Lipid peroxidation of cell membrane is one of the major consequences of ROS overproduction,
which leads to the production of conjugated diene hydroperoxides and unstable substances
that disintegrate into various aldehydes like HNE and MDA, the biomarkers for lipid oxidative
damage. As evaluated by Western blot analysis, the protein production levels of HNE and
MDA in HCECs were significantly increased at an osmolarity-dependent manner after exposure
to hyperosmotic media with 400 and 450 mOsM for 24 h (Fig 2A). In contrast, the protein
levels of house-keeping protein β-actin were relatively stable without changes.
Immunofluorescent staining further showed that the immunoreactivity of HNE and MDA
was mainly present at cytoplasm or membrane, respectively, and their staining intensity dramatically
increased in HCECs exposed to 450 mOsM (Fig 2B).