Chlamydia trachomatis infection induces epithelial-mesenchymal transition in conjunctival epithelial cells

Abstract

Purpose : Epithelial-mesenchymal transition (EMT) of conjunctival epithelium may contribute to trachoma-associated conjunctival fibrosis. Epigenetic mechanisms have an important role in different disease etiologies, however, it is unknown whether they play a role during Chlamydia trachomatis (Ct) ocular infection. The aim of this study was to test the ability of Ct to induce EMT in human conjunctival epithelial (HCjE) cells in vitro and investigate whether alterations in gene expression after Ct infection are correlated with DNA methylation of EMT related marker genes.

Methods : mRNA and protein expression levels of EMT markers, including TGFβ1 and TGFβ2, E-cadherin (CDH1), fibronectin (FN1) and α-smooth muscle actin (ACTA2) were assayed by qPCR and Western blot analysis in HCjE cells infected with Ct serovar B. Promoters and gene-bodies of CDH1, FN1 and ACTA2 were analyzed for the presence of CpG islands using CpG Island Searcher software and thereafter changes in DNA methylation patterns of selected regions were examined with MSP, HRM and bisulfite sequencing.

Results : Infection of HCjE cells with Ct resulted in a statistically significant increase in TGFβ1 and TGFβ2 mRNA and significant reduction of mRNA expression of E-cadherin gene (CDH1), wherein CDH1 mRNA expression was 2.7-fold higher in a control than in Ct infected HCjE cells. Decrease in mRNA expression was accompanied with diminishment in protein level of E-cadherin after Ct infection. HRM data showed convincing difference between control and Ct infected level of DNA methylation with increase from 12.8% to 21.8% suggesting that CDH1 promoter region is subjected to alterations in DNA methylation in HCjE cells because of Ct infection.

Conclusions : Our data for the first time indicate a direct role of Ct infection in epigenetic regulation of human conjunctival epithelium. Studying the pathways of EMT in HCjE cells may help to develop new therapeutic strategies to treat conjunctival fibrosis.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.