title: ICAM-1 cytoplasmic tail regulates endothelial glutathione synthesis through a NOX4/PI3-kinase-dependent pathway.
abstract: We previously reported that ICAM-1 expression modulates endothelial intracellular glutathione (GSH) metabolism through unknown mechanisms. Here we report that the cytoplasmic tail of ICAM-1 is critically involved in governing intracellular GSH production. Peptides containing the antennapedia cell-permeative sequence (AP) or an AP peptide linked to the transmembrane and cytosolic tail of ICAM-1 (AP-ICAM) were synthesized and used to measure alterations in redox status in cultured endothelial cells and determine their biological effect. Treatment with AP-ICAM significantly increased GSH concentrations and glutamate-cysteine ligase (GCL) activity over time. Measuring reactive oxygen species (ROS) production with DCF revealed a rapid increase in ROS generation after AP-ICAM treatment. Measurement of superoxide production with hydroethidium revealed biphasic production at 30 min and 6h after treatment with AP-ICAM. Apocynin, DPI, catalase, or SOD attenuated AP-ICAM-dependent ROS production, GCL activity, and GSH production, implicating superoxide production and dismutation to peroxide. Consistent with these findings, NOX4 siRNA knockdown blocked AP-ICAM peptide increases in GSH or GCL activity, demonstrating the importance of NADPH oxidase. Last, inhibition of PI3-kinase activity with LY 294002 or wortmannin blocked AP-ICAM GSH induction and ROS production. These data reveal that the ICAM-1 cytoplasmic tail regulates production of endothelial GSH through a NOX4/PI3-kinase-dependent redox-sensitive pathway.
authors.affiliation: Department of Pathology, Louisiana State University Health Sciences Center at Shreveport, Shreveport, LA 71130, USA.
authors.forename: Christopher B
authors.initials: CB
authors.lastname: Pattillo
authors.forename: Sibile
authors.initials: S
authors.lastname: Pardue
authors.forename: Xinggui
authors.initials: X
authors.lastname: Shen
authors.forename: Kai
authors.initials: K
authors.lastname: Fang
authors.forename: Will
authors.initials: W
authors.lastname: Langston
authors.forename: David
authors.initials: D
authors.lastname: Jourd'heuil
authors.forename: Terrance J
authors.initials: TJ
authors.lastname: Kavanagh
authors.forename: Rakesh P
authors.initials: RP
authors.lastname: Patel
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Kevil
mesh_terms: D000098:Acetophenones / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D004789:Enzyme Activation / Q000187:drug effects; D005721:Glutamate-Cysteine Ligase / Q000235:genetics / Q000378:metabolism*; D005978:Glutathione / Q000096:biosynthesis; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000494:pharmacology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017434:Protein Structure, Tertiary / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D000077191:Wortmannin
chemical_list: D000098:Acetophenones; D000730:Androstadienes; D002867:Chromones; D009025:Morpholines; D010446:Peptide Fragments; D000081082:Phosphoinositide-3 Kinase Inhibitors; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D018799:Intercellular Adhesion Molecule-1; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C056165:acetovanillone; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C422943:Nox4 protein, mouse; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D000077191:Wortmannin