Objective
The purpose of this study was to examine the relationship between second-trimester maternal serum biomarkers and the development of early- and late-onset severe preeclampsia in euploid pregnancies.
Study Design
Included were 136,139 pregnancies that obtained second-trimester prenatal screening through the California Prenatal Screening Program with live births in 2006-2008. We identified severe preeclampsia diagnoses from hospital discharge records. We used log binomial regression to examine the association between abnormal second-trimester maternal serum biomarkers and the development of severe preeclampsia.
Results
Approximately 0.9% of all women (n = 1208) in our sample experienced severe preeclampsia; 329 women at <34 weeks’ gestation and 879 women ≥34 weeks’ gestation. High levels of alpha fetoprotein (AFP), human chorionic gonadotropin, inhibin (multiple of the median, ≥95th percentile), and low unconjugated estriol (multiple of the median, ≤5th percentile), were associated with severe preeclampsia (relative risk, 2.5-11.7). Biomarkers were more predictive of early-onset severe preeclampsia (relative risk, 3.8-11.7). One in 9.5 pregnancies with combined high AFP, inhibin, and low unconjugated estriol levels experienced severe early-onset preeclampsia compared with 1 in 680.5 pregnancies without any abnormal biomarkers.
Conclusion
The risk of the development of severe preeclampsia increases for women with high second-trimester AFP, human chorionic gonadotropin, inhibin, and/or low unconjugated estriol; this is especially true for early-onset severe preeclampsia. When abnormal biomarkers co-occur, risk dramatically increases. Although the screening value of second-trimester biomarkers is low, abnormal biomarkers, especially when occurring in combination, appear to indicate placental dysfunction that is associated with the development of severe preeclampsia.
Abnormal maternal serum analytes that were obtained for the purpose of prenatal screening for fetal anomalies are associated with adverse pregnancy outcomes; this is particularly true when their values are at extreme levels. Preeclampsia, a placental-based disease, is one such adverse pregnancy outcome. Preeclampsia occurs in approximately 3-5% of births; most cases occur at term. Approximately 10% of preeclampsia disorders have early-onset disease, which is defined as occurring at <34 weeks’ gestation. Although early-onset preeclampsia represents the minority of cases, it is associated more closely with significant maternal and neonatal morbidity and mortality rates.
Although routine markers may be useful in the identification of women whose pregnancies are at increased risk for severe preeclampsia, the identification of those who experience early-onset severe preeclampsia potentially could impact maternal and fetal outcomes. A few studies routinely have used collected maternal serum analytes to identify pregnancies at increased risk for severe preeclampsia while also differentiating between early and late onset disease. However these studies have tended to be limited by small sample size (n <460 pregnancies).
We examined the association between routinely collected second-trimester maternal serum analytes (alpha fetoprotein [AFP], human chorionic gonadotropin [hCG], unconjugated estriol [uE3], inhibin) and the development of early- and late-onset severe preeclampsia in a population-based sample.
Materials and Methods
We included women with singleton pregnancies who underwent second-trimester prenatal screening through the California Prenatal Screening Program within the Genetic Disease Screening Program at the California Department of Public Health with live births in 2006 through 2008 for whom there were linked maternal and baby outcome data available from the Office of Statewide Health Planning and Development hospital discharge records. We excluded pregnancies with Genetic Disease Screening Program records (prenatal screening records, newborn infant screening records, and chromosomal and neural tube defect registries) that indicated a chromosomal or neural tube defect. Severe preeclampsia diagnosis was based on International Classification of Diseases, 9th Revision, Clinical Modifications (ICD-9-CM) code 642.5, which defines severe preeclampsia as hypertension in pregnancy, childbirth, or puerperium, not specified as preexisting, with albuminuria, edema (or both) characterized as severe. Control subjects had no severe preeclampsia or any other preeclampsia disorder (ICD-9-CM code 642.4 [mild preeclampsia] or 642.6 [eclampsia]). Early-onset was defined as severe preeclampsia and delivery at <34 weeks’ gestation or delivery in gestational week 34 with hospitalization at <34 weeks. Late-onset was defined as severe preeclampsia and delivery in gestational week 34 without continuous hospitalization at <34 weeks’ gestation or delivery at >34 weeks’ gestation.
Second-trimester maternal blood samples were collected from 15-20 completed weeks’ gestation and were sent to California state-designated regional laboratories for serum testing of AFP, hCG, uE3, and inhibin levels. Regional laboratories all adhered to the same protocols for measuring these analytes with fully automated equipment (Auto DELFIA; Perkin Elmer Life Sciences, Waltham, MA). Analyte levels were reported directly into the state database along with patient information. Information provided by the regional laboratories was used to convert the analyte values into a multiple of the median (MoM) that was used for interpretation of the final result. All women in our sample had AFP, hCG, uE3, and inhibin level MoMs adjusted for gestational age, maternal weight, smoking status, preexisting diabetes mellitus, and race/ethnicity.
We obtained hospital discharge records for cases with severe preeclampsia diagnoses and control subjects. We obtained race/ethnicity, age, weight, and smoking variables from prenatal screening records and diabetic status from hospital discharge diagnoses (ICD-9-CM code 648.0 for preexisting diabetes mellitus, 648.8 for gestational diabetes mellitus). We did not have the date of diagnosis of preeclampsia in the hospital discharge records. Because the standard of care is to deliver patients who experience severe preeclampsia, we used the gestation of delivery as indicator of early and late onset.
The analyses used logistic binomial regression methods to estimate relative risks (RRs) of developing early- and late-onset severe preeclampsia in pregnancies with abnormal levels of second-trimester AFP, hCG, inhibin, and/or uE3 relative to pregnancies without any marker abnormalities. A biomarker was considered abnormally high if the MoM was ≥95th percentile and abnormally low if the MoM was ≤5th percentile. Pregnancies with normal biomarkers were considered to be those who had all of the associated MoMs between the 5th and 95th percentiles. Biomarker analyses controlled for the maternal characteristics that were found to be significantly different in those who experienced severe preeclampsia vs those who did not. The performance of biomarkers that were found to be significantly predictive of early- or late-onset severe preeclampsia (considered in isolation and in combination) was tested with sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) statistics.
All analyses were performed with Statistical Analysis Software (version 9.3; SAS Institute Inc, Cary, NC). Methods and protocols for the study were approved by the Committee for the Protection of Human Subjects within the Health and Human Services Agency of the State of California and the Institutional Review Board of the University of California, Davis.
Results
A total of 136,139 pregnancies met entry criteria for evaluation of which 1208 pregnancies (0.9%) were classifiable as cases having severe preeclampsia or control subjects (n = 134,931). Early-onset and late-onset preeclampsia developed in 329 (0.2%) and 879 (0.7%) of all women. Maternal demographics that were associated with an increased risk for early- and late-onset severe preeclampsia included black race/ethnicity and diabetes mellitus (any, preexisting, and gestational; RR, 1.5-6.9). Hispanic race/ethnicity, maternal age ≤17 or ≥35 years and weight at testing >the 95th percentile (by race/ethnicity at gestational age at testing) were associated with an increased risk for late-onset preeclampsia only (RR, 1.2-2.1; Table 1 ).
| Maternal characteristic | Severe preeclampsia | ||||
|---|---|---|---|---|---|
| No preeclampsia or eclampsia a | Early onset | Late onset | |||
| n (%) | n (%) | Relative risk (95% CI) | n (%) | Relative risk (95% CI) | |
| Sample | 134,931 (100.0) | 319 (100.0) | 889 (100.0) | ||
| Race/ethnicity | |||||
| White, not Hispanic | 36,738 (27.2) | 79 (24.8) | 219 (24.6) | ||
| Reference | |||||
| Hispanic | 77,476 (57.4) | 198 (62.1) | 554 (62.3) | ||
| 1.2 (0.9–1.5) | 1.2 (1.0–1.4) b | ||||
| Black | 6,806 (5.0) | 30 (9.4) | 2.0 (1.3–3.1) c | 69 (7.8) | 1.7 (1.3–2.2) c |
| Asian | 9,605 (7.1) | 4 (1.3) | 0.2 (0.1–0.5) d | 31 (3.5) | 0.5 (0.4–0.8) d |
| Other e | 4,306 (3.2) | 8 (2.5) | 0.9 (0.4–1.8) | 16 (1.8) | 0.6 (0.4–1.0) |
| Age, y | |||||
| ≤17 | 2,272 (1.7) | 2 (0.6) | 0.4 (0.1–1.5) | 29 (3.3) | 2.0 (1.4–2.9) c |
| 18-34 | 109,225 (81.0) | 256 (80.3) | 681 (76.6) | ||
| Reference | |||||
| ≥35 | 23,434 (17.4) | 61 (19.1) | 1.1 (0.8–1.5) | 179 (20.1) | 1.2 (1.0–1.4) b |
| Weight f | |||||
| <5th percentile | 6259 (4.6) | 13 (4.1) | 0.9 (0.5–1.6) | 46 (5.2) | 1.2 (0.9–1.6) |
| 5th-95th percentile | 121,699 (90.2) | 284 (89.0) | 767 (86.3) | ||
| Reference | |||||
| >95th percentile | 6973 (5.2) | 22 (6.9) | 1.4 (0.9–2.1) | 76 (8.6) | 1.7 (1.4–2.2) c |
| Diabetes mellitus | |||||
| No | 124,617 (92.4) | 274 (85.9) | 757 (85.2) | ||
| Reference | |||||
| Yes | 10,314 (7.6) | 45 (14.1) | 2.0 (1.4–2.7) c | 132 (14.9) | 2.1 (1.7–2.5) c |
| Pregestational | 1049 (0.8) | 12 (3.8) | 5.2 (2.9–9.2) c | 45 (5.1) | 6.8 (5.1–9.1) c |
| Gestational | 9265 (6.9) | 33 (10.3) | 1.7 (1.1–2.3) d | 87 (9.8) | 1.5 (1.2–1.9) c |
| Smoked | |||||
| No | 132,847 (98.5) | 318 (99.7) | 876 (98.5) | ||
| Reference | |||||
| Yes | 2084 (1.5) | 1 (0.3) | 0.2 (0.0–1.4) | 13 (1.5) | 10.0 (0.5–1.6) |
a No mild or severe preeclampsia or eclampsia
e Includes Asian East Indian, Pacific Islander, Native American, Middle Eastern, other race/ethnicity, and unknown race/ethnicity
f Percentile by race/ethnicity at gestational age at testing.
Single factor biomarker models for severe preeclampsia indicated an increased risk for early- and late-onset severe preeclampsia among pregnancies with AFP, hCG, and inhibin MoMs ≥95th percentile or a uE3 MoM ≤5th percentile (RR, 2.5-11.7; Table 2 ). Pregnancies with any of the at-risk biomarkers (elevated AFP, hCG, inhibin, and/or low uE3 levels) had a 5-fold increased risk of experiencing early-onset severe preeclampsia compared with pregnancies without any of these biomarker patterns (RR, 5.0; 95% confidence interval [CI], 3.4–7.4; sensitivity, 49.5%; specificity, 84.4%; PPV, 0.8%; NPV, 99.9%). This same direction of risk was observed for late-onset severe preeclampsia, wherein pregnancies with any at-risk biomarker had a >2-fold increased risk compared with those without any of these marker patterns (RR, 2.3; 95% CI, 1.6–3.3; sensitivity, 25.9%; specificity, 84.4%; PPV, 1.1%; NPV, 99.4%).
| Variable | No preeclampsia or eclampsia a | Severe preeclampsia | |||
|---|---|---|---|---|---|
| Early onset b | Late onset c | ||||
| n (%) | n (%) | Relative risk (95% CI) | n (%) | Relative risk (95% CI) | |
| No abnormal biomarkers d (n = 93,228) | 92,562 (99.3) | 133 (0.1) | 533 (0.6) | ||
| Referent | |||||
| High biomarker (MoM ≥ 95th percentile) | |||||
| Alpha-fetoprotein (n = 6833) | 6,687 (97.9) | 74 (1.1) | 7.1 (4.3–11.7) e | 72 (1.1) | 2.5 (1.5–4.3) e |
| Human chorionic gonadotropin (n = 6863) | 6,709 (97.8) | 68 (1.0) | 6.9 (4.3–11.0) e | 86 (1.3) | 3.5 (2.3–5.4) e |
| Unconjugated estriol (n = 7179) | 7,112 (99.1) | 13 (0.2) | 1.3 (0.5–3.2) | 54 (0.8) | 1.0 (0.5–2.1) |
| Inhibin-A (n = 6719) | 6,494 (96.7) | 106 (1.6) | 11.4 (7.5–17.4) e | 119 (1.8) | 3.5 (2.2–5.5) e |
| Low biomarker (MoM ≤5th percentile) | |||||
| Alpha-fetoprotein (n = 6789) | 6,739 (99.3) | 9 (0.1) | 0.6 (0.2–2.6) | 41 (0.6) | 0.5 (0.2–1.6) |
| Human chorionic gonadotropin (n = 6321) | 6,273 (99.2) | 11 (0.2) | 0.3 (0.0–2.5) | 37 (0.6) | 0.8 (0.3–2.0) |
| Unconjugated estriol (n = 5450) | 5,360 (98.4) | 34 (0.6) | 3.8 (2.0–7.3) e | 56 (1.0) | 2.8 (1.6–4.9) e |
| Inhibin-A (n = 7155) | 7,111 (99.4) | 8 (0.1) | 0.7 (0.2–2.3) | 36 (0.5) | 0.6 (0.4–1.6) |
a No mild or severe preeclampsia or eclampsia
b Binomial analyses included black race/ethnicity and any diabetes mellitus (all dichotomized as yes vs no)
c Binomial analyses included Hispanic and black race/ethnicity, maternal age ≤17 years, maternal age ≥35 years, weight at testing >95th percentile, and any diabetes mellitus (all dichotomized as yes vs no)
d Alpha-fetoprotein, human chorionic gonadotropin, unconjugated estriol, and inhibin-A multiples of the median all between the 5th and 95th percentile (alpha-fetoprotein, >0.60, <1.74; human chorionic gonadotropin, >0.42, <2.35; unconjugated estriol, >0.61, < 1.49; inhibin-A, >0.48, <1.95)
When at-risk biomarker patterns co-occurred, risks were higher for both early- and late-onset severe preeclampsia. For pregnancies with early-onset severe preeclampsia, high AFP and inhibin with low uE3 levels had the highest risk for development of the disease, with a 1 in 9.5 chance of this diagnosis compared with a 1 in 680.5 chance among pregnancies without any at-risk biomarker pattern (RR, 36.9; 95% CI, 5.6–244.3; Table 3 ). For pregnancies with late-onset severe preeclampsia, the highest risk biomarker pattern was high AFP, hCG, and inhibin levels with low uE3 levels, with a 1 in 20.0 chance of having late-onset severe preeclampsia compared with a 1 in 176.2 chance among pregnancies without any at-risk biomarker pattern (RR, 36.9; 95% CI, 5.6–244.3; Table 4 ). Overall, pregnancies with any at-risk biomarker pattern were nearly 3 times as likely to be diagnosed with severe preeclampsia compared with those without any risk pattern (RR, 2.7; 95% CI, 2.0–3.6). The highest risks for severe preeclampsia were also observed when ≥3 biomarker abnormalities were observed (RR, 13.0–34.2; Table 5 ).
| Variable | Severe early onset preeclampsia | ||||||
|---|---|---|---|---|---|---|---|
| n (%) | Rate (1/x) | Relative risk (95% CI) a | Sensitivity, % | Specificity, % | Positive predictive value, % | Negative predictive value, % | |
| Sample (n = 136,139) | 319 (0.2) | 426.8 | — | ||||
| No abnormal biomarkers b (n = 93,228) | 133 (0.1) | 701.0 | Reference | ||||
| Any early onset preeclampsia “at risk” biomarker c (n = 21,290) | 160 (0.8) | 133.1 | 4.9 (3.3–7.3) d | 50.2 | 84.4 | 0.8 | 99.9 |
| One “at risk” biomarker | |||||||
| High AFP (n = 5270) | 27 (0.5) | 195.2 | 1.7 (0.6–4.7) | 8.5 | 96.1 | 0.5 | 99.8 |
| High hCG (n = 3902) | 10 (0.3) | 390.2 | 2.3 (1.0–5.5) | 3.1 | 97.1 | 0.3 | 99.8 |
| High INH (n = 3845) | 29 (0.8) | 132.6 | 4.9 (2.5–9.7) d | 9.1 | 97.2 | 0.8 | 99.8 |
| Low uE3 (n = 4471) | 14 (0.3) | 319.4 | 0.4 (0.1–3.2) | 4.4 | 96.7 | 0.3 | 99.8 |
| Two “at risk” biomarkers | |||||||
| High AFP and hCG (n = 470) | 2 (0.4) | 235.0 | 6.5 (1.6–26.9) e | 0.6 | 99.7 | 0.4 | 99.8 |
| High AFP and INH (n = 424) | 10 (2.4) | 42.4 | 25.0 (10.8–57.9) d | 3.1 | 99.7 | 2.4 | 99.8 |
| High hCG and INH (n = 1526) | 20 (1.3) | 76.3 | 10.0 (4.9–20.3) d | 6.3 | 98.9 | 1.3 | 99.8 |
| High AFP and low uE3 (n = 144) | 1 (0.7) | 144.0 | 12.8 (1.8–90.5) e | 0.3 | 99.9 | 0.7 | 99.8 |
| High hCG and low uE3 (n = 290) | 0 | — | — | — | — | — | — |
| High INH and low uE3 (n = 235) | 7 (3.0) | 33.6 | 21.9 (7.0–68.8) d | 2.2 | 99.8 | 3.0 | 99.8 |
| Three or more “at risk” biomarkers | |||||||
| High AFP, hCG, and INH (n = 403) | 28 (7.0) | 14.4 | 37.6 (17.4–81.3) d | 8.8 | 99.7 | 6.9 | 99.8 |
| High AFP and hCG; low uE3 (n = 24) | 0 | — | — | — | — | — | — |
| High AFP and INH; low uE3 (n = 38) | 4 (10.5) | 9.5 | 36.9 (5.6–244.3) d | 1.3 | 100.0 | 10.5 | 99.8 |
| High hCG and INH; low uE3 (n = 188) | 6 (3.2) | 31.3 | 27.4 (8.8–85.5) d | 1.9 | 99.9 | 3.2 | 99.8 |
| High AFP, hCG, and INH; low uE3 (n = 60) | 2 (3.3) | 30.0 | 79.0 (21.3–293.4) d | 0.6 | 100.0 | 3.3 | 99.8 |
a Binomial analyses included black race/ethnicity and any diabetes mellitus (all dichotomized as yes vs no)
b AFP, hCG, uE3, and INH multiples of the median all between the 5th and 95th percentile (AFP, >0.60, <1.74; hCG, >0.42, <2.35; uE3, >0.61, <1.49; INH, >0.48, <1.95); 21,621 pregnancies who had neither “at risk” biomarkers nor “no abnormal” biomarkers were not included
c Any high biomarker and/or low uE3 (all biomarkers found to be predictive in Table 2 )
| Variable | Severe late-onset preeclampsia | ||||||
|---|---|---|---|---|---|---|---|
| n (%) | Rate (1/x) | Relative risk (95% CI) a | Sensitivity, % | Specificity, % | Positive predictive value, % | Negative predictive value, % | |
| Sample (n = 136,139) | 889 (0.7) | 153.1 | — | ||||
| No abnormal biomarkers b (n = 93,228) | 533 (0.6) | 174.9 | Reference | ||||
| Any early onset preeclampsia “at risk” biomarker c (n = 21,290) | 231 (1.1) | 92.2 | 2.3 (1.6–3.3) d | 26.0 | 84.4 | 1.1 | 99.4 |
| One “at risk” biomarker | |||||||
| High AFP (n = 5270) | 340 (0.8) | 135.1 | 1.3 (0.6–2.9) | 4.5 | 96.1 | 0.8 | 99.4 |
| High hCG (n = 3902) | 25 (0.6) | 156.1 | 1.6 (0.8–3.5) | 2.8 | 97.1 | 0.6 | 99.3 |
| High INH (n = 3845) | 52 (1.4) | 73.9 | 2.0 (1.0–4.2) | 5.8 | 97.2 | 1.4 | 99.4 |
| Low uE3 (n = 4471) | 33 (0.7) | 135.5 | 1.5 (0.7–3.5) | 3.7 | 96.7 | 0.7 | 99.3 |
| Two “at risk” biomarkers | |||||||
| High AFP and hCG (n = 470) | 8 (1.7) | 58.8 | 7.2 (2.3–22.3) d | 0.9 | 99.7 | 1.7 | 99.4 |
| High AFP and INH (n = 424) | 11 (2.6) | 38.5 | 2.9 (0.4–20.2) | 1.2 | 99.7 | 2.6 | 99.4 |
| High hCG and INH (n = 1526) | 32 (2.1) | 50.9 | 3.3 (1.4–8.1) e | 3.6 | 98.9 | 2.1 | 99.4 |
| High AFP and low uE3 (n = 144) | 1 (0.7) | 144.0 | 1.2 (0.2–8.6) f | 0.1 | 99.9 | 0.7 | 99.3 |
| High hCG and low uE3 (n = 290) | 4 (1.4) | 72.5 | 5.6 (1.4–22.4) g | 0.4 | 99.8 | 1.4 | 99.3 |
| High INH and low uE3 (n = 235) | 7 (3.0) | 33.6 | 4.5 (0.6–31.7) | 0.8 | 99.8 | 3.0 | 99.4 |
| Three or more “at risk” biomarkers | |||||||
| High AFP, hCG, and INH (n = 403) | 7 (1.7) | 57.6 | 9.3 (3.0–28.7) d | 0.8 | 99.7 | 1.7 | 99.4 |
| High AFP and hCG; low uE3 (n = 24) | 1 (4.2) | 24.0 | 7.3 (1.1–50.0) f,g | 0.1 | 100.0 | 4.2 | 99.3 |
| High AFP and INH; low uE3 (n = 38) | 1 (2.6) | 38.0 | 5.2 (0.7–35.5) f | 0.1 | 100.0 | 2.6 | 99.3 |
| High hCG and INH; low uE3 (n = 188) | 6 (3.2) | 31.3 | 13.0 (4.3–39.3) d | 0.7 | 99.9 | 3.2 | 99.4 |
| High AFP, hCG, and INH; low uE3 (n = 60) | 3 (5.0) | 20.0 | 44.8 (12.9–155.1) d | 0.3 | 100.0 | 5.0 | 99.3 |
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