Three MMPs especially characteristic of the EMT proteome are MMP-2, MMP-7, and MMP-9 (De Wever et al., 2008, Turley et al., 2008, Thiery, 2002). MMPs are secreted as inactive proenzymes that require activation or cleavage by other MMPs to become active (Egeblad and Werb, 2002). The lower molecular weight form is the activated form for each MMP shown (MMP-9:92/88kd;MMP-7:28/18kd;MMP-2:72/63kd). Equal numbers of Caco-2, MDA-MB-231, and IEC-6 cells (1 × 106/well) were stimulated with alcohol in serum free media (24h,1ml) and the levels of MMP-2 and MMP-9 (upper sections) and MMP-7 (lower sections) were determined in 50μl aliquots of conditioned media by western blotting as described (Forsyth et al., 2002). As seen in Figure 2A–C, alcohol robustly and significantly stimulated the production of all three MMPs over 24h by both transformed, cancer cell lines (colon cancer and breast cancer cells) and the non-transformed (“normal”) intestinal cell line IEC-6. Fold increases for Caco-2, MDA-MB-231, and IEC-6 (respectively) were: MMP-2: 3,6,2; MMP-7: 10,8,6; and MMP-9: 8,8, and 5 fold (p ≤ .05 for all MMPs vs. control, t-tests). We also assessed Caco-2 cell
