Sterilized, freeze-dried amniotic membrane: a useful substrate for ocular surface reconstruction.
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.
Nakamura
Takahiro
T
Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan. tnakamur@ophth.kpu-m.ac.jp
Yoshitani
Makoto
M
Rigby
Helen
H
Fullwood
Nigel J
NJ
Ito
Wakana
W
Inatomi
Tsutomu
T
Sotozono
Chie
C
Nakamura
Tatsuo
T
Shimizu
Yasuhiko
Y
Kinoshita
Shigeru
S
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
D016326:Extracellular Matrix Proteins