TRPA1 channels: expression in non-neuronal murine lung tissues and dispensability for hyperoxia-induced alveolar epithelial hyperplasia
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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.
KeywordsTRPA1 mRNA expression Human lung tissues Murine lung tissues Hyperoxia Alveolar epithelial hyperplasia
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.
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.
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