Chronic nicotine exposure augments renal oxidative stress and injury through transcriptional activation of p66shc.

BACKGROUND Chronic nicotine (Ch-NIC) exposure exacerbates ischemia/reperfusion (I/R)-induced oxidative stress and acute kidney injury (AKI), and mitochondrial production of reactive oxygen species (ROS) in cultured renal proximal tubule cells (RPTCs). Because Ser36-phosphorylated p66shc modulates mitochondrial ROS production and injury of RPTCs, we hypothesized that Ch-NIC exacerbates AKI by increasing stress-induced phosphorylation of p66shc. METHODS We first tested whether Ch-NIC augments I/R-AKI-induced expression and phosphorylation of p66shc in vivo. We then examined whether knocking down p66shc, or impairing its Ser36 phosphorylation or binding to cytochrome c, alters the effects of Ch-NIC on oxidative stress (H₂O₂)-induced production of ROS, mitochondrial depolarization and injury in RPTCs in vitro. RESULTS We found that Ch-NIC increased the expression of p66shc in the control and ischemic kidneys, but only increased its Ser36 phosphorylation after renal I/R. Knocking down p66shc or impairing phosphorylation of its Ser36 residue, via the S36A mutation (but not the phosphomimetic S36D mutation), blunted Ch-NIC + H2O2-dependent ROS production, mitochondrial depolarization and injury in RPTCs. Additionally, Ch-NIC + H2O2-dependent binding of p66shc to mitochondrial cytochrome c was attenuated by S36A mutation of p66shc, and impairing cytochrome c binding (via W134F mutation) abolished ROS production, mitochondrial depolarization and injury, while ectopic overexpression of p66shc (which mimics Ch-NIC treatment) augmented oxidant injury. We determined that Ch-NIC stimulates the p66shc promoter through p53- and epigenetic modification (promoter hypomethylation). CONCLUSIONS Ch-NIC worsens oxidative stress-dependent acute renal injury by increasing expression and consequent oxidative stress-dependent Ser36 phosphorylation of p66shc. Thus, targeting this pathway may have therapeutic relevance in preventing/ameliorating tobacco-related kidney injury.