Gene expression profile of the human trabecular meshwork: NEIBank sequence tag analysis.
OBJECTIVE
To characterize the gene expression pattern in the human trabecular meshwork (TM) and identify candidate genes for glaucoma by expressed sequence tag (EST) analysis as part of the NEIBank project.


METHODS
RNA was extracted from dissected human TM and used to construct unamplified, un-normalized cDNA libraries in the pSPORT1 vector. More than 4000 clones were sequenced from the 5' end. Clones were clustered and identified using GRIST software. In addition, the expression patterns of genes encoding olfactomedin-domain proteins were analyzed by RT-PCR.


RESULTS
After non-mRNA contaminants were removed, 3459 independent TM-expressed clones were obtained. These were grouped in 1888 clusters, potentially representing individual expressed genes. Transcripts for the myocilin gene, a locus for inherited glaucoma, formed the third most abundant cluster in the TM collection, and several other genes implicated in glaucoma (PITX2, CYP1B1, and optineurin) were also represented. One abundant TM transcript was from the gene for the angiopoietin-like factor CTD6, which is located at on the long arm of chromosome 1, area 36.2-36.1 in the region of the glaucoma locus GLC3B, whereas other transcripts were from genes close to known glaucoma loci. The TM collection contains cDNAs for genes that are preferentially expressed in the lymphatic endothelium (matrix Gla protein, apolipoprotein D precursor, and selenoprotein P precursor). In addition to EST profiling, RT- PCR was used to detect transcripts of the olfactomedin-domain proteins latrotoxin receptor Lec3 and optimedin in the TM.


CONCLUSIONS
The TM libraries are a good source of molecular markers for TM and candidate genes for glaucoma. The abundance of myocilin cDNAs corresponds to the critical role of this gene in glaucoma and contrasts with libraries derived from cultured tissue. The expression profile raises the possibility that cells of the TM and Schlemm's canal may be more similar to lymphatic, rather than blood vascular endothelium.
Tomarev
Stanislav I
SI
Laboratory of Molecular and Developmental Biology, National Institutes of Health, Bethesda, Maryland 20892-2730, USA. tomarevs@nei.nih.gov
Wistow
Graeme
G
Raymond
Vincent
V
Dubois
Stéphane
S
Malyukova
Irina
I
D000368:Aged; D000369:Aged, 80 and over; D018076:DNA, Complementary / Q000235:genetics; D030541:Databases, Genetic; D020224:Expressed Sequence Tags; D005136:Eye Proteins / Q000235:genetics*; D015870:Gene Expression; D020869:Gene Expression Profiling; D015723:Gene Library; D005901:Glaucoma / Q000235:genetics*; D006801:Humans; D008875:Middle Aged; D009316:National Institutes of Health (U.S.); D009885:Ophthalmology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017421:Sequence Analysis; D016324:Sequence Tagged Sites; D014129:Trabecular Meshwork / Q000378:metabolism*; D014481:United States
D018076:DNA, Complementary; D005136:Eye Proteins; D012333:RNA, Messenger