Inflammatory Gene Regulatory Networks in Amnion Cells Following Cytokine Stimulation: Translational Systems Approach to Modeling Human Parturition

Abstract

A majority of the studies examining the molecular regulation of human labor have been conducted using single gene approaches. While the technology to produce multi-dimensional datasets is readily available, the means for facile analysis of such data are limited. The objective of this study was to develop a systems approach to infer regulatory mechanisms governing global gene expression in cytokine-challenged cells in vitro, and to apply these methods to predict gene regulatory networks (GRNs) in intrauterine tissues during term parturition. To this end, microarray analysis was applied to human amnion mesenchymal cells (AMCs) stimulated with interleukin-1β, and differentially expressed transcripts were subjected to hierarchical clustering, temporal expression profiling, and motif enrichment analysis, from which a GRN was constructed. These methods were then applied to fetal membrane specimens collected in the absence or presence of spontaneous term labor. Analysis of cytokine-responsive genes in AMCs revealed a sterile immune response signature, with promoters enriched in response elements for several inflammation-associated transcription factors. In comparison to the fetal membrane dataset, there were 34 genes commonly upregulated, many of which were part of an acute inflammation gene expression signature. Binding motifs for nuclear factor-κB were prominent in the gene interaction and regulatory networks for both datasets; however, we found little evidence to support the utilization of pathogen-associated molecular pattern (PAMP) signaling. The tissue specimens were also enriched for transcripts governed by hypoxia-inducible factor. The approach presented here provides an uncomplicated means to infer global relationships among gene clusters involved in cellular responses to labor-associated signals.

Inflammatory and steroid receptor gene methylation in the human amnion and decidua

“…The expression of these genes, PTGS2, BMP2, NAMPT and CXCL2, increases during fetal membrane activation at labour (Haddad et al 2006, Johnson et al 2006, Mazaki-Tovi et al 2008, Li et al 2011, Lim et al 2012. Their promoters are rich in CpG motifs, which form clusters called CpG islands.…”

“…Array-based studies assessing the expression of a large number of genes simultaneously as well as studies determining the expression level of proinflammatory candidate genes have shown that the activity of many inflammatory genes increases at term even in the absence of histological inflammation or detectable infectious insult (Haddad et al 2006, Johnson et al 2006, Mazaki-Tovi et al 2008, Li et al 2011, Lim et al 2012. This changing pattern of gene expression has been described as an 'acute inflammation gene expression signature' (Haddad et al 2006), which transforms the membranes into an activated state producing uterotonic stimulants such as prostaglandins and undergoing structural remodelling in preparation for rupture during labour.…”

“…The aim of the present investigation was to explore this possibility by determining the CpG methylation density in the promoters of four inflammatory genes that show increased expression in the fetal membranes and decidua at term labour. These genes encode prostaglandin endoperoxide synthase-2 (PTGS2, the key prostaglandin biosynthetic enzyme), bone morphogenetic protein-2 (BMP2), nicotinamide phosphoribosyltransferase (NAMPT also called PBEF, or visfatin) and chemokine (C-X-C motif) ligand-2 (CXCL2) (Haddad et al 2006, Li et al 2011. We have assessed the methylation status of these genes in amnion collected from normal pregnancies at 11-17 weeks of gestation, at term (37-41 weeks) in the absence of labour and following term labour.…”

A Computational Model of Lipopolysaccharide-Induced Nuclear Factor Kappa B Activation: A Key Signalling Pathway in Infection-Induced Preterm Labour

“…The novel paradigm of “systems biology” provides a promising opportunity to overcome these restrictions and improve our understanding of the key molecular pathways that initiate labour in women [6]. Systems biology provides useful strategies to integrate the complex interactions within biological systems through building computational models.…”

The Transcription Factor Interferon Regulatory Factor-1 (IRF1) Plays a Key Role in the Terminal Effector Pathways of Human Preterm Labor1

“…There is a paucity of data on IRFs and human labor; although a recent publication demonstrated that using microarray analysis, IRF1 mRNA expression was increased following IL1B challenge in primary human amnion cells [33]. We hypothesized that 1) human labor and delivery would be associated with increased IRF1 expression in fetal membranes and/or myometrium, 2) proinflammatory stimuli would increase IRF1 transcriptional activity, and 3) inhibition of IRF1 using small interfering RNA (siRNA) would be associated with decreased expression and secretion of proinflammatory cytokines in the presence of infection or inflammation.…”