Placental elasticity in trisomy 21: prenatal assessment with shear-wave elastography

Introduction

Trisomy 21 (T21), also known as Down syndrome, is the most prevalent chromosomal aneuploidy in live births, with a frequency of approximately 1/700 [1]. It is characterized by the presence of a partial or complete extra copy of chromosome 21 [2]. This genetic disorder usually causes miscarriage, stillbirth, and fetal structural anomalies in the prenatal period. It is also associated with intellectual disability, various organ dysfunctions, and increased cancer risk after live birth [3], [4], [5]. Due to its frequency and these adverse consequences, prenatal diagnosis of T21 is important. Although advanced maternal age (≥35 years) has traditionally been used as a primary screening criterion for T21, prenatal screening methods have significantly improved over the past few decades, and screening tests that provide highly accurate results have been obtained. Non-invasive prenatal testing (NIPT) for T21 screening is a new method that has been introduced into clinical practice since 2011 [6]. However, its global implementation remains limited due to high costs [7], 8]. The detection rate (DR) and positive predictive value (PPV) of T21 in first-line screening with NIPT were reported as 98% and 96 % [9], [10], [11]. The first trimester screening test, which includes a combination of maternal age, fetal nuchal translucency (NT) measurement, and two placental hormones (human chorionic gonadotropin; β-hCG) and (pregnancy-associated plasma protein-A; PAPP-A), remains the most commonly utilized screening tool for T21 [1], 2]. This combined screening test has a sensitivity of 92.16 % (81.5–96.9) and a specificity of 80.86 % (79.7–81.9) with a cut-off value of 1/300 [12].

In addition to fetal structural abnormalities, placental developmental defects have also been identified in T21 pregnancies. Histopathological evaluations of placentas affected by T21 reported enlargement, asymmetry, and irregular arrangement of chorionic villi [13]. Additionally, defective trophoblast fusion and differentiation have been documented, leading to an increased percentage of double-layered trophoblasts and excessive villous capillaries [14]. Inadequate differentiation from cytotrophoblast to syncytiotrophoblast leads to abnormal hCG secretion, resulting in faulty regulation of placental development [15].

Shear wave elastography (SWE) is a non-invasive, ultrasound-based technique that quantitatively measures tissue elasticity and stiffness [16]. Studies have been reported that histopathological changes, such as fibrosis, inflammation, or neoplasia are also related to tissue elasticity [17]. SWE basically works on the principle of generating shear waves with a focused ultrasound beam via an ultrasound transducer, followed by real-time imaging of these shear waves, allowing for a quantitative assessment of tissue stiffness [18]. Although there is no evidence yet regarding potential adverse fetal effects of SWE, it is recommended that examinations be used with caution, ensuring that thermal and mechanical index values ​​remain within safe limits during examinations [19]. Recent studies have reported that placental histopathological changes in diseases such as preeclampsia and gestational diabetes can be effectively reflected through ultrasonographic quantitative elastographic measurements [20], [21], [22].

The primary aim of this study was to quantitatively investigate placental tissue stiffness pregnancies affected by T21 using SWE. The secondary aim was to explore the potential contribution of placental tissue stiffness measurements in predicting T21 fetuses.

Materials and methods

Study participants

The case group was defined as singleton pregnancies with karyotype analysis reported as T21 after chorionic villus sampling (CVS) or amniocentesis (AS) procedure, regardless of the indication for prenatal diagnostic testing. The control group included singleton pregnancies who underwent CVS or AS procedures because of one or more of the following criteria: an increased risk (>1:1,000) on first- or second-trimester screening tests, advanced maternal age (≥35 years), parental anxiety, or positive ultrasonographic soft markers for aneuploidy, with whose fetal genetic examination reported a normal karyotype. Women with chronic diseases such as hypertension, diabetes, chronic kidney disease, and autoimmune disease that could affect placental elasticity were excluded from the study (Figure 1).

Figure 1:

Flow chart of participants.

CVS procedures were performed transabdominally using a double-needle technique at 11–14 weeks of gestation. The procedure was performed with a 17-gauge external needle and a 19-gauge internal aspiration needle, and no more than two attempts were performed. AS procedures were performed transabdominally with a 22-gauge needle after the 15th week of gestation. All procedures were performed under continuous ultrasonographic guidance to ensure precision and safety.

Ultrasonographic examination

Ultrasonographic examinations were performed using the Samsung Ultrasound System HS70A (Samsung Medison Corporation, Republic of Korea), equipped with a 1–7 MHz curvilinear abdominal transducer, following the fetal genetic examination. Participants were positioned in the supine position and instructed to breathe lightly and minimize movement during the procedure. After determining the location of the placenta and fetus with B-mode imaging, the imaging plane was adjusted according to the location of the placenta. Maternal abdominal subcutaneous fat thickness was measured using the method described by Suresh et al. [23].

SWE measurements were performed using the point SWE (pSWE) application according to the manufacturer’s recommended protocol (S-shear wave imaging non-invasive quantification of tissue stiffness, Samsung Medison Co., Ltd. 2018). SWE measurements were performed in two separate areas of the placenta: central and peripheral. The central measurement was taken from the middle of the thickest part of the placenta, carefully avoiding vascular areas and lacunae, and placental thickness was also recorded from the same area. The peripheral measurement was taken in the same manner, 2 cm medial to the lateral border of the placenta. To ensure fetal safety, particularly when the placenta was located on the posterior uterine wall, measurements were performed from image sections where the fetus was not located in front of the region of interest (ROI) box (Figure 2). The Reliability Measurement Index (RMI) was provided in the S-Shear wave profile. RMI is a quality control parameter calculated by the weighted sum of two factors: the residual of the wave equation and the size of the wave shear wave. To enhance measurement reliability, measurements with RMI<0.4 were removed according to the manufacturer’s recommendations. The median value and interquartile range (IQR) of valid measurements are calculated automatically by the application. Reliable pSWE measurements were defined as those with a median value derived from at least four measurements with an IQR/median (M) ≤30 % and RMI≥0.4. Measurement values were recorded in kilopascals (kPa). The application also automatically calculated the depth of measurements, defined as the distance between the ROI and the ultrasound probe. In this study, these values ranged from 3.5 to 8.7 cm in the peripheral placenta and 3.5 to 8.9 cm in the central placenta.

Figure 2:

Placental share wave elastography. (A) Central (on the control placenta). (B) Peripheral (on the trisomy 21 placenta).

Results

A total of 60 pregnant women were included in the study, with 30 women in the T21 group and 30 in the control group. The demographic and clinical characteristics of both groups are presented in Table 1. The mean maternal age was significantly higher in the T21 group (33 ± 6 years) compared to the control group (30 ± 6 years) (p=0.024). There were no significant differences between the two groups in terms of parity, body mass index (BMI), smoking status, or the diagnostic method used (CVS or AS) (p>0.05, for all).

The sonographic and SWE characteristics of the study groups are presented in Table 2. The mean gestational age at the time of ultrasonographic measurement in both groups was 16 ± 2 weeks, with no significant difference was found between them (p=0.573). There was also no significant difference in maternal abdominal subcutaneous fat thickness, placental location, or placental thickness between the two groups (p>0.05, for all). Peripheral placental SWE velocity was significantly higher in the T21 group (7.4 ± 3.7 kPa vs. 4.8 ± 3.6 kPa, p=0.004). Similarly, central placental SWE velocity was also significantly higher in the T21 group (6.5 ± 2.1 kPa vs. 4.1 ± 2.6 kPa, p<0.001). Peripheral and central placental depths were similar in both groups (p>0.05, for all).

The ROC curve analysis to evaluate the usefulness of peripheral and central placental SWE velocity in the diagnosis of Trisomy 21 is shown in Table 3. The area under the curve (AUC) for peripheral placental SWE velocity was 0.715 (cut-off, ≥4.65 kPa; 95 % CI: 0.576–0.854, p=0.004), with a sensitivity of 70 % and a specificity of 66.7 %. The AUC for central placental SWE velocity was 0.837 (cut-off, ≥4.35 kPa; 95 % CI: 0.728–0.946, p<0.001), with a sensitivity of 76.7 % and a specificity of 73.3 % (Figure 3).

Figure 3:

The ROC curve analysis of predictive performance for peripheral and central placental SWE velocity in the diagnosis of Trisomy 21.

The univariate and multivariate analysis on independent predictors of T21 are shown in Table 4. In univariate analysis, maternal age ≥35 years (OR=1.10, 95 % CI: 1.01–1.21, p=0.030) and peripheral placental SWE velocity ≥4.65 kPa (OR=4, 95 % CI: 1.36–11.70, p=0.011) were significant predictors of T21. However, neither maternal age nor peripheral placental SWE velocity remained significant in multivariate analysis (p>0.05, for both). In contrast, central placental SWE velocity ≥4.35 kPa was a significant predictor of T21 in both univariate (OR=9.03, 95 % CI: 2.80–29.13, p<0.001) and multivariate analyses (OR=6.64, 95 % CI: 1.93–22.75, p=0.003).

In Pearson’s correlation analysis, there was no significant association between placental SWE velocities and BMI, gestational age at measurement, maternal abdominal subcutaneous fat thickness, and placental thickness (p>0.05, for all) (Table 5).

Discussion

In this study, we comprehensively investigated placental stiffness in pregnancies affected by T21 using SWE, a novel and non-invasive ultrasound-based technique for assessing tissue elasticity. Our results demonstrated that both central and peripheral placental stiffness were significantly increased in T21 cases. Moreover, we found that placental SWE measurements could serve as a predictive tool for T21, with a cut-off value of ≥4.35 kPa, central placental SWE velocity was associated with a 6.64-fold increased risk of T21, even after adjusting for maternal age a well-established risk factor for T21.

Previous studies have documented various developmental defects in the placentas of T21 pregnancies [24]. Although many different theories and evidence have been presented, this defect in placental development leads to an abnormal increase in the β subunit of hCG [15]. Syncytiotrophoblasts, which are formed by the aggregation and differentiation of villous cytotrophoblasts, are the active endocrine unit of the placenta and secrete placental hormones such as hCG, human placental lactogen (hPL), and human placental growth hormone (hPGH). In T21-affected placentas, the formation of syncytiotrophoblasts is both delayed and defective, leading to impaired fusion and differentiation of cytotrophoblasts into mature syncytiotrophoblasts [14]. This delayed maturation in T21 chorionic villi resulting in persistent cytotrophoblastic layers, an increased percentage of double-layered trophoblasts, and a higher proportion of villous capillaries, as confirmed by histomorphometric analysis [24]. In addition, while disrupted trophoblast transformation results in syncytiotrophoblastic hypoplasia, studies have shown a significant reduction in mature hCG receptors expressed on the cytotrophoblast surface [25]. Although syncytiotrophoblast formation is defective and delayed in T21-affected placentas, the existing syncytiotrophoblasts have been found to exhibit hyperactivity [15]. The molecular mechanisms underlying these changes in trophoblast differentiation is still not fully elucidated. Recently, evidence was reported that overexpression of the amyloid precursor protein, which is located on chromosome 21 and is also associated with Alzheimer’s disease, may contribute to disrupted trophoblast development in T21 pregnancies [26].

Previous studies have shown that SWE, an ultrasound-based technique for evaluating tissue elasticity and stiffness, can non-invasively demonstrate histopathological changes in tissues affected by conditions such as fibrosis, inflammation, and malignancy [17]. In gynecology and obstetrics, SWE has been studied in conditions such as polycystic ovary syndrome, cervical stiffness, fetal lung maturity, and placental abnormalities [22], [27], [28], [29]. Studies have reported that cervical elasticity measurement can predict preterm birth, while fetal lung elasticity correlates with lung maturation [22], 27], 29]. Research on placental elastography has primarily focused on conditions such as preeclampsia and, more recently, gestational diabetes. These studies showed that there was a significant increase in placental stiffness in preeclamptic and diabetic pregnancies and that this increase was also associated with adverse pregnancy outcomes [30], [31], [32], [33], [34], [35]. There is no study yet evaluating placental elasticity in T21 cases. In our study, we used SWE to examine placentas from T21 pregnancies, assessing elasticity in two distinct regions – central and peripheral – and our findings showed that placental stiffness in T21 cases was significantly higher than in controls. This increase in tissue stiffness was higher in the center of the placenta compared to the periphery. Previous studies have reported inconsistent findings regarding the relationship between placental stiffness and placental regions. Two studies investigating placental elasticity in gestational diabetes found that the peripheral region of the placenta was stiffer in the diabetic group [32], 36]. However, in the control group, Yuksel et al. reported greater stiffness in the central region, whereas Liu et al. observed higher stiffness in the peripheral region [32], 36]. Habibi et al. showed that in cases of fetal growth restriction, the central placental area on the maternal side exhibited greater stiffness compared to the peripheral area, while no significant differences were observed between these regions on the fetal side [37]. Although the exact molecular mechanisms underlying the developmental defects in T21 placentas remain unclear, there is sufficient evidence for impaired differentiation of villous trophoblasts. The abnormal increase in the number of untransformed cytotrophoblasts and the formation of layers may be the main histopathological reason that may affect placental tissue elasticity. Furthermore, the increased number of capillaries in T21 placentas, which occurs secondary to the increase in cytotrophoblasts, may contribute to the greater effect observed in the central placental region, where vascular development is more intense. However, further research, including simultaneous placental pathology assessments, will be necessary to confirm and fully understand this relationship.

T21 is the most common genetic disorder that causes intellectual disability and structural anomalies in live births [1]. Prenatal diagnosis of T21 provides parents with the option of making informed decisions regarding pregnancy management. Although T21 can result in anomalies affecting various fetal structures, including the heart, gastrointestinal tract, or brain, these abnormalities may not always be present or detectable during the prenatal period [3], 5]. To improve prenatal detection of T21, various screening tests have been developed, with their accuracy increasing over time. NIPT is the most recent development and offers a high detection rate of approximately 98 % for T21. However, due to its high cost, NIPT is not universally used as the first-line prenatal screening option in many countries [38]. As a result, the first-trimester screening test, which includes maternal age, fetal NT measurement, and serum biomarkers such as hCG and PAPP-A, remains a cornerstone of prenatal screening. In addition to biochemical markers, several sonographic markers have been identified to increase the likelihood of diagnosing T21. These include increased nuchal fold thickness, nasal bone hypoplasia, and the presence of an aberrant right subclavian artery, all of which significantly raise the likelihood of T21 [1], 2]. Building on these diagnostic advancements, our study introduced an additional screening tool by investigating placental elasticity using SWE. Our findings demonstrated that central placental SWE velocity, with a cut-off value of ≥4.35 kPa, predicted T21 with 76.7 % sensitivity and 73.3 % specificity and was associated with a 6.64-fold increased risk of T21, even after adjusting for maternal age. Although peripheral placental elasticity was also evaluated, its predictive value for T21 was lower, with a sensitivity of 70 % and a specificity of 66.7 % at a cut-off value of ≥4.65 kPa. Importantly, peripheral placental stiffness did not significantly increase the risk of T21 when maternal age was considered in the analysis. These findings highlight the greater diagnostic potential of central placental elasticity compared to peripheral measures.

There are several limitations to this study that should be acknowledged. First, although the sample size was sufficient to detect modest effect size differences, it is unclear whether elastography is a useful adjunct to initial screening for T21. Larger sample sizes in future studies would increase the generalizability of the findings and provide more robust statistical power to confirm these results. Additionally, this study assessed placental elasticity only at specific gestational ages, so potential changes in placental elasticity throughout pregnancy were not investigated. Understanding how placental rigidity evolves over time may provide deeper insights into the progression of placental abnormalities associated with T21, and analysis of long-term outcomes in T21 fetuses with high elastography measurements may be useful. Second, the absence of histopathological examinations limits our understanding of the underlying biological mechanisms contributing to the increased placental stiffness observed in T21 cases. Without histological data, the precise anatomical and cellular factors influencing placental elasticity remain unclear. Despite these limitations, this study offers important preliminary evidence on the potential role of placental elasticity in the prenatal diagnosis of T21. A notable strength of this study is that it is the first study to assess placental elasticity in T21-affected pregnancies using SWE. Moreover, by ensuring that median values were derived from at least four reliable pSWE measurements – with an IQR/M ratio ≤30 % and a reliability RMI≥0.4 – we adhered to rigorous measurement standards. This methodological approach enhances the reliability of the data and strengthens the validity of the study’s findings.

In conclusion, our study suggests that increased central placental stiffness may be a potential marker for T21 and may complement current prenatal screening methods. Future studies including histopathological analyses are important to better understand the underlying mechanisms driving this increased stiffness in T21-affected placentas. If validated in larger studies, SWE could be incorporated into prenatal screening protocols and potentially improve early detection and risk assessment for T21 pregnancies.