Effect of HNE and Fenton reagents on the cytoskeleton of lens epithelial cell

Objective: To investigate the 4-hydroxynonenal(HNE) and Fenton reagent induced early eytoskeleton changes of human lens epithelial cells (HLECs) and rat lens epithelia. Methods: HLECs line were isolated and subcultured in DMEM medium with 20% fetal bovine serum and 20μg/ml gentamicin, and generation 20-30 of HLECs were collected for this experiment. Rat lens were treated with HNE and Fenton reagents. Apoptosis of HLECs was determined using cell death detection ELISA Kit. The opacity of rat lens was observed using image system. Antibodies of anti-cytoskeleton such as anti-tubulin, actin and vimentin were applied for the immunohistochemical staining and Western blot analysis in HLECs and rat lens epithelia. Results: HNE mediated apoptosis of HLECs at 1 hour and got its highest level at 2 hours. However, Fenton reagents resulted in apoptosis at 4 hours with its peak at 24 hours. 80% of cells death occurred at 4 hours in HNE and 27% at 8 hours in Fenton reagents. Opacity of rat lens was gradually increased during the incubation course with HNE and fenton reagents. After exposure to HNE and Fenton reagents, HLECs shrunk and transformed into round shape with the loss of cytoskeleton and its networks. Immunoreactive bands of vimentin, tubulin and actin were decreased in different time points. Disorder of cytoskeletons of the rat lens epithelia and decrease of staining intensity of tubulin and actin were detected. Conclusion: HNE and Fenton induce HLECs apoptosis and formation of rat cataract. Lens cytoskeleton may play a role in genesis of cataract.