[Menthol enhances interleukin-13-induced synthesis and secretion of mucin 5AC in human bronchial epithelial cells].

OBJECTIVE

To investigate the effect of interleukin (IL) -13 combined with cold stimulation on synthesis and secretion of mucin (MUC) 5AC in human bronchial epithelial cell line 16HBE and explore the role of transient receptor potential 8 (TRPM8) and anti-apoptotic factor B-cell lymphoblast-2 (Bcl-2) in this process.

METHODS

16HBE cells were stimulated with 10 ng/mL IL-13, 1 mmol/L menthol, or both (1 mmol/L menthol was added after 6 days of IL-13 stimulation), and the changes in the expression of MUC5AC, intracellular Ca concentration and Bcl-2 expression were evaluated. The effects of ABT-263 (a Bcl-2 inhibitor) and BCTC (a TRPM8 ion channel inhibitor), alone or in combination, on MUC5AC expression in the cells were tested, and the changes in intracellular Ca and Bcl-2 expression following BCTC treatment were observed. The cell viability was assessed using CCK-8 assay, the mRNA expressions of MUC5AC and Bcl-2 were detected with real-time quantitative PCR, the level of MUC5AC in the culture medium was measured with ELISA, and the intracellular Ca fluorescence intensity was determined with flow cytometry.

RESULTS

The mRNA and protein expressions of MUC5AC increased significantly in 16HBE cells following stimulation with IL-13, menthol, and both ( < 0.05), and were the highest in the combined treatment group with its peak level occurred at 24 h ( < 0.01). The intracellular Ca fluorescence intensity and Bcl-2 mRNA expression were also increased in 16HBE cells after the stimulations ( < 0.05), and the increments were the most obvious in the combined treatment group ( < 0.01). Treatment with BCTC significantly lowered intracellular Ca fluorescence intensity and the expressions of Bcl-2 and MUC5AC mRNA and protein in the cells stimulated with menthol or with both IL-13 and menthol ( < 0.05), but caused no significant changes in IL-13-stimulated cells (P > 0.05). Treatment with ABT-263 significantly lowered the mRNA and protein expressions of MUC5AC in the cells stimulated with IL-13 and menthol either alone or in combination ( < 0.05).

CONCLUSIONS

Menthol combined with IL-13 produces a synergistic effect to promote the synthesis and secretion of MUC5AC in 16HBE cells possibly by activating TRPM8 receptor to upregulate the expression of Bcl-2.