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Jesintha Navaratnam, Kristiane Haug, Geir V. Qvale, Jon K. Slettedal, Morten C. Moe, Liv K. Drolsum, Bjørn Nicolaissen, Aboulghassem Shahdadfar; Cell Cycle Gene Study Of Human Corneal Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):6442.
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© ARVO (1962-2015); The Authors (2016-present)
Human corneal endothelial cells (HCEC) have limited regenerative capacity in vivo (1) but proliferate in vitro under appropriate conditions (2). In the present study we examine the expression of selected cell cycle related genes in vivo and alterations in this profile induced by in vitro transfer and monolayer culture on Descemet’s membrane and on tissue culture coated plastic dishes.
Descemet’s membrane with the attached endothelium was carefully dissected from corneas in small strips. One part harvested as noncultured cells and the other cultured on Descemet's mambrane and in monolayer from each donor. All cultures were incubated for 3 weeks at 37º C with 5% humidified CO2 and medium was changed every 2-3 days. Cultured and noncultured HCEC were analyzed by qRT-PCR for genes associated to cell cycle.
mRNA analysis of genes associated with cell cycle such as KI 67, CHK2, CDC25A, CDK1, CCNA2, CCNE1 and CCNB1 by qRT-PCR show that uncultured HCEC are resting cells, but monolayer and cells cultured on Descement's membrane undergo all cell cycle phases. The studied genes associated with G1/S transition such as CHK2, CDC25A and CCNE1 are expressed in all conditions, in contrast to the expression of G2/M transition genes such as CDK1, CCNA2 and CCNB1detected in HCEC cultured on Descemet's membrane and in coated plastic dishes, but not in uncultured cells.
CDK1 activity depends on CCNA2 and CCNB1 association in mice (3) and CDK1 activity is required for entry into M phase (4). In our study, noncultured HCEC neither express associated CCNA2 and CCNB1 nor CDK1, while G1/S related genes are expressed. Our results could suggest that a subpopulation of cells in noncultured HCEC is inhibited at the G2/M transition stage of cell cycle
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