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The novel triterpenoid RTA 408 protects human retinal pigment epithelial cells against H2O2-induced cell injury via NF-E2-related factor 2 (Nrf2) activation.

ABSTRACT: Oxidative stress-induced retinal pigment epithelial (RPE) cell damage is an important factor in the pathogenesis of age-related macular degeneration (AMD). Previous studies have shown that RTA 408, a synthetic triterpenoid compound, potently activates Nrf2. This study aimed to investigate the protective effects of RTA 408 in cultured RPE cells during oxidative stress and to determine the effects of RTA 408 on Nrf2 and its downstream target genes. Primary human RPE cells were pretreated with RTA 408 and then incubated in 200?M H2O2 for 6h. Cell viability was measured with the WST-8 assay. Apoptosis was quantitatively measured by annexin V/propidium iodide (PI) double staining and Hoechst 33342 fluorescent staining. Reduced (GSH) and oxidized glutathione (GSSG) were measured using colorimetric assays. Nrf2 activation and its downstream effects on phase II enzymes were examined by Western blot. Treatment of RPE cells with nanomolar ranges (10 and 100nM) of RTA 408 markedly attenuated H2O2-induced viability loss and apoptosis. RTA 408 pretreatment significantly protected cells from oxidative stress-induced GSH loss, GSSG formation and decreased ROS production. RTA 408 activated Nrf2 and increased the expression of its downstream genes, such as HO-1, NQO1, SOD2, catalase, Grx1, and Trx1. Consequently, the enzyme activities of NQO1, Grx1, and Trx1 were fully protected by RTA 408 pretreatment under oxidative stress. Moreover, knockdown of Nrf2 by siRNA significantly reduced the cytoprotective effects of RTA 408. In conclusion, our data suggest that RTA 408 protect primary human RPE cells from oxidative stress-induced damage by activating Nrf2 and its downstream genes.

SUBMITTER: Liu X

PROVIDER: S-EPMC4731949 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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The novel triterpenoid RTA 408 protects human retinal pigment epithelial cells against H2O2-induced cell injury via NF-E2-related factor 2 (Nrf2) activation.

Liu Xiaobin X Ward Keith K Xavier Christy C Jann Jamieson J Clark Abbot F AF Pang Iok-Hou IH Wu Hongli H

Redox biology 20151219

Oxidative stress-induced retinal pigment epithelial (RPE) cell damage is an important factor in the pathogenesis of age-related macular degeneration (AMD). Previous studies have shown that RTA 408, a synthetic triterpenoid compound, potently activates Nrf2. This study aimed to investigate the protective effects of RTA 408 in cultured RPE cells during oxidative stress and to determine the effects of RTA 408 on Nrf2 and its downstream target genes. Primary human RPE cells were pretreated with RTA ...[more]

PMID: 26773873

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Project description:Objective/Hypothesis: To assess the efficacy and mechanism of action of a novel approach to mitigate acute and chronic radiation toxicity in a validated animal model. We propose that synthetic triterpenoid RTA-408, an Nrf2 activator, activates additional protective pathways which stabilize soft tissue during radiation, thereby reducing necrosis and improves surgical outcomes in a subsequently created axial rotational flap. Methods: Experimental animal study utilizing Sprague-Dawley rats were divided into 3 cohorts: (i) radiation + DMSO (inert vehicle), (ii) radiation + RTA-408 (therapeutic drug), and (iii) no radiation + DMSO. Two main groups of rats were utilized in these experiments: (i) rotational flap experiment (n=40) and (ii) transcriptome analysis (n=9). In the first experiment all animals in the radiation cohorts underwent 40 Gy of radiation to the soft tissue of the abdomen. Then all three cohorts underwent surgery with construction of an inferior epigastric axial rotational flap, which was performed 30 days after resolution of acute injury from radiation in the radiation groups. Percentage of flap necrosis, the primary endpoint, was calculated by standardized measurement of tissue necrosis by blinded observers and vascular density measured by CD31 staining. In a second experiment, an additional three cohorts, with identical treatment as described above underwent serial punch biopsies of the abdominal skin before, during, and after radiation and drug/vehicle control treatment. Transcriptome analysis utilizing gene set enrichment analysis and digital PCR were performed at the various timepoints. Results: In the first experiment, 40 rats were divided into the three groups and underwent treatment accordingly. The average flap necrosis was 20% (95% CI, 16-45%) in the radiation control group, 3% (95% CI, 0-11%) in the non-irradiated control, and 3% (95% CI, 0.2-10%) in the radiation group treated with RTA-408. Vascular density was preserved in the treatment group as compared to the radiated control. Nine rats were included in the second experiment, and transcriptome analyses in the treatment group revealed robust activation of antioxidant pathways with induced expression of genes associated with hypoxia and adipogenesis/angiogenesis. Conclusions: Administration of RTA-408 during radiation treatment in a rat model resulted in transcriptome changes which appear to mitigate the toxic effects of radiation, preserving capillary networks and improving flap survival and tissue healing after subsequent surgery. Such reductions in toxicity may have broad implications for functional outcomes and salvage surgery following therapeutic irradiation.

Dataset Information

The novel triterpenoid RTA 408 protects human retinal pigment epithelial cells against H2O2-induced cell injury via NF-E2-related factor 2 (Nrf2) activation.

Publications

The novel triterpenoid RTA 408 protects human retinal pigment epithelial cells against H2O2-induced cell injury via NF-E2-related factor 2 (Nrf2) activation.

Similar Datasets

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