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S. Proulx, P. Carrier, I. Brunette, F. A. Auger, L. Germain; Characterization of a Posterior Corneal Substitute Reconstructed Using the Self-Assembly Approach of Tissue Engineering. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2836. doi: https://doi.org/.
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To characterize a tissue-engineered posterior cornea reconstructed using the self-assembly approach.
Human corneal keratocytes were cultured in the presence of serum and ascorbic acid, forming sheets of extracellular matrix that were then superposed to reconstruct a corneal stroma. Afterward, human corneal endothelial cells were seeded on top and cultured for 28 days (n=14). These reconstructed posterior corneas were analysed using histology, scanning and transmission electron microscopy and immunofluorescent staining of various proteins. Alizarin red staining was used to evaluate final endothelial cell densities (ECD).
The reconstructed endothelium adhered on the self-assembled stromal matrix and formed a monolayer of flattened cells with a normal ultrastructure and endothelial cell morphology. Endothelial cells expressed Na+/K+-ATPase α1. The reconstructed corneal stroma expressed collagen types I, IV, V, VI, XII, XIV and did not express collagen types II and III. Morphometric analysis of alizarin red stained reconstructed endothelium revealed final ECD between 732 and 1434 cells/mm2 (mean: 982±226) and 43±4% of 6-sided cells.
This study shows the feasibility of reconstructing a posterior corneal substitute with the self-assembly approach, using untransformed human cells and without adding exogenous biomaterials. Potential applications of this model are numerous, including various in vitro pharmacological studies and in vivo grafting. We consider that such results represent an additional step towards the future development bioengineered corneas.
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