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S-E Ti, D Anderson, A Touhami, C Kim, SC G Tseng; Factors Affecting Outcome Following Transplantation of Ex Vivo Expanded Limbal Epithelium on Amniotic Membrane for Total Limbal Deficiency in Rabbits . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4188.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To determine factors affecting the outcome of corneal surface reconstruction in rabbits with total limbal stem cell deficiency (LSCD) using autologous limbal epithelial stem cells (LSC) ex vivo expanded on rabbit amniotic membrane (AM). Methods: Left eyes of 52 rabbits were rendered total LSCD by n-haptanol debridement of the entire corneal epithelium followed by surgical removal of 360º limbal rim. After cytological verification of LSCD, the fibrovascular pannus of each cornea was removed. Group I (n=10) received rabbit AM transplantation, while Groups II-IV (n=42) underwent transplantation of LSC cultured on rabbit AM (LSC-AM graft) derived from a small limbal biopsy taken from the right eye. Clinical outcome was graded as "success" if a smooth, avascular corneal surface was restored, "partial success" if more than two quadrants of corneal surface were smooth, or "failure" if the corneal surface was revascularized and irregular. Results: A long-term follow-up of more than one year was achieved. Compared to 100% failure rate in Group I, inclusion of expanded LSC resulted in variable success rates in Groups II-IV. Kaplan-Meier survival analysis showed that different suturing techniques, subconjunctival injection of long-acting steroid, and tarsorrhaphy used in Groups II (n=17) and III (n=13) did not significantly alter the outcome. However, the use of a larger graft and human AM as a temporary patch with a retained explant for one week in Group IV (n=12) significantly improved the success rate (P<0.002). Among eyes showing clinical failure, there was a significant correlation between the logarithm of the first day when an epithelial defect was noted and the time of graft failure (r2 = 0.60, P<0.001). Furthermore, the presence of severe lid deformity was borderline significant when correlated with failure cases in all four groups (P=0.069). Conclusion: Ex vivo expansion of LSC can be achieved using rabbit AM culture in rabbits. Such expanded LSC can successfully reconstruct corneal surfaces with total LSCD. This animal model is useful to investigate cultivating variables affecting epithelial stemness so that surgical reconstruction of corneas with total LSCD can be successfully performed. This model can also be used to test the feasibility of gene therapies targeted at LSC in the future.
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