Epithelial IL-33 appropriates exosome trafficking for secretion in chronic airway disease
Ella Katz-Kiriakos, … , Mark J. Miller, Jennifer Alexander-Brett
Ella Katz-Kiriakos, … , Mark J. Miller, Jennifer Alexander-Brett
Published January 28, 2021
Citation Information: JCI Insight. 2021;6(4):e136166. https://doi.org/10.1172/jci.insight.136166.
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Research Article Immunology Pulmonology

Epithelial IL-33 appropriates exosome trafficking for secretion in chronic airway disease

Abstract

IL-33 is a key mediator of chronic airway disease driven by type 2 immune pathways, yet the nonclassical secretory mechanism for this cytokine remains undefined. We performed a comprehensive analysis in human airway epithelial cells, which revealed that tonic IL-33 secretion is dependent on the ceramide biosynthetic enzyme neutral sphingomyelinase 2 (nSMase2). IL-33 is cosecreted with exosomes by the nSMase2-regulated multivesicular endosome (MVE) pathway as surface-bound cargo. In support of these findings, human chronic obstructive pulmonary disease (COPD) specimens exhibited increased epithelial expression of the abundantly secreted IL33Δ34 isoform and augmented nSMase2 expression compared with non-COPD specimens. Using an Alternaria-induced airway disease model, we found that the nSMase2 inhibitor GW4869 abrogated both IL-33 and exosome secretion as well as downstream inflammatory pathways. This work elucidates a potentially novel aspect of IL-33 biology that may be targeted for therapeutic benefit in chronic airway diseases driven by type 2 inflammation.

Authors

Ella Katz-Kiriakos, Deborah F. Steinberg, Colin E. Kluender, Omar A. Osorio, Catie Newsom-Stewart, Arjun Baronia, Derek E. Byers, Michael J. Holtzman, Dawn Katafiasz, Kristina L. Bailey, Steven L. Brody, Mark J. Miller, Jennifer Alexander-Brett

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Figure 4

IL33 isoforms in COPD specimens.

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IL33 isoforms in COPD specimens. (A) IL33 isoform–specific qPCR in lung ...
(A) IL33 isoform–specific qPCR in lung tissue specimens for IL33full (non-COPD, n = 13; COPD, n = 27) and IL33Δ34 (non-COPD, n = 12; COPD, n = 23). (B) In situ hybridization in non-COPD and COPD tissues targeting the IL33Δ34 isoform with increased staining in COPD tissue (violet). Nuclear fast red counterstain was also used. Scale bar: 50 μm (gray); 10 μm (white). (C) Immunostaining of non-COPD and COPD tissue with IL-33 (green) and cytokeratin 5 (Krt5, red), showing variable cytoplasmic IL-33 staining prominent in COPD 3; DAPI counterstain was also used. Scale bar: 10 μm (white). (D) Epitopes for IL-33 NTD and CTD targeting antibodies and reactivity validated with IL-33 variants expressed in HEK293T cells: IL-33full 38 kDa (red, 30 kDa degraded band), IL-33Δ34 28 kDa (blue), and IL-33 CTD 17 kDa (black, Peprotech). (E) Matched tissue and bronchial wash (BW) non-COPD (n = 3) and COPD (n = 6) specimens. Western blot shows multiple fragments detected by CTD antibody in tissue and a 28 kDa band detected by CTD but not NTD antibody in BW. ELISA-quantified IL-33 in the same samples, normalized to total protein. IL-33 protein quantity was increased and soluble ST2 (sST2) was decreased in COPD. COPD 3 with the highest level of BW IL-33 protein is highlighted in yellow. (F) IL33 isoform–specific qPCR in airway basal cells for IL33full (non-COPD, n = 12; COPD, n = 26) and IL33Δ34 (non-COPD, n = 11; COPD, n = 24) (mean ± SEM). (G) Western blot on a subset of airway basal cell lysates showing a 28 kDa band detected in COPD by CTD but not NTD antibody. ELISA-quantified normalized IL-33 protein in non-COPD (n = 12) and COPD (n = 26) cells, showing that IL-33 is increased in COPD. Data are shown as the mean ± SEM. Statistical analysis: t test (A, E, F, and G); **P < 0.01.