Microarray analysis identified differentially expressed genes in keratocytes from keratoconus patients.
OBJECTIVE
To identify differentially expressed genes in human keratoconus keratocyte by cDNA microarray.


METHODS
Normal and keratoconic cornea were cultured for keratocytes. RNA was extracted. cDNA probe labeled with fluorescence dye was made from Poly A+ RNA, hybridized with microarray slide, containing 164 human apoptosis genes. Signal intensity was measured. Expression ratio between keratoconus and normal was determined using ImaGene Ver.3.0. Identified genes were further evaluated by RT-PCR and real-time PCR.


RESULTS
Five over-expressed and four under-expressed genes were identified. Of these, differential expression of tumor necrosis factor alpha-induced protein 6 (TNFAIP6), human insulin-like growth factor binding protein 5 (IGFBP5), and IGFBP3 were verified and confirmed by RT-PCR. Real-time PCR showed TNFAIP6 increased by 3.3 folds, while IGFBP5, IGFBP3 decreased by 14 and 11 folds respectively.


CONCLUSIONS
The identified genes could be important and deserve further investigation. Significant differential expression of TNFAIP6, IGFBP5, and IGFBP3 may indicate an important role of these genes in the mechanism underlying stromal thinning.
Ha
Nguyen Thanh
NT
Department of Ophthalmology, Juntendo University School of Medicine, Tokyo, Japan. hant02@yahoo.com
Nakayasu
Kiyoo
K
Murakami
Akira
A
Ishidoh
Kazumi
K
Kanai
Atsushi
A
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D017209:Apoptosis / Q000235:genetics*; D015815:Cell Adhesion Molecules / Q000235:genetics; D002478:Cells, Cultured; D003319:Corneal Stroma / Q000166:cytology*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000235:genetics; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000235:genetics; D007640:Keratoconus / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012313:RNA / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015815:Cell Adhesion Molecules; D018972:Insulin-Like Growth Factor Binding Protein 3; D018975:Insulin-Like Growth Factor Binding Protein 5; C072273:TNFAIP6 protein, human; D012313:RNA