Targeting integrin α5β1 ameliorates severe airway hyperresponsiveness in experimental asthma
Aparna Sundaram, … , Xiaozhu Huang, Dean Sheppard
Aparna Sundaram, … , Xiaozhu Huang, Dean Sheppard
Published December 5, 2016
Citation Information: J Clin Invest. 2017;127(1):365-374. https://doi.org/10.1172/JCI88555.
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Research Article Muscle biology Pulmonology

Targeting integrin α5β1 ameliorates severe airway hyperresponsiveness in experimental asthma

Abstract

Treatment options are limited for severe asthma, and the need for additional therapies remains great. Previously, we demonstrated that integrin αvβ6-deficient mice are protected from airway hyperresponsiveness, due in part to increased expression of the murine ortholog of human chymase. Here, we determined that chymase protects against cytokine-enhanced bronchoconstriction by cleaving fibronectin to impair tension transmission in airway smooth muscle (ASM). Additionally, we identified a pathway that can be therapeutically targeted to mitigate the effects of airway hyperresponsiveness. Administration of chymase to human bronchial rings abrogated IL-13–enhanced contraction, and this effect was not due to alterations in calcium homeostasis or myosin light chain phosphorylation. Rather, chymase cleaved fibronectin, inhibited ASM adhesion, and attenuated focal adhesion phosphorylation. Disruption of integrin ligation with an RGD-containing peptide abrogated IL-13–enhanced contraction, with no further effect from chymase. We identified α5β1 as the primary fibronectin-binding integrin in ASM, and α5β1-specific blockade inhibited focal adhesion phosphorylation and IL-13–enhanced contraction, with no additional effect from chymase. Delivery of an α5β1 inhibitor into murine airways abrogated the exaggerated bronchoconstriction induced by allergen sensitization and challenge. Finally, α5β1 blockade enhanced the effect of the bronchodilator isoproterenol on airway relaxation. Our data identify the α5β1 integrin as a potential therapeutic target to mitigate the severity of airway contraction in asthma.

Authors

Aparna Sundaram, Chun Chen, Amin Khalifeh-Soltani, Amha Atakilit, Xin Ren, Wenli Qiu, Hyunil Jo, William DeGrado, Xiaozhu Huang, Dean Sheppard

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Figure 5

Blockade of α5β1 inhibits focal adhesion phosphorylation and impairs IL-13–enhanced tracheal ring contraction.

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Blockade of α5β1 inhibits focal adhesion phosphorylation and impairs IL-...
(A) Representative Western blot and quantitative densitometry for phosphorylated and total vinculin and FAK in human ASM cells treated for 20 minutes with integrin α5–blocking antibody (P1D6, 10 μg/ml) or mouse IgG control, then plated on poly-l-lysine (300 μg/ml) or poly-l-lysine with either fibronectin (1 μg/ml) or collagen I (1 μg/ml). GAPDH was used as a loading control. *P < 0.05 and **P < 0.01 versus Poly; #P < 0.05 versus Poly/FN. n = 3 distinct experiments. GAPDH was used as a loading control. *P < 0.05 and **P <0.01 versus Poly; #P < 0.05 versus Poly/FN, by 2-way ANOVA n = 3 distinct experiments. Data represent the mean ± SEM. (B) Contractile force of mouse tracheal rings measured after incubation for 12 hours in DMEM with IL-13 (100 ng/ml) or saline and for 12 hours with α5-blocking antibody (300 μg/ml) or rat IgG in response to a range of concentrations of the contractile agonist Mch. ***P < 0.001, for IL-13/IgG versus IL-13/anti-α5. (C) Contractile force of mouse tracheal rings measured after incubation for 12 hours in DMEM with IL-13 (100 ng/ml) or saline, then for 1 hour with a small-molecule inhibitor directed against the α5β1 synergy site of fibronectin (ATN-161, 100 μg/ml) or vehicle in response to a range of concentrations of Mch. **P < 0.01 and ***P < 0.001, for IL-13/vehicle versus IL-13/ATN-161. (D) Contractile force of mouse tracheal rings measured after incubation for 12 hours in DMEM with IL-13 (100 ng/ml) or saline (control), then for 1 hour with ATN-161 (100 μg/ml), 20 minutes with rhChy (30 nM), or both, in response to a range of concentrations of Mch. ***P < 0.001, for IL-13/vehicle versus IL-13/ATN-161 plus rhChy. NS, for IL-13/ATN-161 versus IL-13/rhChy versus IL-13/ATN-161 plus rhChy. (B–D) Data represent the mean ± SEM; n = 3–5 rings per group; significance was determined by repeated measures of variance.