{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T11:10:16Z","timestamp":1785409816340,"version":"3.56.0"},"reference-count":47,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2026,4,21]],"date-time":"2026-04-21T00:00:00Z","timestamp":1776729600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"College of Medicine at the University of Saskatchewan"},{"id":[{"id":"https:\/\/ror.org\/010x8gc63","id-type":"ROR","asserted-by":"publisher"}]},{"name":"two College of Medicine Research Awards"},{"DOI":"10.13039\/501100000038","name":"Natural Sciences and Engineering Research Council of Canada","doi-asserted-by":"publisher","award":["DGECR-2023-00003"],"award-info":[{"award-number":["DGECR-2023-00003"]}],"id":[{"id":"10.13039\/501100000038","id-type":"DOI","asserted-by":"publisher"}]},{"award":["DGECR-2023-00003"],"award-info":[{"award-number":["DGECR-2023-00003"]}],"id":[{"id":"https:\/\/ror.org\/01h531d29","id-type":"ROR","asserted-by":"publisher"}]},{"name":"Canadian Foundation for Innovation (CFI) John R. Evans Leaders Fund","award":["42810"],"award-info":[{"award-number":["42810"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biomolecules"],"abstract":"<jats:p>Cigarette smoke (CS) is the primary risk factor for the development of chronic obstructive pulmonary disease (COPD). Investigating the impact of CS on human airway epithelium is important for understanding COPD development and combating its effects. While some studies show that long exposure to CS activates inflammasome formation in airway epithelium, leading to cytokines\u2019 maturation and release, its acute effect on inflammation regulation requires further elucidation. Due to the importance of acute cellular responses in modulating cell survival and controlling inflammatory outcomes, we examined the effect of acute cigarette smoke extract exposure on human bronchial epithelial cells. Due to the high reactive oxygen species content in CS, we hypothesize that acute CS exposure activates the integrated stress response (ISR) pathway leading to stress granules (SG) formation to facilitate oxidative stress resolution and promote cell survival. Immunostaining, fluorescence confocal imaging, quantitative analyses, and immunoblotting were performed to test our hypothesis. We report here that acute exposure to CS extract triggers canonical SG formation by activating the ISR pathway via the PERK\/eIF2\u03b1 arm in a reactive oxygen species-dependent manner. SG formation is abolished upon inhibiting PERK or eIF2\u03b1 function, or by scavenging oxidants prior to smoke exposure. Characterizing SG formation in terms of measuring SG size and abundance and the sequestration of the SG marker G3BP1 reveals that SG formation is maximal at 15% CS extract exposure for 2 h and undergoes gradual disassembly at longer exposure times. This is closely dependent on cytoplasmic p-eIF2\u03b1 levels. These results demonstrate that acute exposure to CS activates the protective ISR pathway to potentially reduce the detrimental effects of CS and promote stress resolution and cell survival.<\/jats:p>","DOI":"10.3390\/biom16040615","type":"journal-article","created":{"date-parts":[[2026,4,21]],"date-time":"2026-04-21T11:42:45Z","timestamp":1776771765000},"page":"615","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Cigarette Smoke Induces Canonical Stress Granule Formation in Human Bronchial Epithelial Cells in Reactive Oxygen Species- and PERK-Dependent Manners"],"prefix":"10.3390","volume":"16","author":[{"given":"Mousumi","family":"Bhowmik","sequence":"first","affiliation":[{"name":"Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chenkun","family":"Zheng","sequence":"additional","affiliation":[{"name":"Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8773-7467","authenticated-orcid":false,"given":"Bisrat","family":"Bekele","sequence":"additional","affiliation":[{"name":"Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jessica","family":"Failler","sequence":"additional","affiliation":[{"name":"Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carlie","family":"Klatt","sequence":"additional","affiliation":[{"name":"Regina General Hospital, Regina Campus, University of Saskatchewan, Regina, SK S4P 0W5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Souren","family":"Farimani","sequence":"additional","affiliation":[{"name":"Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, QC H3A 2B4, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bryant","family":"Jones","sequence":"additional","affiliation":[{"name":"School of Rehabilitation Science, University of Saskatchewan, Saskatoon, SK S7N 2Z4, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chung-Chun","family":"Tyan","sequence":"additional","affiliation":[{"name":"Division of Respirology, Critical Care and Sleep Medicine, University of Saskatchewan, Saskatoon, SK S7N 0W8, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7069-8836","authenticated-orcid":false,"given":"Asmahan","family":"Abu-Arish","sequence":"additional","affiliation":[{"name":"Department of Anatomy, Physiology and Pharmacology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,21]]},"reference":[{"key":"ref_1","unstructured":"WHO (2019). The Top 10 Causes of Death, WHO."},{"key":"ref_2","unstructured":"Hermus, G., Stonebridge, C., Goldfarb, D., Th\u00e9riault, L., and Bounajm, F. (2012). Cost Risk Analysis for Chronic Lung Disease in Canada, Conference Board of Canada."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"497","DOI":"10.1016\/S2213-2600(21)00506-3","article-title":"Chronic obstructive pulmonary disease in never-smokers: Risk factors, pathogenesis, and implications for prevention and treatment","volume":"10","author":"Yang","year":"2022","journal-title":"Lancet Respir. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1","DOI":"10.2174\/1874306401206010001","article-title":"Asthma and COPD\u2014The C\/EBP Connection","volume":"6","author":"Miglino","year":"2012","journal-title":"Open Respir. Med. J."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"C1374","DOI":"10.1152\/ajpcell.00356.2022","article-title":"COPD and the modulation of CFTR by acute exposure to cigarette smoke","volume":"323","author":"Hanrahan","year":"2022","journal-title":"Am. J. Physiol. Cell Physiol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"C118","DOI":"10.1152\/ajpcell.00099.2017","article-title":"Cigarette smoke activates CFTR through ROS-stimulated cAMP signaling in human bronchial epithelial cells","volume":"314","author":"Wong","year":"2018","journal-title":"Am. J. Physiol. Cell Physiol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"2200898","DOI":"10.1183\/13993003.00898-2022","article-title":"Cystic fibrosis transmembrane conductance regulator (CFTR): A missing link between smoking and chronic airway diseases?","volume":"60","author":"Burgel","year":"2022","journal-title":"Eur. Respir. J."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"L530","DOI":"10.1152\/ajplung.00039.2013","article-title":"Cigarette smoke and CFTR: Implications in the pathogenesis of COPD","volume":"305","author":"Rab","year":"2013","journal-title":"Am. J. Physiol. Lung Cell Mol. Physiol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1321","DOI":"10.1164\/rccm.201304-0733OC","article-title":"Cigarette smoke induces systemic defects in cystic fibrosis transmembrane conductance regulator function","volume":"188","author":"Raju","year":"2013","journal-title":"Am. J. Respir. Crit. Care Med."},{"key":"ref_10","first-page":"1309746","article-title":"Airway Epithelium Dysfunction in Cystic Fibrosis and COPD","volume":"2018","author":"Molloy","year":"2018","journal-title":"Mediators Inflamm."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"147","DOI":"10.1016\/j.ccm.2015.11.003","article-title":"Acquired Cystic Fibrosis Transmembrane Conductance Regulator Dysfunction in Chronic Bronchitis and Other Diseases of Mucus Clearance","volume":"37","author":"Raju","year":"2016","journal-title":"Clin. Chest Med."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"150","DOI":"10.1513\/AnnalsATS.201805-325RL","article-title":"Acquired Cystic Fibrosis Transmembrane Conductance Regulator Dysfunction and Radiographic Bronchiectasis in Current and Former Smokers: A Cross-Sectional Study","volume":"16","author":"Teerapuncharoen","year":"2019","journal-title":"Ann. Am. Thorac. Soc."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1224","DOI":"10.1016\/j.bbagen.2015.02.004","article-title":"Cigarette smoke exposure reveals a novel role for the MEK\/ERK1\/2 MAPK pathway in regulation of CFTR","volume":"1850","author":"Xu","year":"2015","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"631","DOI":"10.1164\/rccm.202109-2064TR","article-title":"Cystic Fibrosis Transmembrane Conductance Regulator: Roles in Chronic Obstructive Pulmonary Disease","volume":"205","author":"Dransfield","year":"2022","journal-title":"Am. J. Respir. Crit. Care Med."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"119090","DOI":"10.1016\/j.lfs.2021.119090","article-title":"Cigarette smoke extract induces pyroptosis in human bronchial epithelial cells through the ROS\/NLRP3\/caspase-1 pathway","volume":"269","author":"Zhang","year":"2021","journal-title":"Life Sci."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Huot-Marchand, S., Nascimento, M., Culerier, E., Bourenane, M., Savigny, F., Panek, C., Serdjebi, C., Le Bert, M., Quesniaux, V.F.J., and Ryffel, B. (2022). Cigarette smoke-induced gasdermin D activation in bronchoalveolar macrophages and bronchial epithelial cells dependently on NLRP3. Front. Immunol., 13.","DOI":"10.3389\/fimmu.2022.918507"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1186\/s12950-021-00286-4","article-title":"AIM2 nuclear exit and inflammasome activation in chronic obstructive pulmonary disease and response to cigarette smoke","volume":"18","author":"Tran","year":"2021","journal-title":"J. Inflamm."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"111832","DOI":"10.1016\/j.intimp.2024.111832","article-title":"The activation of the AIM2 inflammasome after cigarette smoke exposure leads to an immunosuppressive lung microenvironment","volume":"131","author":"Colarusso","year":"2024","journal-title":"Int. Immunopharmacol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"155220","DOI":"10.1016\/j.cyto.2020.155220","article-title":"Cigarette smoke and extracellular Hsp70 induce secretion of ATP and differential activation of NLRP3 inflammasome in monocytic and bronchial epithelial cells","volume":"135","author":"Rumora","year":"2020","journal-title":"Cytokine"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1261","DOI":"10.4049\/jimmunol.1301676","article-title":"Apoptosis-associated speck-like protein containing a CARD forms specks but does not activate caspase-1 in the absence of NLRP3 during macrophage swelling","volume":"194","author":"Compan","year":"2015","journal-title":"J. Immunol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"792","DOI":"10.1016\/j.cell.2016.03.046","article-title":"Inflammasome Complexes: Emerging Mechanisms and Effector Functions","volume":"165","author":"Rathinam","year":"2016","journal-title":"Cell"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"345","DOI":"10.1289\/ehp.8347345","article-title":"Electron-spin resonance study of mainstream and sidestream cigarette smoke: Nature of the free radicals in gas-phase smoke and in cigarette tar","volume":"47","author":"Pryor","year":"1983","journal-title":"Environ. Health Perspect."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1111\/resp.13722","article-title":"Hypoxia-inducible factor and bacterial infections in chronic obstructive pulmonary disease","volume":"25","author":"Shukla","year":"2020","journal-title":"Respirology"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"524","DOI":"10.1165\/rcmb.2009-0054OC","article-title":"Deletion of Keap1 in the lung attenuates acute cigarette smoke-induced oxidative stress and inflammation","volume":"42","author":"Blake","year":"2010","journal-title":"Am. J. Respir. Cell Mol. Biol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1374","DOI":"10.15252\/embr.201642195","article-title":"The integrated stress response","volume":"17","author":"Koryga","year":"2016","journal-title":"EMBO Rep."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1095","DOI":"10.1038\/s41401-023-01225-0","article-title":"Mammalian integrated stress responses in stressed organelles and their functions","volume":"45","author":"Lu","year":"2024","journal-title":"Acta Pharmacol. Sin."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"16129","DOI":"10.1038\/s41598-017-16089-8","article-title":"Inhibition of the inflammatory response to stress by targeting interaction between PKR and its cellular activator PACT","volume":"7","author":"Dabo","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"15427","DOI":"10.1074\/jbc.274.22.15427","article-title":"RAX, a cellular activator for double-stranded RNA-dependent protein kinase during stress signaling","volume":"274","author":"Ito","year":"1999","journal-title":"J. Biol. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"105","DOI":"10.1016\/j.tox.2011.06.005","article-title":"Induction of CCAAT\/enhancer-binding protein-homologous protein by cigarette smoke through the superoxide anion-triggered PERK-eIF2\u03b1 pathway","volume":"287","author":"Tagawa","year":"2011","journal-title":"Toxicology"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Park, J.M., Jeong, H., Seo, Y.S., Do, V.Q., Choi, S.J., Lee, K., Choi, K.C., Choi, W.J., and Lee, M.Y. (2021). Cigarette Smoke Extract Produces Superoxide in Aqueous Media by Reacting with Bicarbonate. Toxics, 9.","DOI":"10.3390\/toxics9110316"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Zhao, C., Guo, H., Hou, Y., Lei, T., Wei, D., and Zhao, Y. (2023). Multiple Roles of the Stress Sensor GCN2 in Immune Cells. Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms24054285"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"37993","DOI":"10.1074\/jbc.M004762200","article-title":"PACT, a stress-modulated cellular activator of interferon-induced double-stranded RNA-activated protein kinase, PKR","volume":"275","author":"Patel","year":"2000","journal-title":"J. Biol. Chem."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1128\/MCB.01030-08","article-title":"TRBP control of PACT-induced phosphorylation of protein kinase R is reversed by stress","volume":"29","author":"Daher","year":"2009","journal-title":"Mol. Cell Biol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"e2788","DOI":"10.1038\/cddis.2017.199","article-title":"Chronically stressed or stress-preconditioned neurons fail to maintain stress granule assembly","volume":"8","author":"Shelkovnikova","year":"2017","journal-title":"Cell Death Dis."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"4827","DOI":"10.1007\/s00018-020-03565-0","article-title":"Stress granule subtypes: An emerging link to neurodegeneration","volume":"77","author":"Advani","year":"2020","journal-title":"Cell Mol. Life Sci."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"e121159","DOI":"10.1172\/jci.insight.121159","article-title":"Residual function of cystic fibrosis mutants predicts response to small molecule CFTR modulators","volume":"3","author":"Han","year":"2018","journal-title":"JCI Insight"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"444","DOI":"10.1038\/nchembio.1253","article-title":"Mechanism-based corrector combination restores \u0394F508-CFTR folding and function","volume":"9","author":"Okiyoneda","year":"2013","journal-title":"Nat. Chem. Biol."},{"key":"ref_38","first-page":"617","article-title":"Possible mechanisms of emphysema in smokers: In vitro suppression of serum elastase-inhibitory capacity by fresh cigarette smoke and its prevention by antioxidants","volume":"118","author":"Carp","year":"1978","journal-title":"Am. Rev. Respir. Dis."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"834","DOI":"10.1085\/jgp.201812143","article-title":"Agonists that stimulate secretion promote the recruitment of CFTR into membrane lipid microdomains","volume":"151","author":"Kim","year":"2019","journal-title":"J. Gen. Physiol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1967","DOI":"10.1016\/j.cub.2023.04.012","article-title":"Stress granule formation inhibits stress-induced apoptosis by selectively sequestering executioner caspases","volume":"33","author":"Fujikawa","year":"2023","journal-title":"Curr. Biol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"e05033","DOI":"10.7554\/eLife.05033","article-title":"The small molecule ISRIB reverses the effects of eIF2\u03b1 phosphorylation on translation and stress granule assembly","volume":"4","author":"Sidrauski","year":"2015","journal-title":"eLife"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"110559","DOI":"10.1016\/j.vetmic.2025.110559","article-title":"PERK-mediated eIF2\u03b1 phosphorylation suppresses porcine epidemic diarrhea virus replication by attenuating global protein synthesis and inducing IFN-I production","volume":"307","author":"Yang","year":"2025","journal-title":"Vet. Microbiol."},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Hicks, D., Giresh, K., Wrischnik, L.A., and Weiser, D.C. (2023). The PPP1R15 Family of eIF2-alpha Phosphatase Targeting Subunits (GADD34 and CReP). Int. J. Mol. Sci., 24.","DOI":"10.3390\/ijms242417321"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"397","DOI":"10.1038\/s41419-020-2601-2","article-title":"Fine tuning of the unfolded protein response by ISRIB improves neuronal survival in a model of amyotrophic lateral sclerosis","volume":"11","author":"Bugallo","year":"2020","journal-title":"Cell Death Dis."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"763","DOI":"10.1016\/j.bbrc.2012.06.033","article-title":"Hydrogen peroxide induces stress granule formation independent of eIF2\u03b1 phosphorylation","volume":"423","author":"Emara","year":"2012","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1091\/mbc.01-05-0221","article-title":"Evidence that ternary complex (eIF2-GTP-tRNA(i)(Met))-deficient preinitiation complexes are core constituents of mammalian stress granules","volume":"13","author":"Kedersha","year":"2002","journal-title":"Mol. Biol. Cell"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"113769","DOI":"10.1016\/j.celrep.2024.113769","article-title":"hnRNPA2B1 represses the disassembly of arsenite-induced stress granules and is essential for male fertility","volume":"43","author":"Wang","year":"2024","journal-title":"Cell Rep."}],"container-title":["Biomolecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2218-273X\/16\/4\/615\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,21]],"date-time":"2026-04-21T12:29:42Z","timestamp":1776774582000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2218-273X\/16\/4\/615"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4,21]]},"references-count":47,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2026,4]]}},"alternative-id":["biom16040615"],"URL":"https:\/\/doi.org\/10.3390\/biom16040615","relation":{},"ISSN":["2218-273X"],"issn-type":[{"value":"2218-273X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,21]]}}}