductus venosus, were recorded. Fetal liver volume measurements were obtained using three-dimensional ultrasound. Fetal brain volume was estimated from fetal head volume following comparison with published postmortem data on fetal brain weight. RESULTS: A significant correlation was observed between prenatally estimated fetal head volume and postmortem fetal brain volume. Fetal brain volume was approximately half that of fetal head volume. The normal fetal brain/liver volume ratio demonstrated a significant reduction with gestational age (R = -0.54; P < 0.001). The normal mean +/- standard deviation (SD) fetal brain/liver volume ratio (3.4 +/- 0.7) was significantly different (P < 0.001) from the mean fetal brain/liver volume ratio in the SGA group (n = 23) (5.9 +/- 1.9). A significant difference existed for mean umbilical venous volume flow between AGA (104.7 +/- 26.9 mL/min/kg) and SGA (59.6 +/- 22.7 mL/min/kg) fetuses. In the SGA fetus, there was a significant inverse relationship (P < 0.001) between fetal weight-related umbilical venous volume flow and fetal brain/liver volume ratio. In a subset of 16 SGA and 16 AGA fetuses matched for gestational age, a significant difference existed for umbilical artery pulsatility index (2.30 +/- 1.52 vs. 0.99 +/- 0.19), fetal middle cerebral artery pulsatility index (1.3 +/- 0.4 vs. 2.1 +/- 0.3) and late diastolic flow velocity in the fetal ductus venosus (6.9 +/- 14.2 cm/s vs. 23.9 +/- 8.8 cm/s), but not for peak systolic, early diastolic and time-averaged velocity in the ductus venosus. CONCLUSION: Sonographic estimates of fetal brain volume can be obtained. The fetal brain/liver volume ratio is a predictor of fetal outcome in the growth-restricted fetus. An inverse relationship exists in SGA fetuses between brain/liver volume ratio and fetal weight-related umbilical venous blood flow.

Umbilical venous volume flow in the normally developing and growth-restricted human fetus.

OBJECTIVE: To determine the reproducibility of measurement of umbilical venous volume flow components and to calculate umbilical venous volume flow in normal and growth-restricted (small-for-gestational age) fetuses in a cross-sectional study. METHOD: Using Labview and Imaq-vision software, the cross-sectional inner area of the umbilical vein was traced. Vessel area (mm2) and Doppler-derived time-averaged flow velocity (mm/s) were multiplied to calculate volume flow (mL/min) including flow per kg fetal weight. The coefficient of variation for vessel area and flow velocity scans and tracings were determined (n = 13; 26-35 weeks). Normal charts for components and volume flow were constructed (n = 100; 20-36 weeks) and related to data from growth restricted fetuses (birth weight < 5th centile) (n = 33; 22-36 weeks). In growth-restricted fetuses the umbilical artery pulsatility index was also obtained. RESULTS: Reproducibility: The coefficient of variation was 5.4% (vessel area) and 7.3% (time-averaged velocity) for scans and 6.6% and 10.5% for measurements, resulting in a coefficient of variation of 8.1% (scans) and 11.9% (measurements) for volume flow. A gestational age-related increase exists for vessel area, time-averaged flow velocity and umbilical venous volume flow from 33.2 (SD, 15.2) mL/min at 20 weeks to 221.0 (SD, 32.8) mL/min at 36 weeks of gestation, but there is a reduction from 117.5 (SD, 33.6) mL/min to 78.3 (SD, 12.4) mL/min for volume flow per kg fetal weight. In small-for-gestational age fetuses, the values were below the normal range in 31 of 33 cases for volume flow and in 21 of 33 cases for volume flow per kg fetal weight. Umbilical artery pulsatility index was significantly different between the subsets with normal and those with reduced volume flow per kg fetal weight. CONCLUSIONS: Measurements of umbilical venous vessel area and time-averaged velocity resulted in acceptable reproducibility of volume flow calculations, which show a seven-fold increase at 20-36 weeks of gestation. In growth-restricted fetuses, volume flow is significantly reduced. When calculated per kg/fetus, the values were reduced in 21 (63.6%) out of 33 cases.