Lens endogenous peptide aA66-80 generates hydrogen peroxide and induces cell apoptosis

Abstract

In previous studies, we reported the presence of a large number of low-molecular-weight (LMW) peptides in aged and cataract human lens tissues. Among the LMW peptides, a peptide derived from aA-crystallin, aA66-80, was found in higher concentration in aged and cataract lenses. Additional characterization of the aA66-80 peptide showed beta sheet signature, and it formed well-defined unbranched fibrils. Further experimental data showed that aA66-80 peptide binds a-crystallin, impairs its chaperone function, and attracts additional crystallin proteins to the peptide a-crystallin complex, leading to the formation of larger light scattering aggregates. It is well established that Aß peptide exhibits cell toxicity by the generation of hydrogen peroxide. The aA66-80 peptide shares the principal properties of Aß peptide. Therefore, the present study was undertaken to determine whether the fibril-forming peptide aA66-80 has the ability to generate hydrogen peroxide. The results show that the aA66-80 peptide generates hydrogen peroxide, in the amount of 1.2 nM H2O2 per μg of aA66-80 peptide by incubation at 37oC for 4h. We also observed cytotoxicity and apoptotic cell death in aA66-80 peptide-transduced Cos7 cells. As evident, we found more TUNEL-positive cells in aA66-80 peptide transduced Cos7 cells than in control cells, suggesting peptide-mediated cell apoptosis. Additional immunohistochemistry analysis showed the active form of caspase-3, suggesting activation of the caspase-dependent pathway during peptide-induced cell apoptosis. These results confirm that the aA66-80 peptide generates hydrogen peroxide and promotes hydrogen peroxide-mediated cell apoptosis.