Metformin-induced protection against oxidative stress is associated with AKT/mTOR restoration in PC12 cells.

AIMS: Reactive oxygen species have been recognized to impair cell function through suppressing Akt the well-known pro-survival molecule. Pile of concrete evidence imply metformin as an Insulin sensitizer may enhance Akt/mTOR activity however the significance of Akt/mTOR recruitment has not yet been revealed in metformin induced neuroprotection against oxidative stress. MAIN METHODS: In the current study using H2O2 induced injury in PC12 cells; we first examined metformin impact on cell death by MTT assay and visual assessment. Metformin pretreated cells were then subjected to immunoblotting as well as real time PCR to find PI3K, Akt, mTOR and S6K concurrent transcriptional and post-transcriptional changes. The proportions of phosphorylated to non-phosphorylated constituents of PI3K/Akt/mTOR/S6K were determined to address their activation upon metformin treatment. KEY FINDINGS: According to cells morphology and MTT data metformin led to significant protection against H2O2 induced injury in 0.1 and 0.5mM concentrations. Metformin induced protection concurred with elevated PI3K/Akt/mTOR/S6K activity as well as enhanced GSH levels. These changes paralleled with a profound decline in the corresponding transcripts as determined by real time PCR. SIGNIFICANCE: Taken together our experimentation supports the hypothesis that Akt/mTOR/S6K cascade may contribute to metformin alleviating effect. The present work while highlighting metformin anti-oxidant characteristics, concludes that Akt/mTOR signaling might be central to the drug's alleviating effects.

Activation of S1P1 receptor regulates PI3K/Akt/FoxO3a pathway in response to oxidative stress in PC12 cells.

FTY720 (fingolimod) is a sphingosine analogue that, when phosphorylated, becomes a prototypical sphingosine-1-phosphate (S1P) receptor modulator. It can enter the CNS and act on S1PRs expressed by most neural lineages. Recently, FTY720 neuroprotective and regenerative actions in the CNS have been demonstrated. In the present study, we have investigated whether the PI3K-Akt-FoxO3a axis is downstream to the S1P1 receptor modulation and involved in the cytoprotective effect of FTY720 in PC12 cells exposed to hydrogen peroxide (H2O2). The data showed that oxidative stress induces cell death in parallel with a significant decrease in PI3K, Akt and Akt target, and FoxO3a phosphorylation. FTY720 pretreatment increased cell survival which can be attributed to enhanced levels of inactive phosphorylated FoxO3a, a transcription factor playing critical role in oxidative stress-induced cell death. FTY720-phosphate (p-FTY720), a pan agonist of S1P receptors, as well as SEW2871, a selective S1P1 receptor agonist, similarly exerted cytoprotective effects. W123, a S1P1 receptor antagonist, abolished the effects of all three drugs, and concomitant application of DMS, a sphingosine kinase inhibitor, prevented the protective effects of FTY720. The data suggests that S1P1 receptor activation in the context of oxidative stress maintains PI3K/Akt signaling to prevent activation of FoxO3a, thereby promoting PC12 cell survival.