UCLA

Purpose

To elucidate the unique molecular factors and biological processes that are differentially expressed in the limbal stem cell microenvironment by comparing directly to that of its immediate adjacent structures, the cornea and conjunctiva.

Methods

Total RNA was isolated and amplified from the limbus, cornea, and conjunctiva. A gene expression profile of each tissue type was obtained by using microarray technique. The transcripts in which the expression level was at least twofold higher than that in the other two tissue types were identified. The expression levels of selected genes were confirmed by quantitative reverse transcription polymerase chain reaction (QRT-PCR). Protein expression of selected genes were confirmed by an immunohistochemistry study in normal human ocular tissue.

Results

There were 186 preferentially overexpressed transcripts in the limbus in direct comparison to that in the cornea and conjunctiva. Many signature genes in the cornea and conjunctiva were among the preferentially expressed transcripts obtained by the microarray data. In addition, a significant number of new genes were identified, and the expression level of all nine selected genes was verified by QRT-PCR. Protein expression levels of keratin 13, tenascin c, homeodomain only protein (HOP), and TP53 apoptosis effector (PERP) were confirmed in human ocular tissues. Functional analysis of the preferentially expressed genes in the limbus reviewed that melanin metabolism and cell-cell adhesion were among the noticeable biological processes. Chromosomal distribution of the overexpressed genes in the limbus was disproportional to that of all known human genes.

Conclusions

These findings may shed light on the unique molecular components and regulation of limbal stem cells and their niche.