Purpose Melatonin is hormone produced by pineal gland and has potent antioxidant function. The goal of this study is to investigate whether melatonin can protect hyperglycemia-induced oxidative injury to müller cells and to explore the mechanisms of its protection.

Methods Rat müller cells were treated with high glucose and different concentrations of melatonin for various time points. Phosphatidylinositol 3-kinase (PI3k) inhibitor LY294002 and melatonin receptor antagonist Luzindole were used under the administration of melatonin. Distributions of melatonin membrane receptors MT1, MT2 and nuclear translocation of the nuclear factor erythroid 2-related factor 2(Nrf2) were detected by immunocytochemistry. Expressions of MT1, MT2, Akt phosphorylation, Nrf2 including its downstream, glutamate-cysteine ligase(GCL) , and Heme oxygenase 1 (HO-1) were measured by Real time PCR and Western Blot analyses. Intracellular glutathione(GSH) content was detected by GSH Assay Kit. The vascular endothelial growth factor(VEGF) protein concentration was measured by Enzyme-Linked Immunoassay.

Results Both of the melatonin membrane receptors located in müller cells. High glucose decreased GSH synthesis while elevated MT1, MT2, HO-1 expressions and VEGF levels. Akt phosphorylation, HO-1 and Nrf2 expressions were induced by melatonin in a time and dose-dependent manner, but the effects decreased significantly by LY294002(PI3k inhibitor) or Luzindole (melatonin receptor antagonist). LY294002 also inhibited the induction of GSH synthesis by melatonin. The elevated VEGF secretion under high glucose could be reduced by melatonin which shown a concentration-dependent manner. Meanwhile, LY294002 further decreased the secretion.

Conclusion Melatonin augmented cellular antioxidant defense capacity through receptor-dependent antioxidant enzymes induction via the PI3K/Akt-Nrf2 signaling pathway, thereby protecting müller cells from oxidative injury. It may be a primary mechanism of antioxidative action of melatonin concerned with its therapeutic effectiveness in retinopathy.