TRPC6 modulates adhesion of neutrophils to airway epithelial cells via NF-κB activation and ICAM-1 expression with ozone exposure.

Ozone (O3) is a major component of air pollution, which has been associated with airway inflammation characterized by the influx of neutrophils in asthmatic subjects. Canonical transient receptor potential 6 (TRPC6) channel is recently identified as a target of oxidative stress which is involved in airway inflammation. However, the regulatory role of TRPC6 in airway epithelial cells and neutrophils has not yet been illuminated in detail. In this study, we investigated the role of TRPC6 in neutrophil adhesion to airway epithelial cells exposed to O3 in vivo and in vitro approaches. Using transgenic mice, the results showed that TRPC6-deficiency attenuated O3-induced neutrophil recruitment to airway epithelial cells and intercellular adhesion molecule-1 (ICAM-1) expression. In vitro, O3 induced ICAM-1 expression and neutrophil adhesion to 16HBE cells (human airway epithelial cell line) and which were reduced by both TRPC6 silencing short hairpin RNA (shRNA) and TRPC6 inhibitor Larixyl Acetate (LA). We also confirmed that TRPC6-dependent Ca2+ entry and NF-κB activation in 16HBE cells were required for ICAM-1-mediated neutrophil adhesion exposed to O3. In conclusion, this study demonstrated the contribution of TRPC6 to O3-induced neutrophil adhesion to airway epithelial cells via NF-κB activation and ICAM-1 expression, which may provide new potential concepts for preventing and treating air pollutant-related inflammatory lung diseases.

TRPC6 contributes to LPS-induced inflammation through ERK1/2 and p38 pathways in bronchial epithelial cells.

receptor potential canonical (TRPC) channels are presently an emerging target for airway disorders. Recent evidence has indicated that TRPC6 as a member of the TRPC family plays an important role in airway inflammation, but its precise function in bronchial epithelial cells remains unclear. The aim of this study was to investigate the role of TRPC6 in Toll-like receptor 4 (TLR4)-mediated inflammation in human bronchial epithelial cells stimulated by endotoxin [lipopolysaccharide (LPS)]. Hyp9 is a simplified phloroglucinol derivative of hyperforin that highly selectively activates TRPC6 channels. The results show that the activation of TRPC6 by Hyp9 induced the production of interleukin (IL)-8 and IL-6. LPS was also able to induce the release of IL-8 and IL-6, which was significantly aggravated by Hyp9 and reduced by knockdown of TRPC6. Treatment with LPS not only chronically induced the expression of TRPC6 mRNA and protein in a TLR4-dependent manner but also acutely increased Ca2+ influx through TRPC6 channels. In addition, LPS-induced overexpression of TRPC6 and Ca2+ influx were associated with the phosphorylation of phosphatidylinositol 3-kinase (PI3K) and Akt. Importantly, TRPC6 was required for the activation of ERK1/2, p38, and NF-κB. In conclusion, these data reveal that LPS induced the overexpression of TRPC6 and TRPC6-dependent Ca2+ influx via the TLR4/PI3K/Akt pathway resulting in Ca2+ mobilization, which subsequently promoted the activation of ERK1/2, p38, and NF-κB and the inflammatory response in bronchial epithelial cells.

[N-acetyl-L-cysteine reduces the ozone-induced lung inflammation response in mice].

In this study, we investigated the protective effect of the antioxidant N-acetyl-L-cysteine (NAC) on the lung inflammation caused by ozone (O3) exposure in mice. Thirty-two C57BL/6 mice were randomly divided into control group, O3 group, O3+NAC group and NAC group. Mice were exposed to O3 (1.0 ppm) or fresh air for 3 h on the day 1, day 3 and day 5, respectively. NAC (100 mg/kg) was intraperitoneally applied to the mice 1 h before each exposure. At 24 h after the 3-time exposure, the alveolar wall structure was severely damaged and the infiltrated inflammatory cells were apparent perivascularly and peribronchiolarly. Significant increases in the total white blood cell count, macrophage, lymphocyte and neutrophil counts, as well as total protein concentration were observed in the bronchoalveolar lavage fluid (BALF) (P < 0.05). The IL-6, IL-8 (P < 0.01) and MDA levels (P < 0.05) in the lung homogenates were elevated coherently. Administration of NAC could attenuate the alveolar wall structure damage induced by O3 exposure and reduce the amount of infiltrated inflammatory cells, total and differential leukocyte counts (P < 0.05), as well as the IL-6, IL-8 (P < 0.01) and MDA release (P < 0.05). Western blotting results showed that the O3 exposure up-regulated the p38 MAPK and NF-κB p65 protein expression in the lung tissue of mice (P < 0.05), which could be alleviated by NAC (P < 0.05). These results indicated that NAC could protect against O3-induced pulmonary inflammation in mice. The beneficial effect of NAC might be related with the p38 MAPK and NF-κB p65 signal pathway.