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Oliver J. Baylis, Francisco C. Figueiredo, Majlinda Lako, Sajjad Ahmad; Cadaveric Limbal Epithelial Outgrowth Is Delayed By Increased Organ Culture Times. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3502.
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To evaluate the epithelial outgrowth characteristics of limbal explants cultured on human amniotic membranes, and to correlate these factors with donor details, including donor age and time between death and limbal epithelial culture, in order to improve limbal culture outcomes.
Cadaveric corneoscleral rings in organ culture medium were provided by the UK Eye Banks. Donor details recorded included age, sex, time between death and enucleation, time between enucleation and organ culture and the time spent in organ culture. Ten limbal explants from different donors were cultured on cryopreserved human amniotic membranes in limbal epithelial culture medium, and the cultures were fed every 2-3 days. The epithelial outgrowth from the limbal explants was measured at each feed. The growth characteristics from the explants were assessed and correlations with donor tissue factors were analysed statistically using Kendall’s tau-b test.
The mean donor age was 63.8 years (43 to 90 years). The epithelial outgrowths consisted of a flat lag phase followed by a steeper linear phase. The lag phase duration and the gradient of the linear phase (i.e. the growth rate) were recorded and used for the correlation analysis. The mean lag time was 10.8 days (6.8 to 15.9 days) and the mean growth rate was 5.1 mm2/day (0.6 to 8.4 mm2/day). The lag phase was significantly correlated with the time the tissue spent in organ culture (p=0.003). However, it was not correlated with any other donor variables, and of particular note not to donor age. The growth rate was not correlated with any of the donor variables studied.
The results suggest that increased duration of organ culture delays epithelial outgrowth from limbal explant cultures. Interestingly, although a wide range of lag times and growth rates were observed, the other donor variables studied did not appear to significantly influence the growth, although this needs to be confirmed with a larger sample size. Autologous explant cultures are used in the treatment of limbal stem cell deficiency and in some cases organ-cultured limbal tissue may be used as allogeneic donor material. Standard organ culture conditions are designed to maintain corneal endothelial health; this study indicates that limbal and epithelial health should also be monitored if this tissue is to be used for limbal culture. Further work is also needed to identify and optimise other factors that affect limbal explant cultures in order to improve patient outcomes.
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