Progenitor cells from limbal, conjunctival and oral mucosal biopsies cultivated on silicone hydrogel contact lenses share CD90, CK15, nestin, CXCR4 and SDF-1 niche markers but preserve tissue-specific characteristics and differentiation directions.

BACKGROUND: Limbal, conjunctival or oral mucosal (OM) epithelial explant cultures are used to treat limbal epithelial stem cell deficiency (LSCD), however, their characteristics remain elusive.

METHODS: Explant cultures established on silicon hydrogel contact lenses and source tissue samples were examined by immunofluorescence microscopy for limbal epithelial progenitor, corneal differentiation, and putative limbal niche markers. Quantitative analysis was done using confocal images of whole slides.

RESULTS: In all source tissue samples, many p63+ and some vimentin + basal epithelial as well as CD90+ stromal cells were seen. All epithelia expressed CXCR4. While SDF-1 was mostly in the epithelium and only weakly in the stroma of the limbus and conjunctiva, it was restricted to the stroma in OM. In all explant cultures, vimentin+/p63α+ small cells were abundant. For limbal explants, their ratio was highest at the source (68%) and decreased significantly with distance indicating highest regenerative and differentiation potential. OM showed the lowest ratio (38%) and no change by distance, while conjunctival explants were in the interim range. The density of both corneal-type CK3+ and less differentiated CK19+ epithelial cells increased with distance from the source for limbal and conjunctival samples (R2:0.3-0.44, p ≤ 0.05) but did not change for OM (R2 < 0.02, p > 0.47). In addition to the dominance of source-specific cytokeratin patterns, all CKs of mature stratified epithelium could be detected, while CK14 and CK15 evidenced the occurrence of less mature epithelial cells. Spatial maturation gradients were only characteristic of limbal sources. Some CD90+ putative niche cells, surrounded by CK15+ and nestin + cells formed distinct clusters in all samples. SDF-1 and CXCR4 were present in all cultures, and their distribution patterns showed similarities with the tissue of origin.

CONCLUSION: Despite similarities, including putative niche-associated cell clusters, explant cultures on silicon hydrogel surfaces from all three epithelia maintain characteristics of their original source. Their SDF-1 and CXCR4 content indicates the ability to establish contact with the host limbal niche that may be a prerequisite for long-term survival and site-specific differentiation after transplantation. Limbal explants show the highest proliferative and differentiation potential, while the generation of corneal epithelial-like cells is increasingly less efficient from conjunctival and OM sources.