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Differential Gene Expression in Mouse Sclera during Ocular Development

¹From the Divisions of Ophthalmology and Genetics, Children’s Hospital of Philadelphia, ²Children’s Hospital of Philadelphia Nucleic Acid and Protein Core, and the ³University of Pennsylvania Bioinformatics Core, University of Pennsylvania, Philadelphia, Pennsylvania.

PURPOSE. Ocular development involves changes in extracellular matrix components of the scleral wall as it expands. This study was conducted to determine scleral gene expression profiles during mouse ocular development to identify genes involved in normal scleral growth.

METHODS. Sample sets of pooled sclerae of 3- and 8-week-old mice were microdissected, and total RNA was isolated. After reverse transcription, the cDNA was in vitro transcribed to produce biotin-labeled cRNA. The purified biotin-labeled cRNA samples were hybridized to microarray chips (GeneChip Mouse Genome 430 2.0; Affymetrix, Santa Clara, CA). Gene transcript expression profiles were determined, and eight differentially expressed genes between the two age groups underwent further confirmation by real-time PCR analysis.

RESULTS. Differential regulation of 4884 gene transcripts in mouse sclera with less than 5% false-discovery rate (FDR) was identified. The top 1000 with the lowest FDR among the 4884 probe sets were filtered for threefold changes between the two age groups, and 718 gene transcripts were identified. Among these 718 gene transcripts, 210 were upregulated and 508 downregulated in adult relative to juvenile mouse sclera. TGF-ß1 and several collagen genes were significantly downregulated. Microarray differential expression by real-time PCR validation of eight extracellular matrix–associated gene transcripts was confirmed.

CONCLUSIONS. This is the first study to demonstrate gene expression profiles in mouse sclera during ocular growth. These findings support the role of TGFß1 as a signaling molecule in modulating extracellular matrix during ocular development. This endeavor may be helpful in furthering understanding of how scleral remodeling is regulated during eye growth.

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