Effect of exogenous stress on the tissue-cultured mouse lens epithelial cells

The effects of heat, oxidative and osmotic stress on heat shock proteins (HSP-70(I), HSC-70, and HSP-40 of tissue cultured mouse lens epithelial cells (alphaTN-4) were investigated. The effect of stress on the heat shock transcription factor (HSF-1), and a nuclear matrix protein (NM-60) of alphaTN-4 cells was also examined. Cells were exposed to heat (45 degrees C), oxidative stress (50 mM H(2)0(2)) and osmotic (600 mM medium) shock for 30 min, and then allowed to recover for 18 h in physiological medium. Control cells were maintained at 37 degrees C in an isosmolar medium. Cellular proteins were isolated and fractionated by SDS-PAGE. Western blot was used to determine the levels of HSP and nuclear proteins. Heat stressed cells were also examined, by immunofluorescence, for the specific localization of NM-60 and HSF-1. The results revealed that both NM-60 and HSF-I were present in specific locations in normal and heat-stressed cell nuclei. Nuclei isolated immediately after stress showed localization of fluorescence near the nuclear membrane. When heat stressed cells were allowed to recuperate at 37 degrees C, most of the fluorescence were relocated in discrete areas of the nucleus. These experiments showed that alphaTN-4 cells responded to stress by overexpression of HSP-70(I) and HSP-40. This increase was not present immediately after the end of the stress period, but clearly evident at 18 h of recovery in physiological medium. Immunofluorescent data suggest that heat stress induced the localization of NM-60 and HSF-1 near the nuclear membrane.