Activation of Human Basophils by A549 Lung Epithelial Cells Reveals a Novel IgE-Dependent Response Independent of Allergen

Abstract

Evidence for epithelial cell (EC)-derived cytokines (e.g. TSLP) activating human basophils remains controversial. We therefore hypothesize that ECs can directly activate basophils via cell-to-cell interaction. Basophils in medium alone or with IL-3±anti-IgE, were co-incubated with TSLP, IL-33, or IL-25. Analogous experiments co-cultured basophils (1–72h) directly with EC lines. Supernatants were tested for mediators and cytokines. Antibodies targeting receptors were tested for neutralizing effects. Lactic acid (pH 3.9) treatment combined with passive sensitization tested the role of IgE. Overall, IL-33 augmented IL-13 secretion from basophils co-treated with IL-3, with minimal effects on histamine and IL-4. Conversely, basophils (but not mast cells) released histamine and marked levels of IL-4/IL-13 (10-fold) when co-cultured with A549 EC and IL-3, without exogenous allergen or IgE cross-linking stimuli. The inability to detect IL-33/TSLP, or to neutralize their activity, suggested a unique mode of basophil activation by A549 EC. Half-maximal rates for histamine (4h) and IL-4 (5h) secretion were slower than observed with standard IgE-dependent activation. Immunoglobulin stripping combined with passive sensitization±omalizumab showed a dependency for basophil-bound IgE, substantiated by requirement for cell-to-cell contact, aggregation, and FcεRI-dependent signaling. A yet unidentified IgE-binding lectin associated with A549 EC is implicated after discovering that n-acetyllactosamine suppressed basophil activation in co-cultures. These findings point to a lectin-dependent activation of basophil requiring IgE but independent of allergen or secreted cytokine. Pending further investigation, we predict this unique mode of activation is linked to inflammatory conditions whereby IgE-dependent activation of basophils occurs despite absence of any known allergen.

Epithelial Cell-Associated Galectin-3 Activates Human Dendritic Cell Subtypes for Pro-Inflammatory Cytokines

“…Cultures were split twice weekly during the duration of these experiments, with no detectable changes in their capacity to activate DC. Seeding of EC for co-culture with basophils has been previously described ( 9 ) and was similarly done herein for DC co-cultures. Briefly, EC were plated into wells of 96-well plates in 0.100 ml of the medium.…”

“…Surface Gal-3 protein was detected using flow cytometry, as previously described ( 9 ). The expression of various markers on DC subtypes isolated from blood and those expressed by CDDC were also assessed using flow cytometry.…”

“…Naturally, the interaction of allergen with specific IgE bound to FcεRI is the mode of stimulation most often associated with facilitating these responses. However, we recently reported evidence for a unique mode of IgE-dependent activation in basophils whereby these granulocytes released histamine and secreted large quantities of IL-4/IL-13 when co-cultured with the A549 epithelial cell (EC) line –an adenocarcinoma of lung origin ( 9 ). These responses proved dependent on cell-to-cell contact and on the expression of IgE by basophils.…”

Human mast cells and basophils—How are they similar how are they different?

“…Recent evidence shows that human basophils secrete high levels of histamine, IL‐4 and IL‐13 when co‐cultured with A549 lung epithelial cells in a reaction requiring cell‐to‐cell interactions but independent of known epithelial‐derived cytokines (eg, TSLP, IL‐33). The underlying mechanism of this response appears dependent on an A549‐associated lectin (possibly galectin‐3) interacting with basophil‐bound IgE to induce a unique FcεRI‐dependent activation that differs from that mediated by allergens …”

Possible roles of basophils in chronic itch

“…This histamine release is provoked through the cross‐linking of the IgE receptors on basophils by anti‐IgG treatment . Interestingly, human basophils with IgE are also able to release histamine in the absence of an antigen when they interact with IgE‐binding lectin that is expressed by a certain type of epithelial cells . NGF, a neurotrophin that may also be involved in itch through multifocal pathways, enhances histamine release from human basophils via its receptor tropomyosin receptor kinase A (TrkA) …”