TRPA1 channel is expressed in non-neuronal pulmonary cells and promotes non-neurogenic inflammation.

Serena Materazzi1, Pamela Pedretti1,2, Camilla Fusi1, Nadia Moretto0,2, Chiara Carnini0,2, Fabrizio Facchinetti0,2, Riccardo Patacchini0,2, Pierangelo Geppetti1, Romina Nassini1. 1University of Florence, Dept. of Preclinical and Clinical Pharmacology 50139, Italy, 2Chiesi Farmaceutici SpA, Pharmacology Department 43100, Italy

The transient receptor potential ankyrin 1 (TRPA1) channel is localized to airway sensory nerves and has been recently proposed to mediate airway inflammation evoked by allergen and cigarette smoke (CS) in rodents, via neurogenic mechanisms. However the limited clinical evidence for the role of neurogenic inflammation in asthma or chronic obstructive pulmonary disease raises an alternative possibility that airway inflammation is promoted by non-neuronal TRPA1. Here, we report that TRPA1 channel if functionally expressed in non-neuronal districts of human and mouse airway, where it produces inflammatory responses.

In this study C57BL/6 mice, wild-type or TRPA1-deficient mice generated by heterozygous mice on a C57BL/6 background were used. By using Real-Time PCR and calcium imaging, we found that cultured human airway cells, including fibroblasts (IMR90 and NHLF), epithelial (A549 and SAEC) and smooth muscle (HBSMC) cells express functional TRPA1 channels. For functional characterization we used selective exogenous and endogenous TRPA1 agonists, cinnamaldehyde and acrolein respectively, and cigarette smoke extract (CSE). By using immunohistochemistry TRPA1 staining was observed in airway epithelial and smooth muscle cells in sections taken from human airways and lung and from airways and lung of wild-type, but not TRPA1-deficient mice. In addition, acrolein and CSE exposure (16 h) evoked IL-8 release in cultured human airway epithelial and smooth muscle cells and fibroblasts in a concentration dependent manner, an effect selectively reduced by TRPA1 antagonists, HC-030031 and AP18 (both 10 µM). Stimulation of TRPV1 (by capsaicin, 100 µM/30 µl i.t.), co-expressed with TRPA1 in the same airway sensory nerves, and TRPA1 (by acrolein 5 mM/30 µl i.t or CS 1 OD/30 µl i.t ), or the neuropeptide, substance P (SP, 25 nM/30 µl i.t ), which is released from sensory nerve terminals stimulation, provoked neurogenic inflammation in mouse airways. However, TRPA1 activation but not TRPV1 or SP activation, was able to increase release of keratinocyte chemoattractant (CXCL-1/KC, IL-8 analogue) in BAL of wild-type mice, and this effect was attenuated by TRPA1 antagonism or in TRPA1-deficient mice. While TRPV1 or TRPA1 activation causes airway neurogenic inflammation, non-neuronal TRPA1 produces an additional, prominent non-neurogenic inflammatory response, which may contribute to inflammatory airway diseases. Statistical significance was determined by using one- or two-way ANOVA, followed by Bonferroni’s post hoc analysis for comparison of multiple groups and the unpaired 2-tailed Student’s t-test between 2 groups. P < 0.05 was considered significant. For in vivo studies at least of 6 mice for group have been used.

Thus, present data, proposing that while TRPV1 or TRPA1 activation causes airway neurogenic inflammation, non-neuronal TRPA1 produces an additional, prominent non-neurogenic inflammatory response, which may contribute to inflammatory airway diseases. In addition, data suggest that non-neuronal TRPA1, rather than the channel expressed in sensory nerve terminals, contributes to the mechanism of inflammatory airway diseases, offering a novel interpretation of the role of TRPA1 that could be a novel target for the treatment of inflammatory respiratory diseases.