Protease-activated receptor-2 (PAR2) mutation attenuates airway fibrosis in mice during the exacerbation of house dust mite‑induced allergic lung disease by multi‑walled carbon nanotubes.

BACKGROUND

Pulmonary exposure to multi-walled carbon nanotubes (MWCNTs) induces potent pro-inflammatory and pro-fibrotic responses in mouse models of allergic lung disease. We recently reported that MWCNTs exacerbated components of house dust mite (HDM)-induced allergic lung disease, including eosinophilic inflammation, mucous cell metaplasia and airway fibrosis. Protease-activated receptor 2 (PAR2) plays a significant role in the development of various respiratory diseases, including asthma and pulmonary fibrosis. However, studies investigating the function of PAR2 in allergic lung disease have produced variable results. To further define the role of PAR2 in pulmonary pathology, we investigated the effects of MWCNTs on HDM-induced allergic lung disease in PAR2-mutant mice.

METHODS

The PAR2-mutant mice used were previously generated by replacing a 1.8-kb region of the PAR2 coding sequence with a neomycin resistance gene, which did not entirely delete the gene. Wild-type (WT) male C57BL/6J mice and PAR2-mutant male mice were exposed to a vehicle solution, MWCNTs, HDM extract, or both via oropharyngeal aspiration six times over 3 weeks. Bronchoalveolar lavage fluid (BALF) was collected to measure changes in inflammatory cells, total protein, and lactate dehydrogenase (LDH). Lung protein and mRNA were assayed for pro-inflammatory and profibrotic mediators, and formalin-fixed lung sections were evaluated for histopathology.

RESULTS

In WT and PAR2-mutant mice, co-exposure to MWCNTs and HDM extract significantly increased eosinophilic lung inflammation, mucous cell metaplasia, increased BALF cellularity, BALF total protein, and LDH levels. These results were not significantly different between genotypes. Additionally, MWCNTs and HDM extract co-exposure significantly increased airway fibrosis in WT and PAR2-mutant mice, characterized by increased airway collagen deposition and Col1a1 mRNA expression. Quantitative morphometry revealed a significant decrease in airway fibrosis in PAR2-mutant mice compared to WT mice, accompanied by reduced Col1a1 mRNA as detected by PCR. Despite this reduction, the pro-fibrotic mediator arginase 1 (Arg-1) protein and mRNA levels were significantly upregulated in PAR2-mutant mice.

CONCLUSION

Our study demonstrates that PAR2 mediates airway fibrosis but does not influence eosinophilic lung inflammation or mucous cell metaplasia caused by co-exposure to MWCNTs and HDM allergen.