title: Identification of TGF-β receptor-1 as a key regulator of carbon nanotube-induced fibrogenesis.
abstract: Carbon nanotubes (CNTs) induce rapid interstitial lung fibrosis, but the underlying mechanisms are unclear. Previous studies indicated that the ability of CNTs to penetrate lung epithelium, enter interstitial tissue, and stimulate fibroblasts to produce collagen matrix is important to lung fibrosis. In this study, we investigated the activation of transforming growth factor-β receptor-1 [TGF-β R1; i.e., activin receptor-like kinase 5 (ALK5) receptor] and TGF-β/Smad signaling pathway in CNT-induced collagen production in human lung fibroblasts. Human lung fibroblasts and epithelial cells were exposed to low, physiologically relevant concentrations (0.02-0.6 μg/cm(2)) of single-walled CNTs (SWCNT) and multiwalled CNTs (MWCNT) in culture and analyzed for collagen, TGF-β1, TGF-β R1, and SMAD proteins by Western blotting and immunofluorescence. Chemical inhibition of ALK5 and short-hairpin (sh) RNA targeting of TGF-β R1 and Smad2 were used to probe the fibrogenic mechanism of CNTs. Both SWCNT and MWCNT induced an overexpression of TGF-β1, TGF-β R1 and Smad2/3 proteins in lung fibroblasts compared with vehicle or ultrafine carbon black-exposed controls. SWCNT- and MWCNT-induced collagen production was blocked by ALK5 inhibitor or shRNA knockdown of TGF-β R1 and Smad2. Our results indicate the critical role of TGF-β R1/Smad2/3 signaling in CNT-induced fibrogenesis by upregulating collagen production in lung fibroblasts. This novel finding may aid in the design of mechanism-based risk assessment and development of rapid screening tests for nanomaterial fibrogenicity.
authors.affiliation: Health Effects Laboratory Division (HELD), National Institute for Occupational Safety and Health, Morgantown, West Virginia; and Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia.
authors.forename: Anurag
authors.initials: A
authors.lastname: Mishra
authors.affiliation: Health Effects Laboratory Division (HELD), National Institute for Occupational Safety and Health, Morgantown, West Virginia; and Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia.
authors.forename: Todd A
authors.initials: TA
authors.lastname: Stueckle
authors.affiliation: Health Effects Laboratory Division (HELD), National Institute for Occupational Safety and Health, Morgantown, West Virginia; and.
authors.forename: Robert R
authors.initials: RR
authors.lastname: Mercer
authors.affiliation: Health Effects Laboratory Division (HELD), National Institute for Occupational Safety and Health, Morgantown, West Virginia; and.
authors.forename: Raymond
authors.initials: R
authors.lastname: Derk
authors.affiliation: Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia.
authors.forename: Yon
authors.initials: Y
authors.lastname: Rojanasakul
authors.affiliation: Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia.
authors.forename: Vincent
authors.initials: V
authors.lastname: Castranova
authors.affiliation: Health Effects Laboratory Division (HELD), National Institute for Occupational Safety and Health, Morgantown, West Virginia; and Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia lmw6@cdc.gov.
authors.forename: Liying
authors.initials: L
authors.lastname: Wang
mesh_terms: D000818:Animals; D002460:Cell Line; D024042:Collagen Type I / Q000096:biosynthesis; D005347:Fibroblasts / Q000378:metabolism; D055785:Gene Knockdown Techniques; D006801:Humans; D008168:Lung / Q000378:metabolism; D051379:Mice; D037742:Nanotubes, Carbon / Q000633:toxicity* / Q000648:ultrastructure; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011658:Pulmonary Fibrosis / Q000209:etiology* / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D020545:Respiratory Mucosa / Q000378:metabolism; D012307:Risk Factors; D015398:Signal Transduction; D051899:Smad2 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000378:metabolism
chemical_list: D024042:Collagen Type I; D037742:Nanotubes, Carbon; D034741:RNA, Small Interfering; D018125:Receptors, Transforming Growth Factor beta; C497782:SMAD2 protein, human; D051899:Smad2 Protein; C508960:TGFB1 protein, human; D053773:Transforming Growth Factor beta1; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human; C000625842:Tgfbr1 protein, mouse
keywords: carbon nanotubes; collagen I; fibrosis; lung; receptor; transforming growth factor