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Pflügers Archiv - European Journal of Physiology

, Volume 470, Issue 8, pp 1231–1241 | Cite as

TRPA1 channels: expression in non-neuronal murine lung tissues and dispensability for hyperoxia-induced alveolar epithelial hyperplasia

  • Martina Kannler
  • Robin Lüling
  • Ali Önder Yildirim
  • Thomas Gudermann
  • Dirk Steinritz
  • Alexander Dietrich
Ion channels, receptors and transporters
Part of the following topical collections:
  1. Ion channels, receptors and transporters

Abstract

Transient receptor potential A1 (TRPA1) channels were originally characterized in neuronal tissues but also identified in lung epithelium by staining with fluorescently coupled TRPA1 antibodies. Its exact function in non-neuronal tissues, however, is elusive. TRPA1 is activated in vitro by hypoxia and hyperoxia and is therefore a promising TRP candidate for sensing hyperoxia in pulmonary epithelial cells and for inducing alveolar epithelial hyperplasia. Here, we isolated tracheal, bronchial, and alveolar epithelial cells and show low but detectable TRPA1 mRNA levels in all these cells as well as TRPA1 protein by Western blotting in alveolar type II (AT II) cells. We quantified changes in intracellular Ca2+ ([Ca2+]i) levels induced by application of hyperoxic solutions in primary tracheal epithelial, bronchial epithelial, and AT II cells isolated from wild-type (WT) and TRPA1-deficient (TRPA1−/−) mouse lungs. In all cell types, we detected hyperoxia-induced rises in [Ca2+]i levels, which were not significantly different in TRPA1-deficient cells compared to WT cells. We also tested TRPA1 function in a mouse model for hyperoxia-induced alveolar epithelial hyperplasia. A characteristic significant increase in thickening of alveolar tissues was detected in mouse lungs after exposure to hyperoxia, but not in normoxic WT and TRPA1−/− controls. Quantification of changes in lung morphology in hyperoxic WT and TRPA1−/− mice, however, again revealed no significant changes. Therefore, TRPA1 expression does neither appear to be a key player for hyperoxia-induced changes in [Ca2+]i levels in primary lung epithelial cells, nor being essential for the development of hyperoxia-induced alveolar epithelial hyperplasia.

Keywords

TRPA1 mRNA expression Human lung tissues Murine lung tissues Hyperoxia Alveolar epithelial hyperplasia 

Notes

Acknowledgements

We thank Bettina Braun, Benjamin Lukas Heinz, Christine Hollauer, Daniel Hofmann, and Heinz Janser for excellent technical expertise as well as Drs. Thomas Büch and Eva Schäfer for kindly providing the TRPA1 expressing HEK293 cell line. We are grateful for Dr. Jiong Zhang’s expertise in preparing dorsal root ganglia (DRG) and thank Dr. Claudia Staab-Weijnitz for her help in identifying lung epithelial cells.

Author’s contributions

M.K. and A.D. designed the study, R.L. and D.S. designed and performed the proteomic analysis, A.Ö.Y. designed and supervised the quantification of lung tissue sections, M.K. performed all other experiments, M.K. and A.D. analyzed the data, M.K., A.D., and T.G. wrote the manuscript, all authors edited and approved the manuscript.

Compliance with ethical standards

All animal experiments were approved by the local authorities (Regierung Oberbayern) and are in compliance with ethical standards.

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

424_2018_2148_MOESM1_ESM.jpg (54 kb)
ESM 1 Supplementary Fig. 1 Quantification of TRPM7 mRNA expression in murine lung cells. M-RNA expression levels of TRPM7 in murine lung tissues and dorsal root ganglions (DRG) as a positive control quantified by the nanostring® technology. Counts from fluorescent oligonucleotides detecting TRPM7 mRNAs after subtraction of counts from negative controls obtained by fluorescent probes - specific for mRNAs not present in this species - and normalized to data from 4 housekeeping genes are presented in black bars. tEPC, tracheal epithelial cells; pmLF, primary murine lung fibroblasts; ATII, alveolar type II cells; ATII-ATI, ATI cells differentiated from ATII cells; PASMC, precapillary pulmonary arterial smooth muscle cells; EC, endothelial cells from lung tissues. Supplementary Fig. 2 Analysis of intracellular Ca2+ [Ca2+]i levels after application of allyl-isothiocyanate (AITC) to primary murine alveolar type II (AT II) cells. Increases in [Ca2+]i levels were detected by analysis of fluorescence ratios (Ratios 340/380 nm). The lines represent calculated means, whereas the light gray areas indicate standard errors of the mean (S.E.M.) (JPG 54 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Walther Straub Institute of Pharmacology and Toxicology, Member of the German Center for Lung Research (DZL)LMU MunichMunichGermany
  2. 2.Bundeswehr Institute of Pharmacology and ToxicologyMunichGermany
  3. 3.Comprehensive Pneumology Center, Institute of Lung Biology and Disease, Member of the German Center for Lung Research (DZL)Helmholtz Center MunichMunichGermany
  4. 4.German Center for Lung Research, Comprehensive Pneumology CenterMunichGermany
  5. 5.German Centre for Cardiovascular ResearchMunich Heart AllianceMunichGermany

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