Molecular mechanism of oxalate-induced free radical production and glutathione redox imbalance in renal epithelial cells: effect of antioxidants

Am J Nephrol. Sep-Oct 2004;24(5):557-68. doi: 10.1159/000082043. Epub 2004 Nov 10.

Abstract

Background: Peroxidation of renal cells is a critical event in the nucleation and formation of calcium oxalate crystals under hyperoxaluric conditions. We previously demonstrated that oxalate-induced peroxidative injury is one of the major mechanisms in promoting crystal attachment to renal epithelial cells.

Methods: In this study we have demonstrated that the mechanism of oxalate-induced peroxidative injury is through the induction of TGF-beta1 and glutathione (GSH) redox imbalance in LLC-PK1 cells.

Results: LLC-PK1, renal epithelial cells exposed to oxalate had significantly higher reactive oxygen species (ROS) production; higher TGF-beta1 levels, as measured by ELISA (1.89 +/- 0.035 fold increase) or Western blot (1.65 +/- 0.01 fold increase); increased malondialdehyde formation; increased LDH release, and loss of cell viability. In addition, oxalate exposure significantly decreased GSH content, glutathione reductase, glucose-6-phosphate dehydrogenase activities, and increased oxidized GSH content. Treatment with vitamin E, neutralizing anti-TGF-beta antibody, or diphenylene iodium, an inhibitor of NAD(P)H oxidase, significantly inhibited oxalate-induced ROS production and prevented peroxidative injury and cytolysis. Vitamin E, catalase, or desferoxamine treatment also significantly restored the oxalate-induced cellular GSH redox status toward the control level, and vitamin E treatment significantly attenuated the oxalate-mediated increase in TGF-beta1 protein in cultured LLC-PK1 cells.

Conclusions: This is the first study to demonstrate that the mechanism of oxalate-induced free radical production in renal tubular epithelial cells is through the activation of NAD(P)H oxidase via cytokine TGF-beta1 induction. These results also provide direct evidence that antioxidant therapy might prevent calcium oxalate nucleation and kidney stone formation by preventing oxalate-mediated peroxidative injury and GSH redox imbalance.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antioxidants / pharmacology*
  • Cell Survival / drug effects
  • Cells, Cultured
  • Epithelial Cells / drug effects*
  • Free Radicals / metabolism
  • Glutathione / metabolism*
  • Humans
  • Kidney / cytology*
  • Kidney / drug effects*
  • Oxalates / pharmacology*
  • Oxidation-Reduction / drug effects
  • Transforming Growth Factor beta / metabolism*
  • Transforming Growth Factor beta1
  • Urothelium / cytology
  • Urothelium / drug effects

Substances

  • Antioxidants
  • Free Radicals
  • Oxalates
  • TGFB1 protein, human
  • Transforming Growth Factor beta
  • Transforming Growth Factor beta1
  • Glutathione