title: Expression of cellular FLICE-inhibitory protein in human coronary arteries and in a rat vascular injury model.
abstract: We previously isolated MACH-related inducer of toxicity (MRIT), a homolog of caspase 8. MRIT, also known as c-FLICE-inhibitory protein (c-FLIP), is an enzymatically inactive homolog of caspase 8 with homology to viral FLIP (v-FLIP). Because of this homology and resemblance to dominant negative proteins, c-FLIP is widely believed to be an antagonist to the death receptor-initiated apoptotic pathways that use caspase 8. We generated a polyclonal antibody, MAG1, and show that this antibody specifically recognizes two splice forms, long form (c-FLIPL) and short form (c-FLIPS). By in situ hybridization and immunohistochemistry, we demonstrate that c-FLIP is expressed in endothelial cells, macrophages, and smooth muscle cells (SMCs) both in human coronary arteries and in cultured cells. In an uninjured rat carotid arteries, c-FLIP protein is abundant in the vascular media. After balloon angioplasty, c-FLIP protein is rapidly down-regulated in medial SMCs for 2 weeks and regains expression by 4 weeks. In contrast, the neointima is strongly immunoreactive to c-FLIP from day 7 after the initial injury and remains strongly immunoreactive until 4 to 6 weeks. Similarly there is strong c-FLIP immunoreactivity in SMCs from nonatherosclerotic diffuse intimal thickening and in the overlying endothelial cells. In contrast, c-FLIP immunoreactivity is uneven and often absent in SMCs within the atherosclerotic plaque. Double labeling with c-FLIP antibody and terminal deoxynucleotidyltransferase-mediated UDP end labeling (TUNEL) in the injured rat common carotid artery show that TUNEL-positive cells in the first 2 days after injury lack detectable c-FLIP, suggested a role for caspase 8 in this form of death. In contrast, there is no correlation of c-FLIP with the spontaneous elevation in death of intima seen at 7 days after injury. For human atherosclerotic plaques, the majority of TUNEL-positive cells lack detectable c-FLIP. The expression pattern of c-FLIP and the relation between c-FLIP and TUNEL suggest a role for c-FLIP- and caspase 8-driven death in control of viability of the cells of the atherosclerotic intima.
authors.affiliation: Departments of Pathology, Medicine (Cardiology), and Molecular Biotechnology, University of Washington, Seattle, Washington 98195, USA.
authors.forename: T
authors.initials: T
authors.lastname: Imanishi
authors.forename: J
authors.initials: J
authors.lastname: McBride
authors.forename: Q
authors.initials: Q
authors.lastname: Ho
authors.forename: K D
authors.initials: KD
authors.lastname: O'Brien
authors.forename: S M
authors.initials: SM
authors.lastname: Schwartz
authors.forename: D K
authors.initials: DK
authors.lastname: Han
mesh_terms: D000328:Adult; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D020212:Carotid Artery Injuries / Q000378:metabolism* / Q000473:pathology; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D003324:Coronary Artery Disease / Q000378:metabolism / Q000473:pathology; D003331:Coronary Vessels / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology; D005260:Female; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008264:Macrophages / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009131:Muscle, Smooth, Vascular / Q000378:metabolism / Q000473:pathology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D002352:Carrier Proteins; D047908:Intracellular Signaling Peptides and Proteins