Trimester-specific reference values of ultrasound-derived fat fraction and automated point shear-wave elastography for liver assessment in healthy singleton pregnancies

Introduction

During pregnancy, the maternal liver undergoes substantial physiological adaptations, including trimester-specific changes in function, lipid metabolism, and tissue biomechanics (1). Conventional approaches for evaluating liver health—such as serum biochemical tests and standard ultrasonography—provide valuable clinical information but are limited in their ability to capture the dynamic hepatic changes that occur throughout gestation. Recently, ultrasound-derived fat fraction (UDFF) and automated point shear-wave elastography (Auto-pSWE) have emerged as innovative, noninvasive imaging techniques that can quantitatively assess hepatic steatosis and tissue stiffness, respectively (2,3). UDFF estimates liver fat content by analyzing ultrasound attenuation and backscatter, providing a rapid, bedside alternative to magnetic resonance imaging (MRI)-based fat quantification without radiation exposure (4). Auto-pSWE measures liver elasticity via shear-wave velocity (SWV), offering insights into liver fibrosis and structural alterations (5).

Although these technologies have demonstrated diagnostic value in general hepatology—especially for detecting and staging nonalcoholic fatty liver disease (NAFLD) and fibrosis (6)—their application during pregnancy remains limited. The unique hormonal, hemodynamic, and metabolic shifts of gestation can influence hepatic structure and function, yet standard liver tests and imaging modalities lack the sensitivity to differentiate normal pregnancy-related changes from early pathological conditions such as gestational cholestasis or steatosis. Furthermore, few studies have established gestational age-specific reference values for liver fat fraction and stiffness measured via quantitative ultrasound. For example, although transient elastography studies have noted changes in liver stiffness during pregnancy (1), they have not provided comprehensive trimester-specific normative data or incorporated fat quantification (7). Moreover, evidence on the correlation between hepatic elastography parameters and biochemical markers in pregnant populations remains scarce.

These deficiencies in research limit the clinical utility of advanced ultrasound biomarkers for prenatal liver assessment. Establishing trimester-specific reference ranges for UDFF and Auto-pSWE in healthy pregnancies is essential for distinguishing normal hepatic adaptations from early disease states and thus enabling timely diagnosis and management. We conducted a prospective cohort study to generate longitudinal normative data and clarify the associations with maternal clinical and biochemical parameters, with the overall aim being to address these unmet needs and support the integration of these novel ultrasound techniques into routine obstetric care. We present this article in accordance with the STROBE reporting checklist (available at https://qims.amegroups.com/article/view/10.21037/qims-2025-1264/rc).

Methods

Study design and participants

This prospective, longitudinal cohort study consecutively enrolled normotensive women with singleton pregnancies admitted to prenatal care at Gansu Provincial Maternity and Child-care Hospital between January 2023 and November 2024. The final analytic cohort comprised 30 gravidae with complete triphasic assessments [first (11–13⁶⁄₇ weeks), second (20–24 weeks), and third (32–36 weeks) trimesters]. All participants underwent standardized protocols including the following: documentation of comprehensive medical history; anthropometric profiling [preconception body mass index (BMI): 18.5–24.9 kg/m²; gestational weight gain: 8.0–14.0 kg]; and multimodal hepatic evaluation, including (I) conventional B-mode ultrasonography, (II) UDFF quantification, (III) Auto-pSWE, and (IV) a serum biochemical panel [alanine aminotransferase (ALT), aspartate aminotransferase, gamma-glutamyl transferase, and alkaline phosphatase]. Ethical approval was obtained from the Institutional Review Board (IRB) of Gansu Provincial Maternity and Child-care Hospital (Gansu Provincial Central Hospital; Approval No. 2022-66). All participants provided written informed consent prior to participation in the study. The study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments.

Inclusion and exclusion criteria

Inclusion criteria

Participants were included if they met all of the following criteria: singleton pregnancy with confirmed gestational age based on crown-rump length measurement during the first trimester, no major obstetric complications throughout the surveillance period, pre-pregnancy BMI within the World Health Organization (WHO)-defined normal range (18.5–24.9 kg/m²), gestational weight gain within the guidelines issued by the Institute of Medicine (IOM), and completion of all imaging and biochemical assessments across all three trimesters.

Exclusion criteria

Participants were excluded if any of the following conditions were present: development of gestational comorbidities, including hypertensive disorders of pregnancy (HDP), gestational diabetes mellitus (diagnosed as per the American Diabetes Association criteria), and intrahepatic cholestasis of pregnancy (ICP; defined as serum bile acids >10 µmol/L); sonographic evidence of liver abnormalities, including focal hepatic lesions ≥5 mm in diameter and advanced hepatic fibrosis [defined as liver stiffness measurement (LSM) ≥9.5 kPa as assessed by FibroScan (Echosens, Paris, France)]; technical inadequacy in imaging assessment, including acoustic window failure (≥3 unsuccessful measurement attempts) and nondiagnostic image quality (SWE quality control score <85); and loss to follow-up or incomplete data collection/documentation.

Ultrasound equipment

All ultrasound examinations were performed with a color Doppler ultrasound system (ACUSON Sequoia, Siemens Healthineers, Erlangen, Germany), equipped with a 5C1 abdominal transducer (frequency: 3–5 MHz) and a low-frequency convex DAX probe (frequency: 1–3.5 MHz).

UDFF and Auto-pSWE data acquisition

Participants were required to fast for at least 8 hours before the examination. To standardize conditions and reduce potential variation in the results, fasting was confirmed prior to measurement. Initial scanning was performed with the 5C1 transducer for the evaluation of liver size, morphology, parenchymal echotexture, intrahepatic biliary structures, and vasculature. The oblique diameter of the right hepatic lobe and the liver length along the midclavicular line below the costal margin were measured. Additional abdominal organs were examined for any abnormalities or masses.

Subsequently, a low-frequency convex DAX probe was used. Participants were positioned in the supine or left lateral decubitus position with the right arm fully abducted to optimize intercostal access. The probe was placed in the right intercostal space with the ultrasound beam perpendicular to the liver capsule. A homogeneous area in hepatic segment V (S5) or segment VIII (S8) was selected, with large vessels, rib shadowing, and echogenic lesions being avoided. A fixed-size region of interest (ROI) 3 cm × 3 cm in size was aligned parallel to the liver capsule with the guideline positioned at the capsule surface.

Participants were instructed to hold their breath during UDFF and Auto-pSWE acquisition. Each sampling frame included 15 sub-ROIs. For those unable to hold their breath, measurements were performed during the quiet respiration. Each measurement was repeated four times, and the average value was calculated. UDFF results are expressed as percentages (%), and Auto-pSWE results are reported in terms of liver stiffness (Young modulus in kilopascals) and shear wave velocity (SWV; in meters per second). A result marked as “” indicates measurement failure. Measurements were considered valid only if the ratio of interquartile range to the median was less than 60%. All procedures were performed by maternal-fetal medicine-certified sonographers (minimum 5 years of hepatic ultrasound experience) in accordance with the consensus guidelines from the World Federation for Ultrasound in Medicine and Biology[5]. Formal inter- and intraoperator repeatability testing was not undertaken in this pilot study, and reliability was supported by standardized presets, fixed ROI depth, and on-scanner quality metrics. The experimental design process is illustrated in Figure 1. Figure 2 provides representative ultrasound images of liver UDFF and Auto-pSWE measurements.

Figure 1 Study flowchart outlining participant recruitment, eligibility screening, and data-processing procedures. ADA, American Diabetes Association; Auto-pSWE, automated point shear-wave elastography; BMI, body mass index; CRL, crown-rump length; GDM, gestational diabetes mellitus; HDP, hypertensive disorders of pregnancy; ICP, intrahepatic cholestasis of pregnancy; IOM, Institute of Medicine; LSM, liver stiffness measurement; SWE-QC, shear wave elastography quality criteria; UDFF, ultrasound-derived fat fraction; WHO, World Health Organization.

Figure 2 Representative ultrasound images demonstrating liver UDFF and Auto-pSWE measurement techniques. The probe is positioned perpendicular to the liver capsule in the intercostal space to acquire an optimal acoustic window of the right hepatic lobe. The horizontal guideline (yellow arrow) is aligned with the liver capsule. (A) During breath-hold, the region of interest is placed within homogeneous liver parenchyma. The median shear-wave velocity is 1.34 m/s, the median liver stiffness is 5.4 kPa, and the UDFF is 2%. (B,C) During UDFF acquisition, areas near large intrahepatic vessels, the capsule, ligaments, and the gallbladder are avoided (white arrows) to minimize measurement artifacts and ensure consistency. Auto-pSWE, automated point shear-wave elastography; UDFF, ultrasound-derived fat fraction.

Blood biochemical parameters

Venous blood samples were collected after a minimum of 8 hours of fasting. Laboratory analyses were performed with a fully automated biochemical analyzer (ADVIA 2400, Siemens Healthineers). The following biochemical parameters were measured: ALT, albumin (ALB), international normalized ratio (INR), total bile acids (TBA), total bilirubin (T-Bil), and prothrombin time (PT).

Anthropometric measurements

Physical examinations were performed in accordance with the anthropometric guidelines recommended by the WHO (8). Blood pressure was measured with an automated upper-arm electronic sphygmomanometer (Omron HEM-7051, Medaval, Dublin, Ireland). Height and weight were measured with a portable ultrasound-based device, while body composition was assessed with a body fat analyzer. The collected anthropometric indices included body weight and BMI.

Statistical analysis

All statistical analyses were conducted with SPSS version 27.0 (IBM Corp., Armonk, NY, USA). Normality of continuous variables was assessed with the Shapiro-Wilk test. Continuous variables with skewed distributions are presented as the median and IQR. The Mann-Whitney test was employed for comparisons of two groups, and the Kruskal-Wallis test was used for comparisons of more than two groups.

Multivariate linear regression analysis was performed to identify factors associated with UDFF and Auto-pSWE values. The Spearman rank correlation coefficient was used to assess correlations between variables. The strength of correlation was defined based on r values as follows: r<0.2, no significant correlation; 0.2≤r<0.5, weak correlation; 0.5≤r<0.7, moderate correlation; 0.7≤r<0.9, strong correlation; and r≥0.9, very strong correlation.

Results

Comparison of general clinical characteristics across trimesters

A total of 30 healthy pregnant women were included in the analysis, with data collected for each participant during the first, second, and third trimesters. Maternal age ranged from 20 to 41 years, with a mean age of 31.29±3.75 years. Gestational age across the dataset ranged from 15 to 40 weeks, with a mean of 31.92±5.05 weeks.

Comparative analysis was performed to assess trimester-related differences in physiological and biochemical parameters. Results indicated a statistically significant difference in BMI across the three trimesters (P<0.05), with a progressive increase observed with more advanced gestational stage. In contrast, serum biochemical markers—including ALT, TBA, T-Bil, PT, INR and ALB—remained relatively stable throughout gestation. Detailed results are presented in Table 1.

Trimester-based comparison of UDFF and Auto-pSWE parameters

The results demonstrated statistically significant differences in both UDFF and Auto-pSWE parameters—including elasticity and SWV—across the three trimesters (P<0.05). A clear upward trend was observed in all three parameters as pregnancy progressed.

In particular, the marked elevation in SWV suggests that hepatic tissue stiffness may increase with gestational advancement. This phenomenon may be associated with dynamic hormonal changes during pregnancy or other physiological adaptations, such as increased maternal blood volume and hepatic perfusion. Detailed data are provided in Table 2.

Correlation between UDFF, Auto-pSWE measurements, and other parameters

Correlation analysis results (Table 3) revealed that UDFF was strongly positively correlated with SWV, BMI, and ALT levels (r>0.7; P<0.01). A weak positive correlation was observed between UDFF and maternal age (0.3<r<0.5; P<0.05). No significant correlations were found between UDFF and TBA, T-Bil, PT, INR, or ALB (P>0.05).

Discussion

During pregnancy, the maternal body undergoes a series of complex physiological adaptations. Traditional methods of liver function assessment often fail to differentiate between physiological changes and pathological liver injury. Therefore, establishing trimester-specific reference values for hepatic UDFF and Auto-pSWE is of considerable clinical value in optimizing liver health monitoring during pregnancy. Given the dynamic nature of hepatic lipid metabolism across gestation, stratified reference values may assist in distinguishing normal physiological changes from early pathological alterations, thereby reducing the risk of misdiagnosis. This study used longitudinal data across multiple gestational weeks to establish reference ranges specific to pregnancy.

In this study, data from 30 healthy pregnant women were analyzed across trimesters. Results showed a statistically significant increase in BMI as gestation progressed. The mean BMI increased from 20.684±1.792 kg/m² in the first trimester to 22.524±1.805 kg/m² in the second trimester and further to 23.767±1.816 kg/m² in the third trimester. This trend is likely attributed to physiological changes such as increased fetal-placental mass, plasma volume expansion, interstitial fluid retention, enhanced fat storage, and hormonal modulation of metabolism. Numerous studies have demonstrated that dynamic changes in maternal BMI are closely related to fetal development and maternal health status (9). Abnormal BMI during pregnancy is associated with elevated risks of gestational diabetes mellitus and preeclampsia (10,11), highlighting its importance as a clinical monitoring index.

Our results indicated significant trimester-based differences in UDFF and Auto-pSWE parameters, which demonstrated an upward linear trend throughout pregnancy. UDFF increased from 1.92%±0.30% in the first trimester to 4.58%±1.48% in the third trimester—an increase of 139%. Concurrently, SWV values rose from 1.21±0.14 m/s (first trimester) to 1.47±0.18 m/s (third trimester). Liver stiffness (Young modulus) values exhibited a similar increasing pattern, with statistically significant intergroup differences. These findings suggest that hepatic fat content and tissue stiffness progressively increase during gestation, especially in the late trimester.

Several physiological mechanisms may underlie these changes, including increased levels of progesterone and estrogen (which promote collagen deposition), establishment of the uteroplacental circulation (leading to increased portal vein flow), enhanced lipid metabolism, and hepatic cell hypertrophy (12,13), along with known determinants of liver stiffness—such as fluid overload, cardiac output, body weight, and intra-abdominal pressure. Mechanical pressure from uterine enlargement in late pregnancy could further impact hepatic stiffness and fat deposition (14,15).

Multivariate linear regression analysis demonstrated that UDFF was significantly positively correlated with SWV, suggesting a potential synergy between hepatic fat accumulation and tissue stiffening. Possible related mechanisms include hepatic cell hypertrophy increasing tissue tension, triglyceride-mediated changes to the extracellular matrix, and lipotoxicity-induced Kupffer cell activation promoting collagen synthesis. Moreover, BMI had a strong positive correlation with UDFF, underscoring the impact of adiposity on hepatic metabolism. A significant correlation was also observed between liver stiffness and TBA levels, consistent with previous findings. For example, Kubota et al. reported that elevated bile acids during pregnancy may activate hepatic stellate cells through the FXR signaling pathway, thereby altering liver mechanical properties (16). A moderate correlation between elasticity and INR suggests that an increased hepatic metabolic demand for coagulation factor synthesis may influence tissue elasticity.

Conversely, several biochemical markers traditionally associated with liver function showed weak or no significant correlation with UDFF and Auto-pSWE parameters. T-Bil levels remained relatively stable throughout pregnancy and did not correlate significantly with liver fat or stiffness. This stability may be explained by the upregulation of hepatic glucuronosyltransferase enzymes such as UGT1A1 enhancing bilirubin conjugation and clearance during gestation (17). Coagulation markers including PT and INR also were not significantly correlated with hepatic stiffness or fat content. Pregnancy is characterized by a procoagulant state with increased synthesis of various coagulation factors, with PT and INR being maintained within normal ranges despite underlying hepatic adaptations (18). Similarly, serum ALB concentrations decreased modestly but were not associated with imaging findings; this is likely due to physiological plasma volume expansion causing dilutional hypoalbuminemia rather than impaired hepatic synthetic function (19). Thus, these conventional biochemical tests may be insufficient for reflecting subtle changes in liver structure or function during pregnancy.

UDFF is an emerging noninvasive ultrasound-based technique for quantifying hepatic fat content, offering a promising alternative to MRI proton density fat fraction (MRI-PDFF) and liver biopsy in diagnosing NAFLD and metabolic dysfunction-associated fatty liver disease (20,21). UDFF measures the attenuation coefficient and backscatter coefficient to estimate liver fat percentage, leveraging the distinct acoustic impedance of fat versus that of normal hepatocytes (4,22). Multiple studies have validated the accuracy, repeatability, and clinical utility of UDFF. For instance, Hall et al. reported a strong correlation between UDFF and MRI-PDFF in patients with metabolic liver disease (4,6). In comparative studies, UDFF outperformed conventional ultrasound and computed tomography in stratifying liver steatosis severity, offering enhanced sensitivity and reproducibility (3). These findings highlight the potential of UDFF for individualized and precision liver assessment.

Similarly, ultrasound elastography has become an essential noninvasive tool for evaluating liver stiffness, with applications in staging fibrosis, monitoring therapeutic responses, differentiating focal hepatic lesions, and assessing portal hypertension. In this study, we observed a progressive increase in liver stiffness in healthy pregnant women, particularly in the third trimester. These findings align with prior research suggesting that pregnancy-associated hemodynamic and hormonal changes may transiently alter hepatic mechanical properties (15,23). The establishment of gestation-specific reference values in our study may constitute valuable normative data for clinical interpretation.

Although our study provides preliminary trimester-specific reference values for UDFF and Auto-pSWE in healthy singleton pregnancies, several limitations should be acknowledged. The small sample size (n=30) and single-center design limit statistical precision and external validity, and these trimester-specific reference values for UDFF and Auto-pSWE should only be considered preliminary. Moreover, because the cohort included only healthy singleton pregnancies with normal BMI, the findings may not generalize to women with different BMI ranges, multiple gestations, or underlying health conditions. The limited sample also restricted evaluation of potential confounders such as maternal age, BMI, and comorbidities, and these should be investigated to establish more robust and broadly applicable reference intervals. In addition, the absence of postpartum follow-up precluded assessment of whether changes in UDFF and liver stiffness are reversible after delivery. Larger, multicenter studies with more diverse populations and postpartum follow-up are needed to refine these reference values and confirm their clinical utility.

To our knowledge, no previous study has established trimester-specific reference values for UDFF and Auto-pSWE in healthy singleton pregnancies. Other research has largely evaluated liver stiffness with transient elastography or two-dimensional SWE, examined only late gestation or pregnancy-related liver disorders, and lacked trimester-stratified data in healthy populations. Data on hepatic fat quantification during pregnancy is scarce, antenatal MRI-based fat mapping has not been extensively applied, and UDFF studies have primarily involved nonpregnant cohorts. In this context, our findings uniquely provide preliminary trimester-specific reference ranges that may inform point-of-care interpretation in obstetric practice, although validation in larger, multicenter studies with postpartum follow-up is required.

Conclusions

The data generated from this study provide practical benchmarks for noninvasive liver health monitoring during pregnancy. UDFF and Auto-pSWE are real-time, efficient, and well-suited tools for routine obstetric assessment. They can detect early subclinical changes in liver fat and stiffness, helping differentiate between normal physiological adaptations and pathological conditions. The trimester-specific reference values established in this study offer a clearer understanding of normal hepatic changes and may help healthcare providers to identify deviations that may signal underlying liver conditions. By facilitating early detection, these biomarkers can guide timely interventions, optimizing both maternal and fetal health outcomes.

Acknowledgments

None.

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at https://qims.amegroups.com/article/view/10.21037/qims-2025-1264/rc

Data Sharing Statement: Available at https://qims.amegroups.com/article/view/10.21037/qims-2025-1264/dss

Funding: The study was supported by the Gansu Province Youth Talent Cultivation Program (No. GSWSQNPY-2024-05).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://qims.amegroups.com/article/view/10.21037/qims-2025-1264/coif). All authors report the funding from the Gansu Province Youth Talent Cultivation Program (No. GSWSQNPY-2024-05). The authors have no other conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. This study was conducted in accordance with the Declaration of Helsinki and its subsequent amendments. Ethical approval has been obtained from the Institutional Review Board (IRB) at Gansu Provincial Maternity and Child-care Hospital (Gansu Provincial Central Hospital) (No. 2022-66). All participants provided written informed consent prior to participation in the study.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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