Abstract
Autophagy protects cells from intracellular pathogens, but can be exploited by some infectious agents to their benefit. Currently it is not known if bacteria induce autohpagy in cells of the cornea. The goal of this study was to develop an ocular surface autophagy reporter cell line and determine whether ocular bacterial pathogens influence host responses through autophagy induction. The cell line was made using lentivirus transduction of an LC3-GFP fusion protein in human corneal limbal epithelial (HCLE) cells. LC3-GFP puncta in HCLEs were induced by rapamycin and ammonium chloride treatments, and prevented by the autophagy inhibitors 3-methyladenine (3'MA) and bafilomycin. Importantly, secretomes from Escherichia coli, Serratia marcescens, Staphylococcus aureus, methicillin sensitive (MSSA) and resistant (MRSA), were found to induce autophagy, whereas other bacteria, including Acinetobacter baumannii, Achromobacter xylosoxidans, Enterococcus faecalis, Klebsiella pneumoniae, Moraxella sp., and Stenotrophomonas maltophilia, did not. Our data indicates differences between tested ocular isolates of MRSA and MSSA in the activation of autophagy. HCLEs treated with 3'MA were slightly more susceptible to cytotoxic factors produced by S. marcescens and MRSA keratitis isolates, by contrast, bafilomycin A1 treatment caused no difference. This work demonstrates the successful development and validation of an autophagy reporter corneal cell line and indicates differences between ocular bacterial isolates in the activation of autophagy.