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Toluene diisocyanate exposure induces airway inflammation of bronchial epithelial cells via the activation of transient receptor potential melastatin 8

View Article: PubMed Central - PubMed

ABSTRACT

Toluene diisocyanate (TDI) is the most important cause of occupational asthma (OA), and various pathogenic mechanisms have been suggested. Of these mechanisms, neurogenic inflammation is an important inducer of airway inflammation. Transient receptor potential melastatin 8 (TRPM8) is a well-established cold-sensing cation channel that is expressed in both neuronal cells and bronchial epithelial cells. A recent genome-wide association study of TDI-exposed workers found a significant association between the phenotype of TDI-induced OA and the single-nucleotide polymorphism rs10803666, which has been mapped to the TRPM8 gene. We hypothesized that TRPM8 located in airway epithelial cells may be involved in the pathogenic mechanisms of TDI-induced OA and investigated its role. Bronchial epithelial cells were treated with TDI in a dose- and time-dependent manner. The expression levels of TRPM8 mRNA and protein were determined by quantitative real-time polymerase chain reaction and western blotting. TDI-induced morphological changes in the cells were evaluated by immunocytochemistry. Alterations in the transcripts of inflammatory cytokines were examined in accordance with TRPM8 activation by TDI. TRPM8 expression at both the mRNA and protein levels was enhanced by TDI in airway epithelial cells. TRPM8 activation by TDI led to significant increases in the mRNA of interleukin (IL)-4, IL-13, IL-25 and IL-33. The increased expression of the cytokine genes by TDI was partly attenuated after treatment with a TRPM8 antagonist. TDI exposure induces increased expression of TRPM8 mRNA in airway epithelial cells coupled with enhanced expression of inflammatory cytokines, suggesting a novel role of TRPM8 in the pathogenesis of TDI-induced OA.

No MeSH data available.


Immunocytochemical detection of TRPM8 protein in BEAS-2B cells. Immunocytochemical staining using a polyclonal antibody of TRPM8 demonstrated protein expression in BEAS-2B cells. The nuclei were counterstained blue using DAPI. (a) 1 mM of TDI intensified green staining in the plasma membranes in a time-dependent manner. (b) Pretreatment with TDI (0.25–1 mM) or (c) menthol (2–4 mM) for 3 h led to increased green staining in a dose-dependent manner. DAPI, 4′, 6-diamidino-2-phenylindole; TDI, toluene diisocyanate.
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fig3: Immunocytochemical detection of TRPM8 protein in BEAS-2B cells. Immunocytochemical staining using a polyclonal antibody of TRPM8 demonstrated protein expression in BEAS-2B cells. The nuclei were counterstained blue using DAPI. (a) 1 mM of TDI intensified green staining in the plasma membranes in a time-dependent manner. (b) Pretreatment with TDI (0.25–1 mM) or (c) menthol (2–4 mM) for 3 h led to increased green staining in a dose-dependent manner. DAPI, 4′, 6-diamidino-2-phenylindole; TDI, toluene diisocyanate.

Mentions: The expression of TRPM8 protein in BEAS-2B cells was evaluated by immunocytochemical analysis using a polyclonal antibody for human TRPM8 (Figure 3). Without any treatment, TRPM8 protein was not detected. However, after treatment with TDI or menthol, time- and dose-dependent intense green staining was detected. The staining pattern suggested localization of the TRPM8 to the plasma membrane (peripheral staining) and endoplasmic reticulum (perinuclear staining).


Toluene diisocyanate exposure induces airway inflammation of bronchial epithelial cells via the activation of transient receptor potential melastatin 8
Immunocytochemical detection of TRPM8 protein in BEAS-2B cells. Immunocytochemical staining using a polyclonal antibody of TRPM8 demonstrated protein expression in BEAS-2B cells. The nuclei were counterstained blue using DAPI. (a) 1 mM of TDI intensified green staining in the plasma membranes in a time-dependent manner. (b) Pretreatment with TDI (0.25–1 mM) or (c) menthol (2–4 mM) for 3 h led to increased green staining in a dose-dependent manner. DAPI, 4′, 6-diamidino-2-phenylindole; TDI, toluene diisocyanate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5382555&req=5

fig3: Immunocytochemical detection of TRPM8 protein in BEAS-2B cells. Immunocytochemical staining using a polyclonal antibody of TRPM8 demonstrated protein expression in BEAS-2B cells. The nuclei were counterstained blue using DAPI. (a) 1 mM of TDI intensified green staining in the plasma membranes in a time-dependent manner. (b) Pretreatment with TDI (0.25–1 mM) or (c) menthol (2–4 mM) for 3 h led to increased green staining in a dose-dependent manner. DAPI, 4′, 6-diamidino-2-phenylindole; TDI, toluene diisocyanate.
Mentions: The expression of TRPM8 protein in BEAS-2B cells was evaluated by immunocytochemical analysis using a polyclonal antibody for human TRPM8 (Figure 3). Without any treatment, TRPM8 protein was not detected. However, after treatment with TDI or menthol, time- and dose-dependent intense green staining was detected. The staining pattern suggested localization of the TRPM8 to the plasma membrane (peripheral staining) and endoplasmic reticulum (perinuclear staining).

View Article: PubMed Central - PubMed

ABSTRACT

Toluene diisocyanate (TDI) is the most important cause of occupational asthma (OA), and various pathogenic mechanisms have been suggested. Of these mechanisms, neurogenic inflammation is an important inducer of airway inflammation. Transient receptor potential melastatin 8 (TRPM8) is a well-established cold-sensing cation channel that is expressed in both neuronal cells and bronchial epithelial cells. A recent genome-wide association study of TDI-exposed workers found a significant association between the phenotype of TDI-induced OA and the single-nucleotide polymorphism rs10803666, which has been mapped to the TRPM8 gene. We hypothesized that TRPM8 located in airway epithelial cells may be involved in the pathogenic mechanisms of TDI-induced OA and investigated its role. Bronchial epithelial cells were treated with TDI in a dose- and time-dependent manner. The expression levels of TRPM8 mRNA and protein were determined by quantitative real-time polymerase chain reaction and western blotting. TDI-induced morphological changes in the cells were evaluated by immunocytochemistry. Alterations in the transcripts of inflammatory cytokines were examined in accordance with TRPM8 activation by TDI. TRPM8 expression at both the mRNA and protein levels was enhanced by TDI in airway epithelial cells. TRPM8 activation by TDI led to significant increases in the mRNA of interleukin (IL)-4, IL-13, IL-25 and IL-33. The increased expression of the cytokine genes by TDI was partly attenuated after treatment with a TRPM8 antagonist. TDI exposure induces increased expression of TRPM8 mRNA in airway epithelial cells coupled with enhanced expression of inflammatory cytokines, suggesting a novel role of TRPM8 in the pathogenesis of TDI-induced OA.

No MeSH data available.