Delay of Diabetic Cataract Progression by a Chemical Chaperone Trimethylamine N-Oxide through Alleviating Unfolded-Protein-Induced ER Stress

Oral administration of TMAO elevated aqueous humor TMAO levels and effectively delayed cataract progression in diabetic rats. • TMAO inhibited the upregulated migration, apoptosis, ROS production and epithelial-mesenchymal transition levels induced by high glucose in lens epithelial cells. • RNA-seq confirmed that the attenuation of endoplasmic reticulum stress is a key mechanism underlying the protective effects of TMAO against high-glucose-induced phenotypic changes in lens epithelial cells. • TMAO protected lens epithelial cells against high-glucose-induced endoplasmic reticulum stress by improving the solubility of unfolded protein. • TMAO shows potential as a safe and effective treatment for diabetic cataract by targeting endoplasmic reticulum stress, without accelerating diabetic disease progression. This study aims to investigate the protective effects of trimethylamine N-oxide (TMAO), a natural occurring chaperone, on delaying diabetic cataract (DC) progression. Rats were divided into 4 experimental groups: untreated controls, TMAO treatment only, streptozotocin-induced diabetic mellitus (DM), and DM with TMAO therapy. After 10 weeks, aqueous humor TMAO levels and lens opacity were assessed. Immunofluorescence analyses and Western blotting were utilized to evaluate epithelial-mesenchymal transition (EMT)-related genes in lens epithelial cells (LECs). In vitro, HLE-B3 cells were treated with or without TMAO under high glucose (HG) environment (50 mmol/L), then collected for RNA-seq. Apoptosis and reactive oxygen species (ROS) were evaluated by flow cytometry while endoplasmic reticulum stress (ERS)-related genes were observed by western blotting. Elevated aqueous humor TMAO levels were detected in DC rats, compared to controls. In vivo and ex vivo, TMAO reduced HG-induced lens opacity and EMT in LECs. In vitro experiments demonstrated that HG resulted in enhanced apoptosis, migration, ROS and EMT-related genes in HLE-B3 cells while TMAO addition diminished these effects. RNA-seq revealed that the inhibition of ERS was the key factor to reverse the impacts of HG on LECs by TMAO and mechanistic investigations suggested that ERS-related pathway were significantly inhibited in TMAO treated LECs under HG or tunicamycin-induced ERS. Moreover, TMAO improved the solubility of lens protein in DC rats. Our study pioneered the confirmation of the protective role of TMAO in DC through inhibiting lens protein unfolding, suppressing ERS, and alleviating ERS-induced apoptosis, ROS and EMT in LECs.