title: Pro-inflammatory TNFα and IL-1β differentially regulate the inflammatory phenotype of brain microvascular endothelial cells.
abstract: BACKGROUND
  The vasculature of the brain is composed of endothelial cells, pericytes and astrocytic processes. The endothelial cells are the critical interface between the blood and the CNS parenchyma and are a critical component of the blood-brain barrier (BBB). These cells are innately programmed to respond to a myriad of inflammatory cytokines or other danger signals. IL-1β and TNFα are well recognised pro-inflammatory mediators, and here, we provide compelling evidence that they regulate the function and immune response profile of human cerebral microvascular endothelial cells (hCMVECs) differentially.
  
  
  METHODS
  We used xCELLigence biosensor technology, which revealed global differences in the endothelial response between IL-1β and TNFα. xCELLigence is a label-free impedance-based biosensor, which is ideal for acute or long-term comparison of drug effects on cell behaviour. In addition, flow cytometry and multiplex cytokine arrays were used to show differences in the inflammatory responses from the endothelial cells.
  
  
  RESULTS
  Extensive cytokine-secretion profiling and cell-surface immune phenotyping confirmed that the immune response of the hCMVEC to IL-1β was different to that of TNFα. Interestingly, of the 38 cytokines, chemokines and growth factors measured by cytometric bead array, the endothelial cells secreted only 13. Of importance was the observation that the majority of these cytokines were differentially regulated by either IL-1β or TNFα. Cell-surface expression of ICAM-1 and VCAM-1 were also differentially regulated by IL-1β or TNFα, where TNFα induced a substantially higher level of expression of both key leukocyte-adhesion molecules. A range of other cell-surface cellular and junctional adhesion molecules were basally expressed by the hCMVEC but were unaffected by IL-1β or TNFα.
  
  
  CONCLUSIONS
  To our knowledge, this is the most comprehensive analysis of the immunological profile of brain endothelial cells and the first direct evidence that human brain endothelial cells are differentially regulated by these two key pro-inflammatory mediators.
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. s.ocarroll@auckland.ac.nz.|Department of Anatomy, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. s.ocarroll@auckland.ac.nz.
authors.forename: Simon J
authors.initials: SJ
authors.lastname: O'Carroll
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. cd.kho@auckland.ac.nz.|Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. cd.kho@auckland.ac.nz.
authors.forename: Dan Ting
authors.initials: DT
authors.lastname: Kho
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. rachaelwiltshire@xtra.co.nz.
authors.forename: Rachael
authors.initials: R
authors.lastname: Wiltshire
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. vicky.nelson@windowslive.com.|Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. vicky.nelson@windowslive.com.
authors.forename: Vicky
authors.initials: V
authors.lastname: Nelson
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. odunayo.rotimi@auckland.ac.nz.|Department of Anatomy, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. odunayo.rotimi@auckland.ac.nz.
authors.forename: Odunayo
authors.initials: O
authors.lastname: Rotimi
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. rebecca.johnson@auckland.ac.nz.|Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. rebecca.johnson@auckland.ac.nz.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Johnson
authors.affiliation: School of Biological Sciences, Faculty of Science, University of Auckland, Auckland, New Zealand. c.angel@auckland.ac.nz.
authors.forename: Catherine E
authors.initials: CE
authors.lastname: Angel
authors.affiliation: Centre for Brain Research, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. s.graham@auckland.ac.nz.|Department of Pharmacology and Clinical Pharmacology, School of Medical Sciences, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand. s.graham@auckland.ac.nz.
authors.forename: E Scott
authors.initials: ES
authors.lastname: Graham
mesh_terms: D001921:Brain / Q000098:blood supply; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism; D004660:Encephalitis / Q000378:metabolism / Q000473:pathology*; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D016130:Immunophenotyping; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D053583:Interleukin-1beta / Q000494:pharmacology*; D055806:Microvessels / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D010641:Phenotype; D062725:Tight Junction Proteins / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
chemical_list: D016207:Cytokines; D053583:Interleukin-1beta; D062725:Tight Junction Proteins; D014409:Tumor Necrosis Factor-alpha; D019010:Vascular Cell Adhesion Molecule-1; D018799:Intercellular Adhesion Molecule-1