Comparative effects of sulfhydryl compounds on target organellae, nuclei and mitochondria, of hydroxylated fullerene-induced cytotoxicity in isolated rat hepatocytes.

Abstract

DNA damage and cytotoxicity induced by a hydroxylated fullerene [C60 (OH)24 ], which is a spherical nanomaterial and/or a water-soluble fullerene derivative, and their protection by sulfhydryl compounds were studied in freshly isolated rat hepatocytes. The exposure of hepatocytes to C60 (OH)24 at a concentration of 50 μM caused time (0 to 3 h)-dependent cell death accompanied by the formation of cell surface blebs, the loss of cellular levels of ATP and reduced glutathione, accumulation of glutathione disulfide, and induction of DNA fragmentation assayed using alkali single-cell agarose-gel electrophoresis. C60 (OH)24 -induced cytotoxicity was effectively prevented by pretreatment with sulfhydryl compounds. N-acetyl-L-cysteine (NAC), L-cysteine and L-methionine, at a concentration of 2.5 mM, ameliorated cell death, accompanied by a decrease in cellular ATP levels, formation of cell surface blebs, induction of reactive oxygen species (ROS) and loss of mitochondrial membrane potential caused by C60 (OH)24 . In addition, DNA fragmentation caused by C60 (OH)24 was also inhibited by NAC, whereas an antioxidant ascorbic acid did not affect C60 (OH)24 -induced cell death and DNA damage in rat hepatocytes. Taken collectively, these results indicate that incubation of rat hepatocytes with C60 (OH)24 elicits DNA damage, suggesting that nuclei as well as mitochondria are target sites of the hydroxylated fullerene; and induction of DNA damage and oxidative stress is ameliorated by an increase in cellular GSH levels, suggesting that the onset of toxic effects may be partially attributable to a thiol redox-state imbalance caused by C60 (OH)24 .