Abstract
Background
Oxidative stress-induced cell ferroptosis occurs during the pathogenesis of diabetic retinopathy (DR), but the detailed molecular mechanisms are still unclear. The present study aimed to investigate this issue.
Materials and methods
The retinal pigment epithelium (RPE) was treated with high glucose (30 mM) in vitro to mimic the realistic conditions of DR progression in vivo. Cell viability was determined by MTT assay and trypan blue staining assay. Gene expressions were examined by Real-Time qPCR and Western Blot analysis. FCM was used to detect cell apoptosis and ROS generation. Dual-luciferase reporter gene system assay was used to verify the targeting sites.
Results
High glucose increased reactive oxygen species (ROS) levels, promoted cell ferroptosis, and suppressed cell proliferation and viability in RPE, which were reversed by co-treating cells with both a ferroptosis inhibitor ferrostatin-1 and an ROS scavenger, N-acetyl-L-Cysteine (NAC). In addition, we screened out a miR-338-3p/ASCT2 (SLC1A5) axis that played an important role in this process. Mechanistically, miR-338-3p targeted the 3’ untranslated regions (3’UTR) of SLC1A5 for its inhibition and degradation, and high glucose downregulated SLC1A5 by upregulating miR-338-3p in RPE cells. Next, the miR-338-3p inhibitor and SLC1A5 overexpression vectors were delivered into the RPE cells, and the following gain- and loss-of-function experiments validated that both miR-338-3p ablation and SLC1A5 upregulation abrogated the regulating effects of high glucose on cell proliferation, viability, ferroptosis and ROS production in RPE cells.
Conclusions
Collectively, data in the present study indicated that targeting the miR-338-3p/SLC1A5 axis could block high glucose-induced ferroptosis in RPE cells.
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The dataset(s) supporting the conclusions of this article is(are) included within the article (and its additional file(s)).
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Dr. Jing Zhou was responsible for the conception of this paper, and also conducted most of the investigation and drafted the manuscript. Dr. Caoyu Sun and Xu Dong provided technical supports, and helped to collect, analyze and visualize the data. Dr. Hui Wang proofread the manuscript, and was responsible for manuscript submission. All authors reviewed the manuscript.
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Zhou, J., Sun, C., Dong, X. et al. A novel miR-338-3p/SLC1A5 axis reprograms retinal pigment epithelium to increases its resistance to high glucose-induced cell ferroptosis. J Mol Histol 53, 561–571 (2022). https://doi.org/10.1007/s10735-022-10070-0
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DOI: https://doi.org/10.1007/s10735-022-10070-0