Ultrasound Obstet Gynecol 2006; 27: 640–646 Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/uog.2794
Three-dimensional ultrasound volume calculations of human embryos and young fetuses: a study on the volumetry of compound structures and its reproducibility H.-G. K. BLAAS*, P. TAIPALE†, H. TORP‡ and S. H. EIK-NES* *National Center for Fetal Medicine, St Olav’s Hospital, Trondheim University Hospital, †Department of Obstetrics & Gynecology, ¨ Finland and ‡Department of Physiology and Biomedical Engineering, University of Trondheim, Norway Hyvinka¨ a¨ Hospital, Hyvinka¨ a,
K E Y W O R D S: embryonic/fetal limb volume; embryonic volume/weight; fetal volume/weight; geometry visualization; three-dimensional ultrasound; volume measurements
ABSTRACT Objective To evaluate volumetry with three-dimensional (3D) ultrasonography in the assessment of the size of human embryos and fetuses. Methods Forty-four healthy embryos/fetuses with crown–rump length (CRL) ranging from 9 mm to 58 mm were studied using a 7.5-MHz annular array transvaginal 3D probe. EchoPAC 3D software was used to calculate the volumes of the head, body and limbs in the same data set by two observers working independently of each other. Regression analysis was used to assess the relationship between estimated volumes and CRL. Results The embryonic and fetal volume estimates of both observers ranged from a mean of 93 mm3 at 10 mm CRL to a mean of 11 169 mm3 at 55 mm CRL. The volume of the limbs as a proportion of the mean whole-body volume increased from 4.7% at a CRL of 15 mm to 9.3% at a CRL of 55 mm. Limits of agreement between the observers were calculated to be −0.12 ± 9.2%. Conclusion It is possible to reconstruct complex small anatomic structures and calculate the volumes of human embryos and fetuses in vivo by using dedicated 3D ultrasound equipment. The reproducibility of whole-body volume estimates seems to be high. The limbs represent a significant proportion of the size of the embryonic/fetal body. Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
INTRODUCTION Research on three-dimensional (3D) ultrasonography in obstetrics started about 25 years ago when Brinkley and
co-workers performed in vitro measurements of fetuses1,2 . However, it was not until improvements in computer technology came at the end of the 1980s that obstetric 3D ultrasonography made major steps forward. During the past decade, attention has been paid mainly to surface rendering of the fetus, although estimation of volume is also of obvious interest. So far, whole-body volume estimates in the second and third trimesters have not been done, partly because of the limitations in acquiring 3D volumes of large objects, and partly because commercially available 3D ultrasound machines have not previously had the software necessary for simultaneously performing segmentations of the fetal head, body, and limbs3 , thus facilitating calculation of the entire fetal volume in the same data set. In the first trimester whole-body 3D ultrasound volume acquisition of small embryos and young fetuses including the limbs is possible. Recently, volumetry of firsttrimester specimens with analysis of the fetal trunk and head has been described4,5 . By using specially designed software, we were able to geometrically reconstruct the various parts of the body (trunk, head and limbs) of embryos and fetuses in the same data set6,7 and calculate the total volume and weight of the specimens. In vitro6 – 12 and in vivo6,13 studies have been done using a variety of different acquisition techniques that include electromagnetic tracking devices or mechanical devices to test the validity of such 3D volume estimates. In these studies, volume calculations were principally done by drawing contours around the objects of interests in various two-dimensional (2D) slices of the 3D data sets, interpolating the space between the contours and then calculating the volume using dedicated volumetry software.
Correspondence to: Dr H.-G. K. Blaas, St Olav’s Hospital, Department of Obstetrics and Gynecology, University Hospital, National Center for Fetal Medicine, Norwegian University of Science and Technology, Sør-Trøndelag, Norway (e-mail:
[email protected]) Accepted: 5 October 2005
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
ORIGINAL PAPER
Volumetry of embryos and fetuses by 3D ultrasound
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The aim of the present study was to evaluate 3D ultrasound volumetry of the bodies of embryos and fetuses including the limbs, to study interobserver reproducibility of such measurements and to establish reference data for embryonic and early fetal volumes.
PATIENTS AND METHODS We recruited 44 healthy pregnant women without any previous pregnancy complications from women who were referred early for a routine ultrasound scan. Written informed consent was obtained, and the regional committee for medical ethics approved the study. The gestational age of the pregnancies ranged from 7 to 12 weeks. For the ultrasound examinations a 7.5-MHz annular array transvaginal 3D-probe was used. This transducer had a large aperture (11.5 mm) and its symmetric focusing generated thin ultrasound tomograms. The 3D system and the method of data acquisition have been described previously6 . For the examination we used System Five, GE Vingmed Ultrasound, Horten, Norway, and Vingmed EchoPAC-3D software, version 1.1. After an initial 2D ultrasound examination by one of two observers, 3D ultrasound data sets were acquired with the fetus in a quiet phase by one of the observers and stored off-line on an external computer. Some time later, at different times and blinded from each other’s measurements, each of the two observers performed volume estimations from these 3D data sets. ‘Anyplane’ slices through the embryo or fetus were magnified on the screen of the computer, and contours were drawn using the computer mouse. The volumes were determined by drawing segmentation lines close to, but not upon, the brightest depiction of the embryonic/fetal surface of the skin (Figure 1). The embryonic/fetal body can be regarded as comprising several sections, namely head, trunk and limbs. All segmentations of the various parts of the
Figure 2 Geometric reconstruction of a fetus with a crown–rump length of 30 mm; (a) segmentation lines used in the reconstruction; (b) surface rendering obtained. Note that the body compartments (head, trunk, arms, legs) are outlined separately.
embryos/fetuses were made in parallel slices (Figure 2). At the junction of different body parts such as the body and the extremities the borders were outlined in detail to avoid overlapping or dropouts (Figures 2–4). The segmentation process took between 20 and 30 minutes. The point-spread function of ultrasound blurs the outline of an object such as the skin of an embryo, thus the manually drawn segmentation lines lie somewhat outside the true surface of the object of interest. The estimated volumes in the present study were corrected for the pointspread function, assuming a deviation in the segmentation process of 0.1 mm, in accordance with the method of Berg et al.7 . Each observer chose the ultrasound planes in which the specimens were analyzed randomly (Figures 2–4).
Statistical analysis Figure 1 Anyplane slice in a three-dimensional ultrasound volume of an embryo of 14.7 mm crown–rump length. A segmentation line is drawn at the surface of the embryo.
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
We used regression analysis to assess the relationship between estimated volume and crown–rump length
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Figure 3 Geometric reconstruction from segmentation lines of an embryo with a crown–rump length of 24 mm by Observer A (a) and Observer B; (b) it can be seen that Observer B used more segmentations than A.
(CRL) in a cross-sectional study. When analyzing the volume of a series of specimens varying from very small to relatively large, one will obtain an increasing variance of the residuals (heteroscedasticity). Square root transformation of the responses was done to remove heteroscedasticity. The normality of the residuals was confirmed by visual inspection and by Shapiro–Francia W-test. The residual plots confirmed the assumption of independency and constant variance of the residuals. Interobserver reproducibility was evaluated by the method proposed by Bland and Altman14 .
RESULTS Clinical data and outcome The mean CRL of the 44 specimens on the day of the ultrasound examination was 21.3 (range, 9–58) mm. Seven cases were examined at 7 weeks’ gestation
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
(although it was possible to examine the limbs in only four of these), 14 at 8 weeks, nine at 9 weeks, seven at 10 weeks, four at 11 weeks, and three at 12 weeks. The mean gestational age at delivery was 40 + 4 (range, 38 + 0 to 43 + 0) weeks. The mean birth weight was 3667 (range, 2770–4990) g. Postnatally, all 22 boys and 22 girls were healthy and thriving.
Volume reconstruction and interobserver variation As part of the evaluation of embryonic/fetal volumetry, the size of the limbs was compared with the total volume of the specimens (Table 1). Observer A estimated that the embryonic and fetal volumes ranged from 132 mm3 at a CRL of 9 mm to 12 861 mm3 at a CRL of 58 mm, while Observer B calculated that the corresponding volumes ranged from 130 mm3 to 11 936 mm3 , respectively (Figures 3 and 4). Limb measurements were obtained in 41 of 44 cases. In three of the smaller embryos, with
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Figure 4 Geometric reconstruction from segmentation lines of a fetus with a crown–rump length of 41 mm by Observer A (a) and Observer B; (b) it can be seen that Observer B used more segmentations than A.
CRLs of 10, 13 and 14 mm, the limbs were included in the segmentation of the bodies. The estimated volumes of the limbs by Observer A ranged from 1.8 mm3 (at a CRL of 9 mm) to 1159 mm3 (at a CRL of 58 mm), and the corresponding estimates of Observer B ranged from 1.9 mm3 to 1493 mm3 . The mean difference in the interobserver variation of the whole-body measurements between A and B was 58.4 (range, −361.2 to 924.6) mm3 , SD 251.9, SE
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
37.98. The limits of agreement14 (mean ± 1.96 SD) were 58.4 ± 493.7 mm3 . The mean percentage difference was −0.12 (range, −8.9 to 17.1), SD 4.7, SE 0.7, giving limits of agreement of −0.12 ± 9.2% (Figure 5). With regard to the limbs, the corresponding calculations were: mean difference in the measurements of Observers A and B, −36.5 (range, −352.3 to 151.6) mm3 , SD 98.1, SE 15.3. The limits of agreement14 (mean ± 1.96 SD) were −36.5 ± 192.3 mm3 . The mean
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644 Table 1 Estimates (SE) of regression coefficients by linear regression analysis (regressions made by SPSS); square root transformations, residuals and R2 are shown
Observer A – total Observer A – limbs Observer B – total Observer B – limbs Mean, whole-body Mean, limbs
Intercept
CRL
324.0 35.6 −78.2 54.0 135.9 22.7
−69.0 −8.1 −30.9 −10.6 −49.8 −9.0
4.95 0.48 4.20 0.57 4.55 0.63
150
Residual SE
R2
322.8 57.0 314.0 88.9 0.15 35.3
0.99 0.97 0.99 0.95 1.00 0.99
% difference A−B
Response
CRL2
200 100 50 0 − 50 − 100 − 150 − 200
10
1
CRL, crown–rump length.
1000
10000
Figure 6 Difference in volume of limbs obtained by Observer A and Observer B expressed as a percentage of the average limb volume calculations of both observers.
20.00 15.00 10.00
14 000
5.00
12 000
0.00 − 5.00 −10.00
Volume (mm3)
% difference A−B
100
Average volume of limbs (mm3)
−15.00 −2 0.00 100
1000 10 000 Average whole-body volume (mm3)
100 000
10 000 8000 6000 4000
Figure 5 Difference in volume of bodies obtained by Observer A and Observer B expressed as a percentage of the average whole-body volume calculations of both observers.
2000 0
percentage difference was −12.2 (range, −173.6 to 46.3), SD 34.49, SE 5.4, giving limits of agreement of −8.8 ± 67.6% (Figure 6). Regression analysis of the relationship of estimated volume to CRL of both observers showed that square root transformation of all data (whole-body volumes, volumes of limbs) fitted best (Table 1); the mean estimates are shown in Table 2. We computed square root transformed regressions of the average whole-body volumes and average volumes of
5
10
15
20
25
30
35
40
45
50
55
60
Crown−rump length (mm)
Figure 7 Estimates of total body volume (average of calculations by Observers A and B), regression line, and lines showing average ± 1.96 SD. , Average volume as deduced by Observers A , mean; , +1.96 SD; , −1.96 SD. and B;
the limbs from the data of Observers A and B (Figures 7 and 8). The volume of the limbs as a proportion of the mean whole-body volume increased from 4.7% at a CRL of 15 mm to 9.3% at a CRL of 55 mm (Table 2).
Table 2 Mean volume estimates (mm3 ) of Observers A and B, and mean estimates of the averages. Regression by square root transformation fitted best. The mean volume of the limbs as a proportion of the mean whole-body volume of embryonic/fetal bodies for Observers A and B (after regression made by SPSS; square root transformation) is also shown
CRL 10 15 20 25 30 35 40 45 50 55
A + B volume (mm3 )
A volume (mm3 )
B volume (mm3 )
A + B limbs (mm3 )
A limbs (mm3 )
B limbs (mm3 )
Limb volume as % of whole-body volume
93 415 962 1737 2738 3973 5434 6948 9040 11 169
88 407 958 1739 2756 4011 5488 7214 9167 11 336
100 423 967 1732 2719 3936 5372 7034 8909 11 002
19.4 56.7 129.1 224.0 336.0 476.0 642.0 830.0 1042.0
18.5 58.3 120.7 205.7 313.5 443.7 596.8 771.7 968.0
19.9 65.0 136.3 233.6 357.9 507.4 683.8 885.4 1112.0
4.7 5.9 7.4 8.2 8.5 8.8 9.2 9.2 9.3
A limbs, mean volume of limbs as deduced by Observer A; A volume, mean volume of whole body as deduced by Observer A; B limbs, mean volume of limbs as deduced by Observer B; B volume, mean volume of whole body as deduced by Observer B; A + B limbs, mean volume of limbs as deduced by combining results of both observers; A + B volume, mean volume of whole body as deduced by combining results of both observers; CRL, crown–rump length.
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
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Volume (mm3)
1200 1000 800 600 400 200 0
0
5
10 15 20 25 30 35 40 45 50 55 60 Crown−rump length (mm)
Figure 8 Estimates of limb volume (average of calculations by Observers A and B), regression line, and lines showing average ± 1.96 SD. , Average volume as deduced by Observers A , mean; , +1.96 SD; , −1.96 SD. and B;
DISCUSSION For more than three decades obstetricians have been attempting with varying degrees of success to estimate the weight of the human conceptus by using 2D ultrasonography. 3D ultrasonography, however, has the potential for improving weight estimations using volumetry. In vitro6 – 12 and in vivo6,13 studies have been done to test the validity of 3D volume estimation. The validity of in vivo measurements in clinical situations cannot be assessed. Therefore in vitro studies may be useful for evaluating the accuracy of volume estimation. However, in vitro studies must be carefully set up to compensate for the environment. For example, when the volume of an object is estimated by 3D ultrasonography in an in vitro setting such as an object lying in a water bath, one has to consider the effect of the temperature of the water on the ultrasound velocity, because ultrasound propagates more slowly at lower temperatures6 . It is also important to consider the point-spread function of the imaging system, which makes the surface of an object appear blurred in an ultrasound image6,7 . The point-spread function is dependent on the resolution of the ultrasound beam. A high-frequency annular array transducer as used in our study produces very thin ultrasound slices of embryos and small fetuses and is particularly suited for creating 2D-images and thus 3D-images of high quality, reducing the effect of the pointspread function. The exact placement of the contours around an embryo or a fetus is subject to individual variation, as shown by the small differences between two observers in an in vitro study7 . Usually, the observer draws the segmentation line slightly outside the real surface in the geometric visualization mode, and volume estimations with outer surfaces tend to become larger and cavities with inner surfaces to become smaller6,7 . In recent years an increasing number of in vivo studies of 3D ultrasound volume calculations of embryos and fetuses and their organs have been published3 – 6,15 – 23 . The age and size of the fetus have a significant impact on such estimations. Because third-trimester fetuses are
Copyright 2006 ISUOG. Published by John Wiley & Sons, Ltd.
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too large for whole-body calculations, 3D volume acquisitions of only parts of the fetal body (upper arm, thigh and abdomen) have been done to construct a best-fit formula for predicting fetal weight15 . Volume measurements have been performed in placentas and fetuses at the beginning of the second trimester (15–17 weeks’ gestation) by Hafner et al.3 . They did not include measurements of the limbs for the volumetry of fetuses, because the 3D software was not able to handle more than one volume in the data set at a time (virtual organ computer aided analysis (VOCAL), Voluson 650, Kretztechnik). In the VOCAL technique, longitudinal sections of a fetus around a fixed axis are obtained, and contours of the trunk and head are traced manually. It is not possible to include traces around the limbs. The VOCAL technique was also applied in a recently published study of first-trimester volumetry, which presented measurements of fetal heads and trunks in fetuses from 45 to 84 mm CRL (VOCAL, Voluson 730 Expert, GE Medical Systems)5 . A different study analyzing 72 embryos and fetuses from 6 to 12 weeks’ gestation reported that limbs were included in the volumetry using the same software (VOCAL, Voluson 730, Kretztechnik); however the authors did not describe how this was performed, nor did they present images of the reconstructed volumes4 . That limbs are not a negligible part of the fetal volume has been shown in the present study; the volume of the limbs as a percentage of the total volume increases from approximately 5% at 7 weeks’ gestation to approximately 9–10% at the end of the first trimester. Based on our knowledge of embryonic/fetal development we can assume that the percentage volume contribution of the limbs in the second trimester is approximately 10% of the whole-body volume or weight. Therefore we recommend including the limbs in 3D estimates. The ultrasound volumetry in the present study was performed using a high-frequency annular array transvaginal transducer and specially designed software that enabled the analysis of head, trunk and limbs in the same data set. Thus, it was possible to make volume reconstructions with a high degree of accuracy. The mean of the average measurements of whole-body volume and limb volume of 44 specimens obtained by the two observers is probably the best available volume/weight estimate of living embryos and young fetuses (Figure 7, Table 2). The percentage error of −0.12 (range, −8.9 to 17.1), SD 4.7, in the whole-body volume calculations between the observers showed good reproducibility. The corresponding results for the limbs (−12.2 (range, −173.6 to 46.3), SD 34.49) were significantly poorer. One reason for the discrepancies in the volume estimation of the limbs was probably that the observers defined the borders between bodies and limbs at the shoulders and hip girdles differently. Because the absolute volumes of the limbs were very small, differences between the observers became relatively large.
CONCLUSIONS There are many sources of error that may affect volume estimation, such as poor image quality of the original
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2D data set, movement artifacts, and incorrect tracing of contours. Optimization of the original 2D image by using ultrasound equipment that generates high resolution and thin ultrasound tomograms is a prerequisite for acceptable 3D ultrasonography. The segmentation process in the ‘anyplane’ 2D slices of the 3D data set is an essential part of 3D volumetry. Therefore it is important to specify precisely how and where the segmentation lines are placed, because erroneous placement will lead to significant errors in the estimation of small volumes, such as those of embryos or young fetuses. The present study showed that it is possible to reconstruct complex small anatomic structures and calculate the volume of human embryos and fetuses in vivo by using dedicated 3D-ultrasound equipment. The reproducibility of whole-body volume estimates seems to be high. We also showed that the limbs represent a significant proportion of the volume of the body. We recommend that limb measurements be included when carrying out embryonic/fetal volumetry. It is desirable that commercially available 3D ultrasound machines have 3D software that makes it possible to combine several volume measurements from various anatomic regions of the embryo/fetus in the same data set at the same time. Volumetry represents new basic research on the living human embryo. Through it we obtain information about embryonic size and thus weight, and it becomes possible to analyze growth24 . Volumetry in abnormal embryonic/fetal development has the potential to differentiate between normal and abnormal size of embryos/fetuses and their organs18 .
ACKNOWLEDGMENTS We thank Nancy Lea Eik-Nes for revision of the paper.
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