title: Isthmin is a novel vascular permeability inducer that functions through cell-surface GRP78-mediated Src activation.
abstract: OBJECTIVE
  Isthmin (ISM) is a recently identified 60 kDa secreted angiogenesis inhibitor. Two cell-surface receptors for ISM have been defined, the high-affinity glucose-regulated protein 78 kDa (GRP78) and the low-affinity αvβ5 integrin. As αvβ5 integrin plays an important role in pulmonary vascular permeability (VP) and ISM is highly expressed in mouse lung, we sought to clarify the role of ISM in VP.
  
  
  RESULTS
  Recombinant ISM (rISM) dose-dependently enhances endothelial monolayer permeability in vitro and local dermal VP when administered intradermally in mice. Systemic rISM administration through intravenous injection leads to profound lung vascular hyperpermeability but not in other organs. Mechanistic investigations using molecular, biochemical approaches and specific chemical inhibitors revealed that ISM-GRP78 interaction triggers a direct interaction between GRP78 and Src, leading to Src activation and subsequent phosphorylation of adherens junction proteins and loss of junctional proteins from inter-endothelial junctions, resulting in enhanced VP. Dynamic studies of Src activation, VP and apoptosis revealed that ISM induces VP directly via Src activation while apoptosis contributes indirectly only after prolonged treatment. Furthermore, ISM is significantly up-regulated in lipopolysaccharide (LPS)-treated mouse lung. Blocking cell-surface GRP78 by systemic infusion of anti-GRP78 antibody significantly attenuates pulmonary vascular hyperpermeability in LPS-induced acute lung injury (ALI) in mice.
  
  
  CONCLUSIONS
  ISM is a novel VP inducer that functions through cell-surface GRP78-mediated Src activation as well as induction of apoptosis. It induces a direct GRP78-Src interaction, leading to cytoplasmic Src activation. ISM contributes to pulmonary vascular hyperpermeability of LPS-induced ALI in mice.
authors.affiliation: Department of Biological Sciences, Faculty of Science, 14 Science Drive 4, National University of Singapore, Singapore, Singapore 117543.
authors.forename: Shruthi
authors.initials: S
authors.lastname: Venugopal
authors.affiliation: Department of Biological Sciences, Faculty of Science, 14 Science Drive 4, National University of Singapore, Singapore, Singapore 117543.
authors.forename: Mo
authors.initials: M
authors.lastname: Chen
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University Health System, Singapore, Singapore Immunology Program, Life Science Institute, National University of Singapore, Singapore, Singapore 117456.
authors.forename: Wupeng
authors.initials: W
authors.lastname: Liao
authors.affiliation: Department of Biological Sciences, Faculty of Science, 14 Science Drive 4, National University of Singapore, Singapore, Singapore 117543.
authors.forename: Shi Yin
authors.initials: SY
authors.lastname: Er
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University Health System, Singapore, Singapore Immunology Program, Life Science Institute, National University of Singapore, Singapore, Singapore 117456.
authors.forename: Wai-Shiu Fred
authors.initials: WS
authors.lastname: Wong
authors.affiliation: Department of Biological Sciences, Faculty of Science, 14 Science Drive 4, National University of Singapore, Singapore, Singapore 117543 dbsgerw@nus.edu.sg.
authors.forename: Ruowen
authors.initials: R
authors.lastname: Ge
mesh_terms: D055371:Acute Lung Injury / Q000209:etiology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002199:Capillary Permeability / Q000187:drug effects; D002478:Cells, Cultured; D000091342:Endoplasmic Reticulum Chaperone BiP; D004789:Enzyme Activation; D005260:Female; D006360:Heat-Shock Proteins / Q000502:physiology*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides / Q000633:toxicity; D051379:Mice; D008807:Mice, Inbred BALB C; D010766:Phosphorylation; D011506:Proteins / Q000502:physiology*; D011994:Recombinant Proteins / Q000494:pharmacology; D019061:src-Family Kinases / Q000378:metabolism*
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; C000717248:Hspa5 protein, mouse; D036341:Intercellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides; D011506:Proteins; D011994:Recombinant Proteins; C571340:isthmin protein, mouse; D019061:src-Family Kinases
keywords: Acute lung injury; GRP78; Isthmin; Src; Vascular permeability