Exosomes derived from LPS-stimulated macrophages promote TGF-β1-induced epithelial-mesenchymal transition of human type II alveolar epithelial A549 cells

Objective To investigate the impact of the macrophage-derived exosomes on transforming growth factor-β1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) of lung epithelial cells in an inflammatory environment. Methods The morphology of exosomes derived from THP-1 macrophages was evaluated by transmission electron microscopy, and the biochemistry properties of exosomes were identified by accessing exosome-specific markers including tumor susceptibility gene 101 (TSG101), accessory protein ALG-2 interacting protein X (ALIX), CD81 and CD9 protein, and the calnexin, a negative control marker absent in exosomes, using an immunoblotting assay. The EMT of alveolar epithelial A549 cells was induced by TGF-β1, and the impacts of exosomes on the EMT of A549 cells was ascertained by comparing cells treated with exsomes derived from LPS-primed THP-1 macrophages and naive THP-1 cells. Results We successfully established an A549 cell EMT model by TGF-β1 induction and isolated exosomes derived from THP-1 macrophages. In comparison with the exosomes derived from untreated naive THP-1 macrophages, exosomes derived from LPS-primed THP-1 cells exhibited an ability to significantly promote TGF-β-induced EMT of A549 cells, as determined by a significantly down-regulated E-cadherin, and an dramatically increased expression of proteins in EMT-related signaling including vimentin, alpha smooth muscle actin (α-SMA), TGF-β1/Smad2/3 signaling proteins Smad2/3 protein and phosphorylated Smad2/3 (p-Smad2/3) and type 1 collagen (Col1). Conclusion Exosomes derived from LPS-stimulated macrophages are able to activate TGF-β/Smad2/3 signaling, which in turn increase the expression of EMT-related proteins vimentin, α-SMA and Col1 in A549 cells, and subsequently promote EMT in A549 cells.