title: Tumor necrosis factor stimulates matrix metalloproteinase 9 secretion from cultured human chorionic trophoblast cells through TNF receptor 1 signaling to IKBKB-NFKB and MAPK1/3 pathway.
abstract: The identification of proinflammatory signal transduction pathways may suggest new therapeutic targets. In this study, we examine which signaling pathways are involved in tumor necrosis factor (TNF)-induced matrix metalloproteinase 9 (MMP9) secretion in human chorionic trophoblast (CT) cells. Purified CT cells were cultured in the presence of antibodies or chemical inhibitors that specifically block/inhibit distinct TNF receptors and kinase pathways. TNF-induced proMMP9 production, as measured by zymography, was significantly blocked/inhibited by TNF receptor 1 (TNFRSF1A) antibody, NFKB activation inhibitor (NFKBAI), and MAPK1/3 (ERK) inhibitor (U0126) (P < 0.01), but not by TNF receptor 2 (TNFRSF1B) antibody, MAPK14 (p38 MAPK) inhibitor (SB203580), and MAPK8/9/10 (JNK) inhibitor (SP600125). By Western blot analysis, we found that TNF rapidly and significantly increased phosphorylation of IKBKB, MAPK1/3, and MAPK8/9/10 and that the phosphorylation of these kinases by TNF was reduced significantly by TNFRSF1A neutralizing antibody, but not by TNFRSF1B neutralizing antibody. Moreover, we found that TNF increased TNF receptor-associated factor (TRAF) 1 and decreased TRAF2 protein expression through TNFRSF1A, but not TNFRSF1B. The CT cells that had increased TRAF1 and decreased TRAF2 after an initial TNF treatment demonstrated a dramatic deficiency in phosphorylation of the above protein kinases following a secondary TNF treatment. Localization of RELA subunit by immunocytochemistry was shifted to the nuclei after TNF treatment compared to cytosol in untreated controls. We also found cross-talk between the phosphoinositide 3-kinase pathway and ERK pathway. In summary, we have demonstrated that TNF stimulates proMMP9 production in CT cells through TNFRSF1A-TRAFs-IKBKB-NFKB and ERK signaling pathways, but not through TNFRSF1B and JNK/p38-AP-1 pathways.
authors.affiliation: CIHR Group in Fetal Development and Health, Department of Physiology, University of Toronto, Toronto, Ontario, Canada. weisun.li@utoronto.ca
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Han
authors.initials: H
authors.lastname: Li
authors.forename: Alan D
authors.initials: AD
authors.lastname: Bocking
authors.forename: John R G
authors.initials: JR
authors.lastname: Challis
mesh_terms: D000704:Analysis of Variance; D057134:Antibodies, Neutralizing; D015153:Blotting, Western; D002478:Cells, Cultured; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism*; D007150:Immunohistochemistry; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects*; D014327:Trophoblasts / Q000187:drug effects / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D057134:Antibodies, Neutralizing; D016328:NF-kappa B; D047888:Receptors, Tumor Necrosis Factor, Type I; D014409:Tumor Necrosis Factor-alpha; D051550:I-kappa B Kinase; D020928:Mitogen-Activated Protein Kinases; D020780:Matrix Metalloproteinase 9