title: Sterilized, freeze-dried amniotic membrane: a useful substrate for ocular surface reconstruction.
abstract: OBJECTIVE
  To examine the feasibility of using sterilized, freeze-dried amniotic membrane (FD-AM) as a substrate for cultivating autologous corneal epithelial cells for ocular surface reconstruction.
  
  
  METHODS
  Human AM deprived of amniotic epithelial cells by incubation with EDTA was freeze dried, vacuum packed, and sterilized with gamma-irradiation. The resultant FD-AM was characterized for its physical, biological, and morphologic properties by stretch stress tests, immunohistochemistry, electron microscopy, and cell culture. In addition, 3 weeks after an ocular surface injury, the conjunctivalized corneal surfaces of eyes in eight rabbits were surgically reconstructed by transplantation of autologous cultivated corneal epithelial cells on FD-AM.
  
  
  RESULTS
  A stretch stress test revealed no significant differences between sterilized FD-AM and cryopreserved AM. Immunohistochemistry for several extracellular matrix molecules and electron microscopic analysis of FD-AM revealed that the process of drying and irradiation did not affect its biological and morphologic properties. The corneal epithelial cells cultivated on FD-AM had four to five stratified, well-differentiated cell layers. Corneas that were grafted with the cultivated corneal epithelial cells on FD-AM were clear and were all epithelialized at 10 days after surgery.
  
  
  CONCLUSIONS
  The sterilized, freeze-dried AM retained most of the physical, biological, and morphologic characteristics of cryopreserved AM; consequently, it is a useful biomaterial for ocular surface reconstruction.
authors.affiliation: Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan. tnakamur@ophth.kpu-m.ac.jp
authors.forename: Takahiro
authors.initials: T
authors.lastname: Nakamura
authors.forename: Makoto
authors.initials: M
authors.lastname: Yoshitani
authors.forename: Helen
authors.initials: H
authors.lastname: Rigby
authors.forename: Nigel J
authors.initials: NJ
authors.lastname: Fullwood
authors.forename: Wakana
authors.initials: W
authors.lastname: Ito
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Inatomi
authors.forename: Chie
authors.initials: C
authors.lastname: Sotozono
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Nakamura
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Shimizu
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kinoshita
mesh_terms: D000650:Amnion / Q000648:ultrastructure; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D017690:Cell Transplantation / Q000379:methods*; D018920:Coculture Techniques; D003316:Corneal Diseases / Q000601:surgery*; D004847:Epithelial Cells / Q000637:transplantation*; D019573:Epithelium, Corneal / Q000166:cytology*; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005240:Feasibility Studies; D005612:Freeze Drying; D006801:Humans; D007150:Immunohistochemistry; D008855:Microscopy, Electron, Scanning; D011817:Rabbits; D019651:Plastic Surgery Procedures; D013242:Sterilization
chemical_list: D016326:Extracellular Matrix Proteins