Modification of thymocytes membrane radiooxidative damage and apoptosis by eugenol

Radiation-induced membrane oxidative damage, generation of reactive oxygen species (ROS), and cellular apoptosis, as well as their modification by a natural antioxidant from plant source, eugenol, have been investigated in mouse thymocytes. Thymocyte suspension was irradiated by gamma-rays, and the malondialdehyde (MDA) formation was measured with the thiobarbituric acid reactive species (TBARS) method. The results showed an increase in MDA in irradiated (2 Gy) thymocytes, which was inhibited in samples treated with increasing concentrations of eugenol (10-200 microM) prior to irradiation. The concentration of eugenol required to inhibit half of the MDA formation (IC(50)) in irradiated thymocytes was 100 microM. A dose-dependent increase in the generation of ROS was observed in irradiated thymocytes (0.5-200 cGy) as measured by 2,7-dichlorodihydro fluorescein diacetate (DCH-FDA), which was, however, inhibited by eugenol administered before irradiation. Furthermore, experiments showed a significant decrease in thymocytes apoptosis by antioxidant as measured by annexin-V labeling protocol. The inhibition of apoptosis by membrane-localized antioxidants such as eugenol, isoeugenol, and alpha-tocopherol acetate was more effective than a cytosolic antioxidant such as ascorbic acid. The results suggest an effective prevention of membrane and cellular damage in irradiated thymocytes by eugenol. It is inferred that damage to membrane played a significant role in radiation-induced cellular apoptotic death, which was markedly modified by membrane-specific antioxidants.