Global Metabonomic and Proteomic Analysis of Human Conjunctival Epithelial Cells (IOBA-NHC) in Response to Hyperosmotic Stress

Dry eye is a multifactorial inflammatory disease affecting the ocular surface. Tear hyperosmolarity in dry eye contributes to inflammation and cell damage. Recent research efforts on dry eye have been directed toward biomarker discovery for diagnosis, response to treatment, and disease mechanisms. This study employed a spontaneously immortalized normal human conjunctival cell line, IOBA-NHC, as a model to investigate hyperosmotic stress-induced changes of metabolites and proteins. Global and targeted metabonomic analyses as well as proteomic analysis were performed on IOBA-NHC cells incubated in serum-free media at 280 (control), 380, and 480 mOsm for 24 h. Twenty-one metabolites and seventy-six iTRAQ-identified proteins showed significant changes under at least one hyperosmotic stress treatment as compared with controls. SWATH-based proteomic analysis further confirmed the involvement of inflammatory pathways such as prostaglandin 2 synthesis in IOBA-NHC cells under hyperosmotic stress. This study is the first to identify glycerophosphocholine synthesis and O-linked beta-N-acetylglucosamine glycosylation as key activated pathways in ocular surface cells under hyperosmotic stress. These findings extend the current knowledge in metabolite markers of dry eye and provide potential therapeutic targets for its treatment.