p66SHC-mediated mitochondrial dysfunction in renal proximal tubule cells during oxidative injury

Abstract

Mitochondrial dysfunction is involved in pathopysiology of ischemia-reperfusion-induced acute kidney injury (AKI). The p66shc adaptor protein is a newly recognized mediator of mitochondrial dysfunction, which might play a role in AKI-induced renal tubular injury. Oxidative stress-mediated Serine36 phosphorylation of p66shc facilitates its transportation to the mitochondria where it oxidizes cytochrome c and generates excessive amount of reactive oxygen species (ROS). The consequence is mitochondrial depolarization and injury. Earlier we determined that p66shc plays an essential role in injury of cultured mouse renal proximal tubule cells during oxidative stress. Here, we studied the role of p66shc in ROS generation and consequent mitochondrial dysfunction during oxidative injury in renal proximal tubule cells. We employed p66shc knockdown renal proximal tubule cells and cells that overexpress wild-type, Serine phosphorylation (S36A), or cytochrome c-binding (W134F) mutants of p66shc. Inhibition of the mitochondrial electron transport chain or the mitochondrial permeability transition revealed that hydrogen peroxide-induced injury is mitochondrial ROS and consequent mitochondrial depolarization dependent. We also found that through Ser36 phosphorylation and mitochondria/cytochrome c binding, p66shc mediates those effects. We propose a similar mechanism in vivo as we demonstrated mitochondrial binding of p66shc as well as its association with cytochrome c in the postischemic kidneys of mice. Thus, manipulating p66shc might offer a new therapeutic modality to ameliorate renal ischemic injury.

p66Shc mediates high-glucose and angiotensin II-induced oxidative stress renal tubular injury via mitochondrial-dependent apoptotic pathway

“…They are also protected from the development of diabetic glomerulopathy, presumably due to a blockade of hyperglycemia-induced ROS production (23). Furthermore, it has been reported that the p66Shc mediates mitochondrial dysfunction in renal proximal tubule cells in the ischemia-reperfusion injury model (1), and ANG II induced apoptosis in myocardial cells (10). These literature reports certainly establish the role of p66Shc in states of oxidant stress.…”

Nicotine Exposure Augments Renal Toxicity of 5-azacytidine Through p66shc: Prevention by Resveratrol

“…p66sch is known as a pro-oxidant enzyme that links oxidative stress to renal injury (2,8,23,24). Interestingly, lately some studies -including ours (17)-revealed that under certain circumstances p66shc can serve as an antioxidant (25).…”

“…Extent of cell injury was determined by LDH release using the fluorescent "Cytotox-One Homogeneous Membrane Integrity" kit (Promega, Madison, WI) as described earlier (13). Briefly, cells grown in 96-well-plates were transfected and treated as described in the appropriate section and 24 h later LDH content of the supernatants were determined and compared to total 4075 This article is freely accessible online.…”

Resveratrol Attenuates Nicotine-mediated Oxidative Injury by Inducing Manganese Superoxide Dismutase in Renal Proximal Tubule Cells

“…Briefly, cells grown in T25 flasks and treated with 20 μM RES or vehicle overnight were collected, counted and loaded with 100 μM DCFDA for 30 min. After washing away the excess dye equal number (0.5×10 6 ) of cells were distributed in a 96-well plate; 200 μM NIC was added and ROS production was immediately determined as described elsewhere (15). ROS production was calculated as the increase in fluorescence/30 min/0.5×10 6 cells and expressed as a percentage of that of corresponding untreated cells.…”