title: Loss of neutral ceramidase protects cells from nutrient- and energy -deprivation-induced cell death.
abstract: Sphingolipids are a family of lipids that regulate the cell cycle, differentiation and cell death. Sphingolipids are known to play a role in the induction of apoptosis, but a role for these lipids in necroptosis is largely unknown. Necroptosis is a programmed form of cell death that, unlike apoptosis, does not require ATP. Necroptosis can be induced under a variety of conditions, including nutrient deprivation and plays a major role in ischaemia/reperfusion injury to organs. Sphingolipids play a role in ischaemia/reperfusion injury in several organs. Thus, we hypothesized that sphingolipids mediate nutrient-deprivation-induced necroptosis. To address this, we utilized mouse embryonic fibroblast (MEFs) treated with 2-deoxyglucose (2DG) and antimycin A (AA) to inhibit glycolysis and mitochondrial electron transport. 2DG/AA treatment of MEFs induced necroptosis as it was receptor- interacting protein (RIP)-1/3 kinase-dependent and caspase-independent. Ceramides, sphingosine (Sph) and sphingosine 1-phosphate (S1P) were increased following 2DG/AA treatment. Cells lacking neutral ceramidase (nCDase(-/-)) were protected from 2DG/AA. Although nCDase(-/-) cells generated ceramides following 2DG/AA treatment, they did not generate Sph or S1P. This protection was stimulus-independent as nCDase(-/-) cells were also protected from endoplasmic reticulum (ER) stressors [tunicamycin (TN) or thapsigargin (TG)]. nCDase(-/-) MEFs had higher autophagic flux and mitophagy than wild-type (WT) MEFs and inhibition of autophagy sensitized them to necroptosis. These data indicate that loss of nCDase protects cells from nutrient- deprivation-induced necroptosis via autophagy, and clearance of damaged mitochondria. Results suggest that nCDase is a mediator of necroptosis and might be a novel therapeutic target for protection from ischaemic injury.
authors.affiliation: Department of Pharmacology & Toxicology, University of Louisville, Louisville, KY 40202, U.S.A.
authors.forename: Kumaran
authors.initials: K
authors.lastname: Sundaram
authors.affiliation: University of South Carolina Medical School, Columbia, SC 29209, U.S.A.
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Mather
authors.affiliation: Department of Pharmacology & Toxicology, University of Louisville, Louisville, KY 40202, U.S.A.
authors.forename: Subathra
authors.initials: S
authors.lastname: Marimuthu
authors.affiliation: Department of Pharmacology & Toxicology, University of Louisville, Louisville, KY 40202, U.S.A. James Graham Brown Cancer Center, University of Louisville, KY 40202, U.S.A.
authors.forename: Parag P
authors.initials: PP
authors.lastname: Shah
authors.affiliation: Department of Medicine, Stony Brook Cancer Center, Stony Brook University, NY 11794, U.S.A. ∥Northport Veterans Affairs Medical Center, Northport, NY 11768, U.S.A.
authors.forename: Ashley J
authors.initials: AJ
authors.lastname: Snider
authors.affiliation: Department of Medicine, Stony Brook Cancer Center, Stony Brook University, NY 11794, U.S.A. ∥Northport Veterans Affairs Medical Center, Northport, NY 11768, U.S.A.
authors.forename: Lina M
authors.initials: LM
authors.lastname: Obeid
authors.affiliation: Department of Medicine, Stony Brook Cancer Center, Stony Brook University, NY 11794, U.S.A.
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
authors.affiliation: Department of Pharmacology & Toxicology, University of Louisville, Louisville, KY 40202, U.S.A. James Graham Brown Cancer Center, University of Louisville, KY 40202, U.S.A. Department of Medicine, University of Louisville, KY 40202, U.S.A.
authors.forename: Levi J
authors.initials: LJ
authors.lastname: Beverly
authors.affiliation: Department of Pharmacology & Toxicology, University of Louisville, Louisville, KY 40202, U.S.A. James Graham Brown Cancer Center, University of Louisville, KY 40202, U.S.A. leah.siskind@louisville.edu.
authors.forename: Leah J
authors.initials: LJ
authors.lastname: Siskind
mesh_terms: D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003847:Deoxyglucose / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects*; D017353:Gene Deletion; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D008246:Lysophospholipids / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D055576:Neutral Ceramidase / Q000235:genetics / Q000378:metabolism*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D008246:Lysophospholipids; C032456:antimycin; C060506:sphingosine 1-phosphate; D000968:Antimycin A; D003847:Deoxyglucose; C526747:Asah2 protein, mouse; D055576:Neutral Ceramidase; D013110:Sphingosine
keywords: autophagy flux; endoplasmic reticulum (ER) stress; mitophagy; necroptosis; neutral ceramidase; sphingolipids