Meta-Analysis of Maternal and Fetal Transcriptomic Data Elucidates the Role of Adaptive and Innate Immunity in Preterm Birth

Abstract

Preterm birth (PTB) is the leading cause of newborn deaths around the world. Spontaneous preterm birth (sPTB) accounts for two-thirds of all PTBs; however, there remains an unmet need of detecting and preventing sPTB. Although the dysregulation of the immune system has been implicated in various studies, small sizes and irreproducibility of results have limited identification of its role. Here, we present a cross-study meta-analysis to evaluate genome-wide differential gene expression signals in sPTB. A comprehensive search of the NIH genomic database for studies related to sPTB with maternal whole blood samples resulted in data from three separate studies consisting of 339 samples. After aggregating and normalizing these transcriptomic datasets and performing a meta-analysis, we identified 210 genes that were differentially expressed in sPTB relative to term birth. These genes were enriched in immune-related pathways, showing upregulation of innate immunity and downregulation of adaptive immunity in women who delivered preterm. An additional analysis found several of these differentially expressed at mid-gestation, suggesting their potential to be clinically relevant biomarkers. Furthermore, a complementary analysis identified 473 genes differentially expressed in preterm cord blood samples. However, these genes demonstrated downregulation of the innate immune system, a stark contrast to findings using maternal blood samples. These immune-related findings were further confirmed by cell deconvolution as well as upstream transcription and cytokine regulation analyses. Overall, this study identified a strong immune signature related to sPTB as well as several potential biomarkers that could be translated to clinical use.

Keywords: immunology; meta-analysis; pregnancy; preterm birth; transcriptomics.

Figure 1

Analysis of relationship of gene expression differences in term vs. preterm birth. We identified three independent studies from the Gene Expression Omnibus database (in yellow) to perform a meta-analysis using third trimester maternal blood samples (in green), an additional differential expression analysis with second trimester samples from GSE59491 (in orange), and a tissue-specific analysis with samples from GSE73685 (in blue).

Figure 2

Results from the cross-study meta-analysis and distribution of gestational age at sampling. (A,B) Principal component analysis plots with all genes before (A) and after (B) ComBat. (C,D) Principal component analysis plot (C) and heatmap (D) of all samples based on 210 significant differentially expressed genes. (E) Gestational age at sampling was not significantly different between preterm and term maternal whole blood samples (n = 315, p-value = 0.125).

Figure 3

STRING connectivity networks based on 210 differentially expressed genes. (A,B) Connectivity networks for significantly downregulated (A) and upregulated (B) genes from meta-analysis.

Figure 4

(A–D) Network visualization of functionally enriched GO biological processes in significantly downregulated genes from the meta-analysis.

Figure 5

Cell deconvolution of 339 meta-analysis samples. Boxplot (A) and heatmap (B) of average xCell scores for enriched cell types.

Figure 6

Results from additional second trimester analysis. (A) Heatmap of significant genes from second trimester analysis; genes which are secreted as proteins are boxed. (B,C) Boxplots of genes that encode secreted proteins at second (T2) and third (T3) trimester; raw gene expression values from GSE59491 are plotted.

Figure 7

Regulatory networks for second and third trimester differentially expressed genes. Transcription regulation networks for differentially expressed genes in the second trimester (A) and third trimester (B), where the transcription factors are represented with a purple round node and the differentially expressed targets are represented with a gray square node. Cytokine networks for second trimester (C) and third trimester (D), where the transcription factors are represented with an orange hexagon node and the differentially expressed targets are represented with a gray square node.

Figure 8

Significant genes from cord blood (CB) tissue analysis and maternal–cord gene signature comparison. (A) Heatmap of significant differentially expressed genes from CB analysis. (B) Boxplot of overlapping significant genes from meta-analysis and CB analysis; raw gene expression values from GSE73685 plotted.

Figure 8

Significant genes from cord blood (CB) tissue analysis and maternal–cord gene signature comparison. (A) Heatmap of significant differentially expressed genes from CB analysis. (B) Boxplot of overlapping significant genes from meta-analysis and CB analysis; raw gene expression values from GSE73685 plotted.