Article

Respiratory epithelial cell responses to cigarette smoke: The unfolded protein response

Division of Pulmonary, Allergy and Critical Care Medicine, Department of Medicine, Temple University School of Medicine, USA.
Pulmonary Pharmacology &amp Therapeutics (Impact Factor: 2.57). 07/2012; 25(6). DOI: 10.1016/j.pupt.2012.07.005
Source: PubMed

ABSTRACT Cigarette smoking exposes the respiratory epithelium to highly toxic, reactive oxygen nitrogen species which damage lung proteins in the endoplasmic reticulum (ER), the cell organelle in which all secreted and membrane proteins are processed. Accumulation of damaged or misfolded proteins in the ER, a condition termed ER stress, activates a complex cellular process termed the unfolded protein responses (UPR). The UPR acts to restore cellular protein homeostasis by regulating all aspects of protein metabolism including: protein translation and syntheses; protein folding; and protein degradation. However, activation of the UPR may also induce signaling pathways which induce inflammation and cell apoptosis. This review discusses the role of UPR in the respiratory epithelial cell response to cigarette smoke and the pathogenesis of lung diseases like COPD.

0 Followers
 · 
83 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Age-related macular degeneration is a complex disease caused by genetic and environmental factors, including genetic variants in complement components and smoking. Smoke exposure leads to oxidative stress, complement activation, endoplasmic reticulum (ER) stress, and lipid dysregulation, which have all been proposed to be associated with AMD pathogenesis. Here we examine the effects of smoke exposure on the retinal pigment epithelium (RPE). Mice were exposed to cigarette smoke or filtered air for 6 months. RPE cells grown as stable monolayers were exposed to 5% cigarette smoke extract (CSE). Effects of smoke were determined by biochemical, molecular and histological measures. Effects of the alternative pathway (AP) of complement and complement C3a anaphylatoxin receptor signaling were analyzed using knockout mice or specific inhibitors. ER stress markers were elevated after smoke exposure in RPE of intact mice, which was eliminated in AP-deficient mice. To examine this relationship further, RPE monolayers were exposed to CSE. Short-term smoke exposure resulted in production and release of complement C3, the generation of C3a, oxidative stress, complement activation on the cell membrane, and ER stress. Long-term exposure to CSE resulted in lipid accumulation, and secretion. All measures were reversed by blocking C3a complement receptor (C3aR), alternative complement pathway signaling, and anti-oxidant therapy. Taken together, our results provide clear evidence that smoke exposure results in oxidative stress and complement activation via the AP, resulting in ER stress-mediated lipid accumulation, and further suggesting that oxidative stress and complement act synergistically in the pathogenesis of AMD.
    Journal of Biological Chemistry 04/2014; 289(21). DOI:10.1074/jbc.M114.564674 · 4.60 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: A grape pomace extract enhanced antioxidant mechanisms in muscle and endothelial cells both in the absence and in the presence of oxidative stress-induced agent tert-butyl hydroperoxide (tBHP). In particular, muscle (C2C12) and endothelial (EA.hy926) cells were treated with the extract at noncytotoxic concentrations for 24 h, and the oxidative stress markers, total reactive oxygen species (ROS), glutathione (GSH), thiobarbituric reactive substances (TBARS), and protein carbonyl levels were assessed. The results showed that the grape extract treatment reduced significantly ROS, TBARS, and protein carbonyl levels and increased GSH in C2C12 cells, while it increased GSH and decreased protein carbonyl levels in EA.hy926 cells. In the presence of tBHP, the grape extract treatment in C2C12 cells reduced significantly ROS, TBARS, and protein carbonyls and increased GSH compared with tBHP alone treatment, while, in EA.hy926 cells, the extract decreased significantly TBARS and protein carbonyls but increased GSH. The antioxidant potency of the extract was different between muscle and endothelial cells suggesting that the antioxidant activity depends on cell type. Moreover, the antioxidant activity of the grape extract, in both cell lines, exerted, at least in part, through increase in GSH levels. The present work is the first to report the effects of grape extract shown for skeletal muscle cells.
    Human &amp Experimental Toxicology 09/2014; 33(11). DOI:10.1177/0960327114533575 · 1.41 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Endoplasmic reticulum stress plays a critical role to restore the homeostasis of protein production in eukaryotic cells. This vital process is hence involved in many types of diseases including COPD. PERK, one branch in the ER stress signaling pathways, has been reported to activate NRF2 signaling pathway, a known protective response to COPD. Based on this scientific rationale, we aimed to identify PERK activators as a mechanism to achieve NRF2 activation. In this report, we describe a phenotypic screening assay to identify PERK activators. This assay measures phosphorylation of GFP-tagged eIF2α upon PERK activation via a cell-based LanthaScreen technology. To obtain a robust assay with sufficient signal to background and low variation, multiple parameters were optimized including GFP-tagged eIF2α BacMam concentration, cell density and serum concentration. The assay was validated by a tool compound, Thapsigargin, which induces phosphorylation of eIF2α. In our assay, this compound showed maximal signal window of approximately 2.5-fold with a pEC50 of 8.0, consistent with literature reports. To identify novel PERK activators through phosphorylation of eIF2α, a focused set of 8,400 compounds was screened in this assay at 10 µM. A number of hits were identified and validated. The molecular mechanisms for several selected hits were further characterized in terms of PERK activation and effects on PERK downstream components. Specificity of these compounds in activating PERK was demonstrated with a PERK specific inhibitor and in PERK knockout mouse embryonic fibroblast (MEF) cells. In addition, these hits showed NRF2-dependent anti-oxidant gene induction. In summary, our phenotypic screening assay is demonstrated to be able to identify PERK specific activators. The identified PERK activators could potentially be used as chemical probes to further investigate this pathway as well as the link between PERK activation and NRF2 pathway activation.
    PLoS ONE 10(3):e0119738. DOI:10.1371/journal.pone.0119738 · 3.53 Impact Factor