The superoxide dismutase mimetic tempol blunts diabetes-induced upregulation of NADPH oxidase and endoplasmic reticulum stress in a rat model of diabetic nephropathy

Endoplasmic reticulum (ER) stress contributes to progression of diabetic nephropathy, which promotes end-stage renal failure in diabetic patients. This study was undertaken to investigate the actions of tempol and ramipril, pharmacological agents that target the consequences of NADPH oxidase, on diabetic nephropathy in a rat model of type 1 diabetes, with an emphasis on markers of ER stress. Male Sprague-Dawley rats were injected intravenously with a single bolus of streptozotocin (55 mg/kg) to induce type 1 diabetes. An additional age-matched group of rats was administered with citrate vehicle as controls. After 4 weeks of untreated diabetes, rats received tempol (1.5 mM/kg/day subcutaneously, n=8), ramipril (1 mg/kg/day in drinking water, n=8) or remained untreated for an additional 4 weeks (n=7). After 8 weeks of diabetes in total, kidneys were collected for histological analysis, gene expression and protein abundance. Tempol and ramipril blunted diabetes-induced upregulation of NADPH oxidase isoforms (Nox4, Nox2, p47phox), accompanied by an amelioration of diabetes-induced glomerular injury (podocin, nephrin, Kim-1), tubulo-interstitial fibrosis (TGFβ1, TGFβ-R2, pSMAD3, α-SMA) and pro-inflammatory cytokines (TNFα, MCP-1, ANX-A1, FPR2) expression. In addition, the diabetes-induced renal ER stress, evidenced by increased expression of GRP-78 chaperone and stress-associated markers ATF4, TRB3, as well as XBP1s, phospho-p38 mitogen-activated protein kinase (MAPK) and 3-nitrotyrosination, were all attenuated by tempol and ramipril. These observations suggest that antioxidant approaches that blunt NADPH upregulation may attenuate diabetic nephropathy, at least in part by negatively regulating ER stress and inflammation, and hence ameliorating kidney damage.

157Schematic illustration of the potential mechanisms by which tempol and ramipril limit diabetic nephropathy. Diabetes increases activity of NADPH oxidase leading to diabetic nephropathy via an increase in generation of ROS, p38 MAPK, ER stress, inflammation and glomerular injury and fibrosis. Tempol inhibits gene expression of NADPH oxidase isoforms, limits ER stress and inflammation, and kidney damage and ROS while ramipril inhibits Angiotensin II, thus ameliorating diabetes-induced nephropathy.