Integrated proteomics and metabolomics network analysis across different delivery modes in human pregnancy: a pilot study

Abstract

Background: Delivery mode has been linked to child health, e.g., allergic disease. However, it remains unclear whether protein and metabolite differences across different delivery modes may underlie child development.

Methods: A cohort comprising 16 spontaneous onset vaginal delivery (VD), 16 prelabor cesarean delivery on maternal request (CS), and 8 intrapartum cesarean section (Intra_CS) women were analyzed using label-free proteomic and untargeted metabolomics assays on amniotic fluid and cord blood samples, respectively. We used weighted gene co-expression network analyses (WGCNA) to identify modules of highly correlated proteins or metabolites that associated with delivery modes and related clinical traits. KEGG enrichment analyses were performed to investigate the biological function of the identified modules. Integrative multiomics analysis was employed to examine the biological interplay between proteomic and metabolic interactions.

Results: Compared to the CS group, the proteomic and metabolomic profiles were similar between the Intra_CS and VD groups in our study. We did not identify any enriched protein or metabolite pathways related to immune development that could influence the risk of allergic diseases in offspring across different delivery modes. However, we identified seven protein modules correlated with the duration from the rupture of the membranes to full dilation of the cervix, with the actin cytoskeleton module significantly enriched. A metabolic module in cord blood that correlated with VD was enriched in subclasses including C21 steroids, steroid sulfates, and oxysterols. Integrative analysis of proteomic and metabolomic data suggested pathways related to mode of delivery and duration of labor, encompassing the actin cytoskeleton, NADP metabolic process, nicotinate, and nicotinamide metabolism in amniotic fluid, and the steroid hormone biosynthesis pathway in cord blood.

Conclusions: Differences in steroid hormones and the actin cytoskeleton pathway according to proteomics and metabolomics in amniotic fluid and cord blood were more indicative of the labor process. These findings could guide future studies on delivery-associated biochemical pathways.

Keywords: Cesarean section; Delivery mode; Metabolomics; Proteomics; WGCNA.

Fig. 1

KEGG enrichment analysis for proteomics between groups of delivery mode. (a) KEGG pathway enriched in amniotic fluid proteins between CS vs. VD and CS vs. Intra_CS. (b) KEGG pathway enriched in cord blood proteins between CS vs. VD and CS vs. Intra_CS. VD, spontaneous onset vaginal delivery group. CS, prelabor cesarean delivery on maternal request, Intra_CS, intrapartum cesarean section

Fig. 2

Volcano plot for the proteome and metabolome analysis in amniotic fluid and cord blood sample among the merged vaginal delivery (merged_labor) group and prelabor cesarean delivery on maternal request (CS) group. (a) Volcano plots of the proteins in the amniotic fluid. (b) Volcano plots of the proteins in the cord blood. (c) Volcano plots of the metabolites in the amniotic fluid. (d) Volcano plots of the metabolites in the cord blood. Benjamini-Hochberg (BH) corrected p-value < 0.05 and log2FC > 0.585 or < -0.585 were used as cutoffs to define up and down regulation

Fig. 3

Quantitative proteomics and metabolomics of amniotic fluid and cord blood sample among the merged vaginal delivery (merged_labor) group and prelabor cesarean delivery on maternal request (CS) group a. Hierarchical clustering based on differential proteins in amniotic fluid; b. Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis of differentially expressed proteins in amniotic fluid; c. Hierarchical clustering based on differential metabolites in amniotic fluid; d. KEGG pathway enrichment analysis of differentially expressed metabolites in amniotic fluid; e. Hierarchical clustering based on differential metabolites in cord blood; f. KEGG pathway enrichment analysis of differentially expressed metabolites in cord blood. ROM_birth, duration of rupture of the membranes (ROM) to childbirth (hours). For hierarchical clustering, the differentially expressed proteins and metabolites were identified with Benjamini-Hochberg (BH) corrected p-value < 0.05 and log2FC > absolute value of log2|1.5|. Significantly altered proteins and metabolites are highlighted in red (increased) and blue (decreased)

Fig. 4

Correlations among protein networks and metabolic networks and clinical traits in amniotic fluid. a. Circus plot showing the correlations among metabolite modules, protein modules and clinical traits in amniotic fluid. Only significant associations after FDR correction are shown, and the width of the lines represents the strength of the correlations. b. Protein-protein interaction plot of top drivers for turquoise module in amniotic fluid. BP, MF, and CC represent Biological Process, Molecular Function, and Cellular Component groups of gene ontology (GO), respectively. c. Top drivers of turquoise module between merged_labor and CS group (top), and correlation between top drivers of turquoise module with the duration of ROM to childbirth. d. Heat map showing the Pearson correlations and FDR-adjusted P values (in bracket) between metabolite modules (rows) and protein modules (columns) associated with clinical traits in amniotic fluid

Fig. 5

Correlations among protein networks and metabolic networks and clinical traits in cord blood. (a) Circus plot showing the correlations among metabolite modules, protein modules and clinical traits in cord blood. Only significant associations after FDR correction are shown, and the dotted edges represent a significant association between the protein module and the metabolite module. The width of the lines represents the strength of the correlations. (b) Protein-protein interaction plot of top drivers for turquoise module in cord blood. CC represents Cellular Component groups of gene ontology (GO). (c) Levels of Cofilin-1, top driver of turquoise module of proteins, between the merged_labor and CS group. (d) Correlation between Cofilin-1 in cord blood with the duration of second stage of labor. (e) Levels of metabolic green module between the merged_labor and CS group. (f) Levels of metabolic magenta module between the merged_labor and CS group. (g) Heat map showing the Pearson correlations and FDR-adjusted P values (in bracket) between metabolite modules (rows) and protein modules (columns) associated with clinical traits in cord blood

Fig. 6

Pathway enrichment analysis for metabolic and proteomic top drivers of identified modules. (a) joint pathway analysis for protein (turquoise) and metabolites (turquoise) modules, borderline indicate pathway that enriched in the joint pathway analysis in amniotic fluid. (b) joint pathway analysis for protein (black) and metabolites (turquoise) modules in amniotic fluid. (c) joint pathway analysis for protein (greenyellow) and metabolites (green) modules in cord blood