Tissue Inhibitor of Metalloproteinase–3 (TIMP-3) induces FAS dependent apoptosis in human vascular smooth muscle cells

English, William R. ORCID: https://orcid.org/0000-0003-3024-2441, Ireland-Zecchini, Heather, Baker, Andrew H., Littlewood, Trevor D., Bennett, Martin R. and Murphy, Gillian (2018) Tissue Inhibitor of Metalloproteinase–3 (TIMP-3) induces FAS dependent apoptosis in human vascular smooth muscle cells. PLoS One, 13 (4). ISSN 1932-6203

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Over expression of Tissue Inhibitor of Metalloproteinases-3 (TIMP-3) in vascular smooth muscle cells (VSMCs) induces apoptosis and reduces neointima formation occurring after saphenous vein interposition grafting or coronary stenting. In studies to address the mechanism of TIMP-3-driven apoptosis in human VSMCs we find that TIMP-3 increased activation of caspase-8 and apoptosis was inhibited by expression of Cytokine response modifier A (CrmA) and dominant negative FAS-Associated protein with Death Domain (FADD). TIMP-3 induced apoptosis did not cause mitochondrial depolarisation, increase activation of caspase-9 and was not inhibited by over-expression of B-cell Lymphoma 2 (Bcl2), indicating a mitochondrial independent/type-I death receptor pathway. TIMP-3 increased levels of the First Apoptosis Signal receptor (FAS) and depletion of FAS with shRNA showed TIMP-3-induced apoptosis was FAS dependent. TIMP-3 induced formation of the Death-Inducing Signalling Complex (DISC), as detected by immunoprecipitation and by immunofluorescence. Cellular-FADD-like IL-1 converting enzyme-Like Inhibitory Protein (c-FLIP) localised with FAS at the cell periphery in the absence of TIMP-3 and this localisation was lost on TIMP-3 expression with c-FLIP adopting a perinuclear localisation. Although TIMP-3 inhibited FAS shedding, this did not increase total surface levels of FAS but instead increased FAS levels within localised regions at the cell surface. A Disintegrin And Metalloproteinase 17 (ADAM17) is inhibited by TIMP-3 and depletion of ADAM17 with shRNA significantly decreased FAS shedding. However ADAM17 depletion did not induce apoptosis or replicate the effects of TIMP-3 by increasing localised clustering of cell surface FAS. ADAM17-depleted cells could activate caspase-3 when expressing levels of TIMP-3 that were otherwise sub-apoptotic, suggesting a partial role for ADAM17 mediated ectodomain shedding in TIMP-3 mediated apoptosis. We conclude that TIMP-3 induced apoptosis in VSMCs is highly dependent on FAS and is associated with changes in FAS and c-FLIP localisation, but is not solely dependent on shedding of the FAS ectodomain.

Item Type: Article
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
Faculty of Science > School of Biological Sciences
UEA Research Groups: Faculty of Medicine and Health Sciences > Research Centres > Metabolic Health
Depositing User: LivePure Connector
Date Deposited: 15 Dec 2022 17:31
Last Modified: 19 Oct 2023 03:30
URI: https://ueaeprints.uea.ac.uk/id/eprint/90237
DOI: 10.1371/journal.pone.0195116

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