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. Author manuscript; available in PMC: 2014 Apr 1.
Published in final edited form as: Exp Cell Res. 2012 Dec 10;319(6):850–859. doi: 10.1016/j.yexcr.2012.12.002

Figure 3. Stimulation of esophageal epithelial cells with fibroblast conditioned media (FCM) leads to features of EMT, in an IL-1β and TNFα-dependent fashion.

Figure 3

A: Schematic of experimental design: Following stimulation of FEF3 cells with CEM (2 days), this “fibroblast conditioned media” (FCM) was harvested and transferred to fresh EPC2-hTERT cells. After 3 weeks of FCM stimulation in the presence of absence of competitive inhibitors of IL-1β and/or TNFα, EPC2-hTERT cells were harvested for mRNA isolation, or immunolocalization of mesenchymal/epithelial markers. B, C, D: mRNA expression of E-cadherin, vimentin, and αSMA by EPC2-hTERT cells after stimulation with FCM in the presence or absence of anti-TNFα mAb (Remicade) and/or anti-IL-1R (Anakinra). E, J: Constitutive expression of epithelial E-cadherin (red) and αSMA (green) by EPC2-hTERT cells. Nuclei are counterstained with DAPI (blue). F, K: Loss of E-cadherin expression (red) and enhanced αSMA (green) expression by EPC2-hTERT cells following 3 weeks of stimulation with FCM. G, H: Partial recovery of E-cadherin expression by EPC2-hTERT cells stimulated with FCM and anti-TNFα mAb (Remicade) or anti-IL-1R (Anakinra). L, M: Absence of αSMA in EPC2-hTERT cells treated with FCM in the presence anti-TNFα mAb (Remicade) or anti-IL-1R (Anakinra). I, N: Combination of anti-TNFα mAb (Remicade) and anti-IL-1R (Anakinra) protects EPC2-hTERT cells from effects of FCM stimulation. O, P: Morphology of EPC2-hTERT cells before and after FCM stimulation. Q: Effect of anti-TNFα mAb and anti-IL-1R upon EPC2-hTERT morphology. p-values were calculated based upon comparisons to unstimulated conditions. *p<0.05, **p<0.01, ***p<0.001, NS = not significant.