English, W. R., Ireland-Zecchini, H., Baker, A. H., Littlewood, T. D., Bennett, M. R. and Murphy, G. (2018) Tissue Inhibitor of Metalloproteinase–3 (TIMP-3) induces FAS dependent apoptosis in human vascular smooth muscle cells. PLoS ONE, 13(4), e0195116. (doi: 10.1371/journal.pone.0195116) (PMID:29617412) (PMCID:PMC5884528)
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Abstract
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: | Articles |
|---|---|
| Keywords: | Research article, biology and life sciences, medicine and health sciences, research and analysis methods |
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Baker, Professor Andrew |
| Creator Roles: | Baker, A. H.Conceptualization, Funding acquisition, Methodology, Resources, Supervision, Writing – review and editing |
| Authors: | English, W. R., Ireland-Zecchini, H., Baker, A. H., Littlewood, T. D., Bennett, M. R., and Murphy, G. |
| College/School: | College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health |
| Journal Name: | PLoS ONE |
| Publisher: | Public Library of Science |
| ISSN: | 1932-6203 |
| ISSN (Online): | 1932-6203 |
| Published Online: | 04 April 2018 |
| Copyright Holders: | Copyright © 2018 The Authors |
| First Published: | First published in PLoS ONE 13(4):e0195116 |
| Publisher Policy: | Reproduced under a Creative Commons License |
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