Demographics of the study population are shown in Table 1. The majority of mothers were under the age of 30 (67.1%) [median(IQR): 27 (23, 31)], had at least some college education (79%), were employed (63%), had an annual household income under $30,000 (63.1%), were married (53.1%), had never smoked (86%) or been exposed to environmental tobacco smoke (88.7%), did not drink alcohol during pregnancy (93.6%), had given birth to less than 2 previous children (86.9%), and had a pre-pregnancy BMI of less than 25 (56.1%) [median(IQR): 24.1 (21.4, 28.1)].
Table 1
Maternal demographic characteristics of the study population (N = 976)
| N (%) |
Maternal Age (years) | |
18–24 | 354 (36.3%) |
25–29 | 301 (30.8%) |
30–34 | 206 (21.1%) |
35–41 | 115 (11.8%) |
Missing | 0 (0%) |
Maternal Education | |
GED or less | 203 (21%) |
Some College | 331 (34.2%) |
Bachelors or Higher | 433 (44.8%) |
Missing | 9 (0.9%) |
Employment Status | |
No | 357 (37%) |
Yes | 608 (63%) |
Missing | 11 (1.1%) |
Annual Household Income | |
< 10 k | 269 (31.6%) |
10 k-<30 k | 268 (31.5%) |
30 k-<50 k | 203 (23.8%) |
>=50 k | 112 (13.1%) |
Missing | 124 (12.7%) |
Marital Status | |
Single | 197 (20.4%) |
Married | 521 (53.9%) |
Cohabitating | 249 (25.7%) |
Missing | 9 (0.9%) |
Smoking Status | |
Never | 833 (86%) |
Ever | 121 (12.5%) |
Current | 15 (1.55%) |
Missing | 7 (0.7%) |
Daily Environmental Tobacco Smoke Exposure | |
Never | 808 (88.7%) |
1 Hour or less | 40 (4.39%) |
> 1 Hour | 63 (6.92%) |
Missing | 65 (6.7%) |
Alcohol Use | |
Never | 504 (52.2%) |
Yes, before Pregnancy | 400 (41.4%) |
Yes, currently | 62 (6.42%) |
Missing | 10 (1.0%) |
Number of Previous Children | |
0 | 355 (42.7%) |
1 | 367 (44.2%) |
2 to 5 | 109 (13.1%) |
Missing | 145 (14.9%) |
Pre-Pregnancy BMI | |
[0,25] | 520 (56.1%) |
(25, 30] | 240 (25.9%) |
Above 30 | 167 (18%) |
Missing | 49 (5.0%) |
Fetal Sex | |
Female | 464 (48%) |
Male | 502 (52%) |
Missing | 10 (1.0%) |
Distributions of hormone concentrations are shown in Table 2. Most hormone concentrations were significantly different at 18 and 26 weeks gestation, with notable increases occurring with estriol (median 15.1 and 38.2 ng/mL at 18 and 26 weeks, respectively) and progesterone (median 39.3 and 73.5 ng/mL at 18 and 26 weeks, respectively). ICCs, which reflect the degree of correlation and agreement between measurements, for all other hormones ranged from 0.647 (T4) to 0.856 (testosterone).
Table 2
Distributions of gestational average (GA) and visit specific hormone concentrations
| | N | min | 25th | 50th | 75th | 90th | 95th | Max | Geo. Mean | Geo. Stdv | IQR | Visit P-value* | ICC (95% CI) |
CRH (pg/mL) | GA | 976 | 3.50 | 15.4 | 43.2 | 86.3 | 118 | 148 | 243 | 35.7 | 2.77 | 70.9 | 0.914 | 0.71 (0.66, 0.74) |
Visit 1 | 818 | 3.50 | 15.1 | 37.6 | 84.3 | 121 | 156 | 254 | 34.4 | 2.89 | 69.2 | | |
Visit 2 | 602 | 3.50 | 14.7 | 39.3 | 88.2 | 130 | 159 | 249 | 34.2 | 2.95 | 73.4 | | |
Estriol (mg/mL) | GA | 971 | 0.74 | 15.6 | 23.1 | 33.0 | 44.7 | 57.5 | 265 | 22.7 | 1.80 | 17.4 | 0.000 | -0.22 (-0.35, -0.11) |
Visit 1 | 812 | 0.74 | 11.3 | 15.1 | 22.2 | 31.8 | 41.5 | 108 | 15.8 | 1.75 | 10.9 | | |
Visit 2 | 600 | 6.90 | 29.3 | 38.2 | 50.5 | 64.4 | 74.6 | 265 | 38.7 | 1.55 | 21.2 | | |
SHBG (pg/mL) | GA | 976 | 47.6 | 413 | 538 | 668 | 818 | 895 | 1404 | 522 | 1.45 | 254 | 0.000 | 0.76 (0.72, 0.79) |
Visit 1 | 820 | 47.6 | 389 | 516 | 630 | 775 | 850 | 1461 | 491 | 1.47 | 241 | | |
Visit 2 | 602 | 123 | 434 | 566 | 723 | 898 | 979 | 1428 | 558 | 1.45 | 289 | | |
Prog. (ng/mL) | GA | 973 | 10.1 | 36.6 | 50.4 | 71.0 | 99.4 | 124 | 1037 | 51.8 | 1.68 | 34.5 | 0.000 | 0.07 (-0.04, 0.17) |
Visit 1 | 815 | 10.1 | 29.2 | 39.3 | 54.5 | 71.9 | 85.0 | 301 | 40.1 | 1.59 | 25.3 | | |
Visit 2 | 601 | 19.4 | 51.2 | 73.5 | 104 | 146 | 179 | 1037 | 74.4 | 1.70 | 53.2 | | |
TSH (uIU/mL) | GA | 971 | 0.03 | 0.71 | 1.10 | 1.72 | 2.38 | 2.99 | 32.4 | 1.08 | 1.96 | 1.02 | 0.031 | 0.72 (0.67, 0.75) |
Visit 1 | 812 | 0.02 | 0.67 | 1.05 | 1.66 | 2.38 | 2.88 | 40.9 | 1.03 | 2.06 | 0.99 | | |
Visit 2 | 600 | 0.11 | 0.72 | 1.15 | 1.75 | 2.43 | 3.23 | 25.7 | 1.12 | 1.96 | 1.03 | | |
fT4 (ng/dL) | GA | 976 | 0.11 | 1.09 | 1.62 | 2.02 | 2.32 | 2.50 | 8.35 | 1.41 | 1.68 | 0.93 | 0.452 | 0.75 (0.71, 0.79) |
Visit 1 | 818 | 0.11 | 1.03 | 1.57 | 2.01 | 2.30 | 2.48 | 8.35 | 1.34 | 1.84 | 0.98 | | |
Visit 2 | 602 | 0.11 | 1.10 | 1.61 | 2.03 | 2.33 | 2.49 | 4.68 | 1.39 | 1.75 | 0.93 | | |
T4 (ug/dL) | GA | 975 | 0.35 | 0.89 | 1.00 | 1.10 | 1.21 | 1.28 | 1.72 | 0.99 | 1.19 | 0.21 | 0.000 | 0.65 (0.59, 0.69) |
Visit 1 | 818 | 0.35 | 0.90 | 1.01 | 1.12 | 1.21 | 1.28 | 1.72 | 1.00 | 1.19 | 0.22 | | |
Visit 2 | 602 | 0.44 | 0.83 | 0.96 | 1.08 | 1.19 | 1.23 | 1.43 | 0.94 | 1.21 | 0.25 | | |
T3 (mg/mL) | GA | 971 | 6.20 | 10.5 | 11.8 | 13.2 | 14.4 | 15.2 | 19.0 | 11.7 | 1.18 | 2.70 | 0.008 | 0.72 (0.67, 0.75) |
Visit 1 | 812 | 6.80 | 10.6 | 11.9 | 13.3 | 14.4 | 15.3 | 19.0 | 11.8 | 1.19 | 2.70 | | |
Visit 2 | 600 | 5.30 | 10.3 | 11.6 | 13.0 | 14.2 | 14.9 | 20.6 | 11.5 | 1.19 | 2.75 | | |
Test. (pg/mL) | GA | 973 | 2.80 | 53.0 | 107 | 557 | 819 | 992 | 2868 | 160 | 3.55 | 504 | 0.012 | 0.86 (0.83, 0.88) |
Visit 1 | 815 | 1.10 | 50.1 | 105 | 544 | 789 | 952 | 2500 | 156 | 3.66 | 493 | | |
Visit 2 | 601 | 9.20 | 59.3 | 121 | 650 | 933 | 1092 | 3291 | 185 | 3.64 | 591 | | |
*P-value from a univariate linear model for association between hormone concentrations and study visit. Boldface p-values are < 0.05. Gestational average values were calculated as arithmetic means for normally distributed hormones and geometric means for log-normally distributed hormones. |
Distributions of birth outcomes are shown in Table 3. PTB and spontaneous PTB occurred in 9.9% and 5.8% of the study population, respectively. As expected, preeclampsia and GDM were less prevalent (2.9% and 1.9%, respectively). Occurrences of SGA and LGA births were similar (8.9% and 9.6%, respectively). Median gestational age of the study population was 39.1 weeks (IQR: 38.1–40).
Table 3
Distributions of continuous and binary birth outcomes
| Min | 10th | 25th | 50th | 75th | 90th | Max |
Gestational Age (wks) | 20.3 | 36.7 | 38.1 | 39.1 | 40 | 40.7 | 42.7 |
Birth Weight Z-Score (ounces) | -5.34 (19.0) | -1.19 (91.0) | -0.571 (102) | -0.00005 (113) | 0.707 (123) | 1.25 (133) | 9.70 (224) |
| N (%) | | | | | | |
Preterm Birth | | | | | | | |
No | 867 (90.1%) | | | | | | |
Yes | 95 (9.88%) | | | | | | |
Spontaneous Preterm Birth | | | | | | | |
No | 883 (94.2%) | | | | | | |
Yes | 54 (5.76%) | | | | | | |
Preeclampsia | | | | | | | |
No | 947 (97.1%) | | | | | | |
Yes | 28 (2.87%) | | | | | | |
Gestational Diabetes | | | | | | | |
No | 900 (98.1%) | | | | | | |
Yes | 17 (1.85%) | | | | | | |
Small for Gestational Age | | | | | | | |
No | 842 (91.1%) | | | | | | |
Yes | 82 (8.87%) | | | | | | |
Large for Gestational Age | | | | | | | |
No | 835 (90.4%) | | | | | | |
Yes | 89 (9.63%) | | | | | | |
Additional File 1: Hormone concentrations between cases and controls for binary outcomes, comparing women carrying females versus males. |
Solid lines are trends for cases, while dashed lines are trends for normal pregnancies. All red lines correspond to mothers carrying a female fetus, and all blue lines correspond to mothers carrying a male fetus. Background dots represent cases for each birth outcome. |
Additional File 1 depicts the differences in hormone concentrations between cases and controls over the study period for each birth outcome, stratified by fetal sex. CRH, progesterone, and T3 concentrations were higher across pregnancy among women presenting with PTB, spontaneous PTB, and GDM only when the fetal sex was male, with the difference in progesterone concentrations becoming minimal later in pregnancy. When mothers were carrying a female fetus, CRH concentrations among PTB and spontaneous PTB cases were lower than concentrations for control women, particularly later in pregnancy (around 26 weeks). Conversely, testosterone concentrations were higher among PTB, spontaneous PTB, and GDM cases when fetal sex was female compared to male across pregnancy. Among women with preeclampsia, those carrying a female fetus had lower concentrations of SHBG and progesterone but higher concentrations of TSH and testosterone compared to women carrying males across pregnancy. Among women with GDM, those carrying a male fetus had higher SHBG, progesterone, fT4, T4, and prog/e3 concentrations later in pregnancy relative to those carrying females. Among GDM cases, T3/T4 was higher earlier in pregnancy (around 18 weeks) when the fetal sex was male, but later in pregnancy (around 26 weeks) when fetal sex was female.
Associations between gestational average hormone concentrations and birth outcomes are shown in Fig. 1. PTB was not significantly associated with any hormone concentrations, however gestational age at birth was reduced by 1.81 (95% CI: 0.36, 3.26), 1.78 (95% CI: 0.18, 3.39) and 2.55 (95% CI: 1.16, 3.93) days with IQR increases in fT4, T4, and prog/e3, respectively. An IQR increase in fT4 was also associated with 1.57 (95% CI: 1.11, 2.24) times greater odds of spontaneous PTB, while an IQR increase in prog/e3 was also associated with 1.63 (95% CI: 1.00, 2.65) times greater odds of preeclampsia and 1.43 (95% CI: 1.10, 1.88) times greater odds of SGA. Increased progesterone was associated with both greater odds of SGA (OR: 1.47, 95% CI: 1.10, 1.95) and with lower birth weight z-score (β: 0.09, 95% CI: -0.19, 0.00). Finally, the odds of GDM were reduced (OR: 0.37, 95% CI: 0.13, 1.01) with an IQR increase in testosterone concentration.
We also examined the differences in hormone associations with birth outcomes by fetal sex and the timing of hormone concentration measurement. Figure 2 shows the associations between gestational average hormone concentrations and birth outcomes, by fetal sex. Among women carrying a male fetus, the odds of PTB were significantly increased with IQR increases in CRH (OR: 1.87, 95%CI: 1.08, 3.24), estriol (OR: 1.48, 95%CI: 0.99, 2.20), progesterone (OR:1.60, 95% CI: 1.11, 2.29), and fT4 (OR: 1.48, 95%CI: 1.00, 2.20), whereas the odds of PTB were decreased with an IQR increase in testosterone (OR: 0.56, 95% CI: 0.31, 1.00). Similar results were found with spontaneous PTB, with the exception of estriol which showed a null association. Additionally, an IQR increase in T3 was associated with 1.80 (95% CI: 1.02, 3.16) times greater odds of spontaneous PTB among women carrying a male fetus. Similarly, gestational age at birth decreased by 2.88 (95%CI: 0.97, 4.79), 2.24 (95%CI: 0.26, 4.21), and 2.74 (95%CI: 0.84, 4.63) days with IQR increases in progesterone, fT4, and prog/e3, respectively, when fetal sex was male.
The odds of GDM increased with an IQR increase in fT4 (OR: 2.80, 95%CI: 1.06, 7.41) and decreased with an IQR increase in testosterone (OR: 0.23, 95%CI: 0.06, 0.86) when fetal sex was male. Odds of GDM also decreased with an IQR increase in prog/e3 (OR: 0.34, 95%CI: 0.12, 0.99) when fetal sex was female.
Increases in progesterone (OR: 1.50, 95%CI: 1.02, 2.21) and prog/e3 (OR: 2.08, 95%CI: 1.36, 3.19) were associated with increased odds of delivering an SGA male, but not female infant. Birth weight z-score was reduced with increasing concentrations of progesterone (β: -0.17, 95%CI: -0.31, -0.02) and T4 (β: -0.16, 95%CI: -0.30, -0.02) among women carrying a female fetus, but only prog/e3 (β: -0.14, 95%CI: -0.27, -0.01) among women carrying a male fetus.
Some hormone relationships with birth outcomes differed when measured around 18 versus 26 weeks gestation (Fig. 3). Regardless of fetal sex, there were greater odds of spontaneous PTB with increasing progesterone concentrations around 26 weeks (OR: 2.12, 95% CI: 1.26, 3.55) and fT4 concentrations at both study visits (18wk OR: 1.60, 95% CI: 1.06, 2.41; 26wk OR: 1.73, 95% CI: 1.00, 2.97). Reductions in gestational age at birth were observed with increased concentrations of progesterone (β: -3.56 days, 95% CI: -5.51, -1.60), fT4 (β: -2.22 days, 95% CI: -3.87, -0.58), and T4 (β: -1.87 days, 95% CI: -3.47, -0.27) around 18 weeks, and with prog/e3 at both study visits (18wk β: -1.77 days, 95% CI: -3.20, -0.34; 26wk β: -1.98 days, 95% CI: -3.83, -0.12).
Increases in TSH (OR: 2.18, 95% CI: 1.08, 4.40) and prog/e3 (OR: 1.78, 95% CI: 0.99, 3.19) at 26 weeks were associated with greater odds of preeclampsia, while SHBG at both time points was inversely associated with preeclampsia (18wk OR: 0.55, 95% CI: 0.31, 0.96; 26wk OR: 0.46, 95% CI: 0.22, 0.94). Increases in T3 (OR: 2.83, 95% CI: 1.09, 7.34) and T3/T4 (OR: 2.97, 95% CI:1.19, 7.43) at 18 weeks were associated with greater odds of GDM, while increases in estriol (OR: 5.95, 95% CI: 1.45, 24.5) and progesterone (OR: 2.71, 95% CI: 1.09, 6.69) were associated with greater odds of GDM around 26 weeks.
Increased odds of SGA were observed with increasing progesterone concentrations around 18 weeks (OR: 1.53, 95% CI: 1.00, 2.34), and with increasing prog/e3 at both study visits (18wk OR: 1.77, 95% CI: 1.29, 2.43; 26wk OR: 1.53, 95% CI: 1.06, 2.19). Concordantly, increased estriol around 26 weeks (β: 0.21, 95% CI: 0.00, 0.42), and decreased prog/e3 around 18 weeks (β: -0.12, 95% CI: -0.22, -0.03), were associated with increased birth weight z-score.