TIMP-3 INDUCES CELL-DEATH BY STABILIZING TNF-ALPHA RECEPTORS ON THE SURFACE OF HUMAN COLON-CARCINOMA CELLS

Matrix metalloproteinases (MMPs) and tissue inhibitor of metalloprotei nases (TIMPs) regulate the structural integrity of the extracellular m atrix (ECM). Constitutive expression of human TIMP3 in human DLD colon carcinoma cells renewed serum-responses and inhibited tumour formatio n in nude mice, To elucidate the mechanism of TIMP-3-mediated tumour s uppression, we compared parental DLD and TIMP-3 expressing DLD cells ( TIMP-3/DLD), finding them to be significantly different, TIMP-3/DLD cu ltures have fewer mitotic cells, are delayed in G(1), and die after se rum starvation, TIMP-3/DLD conditioned media activates cell death on f ibroblast cells, The cell death induced by serum starvation and condit ioned media was inhibited by 70%, in the presence of neutralizing tumo ur necrosis factor alpha (TNF-alpha) antibody, TIMP-3/DLD whole cell l ysate contained p55 TNF-alpha receptor, while vector/DLD lysate had p5 5 TNF-alpha receptor and p46 soluble TNF-alpha inhibitor, Vector/DLD c onditioned media had p46, while no soluble TNF-alpha receptor,vas dete cted in TIMP-3/DLD conditioned media, In addition, FAGS analysis revea led that TIMP-3/DLD cells have more TNF-alpha surface binding sites, s uggesting a direct correlation between TIMP-3 expression and surface r eceptors, The mechanism of tumorigenic reversion induced by TIMP-3 in DLD cells may involve protection of receptors from the proteolytic act ivity of MMPs. Putative TIMP-3-mediated inhibition of MMPs restores th e TNF-alpha p55 signalling pathway and the carcinoma cell is killed by autocrine TNF-alpha, Thus, DLD cells have specific ECM MMPs that clea ve cytokines and cytokine receptors, TIMP-3 specifically inhibits MMPs involved in receptor shedding. (C) 1997 Academic Press Limited.