Ancy Leroy

Helicobacter pylori induces gastric epithelial cell invasion in a c-Met and type IV secretion system-dependent manner

Helicobacter pylori interacts with gastric epithelial cells, activating signaling pathways important for carcinogenesis. In this study we examined the role of H. pylori on cell invasion and the molecular mechanisms underlying this process. The relevance of H. pylori cag pathogenicity island-encoded type IV secretion system (T4SS), CagA, and VacA for cell invasion was also investigated. We found that H. pylori induces AGS cell invasion in collagen type I and in Matrigel invasion assays. H. pylori-induced cell invasion requires the direct contact between bacteria and cancer cells. H. pylori-mediated cell invasion was dependent on the activation of the c-Met receptor and on increased MMP-2 and MMP-9 activity. The abrogation of the c-Met receptor using the specific NK4 inhibitor or the silencing of c-Met expression with small interference RNA suppressed both cell invasion and MMP activity. Studies with different H. pylori strains revealed that cell invasion, c-Met tyrosine phosphorylation, and increased MMP-2 and MMP-9 activity were all dependent on the presence of a functional bacterial T4SS, but not on VacA cytotoxicity. Our findings demonstrate that H. pylori strains with a functional T4SS stimulate gastric epithelial cell invasion through a c-Met-dependent signaling pathway that comprises an increase in MMP-2 and MMP-9 activity.

Entamoeba histolytica disturbs the tight junction complex in human enteric T84 cell layers

Entamoeba (E.) histolytica trophozoites initiate amebiasis through invasion into the enteric mucosa. It was our aim to understand the molecular interactions between amebic trophozoites and en‐terocytes during the early steps of invasion. Tropho‐zoites of E. histolytica strain HM1:IMSS were seeded on the apical side of enteric T84 cell layers, which were established on filters in two‐compartment culture chambers. Cocultures were analyzed for para‐cellular permeability by measurement of transepithe‐lial electrical resistance (TER) and for the tight junction proteins ZO‐1, ZO‐2, occludin, and cingulin by immunocytochemistry and immunoprecipitation. On direct contact with the apical side of the enteric cells, trophozoites caused an increase in paracellular permeability as evidenced by a decrease of TER associated with an increase in [3H]mannitol flux. Immunoprecipitation of cocultures revealed dephos‐phorylation of ZO‐2, loss of ZO‐1 from ZO‐2, and degradation of ZO‐1 but less so of ZO‐2 and none of occludin or E‐cadherin. In conclusion, trophozoite‐associated increase in paracellular permeability of enteric cell layers is ascribed to disturbance of the molecular organization of tight junction proteins.—Leroy, A., Lauwaet, T., De Bruyne, G., Cornelissen, M., Mareel, M. Entamoeba histolytica disturbs the tight junction complex in human enteric T84 cell layers. FASEB J. 14, 1139–1146 (2000)

Molecular mechanisms of invasion by cancer cells, leukocytes and microorganisms

Invasion is a phenotype common to cancer cells, leukocytes, parasites, bacteria and viruses, involving cell-cell adhesion, cell-matrix adhesion, proteolysis and motility. These activities are regulated by the cross talk between invaders and host. We discuss the invasion-related molecular interactions of E-cadherin, integrins, matrix metalloproteinases and the chemokine receptor RANTES.

Listeria monocytogenes produces a pro-invasive factor that signals via ErbB2/ErbB3 heterodimers

Purpose We have previously demonstrated that conditioned medium from bacteria, some of which were isolated from the colon of cancer patients, stimulate cancer cell invasion in vitro through a 13-mer β-casein-derived peptide. Since invasion signalling pathways are coordinated by the balance between protein kinases and phosphatases, we investigated the effect of conditioned medium from bacteria on the overall cellular tyrosine phosphorylation.Methods The tyrosine phosphorylation level of HCT-8/E11 human colon cancer cells treated with the pro-invasive conditioned medium of Listeria, prepared on top of collagen type I gels (CMCollListeria/TSB), were analysed by means of immunoprecipitation and Western blot, with specific anti-phosphotyrosine antibodies.Results We demonstrated that CMCollListeria/TSB increases the tyrosine phosphorylation level of ErbB2 and ErbB3, members of the epidermal growth factor receptor (EGFR) family, and the association between ErbB3 and the phosphatidylinositol 3-kinase (PI3K) regulatory subunit (p85α). CMCollListeria/TSB-stimulated ErbB3 tyrosine phosphorylation and cancer cell invasion were independent from EGFR expression and activity but dependent on ErbB2 activity.Conclusions The interaction between Listeria and collagen type I produces, next to the 13-mer peptide, at least another pro-invasive factor that signals via ErbB2/ErbB3 heterodimers.

Do Entamoeba histolytica trophozoites signal via enteric microvilli

Entamoeba histolytica trophozoites initiate amebiasis through invasion into the enteric mucosa. We have examined the molecular interactions between trophozoites and enteric cells during the early steps of invasion (1). Upon direct contact with enteric cells, trophozoites cause disturbance of microvilli, as shown previously by Martinez-Palomo et al. (2). Analysis of molecules implicated in the formation of microvilli might help understand the mechanism by which trophozoites cause disturbance of microvilli. Because microvilli contain bundles of actin filaments, we selected to analyze the actin-binding proteins villin and ezrin. Villin is a 92.5-kDa protein that caps, severs, and bundles actin filaments in microvilli. Ezrin is an 81-kDa protein belonging to the family of ERM (ezrin, radixin, moesin) proteins. The inactive form of ezrin is abundantly present in the cell cytoplasm, whereas only a small fraction is transported to the apical membrane, where it directly crosslinks actin filaments and transmembrane proteins such as CD44, CD43, ICAM-1, ICAM-2, and indirectly NHE3 via EBP50 (3). Our hypothesis is that the transmembrane molecule to which ezrin binds could serve as a receptor for an amebic ligand. Such ligand–receptor binding could initiate signal transduction from microvilli into the cytoplasm of enteric cells.