title: Advanced glycation end products in Descemet's membrane and their effect on corneal endothelial cell.
abstract: OBJECTIVE
  [corrected] The purpose of this study was to evaluate the effect of advanced glycation end products (AGEs) in Descemet's membrane on the attachment and spreading of the corneal endothelial cells.
  
  
  METHODS
  An anti-AGEs monoclonal antibody (6D12), which recognizes a N(epsilon)-carboxymethyl lysine (CML)-protein adduct as an epitope, was used for immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). Fresh bovine Descemet's membrane was incubated for 4 weeks in the buffered solution with 500 mM of glucose-6-phosphate (G-6-P). In the incubated Descemet's membrane, the immunohistochemical localization of CML was examined. Type I collagen-, type IV collagen-, fibronectin-, or laminin-coated 96-well plates were glycated by G-6-P. The amount of CML was determined by ELISA using 6D12. Cultured bovine corneal endothelial cells were seeded onto glycated or non-glycated extracellular matrix (ECM) in 96-well plates and allowed to attach for 3 hours. The number and the surface area of the attached cells were examined.
  
  
  RESULTS
  Immunoreactivity to CML was detected in Descemet's membrane incubated in the buffered solution containing G-6-P. Glycation of fibronectin and laminin decreased the number and the surface area of the attached corneal endothelial cells. Aminoguanidine in the incubation mixture inhibited CML formation of ECM components and increased the number and the surface area of the attached corneal endothelial cells in a dose-dependent manner.
  
  
  CONCLUSIONS
  AGE formation on fibronectin and laminin attenuated the attachment and spreading of the corneal endothelial cells. AGEs' formation in Descemet's membrane may be responsible for the corneal endothelial cell loss with aging and corneal endothelial abnormalities in diabetic patients
authors.affiliation: Department of Ophthalmology, University of Tokyo Graduate School of Medicine, Tokyo, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Kaji
authors.forename: S
authors.initials: S
authors.lastname: Amano
authors.forename: T
authors.initials: T
authors.lastname: Usui
authors.forename: K
authors.initials: K
authors.lastname: Suzuki
authors.forename: S
authors.initials: S
authors.lastname: Tanaka
authors.forename: T
authors.initials: T
authors.lastname: Oshika
authors.forename: R
authors.initials: R
authors.lastname: Nagai
authors.forename: S
authors.initials: S
authors.lastname: Horiuchi
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D002417:Cattle; D002448:Cell Adhesion; D002455:Cell Division; D002478:Cells, Cultured; D024042:Collagen Type I / Q000378:metabolism; D024141:Collagen Type IV / Q000378:metabolism; D003886:Descemet Membrane / Q000378:metabolism*; D004728:Endothelium, Corneal / Q000166:cytology* / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D005353:Fibronectins / Q000378:metabolism; D017127:Glycation End Products, Advanced / Q000378:metabolism*; D006031:Glycosylation; D006146:Guanidines / Q000494:pharmacology; D007124:Immunoenzyme Techniques; D007797:Laminin / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives / Q000276:immunology / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D024042:Collagen Type I; D024141:Collagen Type IV; D005353:Fibronectins; D017127:Glycation End Products, Advanced; D006146:Guanidines; D007797:Laminin; C048496:N(6)-carboxymethyllysine; D008239:Lysine; C004479:pimagedine