SIRT1 alleviates hyperglycemia-induced oxidative stress and dysfunction of limbal niche cells by deacetylating FOXO1

Diabetic keratopathy (DK) is a major cause of vision loss, with a complex pathogenesis. In particular, the mechanisms by which hyperglycemia affects the limbal remain unclear. The purpose of this study was to investigate the impact of hyperglycemia on oxidative stress and the function of limbal niche cells (LNCs), as well as the underlying molecular mechanisms. Human LNCs were cultured under high glucose conditions to simulate the diabetic microenvironment, and cell viability was assessed using CCK8 assay. The localization and expression of proteins were detected by immunofluorescence and immunohistochemistry. Reactive oxygen species (ROS) assay, mitochondrial membrane potential analysis, and senescence-associated β-galactosidase (SA-β-gal) staining were used to evaluate oxidative stress levels, mitochondrial function, and cell senescence, respectively. The expression of SIRT1, FOXO1, Ki67, CD73, CD90, CD105, SCF, PDGFRβ, PAX6, apoptosis and senescence markers were evaluated by reverse-transcription quantitative PCR (RT-qPCR) and western blotting (WB). The expression levels of ABCG2, p63, and PAX6 were decreased in the limbus of diabetic patients. In vitro, hyperglycemia reduced LNCs proliferation, induced oxidative stress and senescence, and decreased the expression of SCF, PDGFRβ, and PAX6. Hyperglycemia inhibited SIRT1 nuclear translocation and acetylation of FOXO1. SIRT1 activation promoted FOXO1 deacetylation, thereby reducing hyperglycemia-induced oxidative stress, senescence, and mitochondrial overload in LNCs. Inhibition of SIRT1 enhanced FOXO1 acetylation and the promotion of cellular senescence. Hyperglycemia regulated the biological and stem cell functions through the SIRT1/FOXO1 signaling pathway in LNCs. These findings reveal a new mechanism in the diabetic limbal niche microenvironment and provide a new direction for the treatment of DK.