Transwell co-culture of RAW 264.7 macrophages induces metastatic gene expression by A549 cells.

<p><b>A</b> A transwell co-culture system of RAW 264.7 macrophages and A549 cells was used in this study. RAW 264.7 cells and A549 cells were separately cultured for 24 h and then co-cultured in serum free medium for further 12 h or 24 h as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054073#s2" target="_blank">Materials and methods</a>. <b>B</b> Control A549 and A549-TPα cells became more scattered and spindle shaped after culture with RAW 264.7 cells. Both types of A549 cells were pictured after 12 h of culture with or without RAW 264.7 cells as described in A. <b>C</b> Co-culture of RAW 264.7 macrophages induced metastatic gene expression by A549 cells. After 12 h of co-culture with macrophages, the expression of several metastatic genes in control A549 and A549-TPα cells, including MMPs, VEGF and MCP-1 mRNA, were examined by RT-PCR as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054073#s2" target="_blank">Materials and methods</a>. <b>D</b> Co-culture of RAW 264.7 macrophages decreased E-cadherin expression by A549 cells. Control A549 and A549-TPα cells were collected after 24 h of culture with or without RAW 264.7 cells as described in A. Protein level of E-cadherin was determined by Western blot as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054073#s2" target="_blank">Material and methods</a>. All data are representative of at least three independent experiments.