title: TRX-ASK1-JNK signaling regulation of cell density-dependent cytotoxicity in cigarette smoke-exposed human bronchial epithelial cells.
abstract: Cigarette smoke is a major environmental air pollutant that injures airway epithelium and incites subsequent diseases including chronic obstructive pulmonary disease. The lesion that smoke induces in airway epithelium is still incompletely understood. Using a LIVE/DEAD cytotoxicity assay, we observed that subconfluent cultures of bronchial epithelial cells derived from both human and monkey airway tissues and an immortalized normal human bronchial epithelial cell line (HBE1) were more susceptible to injury by cigarette smoke extract (CSE) and by direct cigarette smoke exposure than cells in confluent cultures. Scraping confluent cultures also caused an enhanced cell injury predominately in the leading edge of the scraped confluent cultures by CSE. Cellular ATP levels in both subconfluent and confluent cultures were drastically reduced after CSE exposure. In contrast, GSH levels were significantly reduced only in subconfluent cultures exposed to smoke and not in confluent cultures. Western blot analysis demonstrated ERK activation in both confluent and subconfluent cultures after CSE. However, activation of apoptosis signal-regulating kinase 1 (ASK1), JNK, and p38 were demonstrated only in subconfluent cultures and not in confluent cultures after CSE. Using short interfering RNA (siRNA) to JNK1 and JNK2 and a JNK inhibitor, we attenuated CSE-mediated cell death in subconfluent cultures but not with an inhibitor of the p38 pathway. Using the tetracycline (Tet)-on inducible approach, overexpression of thioredoxin (TRX) attenuated CSE-mediated cell death and JNK activation in subconfluent cultures. These results suggest that the TRX-ASK1-JNK pathway may play a critical role in mediating cell density-dependent CSE cytotoxicity.
authors.affiliation: Center for Comparative Respiratory Biology and Medicine, Genome and Biomedical Science Facility, Rm. 6510, Univ. of California at Davis, 451 East Health Sciences Dr., Davis, CA 95616, USA.
authors.forename: Yong Chan
authors.initials: YC
authors.lastname: Lee
authors.forename: Chun-Yu
authors.initials: CY
authors.lastname: Chuang
authors.forename: Pak-Kei
authors.initials: PK
authors.lastname: Lee
authors.forename: Jin-Soo
authors.initials: JS
authors.lastname: Lee
authors.forename: Richart W
authors.initials: RW
authors.lastname: Harper
authors.forename: Alan B
authors.initials: AB
authors.lastname: Buckpitt
authors.forename: Reen
authors.initials: R
authors.lastname: Wu
authors.forename: Karen
authors.initials: K
authors.lastname: Oslund
mesh_terms: D001980:Bronchi / Q000166:cytology; D002452:Cell Count; D016923:Cell Death; D002460:Cell Line; D002470:Cell Survival; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D048848:MAP Kinase Kinase Kinase 5 / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000502:physiology; D008565:Membrane Proteins; D048055:Mitogen-Activated Protein Kinase 8 / Q000235:genetics / Q000378:metabolism*; D048056:Mitogen-Activated Protein Kinase 9 / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering; D020545:Respiratory Mucosa / Q000166:cytology / Q000378:metabolism*; D015208:Smoke Inhalation Injury / Q000378:metabolism* / Q000473:pathology; D014028:Tobacco Smoke Pollution / Q000009:adverse effects*
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D034741:RNA, Small Interfering; D014028:Tobacco Smoke Pollution; C494631:VAC14 protein, human; D048056:Mitogen-Activated Protein Kinase 9; D048055:Mitogen-Activated Protein Kinase 8; D048848:MAP Kinase Kinase Kinase 5; C482858:MAP3K5 protein, human