Directional secretomes reflect polarity-specific functions in an in vitro model of human bronchial epithelium

The polarity of the conducting airway epithelium is responsible for its directional secretion. This is an essential characteristic of lung integrity and function that dictates interactions between the external environment (apical) and subepithelial structures (basolateral). Defining the directional secretomes in the in vitro human bronchial epithelial (HBE) differentiated model could bring valuable insights into lung biology and pulmonary diseases. Normal primary HBE cells (n = 3) were differentiated into respiratory tract epithelium. Apical and basolateral secretions (24 h) were processed for proteome profiling and pathway analysis. A total of 243 proteins were identified in secretions from all HBE cultures combined. Of these, 51% were classified as secreted proteins, including true secreted proteins (36%) and exosomal proteins (15%). Close examination revealed consistent secretion of 69 apical proteins and 13 basolateral proteins and differential secretion of 25 proteins across all donors. Expression of Annexin A4 in apical secretions and Desmoglein-2 in basolateral secretions was validated using Western blot or ELISA in triplicate independent experiments. To the best of our knowledge, this is the first study defining apical and basolateral secretomes in the in vitro differentiated HBE model. The data demonstrate that epithelial polarity directs protein secretion with different patterns of biological processes to the apical and basolateral surfaces that are consistent with normal bronchial epithelium homeostatic functions. Applying this in vitro directional secretome model to lung diseases may elucidate their molecular pathophysiology and help define potential therapeutic targets.