title: TNFα signals via p66(Shc) to induce E-Selectin, promote leukocyte transmigration and enhance permeability in human endothelial cells.
abstract: Endothelial cells participate in inflammatory events leading to atherogenesis by regulating endothelial cell permeability via the expression of VE-Cadherin and β-catenin and leukocyte recruitment via the expression of E-Selectins and other adhesion molecules. The protein p66(Shc) acts as a sensor/inducer of oxidative stress and may promote vascular dysfunction. The objective of this study was to investigate the role of p66(Shc) in tumor necrosis factor TNFα-induced E-Selectin expression and function in human umbilical vein endothelial cells (HUVEC). Exposure of HUVEC to 50 ng/ml TNFα resulted in increased leukocyte transmigration through the endothelial monolayer and E-Selectin expression, in association with augmented phosphorylation of both p66(Shc) on Ser(36) and the stress kinase c-Jun NH2-terminal protein kinase (JNK)-1/2, and higher intracellular reactive oxygen species (ROS) levels. Overexpression of p66(Shc) in HUVEC resulted in enhanced p66(Shc) phosphorylation on Ser(36), increased ROS and E-Selectin levels, and amplified endothelial cell permeability and leukocyte transmigration through the HUVEC monolayer. Conversely, overexpression of a phosphorylation-defective p66(Shc) protein, in which Ser(36) was replaced by Ala, did not augment ROS and E-Selectin levels, nor modify cell permeability or leukocyte transmigration beyond those found in wild-type cells. Moreover, siRNA-mediated silencing of p66(Shc) resulted in marked reduction of E-Selectin expression and leukocyte transmigration. In conclusion, p66(Shc) acts as a novel intermediate in the TNFα pathway mediating endothelial dysfunction, and its action requires JNK-dependent phosphorylation of p66(Shc) on Ser(36).
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Luigi
authors.initials: L
authors.lastname: Laviola
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Maura Roberta
authors.initials: MR
authors.lastname: Orlando
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Maria Angela
authors.initials: MA
authors.lastname: Incalza
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Caccioppoli
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Mariangela
authors.initials: M
authors.lastname: Melchiorre
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Anna
authors.initials: A
authors.lastname: Leonardini
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Angelo
authors.initials: A
authors.lastname: Cignarelli
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: Tortosa
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Rossella
authors.initials: R
authors.lastname: Labarbuta
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Sabina
authors.initials: S
authors.lastname: Martemucci
authors.affiliation: Department of Medical Biochemistry, Biology and Physics, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Consiglia
authors.initials: C
authors.lastname: Pacelli
authors.affiliation: Department of Medical Biochemistry, Biology and Physics, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Tiziana
authors.initials: T
authors.lastname: Cocco
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Sebastio
authors.initials: S
authors.lastname: Perrini
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Annalisa
authors.initials: A
authors.lastname: Natalicchio
authors.affiliation: Department of Emergency and Organ Transplantation - Section of Internal Medicine, Endocrinology, Andrology and Metabolic Diseases, University of Bari Aldo Moro, Bari, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Giorgino
mesh_terms: D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D018920:Coculture Techniques; D019040:E-Selectin / Q000235:genetics* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation; D018922:HL-60 Cells; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D048055:Mitogen-Activated Protein Kinase 8 / Q000235:genetics / Q000378:metabolism; D048056:Mitogen-Activated Protein Kinase 9 / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D057705:Transendothelial and Transepithelial Migration / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D019040:E-Selectin; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014409:Tumor Necrosis Factor-alpha; D048056:Mitogen-Activated Protein Kinase 9; D048055:Mitogen-Activated Protein Kinase 8