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 exposed to long-term oxidative stress at near physiological levels. Rabbit lenses, incubated in 50 microM H2O2 for 1 or 24 h, were checked for clarity and morphological changes in the epithelium. Proteolytic activity was measured in the epithelium using a fluorogenic synthetic substrate; N-succinyl-Leu-Tyr-7-amino-4-methylocoumarin, both in the presence and the absence of calcium (1 mM Ca2+ and 5 mM EDTA respectively). The effect on transparency and morphology of the epithelium following a 1-hour incubation in 100 microM H2O2 was also studied. Lenses incubated in 50 microM H2O2 were clear even after 24h. After a 1-hour incubation in 50 microM H2O2 the epithelium of the exposed lens appeared normal. However, after 24 h the epithelium cells appeared swollen and microscopical examination showed extensive intracellular and subepithelial vacuolization. Incubation in 100 microM H2O2 for 1 h caused loss of transparency; 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 microM H2O2, implying a role for the calcium-activated protease calpain in oxidatively induced cataract.
Andersson
M
M
Department of Histology, University of Göteborg, Sweden. madeleine.andersson@anatcell.gu.se
Sjöstrand
J
J
Petersen
A
A
Karlsson
J O
JO
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002154:Calpain / Q000378:metabolism*; D002386:Cataract / Q000139:chemically induced / Q000201:enzymology / Q000473:pathology; D004848:Epithelium / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D005260:Female; D006861:Hydrogen Peroxide / Q000633:toxicity; D007908:Lens, Crystalline / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D009924:Organ Culture Techniques; D018384:Oxidative Stress; D011817:Rabbits
D006861:Hydrogen Peroxide; D002154:Calpain; D002118:Calcium