Original Contribution
Mechanism for the protective effect of resveratrol against oxidative stress-induced neuronal death

https://doi.org/10.1016/j.freeradbiomed.2010.06.002Get rights and content

Abstract

Oxidative stress can induce cytotoxicity in neurons, which plays an important role in the etiology of neuronal damage and degeneration. This study sought to determine the cellular and biochemical mechanisms underlying resveratrol's protective effect against oxidative neuronal death. Cultured HT22 cells, an immortalized mouse hippocampal neuronal cell line, were used as an in vitro model, and oxidative stress and neurotoxicity were induced in these neuronal cells by exposure to high concentrations of glutamate. Resveratrol strongly protected HT22 cells from glutamate-induced oxidative cell death. Resveratrol's neuroprotective effect was independent of its direct radical scavenging property, but instead was dependent on its ability to selectively induce the expression of mitochondrial superoxide dismutase (SOD2) and, subsequently, reduce mitochondrial oxidative stress and damage. The induction of mitochondrial SOD2 by resveratrol was mediated through the activation of the PI3K/Akt and GSK-3β/β-catenin signaling pathways. Taken together, the results of this study show that up-regulation of mitochondrial SOD2 by resveratrol represents an important mechanism for its protection of neuronal cells against oxidative cytotoxicity resulting from mitochondrial oxidative stress.

Section snippets

Materials

Resveratrol, Dulbecco's modified Eagle's medium (DMEM), fetal bovine serum (FBS), and trypsin–EDTA solution (containing 0.5 g/L trypsin and 0.2 g/L EDTA) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). Resveratrol was dissolved in 200-proof ethanol and stored at − 20°C. The antibiotic solution (containing 10,000 U/ml penicillin and 10 mg/ml streptomycin) was obtained from Gibco (Invitrogen, Grand Island, NY, USA). The inhibitors of mitogen-activated protein kinases (MAPKs) and PI3K

Resveratrol protects HT22 cells against glutamate-induced oxidative cytotoxicity

When HT22 cells were cultured in the presence of increasing concentrations of glutamate (2, 4, 6, 8, and 10 mM) for 24 h, cell viability was decreased in a concentration-dependent manner (Fig. 1A). The presence of 4 mM glutamate reduced cell viability over 80%. The presence of resveratrol alone (at 1, 5, 10, and 20 μM) caused a weak but concentration-dependent decrease in cell viability (MTT assay), which was due to a transient, noncytotoxic S-phase delay induced by resveratrol [26].

Discussion

The results of this study showed that resveratrol could strongly protect HT22 cells from glutamate-induced oxidative cytotoxicity by removing intracellular ROS. To determine whether resveratrol removed ROS directly (i.e., through its direct antioxidant activity) or indirectly (i.e., through the induction of cellular antioxidant capacity), we conducted two parallel experiments: one sought to determine whether the copresence of resveratrol with hydrogen peroxide could protect against hydrogen

Acknowledgments

We thank Dr. Joyce Slusser for technical assistance with the flow cytometric analysis and Stephanie C. Bishop for proofreading the manuscript. This work was supported, in part, by an NIH grant (ES015242). Some of the analytical and imaging instruments employed in this study are part of the COBRE core facility that is supported by NIH Grant P20RR021940 from the National Center for Research Resources.

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