title: Anti-dengue virus nonstructural protein 1 antibodies cause NO-mediated endothelial cell apoptosis via ceramide-regulated glycogen synthase kinase-3β and NF-κB activation.
abstract: Immunopathogenetic mechanisms of dengue virus (DENV) infection are involved in hemorrhagic syndrome resulting from thrombocytopenia, coagulopathy, and vasculopathy. We have proposed a mechanism of molecular mimicry in which Abs against DENV nonstructural protein 1 (NS1) cross-react with human endothelial cells and cause NF-κB-regulated immune activation and NO-mediated apoptosis. However, the signaling pathway leading to NF-κB activation after the binding of anti-DENV NS1 Abs to endothelial cells is unresolved. In this study, we found that anti-DENV NS1 Abs caused the formation of lipid raftlike structures, and that disrupting lipid raft formation by methyl-β-cyclodextrin decreased NO production and apoptosis. Treatment with anti-DENV NS1 Abs elevated ceramide generation in lipid rafts. Pharmacological inhibition of acid sphingomyelinase (aSMase) decreased anti-DENV NS1 Ab-mediated ceramide and NO production, as well as apoptosis. Exogenous ceramide treatment induced biogenesis of inducible NO synthase (iNOS)/NO and apoptosis through an NF-κB-regulated manner. Furthermore, activation of glycogen synthase kinase-3β (GSK-3β) was required for ceramide-induced NF-κB activation and iNOS expression. Notably, anti-DENV NS1 Abs caused GSK-3β-mediated NF-κB activation and iNOS expression, which were regulated by aSMase. Moreover, pharmacological inhibition of GSK-3β reduced hepatic endothelial cell apoptosis in mice passively administered anti-DENV NS1 Abs. These results suggest that anti-DENV NS1 Abs bind to the endothelial cell membrane and cause NO production and apoptosis via a mechanism involving the aSMase/ceramide/GSK-3β/NF-κB/iNOS/NO signaling pathway.
authors.affiliation: Department of Microbiology and Immunology, National Cheng Kung University Medical College, Tainan 701, Taiwan.
authors.forename: Chia-Ling
authors.initials: CL
authors.lastname: Chen
authors.forename: Chiou-Feng
authors.initials: CF
authors.lastname: Lin
authors.forename: Shu-Wen
authors.initials: SW
authors.lastname: Wan
authors.forename: Li-Shiung
authors.initials: LS
authors.lastname: Wei
authors.forename: Mei-Chun
authors.initials: MC
authors.lastname: Chen
authors.forename: Trai-Ming
authors.initials: TM
authors.lastname: Yeh
authors.forename: Hsiao-Sheng
authors.initials: HS
authors.lastname: Liu
authors.forename: Robert
authors.initials: R
authors.lastname: Anderson
authors.forename: Yee-Shin
authors.initials: YS
authors.lastname: Lin
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000276:immunology / Q000502:physiology*; D017209:Apoptosis / Q000502:physiology*; D002460:Cell Line; D002518:Ceramides / Q000096:biosynthesis / Q000502:physiology*; D003429:Cross Reactions; D003716:Dengue Virus / Q000276:immunology*; D042783:Endothelial Cells / Q000276:immunology* / Q000473:pathology; D004789:Enzyme Activation / Q000276:immunology; D005786:Gene Expression Regulation / Q000276:immunology; D038362:Glycogen Synthase Kinase 3 / Q000502:physiology*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D008297:Male; D021962:Membrane Microdomains / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D016328:NF-kappa B / Q000502:physiology*; D009569:Nitric Oxide / Q000502:physiology*; D052247:Nitric Oxide Synthase Type II / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors / Q000502:physiology; D017361:Viral Nonstructural Proteins / Q000276:immunology*; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D000914:Antibodies, Viral; D002518:Ceramides; D007074:Immunoglobulin G; D016328:NF-kappa B; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D009569:Nitric Oxide; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3; C112980:acid sphingomyelinase-1; D013108:Sphingomyelin Phosphodiesterase