title: Inhibition of autophagy rescues palmitic acid-induced necroptosis of endothelial cells.
abstract: Accumulation of palmitic acid (PA) in cells from nonadipose tissues is known to induce lipotoxicity resulting in cellular dysfunction and death. The exact molecular pathways of PA-induced cell death are still mysterious. Here, we show that PA triggers autophagy, which did not counteract but in contrast promoted endothelial cell death. The PA-induced cell death was predominantly necrotic as indicated by annexin V and propidium iodide (PI) staining, absence of caspase activity, low levels of DNA hypoploidy, and an early ATP depletion. In addition PA induced a strong elevation of mRNA levels of ubiquitin carboxyl-terminal hydrolase (CYLD), a known mediator of necroptosis. Moreover, siRNA-mediated knockdown of CYLD significantly antagonized PA-induced necrosis of endothelial cells. In contrast, inhibition and knockdown of receptor interacting protein kinase 1 (RIPK1) had no effect on PA-induced necrosis, indicating the induction of a CYLD-dependent but RIPK1-independent cell death pathway. PA was recognized as a strong and early inducer of autophagy. The inhibition of autophagy by both pharmacological inhibitors and genetic knockdown of the autophagy-specific genes, vacuolar protein sorting 34 (VPS34), and autophagy-related protein 7 (ATG7), could rescue the PA-induced death of endothelial cells. Moreover, the initiation of autophagy and cell death by PA was reduced in endothelial cells loaded with the Ca(2+) chelator 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-(acetoxymethyl) ester (BAPTA-AM), indicating that Ca(2+) triggers the fatal signaling of PA. In summary, we introduce an unexpected mechanism of lipotoxicity in endothelial cells and provide several novel strategies to counteract the lipotoxic signaling of PA.
authors.affiliation: Institute of Molecular Biology and Biochemistry, Center of Molecular Medicine, Medical University of Graz, 8010 Graz, Austria.
authors.forename: Muhammad Jadoon
authors.initials: MJ
authors.lastname: Khan
authors.forename: Muhammad
authors.initials: M
authors.lastname: Rizwan Alam
authors.forename: Markus
authors.initials: M
authors.lastname: Waldeck-Weiermair
authors.forename: Felix
authors.initials: F
authors.lastname: Karsten
authors.forename: Lukas
authors.initials: L
authors.lastname: Groschner
authors.forename: Monika
authors.initials: M
authors.lastname: Riederer
authors.forename: Seth
authors.initials: S
authors.lastname: Hallström
authors.forename: Patrick
authors.initials: P
authors.lastname: Rockenfeller
authors.forename: Viktoria
authors.initials: V
authors.lastname: Konya
authors.forename: Akos
authors.initials: A
authors.lastname: Heinemann
authors.forename: Frank
authors.initials: F
authors.lastname: Madeo
authors.forename: Wolfgang F
authors.initials: WF
authors.lastname: Graier
authors.forename: Roland
authors.initials: R
authors.lastname: Malli
mesh_terms: D001343:Autophagy / Q000187:drug effects*; D000071193:Autophagy-Related Protein 7; D020013:Calcium Signaling; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D058546:Class III Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D000074749:Deubiquitinating Enzyme CYLD; D004533:Egtazic Acid / Q000031:analogs & derivatives; D042783:Endothelial Cells / Q000378:metabolism* / Q000473:pathology; D004791:Enzyme Inhibitors / Q000493:pharmacokinetics*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D055785:Gene Knockdown Techniques; D006801:Humans; D009336:Necrosis; D019308:Palmitic Acid / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins; D044764:Ubiquitin-Activating Enzymes / Q000235:genetics / Q000378:metabolism
chemical_list: D002614:Chelating Agents; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D025521:Tumor Suppressor Proteins; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D019308:Palmitic Acid; D004533:Egtazic Acid; D058546:Class III Phosphatidylinositol 3-Kinases; C506249:RIPK1 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C409984:CYLD protein, human; D000074749:Deubiquitinating Enzyme CYLD; C509349:ATG7 protein, human; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes