The effect of fibroblast phenotype on A549 mesenchymal marker expression and proliferation. (A) Immunofluorescent staining of A549 monoculture, A549-NHLF, and A549-IPF co-cultures on epithelial side showing co-staining e-cadherin/vimentin/DAPI in mesenchymal marker expressing A549 cells at 48 h. (B) A549 cells expressing e-cadherin/vimentin/DAPI was significantly greater in mono-cultures and in IPF co-cultures compared to NHLF co-culture. Additionally, at the 48-h time point, mesenchymal marker expression was significantly greater in IPF co-cultures compared to A549 mono-cultures. However, by 72 h, there were significantly fewer epithelial cells expressing vimentin in IPF co-cultures compared to A549 mono-cultures (not shown on graph). (C) Conditioned medium at 72-h time point was collected and its effect on A549 proliferation was assessed using a DELFIA assay. There was no difference in the induction of proliferation. *P <0.05, **P <0.01, ***P <0.001 compared with A549-NHLF.