ITCH regulates oxidative stress induced by high glucose through thioredoxin interacting protein in cultured human lens epithelial cells.

ABSTRACT: Thioredoxin (Trx) is an important protein that controls oxidative damage in almost all eukaryotic cells. Trx interaction protein (Txnip) has been reported to negatively regulate the bioavailability of Trx and inhibit its biological function. The E3 ubiquitin ligase ITCH can specifically degrade Txnip via ubiquitination. The apoptosis of human lens epithelial cells (HLECs), which are highly sensitive to redox caused by oxidative stress, is a significant factor for the development of sugar cataract in a high?glucose environment. However, whether Trx, Txnip and ITCH contribute to the progression of sugar cataracts and the underlying mechanisms remain unknown, and thus, identifying these were the aims of the present study. The present results suggested that the expression levels of Trx, Txnip and ITCH in HLECs cultured with different glucose concentrations were detected by reverse transcription?quantitative PCR and western blotting, and the apoptotic rate of the cells was detected by flow cytometry and superoxide detection assay. The interaction between ITCH and Txnip was determined by co?localization immunofluorescence and co?immunoprecipitation. In addition, a vector and small interfering RNA of ITCH were transfected to overexpress and knockdown ITCH, respectively, to alter the expression of downstream proteins and cell apoptosis. It was found that Txnip was highly expressed in cultured HLECs in high?glucose environment, and the antioxidative function of Trx was restricted and suppressed, thus promoting apoptosis. The overexpression of ITCH increased the expression of Trx and decreased oxidative stress and apoptosis by decreasing Txnip in cultured HLECs, while downregulation of ITCH significantly decreased the expression of Trx and enhanced oxidative stress and apoptosis. Therefore, the present results indicated that ITCH could regulate the apoptosis of HLECs that were cultured in high?glucose concentration and that it may be a treatment target for sugar cataract.

SUBMITTER: Jiang L 

PROVIDER: S-EPMC7533507 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature