PKR-dependent CHOP induction limits hyperoxia-induced lung injury

Abstract

Lozon TI, Eastman AJ, Matute-Bello G, Chen P, Hallstrand TS, Altemeier WA. PKR-dependent CHOP induction limits hyperoxia-induced lung injury. Am J Physiol Lung Cell Mol Physiol 300: L422-L429, 2011. First published December 24, 2010 doi:10.1152 doi:10. /ajplung.00166.2010 is commonly employed in patients with respiratory failure; however, hyperoxia is also a potential contributor to lung injury. In animal models, hyperoxia causes oxidative stress in the lungs, resulting in increased inflammation, edema, and permeability. We hypothesized that oxidative stress from prolonged hyperoxia leads to endoplasmic reticulum (ER) stress, resulting in activation of the unfolded protein response (UPR) and induction of CCAAT enhancer-binding protein homologous protein (CHOP), a transcription factor associated with cell death in the setting of persistent ER stress. To test this hypothesis, we exposed the mouse lung epithelial cell line MLE-12 to 95% O2 for 8 -24 h and evaluated for evidence of UPR induction and CHOP induction. Hyperoxia caused increased CHOP expression without other evidence of UPR activation. Because CHOP expression is preceded by phosphorylation of the ␣-subunit of the eukaryotic initiation factor-2 (eIF2␣), we evaluated the role of double-stranded RNA-activated protein kinase (PKR), a non-UPR-associated eIF2␣ kinase. Hyperoxia caused PKR phosphorylation, and RNA interference knockdown of PKR attenuated hyperoxia-induced CHOP expression. In vivo, hyperoxia induced PKR phosphorylation and CHOP expression in the lungs without other biochemical evidence for ER stress. Additionally, Ddit3 Ϫ/Ϫ (CHOP-null) mice had increased lung edema and permeability, indicating a previously unknown protective role for CHOP after prolonged hyperoxia. We conclude that hyperoxia increases CHOP expression via an ER stress-independent, PKRdependent pathway and that increased CHOP expression protects against hyperoxia-induced lung injury.CCAAT enhancer-binding protein homologous protein; acute respiratory distress syndrome; endoplasmic reticulum stress; epithelial cell; eukaryotic initiation factor-2␣; activating transcription factor-4; double-stranded RNA-activated protein kinase OXYGEN SUPPLEMENTATION is routinely used in the management of patients with acute respiratory failure. However, prolonged exposure to hyperoxia has long been recognized as a potential contributor to acute lung injury (26). In animals, prolonged exposure to hyperoxia causes lung injury, characterized by cell death, increased lung permeability, edema, and inflammation (5, 29). Although the exact mechanisms by which hyperoxia causes lung injury are incompletely understood, generation of reactive oxygen species (ROS) and cellular apoptosis appear to play important roles (1).One potential mechanism linking ROS generation and cell death is endoplasmic reticulum (ER) stress, resulting in activation of the associated unfolded protein response (UPR) (23). The ER functions to modify proteins for subsequent exposure to the extracellular environment. ER stress occurs ...

Hyperoxia and Interferon-γ–Induced Injury in Developing Lungs Occur via Cyclooxygenase-2 and the Endoplasmic Reticulum Stress–Dependent Pathway

“…However, BiP protein appeared to be increased in MLE-12 cells (Figures E1A and E1B). On the other hand, we noted a marked increase in CHOP protein ( Figure E1B), as also reported in an earlier study with MLE-12 cells (43). We found that CHOP siRNA decreased cell death in MLE-12 cells ( Figure E2).…”

“…In terms of hyperoxic exposure, an earlier study evaluated MLE-12 cells for 24 hours, and noted little change in BiP mRNA and protein concentrations (43). We noted no change in BiP mRNA.…”

“…In that study, the induction of CHOP was considered independent of the ER stress pathway because BiP concentrations did not alter, but was considered dependent on RNA-dependent protein kinase (PKR), because PKR phosphorylation was increased for up to 72 hours (43). In addition, they reported that CHOP was protective in hyperoxia-induced lung injury among adult mice (43).…”

Superoxide Dismutase 3 Dysregulation in a Murine Model of Neonatal Lung Injury

“…Although primary AEII cells would be a more accurate model to study O2 in vitro, the yield is often low during the isolation process, thus making analyzing various readouts difficult. However, there have been a few studies that have isolated murine primary AE II cells (40,49,50) with varying O2 exposure times between 24 to 48 hours. Although most described the overall cell survival and proliferation in such settings, none commented on the antioxidant profile associated with their findings.…”

Dual Functions for Cytosolic α-Mannosidase (Man2C1)

“…CHOP is often used as a marker for ER stress activation, and it is a major player in ER stress-triggered apoptosis (36 -38); nevertheless, the ER stress response can be induced independently of CHOP (44). Conversely, CHOP expression can be up-regulated independently of ER stress (45). Therefore, it appears possible that, following suppression of Man2C1 expression, CHOP is somehow up-regulated independently of ER stress.…”