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Mouse Lung Fibroblast Resistance to Fas-Mediated Apoptosis Is Dependent on the Baculoviral Inhibitor of Apoptosis Protein 4 and the Cellular FLICE-Inhibitory Protein.

ABSTRACT: A characteristic feature of idiopathic pulmonary fibrosis (IPF) is accumulation of apoptotic resistant fibroblasts/myofibroblasts in the fibroblastic foci. As caveolin (Cav)-null mice develop pulmonary fibrosis (PF), we hypothesized that the participating fibroblasts display an apoptosis-resistant phenotype. To test this hypothesis and identify the molecular mechanisms involved we isolated lung fibroblasts from Cav-null mice and examined the expression of several inhibitors of apoptosis (IAPs), of c-FLIP, of Bcl-2 proteins and of the death receptor CD95/Fas. We found significant increase in XIAP and c-FLIP constitutive protein expression with no alteration of Bcl-2 and lower levels of CD95/Fas. The isolated fibroblasts were then treated with the CD95/Fas ligand (FasL) to induce apoptosis. While the morphological and biochemical alterations induced by FasL were similar in wild-type (wt) and Cav-null mouse lung fibroblasts, the time course and the extent of the alterations were greater in the Cav-null fibroblasts. Several salient features of Cav-null fibroblasts response such as loss of membrane potential, fragmentation of the mitochondrial continuum concurrent with caspase-8 activation, and subsequent Bid cleavage, prior to caspase-3 activation were detected. Furthermore, M30 antigen formation, phosphatidylserine expression and DNA fragmentation were caspase-3 dependent. SiRNA-mediated silencing of XIAP and c-FLIP, individually or combined, enhanced the sensitivity of lung fibroblasts to FasL-induced apoptosis. Pharmacological inhibition of Bcl-2 had no effect. Together our findings support a mechanism in which CD95/Fas engagement activates caspase-8, inducing mitochondrial apoptosis through Bid cleavage. XIAP and c-FLIP fine tune this process in a cell-type specific manner.

SUBMITTER: Predescu SA

PROVIDER: S-EPMC5348516 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Mouse Lung Fibroblast Resistance to Fas-Mediated Apoptosis Is Dependent on the Baculoviral Inhibitor of Apoptosis Protein 4 and the Cellular FLICE-Inhibitory Protein.

Predescu Sanda A SA Zhang Jian J Bardita Cristina C Patel Monal M Godbole Varun V Predescu Dan N DN

Frontiers in physiology 20170314

A characteristic feature of idiopathic pulmonary fibrosis (IPF) is accumulation of apoptotic resistant fibroblasts/myofibroblasts in the fibroblastic foci. As caveolin (Cav)-null mice develop pulmonary fibrosis (PF), we hypothesized that the participating fibroblasts display an apoptosis-resistant phenotype. To test this hypothesis and identify the molecular mechanisms involved we isolated lung fibroblasts from Cav-null mice and examined the expression of several inhibitors of apoptosis (IAPs), ...[more]

PMID: 28352235

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Project description:BackgroundCytoplasmic viral double-stranded RNA (dsRNA) is detected by a class of ubiquitous cytoplasmic RNA helicases, retinoic acid inducible gene-I (RIG-I) and melanoma differentiation antigen-5 (MDA5), which initiate a signaling cascade via their common adaptor called interferon-? (IFN-?) promoter stimulator-1 (IPS-1). This leads to the production of proinflammatory and antiviral cytokines, the type I Interferons, via mainly nuclear factor kappa B (NF-?B) and interferon response factor-3 (IRF3) transcription factors. Fas-associated death domain (FADD) protein, receptor-interacting protein (RIP1), caspase-8 and tumor necrosis factor receptor (TNFR)-associated death domain (TRADD) protein, all traditionally associated with death receptor signaling, are also involved in RIG-I/MDA5 signaling pathway. We previously showed that FLIP (Flice-like inhibitory protein), also designated as cflar (CASP8 and FADD-like apoptosis regulator), negatively regulates lipopolysaccharide (LPS)-induced toll-like receptor 4 (TLR4) signaling in endothelial cells and mouse embryonic fibroblasts (MEFs) and protected against TLR4-mediated apoptosis.ResultsIn this study, we investigated the role of FLIP in cellular response to cytoplasmic polyinosinic:polycytidylic acid, poly(I:C), a synthetic analog of dsRNA. Consistent with the previously described role of FADD in RIG-I/MDA5-mediated apoptosis, we found that FLIP-/- MEFs were more susceptible to killing by cytoplasmic poly(I:C). However, FLIP-/- MEFs also exhibited markedly increased expression of NF-?B-and IRF3- dependent genes in response to cytoplasmic poly(I:C). Importantly, reconstitution of FLIP in FLIP-/-MEFs reversed the hyper-activation of IRF3- and NF-?B-mediated gene expression. Further, we found that caspase-8 catalytic activity was not required for cytoplasmic poly(I:C)-mediated NF-?B and IRF3 signaling.ConclusionsThese results provide evidence for a crucial dual role for FLIP in antiviral responses to cytoplasmic dsRNA: it protects from cytoplasmic dsRNA-mediated cell death while down-regulating IRF3-and NF-?B-mediated gene expression. Since the pathogenesis of several viral infections involves a heightened and dysregulated cytokine response, a possible therapy could involve modulating FLIP levels.

Dataset Information

Mouse Lung Fibroblast Resistance to Fas-Mediated Apoptosis Is Dependent on the Baculoviral Inhibitor of Apoptosis Protein 4 and the Cellular FLICE-Inhibitory Protein.

Publications

Mouse Lung Fibroblast Resistance to Fas-Mediated Apoptosis Is Dependent on the Baculoviral Inhibitor of Apoptosis Protein 4 and the Cellular FLICE-Inhibitory Protein.

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