Nasser Al Sabaani1. 1. Ophthalmology Department, College of Medicine, King Khalid University, Abha, Saudi Arabia.
Abstract
PURPOSE: Activation of p66Sch, an adaptor protein, is associated with oxidative stress and apoptosis and has been implicated in the pathogenesis of diabetes-induced retinal pigment epithelial cell damage and diabetic retinopathy. Exendin-4 is a glucagon-like protein that protects against diabetic retinopathy, but the mechanism of action is not well understood. This study aimed to investigate whether Exendin-4 could protect against high glucose-induced oxidative stress and apoptosis in the adult human retinal pigment epithelial-19 cell line by modulating levels and activation of p66Shc and to study the underlying mechanisms. MATERIALS AND METHODS: Adult human retinal pigment epithelial-19 cells were cultured under low (5 µM) or high glucose (100 µM) conditions in the presence or absence of Exendin-4 and with or without pre-incubation with Exendin-9-39, a glucagon-like peptide-1 receptor antagonist. RESULTS: In a dose-dependent manner, Exendin-4 inhibited high glucose-induced cell death and decreased levels of reactive oxygen species, lactate dehydrogenase release, and single single-stranded DNA. At the most effective concentration (100 µM), Exendin-4 reduced mitochondrial levels of phospho-p66Shc (Ser36), cytoplasmic levels of cleaved caspase-3 and cytochrome-c, and NADPH oxidase levels in high glucose-treated cells. It also increased levels of glutathione and magnesium superoxide dismutase and protein levels of magnesium superoxide dismutase but downregulated total protein levels of protein kinase-β and p66Shc and inhibited c-Jun N-terminal kinase phosphorylation in both low- and high glucose-treated cells. All these Exendin-4 effects, however, were inhibited by Exendin-9-39. CONCLUSIONS: Exendin-4 protects against high glucose-induced adult human retinal pigment epithelial-19 cell damage by increasing antioxidants, downregulating NADPH, and inhibiting mitochondria-mediated apoptosis, effects that are associated with the inhibition of c-Jun N-terminal kinase and downregulation of protein kinase-β and p66Shc.
PURPOSE: Activation of p66Sch, an adaptor protein, is associated with oxidative stress and apoptosis and has been implicated in the pathogenesis of diabetes-induced retinal pigment epithelial cell damage and diabetic retinopathy. Exendin-4 is a glucagon-like protein that protects against diabetic retinopathy, but the mechanism of action is not well understood. This study aimed to investigate whether Exendin-4 could protect against high glucose-induced oxidative stress and apoptosis in the adult human retinal pigment epithelial-19 cell line by modulating levels and activation of p66Shc and to study the underlying mechanisms. MATERIALS AND METHODS: Adult human retinal pigment epithelial-19 cells were cultured under low (5 µM) or high glucose (100 µM) conditions in the presence or absence of Exendin-4 and with or without pre-incubation with Exendin-9-39, a glucagon-like peptide-1 receptor antagonist. RESULTS: In a dose-dependent manner, Exendin-4 inhibited high glucose-induced cell death and decreased levels of reactive oxygen species, lactate dehydrogenase release, and single single-stranded DNA. At the most effective concentration (100 µM), Exendin-4 reduced mitochondrial levels of phospho-p66Shc (Ser36), cytoplasmic levels of cleaved caspase-3 and cytochrome-c, and NADPH oxidase levels in high glucose-treated cells. It also increased levels of glutathione and magnesium superoxide dismutase and protein levels of magnesium superoxide dismutase but downregulated total protein levels of protein kinase-β and p66Shc and inhibited c-Jun N-terminal kinase phosphorylation in both low- and high glucose-treated cells. All these Exendin-4 effects, however, were inhibited by Exendin-9-39. CONCLUSIONS: Exendin-4 protects against high glucose-induced adult human retinal pigment epithelial-19 cell damage by increasing antioxidants, downregulating NADPH, and inhibiting mitochondria-mediated apoptosis, effects that are associated with the inhibition of c-Jun N-terminal kinase and downregulation of protein kinase-β and p66Shc.
Entities:
Keywords:
Exendin-4; high glucose; oxidative stress; p66Shc; retina