Limbal stem cell (LSC) deficiency is one of the leading causes of corneal blindness worldwide. LSCs are a population of multipotent cells found in the corneal limbus that regenerate the corneal epithelium. Excessive loss of these cells from various surface diseases results in corneal scarring and ulceration. Transplantation of cultured donor LSCs is one of the few viable treatments for bilateral disease. However, immune rejection of the allogeneic donor cells limits the long-term outcomes of this procedure. Induced pluripotent stem cell (iPSC) technology offers a potential solution. Using iPSC technology, autologous LSCs reprogrammed from a patient’s own somatic calls can be used as an alternative to donor cells, eliminating any risk of rejection. However, more work is still needed to establish the efficacy of existing protocols for differentiating iPSC into LSC. Putative LSC markers are frequently used to compare iPSC-derived LSC (iPSC-LSC) to donor LSC in order to establish the efficacy of various differentiation protocols; but the questionable specificity of these markers for authentic LSC raises concerns about this method. In order to facilitate translation of this technology into clinical practice, more rigorous phenotypic and functional comparison of iPSC-LSC to primary donor LSC is warranted.
The first aim of this research is to perform a phenotypic comparison of iPSC-LSCs to primary donor LSCs using RNA-sequencing for whole transcriptome comparison. The second aim is to perform a functional comparison of iPSC-LSC to primary donor LSC using an organ culture model of LSC deficiency. Phenotypic concordance between primary and iPSC-LSC will help validate current differentiation protocols. Phenotypic discordance will provide insight into gene pathways that define LSC identity and may be good targets for further protocol refinement. Evaluating the functional behavior of iPSC-LSCs will help establish whether the cells obtained from current protocols are appropriate for clinical use.