Purpose: To investigate the role of lens epithelium- derived growth factor (LEDGF) in lens epithelial cells subjected to heat or oxidative stress and to determine the localization of LEDGF in lens epithelial cells at different temperatures.
Methods: Mouse lens epithelial cells overexpressing a fusion protein between a green fluorescent protein (GFP) and LEDGF (GFP-LEDGF) were cultured for up to 7 days at various temperatures or for 24 hours in the presence of hydrogen peroxide. Translocation of GFP-LEDGF in the lens epithelial cells was monitored with a fluorescence microscope. Cell survival was determined with the trypan blue dye exclusion test. Expression of heat shock proteins (Hsps) was studied with protein blot analyses using antibody probes.
Results: LEDGF was found in the cytosol of lens epithelial cells at 4 degrees C, 15 degrees C, and 28 degrees C, and in the nucleus and nucleolus at 37 degrees C. At 41 C, it was apparent at higher levels in the cytosol, nucleus, and nucleolus. Lens epithelial cells overexpressing LEDGF manifested resistance to thermal and oxidative stress. Levels of Hsp 27, and alphaB-crystallin were elevated in these cells. Depriving lens epithelial cells of LEDGF with anti-LEDGF antibodies reduced nuclear localization of LEDGF and induced cell death.
Conclusions: The overexpression of LEDGF in lens epithelial cells conferred resistance to thermal and oxidative stress. The mechanism of this resistance appears to involve the increased expression of Hsp 27 and alphaB-crystallin.