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S. Kolli, S. Ahmad, M. Lako, F. Figueiredo; The Comparison of Limbal Stem Cell Properties of Organ Cultured versus Fresh Corneas. Invest. Ophthalmol. Vis. Sci. 2007;48(13):461.
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© ARVO (1962-2015); The Authors (2016-present)
Organ culture storage techniques for corneas have improved the donor quality of corneas for transplantation and allow for longer storage times. However, the effect of organ culture storage techniques on the viability of limbal stem cells in corneoscleral tissue is less well studied. Limbal stem cell transplantation (either whole tissue or ex-vivo expanded limbal tissue) forms the basis of treatment for significant limbal stem cell deficiency. The aim of this study was to measure and compare the stem cell properties of limbal tissue which had been stored in organ culture with fresh limbal tissue that had been obtained from cadaveric donors.
Limbal epithelial stem cells (LESCs) were obtained from either fresh limbal tissue (eyes donated for research purposes) or organ culture stored limbal tissue (left over corneoscleral rings supplied by UK Transplant for clinical corneal transplantation and research) of comparable donor age. These LESCs were co-cultured on 3T3-J2 mouse fibroblast feeder cells. Near-confluent colonies were obtained after 1 week. The LESCs from the colonies of these two groups were then analysed for the expression of p63 (a putative marker of limbal stem cells) using both real time reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry. In addition, the colony forming efficiencies (CFEs) of both groups of cultured cells were measured.
The expression of p63 was significantly reduced in organ culture stored limbal tissue compared with fresh limbal tissue as measured by both RT-PCR (p<0.05; n=3) and flow cytometry (0<0.0005; n=3). In addition, the CFEs were significantly lower (p<0.05; n=3) with organ culture stored limbal tissue (20%) than with fresh limbal tissue (30%).
This study indicates that the limbal stem cell properties of organ cultured limbal tissue are lower compared with fresh limbal tissue. This has important implications for the treatment of limbal stem cell deficiency with limbal stem cell transplants. There is a need to consider the use of fresh corneal tissue for these transplants and to look at ways of improving limbal stem cell properties of stored corneal tissue.
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