title: High glucose increases lysyl oxidase expression and activity in retinal endothelial cells: mechanism for compromised extracellular matrix barrier function.
abstract: OBJECTIVE
  In diabetes, retinal vascular basement membrane (BM) undergoes significant thickening and compromises vessel function including increased vascular permeability, a prominent lesion of early diabetic retinopathy. In this study we determined whether altered expression and activity of lysyl oxidase (LOX), a cross-linking enzyme, may compromise vascular basement membrane functional integrity under high-glucose (HG) conditions.
  
  
  METHODS
  Rat retinal endothelial cells (RRECs) grown in normal (5 mmol/l) or HG (30 mmol/l glucose) medium for 7 days were assessed for expression of LOX and proLOX by Western blot analysis and LOX enzyme activity. To determine whether HG alters cellular distribution patterns of LOX and proLOX, immunostaining with respective antibodies was performed. Similarly, cells grown in normal or HG medium were subjected to both LOX inhibition with β-aminopropionitrile (BAPN) and by small interfering RNA knockdown, and respectively examined for cell monolayer permeability. Additionally, retinas of streptozotocin (STZ)-induced diabetic rats were analyzed to determine if diabetes altered LOX expression.
  
  
  RESULTS
  Western blot analysis revealed significantly increased LOX and proLOX expression in cells grown in HG medium compared with those grown in normal medium. The increased LOX level was strikingly similar to LOX upegulation in the diabetic retinas. In cells grown in HG medium, LOX activity and cell monolayer permeability was significantly increased, as were LOX and proLOX immunostaining. Small interfering RNA- or BAPN-induced-specific blockage of LOX expression or activity, respectively, reduced cell monolayer permeability.
  
  
  CONCLUSIONS
  HG-induced increased LOX expression and activity compromises barrier functional integrity, a prominent lesion of diabetic retinopathy.
authors.affiliation: Departments of Medicine and Ophthalmology, Boston University School of Medicine, Boston, Massachusetts, USA.
authors.forename: Argyrios
authors.initials: A
authors.lastname: Chronopoulos
authors.forename: Amanda
authors.initials: A
authors.lastname: Tang
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Beglova
authors.forename: Philip C
authors.initials: PC
authors.lastname: Trackman
authors.forename: Sayon
authors.initials: S
authors.lastname: Roy
mesh_terms: D000629:Aminopropionitrile / Q000494:pharmacology; D000818:Animals; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology; D003930:Diabetic Retinopathy / Q000235:genetics / Q000473:pathology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology; D005109:Extracellular Matrix / Q000187:drug effects / Q000502:physiology*; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology*; D008297:Male; D008249:Protein-Lysine 6-Oxidase / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012160:Retina / Q000187:drug effects / Q000201:enzymology; D012172:Retinaldehyde / Q000378:metabolism
chemical_list: D000629:Aminopropionitrile; D008249:Protein-Lysine 6-Oxidase; D005947:Glucose; D012172:Retinaldehyde
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