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Hannah J. Levis, Kah Peng Toh, Rebekah W. Poh, Gary S. Peh, Jodhbir S. Mehta, Julie T. Daniels; Plastic Compressed Collagen As A Novel Carrier For Expanded Human Corneal Endothelial Cells For Transplantation. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1737.
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Descemet’s membrane endothelial keratoplasty is performed to replace failed endothelium in conditions such as Fuch’s endothelial dystrophy. Current methods require the use of one donor per recipient. Human corneal endothelial cells can be successfully expanded in culture but will require a suitable substrate and carrier for transplantation. We describe the use of plastic compressed collagen as a novel substrate for culture and transplantation of human corneal endothelial cells.
Plastic compressed collagen constructs were produced by casting gels of neutralised rat-tail type I collagen, 10x MEM and endothelial medium in 12 well plates. Gels were set at 37°C for 30 min and then compressed for 15 min. Trephined discs of plastic compressed collagen were coated with FNC coating mix. Human corneal endothelial cells were isolated from the Descemet’ s membrane of three human donor corneas (Florida Lions Eye Bank, US) and cultured on FNC coated dishes up to passage three. Human corneal endothelial cells were then seeded onto multiple construct discs at a density of 2000-3000/mm2. Constructs were maintained in culture for 4 days before being fixed for analysis. Human corneal endothelial constructs were analysed with light microscopy, fluorescent immunochemistry and scanning and transmission electron microscopy.
Endothelial cells seeded onto collagen constructs attached overnight and were seen to have a typical hexagonal, cobblestone appearance when viewed using light microscopy. The majority of human corneal endothelial cells on collagen constructs strongly expressed ZO-1 and Na+/K+ ATPase α1 at cell-cell junctions and on the cell surface respectively. Electron microscopy revealed overlapping finger-like projections onto juxtaposed cells with tight junctions, apical microvilli and anchoring filaments into the collagen substrate.
Human corneal endothelial cells cultured on plastic compressed collagen retained their endothelial characteristics on a substrate that is less prone to the curling commonly seen with Descemet’s membrane. This method provides expanded human corneal endothelial cells with an ideal substrate for transplantation so that one donor cornea could potentially treat multiple patients requiring Descemet’s membrane endothelial keratoplasty.
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