N-acetyl tryptophan glucopyranoside (NATG) provides radioprotection to murine macrophage J774A.1 cells

ABSTRACT

Ionizing radiation induced perturbations in cellular redox homeostasis are manifested as cell cycle arrest, inflammatory response, and apoptosis. Present study was focused on determination of radioprotective efficacy of a secondary metabolite N-acetyl-l-tryptophan glucoside (NATG) isolated from radioresistant bacterium Bacillus sp. INM-1. Macrophage J774A.1 cells were treated with NATG (0.025–200?μg/ml) before γ-irradiation (10-50?Gy) and radioprotective efficacy in terms of cellular metabolic activity was assessed using MTT assay. Radiation-induced intracellular ROS generation and its inhibition by NATG (0.25?μg/ml) pretreatment was evaluated using 2',7'-dichlorodihydrofluorescein diacetate (H₂DCFDA) probe. Effects of NATG pretreatment with or without γ-irradiation (20?Gy) on cell cycle perturbations and apoptosis regulation was evaluated using flowcytometry. Results of the study displayed 0.25 and 50?μg/ml as effective dose (ED) and lethal dose (LD₅₀) dose of NATG. A significant (p?<?0.05) decrease in DCFDA fluorescence corresponding to decreased ROS levels was observed in NATG pretreated irradiated cells as compared to irradiated alone cells. Cell cycle progression analysis demonstrated decrease (～10%) in G_1, G₂, and S phase cells at 24?h post-treatment time interval in NATG pretreated cells as compared to control group. However, radiation-induced or NATG-induced treatment did not cause any alterations in G₂/M arrest. APO-BrDU analysis demonstrated significant (p?<?0.05) reduction in apoptosis level at 24?h time interval in NATG pretreated irradiated cells as compared to alone irradiated cells. Conclusively, present study suggests that NATG offers radioprotection by apoptosis inhibition mechanism without altering cell-cycle progression in J774A.1 cells. Further studies to evaluate detailed molecular mechanisms of radioprotection offered by NATG are ongoing.