42 - Critical Role of Peroxiredoxin 6 in the Repair of Peroxidized Lung Membrane Phospholipids

Abstract

Peroxidation of membrane lipids represents a severe threat to cellular integrity and its repair is crucial to prevent cell death. Peroxiredoxin 6 (Prdx6), a protein with both GSH peroxidase and phospholipase A2 (PLA2) activities, plays a major role in antioxidant defense of the lung, an organ with very low expression of GPx4.We investigated the role of Prdx6 in the repair of peroxidized cell membranes in intact mouse lungs and isolated pulmonary microvascular endothelial cells (PMVEC) treated with tert-butylhydroperoxide. Lipid peroxidation was evaluated by measurements of: a) thiobarbituric acid reactive substances (TBARS); b) oxidation of diphenyl-1-pyrenylphosphine (DPPP); and c) ferrous- xylenol orange (FOX). The exposure dose in the different models was varied to give a similar increase of about 4-fold over control in lipid peroxidation measurements at end-exposure. Values for lipid peroxidation returned to control levels within 2 h after oxidant removal in wild type (WT) lungs and PMVEC; however,there was essentially no repair in Pxdx6 null lungs or PMVEC. There also was no recovery in lungs and PMVEC that express H26A-Prdx6; this mutation abolishes the phospholipid hydroperoxidase function of Prdx6 but not its ability to reduce H2O2. An intermediate degree of repair was observed with lungs and PMVEC that express either C47S- or D140A-Prdx6; the former mutantion abolishes the peroxidase activity of Prdx6 while the latter abolishes PLA2 activity. Since repair was not seen with Prdx6 knock-out, GPx4 has no role in recovery from peroxidative stress by lung cells while Prdx6 plays a critical role; both the peroxidase and the PLA2 activities of Prdx6 contribute to the recovery process.