Vitamin C and N-Acetyl-L-cysteine prevent ethanol induced cultured cerebellar granule neurons apoptosis through Nuclear Factor-kappa B pathway

摘要

Objective: Oxidative stress is a candidate mechanism for ethanol neuropathology in fetal alcohol spectrum disorders. Our overall objective was to investigate the hypothesis that ethanol neurotoxicity involves production of reactive oxygen species (ROS) with negative downstream consequences for mitochondrial membrane potential and neuron survival. Methods: We use P4 cerebellar granule neurons (CGNs) as the cell model. Methyl thiazolyl tetrazolium (MTT), lactate dehydrogenase (LDH) assay and Hoechst 33258 staining were used to investigate the apoptosis induced by ethanol. Hydroxyl radical and lipid peroxidation or glutathione peroxidase (GSH-Px) activity assay were used to study the effect of oxidative stress. Luciferase assay and western blot were used to show the mechanism of cell death caused by ethanol exposure or the function of antioxidants in CGNs after ethanol treatment. Results: Ethanol exposure can cause oxidative stress by increasing the content of hydroxyl radical and the level of lipid peroxidation, and decreasing GSH-Px activity in cultured CGNs. Ethanol also decreases the expression of Nuclear Factor-kappa B (NF-KB) and reduces its downstream genes expression such as Bcl-2 and Bcl-xL in CGNs. The inhibition of ethanol can be rescued by anti-oxidative reagent ascorbic acid (Vitamin C) and N-Acetyl-L-cysteine (NAC). It is likely that oxidative stress is the principal mechanism of ethanol neurotoxicity for CGNs during the stage of the brain development. Conclusion: Our data show that ethanol exposure can promote cell death by increasing oxidative stress and lowering the ability of both anti-oxidation and anti-apoptosis in CGNs. This damage can be attenuated by anti-oxidative reagents.