409 The Colonic Adherent-Invasive Escherichia coli Strain HM605 Induces Anti-Apoptotic Responses in Intestinal Epithelial Cells, Reduces Barrier Integrity and Worsens Experimental Colitis

G A A b st ra ct s (MMPs) are zinc-dependent neutral endopeptidases that participate in degradation of extracellular matrix proteins involved in a variety of pathological processes including inflammation and cancer. MMP9 protein expression and activity is undetectable in most healthy adult tissues including the intestine but is highly expressed in inflammatory state. We have shown that despite being a mediator of acute inflammation MMP9 plays a protective role in the development of CAC. Aim of the present study is to determine the necessity and sufficiency of epithelial derived MMP9 mediated p53 activation via Notch1 signaling for its tumor suppressive function in CAC, by using MMP9 transgenic mice (Tg-villin-MMP9) that specifically overexpresses MMP9 in the colonic epithelium. Methods: Age and gender matched Tg-villin-MMP9 and their wild type littermates (WT) mice were used for in vivo experiments and embryonic fibroblasts (MEFs) isolated from WT and MMP9-/mice were used for in vitro experiments. Mice were exposed to dextran sulfate sodium (DSS, 3% in drinking water) and were given 3 DSS cycles of one week each and with two weeks apart and were sacrificed at day 75. Colons were processed for inflammation and polyps. Swiss rolls of colonic tissues were histologically examined. Western blot (WB) analysis was done with the colonic mucosal stripping. Results: Tg-villin-MMP9 mice were more resistant to CAC as evidenced by decreased tumor burden. In addition, decreased erosion of mucosal layer, infiltration of neutrophils and dysplasia were seen in the polyps of Tg-villin-MMP9 mice compared to WTs both inducedwith CAC.WB analysis indicated significantly increased protein expression of NICD (active Notch1), p53, p21WAF1/Cip1, Bax-1 and caspase-3 compared to WTs mice both induced with CAC. TUNEL staining showed increased apoptosis in the colons of Tg-villin-MMP9 mice compared to WTs mice both induced with CAC. MMP9 overexpression (by stable transfection) in MMP9-/MEFs resulted in significantly increased protein expression of NICD, p53 and p21WAF1/Cip1 expressions compared to non-transfected MMP9-/MEFs, and were comparable to WT MEFs. Conclusion: Together, the data show that constitutive expression of MMP9 in colonic epithelium modulates apoptosis and cell cycle arrest, and thereby acts as a tumor suppressor in CAC. This study elucidates the novel tumor suppressive role of MMP9 in malignant transformation of the colonic epithelium in context of chronic inflammation which may provide new diagnostic and therapeutic approaches.