title: Short-term corneal response to cross-linking in rabbit eyes assessed by in vivo confocal laser scanning microscopy and histology.
abstract: OBJECTIVE
  Corneal cross-linking for the treatment of keratoconus has been tested in animal trials and proven clinically. A combination of in vivo confocal laser scanning microscopy (CLSM) and histology was used in rabbit corneas to assess early modifications at the cellular level after corneal cross-linking.
  
  
  METHODS
  Twelve New Zealand male rabbits were tested; in each case, the right eye was the study eye and left eye was the control eye. In vivo CLSM was performed on both eyes before and at 3 days and 1 week after cross-linking. Keratocyte and endothelial cell densities were determined by CLSM before and after cross-linking. After CLSM, the corneas were excised and processed for histology and immunohistochemistry.
  
  
  RESULTS
  Massive edema was observed 3 days after cross-linking. The corneal epithelium had already closed again by day 3. No cellular structures were detected in the stroma and endothelium. One week after cross-linking, normal corneal transparency and thickness were restored. The anterior stroma still lacked nuclei. The number of nuclei in the posterior stroma was significantly lower than that in the intact corneas. Highly reflective spindle-shaped structures were detected in the posterior stroma. The endothelial monolayer had closed again but still showed significantly decreased cell density. At 1 week after cross-linking, immunohistochemical staining revealed the presence of proliferating cells in the corneal epithelium, posterior stroma, and endothelium.
  
  
  CONCLUSIONS
  The early response of the rabbit cornea to cross-linking was successfully characterized at the cellular level by in vivo CLSM and histology, and the results obtained with both techniques correlated positively.
authors.affiliation: Department of Ophthalmology, University of Rostock, Rostock, Germany. marinahovakimyan@yahoo.com
authors.forename: Marine
authors.initials: M
authors.lastname: Hovakimyan
authors.forename: Rudolf
authors.initials: R
authors.lastname: Guthoff
authors.forename: Steffi
authors.initials: S
authors.lastname: Knappe
authors.forename: Andrey
authors.initials: A
authors.lastname: Zhivov
authors.forename: Andreas
authors.initials: A
authors.lastname: Wree
authors.forename: Alexander
authors.initials: A
authors.lastname: Krüger
authors.forename: Alexander
authors.initials: A
authors.lastname: Heisterkamp
authors.forename: Oliver
authors.initials: O
authors.lastname: Stachs
mesh_terms: D000818:Animals; D003094:Collagen / Q000378:metabolism / Q000528:radiation effects; D003315:Cornea / Q000473:pathology*; D003319:Corneal Stroma / Q000378:metabolism / Q000473:pathology; D004728:Endothelium, Corneal / Q000473:pathology; D019573:Epithelium, Corneal / Q000473:pathology; D007640:Keratoconus / Q000188:drug therapy* / Q000473:pathology*; D008297:Male; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000627:therapeutic use*; D011817:Rabbits; D012256:Riboflavin / Q000627:therapeutic use*; D013997:Time Factors; D014466:Ultraviolet Rays
chemical_list: D017319:Photosensitizing Agents; D003094:Collagen; D012256:Riboflavin