Gene Expression in Human Conjunctiva and Cornea

PURPOSE. To determine global mRNA expression levels in the corneal and conjunctival epithelia and identify transcripts that exhibit preferential tissue expression. METHODS. cDNA samples derived from human conjunctival and corneal epithelia were hybridized in three independent experiments to a commercial oligonucleotide array representing > 22,000 transcripts. The resulting signal intensities and transcript present/absent calls by the primary microarray analyzing software were used in conjunction with a statistical method to identify transcripts that are preferentially or exclusively (transcripts absent in all three replicates in the other tissue) and significantly (expression >1% of the ß-actin level) expressed in one of the two tissues. EASE was used to identify biological systems comparatively over-represented in either epithelium. Immuno-, and cyto-histochemistry was performed to validate or expand on selected the results of interest. RESULTS. Our analysis identified 332 preferential and 93 exclusive significant corneal epithelial transcripts. The corresponding conjunctival epithelium values were 592 and 211. The over-represented biological processes in the cornea were related to cell adhesion and oxi-redox equilibria and cytoprotection activities. In the conjunctiva the biological processes that were most prominent were related to innate immunity and to melanogenesis. Immunohistochemistry for antigen presenting cells and melanocytes were consistent with these gene signatures. The transcript comparison identified a substantial number of genes that have either not previously been identified or not known to be highly expressed in these two epithelia, including, testican-1, ECM1, a forming isoform, CRTAC1 and NQO1 in the cornea and sPLA2-IIA, multiple MCH class II proteins, IGFBP3, and Na-Pi cotransporter type IIb in the conjunctiva. CONCLUSION. Comparative gene expression profiling led to the identification of many biological processes and previously unknown genes potentially important for the function of corneal and conjunctival epithelia. Keywords: Cell type comparison Whole human conjunctivae and human corneas were obtained from the National Disease Research Interchange (NDRI, Philadelphia, PA) and processed between 48 - 72 hr of death. No donor details apart from age, sex and cause of death were released. Criteria for inclusion were donor ages between 25 - 65 years old and overt physical integrity of the epithelium over the central cornea as revealed by a quick stain with 0.1 % Trypan blue. For each replicate experiment (Conjunctiva 1 and Cornea 1; Conjunctiva 2 and Cornea 2;Conjunctiva 3 and Cornea 3), RNA from two different corneal or conjunctival specimens was pooled and converted into cDNA. Appropriately fragmented biotin-labeled cRNA was hybridized to the HG-U133A GeneChips® (Affymetrix) oligonucleotide microarray following the manufacture protocol. Three separate experiments each including one conjunctival and one corneal cRNA pool were performed (Conjunctiva 1 and Cornea 1; Conjunctiva 2 and Cornea 2;Conjunctiva 3 and Cornea 3).