Maternal plasma concentrations of angiogenic/antiangiogenic factors in the third trimester of pregnancy to identify the patient at risk for stillbirth at or near term and severe late preeclampsia




Objective


To determine whether maternal plasma concentrations of placental growth factor (PlGF), soluble endoglin (sEng), and soluble vascular endothelial growth factor receptor-1 (sVEGFR-1) at 30-34 weeks of gestation can identify patients at risk for stillbirth, late preeclampsia, and delivery of small-for-gestational-age (SGA) neonates.


Study Design


A prospective cohort study included 1269 singleton pregnant women from whom blood samples were obtained at 30-34 weeks of gestation and who delivered at >34 weeks of gestation. Plasma concentrations of PlGF, sEng, and sVEGFR-1 were determined by enzyme-linked immunosorbent assay.


Results


The prevalence of late (>34 weeks of gestation) preeclampsia, severe late preeclampsia, stillbirth, and SGA was 3.2% (n = 40), 1.8% (n = 23), 0.4% (n = 5), and 8.5% (n = 108), respectively. A plasma concentration of PlGF/sEng <0.3 MoM was associated with severe late preeclampsia (adjusted odds ratio, 16); the addition of PlGF/sEng to clinical risk factors increased the area under the receiver-operating characteristic curve from 0.76 to 0.88 ( P = .03). The ratio of PlGF/sEng or PlGF/sVEGFR-1 in the third trimester outperformed those obtained in the first or second trimester and uterine artery Doppler velocimetry at 20-25 weeks of gestation for the prediction of severe late preeclampsia (comparison of areas under the receiver-operating characteristic curve; each P ≤ .02). Both PlGF/sEng and PlGF/sVEGFR-1 ratios achieved a sensitivity of 74% with a fixed false-positive rate of 15% for the identification of severe late preeclampsia. A plasma concentration of PlGF/sVEGFR-1 <0.12 MoM at 30-34 weeks of gestation had a sensitivity of 80%, a specificity of 94%, and a likelihood ratio of a positive test of 14 for the identification of subsequent stillbirth. Similar findings (sensitivity 80%; specificity 93%) were observed in a separate case-control study.


Conclusion


Risk assessment for stillbirth and severe late preeclampsia in the third trimester is possible with the determination of maternal plasma concentrations of angiogenic and antiangiogenic factors at 30-34 weeks of gestation.


Preeclampsia, a leading cause of maternal and perinatal morbidity/mortality worldwide, affects 2-8% of all pregnancies and has a complex pathophysiology that may include abnormal physiologic transformation of the spiral arteries, intravascular inflammation, endothelial cell dysfunction, excessive thrombin generation, oxidative stress, and/or an antiangiogenic state.




For Editors’ Commentary, see Contents



Preeclampsia may be classified as “early” or “late” according to gestational age at diagnosis or delivery. The gestational age cutoff most frequently used is 34 weeks. Early preeclampsia is associated with multisystemic involvement, a higher frequency of small-for-gestational-age (SGA) neonates, and placental vascular lesions of underperfusion. Because early preeclampsia is a frequent indication for preterm delivery, the condition is also associated with a higher rate of neonatal morbidity. In contrast, preeclampsia at term is associated with better neonatal outcomes than preterm preeclampsia. Although emphasis has focused on early preeclampsia, most cases of preeclampsia occur at or near term. Consequently, late preeclampsia accounts for a substantial proportion of medically indicated late preterm births and severe maternal morbidity, which includes most cases of eclampsia, the form of the disease that accounts for most maternal deaths. Hence, the identification of predictors of late preeclampsia is a healthcare priority.


Stillbirth, another obstetrical syndrome which may or may not be related to preeclampsia, affects >3 million women in the third trimester worldwide each year. The circumstances surrounding stillbirth vary depending on socioeconomic conditions. In high-income countries, stillbirth is associated with fetal growth restriction or placental insufficiency; however, in nearly one-half of the cases, the cause is unknown. Intrapartum complications, preeclampsia, and infection play a more important role in the cause of stillbirth in low-income countries. Currently, there is no effective risk assessment tool for the detection of stillbirth at or near term.


An imbalance of angiogenic/antiangiogenic factors has been implicated in the pathophysiologic description of preeclampsia, pregnancies with SGA neonates, stillbirth, and other obstetrical complications. Changes in the concentrations of the angiogenic factor (placental growth factor [PlGF]) and antiangiogenic factors (soluble vascular endothelial growth factor receptor-1 [sVEGFR-1; also known as soluble fms-like tyrosine kinase-1] and soluble endoglin [sEng]) in maternal circulation precede the clinical diagnosis of preeclampsia, SGA, and stillbirth. Most studies that examined the value of these biomarkers, however, have focused on the prediction of preeclampsia during the first or second trimesters. The results of such studies largely suggest that an imbalance between angiogenic and antiangiogenic factors increases the likelihood of preterm preeclampsia at a higher magnitude than that of term preeclampsia. Yet, not all studies have arrived at the same conclusion. Thus far, no cohort study has evaluated the predictive performance of these biomarkers in the third trimester for the identification of patients at risk for stillbirth at or near term or late-onset preeclampsia.


Recently, a new approach in screening for adverse pregnancy outcomes proposes to focus on the prevention of pregnancy complications at term. Such an approach would identify the more prevalent disease (eg, preeclampsia at term), and predictive models could be applied to low-income settings, where most maternal and perinatal deaths occur.


The objective of this study was to determine whether maternal plasma concentrations of PlGF, sEng, sVEGFR-1, and their ratios at 30-34 weeks of gestation could be used to identify patients at risk for stillbirth, late preeclampsia, severe late preeclampsia, or delivery of SGA neonates.


Methods


Study design


We first designed a cohort study of women who had a venipuncture at 30-34 weeks of gestation and outcome data to examine the value of PlGF, sVEGFR-1, and sEng in the identification of patients who subsequently experienced late preeclampsia, severe late preeclampsia, stillbirth, and SGA. Subsequent to this cohort study, a case-control study was performed to determine whether these biomarkers and their ratios could identify patients at risk for stillbirth at or near term in a different population.


Cohort study


A prospective longitudinal cohort study was conducted between November 2003 and August 2006 to identify biological markers for the prediction of preeclampsia, SGA neonates, and stillbirth. Patients were enrolled in the prenatal clinic of the Sotero del Rio Hospital, a tertiary care center in Santiago, Chile, and followed until delivery. Inclusion criteria were singleton gestation and 6-22 weeks of gestation. Exclusion criteria were (1) preterm labor, preterm prelabor rupture of membranes, preeclampsia, or impaired fetal growth at the time of recruitment; (2) known major fetal anomaly or fetal death; (3) active vaginal bleeding; and (4) serious medical illness (renal insufficiency, congestive heart disease, chronic respiratory insufficiency, or active hepatitis). At enrollment and each subsequent visit, patients underwent a venipuncture for the collection of maternal blood. The protocol consisted of collecting samples every 4 weeks until 24 weeks of gestation and every 2 weeks thereafter until delivery.


We previously reported the predictive performance of angiogenic/antiangiogenic factors at 6-15 and 20-25 weeks of gestation and uterine artery Doppler velocimetry (UADV) at 20-25 weeks of gestation for preeclampsia in this cohort. In summary, 2998 consecutive women were enrolled during the study period; 2495 women had a plasma sample collected in early pregnancy. Of those, an additional plasma sample was obtained in the mid trimester from 1917 women. Subsequently, an additional 204 patients without results of UADV in the second trimester were excluded. Ninety-one patients were lost to follow up; the remaining 1622 patients had been included in a previous article that examined the role of angiogenic/antiangiogenic factors at 6-15 and 20-25 weeks of gestation. The current study involved a subset of this cohort that excluded patients who delivered at ≤34 weeks of gestation (n = 27) and those who did not have a plasma sample collected at 30-34 weeks of gestation (n = 326) to examine the role of angiogenic/antiangiogenic factors at 30-34 weeks for the identification of adverse pregnancy outcomes at >34 weeks of gestation.


All women provided written informed consent before participating in the study. The use of clinical and ultrasound data and the collection and use of maternal blood for research purposes was approved by the institutional review boards of the Sotero del Rio Hospital, Santiago, Chile (an affiliate of the Pontificia Catholic University of Santiago, Chile), and the Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Department of Health and Human Services.


Outcomes of the study


The outcomes of the study included late preeclampsia, severe late preeclampsia, SGA without preeclampsia, and stillbirth. Preeclampsia was defined as new-onset hypertension that developed at >20 weeks of gestation and proteinuria. Hypertension was defined as systolic ≥140 mm Hg and/or diastolic blood pressure ≥90 mm Hg that was measured at 2 occasions, 4 hours to 1 week apart. Proteinuria was defined as a urine protein of ≥300 mg in a 24-hour urine collection or 2 random urine specimens that were obtained 4 hours to 1 week apart and showed ≥1+ by dipstick or 1 dipstick that demonstrated ≥2+ protein. Late preeclampsia was defined as patients with preeclampsia who delivered at >34 weeks of gestation. Severe preeclampsia was diagnosed based on criteria of the American College of Obstetricians and Gynecologists. SGA was defined as a birthweight of <10th percentile for gestational age according to the Chilean birthweight distribution of a Hispanic population. Stillbirth was defined as death of a fetus before delivery that was not a consequence of an induced termination of pregnancy (including intrapartum and antepartum stillbirth). Abnormal UADV was defined as a mean uterine artery Doppler pulsatility index of >1.45.


Data quality


Customized case report forms and a perinatal database were generated. Data were extracted from medical records by trained research nurses. To account for misclassification, abstracters were trained, the data collection methods were verified, and data logic was monitored. Cases of uncertainty were resolved by iterative discussion among 3 of the authors. Gestational age at venipuncture and at delivery was based on best obstetrical estimates with the use of the last menstrual period and the earliest fetal biometric parameters, which were performed at ≤20 weeks of gestation in 98.2% of cases.


Sample collection and immunoassays


Blood was obtained by venipuncture and collected into tubes that contained ethylenediaminetetraacetic acid. Samples were centrifuged and stored at −70°C. Maternal plasma concentrations of sVEGFR-1, PlGF, and sEng were determined by sensitive and specific immunoassays (R&D Systems, Minneapolis, MN) as previously described. The inter- and intra-assay coefficients of variation (CV) were 1.4% and 3.9% for sVEGFR-1, 2.3% and 4.6% for sEng, and 6.02% and 4.8% for PlGF. The sensitivity of the assays was 16.97 pg/mL for sVEGFR-1, 0.08 ng/mL for sEng, and 9.52 pg/mL for PlGF. The laboratory personnel who performed the assays were blinded to the clinical information.


Statistical analysis


Differences in distributions of dichotomous and categorical variables were tested with the χ 2 test or Fisher exact test where appropriate; continuous parameters were compared by analysis of variance or Friedman’s 2-way nonparametric analysis of variance test with Bonferroni correction for multiple comparisons, depending on the distribution of data. Normality was assessed with the Kolmogorov-Smirnov test and visual plot inspection.


Quantile regression was used to calculate median analyte ratio concentrations (PlGF/sVEGFR-1, PlGF/sEng) that were conditional on gestational age among uncomplicated pregnancies (n = 886). Multiple of the median (MoM) values were calculated for both analyte ratios for each patient. MoM cutoffs were determined based on inspection of receiver operating characteristic curves that were calculated for each outcome (stillbirth, late preeclampsia, severe late preeclampsia, and SGA without preeclampsia). Prognostic logistic regression models were constructed for each outcome. Covariables included in adjusted models were selected on the basis of clinical knowledge. Model reduction was performed additionally based on the plausibility of regression coefficients, association with independent variables, and the magnitude of change in the main-effect parameter estimates. To account for potential model over-fitting, when van Houwelingen and le Cessie’s heuristic shrinkage estimator was below 0.85 (indicator of instability), bootstrap estimated linear shrinkage factors and Firth’s penalized maximum likelihood estimation were used to calculate conservative estimates that were less likely to be affected by over-fitting.


Predictive performance metrics were also calculated for each outcome. Paired sample nonparametric statistical techniques were used to compare area under the receiver operating characteristic curves (AUC) of models that were constructed with logistic regression for the identification of selected pregnancy outcomes. A McNemar’s test was also used to test for differences in sensitivity at a fixed false-positive rate of 15%. A 5% threshold for type I error was used to determine statistical significance. Statistical analyses were performed with SAS software (version 9.3; Cary, NC).


Case-control study for stillbirth


Participants were identified from a cohort of 5828 singleton pregnancies who were either enrolled in a similar longitudinal protocol to that used in the Chilean cohort or another cross-sectional protocol from 2007-2009 at Hutzel Women’s Hospital, Detroit, MI. Stillbirth was defined as death of a fetus before delivery (which was not a consequence of an induced termination of pregnancy). In the longitudinal study, plasma samples were obtained from the first or early second trimester and at the time of each prenatal visit (scheduled every 4 weeks until 24 weeks and every 2 weeks thereafter until delivery). In the cross-sectional study, patients were enrolled when they presented to the labor and delivery unit with a suspicion of spontaneous preterm labor or medically-indicated preterm birth. Among 31 cases of stillbirth at ≥34 weeks of gestation, 5 cases were included because they had a plasma sample collected at 30-34 weeks of gestation. Control subjects were identified from uncomplicated pregnancies who both delivered an appropriate weight for gestational age neonate at term and had a plasma sample collected at 30-34 weeks of gestation. Control subjects were matched to cases at a ratio of 6 to 1 on gestational age at venipuncture, parity, ethnicity, tobacco use, and body mass index. Maternal plasma concentrations of sVEGFR-1, sEng, and PlGF were determined by sensitive and specific immunoassays (R&D Systems) similar to those used in the Chilean cohort, as described earlier.


All women provided written informed consent before participating in the study. The use of clinical and ultrasound data and collection and utilization of maternal blood for research purposes was approved by the institutional review boards of the Wayne State University, Detroit, MI, and the Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Department of Health and Human Services.


Differences among cases and control subjects were tested with the use of χ 2 , Fisher exact, or Mann-Whitney U tests where appropriate. AUC was calculated, and sensitivities and specificities were determined with the use of absolute value thresholds for each biomarker ratio that were derived from the inspection of receiver operating characteristic curves.




Results


The cohort study included 1269 pregnant women ( Figure 1 ). The prevalence of late preeclampsia, severe late preeclampsia, stillbirth, and SGA without preeclampsia was 3.2% (n = 40), 1.8% (n = 23); 0.4% (n = 5), and 8.5% (n = 108), respectively. Among 23 patients who received a diagnosis of severe preeclampsia, 6 experienced severe high blood pressure and severe proteinuria; 4 had severe high blood pressure; 4 had severe high blood pressure with severe proteinuria with SGA fetuses; 3 had SGA fetuses; 2 had severe headache with severe proteinuria; 2 had severe proteinuria, 1 had severe high blood pressure, and 1 had severe proteinuria and pulmonary edema. Table 1 displays the demographic and obstetrical characteristics of patients with SGA neonates, preeclampsia, stillbirth, other complications and of patients without any of these complications (uncomplicated pregnancy). There were no significant differences in the mean gestational age at venous sampling or mean duration of sample storage among the 4 groups. The distribution of baseline characteristics did not significantly differ between patients included in the current study compared with the overall cohort (data not shown). Similarly, there were no significant differences in the risk of stillbirth or SGA between the entire cohort and subcohort. However, by design, participants in the subcohort were more likely to deliver at >34 weeks of gestation. Patients in this subcohort had a lower frequency of preeclampsia than those in the entire cohort (3.2% vs 4.8%; P = .03). Three patients who had a diagnosis of gestational hypertension before venipuncture at 30-34 weeks of gestation; however, subsequent preeclampsia did not develop. The median MoM plasma concentration of PlGF/sVEGFR-1 and PlGF/sEng was significantly lower in patients with subsequent stillbirth, preeclampsia, and SGA than in patients without these conditions ( P < .05 for each comparison; Table 1 ).




FIGURE 1


Flow diagram of patients enrolled in the study

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.


TABLE 1

Demographical and obstetrical characteristics of the study population































































































Patient characteristics Uncomplicated pregnancy (n = 886) Pregnancy with small-for-gestational-age neonate (n = 108) Preeclampsia (n = 40) Stillbirth (n = 5) Other complications (n = 230)
Maternal age, y a 26.2 ± 5.9 26.5 ± 7.1 23.6 ± 5.4 b 27 ± 10 28 ± 6.5
Tobacco use, n (%) 93 (10.5) 20 (18.50) 5 (12.50) 0 28 (12.2)
Nulliparous, n (%) 356 (40.2) 48 (44.4) 28 (70) b 2 (40) 76 (33.0)
Multiparous with history of preeclampsia, n (%) 19 (2.1) 1 (0.9) 3 (7.5) b 0 14 (6.1)
Multiparous without history of preeclampsia, n (%) 511 (57.7) 59 (54.60) 9 (22.50) b 3 (60) 140 (60.9)
Body mass index, kg/m 2 a 24.6 ± 4.2 24 ± 4.3 27.4 ± 8 22.4 ± 1.6 27.3 ± 6
Gestational age at venipuncture, wk a 32.2 ± 1.1 32.2 ± 1.1 32.2 ± 1.2 32 ± 0.9 32.2 ± 1.2
Storage time, y a 6.8 ± 0.7 6.8 ± 0.8 6.9 ± 1.1 6.6 ± 0.6 6.8 ± 0.7
Gestational age at delivery, k a 39.6 ± 1.1 39.4 ± 1.1 38.48 ± 1.6 b 36.5 ± 2.3 b 38.6 ± 1.7
Birthweight, g a 3505 ± 399 2710 ± 230 b 3096 ± 550 b 2896 ± 642 3366 ± 521
Placental growth factor/soluble vascular endothelial growth factor receptor-1 (multiple of the median) c , d 1.00 (0.51–1.83) 0.53 (0.21–1.22) b 0.21 (0.08–0.50) b 0.08 (0.07–0.1) b 0.73 (0.33–1.27)
Placental growth factor/soluble endoglin (multiple of the median) c , e 1.00 (0.56–1.78) 0.59 (0.26–1.10) b 0.27 (0.11–0.63) b 0.18 (0.1–0.3) b 0.74 (0.35–1.18)

Other complications included spontaneous preterm delivery (3%; n = 38), chronic hypertension (2.2%; n = 28), gestational hypertension (6.8%; n = 86), gestational and pregestational diabetes mellitus (4.6%; n = 58), placental abruption (0.4%; n = 5), cholestasis of pregnancy (0.9%; n = 12), and placenta previa (0.2%; n = 3).

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.

a Data are given as mean ± SD;


b Indicated significant difference ( P < .05) compared with combined “other + uncomplicated pregnancy” categories;


c Data are given as median (interquartile range) that were calculated among uncomplicated pregnancies (n = 886) by quantile regression:


d Median = 1.8863+ (−0.0508 × gestational week);


e Median = 354.3280 + (−8.9791 × gestational week).



Table 2 displays the magnitude of association between abnormal biomarker profiles and late preeclampsia (overall and severe), delivery of SGA neonates (birthweight: <10%, <3%), and stillbirth. Patients with plasma PlGF/sEng or PlGF/sVEGFR-1 ratio concentrations of <0.3 MoM were significantly more likely to experience late preeclampsia (adjusted odds ratio [aOR], 7.1; 95% confidence interval [CI], 3.6–13.8; and 6.1; 95% CI, 3.1–11.8, respectively) and severe late preeclampsia (aOR, 16.1; 95% CI, 5.8–44.6 and 12.2; 95% CI, 4.6–32, respectively) than those with MoMs at or above the threshold ( Table 2 ). The likelihood ratio of a positive test and sensitivity for either PlGF/sEng or PlGF/sVEGFR-1 ranged from 4.5–4.8 and 74–78%, respectively; both had a specificity of 84% for the identification of patients with severe late preeclampsia ( Table 3 ).



TABLE 2

Likelihood of subsequent stillbirth, preeclampsia, and small-for-gestational-age neonate







































































































































Dependent variable Others (n/N) Outcome (n/N) Unadjusted Adjusted
Odds ratio a 95% CI Odds ratio a 95% CI
Preeclampsia (n = 40)
PlGF/sVEGFR-1 <0.3 MoM 199/1229 (16.2%) 23/40 (57.5%) 7.9 4.1–15.2 6.1 3.1–11.8
PlGF/sEng <0.3 MoM 196/1229 (15.9%) 24/40 (60.0%) 5.9 1.9–18.7 7.1 3.6–13.8
Severe preeclampsia (n = 23)
PlGF/sVEGFR-1 <0.3 MoM 205/1246 (16.5%) 17/23 (73.9%) 11.9 2.2–66.0 12.2 0.6–32.0
PlGF/sEng <0.3 MoM 202/1246 (16.3%) 18/23 (78.3%) 11.7 2.1–64.6 16.1 5.8–44.6
Small-for-gestational-age <10th percentile (n = 108)
PlGF/sVEGFR-1 <0.3 MoM 184/1161 (15.8%) 38/108 (35.2%) 4.2 2.4–7.3 3.0 2.0–4.7
PlGF/sEng <0.3 MoM 189/1161 (16.3%) 31/108 (28.7%) 3.6 2.0–6.4 2.0 1.3–3.1
Small-for-gestational-age <3rd percentile (n = 23)
PlGF/sVEGFR-1 <0.3 MoM 210/1246 (16.9%) 12/23 (52.2%) 6.9 2.4–19.4 5.5 2.3–13.1
PlGF/sEng <0.3 MoM 209/1246 (16.8%) 11/23 (47.8%) 7.0 2.5–19.8 4.4 1.8–10.4
Stillbirth (n = 5) b
PlGF/sVEGFR-1 <0.12 MoM 71/1264 (5.6%) 4/5 (80%) 20.1 4.8–84.3 23.1 5.6–95.4
PlGF/sEng <0.2 MoM 137/1264 (10.8%) 3/5 (60%) 8.4 2.0–35.1 9.1 2.2–37.2

Medians were calculated among uncomplicated pregnancies (n = 886) by quantile regression (PlGF/sVEGFR-1: median = 1.8863 + (−0.0508 × gestational week); PlGF/sEng: median = 354.3280 + (−8.9791 × gestational week); cutoffs were selected based on inspection of receiver operating characteristic curves. The prediction of preeclampsia and small-for-gestational-age was adjusted for maternal age (continuous), combined parity and history of preeclampsia, prepregnancy body mass index (continuous), and tobacco use. PlGF/sVEGFR-1 MoM cutoff: <0.12 (or 5th -6th percentile of uncomplicated pregnancies) for stillbirth and <0.3 (or 17th percentile of uncomplicated pregnancies) for preeclampsia and small-for-gestational-age. PlGF/sEng MoM cutoff: <0.2 (or 11th percentile of uncomplicated pregnancies) for stillbirth and <0.3 (or 17th percentile of uncomplicated pregnancies) for preeclampsia and small-for-gestational-age.

CI , confidence interval; MoM, multiples of the median; PlGF/sEng, placental growth factor/soluble endoglin; PlGF/sVEGFR-1, placental growth factor/soluble vascular endothelial growth factor receptor-1.

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.

a Data represent the likelihood of outcome in subjects with abnormal analyte ratio concentrations (above/below MoM cutoff) relative to patients with normal analyte ratio concentration multiples of the median cutoff;


b Odds ratio was adjusted for gestational age at venipuncture (continuous).



TABLE 3

Predictive performance of maternal plasma concentrations of angiogenic and antiangiogenic factors for stillbirth and preeclampsia screening
































































































































































Diagnostic performance metrics Preeclampsia (n = 40) Severe preeclampsia (n = 23) Stillbirth (n = 5)
Estimate 95% CI Estimate 95% CI Estimate 95% CI
PlGF/sVEGFR-1
Sensitivity, % 58 40–73 74 52–90 80 28–100
Specificity, % 83 82–86 84 81–86 94 93–96
Positive predictive value, % 10 6–15 8 5–12 5 1–13
Negative predictive value, % 98 97–99 99 99–100 100 99–100
False positive rate, % 16 14–18 16 14–19 6 4–7
False negative rate, % 43 27–59 26 10–48 20 0.5–72
Positive likelihood ratio 3.6 2.6–4.8 4.5 3.4–5.9 14.2 8.7–23.3
Negative likelihood ratio 0.5 0.4–0.7 0.3 0.2–0.6 0.2 0.04–1.22
PlGF/sEng
Sensitivity, % 60 43–75 78 56–93 60 15–95
Specificity, % 84 82–86 84 82–86 89 87–91
Positive predictive value, % 11 7–16 8 5–13 2 0.4–6
Negative predictive value, % 98 97–99 99 99–100 99 99–100
False positive rate, % 16 14–18 16 14–18 11 9–13
False negative rate, % 40 25–57 22 7–44 40 5–85
Positive likelihood ratio 3.8 2.8–5.0 4.8 3.8–6.2 5.5 2.7–11.5
Negative likelihood ratio 0.5 0.3–0.7 0.3 0.1–0.6 0.4 0.2–1.3

Medians were calculated among uncomplicated pregnancies (n = 886) by quantile regression (PlGF/sVEGFR-1: median = 1.8863 + (−0.0508 × gestational week); PlGF/sEng: median = 354.3280 + (−8.9791 × gestational week); cutoffs were selected based on inspection of receiver operating characteristic curves. PlGF/sVEGFR-1 multiples of the median cutoff: <0.12 (or 5th -6th percentile of uncomplicated pregnancies) for stillbirth and <0.3 (or 17th percentile of uncomplicated pregnancies) for preeclampsia; PlGF/sEng multiples of the median cutoff: <0.2 (or 11th percentile of uncomplicated pregnancies) for stillbirth and <0.3 (or 17th percentile of uncomplicated pregnancies) for preeclampsia.

CI, confidence interval; MoM, multiples of the median; PlGF/sEng, placental growth factor/soluble endoglin; PlGF/sVEGFR-1, placental growth factor/soluble vascular endothelial growth factor receptor-1.

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.


The addition of the PlGF/sEng or PlGF/sVEGFR-1 ratio to the clinical risk factors increased the AUC from 0.76 to 0.88 and 0.86, respectively, for the prediction of severe late preeclampsia ( P = .03 and .06). With a fixed false-positive rate of 15%, both the PlGF/sEng ratio and PlGF/sVEGFR-1 ratios achieved a sensitivity of 74% in predicting severe PE. These biomarkers in the third trimester outperformed those obtained previously at 6-15 and 20-25 weeks of gestation and UADV that was assessed at 20-25 weeks of gestation for the prediction of severe late preeclampsia (each P ≤ .02; Figure 2 ). Further, the addition of the PlGF/sVEGFR-1 or the PlGF/sEng ratio measured in the third trimester to clinical risk factors (age, body mass index, combined parity and history of preeclampsia, and tobacco use) yielded significantly greater sensitivity (74%) at a fixed false-positive rate of 15%, compared with a model that used the same biomarker ratios measured in the second trimester, clinical risk factors, and abnormal UADV values that were obtained at 20-25 weeks of gestation ( P = .008 and .03, respectively). The direction, magnitude, and significance of these associations also persisted during sensitivity analyses performed excluding patients with a history of preeclampsia (n = 37) based on their elevated a priori risk in the current pregnancy.




FIGURE 2


Comparison of ROC curves for the identification of severe late preeclampsia

Areas under the ROC curves achieved by plasma concentrations of A, placental growth factor ( PlGF )/soluble endoglin ( sEng ) ratio or B, PlGF/soluble vascular endothelial growth factor receptor-1 ( sVEGFR-1 ) ratio. Multiples of the median at 6-15, 20-25, and 30-34 weeks of gestation and uterine ( Ut. ) artery Doppler velocimetry at 20-25 weeks of gestation are shown.

GA , gestational age; ROC , receiver-operating characteristic.

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.


Although patients with plasma PlGF/sVEGFR-1 or PlGF/sEng ratio concentrations of <0.3 MoM were more likely to experience SGA without preeclampsia (aOR, 2-3; Table 2 ), the addition of these biomarkers to demographic/perinatal data did not improve the AUC (0.64 vs 0.62; P = .2 and .6; respectively). Subgroup analysis that focused on patients with severe SGA (birthweight less than the third percentile; n = 23) indicated that the adjusted odds ratio of patients with PlGF/sVEGFR-1 or PlGF/sEng ratio <0.3 MoM to experience severe SGA ranged from 4.4–5.5 ( Table 2 ). However, the addition of these biomarkers to clinical risk factors did not significantly improve the AUC ( P > .05).


Patients with a PlGF/sVEGFR-1 ratio of <0.12 MoM were significantly more likely to have a stillbirth than patients with a MoM ratio at or above the threshold (aOR, 23.1; 95% CI, 5.6–95.4). This cutoff had a sensitivity of 80%, a specificity of 94%, and a likelihood ratio of a positive test of 14.2 for the identification of a subsequent stillbirth at >34 weeks of gestation ( Table 3 ). Compared with a model that included only clinical data (maternal age, combined parity and history of preeclampsia, body mass index, and tobacco use), the addition of the PlGF/sVEGFR-1 ratio or the PlGF/sEng ratio to clinical data increased the AUC from 0.7 to 0.91 ( P = .03 and .06, respectively; Figure 3 ). The association between an abnormal ratio of angiogenic/antiangiogenic factors and stillbirth at or near term was also observed in the subsequent case-control study performed in a different population ( Table 4 ). A maternal plasma concentration of PlGF/sVEGFR-1 ratio ≤0.046 or PlGF/sEng ratio ≤11.7 pg/ng at 30-34 weeks had a sensitivity of 80% and a specificity of 93% for the identification of subsequent stillbirth ( Figure 4 ).




FIGURE 3


Comparison of ROC curves for the identification of stillbirth

Areas under the ROC curves achieved by plasma concentrations of placental growth factor ( PIGF )/soluble vascular endothelial growth factor receptor-1 ( sVEGFR-1 ) ratio or PIGF/soluble endoglin ( sEng ) ratio at 30-34 weeks’ gestation in combination with clinical data were compared with that achieved by clinical data alone.

GA , gestational age; ROC , receiver-operating characteristic.

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.


TABLE 4

Demographic and obstetrical characteristics of the population in the case-control study










































































Variable Control (n = 30) Fetal death (n = 5) P value
Maternal age, y a 21.5 (19.8–23.2) 26 (21.5–36.0) .154
Gestational age at venipuncture, wk a 32.9 (32.1–33.6) 33.4 (32–33.7) .493
Body mass index, kg/m 2 a 26.1 (21.1–35.8) 24.6 (17.9–44.5) .873
Nulliparity, n (%) 16 (53.3) 3 (60) .585
Tobacco use 0 0
African American 80% (24) 80% (4) .90
Gestational age at delivery, wk a 39.5 (38.9–40.6) 37.7 (34.7–38.9) .047
Birthweight, g a 3273 (3165–3478) 2305 (1635–3360) .030
Placental growth factor, pg/mL 646 (279–1108) 97 (63–640) .016
Soluble vascular endothelial growth factor receptor-1, pg/mL a 2779 (1822–4349) 6333 (3740–6908) .016
Soluble endoglin, ng/mL a 7.5 (5.7–10.1) 23.8 (14.4–33.4) .002
Placental growth factor/soluble vascular endothelial growth factor receptor-1 ratio a 0.25 (0.09–0.5) 0.02 (0.009–0.1) .002
Placental growth factor/soluble endoglin ratio, pg/ng a 96 (28–167) 6.9 (2.3–28) .002

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.

a Data are given as median (interquartile range).




FIGURE 4


ROC curves for the identification of subsequent stillbirth in the case-control study

Maternal plasma concentrations of placental growth factor ( PlGF )/soluble vascular endothelial growth factor receptor-1 ( sVEGFR-1 ) or PlGF/soluble endoglin ( sEng ) at 30-34 weeks of gestation were used to identify patients at risk of stillbirth. Area under the ROC curves = 0.91 for both biomarkers.

GA, gestational age; ROC , receiver-operating characteristic.

Chaiworapongsa. Third-trimester screening by angiogenic/antiangiogenic factors for preeclampsia and stillbirth. Am J Obstet Gynecol 2013.


Table 5 displays the obstetrical events at delivery, gestational age at venipuncture, and placental pathologic conditions of each patient with a stillbirth in the cohort and case-control studies. Among patients with a stillbirth in the cohort study, the interval from venipuncture to the diagnosis of stillbirth ranged from 2.2–6.1 weeks (median, 4.5 weeks). One patient was diagnosed with gestational diabetes mellitus, and another had an abruptio placentae. Three patients had histologic placental lesions consistent with maternal underperfusion, according to the criteria of the Society for Pediatric Pathology. Chronic chorioamnionitis and hyalinized avascular villi (consistent with fetal thrombotic vasculopathy), were observed in the other 2 cases. None of the cases included in the cohort study had a fetal autopsy. Of 5 cases included in the case-control study, 2 women had a diagnosis of diabetes mellitus and the remaining 3 had a diagnosis of severe preeclampsia, chronic hypertension, and Marfan’s syndrome, respectively. The interval from venipuncture to the diagnosis of stillbirth ranged from 2.4–5.4 weeks (median, 4 weeks). All 4 cases of stillbirth who had plasma concentrations of angiogenic/antiangiogenic factor ratios below the aforementioned cutoff in the case-control study also had lesions in the placenta suggestive of maternal underperfusion. Two stillbirths had a karyotype performed, and the results were 46 XY. Among 4 cases with available fetal autopsy results, 1 had lesions in the fetal brain consistent with acute hypoxic/ischemic damage in the grey matter.



TABLE 5

Obstetrics events at delivery, gestational age at venipuncture, and placental lesions of patients with stillbirth








































































































































Case Obstetrics events at delivery PlGF/sVEGFR-1 ratio, MoM Gestational age, wk Birthweight in grams (percentile) Placental lesions consistent with maternal underperfusion Placental lesions Fetal autopsy
At venous sampling At delivery
Cohort study
1 Normal blood pressure 0.07 32 1/7 34 3/7 2200 (50%) Yes Diffuse chronic villitis; persistent muscularization of basal plate arteries Not available
2 Gestational diabetes mellitus noncompliance with care 0.10 31 3/7 34 4/7 2280 (58%) Yes Increased syncytial knot Not available
3 Blood pressure 140/90 mm Hg, urine protein dipstick negative, placental abruption 0.04 31 35 4/7 3000 (92%) No Chronic chorioamnionitis Not available
4 Normal blood pressure, decreased fetal movement, thick meconium-stained amniotic fluid 0.08 33 3/7 39 3/7 3650 (74%) Yes Increased intervillous fibrin; prominent nucleated red blood cells, absence of physiologic change of the spiral arteries Not available
5 Normal blood pressure 0.78 32 2/7 38 3/7 3350 (60.5%) No Hyalinized avascular villi; fetal thrombotic vasculopathy Not available
Case-control study PlGF/sVEGFR-1 ratio, pg/pg
1 Gestational diabetes mellitus class A2: poorly controlled glucose 0.02 33 5/7 37 5/7 3620 (81.5%) Yes Microscopic chorionic pseudo cysts in placental membranes No congenital anomalies; no cause found
2 Pregestational diabetes mellitus class B: poorly controlled glucose 0.04 33 5/7 39 1/7 3100 (28.1%) Yes Recent villous infarction; persistent muscularization of basal plate arteries Acute hypoxic/ischemic gray matter damage and subarachnoid hemorrhage; no congenital anomalies
3 Severe preeclampsia 0.01 31 3/7 34 2/7 2040 (15%) Yes Recent villous infarction No congenital anomalies; no cause found
4 Chronic hypertension 0.005 32 4/7 35 1231 (1%) Yes Remote villous infarction; increased syncytial knots Not available
5 Marfan’s syndrome 0.19 33 3/7 38 4/7 2305 (1%) No Normal No congenital anomalies; no cause found

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May 13, 2017 | Posted by in GYNECOLOGY | Comments Off on Maternal plasma concentrations of angiogenic/antiangiogenic factors in the third trimester of pregnancy to identify the patient at risk for stillbirth at or near term and severe late preeclampsia
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