Figure 1.

Chronic cytokine exposure leads to EMT memory in NSCLC. (A) A549 cells underwent EMT upon 48-hour IL-1β (1 ng/ml) exposure. (B) Experimental design for the chronic IL-1β treatment: A549 cells were exposed to IL-1β for 21 days and subsequently cultured for 30 days after IL-1β withdrawal. Cells were split every three to four days with fresh IL-1β in the medium. (C) Morphological changes of the untreated and treated A549 cells with IL-1β by bright-field microscopy at different time points. (D) EMT markers in A549 cells were determined by immunoblotting following the time course. Quantification of each marker was normalized to the untreated samples. (E) EMT was reversed within 6 days upon IL-1β withdrawal following 3 days or 6 days of IL-1β treatment in A549 cells. (F) A549 cells chronically treated with TNF-α (10 ng/ml) or TGF-β (5 ng/ml) for 21 days were examined for EMT and EMT memory. “-d”, days after IL-1β withdrawal; “p”, previously treated with IL-1β. “n.d.”, non-detectable. “Tα”, TNF-α; “Tβ”, TGF-β; “pTα”, previously treated with TNF-α; “pTβ”, previously treated with TGF-β. See also Fig. S1.