Endometrial epithelial cells play a critical role in mediating inflammatory mechanisms key to bacterial clearance and tissue re-modelling postpartum. This study characterised innate immune gene expression by bovine endometrial epithelial cells from three animals in response to Escherichia coli, a common cause of bovine uterine disease. Expression of key innate immune genes, encoding Toll-like receptor 4 (TLR4), the transcription factor NFkB1, the chemokine interleukin 8 (IL8), inflammatory cytokines (interleukins IL1β, IL6; tumour necrosis factor, TNF), β-defensins (lingual antimicrobial peptides LAP, tracheal antimicrobial peptide TAP) and acute phase proteins (haptoglobin, HP; serum amyloid A, SAA3) was examined in endometrial epithelial cells stimulated with E. coli for 6 and 24 h using qRT-PCR. Expression of all genes was increased significantly (P < 0.05) 6 h post-stimulation. Expression of IL1b, TNF and SAA3 genes was increased by 121-, 357-and 721-fold, respectively (P < 0.05). Twenty four hours post-stimulation, IL1b, IL6, IL8, TNF and LAP gene expression was decreased compared to 6 h, whereas TAP and SAA3 expression was further increased to 209-and 3452-fold (P < 0.05). E. coli driven expression of immune effector genes demonstrates potent immune, antimicrobial and regulatory capacity of endometrial epithelial cells to respond to this pathogen.