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T. Senoo, S. Ishimaru, N.C. Joyce, M. Kikuchi, K. Chiba, K. Hasegawa, Y. Obara; Proliferative Capability of Endothelial Cells Derived from Human Bullous Keratopathy Patients . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4725.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The purpose of this ex vivo study was to determine the ability of endothelial cells from human bullous keratopathy patients to progress through the cell cycle upon release from cell-cell contact inhibition. Methods: Corneas were obtained from 9 patients with bullous keratopathy (BK) (mean: 69.2±11.8 yo) and from 2 normal donors (mean: 48.2±5.8 yo). Corneas were cut in quarters, incubated for 24 hrs in medium containing 10% FBS and 10ng/ml EGF, and then incubated with 0.2 mg/ml EDTA in HBSS, pH 7.4, for 30 min. One quarter was returned to the same medium. The other 3 quarters were cultured for 48 hrs in the same medium, but with anti-N-cadherin (10 ng/ml) to release cell-cell contacts. Corneal quarters were stained with propidium iodide (to stain all nuclei) and immunostained for one of the following: Ki67 (a marker of actively cycling cells), ZO-1 (a tight junction marker), or cell cycle-associated proteins (cyclin D, cyclin E, E2F, pRb, PD-1). Proliferative activity and the state of contact inhibition was compared in normal and BK human corneal endothelial cells (HCEC) using previously described methods (IOVS, 2000;41:2930). The clinical history of each BK patient was reviewed and compared with the study results. Results: Cell cycle progression, as indicated by Ki67 positive staining, was detected in only 2 of 9 BK corneas compared with 2 normal controls. Patients whose HCEC initiated proliferation had developed BK due to complications after cataract surgery and underwent full thickness corneal transplantation (PKP) relatively early. In cases where no proliferative activity was observed, follow-up observations were made over a long period of time after PBK, or retransplantation was done. No significant differences in the degree of pre-transplantation BK, age, or intraoperative condition were found. Conclusions: As more is learned about the proliferative process of HCEC, it should become possible to control its regeneration. HCEC in some patients with BK can be induced to proliferate. However, we suggest that it is necessary to start treatment before BK is fully developed rather than to try and induce proliferation in heavily damaged HCEC.
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