Selenite May Cause Mitochondrial Damage and Cell Death in Cultured Lens Epithelial Cells

Abstract

Abstract: : Purpose: Our previous study suggested that calpain-induced proteolysis was an important mid-stage event in the death of lens epithelial cells following injection of a cataractous dose of sodium selenite into rats. However, the initial insult induced by selenite in lens epithelial cells was not clear. Further, activation of one or more calcium influx pathways was predicted. The purpose of the present study was to use lens epithelial cells cultured in selenite to identify pathways leading to lens epithelial cell death. Methods: α-TN4 cells were cultured in medium containing with 100 µM selenite for up to 16 hours. Leakage of lactate dehydrogenase (LDH) from cells to the medium was used as a marker for cell death. Leakage of cytochrome c from mitochondria to the cytosol was used as a marker for mitochondrial damage. Proteolysis of calpain-preferred substrate α-spectrin was detected by immunoblotting. Results: Leakage of LDH into the medium was increased when α-TN4 cells were cultured in selenite. α-spectrin was proteolyzed. Cytochrome c levels increased in the cytosol. The selenite-induced release of cytochrome c from the mitochondria suggested that calcium could also be released form organelles such as mitochondria and endoplasmic reticulum. Thus, α-TN4 cells were next cultured with calcium modulators thapsigargin and calcium ionophore A23187. Both reagents lead to proteolysis of α–spectrin. Conclusions: Results suggested that mitochondrial damage by selenite contributed to calcium accumulation in the cytosol (influx pathway), leading to calpain-induced proteolysis and subsequent cell death in cultured α-TN4 cells.

Keywords: cell death/apoptosis • calcium • proteolysis