Autophagy regulates DUOX1 localization and superoxide production in airway epithelial cells during chronic IL-13 stimulation

John D Dickinson, Jenea M. Sweeter, Kristi J. Warren, Iman M Ahmad, Xavier De Deken, Matthew C Zimmerman, Steven L. Brody

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

The airway epithelium is a broad interface with the environment, mandating well-orchestrated responses to properly modulate inflammation. Classically, autophagy is a homeostatic pathway triggered in response to external cellular stresses, and is elevated in chronic airway diseases. Recent findings highlight the additional role of autophagy in vesicle trafficking and protein secretion, implicating autophagy pathways in complex cellular responses in disease. Th2 cytokines, IL-13 and IL-4, are increased in asthma and other airway diseases contributing to chronic inflammation. Previously, we observed that IL-13 increases reactive oxygen species (ROS) in airway epithelial cells in an autophagy-dependent fashion. Here, we tested our hypothesis that autophagy is required for IL-13-mediated superoxide production via the NADPH oxidase DUOX1. Using a mouse model of Th2-mediated inflammation induced by OVA-allergen, we observed elevated lung amounts of IL-13 and IL-4 accompanied by increased autophagosome levels, determined by LC3BII protein levels and immunostaining. ROS levels were elevated and DUOX1 expression was increased 70-fold in OVA-challenged lungs. To address the role of autophagy and ROS in the airway epithelium, we treated primary human tracheobronchial epithelial cells with IL-13 or IL-4. Prolonged, 7-day treatment increased autophagosome formation and degradation, while brief activation had no effect. Under parallel culture conditions, IL-13 and IL-4 increased intracellular superoxide levels as determined by electron paramagnetic resonance (EPR) spectroscopy. Prolonged IL-13 activation increased DUOX1, localized at the apical membrane. Silencing DUOX1 by siRNA attenuated IL-13-mediated increases in superoxide, but did not reduce autophagy activities. Notably, depletion of autophagy regulatory protein ATG5 significantly reduced superoxide without diminishing total DUOX1 levels. Depletion of ATG5, however, diminished DUOX1 localization at the apical membrane. The findings suggest non-canonical autophagy activity regulates DUOX1-dependent localization required for intracellular superoxide production during Th2 inflammation. Thus, in chronic Th2 inflammatory airway disease, autophagy proteins may be responsible for persistent intracellular superoxide production.

Original languageEnglish (US)
Pages (from-to)272-284
Number of pages13
JournalRedox Biology
Volume14
DOIs
StatePublished - Apr 2018

Fingerprint

Interleukin-13
Autophagy
Superoxides
Epithelial Cells
Interleukin-4
Reactive Oxygen Species
Inflammation
Proteins
Chemical activation
Membranes
Epithelium
NADPH Oxidase
Lung
Allergens
Small Interfering RNA
Paramagnetic resonance
Electron Spin Resonance Spectroscopy
Protein Transport
Spectroscopy
Cytokines

Keywords

  • Asthma
  • Autophagy
  • DUOX1
  • Electron Paramagnetic Resonance Spectroscopy
  • Epithelial cells
  • IL-13
  • IL-4
  • Reactive oxygen species
  • Superoxide

ASJC Scopus subject areas

  • Biochemistry
  • Organic Chemistry

Cite this

Autophagy regulates DUOX1 localization and superoxide production in airway epithelial cells during chronic IL-13 stimulation. / Dickinson, John D; Sweeter, Jenea M.; Warren, Kristi J.; Ahmad, Iman M; De Deken, Xavier; Zimmerman, Matthew C; Brody, Steven L.

In: Redox Biology, Vol. 14, 04.2018, p. 272-284.

Research output: Contribution to journalArticle

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AU - De Deken, Xavier

AU - Zimmerman, Matthew C

AU - Brody, Steven L.

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AB - The airway epithelium is a broad interface with the environment, mandating well-orchestrated responses to properly modulate inflammation. Classically, autophagy is a homeostatic pathway triggered in response to external cellular stresses, and is elevated in chronic airway diseases. Recent findings highlight the additional role of autophagy in vesicle trafficking and protein secretion, implicating autophagy pathways in complex cellular responses in disease. Th2 cytokines, IL-13 and IL-4, are increased in asthma and other airway diseases contributing to chronic inflammation. Previously, we observed that IL-13 increases reactive oxygen species (ROS) in airway epithelial cells in an autophagy-dependent fashion. Here, we tested our hypothesis that autophagy is required for IL-13-mediated superoxide production via the NADPH oxidase DUOX1. Using a mouse model of Th2-mediated inflammation induced by OVA-allergen, we observed elevated lung amounts of IL-13 and IL-4 accompanied by increased autophagosome levels, determined by LC3BII protein levels and immunostaining. ROS levels were elevated and DUOX1 expression was increased 70-fold in OVA-challenged lungs. To address the role of autophagy and ROS in the airway epithelium, we treated primary human tracheobronchial epithelial cells with IL-13 or IL-4. Prolonged, 7-day treatment increased autophagosome formation and degradation, while brief activation had no effect. Under parallel culture conditions, IL-13 and IL-4 increased intracellular superoxide levels as determined by electron paramagnetic resonance (EPR) spectroscopy. Prolonged IL-13 activation increased DUOX1, localized at the apical membrane. Silencing DUOX1 by siRNA attenuated IL-13-mediated increases in superoxide, but did not reduce autophagy activities. Notably, depletion of autophagy regulatory protein ATG5 significantly reduced superoxide without diminishing total DUOX1 levels. Depletion of ATG5, however, diminished DUOX1 localization at the apical membrane. The findings suggest non-canonical autophagy activity regulates DUOX1-dependent localization required for intracellular superoxide production during Th2 inflammation. Thus, in chronic Th2 inflammatory airway disease, autophagy proteins may be responsible for persistent intracellular superoxide production.

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