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S. Mi, B. Chen, B. Wright, C. Connon; Construction of an Artificial Ocular Surface by Combination of Corneal Limbal Epithelial Cells and a Compressed Collagen Scaffold Containing Keratocytes. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1935.
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© ARVO (1962-2015); The Authors (2016-present)
We investigated the use of a laminin coated compressed collagen gel containing corneal keratocytes as a novel scaffold to support the growth of corneal limbal epithelial cells. Such a gel has mechanical properties similar to corneal stroma.
All animal experiments conformed to the ARVO statement for the use of animals in ophthalmic and vision research. The growth of limbal epithelial cells was compared between compressed collagen gel and a clinically proven conventional substrate, denuded amniotic membrane. Using a series of methods including immunostaining, western blotting and real-time PCR to assess the relative phenotype of cells grown on both scaffolds.
Following compression of the collagen gel the encapsulated keratocytes remained viable and scanning electron microscopy showed that fibres within the compressed gel were dense, homogeneous and similar in structure to those within denuded amniotic membrane. Limbal epithelial cells were successfully expanded upon the compressed collagen resulting in stratified layers of cells containing desmosome and hemidesmosome structures. The resulting corneal constructs shared a high degree of transparency, cell morphology and cell stratification. Similar protein expression profiles for K3 and K14 and no significant difference in K12 mRNA expression levels by real time PCR were also observed.
This study provides the first line of evidence that a laminin coated compressed collagen gel containing keratocytes can adequately support limbal epithelial cell expansion, stratification and differentiation to a degree that is comparable to the leading conventional scaffold, denuded amniotic membrane.
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