First-trimester Placenta Volume and Three-dimensional Vascularization

Technique of Acquisition

The entire view of the placenta needs to be identified using a transabdominal volumetric probe by two-dimensional ultrasound, and the volume box was adjusted to include the entire placenta. The angle of volume acquisition varied from 45° to 90° according to placental size (Fig. 1).⁸ The volume acquisition should be obtained in “maximum” quality and setting the duration between 10 and 15 seconds. For posteriorly and laterally located placentas, a slight lateral inclination of the transducer is performed to acquire the entire placenta. The same preestablished instrument settings should be used in all the acquisitions, as reported in Table 1.

After the acquisition from the real-time images, the PD is superimposed (Fig. 2), and a second volume is acquired.⁹ In order to obtain reproducible results the gain is set in all the cases respectively at −0.2 for PD and −0.8 if the high-definition flow is used.

Volume Analysis

After the acquisition, the placental volume was stored for later analysis. The software virtual organ computer-aided analysis is used to evaluate placental volume by obtaining a sequence of 12 sections of the placenta, separated by successive rotations of 15°. In each plane, the contour was traced manually, and the organ was then reconstructed automatically by the software (Figs 3 and 4).

After the estimation of the placental volume, the 3D-PD histogram was used to determine the following vascular indices from computer algorithms⁴—(1) vascularization index, which refers to the color voxel/total voxel ratio, that is, the color percentage within the volume of interest (placenta), and provides an indication of how many vessels can be detected within the placenta (vascularity); (2) flow index, which refers to the weighted color voxel (on a scale of 0–100)/total color voxel ratio and provides an amplitude value for the color signal, thus giving information on how many blood c ells are being transported at the time of the 3D sweep (placental blood flow); (3) vascularization flow index refers to the weighted color voxel/total voxel ratio, combining the information of vessel presence (vascularity) and the amount of transported blood cells (blood flow) (Figs 5 and 6).

Reference Limits for Gestation

Since placental volume increases with gestational age, we constructed reference limits for crown-rump length (CRL) in order to allow proper comparison (Fig. 7).⁸ Similarly, indices of vascularization change with advancing gestation and reference limits for CRL were constructed (Fig. 8).⁹

Significance of Placental Vascular Indices

Although caution is necessary for relating 3D Doppler measurements to anatomical placental characteristics, we related histomorphological analysis of chorionic villi to the ultrasonographic evaluation of 3D placental volume and vascularization in pregnancies undergoing chorionic villus sampling (CVS).¹⁰ A direct relationship was evidenced between the number of capillaries and the 3D Doppler placental vascularization, demonstrating a direct relationship between histological results and 3D ultrasound evaluation of placental vascularization.¹⁰

Changes of Placental Volume and Vascularization in Abnormal Pregnancies

Maternal Smoking

We have investigated the effects of smoking on placental volume and its Doppler-measured vascularization.¹¹ Smoking <10 cigarettes/day did not induce any evident changes in the 3D Doppler placental vascular indices considered, while in mothers who smoked heavily, there was a significant reduction of all these indices, suggesting a reduced placental vascularization already present at 11⁰–13⁶ weeks of gestation. We did not evidence any dose-dependent effect of maternal smoking >10 cigarettes/day on placental vascularization. A possible explanation is the absence of a dose-dependent effect of smoking on placental angiogenesis, as suggested by in vitro studies. Of interest is that placental vascular indices are significantly related to birthweight, and this relationship resulted stronger when the analysis was restricted to heavy smokers.

Prediction of Preeclampsia (PE)

We tested the role of uterine artery Doppler, placental volume assessed by 3D ultrasound, and their combined use in the first trimester for the prediction of PE in an unselected population of nulliparous pregnancies.⁸ We demonstrated how both abnormal uterine artery and reduced placental volume resulted in independent risk factors to develop PE; of interest is the lack of relationship found in our study between uterine Doppler and placental volume suggesting their independent role as risk factors. The lack of relationship may reflect two different pathophysiological mechanisms involved in the genesis of PE. Pregnancies with abnormal Doppler uterine waveforms may reflect an inadequate trophoblastic invasion of the maternal spiral arteries that may occur in presence of a normal size placenta. On the contrary, despite the presence of a normal trophoblastic invasion as indicated by normal uterine Doppler indices, a small placenta, for example, by producing a smaller amount of vasoactive substances or antioxidative agents, may act as a distinct genesis of PE. Irrespective of the underlying mechanisms, our data suggest that combining the first trimester uterine Doppler screening with the assessment of placental volume by 3D ultrasound may improve the detection rate of PE to values similar to those found for the late second trimester. These data encourage further research on uterine Doppler combined with placental volume as well as with other potential biochemical markers in the attempt to detect pregnancies at risk of PE in the first trimester.

Prediction of Fetal Growth Retardation (FGR)

We tested the efficiency of the assessment of placental volume and its vascularization in pregnancies characterized by low maternal serum of pregnancy-associated plasma protein A (PAPP-A) at 11⁰–13 weeks of gestation,¹² a condition associated with a higher incidence of low birth weight.¹³ An interesting aspect of this study was the evidence that among pregnancies with low serum PAPP-A levels, only those resulting in the birth of fetal growth retardation (FGR) neonates showed a significant decrease of the 3D-PD indices of placental vascularization at 11⁰–13⁶ weeks of gestation. On the contrary, pregnancies with low PAPP-A concentrations delivering newborns of appropriate weight or healthy low birthweight neonates did not show differences in placental vascularization during the first trimester. A second finding of this study is the significant association between the degree of reduction in 3D placental Doppler indices and the severity of the growth defect at birth. Although caution is necessary for the interpretation of these Doppler indices as the experimental design of this study does not allow for to investigation of direct links between vascular indices and histomorphometric parameters, the relationships found between the changes in placental vascular indices and birthweight support the hypothesis that the Doppler parameters measured reflect a reduced placental vascularization already present in the first trimester.

Maternal Type 1 Diabetes Mellitus (DM)

We investigated placental volume and vascularization assessed by 3D ultrasound in a population of type 1 DM.¹⁴ We demonstrated that while a diabetic condition does not affect placental volume, its vascularity is significantly increased. An interesting result of this study is the evidence that, among pregnancies with diabetes, those with poorer glycemic control, as expressed by glycated hemoglobin concentrations ≥7%, exhibited significantly higher values of placental vascular indices. This suggests that placental vascular development in women with diabetes can be affected by first-trimester maternal hyperglycemia. Although associations between first-trimester glycemic control and placental angiogenesis have been already recognized by stereological studies on human placentas at term showing an increased capillary volume in DM with poor metabolic control, our data are the first to provide in vivo evidence of increased vascularity already present during the first trimester.

The clinical significance of the modification of placental vascular indices in DM remains to be established. However, there is some evidence that the phenotypic changes in the placental microvascular bed may extend to umbilical vascular cells and even to fetal vasculature and heart, which might produce an overt pathological response in the offspring later life if challenged with additional cardiovascular stresses. The presence of abnormalities in fetal cardiac function occurring already in early gestation in fetuses of type 1 DM supports this hypothesis.

In conclusion, placentas of diabetic mothers exhibited evidence of first-trimester increased vascular indices and these changes become more evident with worsening glycemic control. The ability to document placental vascular changes so early in pregnancy potentially opens new avenues to understanding the range of fetal developmental consequences of maternal glycemic status.

Pregnancies from In Vitro Fertilization (IVF)

Pregnancies achieved with IVF are at increased risk of obstetrical complications such as PE. Further, the incidence of this disease is significantly higher in pregnancies obtained with donor oocytes than in pregnancies achieved with autologous IVF. We analyzed three groups of pregnancies (naturally conceived, autologous IVF, and heterologous IVF); the two IVF groups considered, and the incidence of PE was three-fold higher in donor oocytes recipient pregnancies; thus, suggesting the coexistence of other causative factors. No relationships were found between placental volume and maternal age, and multivariate logistic regression evidenced an independent role of placental volume. Of interest is the significant contribution in the prediction of PE given from donor IVF in association with maternal age and placental volume. An impaired trophoblastic invasion is considered to be the major etiological factor in the development of PE, particularly when at early onset. Indeed, there are several evidence showing that an abnormal impedance to flow in the uteroplacental circulation, manifested by an increased uterine artery pulsatility index (PI), is already present at 11 weeks of gestation in pregnancies that will develop PE. Despite the high prevalence of PE in IVF pregnancies found in this study, we did not find evidence of any differences in mean uterine PI values. This is in agreement with other reports and suggests that the mechanism causing the increased incidence of PE in IVF pregnancies is unlikely to be related to impaired uteroplacental perfusion.

The presence in IVF pregnancies of a reduced placental volume, particularly marked in donor oocytes recipients and its association with the development of PE despite the presence of normal uterine artery PI values found in this study must be pointed out. One of the different hypotheses that have been suggested to explain the increased incidence of PE in IVF pregnancies is a different immune response of the mother to trophoblastic antigens. This abnormal immune response may be either secondary to the different maternal endocrine characteristics of women undergoing IVF or to the presence of foreign antigens, it is the case with donor oocytes (since placental development is influenced by maternal immune response). This hypothesis may well explain the reduced volume hereby found in IVF pregnancies.

The importance of first-trimester identification of IVF pregnancies at higher risk of developing PE should be underlined since this condition is associated with an increased risk of perinatal mortality and morbidity and both short and long-term maternal complications that deserve targeted antenatal care and surveillance. However, the degree of overlap in placental volume, multiple of the median values, evidenced in this study between pregnancies, who did or did not develop PE, suggests a limited role in its isolated use as a screening tool. In naturally conceived pregnancies effective screening for PE was achieved only with the combined use of maternal variables, uterine artery Doppler, and biochemical markers. A similar approach should also be applied to IVF pregnancies adding placental volume data. The results obtained in the multiple logistic models constructed support this hypothesis by showing a potential combined role of placental volume with maternal characteristics such as age or type of conception.

We also analyzed placental volume in IVF pregnancies divided according to the use of fresh or frozen-thawed embryos. We found a reduced placental volume particularly evident when fresh embryos were used, and it was associated with a higher incidence in the development of PE.15,16

The lower placental volume found in IVF with fresh embryos is probably secondary to the inferior endometrial receptivity occurring in these pregnancies rather than in naturally conceived and cryopreservation groups. Indeed, the use of ovarian stimulation in the former group is associated with an altered endometrial development that may impair its receptivity, and therefore, the subsequent placental development. Although the role of endometrial receptivity seems to play a pivotal role, it is not possible to exclude other factors that may explain our findings. A possibility is that the freeze-thaw procedure may filter out flawed embryos, thus selecting a higher percentage of more viable blastocysts to be transferred. A second possibility is that freezing and thawing the embryos may improve their biological ability to reach normal implantation. Irrespective of the underlying cause, which is out of the objectives of this work, we demonstrated that placental volume is reduced in IVF pregnancies achieved with fresh embryos and that these pregnancies are at higher risk of PE.

In conclusion, during the first trimester, placental volume in IVF pregnancies is lower than in naturally conceived pregnancies, and it is particularly reduced in donor oocyte recipients and fresh embryos. Placental volume assessment at 11⁰–13⁶ weeks may be, therefore, useful to identify among IVF pregnancies those destined to develop PE.