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Jin San Choi, Matthew Giegengack, Eun Young Kim, Min Jeong Kim, Ralph D'Agostino, Gilson Khang, Shay Soker; Success Isolation of Human Corneal Endothelial Cells for Clinical Use. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1647.
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© ARVO (1962-2015); The Authors (2016-present)
Corneal transplantation is a common transplant procedure to improve visual acuity by replacing the opaque or distorted host tissue by clear healthy donor tissue. However, its clinical utility is limited due to a lack of donor supply with high quality corneas. Bioengineered neo-corneas, created using an expandable population of human donor-derived corneal endothelial cells (HCECs), could address this shortage. Thus, the objective of this study was to evaluate HCEC sourcing with various isolation methods, including enzymatic digestion, culture medium components, and adhesive proteins.
HCECs were obtained from corneas with various aged donors after endothelial keratoplasty. Under a dissection microscope, the Descemet’s membrane, including the attached corneal endothelium was stripped from the stroma and the cells were isolated and expanded by explant culture and the use of enzymatic digestion with enzyme such as collagenase II, dispase, or trypsin. In order to improve the initial cell attachment, tissue culture plates were coated with collagen IV, fibronectin, or fibronectin-collagen combination coating mix (FNC) before cell plating.
In results, HCECs were successfully isolated from 32% (86/269) of donor corneas. Donor age and isolation method influenced the characteristics of the resulting in vitro HCEC culture. Under all conditions tested, FNC-coated plates showed higher quality cultures than other coatings tested.
The results suggest that donor and isolation method characteristics are the two factors that most directly affect the quality of the resulting HCEC in vitro culture. These factors should guide the clinical expansion of HCECs for the generation of bioengineered neo-corneas.
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