title: Functional expression of the transient receptor potential channel TRPA1, a sensor for toxic lung inhalants, in pulmonary epithelial cells.
abstract: The cation channel TRPA1 functions as a chemosensory protein and is directly activated by a number of noxious inhalants. A pulmonary expression of TRPA1 has been described in sensory nerve endings and its stimulation leads to the acceleration of inflammatory responses in the lung. Whereas the function of TRPA1 in neuronal cells is well defined, only few reports exist suggesting a role in epithelial cells. The aim of the present study was therefore (1) to evaluate the expression of TRPA1 in pulmonary epithelial cell lines, (2) to characterize TRPA1-promoted signaling in these cells, and (3) to study the extra-neuronal expression of this channel in lung tissue sections. Our results revealed that the widely used alveolar type II cell line A549 expresses TRPA1 at the mRNA and protein level. Furthermore, stimulating A549 cells with known TRPA1 activators (i.e., allyl isothiocyanate) led to an increase in intracellular calcium levels, which was sensitive to the TRPA1 blocker ruthenium red. Investigating TRPA1 coupled downstream signaling cascades it was found that TRPA1 activation elicited a stimulation of ERK1/2 whereas other MAP kinases were not affected. Finally, using epithelial as well as neuronal markers in immunohistochemical approaches, a non-neuronal TRPA1 protein expression was detected in distal parts of the porcine lung epithelium, which was also found examining human lung sections. TRPA1-positive staining co-localized with both epithelial and neuronal markers underlining the observed epithelial expression pattern. Our findings of a functional expression of TRPA1 in pulmonary epithelial cells provide causal evidence for a non-neuronal TRPA1-mediated control of inflammatory responses elicited upon TRPA1-mediated registration of toxic inhalants in vivo.
authors.affiliation: Rudolf-Boehm Institute of Pharmacology and Toxicology, Clinical Pharmacology, University Leipzig, Härtelstraße 16-18, 04107 Leipzig, Germany.
authors.forename: Thomas Robert Heinrich
authors.initials: TR
authors.lastname: Büch
authors.forename: Eva Anna Maria
authors.initials: EA
authors.lastname: Schäfer
authors.forename: Maria-Theresia
authors.initials: MT
authors.lastname: Demmel
authors.forename: Ingrid
authors.initials: I
authors.lastname: Boekhoff
authors.forename: Horst
authors.initials: H
authors.lastname: Thiermann
authors.forename: Thomas
authors.initials: T
authors.lastname: Gudermann
authors.forename: Dirk
authors.initials: D
authors.lastname: Steinritz
authors.forename: Annette
authors.initials: A
authors.lastname: Schmidt
mesh_terms: D055371:Acute Lung Injury / Q000139:chemically induced / Q000378:metabolism; D056809:Alveolar Epithelial Cells / Q000187:drug effects* / Q000378:metabolism*; D000818:Animals; D015220:Calcium Channels / Q000235:genetics* / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D015870:Gene Expression; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D009419:Nerve Tissue Proteins / Q000819:agonists / Q000235:genetics* / Q000378:metabolism*; D009676:Noxae / Q000008:administration & dosage / Q000633:toxicity*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013552:Swine; D000074025:TRPA1 Cation Channel; D050051:Transient Receptor Potential Channels / Q000819:agonists / Q000235:genetics* / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D017879:Isothiocyanates; D009419:Nerve Tissue Proteins; D009676:Noxae; D012333:RNA, Messenger; D000074025:TRPA1 Cation Channel; C473350:TRPA1 protein, human; D050051:Transient Receptor Potential Channels; C004471:allyl isothiocyanate
keywords: AITC; Calcium channel signaling; Chemosensation; EGF; ERK1/2; FCS; ICC; IHC; Lung; MAPK; Non-neuronal TRPA1; RR; SDS; TRP; Toxic inhalation hazards; WB; Western blot; allyl isothiocyanate; epithelial growth factor; extracellular signal-regulated protein kinase ½; fetal calf serum HBS, HEPES buffered saline; immunocytochemistry; immunohistochemistry; mitogen activated protein kinase; ruthenium red; sodium dodecyl sulfate; transient receptor potential channel