Background: Mast cell (MC) - eosinophil (Eos) activating cross-talk might be critical for the severity and chronicity of allergy. Among soluble mediators, eosinophil major basic protein (MBP), a hallmark of allergy, is particularly important because it was shown to activate specific MC subtypes. We previously demonstrated that MBP activates IgE-desensitized rat MC and human lung and cord blood-derived MC (CBMC) after priming with fibroblast membranal stem cell factor. However, a distinct mechanism for this activation was missing. Therefore, we aimed to investigate it.
Methods: Major basic protein-1 activation of CBMC primed with fibroblast-derived membranes (FBM) was measured by β-hexosaminidase and tryptase release. Chemical cross-linking followed by micrometric flow cytometry probed direct interactions. Antibodies neutralized integrin-β1 and recognized its active form. Pertussis toxin (Ptx) was used to decrease integrin-β1 active form expression. Hematopoietic cell kinase (Hck) was identified by immunoprecipitation (IP) and silenced by siRNA.
Results: Major basic protein-1-induced CBMC activation is mediated partly by MBP1-integrin-β1 interaction on the MC surface. FBM prime CBMC via a G protein, as confirmed by Ptx, to shift integrin-β1 to its active form. Following MBP1 binding, integrin-β1 binds Hck that further transduces the activation signal. MC priming with FBM leads to up-regulation in Hck protein level. MC integrin-β1 neutralization inhibits MBP1-induced activation and uptake. Hck silencing results with reduced MBP1-induced activation.
Conclusions: Fibroblast-derived membranes, integrin-β1, and Hck are involved in MBP1-induced activation of CBMC and therefore represent a distinct mechanism for this activation. This finding might implicate integrin-β1 and Hck as targets for decreasing MC - Eos activating cross-talk in allergy.
Keywords: Hck; eosinophil major basic protein-1; fibroblast membrane fraction; integrin-β1; mast cells.
© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.