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HeeJung Ju, Yong Soo Byon, Ji Young Kwon, Young-Sik Yoo, Jun-Sub Choi, Jeewon Mok, Choun-Ki Joo; The Blockade of IL-6 counterparts the Osmolar Stress-induced Apoptotic Change and Junctional Instability in Human Conjunctival Epithelial Cells.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):400.
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© ARVO (1962-2015); The Authors (2016-present)
To determine the effect of osmolar stress on cell survival/apoptosis and intercellular junctions of conjunctival epithelial cells and evaluate the possible role of IL-6.
Wong-Kilourne-derived human conjunctival epithelial cell line (WK-hC) was used in this study. Confluent cells were incubated under different osmolarity (control, 400 mOSm and 500 mOSm) with or without neutralizing IL-6 antibody (50 ng/ml). The expression of IL-6 level was measured in the supernatant of each conditioned medium. Cell viability/apoptosis assay was performed using Annexin v/ PI, CCK-8 and TUNEL assay. Western blot was conducted to measure the abundance of apoptic markers, IL-6 related downstream signaling pathway (JAK/STAT/ERK), and junctional markers (ZO-1, Occludin, beta-catenin, E-cadherin). In addition, cellular morphology and expression of junctional molecules were assessed on immunocytochemistry.
The concentration of IL6 showed time, dose-dependent increase in cells treated with 400mOsm, 500mOsm, not in control. Twenty four hours incubation in 400mOsm, 500mOsm decreased conjunctival epithelial cell viability and increased apoptosis. IL-6 neutralizing antibody reduced apoptotic change and junctional instability of cells incubated in 400mOsm and 500mOsm based on the results of TUNEL, CCK assay and flow cytometry with Annexin V/ PI. IL-6 neutralizing antibody inhibited the activation of JAK-STAT signaling pathway and loss of junctional molecules (ZO-1, occludin, beta-catenin, and E-cadherin), which were induced by hyperosmolar stress.
Hyperosmolar condition induced apoptosis and junctional instability in conjunctival epithelial cells, along with increase of IL-6 production. IL-6 neutralizing antibody inhibited apoptosis and JAK-STAT signaling and loss of junctional proteins in hyperosmolar condition. These findings suggested that IL-6 may be involved in apoptotic change and desquamation of conjunctival epithelial cells in hyperosmolar stress.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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