title: Pollen proteases compromise the airway epithelial barrier through degradation of transmembrane adhesion proteins and lung bioactive peptides.
abstract: BACKGROUND
  Allergic disorders, such as seasonal rhinitis and asthma, are increasing causes of morbidity worldwide and often result from exposure to airborne pollen. Pollen allergy has a remarkable clinical impact all over Europe. In fact, epidemiological longitudinal studies confirm that pollen species usually considered with low allergenic potential became more recently responsible for intense allergic reactions. In this study, we aimed to characterize major pollen proteolytic activity and evaluate its contribution to the immunologic and inflammatory response to airborne allergens.
  
  
  METHODS
  Proteolytic activity in four pollen diffusates with distinct allergenicity, Olea europaea, Dactylis glomerata, Cupressus sempervirens and Pinus sylvestris, was evaluated through several enzymatic assays. The action of pollen proteases on the paracellular integrity of Calu-3, grown at the air-liquid interphase, was evaluated through a transepithelial permeability assay. Immunoblot and immunofluorescence experiments were performed to analyse the disruption of intercellular complexes. Degradation of bioactive peptides by pollen crude extracts was assessed by mass spectrometry.
  
  
  RESULTS
  All pollen diffusates were shown to have high molecular weight proteases with serine and/or aminopeptidase activity. These proteases increased Calu-3 transepithelial permeability through disruption of transmembrane adhesion proteins: occludin, claudin-1 and E-cadherin. Moreover, they were able to degrade airway bioactive peptides and were not blocked by endogenous protease inhibitors.
  
  
  CONCLUSIONS
  Pollen grains with distinct allergenic abilities release proteases that might be involved in the sensitization to a range of airborne allergens by facilitating allergen delivery across the epithelium and also contribute directly to the inflammation characteristic of allergic diseases.
authors.affiliation: Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal.
authors.forename: R
authors.initials: R
authors.lastname: Vinhas
authors.forename: L
authors.initials: L
authors.lastname: Cortes
authors.forename: I
authors.initials: I
authors.lastname: Cardoso
authors.forename: V M
authors.initials: VM
authors.lastname: Mendes
authors.forename: B
authors.initials: B
authors.lastname: Manadas
authors.forename: A
authors.initials: A
authors.lastname: Todo-Bom
authors.forename: E
authors.initials: E
authors.lastname: Pires
authors.forename: P
authors.initials: P
authors.lastname: Veríssimo
mesh_terms: D000485:Allergens / Q000378:metabolism; D001692:Biological Transport; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D006801:Humans; D008168:Lung / Q000737:chemistry*; D008565:Membrane Proteins; D010447:Peptide Hydrolases / Q000276:immunology* / Q000378:metabolism; D010455:Peptides / Q000378:metabolism*; D011058:Pollen / Q000201:enzymology*; D020545:Respiratory Mucosa / Q000737:chemistry*; D006255:Rhinitis, Allergic, Seasonal / Q000209:etiology / Q000276:immunology
chemical_list: D000485:Allergens; D015815:Cell Adhesion Molecules; D008565:Membrane Proteins; D010455:Peptides; D010447:Peptide Hydrolases