Abstract

Objective- We investigated the role of cytochrome P4504A (Cyp4a), its metabolites and NAD(P)H oxidases in ROS production and apoptosis of podocytes exposed to high glucose and in OVE 26 mice a model of type1 diabetes.

Research Design and Methods- Apoptosis, albuminuria, ROS generation, NADPH superxide generation, Cyp4a and Nox protein expression and mRNA levels were measured in vitro and in vivo

Results- Exposure of mouse podocytes to high glucose (HG) results in apoptosis with approximately one third of the cells being apoptotic by 72 h. HG treatment increases ROS generation and is associated with sequential upregulation of Cyp4a, increase 20-HETE and Nox oxidases. This is consistent with the observation of delayed induction of NADPH oxidase activity by HG. The effects of HG on NADPH oxidase activity, Noxes protein and mRNA expression and apoptosis are blocked by HET0016, an inhibitor of Cyp4a and are mimicked by 20-HETE. Cyp4a and Nox oxidase expression is upregulated in glomeruli of type 1 diabetic OVE26 mice. Treatment of OVE26 mice with HET0016 decreases NAD(P)H oxidase activity, Nox1 and Nox4 protein expression and ameliorates apoptosis and albuminuria.

Conclusion- Generation of ROS by Cyp4a monooxygenases, 20-HETE and Nox oxidases is involved in podocyte apoptosis in vitro and in vivo. Inhibition of selected cytochrome P450 isoforms prevents podocyte apoptosis and reduces proteinuria in diabetes.