CALCIUM-DEPENDENT PROTEOLYSIS IN RABBIT LENS EPITHELIUM AFTER OXIDATIVE STRESS

The purpose of this study was to examine changes in calcium-dependent proteolytic activity in the lens epithelium from whole rabbit lenses e xposed to long-term oxidative stress at near physiological levels, Rab bit lenses, incubated in 50 mu M H2O2 for 1 or 24 h, were checked for clarity and morphological changes in the epithelium, Proteolytic activ ity was measured in the epithelium using a fluorogenic synthetic subst rate, N-succinyl-Leu-Tyr-7-amino-4-methylcoumarin, both in the presenc e and the absence of calcium (1 mM Ca2+ and 5 mM EDTA respectively), T he effect on transparency and morphology of the epithelium following a 1-hour incubation in 100 mu M H2O2 was also studied, Lenses incubated in 50 mu M H2O2 were clear even after 24 h, After a I-hour incubation in 50 mu M H2O2 the epithelium of the exposed lens appeared normal, H owever: after 24 h the epithelium cells appeared swollen and microscop ical examination showed extensive intracellular and subepithelial vacu olization, Incubation in 100 mu M H2O2 for 1 h caused loss of transpar ency; vacuole formation, globulization of the superficial lens fibers and death of the epithelial cells, There was a 55% increase in calcium -dependent proteolytic activity after 1 h in 50 mu M H2O2, implying a role for the calcium-activated protease calpain in oxidatively induced cataract.