Tiotropium Bromide Attenuated Innate-Type Airway Inflammation Through Reducing Basophil-Derived IL-4 Production

Abstract

Background: Tiotropium bromide (TIO) is a long-acting muscarinic antagonist (LAMA). In moderate to severe asthma patients, adding TIO to inhaled corticosteroid and long-acting betaagonist reduces the frequency of exacerbation. Recent studies demonstrated that TIO acts on airway epithelium and CD4 lymphocytes to attenuate inflammation, but the underlying mechanism is not clear enough. Asthma exacerbation occurs in response to exposure to external agents, such as viral infection, pollen and environmental pollution, and group 2 innate lymphoid cells (ILC2s) are considered to be involved in the pathogenesis of asthma exacerbation. Objective: We investigated whether TIO modulates airway inflammation through ILC2 functions. Methods: Mice were administrated papain intranasally with or without TIO. Bronchoalveolar lavage fluids (BALF) and lung tissues were collected, and the levels of IL-5 and IL-13 in BALF were measured by ELISA. Isolated ILC2s from lungs and bone-marrow-derived basophils were stimulated in vitro with IL-33 in the presence or absence of TIO to determine whether the engagement of TIO altered the cytokine production. The levels of cytokines in the supernatant were measured by ELISA. The expression of M3R on immune cells were assessed by RNA sequence. Results: Papain induced eosinophilic airway inflammation and the pretreatment with TIO reduced the number of eosinophils in BALF compared to vehicle. The concentration of IL-5 and IL-13 in BALF was reduced by TIO treatment. TIO also reduced the frequencies of ILC2s analyzed by flow cytometry in BALF. Stimulation of ILC2s with IL-33 increased IL-5 and IL-13 levels in supernatant, but pretreatment with TIO did not affected IL-33-induced cytokine production from ILC2s, suggesting that TIO does not regulate ILC2s directly. The geneexpression analysis showed that basophils predominantly expressed M3R. Stimulation of basophils with IL-33 increased IL-4 production, and pretreatment with TIO reduced IL-4 production from basophils. Conclusion: TIO attenuated innate-type airway inflammation. TIO regulated the activation of ILC2 by reducing IL-4 production from basophils.