Sterilized, Freeze-Dried Amniotic Membrane: A Useful Substrate for Ocular Surface Reconstruction

Abstract

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.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.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.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.