September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Does hyperosmolarity induce an irreversible process leading to human corneal epithelial cell death?
Author Affiliations & Notes
  • Wendy Kam
    Schepens Eye Research Institute/Massachusetts Eye and Ear, Boston, Massachusetts, United States
  • David A Sullivan
    Schepens Eye Research Institute/Massachusetts Eye and Ear, Boston, Massachusetts, United States
  • Benjamin D Sullivan
    TearLab Corp., San Diego, California, United States
  • Manoj Venkiteshwar
    TearLab Corp., San Diego, California, United States
  • Footnotes
    Commercial Relationships   Wendy Kam, TearLab Corp. (R); David Sullivan, TearLab Corp. (R), TearLab Corp. (S); Benjamin Sullivan, TearLab Corp. (I), TearLab Corp. (E), TearLab Corp. (P); Manoj Venkiteshwar, TearLab Corp. (E)
  • Footnotes
    Support  Supported by an unrestricted gift from TearLab to Schepens Eye Research Institute, and the Margaret S. Sinon Scholar in Ocular Surface Research fund
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 6181. doi:
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    • Get Citation

      Wendy Kam, David A Sullivan, Benjamin D Sullivan, Manoj Venkiteshwar; Does hyperosmolarity induce an irreversible process leading to human corneal epithelial cell death?. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6181.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Tear film hyperosmolarity is a core mechanism of dry eye disease. Hyperosmolarity leads to increased ocular surface stress, friction, inflammation and damage, as well as symptoms of discomfort and visual impairment. These detrimental sequelae are mediated in large part through morphological and cytotoxic effects on corneal epithelial cells. We hypothesized that (1) short-term exposure to a hyperosmolar environment may alter cell morphology but not initiate an irreversible process leading to cell death, and (2) extended exposure to the same conditions would be cytotoxic.

Methods : Human corneal epithelial cells (gift from Dr. James Jester) were cultured in normal (290 mOsm/L) or hyperosmolar (308, 338, 400, 600 mOsm/L) keratinocyte serum-free medium for 1, 3, 6 and/or 24 hours. Cells were evaluated for appearance, apoptosis and death with light and fluorescent microscopes. Experiments included positive controls for DNA fragmentation and dead cells.

Results : Our findings demonstrate that hyperosmolarity induces morphological and cytotoxic effects in human corneal epithelial cells and that these responses are both dose- and time-dependent. Very few cells die after a 24-hour exposure to 290 or 308 mOsm solutions, cells exposed to 338 mOsm medium exhibit membrane blebs (i.e. a sign of impending cell death), the 400 mOsm condition kills a large percentage of cells, and the 600 mOsm medium causes complete cell death after 3, 6 or 24 hours. Cell death does not appear to be mediated primarily through apoptotic DNA fragmentation. Following a 1-hour exposure to the 600 mOsm medium, cells appear rounded, but still adherent and alive. Limiting this 600 mOsm exposure to 1 hour and replacing with 290 mOsm medium for 23 hours increases cell survival.

Conclusions : Our results indicate that chronic exposure to a hyperosmolar challenge will kill the majority of human corneal epithelial cells, whereas changes in cell morphology due to brief exposures can be largely ameliorated by normalization of the surrounding environment. Our findings also raise the possibility that hyperosmolar conditions may induce forms of death other than apoptosis (e.g. necrosis) in these cells.

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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