TY - JOUR T1 - Immunology/Infection/Inflammation free papers 091–104 JF - Gut JO - Gut SP - a25 LP - A29 DO - 10.1136/gut.50.suppl_2.a25 VL - 50 IS - suppl 2 A2 - , Y1 - 2002/04/01 UR - http://gut.bmj.com/content/50/suppl_2/a25.abstract N2 - J.R. Bebb1, D.P. Letley1, R.J. Thomas1, H.M. Collins2, S.A. Watson2, J.C. Atherton1.Division of Gastroenterology and Institute of Infections and Immunity, Academic Unit of Cancer Studies, University Hospital, Nottingham, UKBackground: MMP-7 (matrilysin) is a member of the metalloproteinase family of enzymes, which are important in normal and pathological remodelling of epithelial-matrix interactions. Several studies have shown increased expression of MMP-7 in gastric cancer tissue compared to non-cancer tissue. More virulent strains of H.pylori possess the cagPAI (encoding a Type IV secretion system) and an active form of VacA, a toxin that induces vacuolation in vitro. This study examines the effect of H.pylori on MMP-7 expression in HT29 cells.Methods:H.pylori strains 60190 (cagPAI+, vacA s1/m1), Tx30a (cagPAI-, vacA s2/m2) and the VacA and CagE isogenic mutants of 60190 were co-cultured with HT29 cells for 24 hours. Cell pellets were used for RNA extraction and reverse transcription, and cDNA levels assessed for MMP-7 levels (and GAPDH) by Real Time PCR. HT29 supernatants were assessed for MMP-7 expression by western blot and casein zymography and for other metalloproteinase activity by gelatin zymography. Experiments were performed at least three times.Results:H.pylori pathogenic strain 60190 increased MMP-7 RNA levels (13-fold vs untreated, p=0.06, unpaired t-test) more than non-pathogenic strain Tx30a. Use of isogenic mutants showed this effect to be VacA-independent but CagE-dependent. Western blot gave a 29kDa band for 60190 and its VacA mutant. Casein zymography revealed a 29kDa band of activity for all samples but reduced intensity for strain Tx30a and the 60190 cagE mutant. This band corresponds to the predicted size for pro-MMP-7. Gelatin zymography showed no differences between treatments.Conclusion: In HT29 cells H.pylori co-culture leads to upregulation of MMP-7 at both RNA … ER -