Potency (EC₅₀ and CI) of the various TRPA1 agonists in different non-neuronal cell types of the human respiratory tract.

<p>Potency (EC₅₀ and CI) of the various TRPA1 agonists in different non-neuronal cell types of the human respiratory tract.</p

Functional TRPA1 is expressed in human airway/lung cells.

<p>Intracellular calcium response was used to assess agonist-induced TRPA1 activation in small airways epithelial cells (SAEC) (<b>A</b>), normal human lung fibroblasts (NHLF) (<b>B</b>) and bronchial smooth muscle cells (HBSMC) (<b>C</b>). Typical traces and pooled data of the concentration-dependent calcium response evoked by the selective TRPA1 agonists, cinnamaldehyde (CNM, typical traces and black circles) and acrolein (ACR, grey circles), in all different cell types in primary culture. Similarly to CNM and ACR, cigarette smoke extract (CSE, black triangles) produces in all the different types of cells a concentration-dependent calcium response. Responses to CNM, ACR and CSE are prevented by selective TRPA1 antagonists, HC-030031 (HC, 10 µM) and AP18 (10 µM). The activating peptide (SLIGKV-NH₂) of the PAR-2 receptor (PAR-2 AP, 100 µM) elicits a calcium response that is not modified by TRPA1 antagonists. Veh is a combination of vehicles of HC and AP18. Values represent mean ± SEM of n>25 cells. ^*<i>P</i><0.05 <i>vs.</i> Veh.</p

TRPA1 mediates IL-8 release by acrolein from human airway/lung cells in primary culture.

<p>Overnight exposure to acrolein (ACR) induces IL-8 release from small airway epithelial cells (SAEC) (<b>A</b>), normal human lung fibroblasts (NHLF) (<b>B</b>) and human bronchial smooth muscle cells (HBSMC) (<b>C</b>) in a concentration–dependent manner. IL-8 release evoked by ACR is reduced by HC-030031 (HC, 30 µM) and AP18 (10 µM). Each column represents mean ± SEM of at least 3 independent experiments. ^§<i>P</i><0.05 vs. Basal group or Veh/Veh-ACR; ^*<i>P</i><0.05 vs. Veh/ACR.</p

TRPA1 channel expression in non-neuronal cells of the human and mouse respiratory tract.

<p>(<b>A</b>) Total RNAs were extracted from primary small airways epithelial cells (SAEC), human type II alveolar epithelial cells (A549), human primary smooth muscle cells (HBSMC), human embryonic lung fibroblasts (IMR90) and primary normal human lung fibroblasts (NHLF) and relative TRPA1 mRNA amounts were measured by Taqman Real-Time PCR assay. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0042454#s2" target="_blank">Results</a> are normalized to the reference gene, β-actin. Each column represents mean ± SEM of n>2 independent experiments. Immunohistochemical analysis of TRPA1 expression in samples taken from human (<b>B</b>) or <i>Trpa1^+/+</i> and <i>Trpa1^−/−</i> mouse airways and lung (<b>C</b>). Representative images of TRPA1 immunostaining show intense staining in epithelial and smooth muscle cells in human tissue. No staining is detected in human samples incubated with the normal serum peptide (Negative control). (<b>C</b>) Incubation with TRPA1 antibody shows a strong staining in epithelial and smooth muscle cells in tissues taken from <i>Trpa1^+/+</i> mice, but not in those from <i>Trpa1^−/−</i> mice. Preadsorption of the TRPA1 antibody with the peptide used for immunization abolished staining (Peptide). Staining for cytokeratin and α-smooth muscle actin (α-SMA) overlaps with the TRPA1 staining in the bronchial epithelium and smooth muscle layer in serial section of human and mice airways/lung tissues (<b>B</b> and <b>C</b>). Scale bar 100 µm.</p