Enrichment of Cis-Acting Regulatory Elements in Differentially Methylated Regions Following Lipopolysaccharide Treatment of Bovine Endometrial Epithelial Cells

Endometritis is an inflammatory disease that negatively influences fertility and is common in milk-producing cows. An in vitro model for bovine endometrial inflammation was used to identify enrichment of cis-acting regulatory elements in differentially methylated regions (DMRs) in the genome of in vitro-cultured primary bovine endometrial epithelial cells (bEECs) before and after treatment with lipopolysaccharide (LPS) from E. coli, a key player in the development of endometritis. The enriched regulatory elements contain binding sites for transcription factors with established roles in inflammation and hypoxia including NFKB and Hif-1 alpha. We further showed co-localization of certain enriched cis-acting regulatory motifs including ARNT, Hif-1 alpha, and NRF1. Our results show an intriguing interplay between increased mRNA levels in LPS-treated bEECs of the mRNAs encoding the key transcription factors such as AHR, EGR2, and STAT1, whose binding sites were enriched in the DMRs. Our results demonstrate an extraordinary cis-regulatory complexity in these DMRs having binding sites for both inflammatory and hypoxia-dependent transcription factors. Obtained data using this in vitro model for bacterial-induced endometrial inflammation have provided valuable information regarding key transcription factors relevant for clinical endometritis in both cattle and humans.