Metabolomics of Human Amniotic Fluid and Maternal Plasma during Normal Pregnancy

Abstract

Metabolic profiles of amniotic fluid and maternal blood are sources of valuable information about fetus development and can be potentially useful in diagnosis of pregnancy disorders. In this study, we applied 1H NMR-based metabolic profiling to track metabolic changes occurring in amniotic fluid (AF) and plasma (PL) of healthy mothers over the course of pregnancy. AF and PL samples were collected in the 2nd (T2) and 3rd (T3) trimester, prolonged pregnancy (PP) until time of delivery (TD). A multivariate data analysis of both biofluids reviled a metabolic switch-like transition between 2nd and 3rd trimester, which was followed by metabolic stabilization throughout the rest of pregnancy probably reflecting the stabilization of fetal maturation and development. The differences were further tested using univariate statistics at α = 0.001. In plasma the progression from T2 to T3 was related to increasing levels of glycerol, choline and ketone bodies (3-hydroxybutyrate and acetoacetate) while pyruvate concentration was significantly decreased. In amniotic fluid, T2 to T3 transition was associated with decreasing levels of glucose, carnitine, amino acids (valine, leucine, isoleucine, alanine, methionine, tyrosine, and phenylalanine) and increasing levels of creatinine, succinate, pyruvate, choline, N,N-dimethylglycine and urocanate. Lactate to pyruvate ratio was decreased in AF and conversely increased in PL. The results of our study, show that metabolomics profiling can be used to better understand physiological changes of the complex interdependencies of the mother, the placenta and the fetus during pregnancy. In the future, these results might be a useful reference point for analysis of complicated pregnancies.

Fig 1

A representative example of the ¹H NMR spectrum of amniotic fluid (A) and plasma (B). The numbers in the figure correspond to the numbers in Table A in S1 File. 1 –leucine, 2 –valine, 3 –isoleucine, 4 –threonine, 5 –alanine, 6 –lysine, 7 –methionine, 8 –glutamine, 9 –glutamate, 10 –glycine, 11 –tyrosine, 12 –phenylalanine, 13 –histidine, 14 –N,N—dimethylglycine, 15 –creatinine, 16 –creatine, 17 –choline, 18 –carnitine, 19 –dimethylamine, 20 –sarcosine, 21 –dimethylsulfone, 22 –trimethylamine N-oxide (TMAO), 23 –lactate, 24 –acetate, 25 –pyruvate, 26 –succinate, 27 –citrate, 28 –urocanate, 29 –fumarate, 30 –formate, 31 –acetoacetate, 32–2-hydroxyisovalerate, 33–2-hydroxybutyrate, 34 –isobutyrate, 35–3-hydroxybutyrate, 36–3-hydroxyisovalerate, 37 –proline, 38 –N-acetyl groups, 39 –NAC, 40 –VLDL/LDL, 41 –acetone, 42 –methanol, 43 –glycerol, 44 –mannose, 45 –glucose, 46 –AU1, 47 –AU2, 48 –PU1, 49 –PU2.

Fig 2

The PCA results obtained using quantified metabolites (signal areas) for amniotic fluid (A) and plasma (B). Pregnancy stages. Yellow inverted triangles– 2^nd trimester (T2), red triangles– 3^rd trimester (T3), blue squares–delivery (TD) and green circles–prolonged pregnancy (PP).

Fig 3

Significantly changed metabolites that were detected in both biofluids: pyruvate (A), glucose (B), choline (C), 3-hydroxybutyrate (D) and lactate to pyruvate ratio (E). Bar graphs obtained for amniotic fluid (left) and for plasma (middle). Relationship plot (right)—symbol denotes to average and error bars denotes to standard deviation of the normalized signal values for each group. T2 –second trimester, T3 –third trimester, TD–time of delivery, PP–prolonged pregnancy.