PURPOSE: Cultivated limbal epithelial transplantation using an amniotic membrane (AM) carrier is now widely used for ocular surface reconstruction. The reasons for the exceptional success of AM as a carrier are not fully understood but are believed to be related to its unique composition. In this project we characterize, at the ultrastructural level, the extracellular matrix (ECM) components present in AM. We also compare the distribution of ECM components of cellular AM with that of denuded AM. METHODS: Scanning, transmission, and atomic force microscopy was used to examine the structure of cellular and denuded amniotic membranes. Immunogold labeling with a panel of antibodies against ECM molecules was carried out on cellular and denuded AM. RESULTS: Heparan sulfate, fibronectin, and laminin were present at high concentration in the lamina densa, Collagen IV was the major component of the basal lamina. Type I collagen was confined to the stroma along with significant amounts of keratan and chondroitin sulfate. Both cellular and denuded AMs had similar distributions of the ECM components. CONCLUSIONS: We were able to determine the distribution of ECM molecules in the lamina densa, basal lamina, and stroma of AM at the ultrastructural level. The removal of amniotic epithelial cells using our protocol does not appear to have any significant effects on the structure of the basal lamina or the distribution of ECM components.
PURPOSE: Cultivated limbal epithelial transplantation using an amniotic membrane (AM) carrier is now widely used for ocular surface reconstruction. The reasons for the exceptional success of AM as a carrier are not fully understood but are believed to be related to its unique composition. In this project we characterize, at the ultrastructural level, the extracellular matrix (ECM) components present in AM. We also compare the distribution of ECM components of cellular AM with that of denuded AM. METHODS: Scanning, transmission, and atomic force microscopy was used to examine the structure of cellular and denuded amniotic membranes. Immunogold labeling with a panel of antibodies against ECM molecules was carried out on cellular and denuded AM. RESULTS:Heparan sulfate, fibronectin, and laminin were present at high concentration in the lamina densa, Collagen IV was the major component of the basal lamina. Type I collagen was confined to the stroma along with significant amounts of keratan and chondroitin sulfate. Both cellular and denuded AMs had similar distributions of the ECM components. CONCLUSIONS: We were able to determine the distribution of ECM molecules in the lamina densa, basal lamina, and stroma of AM at the ultrastructural level. The removal of amniotic epithelial cells using our protocol does not appear to have any significant effects on the structure of the basal lamina or the distribution of ECM components.
Authors: Gisele P Valdetaro; Marcela Aldrovani; Ivan R M Padua; Priscila C Cristovam; José A P Gomes; José L Laus Journal: Biomed Opt Express Date: 2016-11-07 Impact factor: 3.732