Abstract
BackgroundIn the airways, mast cells are present in close vicinity to epithelial cells, and they can interact with each other via multiple factors, including extracellular vesicles (EVs). Mast cell-derived EVs have a large repertoire of cargos, including proteins and RNA, as well as surface DNA. In this study, we hypothesized that these EVs can induce epithelial to mesenchymal transition (EMT) in airway epithelial cells.MethodsIn this in-vitro study we systematically determined the effects of mast cell-derived EVs on epithelial A549 cells. We determined the changes that are induced by EVs on A549 cells at both the RNA and protein levels. Moreover, we also analyzed the rapid changes in phosphorylation events in EV-recipient A549 cells using a phosphorylated protein microarray. Some of the phosphorylation-associated events associated with EMT were validated using immunoblotting.ResultsMorphological and transcript analysis of epithelial A549 cells indicated that an EMT-like phenotype was induced by the EVs. Transcript analysis indicated the upregulation of genes involved in EMT, including TWIST1, MMP9, TGFB1, and BMP-7. This was accompanied by downregulation of proteins such as E-cadherin and upregulation of Slug-Snail and matrix metalloproteinases. Additionally, our phosphorylated-protein microarray analysis revealed proteins associated with the EMT cascade that were upregulated after EV treatment. We also found that transforming growth factor beta-1, a well-known EMT inducer, is associated with EVs and mediates the EMT cascade induced in the A549 cells.ConclusionMast cell-derived EVs mediate the induction of EMT in epithelial cells, and our evidence suggests that this is triggered through the induction of protein phosphorylation cascades.
Highlights
In the airways, mast cells are present in close vicinity to epithelial cells, and they can interact with each other via multiple factors, including extracellular vesicles (EVs)
Morphological changes are induced in epithelial cells upon uptake of mast cell-derived EVs Mast cells are known to reside in the proximity of airway epithelial cells and can readily interact with epithelial cells
The expression of several genes involved in epithelial to mesenchymal transition (EMT) such as TGFB1, SMAD2, TWIST1, MMP9, MMP2, WNT5A, FOXC2, BMP7, and VIM were evaluated in A549 cells
Summary
Mast cells are present in close vicinity to epithelial cells, and they can interact with each other via multiple factors, including extracellular vesicles (EVs). Cells that are present in close proximity to each other, including immune cells recruited to epithelial tissues, are in constant communication with each other, both in disease or homeostatic condition [1,2,3,4] This communication can be induced by direct cell-to-cell contact, secreted free proteins, or shuttling of bioactive molecules via extracellular vesicles (EVs). EV-based cellular cross talk is an evolutionarily conserved form of communication, and EVs can induce signaling through their cargoes of protein, DNA, and RNA [5,6,7,8,9] With their varied cargo, EVs have the potential to activate multiple signaling events in recipient cells. In this study we determined the potential of mast cell-derived EVs to regulate EMT, and we identified the potential signaling events in recipient epithelial cells
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