Attenuation of oxygen fluctuation-induced endoplasmic reticulum stress in human lens epithelial cells

Abstract

Cataractogenic stresses are associated with the induction of endoplasmic reticulum (ER) stress. However, little is known about oxygen (O 2)-induced ER stress in the lens. Cataract research has focused on elevated levels of O 2 in lens epithelial cells (LECs). Excessive levels or a lack of O 2 are known to induce ER stress whereas chronic ER stress activates the unfolded protein response (UPR). The present study investigated the hypothesis that the fluctuation of O 2 levels induces a UPR, and may be controlled by maintaining human LECs (hLECs) in a specific concentration of O 2. Human LECs were cultured in different atmospheric levels of O 2. Hypoxic conditions were determined by the level of hypoxia‑inducible factor (HIF)-1α. 2',7'-Dichlorodihydrofluorescein diacetate and ethidium homodimer-1 staining were conducted to detect reactive oxygen species (ROS) and cell death, respectively. Protein blot analyses were performed with antibodies specific to antioxidant and UPR‑specific proteins. Reverse transcription‑quantitatative polymerase chain reaction assays were performed to quantify the mRNA levels of activated NF-E2‑related factor 2 (Nrf2) and kelch-like ECH-associated protein 1 (Keap1). The treatment of human LECs with 0 and 20% atmospheric O 2 activated Nrf2/Keap1. The LECs shifted to 1% atmospheric O 2 from 0, 4 or 20% for 24 h showed decreased levels of Keap1. By contrast, hLECs cultured in 1% atmospheric O 2 for 24 h and then shifted to 0, 4 or 20% O 2 exhibited a significant upregulation of Nrf2. These results suggest that oxidative stress proteins were not expressed in a 1% O 2 environment. The O 2 levels in the culture medium were equilibrated within 2 h in the cell culture plates. These results of LECs is ~1 % atmospheric O 2. Either 0 or 20% of atmospheric O 2 activated the UPR and the Nrf2/Keap1‑mediated antioxidant system in LECs and chronic exposure to O 2 fluctuation led to ROS production and cell death. This study revealed that O 2 fluctuation‑induced UPR/ER stress could be prevented by maintaining the cells in a 1% O 2 environment.