Transcriptome Analysis of the Human Corneal Endothelium. Homo sapiens

Defining the normal and age-dependent HCEnC transcriptome will further refine our understanding of the functional roles that the endothelium plays in the cornea and will provide a basis upon which to compare transcriptomes of normal and dystrophic endothelium for the subsequent development of gene-targeted therapies. We used microarrays to comprehensively characterize human corneal endothelial cell (HCEnC) gene expression, age-dependent differential gene expression and to identify expressed genes mapped to chromosomal loci associated with the corneal endothelial dystrophies PPCD1, FECD4 and XECD Overall design: Eleven corneas from six pediatric (4, 6, 10,11, 17 and 18 years old) and five adult (53, 56, 57, 64 and 70 years old) donors, were obtained from various eye banks . Descemet and corneal endothelium were removed from donor eye bank corneas as intact sheets utilizing a technique that we have adapted for ex vivo donor tissue. Total RNA was isolated from the corneal endothelium using TriReagent and subsequently purified with the RNeasy Clean-Up Kit. The integrity of the isolated RNA was analyzed using the Agilent 2100 Electrophoresis Bioanalyzer System and found to be of sufficient quantity (approximately 1ug) and quality (RNA integrity number in the range of 8.3 to 9.0) for analysis using the Affymetrix 1.1ST gene chip arrays.