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IL-5-stimulated eosinophils adherent to periostin undergo stereotypic morphological changes and ADAM8-dependent
M W Johansson1, M Khanna1, V Bortnov1
1Department of Biomolecular Chemistry, University of Wisconsin, Madison, WI, USA.
Background:
IL-5 causes suspended eosinophils to polarize with filamentous (F)-actin and granules at one pole and the nucleus in a specialized uropod, the "nucleopod," which is capped with P-selectin glycoprotein ligand-1 (PSGL-1). IL-5 enhances eosinophil adhesion and migration on periostin, an extracellular matrix protein upregulated in asthma by type 2 immunity mediators.
Objective:
Determine how the polarized morphology evolves to foster migration of IL-5-stimulated eosinophils on a surface coated with periostin.
Methods:
Blood eosinophils adhering to adsorbed periostin were imaged at different time points by fluorescent microscopy, and migration of eosinophils on periostin was assayed.
Results:
After 10 minutes in the presence of IL-5, adherent eosinophils were polarized with PSGL-1 at the nucleopod tip and F-actin distributed diffusely at the opposite end. After 30-60 minutes, the nucleopod had dissipated such that PSGL-1 was localized in a crescent or ring away from the cell periphery, and F-actin was found in podosome-like structures. The periostin layer, detected with monoclonal antibody Stiny-1, shown here to recognize the FAS1 4 module, was cleared in wide areas around adherent eosinophils. Clearance was attenuated by metalloproteinase inhibitors or antibodies to disintegrin metalloproteinase 8 (ADAM8), a major eosinophil metalloproteinase previously implicated in asthma pathogenesis. ADAM8 was not found in podosome-like structures, which are associated with proteolytic activity in other cell types. Instead, immunoblotting demonstrated proteoforms of ADAM8 that lack the cytoplasmic tail in the supernatant. Anti-ADAM8 inhibited migration of IL-5-stimulated eosinophils on periostin.
Conclusions And Clinical Relevance:
Migrating IL-5-activated eosinophils on periostin exhibit loss of nucleopodal features and appearance of prominent podosomes along with clearance of the Stiny-1 periostin epitope. Migration and epitope clearance are both attenuated by inhibitors of ADAM8. We propose, therefore, that eosinophils remodel and migrate on periostin-rich extracellular matrix in the asthmatic airway in an ADAM8-dependent manner, making ADAM8 a possible therapeutic target.