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Z. Ding, J. Dong, J. Liu, S. Deng; Unique Gene Expression in Limbus Compared to That in Cornea and Conjunctiva. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4305. doi: https://doi.org/.
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The purpose of this study is to identify the unique gene expression profile in limbus compared to its immediate adjacent structures, the cornea and conjunctiva.
The epithelium, along with 1/3 of the underlying stroma from the cornea, conjunctiva, and limbus were dissected from a single velvet monkey eye and were snap-frozen upon collection. RNA was isolated from each of these three tissue types using the Qiagen RNeasy kit. RNA quality was assessed by gel electrophoresis. Probes for hybridization were then prepared from the total RNA according to standard Affymetrix protocol and hybridized on the Affymetrix Human U133 plus 2.0 microarray chip. Genes that were preferentially (>2 fold) or exclusively (<0.5 fold) expressed in the limbus than those in both the cornea and conjunctiva were identified. EASE (Expression analysis systematic explorer software) was used to identify biological processes preferentially expressed in the limbus. Quantitative real time-PCR (QRT-PCR) was performed to confirm the expression level of representative transcripts.
There were 194 transcripts preferentially and 51 exclusively expressed in the limbus as compared to those in the cornea and the conjunctiva. Using gene ontology annotation analysis, these transcripts were classified into many biologic processes, including cell growth and maintenance, signal transduction, cell adhesion, and programmed cell death. The expression levels of six genes preferentially expressed in the limbus, WIF1, HPGD, KRT13, NTNG1, MYBPC1 and TYRP1 were confirmed by QRT-PCR.
Comparative gene profiling has identified a group of new genes that are uniquely expressed in the limbus compared to those in the cornea and conjunctiva. These genes indicate a variety of potential active biological processes that might give some insights into the limbal stem cell niche.
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