title: Nitrosative stress inhibits the aminophospholipid translocase resulting in phosphatidylserine externalization and macrophage engulfment: implications for the resolution of inflammation.
abstract: Macrophage recognition of apoptotic cells depends on externalization of phosphatidylserine (PS), which is normally maintained within the cytosolic leaflet of the plasma membrane by aminophospholipid translocase (APLT). APLT is sensitive to redox modifications of its -SH groups. Because activated macrophages produce reactive oxygen and nitrogen species, we hypothesized that macrophages can directly participate in apoptotic cell clearance by S-nitrosylation/oxidation and inhibition of APLT causing PS externalization. Here we report that exposure of target HL-60 cells to nitrosative stress inhibited APLT, induced PS externalization, and enhanced recognition and elimination of "nitrosatively" modified cells by RAW 264.7 macrophages. Using S-nitroso-L-cysteine-ethyl ester (SNCEE) and S-nitrosoglutathione (GSNO) that cause intracellular and extracellular trans-nitrosylation of proteins, respectively, we found that SNCEE (but not GSNO) caused significant S-nitrosylation/oxidation of thiols in HL-60 cells. SNCEE also strongly inhibited APLT, activated scramblase, and caused PS externalization. However, SNCEE did not induce caspase activation or nuclear condensation/fragmentation suggesting that PS externalization was dissociated from the common apoptotic pathway. Dithiothreitol reversed SNCEE-induced S-nitrosylation, APLT inhibition, and PS externalization. SNCEE but not GSNO stimulated phagocytosis of HL-60 cells. Moreover, phagocytosis of target cells by lipopolysaccharide-stimulated macrophages was significantly suppressed by an NO. scavenger, DAF-2. Thus, macrophage-induced nitrosylation/oxidation plays an important role in cell clearance, and hence in the resolution of inflammation.
authors.affiliation: Center for Free Radical and Antioxidant Health, Department of Environmental and Occupational Health, and Cancer Institute, University of Pittsburgh, Pittsburgh, Pennsylvania 15219, USA.
authors.forename: Yulia Y
authors.initials: YY
authors.lastname: Tyurina
authors.forename: Liana V
authors.initials: LV
authors.lastname: Basova
authors.forename: Nagarjun V
authors.initials: NV
authors.lastname: Konduru
authors.forename: Vladimir A
authors.initials: VA
authors.lastname: Tyurin
authors.forename: Ala I
authors.initials: AI
authors.lastname: Potapovich
authors.forename: Peter
authors.initials: P
authors.lastname: Cai
authors.forename: Hülya
authors.initials: H
authors.lastname: Bayir
authors.forename: Detcho
authors.initials: D
authors.lastname: Stoyanovsky
authors.forename: Bruce R
authors.initials: BR
authors.lastname: Pitt
authors.forename: Anna A
authors.initials: AA
authors.lastname: Shvedova
authors.forename: Bengt
authors.initials: B
authors.lastname: Fadeel
authors.forename: Valerian E
authors.initials: VE
authors.lastname: Kagan
mesh_terms: D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D009584:Nitrogen / Q000737:chemistry*; D010084:Oxidation-Reduction; D010718:Phosphatidylserines / Q000737:chemistry*; D049294:Phospholipid Transfer Proteins / Q000378:metabolism*; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D026422:S-Nitrosoglutathione / Q000737:chemistry
chemical_list: D010718:Phosphatidylserines; D049294:Phospholipid Transfer Proteins; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D026422:S-Nitrosoglutathione; D053148:Caspase 3; D053179:Caspase 7; D009584:Nitrogen