NAD+ Induces Autophagy of Neuro2a Cells by Increasing Oxidative Stress

Abstract

NAD+ plays important roles in various biological processes. Previous studies have shown that NAD+ treatment can significantly decrease genotoxic agent‐induced death of primary neurons and astrocytes, and NAD+ administration can markedly reduce ischemic brain damage in rodents. However, whether NAD+ treatment affects cell autophagy remains unknown. In this study we have found that treatment of NAD+ at concentrations from 100 μM to 10 mM can induce autophagy of Neuro2a cells, as assessed by the immunoblotting of LC 3 I/II conversion,, real‐time qPCR quantification of beclin‐1 expression, and monodansylcadaverine (MDC) staining. Our study has also indicated that oxidative stress mediates the effects of NAD+ on the autophagy of the cells: NAD+ induced increases in the superoxide levels in Neuro2a cells, and treatment of the cells with antioxidants attenuated the effects of NAD+ on the autophagy. We have further indicated the NAD+‐induced autophagy contributes to the NAD+‐induced decrease in the survival of the cells. Our experimental results have also argued against the possibility that NAD+ affects the autophagy through its degradation product ‐‐‐ nicotinamide. In summary, our study has provided the first evidence that NAD+ treatment can induce autophagy of such tumor cells as Neuro2a by generating oxidative stress, which contributes to the NAD+‐induced decrease in cell survival (Supported by a Key Research Grant of Shanghai Municipal Scientific Committee #08JC1415400, a National Key Basic Research ‘973 Program Grant’ #2010CB834306, a Shanghai Engineering Center Grant of Equipment and Technology of Physical Therapy for Major Diseases #08DZ2211200).