title: Second-harmonic imaging microscopy of normal human and keratoconus cornea.
abstract: OBJECTIVE
  The purpose of this study was to evaluate the ability of second-harmonic imaging to identify differences in corneal stromal collagen organization between normal human and keratoconus corneas.
  
  
  METHODS
  Six normal corneas from eye bank donors and 13 corneas of patients with keratoconus, obtained after penetrating keratoplasty were examined. A femtosecond titanium-sapphire laser with 800-nm output was used to generate second-harmonic signals collected at 400 nm from central and paracentral corneal tissue blocks. Three-dimensional (3-D) data sets were collected and reconstructed to evaluate the location and orientation of stromal collagen lamellae.
  
  
  RESULTS
  Imaging of second-harmonic signals combined with 3-D reconstruction of the normal cornea identified a high degree of lamellar interweaving, particularly in the anterior cornea. Of note was the detection of lamellae that inserted into Bowman's layer and were oriented transverse to the corneal surface, penetrating posteriorly approximately 120 mum. In keratoconus corneas, imaging second-harmonic signals identified less lamellar interweaving and a marked reduction or loss of lamellae inserting into Bowman's layer in 12 of 13 cases, particularly in regions associated with cone development without breaks in Bowman's layer or scarring.
  
  
  CONCLUSIONS
  Compared with normal adult corneas, marked abnormalities were detected in the organization of the anterior corneal collagen lamellae of keratoconus corneas by second harmonic imaging. These structural abnormalities are consistent with the known changes in collagen organization and biomechanical strength of keratoconus.
authors.affiliation: The Eye Institute, School of Medicine, University of California Irvine Medical Center, Orange, California.
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Morishige
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Wahlert
authors.forename: M Cristina
authors.initials: MC
authors.lastname: Kenney
authors.forename: Donald J
authors.initials: DJ
authors.lastname: Brown
authors.forename: Koji
authors.initials: K
authors.lastname: Kawamoto
authors.forename: Tai-Ichiro
authors.initials: T
authors.lastname: Chikama
authors.forename: Teruo
authors.initials: T
authors.lastname: Nishida
authors.forename: James V
authors.initials: JV
authors.lastname: Jester
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D003094:Collagen / Q000378:metabolism*; D003319:Corneal Stroma / Q000378:metabolism*; D005125:Eye Banks; D005260:Female; D006801:Humans; D021621:Imaging, Three-Dimensional; D007640:Keratoconus / Q000378:metabolism* / Q000601:surgery; D015948:Keratoplasty, Penetrating; D008297:Male; D018613:Microscopy, Confocal; D036641:Microscopy, Fluorescence, Multiphoton; D008875:Middle Aged; D014019:Tissue Donors
chemical_list: D003094:Collagen