title: Hyperglycaemia promotes cerebral barrier dysfunction through activation of protein kinase C-β.
abstract: OBJECTIVE
  To examine whether protein kinase C (PKC) and associated downstream mechanisms are involved in hyperglycaemia (HG)-evoked blood-brain barrier (BBB) damage.
  
  
  METHODS
  The activities of total PKC (Peptag assay), NADPH oxidase (lucigenin assay) and matrix metalloproteinase-2 (MMP-2; gelatin zymography) were measured in human brain microvascular endothelial cells (HBMEC) exposed to normoglycaemia (5.5 mM) or HG (25 mM) using the specific assays indicated in parentheses. The integrity and function of the in vitro models of human BBB were assessed by measurements of transendothelial electrical resistance and paracellular flux of permeability markers, respectively. Occludin protein expression was studied by immunoblotting.
  
  
  RESULTS
  HG significantly compromised the BBB integrity and enhanced total PKC activity to which increases in PKC-β and PKC-βII isoforms contributed the most. Elevations in NADPH oxidase and MMP-2 activities and decreases in occludin levels contributed to barrier dysfunction. Selective inhibition of PKC-β isoform prevented the changes observed in occludin expression and the aforementioned enzyme activities and thus effectively preserved barrier integrity. Similarly, apocynin, a specific NADPH oxidase inhibitor, also effectively neutralized the effects of HG on barrier integrity, MMP-2 activity, occludin expression and PKC-β activity.
  
  
  CONCLUSIONS
  HG promotes cerebral-barrier dysfunction through activation of PKC-β and consequent stimulations of oxidative stress and tight junction dissolution.
authors.affiliation: Division of Stroke Medicine, School of Clinical Sciences, The University of Nottingham, Nottingham, UK.
authors.forename: B
authors.initials: B
authors.lastname: Shao
authors.forename: U
authors.initials: U
authors.lastname: Bayraktutan
mesh_terms: D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D001812:Blood-Brain Barrier / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000503:physiopathology*; D001921:Brain / Q000098:blood supply / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002199:Capillary Permeability / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D015536:Down-Regulation / Q000187:drug effects; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D006943:Hyperglycemia / Q000201:enzymology / Q000378:metabolism / Q000503:physiopathology*; D020778:Matrix Metalloproteinase 2 / Q000737:chemistry / Q000378:metabolism; D055806:Microvessels / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D019255:NADPH Oxidases / Q000737:chemistry / Q000378:metabolism; D062793:Occludin / Q000378:metabolism; D020286:Pericytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D064546:Protein Kinase C beta / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism*
chemical_list: D062793:Occludin; D047428:Protein Kinase Inhibitors; D019255:NADPH Oxidases; C579054:PRKCB protein, human; D064546:Protein Kinase C beta; C522361:MMP2 protein, human; D020778:Matrix Metalloproteinase 2
keywords: blood-brain barrier; hyperglycaemia; occludin; oxidative stress; protein kinase C