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. Author manuscript; available in PMC: 2011 Dec 12.
Published in final edited form as: Sci Transl Med. 2010 Nov 10;2(57):57ra82. doi: 10.1126/scitranslmed.3001510

Figure 3. CpG-induced EMT in Human A549 Cells.

Figure 3

A. Representative photomicrographs (n=5) of A549 cells cultured in media (DMEM + 10% FCS) (a), TGFβ (b), and increasing CpG concentrations of 5 μg/mL (c), 10 μg/mL (d), 50 μg/mL (e), 100 μg/mL (f), and 200 μg/mL (g) for 96 h. B. qRT-PCR analysis of αSMA (a), vimentin (b), and e-cadherin (c) in A549 cells cultured with increasing concentrations of CpG for 96 hours. C. qRT-PCR analysis of IFNα in A549 cells cultured with increasing concentrations of CpG for 96 h. D. Fluorescent ICC for collagen 1 in A549 cells (40× magnification) that were cultured for 96 h in media (a), 10 μg/mL CpG (b), 50 μg/mL CpG (c), and 100 μg/mL CpG (d). Isotype control for collagen 1 antibody using cells cultured with 100 μg/mL CpG (e). E. siRNA knockdown of TLR9 in A549 cells in a CpG EMT assay: Western Blot analysis of TLR9 protein and β-actin loading control in A549 cell lysates after siRNA treatment with a non targeting control siRNAs, cyclophilin B control siRNAs (a), and TLR9 siRNAs; photomicrographs of A549 cells before CpG-DNA treatment cultured in media and transfection agent alone (b), with non target siRNA (c), and with TLR9 siRNA (d); representative photomicrographs (n=4) of A549 cells after siRNA treatment and stimulated with media and transfection agent alone (e), non target siRNA + 75 μg/ml CpG (f), and TLR9 siRNA + 75 μg/ml CpG-DNA for 72 hrs (g); qRT-PCR analysis of vimentin (h) and e-cadherin (i) in siRNA-treated A549 cells and cultured with 75 μg/ml CpG for 72 hours. Data are mean ± SD. *** p < 0.0001.