This paper conducted research on biomechanical characteristics and biological activity of concavity-convex amniotic membrane (CCAM) and discussed its superiority as ocular surface repair material. Folding and compression with vacuum of fresh amniotic membrane were used to prepare CCAM. After cutting the striga of CCAM, sixteen CCAM tissue section were chosen at random to test their tensile strength using electronic universal testing machine. The bilayer amniotic membrane (BAM), the double-deck amniotic membrane (DAM) and the monolayer amniotic membrane (MAM) were as controls. The test parameters included yield strength, tensile strength, elongation at break, elastic modulus and so on. The cytokines of fresh amniotic membrane (FAM), MAM and CCAM were analyzed by radioimmunoassay method. The CCAM was obviously thicker than MAM and DAM. After 15 min in PBS, the CCAM tissue can recover the normal shape. The tensile strength and the elongation at break of CCAM were higher than those of the MAM and the DAM (P < 0.05). The elastic modulus of the CCAM was smaller than that of the MAM and the DAM (P < 0.05). The content of 10 cytokines [epidermal growth factor (EGF), fibroblast growth factor (FGF), b-fibroblast growth factor b-FGF, hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-beta), insulin-like growth factor (IGF), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), nerve growth factor (NGF), brain-derived nellrotrophic factor (BDNF), ciliary neurotrophic factor (CNTF)] of CCAM decreased significantly compared with the FAM and increased significantly compared with MAM and DAM in 6 cytokines (EGF, FGF, HGF, TGF-betap, PDGF, NGF; P < 0.05). The CCAM composites is thinner and has higher cytokine content than MAM, and better biomechanical properties than the MAM and the DAM, showing the superiority as ocular surface repair material.
Amnion ; chemistry ; physiology ; transplantation ; Biomechanical Phenomena ; Cytokines ; analysis ; Epidermal Growth Factor ; analysis ; Fibroblast Growth Factors ; analysis ; Hepatocyte Growth Factor ; analysis ; Humans ; Tissue Engineering ; methods ; Tissue Scaffolds

Human amniotic membrane isolated from the placenta contained basement membrane components such as type IV collagen, laminin, and 6 and 4 integrins, all of which remained detectable while preserved in glycerin for one week. One month after the n-heptanol removal of the total corneal epithelium and the limbal lamellar keratectomy, all rabbit eyes carried features of limbal deficiency, including conjunctival epithelial ingrowth, vascularization and chronic inflammation. Ten control eyes then received a total keratectomy, and 13 experimental eyes received an additional amniotic membrane transplantation. Three-month follow-ups revealed that all control corneas were revascularized to the center with granuloma and retained a conjunctival phenotype. In contrast, in the experimental groups, 5 corneas became clear with either minimal or no vascularization; the rest had either mild peripheral (5) or total (3) vascularization and more cloudy stroma. Using monoclonal antibodies for epithelial markers and matrix components, we concluded that the success correlated with the return of a cornea-like epithelial phenotype and the preservation of the amniotic membrane, whereas the failure maintained a conjunctival epithelial phenotype and the amniotic membrane was either partially degraded or covered by host fibrovascular stroma. Measures taken to facilitate the former might prove this procedure clinically useful for ocular surface reconstruction.
Amnion/chemistry/*transplantation ; Animals ; Antibodies, Monoclonal ; Basement Membrane/chemistry/pathology ; Cornea/pathology/*surgery ; Corneal Neovascularization/pathology/*prevention & control/surgery ; Epithelium/pathology/surgery ; Extracellular Matrix Proteins/analysis ; Female ; Fluorescent Antibody Technique ; Humans ; Immunophenotyping ; Male ; Rabbits

OBJECTIVETo investigate the proliferation and differentiation of cultured corneal stem cells and determine the effect of corneal stem cells cultured on amniotic membranes on the limbal area for treating corneal burns.

METHODSThe proliferation and differentiation of corneal stem cells in vitro had been examined using colony-forming efficiency and immunohistochemistry. The stem cells had been cultured on amniotic membranes and transplanted to the limbal area for treating corneal burns.

RESULTSCorneal stem cells had a high proliferation capacity in primary and first passage, cytokeratin 3 was not expressed in primary culture but partly in first passage. The stem cells could proliferate to form cell layer on an amniotic membrane. When transplanted, stem cells could survive on limbus. After transplantation, ocular inflammation resolved, the cornea re-epithelialized, the stromal opacity reduced, the superficial neovascularity was lessened and the conjunctival fornix re-established.

CONCLUSIONSOcular surface conditions could be improved by allograft of corneal stem cells cultured on amniotic membranes.

Alkalies ; Amnion ; transplantation ; Animals ; Burns, Chemical ; surgery ; Cell Culture Techniques ; methods ; Cell Differentiation ; Cell Division ; Cell Transplantation ; methods ; Cells, Cultured ; Epithelium, Corneal ; chemistry ; cytology ; Eye Burns ; chemically induced ; surgery ; Graft Survival ; Humans ; Keratins ; analysis ; Limbus Corneae ; chemistry ; cytology ; Rabbits ; Stem Cells ; chemistry ; cytology ; Treatment Outcome