title: Evaluation of UVA Cytotoxicity for Human Endothelium in an Ex Vivo Corneal Cross-linking Experimental Setting.
abstract: OBJECTIVE
  To evaluate endothelial cytotoxicity after exposure of human corneas to ultraviolet-A (UVA) (λ = 365 nm; 5.4 J/cm(2)) in an experimental ex vivo corneal cross-linking setting.
  
  
  METHODS
  Sixteen pairs of human donor corneas were cut into two pieces. One piece of each cornea was treated with 0.025% riboflavin solution prior to UVA irradiation (5 minutes; 18 mW/cm(2)), whereas the other piece was not irradiated but treated with riboflavin (right eye) or preservation medium (left eye). By irradiating from the endothelial side, the UVA dosage applied to endothelial cells exceeded at least eight times the cytotoxic threshold established in animal models (0.65 J/cm(2)). Endothelial cell counts were performed by two independent investigators after storage (4 to 5 days at 31 °C) and staining (trypan blue, alizarin red). Normality (Q-Q plot; Shapiro-Wilk test) and equivalence (mixed-effects modeling with a 10% equivalence threshold) of the endothelial cell counts of the different groups were evaluated.
  
  
  RESULTS
  Equivalence of mean endothelial cell density between both groups was observed: 2,237 ± 208 cells/mm(2) in UVA-irradiated pieces and 2,290 ± 281 cells/mm(2) in control pieces (mean difference of 53 ± 240 cells/mm(2) between both groups).
  
  
  CONCLUSIONS
  Despite direct irradiation of human donor endothelium using the clinical dosage for cross-linking, equivalence in endothelial cell counts was observed between irradiated tissues and controls. Ex vivo human corneal endothelial cells seem to be far more resistant to riboflavin-enhanced UVA irradiation than previously estimated by animal experiments.
authors.forename: Pepijn
authors.initials: P
authors.lastname: Mooren
authors.forename: Laure
authors.initials: L
authors.lastname: Gobin
authors.forename: Nezahat
authors.initials: N
authors.lastname: Bostan
authors.forename: Kristien
authors.initials: K
authors.lastname: Wouters
authors.forename: Nadia
authors.initials: N
authors.lastname: Zakaria
authors.forename: Danny G P
authors.initials: DG
authors.lastname: Mathysen
authors.forename: Carina
authors.initials: C
authors.lastname: Koppen
mesh_terms: D000328:Adult; D000368:Aged; D002452:Cell Count; D003094:Collagen / Q000378:metabolism; D055954:Corneal Endothelial Cell Loss / Q000503:physiopathology*; D003319:Corneal Stroma / Q000378:metabolism; D003432:Cross-Linking Reagents; D004728:Endothelium, Corneal / Q000473:pathology / Q000528:radiation effects*; D005125:Eye Banks; D006801:Humans; D008875:Middle Aged; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000627:therapeutic use; D011832:Radiation Injuries / Q000503:physiopathology*; D012256:Riboflavin / Q000627:therapeutic use; D014019:Tissue Donors; D014466:Ultraviolet Rays / Q000009:adverse effects*
chemical_list: D003432:Cross-Linking Reagents; D017319:Photosensitizing Agents; D003094:Collagen; D012256:Riboflavin