Zymography and Western blot analyses after co -culturing MDA- MB -231 with MDCK. (A) Zymography analysis of cell extracts from an MDA-MB-231 monoculture (MDA), a co-culture of MDA-MB-231 and MDCK (MDA + MDCK), and an MDCK monoculture (MDCK). The amount of MDA-MB-231 cells present was kept the same in the first two lanes (i.e. the MDA and MDA + MDCK lanes). The amount of MDCK cells was also kept the same in the last two lanes (i.e. the MDA + MDCK and MDCK lanes). Hence, the total amount of cells in the MDA + MDCK lane is the sum of the amount of cells in each monoculture (MDA and MDCK lanes). (B) Zymography analysis of conditioned media from the same cultures as in (A). (C) Western blot detecting MMP2 from cell extracts obtained in the same way as A. In (A-C), the MDA + MDCK samples show less gelatinase activity (A, B) and less MMP2 expression (C) compared to the additive amount of MDA alone and MDCK alone. (D) Western blot detecting GFP from the same blot as C, showing the similar amount of MDA-MB-231 cells in the first two lanes.