Expressed sequence tag analysis of adult human optic nerve for NEIBank: identification of cell type and tissue markers.
BACKGROUND
The optic nerve is a pure white matter central nervous system (CNS) tract with an isolated blood supply, and is widely used in physiological studies of white matter response to various insults. We examined the gene expression profile of human optic nerve (ON) and, through the NEIBANK online resource, to provide a resource of sequenced verified cDNA clones. An un-normalized cDNA library was constructed from pooled human ON tissues and was used in expressed sequence tag (EST) analysis. Location of an abundant oligodendrocyte marker was examined by immunofluorescence. Quantitative real time polymerase chain reaction (qRT-PCR) and Western analysis were used to compare levels of expression for key calcium channel protein genes and protein product in primate and rodent ON.


RESULTS
Our analyses revealed a profile similar in many respects to other white matter related tissues, but significantly different from previously available ON cDNA libraries. The previous libraries were found to include specific markers for other eye tissues, suggesting contamination. Immune/inflammatory markers were abundant in the new ON library. The oligodendrocyte marker QKI was abundant at the EST level. Immunofluorescence revealed that this protein is a useful oligodendrocyte cell-type marker in rodent and primate ONs. L-type calcium channel EST abundance was found to be particularly low. A qRT-PCR-based comparative mammalian species analysis reveals that L-type calcium channel expression levels are significantly lower in primate than in rodent ON, which may help account for the class-specific difference in responsiveness to calcium channel blocking agents. Several known eye disease genes are abundantly expressed in ON. Many genes associated with normal axonal function, mRNAs associated with axonal transport, inflammation and neuroprotection are observed.


CONCLUSIONS
We conclude that the new cDNA library is a faithful representation of human ON and EST data provide an initial overview of gene expression patterns in this tissue. The data provide clues for tissue-specific and species-specific properties of human ON that will help in design of therapeutic models.
Bernstein
Steven L
SL
Department of Ophthalmology, University of Maryland School of Medicine, Baltimore, Maryland, USA. slbernst@umaryland.edu
Guo
Yan
Y
Peterson
Katherine
K
Wistow
Graeme
G
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D001369:Axons / Q000378:metabolism*; D015152:Blotting, Northern; D015153:Blotting, Western; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D030541:Databases, Genetic; D020224:Expressed Sequence Tags / Q000378:metabolism*; D005260:Female; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D015723:Gene Library; D006801:Humans; D008251:Macaca; D008297:Male; D018613:Microscopy, Confocal; D008875:Middle Aged; D009836:Oligodendroglia / Q000378:metabolism*; D009900:Optic Nerve / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018076:DNA, Complementary; D012333:RNA, Messenger