An Analysis Characterizing the composition of the Corneal Limbal Epithelial Stem Cell Niche (LSCN)



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Limbal epithelial stem cells (LESCs) are required for the long-term maintenance of the corneal epithelium and are required to regenerate the corneal epithelium after injury.The loss or damage to LESCs or the LSCN can lead to Limbal stem cell deficiency (LSCD). LSCD causes corneal erosions, neovascularization, and conjunctivalization of the cornea, which leads to ocular pain and vision loss. Hyaluronan (HA) is a constituent of the LSC Niche (LSCN) that is necessary for maintaining LESCs in their stem cell state. My research this summer focused on characterizing the HA-specific LSCN by identifying HA-bound proteins and proteoglycans (PGs) within the limbal region of human and porcine corneas. HA bound proteins and proteoglycans (PGs) were extracted from the central corneas and limbal region. Firstly, we dissected the corneas and used chaotropic agents and mechanical trituration to homogenize the tissues. Then, an anion exchange chromatography was used to isolate HA bound proteins and PGs. Finally, samples were desalted and concentrated.Total protein/PG extracts were analyzed by Western Blotting and agarose gel electrophoresis. Additionally, human corneas were processed for histological analysis. Inter-Alpha Inhibitor (IaI), a molecule that forms specific HA matrices with anti-inflammatory properties, was identified in our samples by both Western blotting and immunohistochemistry. Future studies will work on characterizing the role of this HA/IaI or HA/TSG-6/IaI matrix within the corneal limbus. Characterizing the role of the LSCN is vital for establishing novel mechanisms for treating LSCD