Catalytic ferrous iron in amniotic fluid as a predictive marker of human maternal-fetal disorders

Abstract

Amniotic fluid contains numerous biomolecules derived from fetus and mother, thus providing precious information on pregnancy. Here, we evaluated oxidative stress of human amniotic fluid and measured the concentration of catalytic Fe(II). Amniotic fluid samples were collected with consent from a total of 89 subjects in Nagoya University Hospital, under necessary medical interventions: normal pregnancy at term, normal pregnancy at the 2nd trimester, preterm delivery with maternal disorders but without fetal disorders, congenital diaphragmatic hernia, fetal growth restriction, pregnancy-induced hypertension, gestational diabetes mellitus, Down syndrome and trisomy 18. Catalytic Fe(II) and oxidative stress markers (8-hydroxy-2'-deoxyguanosine, 8-OHdG; dityrosine) were determined with RhoNox-1 and specific antibodies, respectively, using plate assays. Levels of 8-OHdG and dityrosine were higher in the 3rd trimester compared with the 2nd trimester in normal subjects, and the abnormal groups generally showed lower levels than the controls, thus suggesting that they represent fetal metabolic activities. In contrast, catalytic Fe(II) was higher in the 2nd trimester than the 3rd trimester in the normal subjects, and overall the abnormal groups showed higher levels than the controls, suggesting that high catalytic Fe(II) at late gestation reflects fetal pathologic alterations. Notably, products of H2O2 and catalytic Fe(II) remained almost constant in amniotic fluid.

Keywords: amniotic fluid; catalytic ferrous iron; oxidative stress; pregnancy.

Fig. 1

8-OHdG levels in amniotic fluid. (A) 8-OHdG levels in amniotic fluid were increased according with the progression of gestational period (mean ± SEM; *p<0.01 vs control with Bonferroni test). (B) Comparison of control with abnormal pregnancies. 8-OHdG levels in abnormal pregnancies were significantly lower than the controls (mean ± SEM; *p<0.05 by Student’s t test; **p<0.01 by Mann-Whitney test). PIH, pregnancy induced hypertension; GDM, gestational diabetes mellitus; FGR, fetal growth restriction; CDH, congenital diaphragmatic hernia.

Fig. 2

Dityrosine levels in amniotic fluid. (A) Dityrosine levels in amniotic fluid were increased as gestation progressed (mean ± SEM; *p<0.01 with Bonferroni test). (B) Comparison of control with abnormal pregnancies. Dityrosine levels abnormal pregnancies were significantly lower than the controls (mean ± SEM; *p<0.05; **p<0.01 with Student’s t test). PIH, pregnancy induced hypertension; GDM, gestational diabetes mellitus; FGR, fetal growth restriction; CDH, congenital diaphragmatic hernia.

Fig. 3

Levels of catalytic Fe(II) in amniotic fluid. (A) Catalytic Fe(II) levels in amniotic fluid of normal pregnancies were decreased according with the progression of gestational periods (mean ± SEM; *p<0.01 with Bonferroni test). (B) Comparison of control with abnormal pregnancies. Catalytic Fe(II) levels in amniotic fluid of abnormal pregnancies were all significantly higher than the controls (mean ± SEM; *p<0.05; **p<0.01 by Mann-Whitney test). PIH, pregnancy induced hypertension; GDM, gestational diabetes mellitus; FGR, fetal growth restriction; CDH, congenital diaphragmatic hernia.

Fig. 4

Concentration of hydrogen peroxide (H₂O₂). (A) The levels of H₂O₂ in amniotic fluid of normal pregnancies showed no significant differences among each pregnant stages (mean ± SEM; Bonferroni test). (B) Comparison of control and abnormal pregnancies. The levels of control were higher than abnormal pregnancies (mean ± SEM; *p<0.05 by Mann-Whitney test, **p<0.05 by Student’s t test).

Fig. 5

Product of H₂O₂ and catalytic Fe(II). (A) Products of normal pregnancy showed no significant differences (mean ± SEM; Bonferroni test). (B) Comparison of control group with abnormal pregnancies. Products also showed no significant differences (mean ± SEM; Mann-Whitney test).