title: Transient receptor potential vanilloid 1 agonists cause endoplasmic reticulum stress and cell death in human lung cells.
abstract: Transient receptor potential vanilloid 1 (TRPV1) is a calcium-selective ion channel expressed in human lung cells. We show that activation of the intracellular subpopulation of TRPV1 causes endoplasmic reticulum (ER) stress and cell death in human bronchial epithelial and alveolar cells. TRPV1 agonist (nonivamide) treatment caused calcium release from the ER and altered the transcription of growth arrest- and DNA damage-inducible transcript 3 (GADD153), GADD45alpha, GRP78/BiP, ATF3, CCND1, and CCNG2) in a manner comparable with prototypical ER stress-inducing agents. The TRPV1 antagonist N-(4-tert-butylbenzyl)-N'-(1-[3-fluoro-4-(methylsulfonylamino)-phenyl]ethyl)thiourea (LJO-328) inhibited mRNA responses and cytotoxicity. EGTA and ruthenium red inhibited cell surface TRPV1 activity, but they did not prevent ER stress gene responses or cytotoxicity. Cytotoxicity paralleled eukaryotic translation initiation factor 2, subunit 1 (EIF2alpha) phosphorylation and the induction of GADD153 mRNA and protein. Transient overexpression of GADD153 caused cell death independent of agonist treatment, and cells selected for stable overexpression of a GADD153 dominant-negative mutant exhibited reduced sensitivity. Salubrinal, an inhibitor of ER stress-induced cytotoxicity via the EIF2alphaK3/EIF2alpha pathway, or stable overexpression of the EIF2alpha-S52A dominant-negative mutant also inhibited cell death. Treatment of the TRPV1-null human embryonic kidney 293 cell line with TRPV1 agonists did not initiate ER stress responses. Likewise, n-benzylnonanamide, an inactive analog of nonivamide, failed to cause ER calcium release, an increase in GADD153 expression, and cytotoxicity. We conclude that activation of ER-bound TRPV1 and stimulation of GADD153 expression via the EIF2alphaK3/EIF2alpha pathway represents a common mechanism for cytotoxicity by cell-permeable TRPV1 agonists. These findings are significant within the context of lung inflammatory diseases where elevated concentrations of endogenous TRPV1 agonists are probably produced in sufficient quantities to cause TRPV1 activation and lung cell death.
authors.affiliation: Department of Pharmacology and Toxicology, University of Utah, Salt Lake City, UT 84112, USA.
authors.forename: Karen C
authors.initials: KC
authors.lastname: Thomas
authors.forename: Ashwini S
authors.initials: AS
authors.lastname: Sabnis
authors.forename: Mark E
authors.initials: ME
authors.lastname: Johansen
authors.forename: Diane L
authors.initials: DL
authors.lastname: Lanza
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Moos
authors.forename: Garold S
authors.initials: GS
authors.lastname: Yost
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Reilly
mesh_terms: D051700:Activating Transcription Factor 3 / Q000235:genetics; D001095:Arachidonic Acids / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002211:Capsaicin / Q000031:analogs & derivatives / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002934:Cinnamates / Q000494:pharmacology; D019938:Cyclin D1 / Q000235:genetics; D056767:Cyclin G2; D016213:Cyclins / Q000235:genetics; D004224:Diterpenes / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D063388:Endocannabinoids; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015852:Eukaryotic Initiation Factor-2 / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D008168:Lung / Q000166:cytology / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D053284:Polyunsaturated Alkamides / Q000494:pharmacology; D050916:TRPV Cation Channels / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000235:genetics; D019284:Thapsigargin / Q000494:pharmacology; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D051742:Transcription Factor CHOP / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: C490449:ATF3 protein, human; D051700:Activating Transcription Factor 3; D001095:Arachidonic Acids; C101593:CCNG2 protein, human; D002934:Cinnamates; D056767:Cyclin G2; D016213:Cyclins; C497249:DDIT3 protein, human; D004224:Diterpenes; D063388:Endocannabinoids; D000091342:Endoplasmic Reticulum Chaperone BiP; D004791:Enzyme Inhibitors; D015852:Eukaryotic Initiation Factor-2; C000717247:HSPA5 protein, human; D053284:Polyunsaturated Alkamides; D050916:TRPV Cation Channels; C494482:TRPV1 protein, human; C496827:salubrinal; D019938:Cyclin D1; D051742:Transcription Factor CHOP; D019284:Thapsigargin; C024353:resiniferatoxin; D013890:Thiourea; D002211:Capsaicin; C040937:nonivamide; D002118:Calcium; D004229:Dithiothreitol; C078814:anandamide