Astaxanthin protects ARPE-19 cells from oxidative stress via upregulation of Nrf2-regulated phase II enzymes through activation of PI3K/Akt

Mol Vis. 2013 Jul 25;19:1656-66. Print 2013.

Abstract

Purpose: Oxidative stress on retinal pigment epithelial (RPE) cells is thought to play a crucial role in the development and progression of age-related macular degeneration. Astaxanthin (AST) is a carotenoid that shows significant antioxidant properties. This study was designed to investigate the protective effect of AST on ARPE-19 cells against oxidative stress and the possible underlying mechanism.

Methods: ARPE-19 cells exposed to different doses of H2O2 were incubated with various concentrations of AST and cell viability subsequently detected with the (4-[3-[4-iodophenyl]-2-4(4-nitrophenyl)-2H-5- tetrazolio-1,3-benzene disulfonate]; WST-1) assay. The apoptosis rate and intracellular levels of reactive oxygen species (ROS) were measured with flow cytometry. NAD(P)H quinine oxidoreductase 1 (NQO1), hemeoxygenase-1 (HO-1), glutamate-cysteine ligase modifier subunit (GCLM), and glutamate-cysteine ligase catalytic subunit (GCLC) expression were examined with real-time PCR and western blotting. The nuclear localization of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) protein and the expression levels of cleaved caspase-3 and protein kinase B proteins were evaluated with western blotting.

Results: AST clearly reduced H2O2-induced cell viability loss, cell apoptosis, and intracellular generation of ROS. Furthermore, treatment with AST activated the Nrf2-ARE pathway by inducing Nrf2 nuclear localization. Consequently, Phase II enzymes NQO1, HO-1, GCLM, and GCLC mRNA and proteins were increased. AST inhibited expression of H2O2-induced cleaved caspase-3 protein. Activation of the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway was involved in the protective effect of AST on the ARPE-19 cells.

Conclusions: AST protected ARPE-19 cells against H2O2-induced oxidative stress via Nrf2-mediated upregulation of the expression of Phase II enzymes involving the PI3K/Akt pathway.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / drug effects
  • Caspase 3 / metabolism
  • Cell Line
  • Cell Survival / drug effects
  • Cytoprotection / drug effects
  • Down-Regulation / drug effects
  • Enzyme Activation / drug effects
  • Epithelial Cells / cytology*
  • Epithelial Cells / drug effects
  • Epithelial Cells / enzymology
  • Gene Expression Regulation / drug effects
  • Glutamate-Cysteine Ligase / genetics
  • Glutamate-Cysteine Ligase / metabolism
  • Heme Oxygenase-1 / genetics
  • Heme Oxygenase-1 / metabolism
  • Humans
  • Hydrogen Peroxide / toxicity
  • Metabolic Detoxication, Phase II*
  • NAD(P)H Dehydrogenase (Quinone)
  • NF-E2-Related Factor 2 / metabolism*
  • Oxidative Stress / drug effects*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Protective Agents / chemistry
  • Protective Agents / pharmacology
  • Proto-Oncogene Proteins c-akt / metabolism
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Retinal Pigment Epithelium / cytology*
  • Signal Transduction / drug effects
  • Up-Regulation / drug effects*
  • Xanthophylls / chemistry
  • Xanthophylls / pharmacology

Substances

  • NF-E2-Related Factor 2
  • Protective Agents
  • RNA, Messenger
  • Xanthophylls
  • astaxanthine
  • Hydrogen Peroxide
  • Heme Oxygenase-1
  • NAD(P)H Dehydrogenase (Quinone)
  • NQO1 protein, human
  • Phosphatidylinositol 3-Kinases
  • Proto-Oncogene Proteins c-akt
  • Caspase 3
  • Glutamate-Cysteine Ligase
  • glutamate-cysteine ligase modifier subunit, human