Chitinase-Induced Airway Hyperreactivity and Inflammation in a Mouse Model of Nonallergic Asthma

Abstract

Introduction: Environmental exposure to mites and fungi has been proposed to critically contribute to the development of IgE-mediated asthma. A common denominator of such organisms is chitin. Human chitinases have been reported to be upregulated by interleukin-13 secreted in the context of Th2-type immune responses and to induce asthma. We assessed whether chitin-containing components induced chitinases in an innate immune-dependent way and whether this results in bronchial hyperresponsiveness.

Materials and methods: Monocyte/macrophage cell lines were stimulated with chitin-containing or bacterial components in vitro. Chitinase activity in the supernatant and the expression of the chitotriosidase gene were measured by enzyme assay and quantitative PCR, respectively. Non-sensitized mice were stimulated with chitin-containing components intranasally, and a chitinase inhibitor was administered intraperitoneally. As markers for inflammation leukocytes were counted in the bronchoalveolar lavage (BAL) fluid, and airway hyperresponsiveness was assessed via methacholine challenge.

Results: We found both whole chitin-containing dust mites as well as the fungal cell wall component zymosan A but not endotoxin-induced chitinase activity and chitotriosidase gene expression in vitro. The intranasal application of zymosan A into mice led to the induction of chitinase activity in the BAL fluid and to bronchial hyperresponsiveness, which could be reduced by applying the chitinase inhibitor allosamidin.

Discussion: We propose that environmental exposure to mites and fungi leads to the induction of chitinase, which in turn favors the development of bronchial hyperreactivity in an IgE-independent manner.

Keywords: Airway inflammation; Chitinase; Fungi; Innate immune system; Mites.

Fig. 1

Elevated chitinase activity after mite or fungi stimulation. Increased chitinase enzyme activity in supernatant (a) and increased chitotriosidase gene expression in THP-1 cells after stimulation with Dermatophagoides pteronyssinus or zymosan (b). No significant difference after LPS stimulation. Data are mean ± SEM, n = 3.

Fig. 2

Decreased gene expression of chitotriosidase mRNA was measured after stimulation of THP-1 cells with zymosan and inhibitory antibodies for Dec 206 (a), TLR-2 (b), or dectin-1 receptors (c). After addition of Dec 205, SIGN, or SIGN-related receptor antibodies, there was no inhibition of chitinase gene expression detected. Data are SEM, n = 3. THP-1, human monocyte cell line.

Fig. 3

BALB/c mice showed a significant increase of chitinase activity in BAL after daily intranasally stimulation with zymosan. After zymosan stimulation and daily intraperitoneal application of chitinase inhibitor allosamidin, we determined a decrease in enzyme activity. Data are mean ± SEM. BAL, bronchoalveolar lavage.

Fig. 4

Increased airway hyperresponsiveness and inflammation after intranasal fungal stimulation. Airway hyperresponsiveness (a) and airway inflammation (b) shown as cells/mL in the BAL fluid of nonallergic BALB/c mice. Results after intranasal and intraperitoneal stimulation with PBS (a: n = 4, b: n = 3) as negative control, intranasal stimulation with zymosan with intraperitoneal PBS (a: n = 8, b: n = 7) or the chitinase inhibitor allosamidin (a: n = 8, b: n = 7). An increase of airway inflammation and hyperresponsiveness after chitin-containing zymosan and a downward trend after intraperitoneal chitinase inhibition could be shown. Data are mean ± SEM. BAL, bronchoalveolar lavage.