Purchase this article with an account.
MK Wirtz, JR Samples, H Xu, T Severson, TS Acott; Expression Profile of Active Genes in Human Trabecular Meshwork Cells . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1021.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: This report describes an expression profile of 1117 genes in human trabecular meshwork cells (HTM) obtained by analyzing a cDNA library prepared from a nonamplified mRNA population. Methods: HTM cell cultures were established from 6 young donors (ages 2 weeks to 2 years). Eyes were obtained from the Oregon Lion's Eye Bank (Portland, OR) and protocols were approved by the institutional human subjects committee and conformed to the Declaration of Helsinki. Cells grown in Dulbelco's media supplemented with 10% fetal calf serum were harvested at passages 3 through 6. The frozen cells (∼96 million)were shipped on dry ice to Invitrogen (Carlsbad, CA) where a unidirectional cDNA library was constructed using a pcDNA3 vector and TOP10F' host cells. The size of the inserts ranged from 0.5 to 1.6 Kb with the average size being 0.85 Kb. Plasmid purifications were performed using the Mini-prep 24HTM from McConnell Research (San Diego, CA) or the Perfectprep Plasmid Mini Kit from Eppendorf (Westbury, NY). The purified plasmid preps were then sequenced at OHSU core facilities. Nucleotide searches were performed using BLASTN (version 2.1.3) at the National Center for Biotechnology Information against the nonredundant nucleic acid sequence and dbEST databases. Results: A total of 1117 ESTs from this nonamplified cDNA library were categorized to determine the fidelity of the library. 877 ESTs (78.7%) were known genes. 139 ESTs (12.5%) showed close identity to EST sequences reported in the public domain database (dbEST). 13 ESTs (1.2%) showed no significant similarity to known genes or ESTs in the public databases and were thus defined as novel ESTs. The most abundant genes expressed by the HTM included ferritin H, eukaryotic translongation factor alpha 1, ferritin L, fibronectin and TIMP-1. Ferritin H was the most abundant transcript making up over 4% of the genes expressed by the HTM. Extracellular matrix proteins were also highly expressed. Conclusion: A gene expression profile of a young HTM cDNA library supports previous work documenting HTM gene expression, including finding high levels of TIMP-1 and fibronectin. An unexpected finding was the high levels of the two subunits of ferritin, H and L, which may be important in mediating stress in the HTM.≷
This PDF is available to Subscribers Only