Evaluation of global differential gene and protein expression in primary Pterygium: S100A8 and S100A9 as possible drivers of a signaling network.
OBJECTIVE
Pterygium is a wing shaped fibrovascular growth on the ocular surface, characterized by fibrosis, angiogenesis, extracellular matrix remodeling, and inflammatory infiltrates. Epidemiologic studies have linked pterygium formation to various chronic inflammatory conditions, such as ultraviolet radiation, sawdust exposure, and dry eye disease. The purpose of this study is to identify proteins that are differentially expressed in primary pterygium by using a combination of gene microarray and proteomic platforms.


METHODS
Paired pterygium and uninvolved conjunctiva tissues of four patients were evaluated for differences in global gene transcript levels using a genechip microarray. Proteins extracted from another four pairs of tissues were quantified by iTRAQ approach. Western blot and immunofluorescent staining on additional patients were used to validate dysregulated protein expression obtained from microarray and proteomics data. In addition, primary conjunctival fibroblasts were treated with recombinant S100A8, S100A9 or both. Transcript level changes of a panel of potential target genes were evaluated by real time-PCR.


RESULTS
The following were up-regulated at both protein and transcript levels S100 A8 and A9, aldehyde dehydrogenase 3 family, member1 (ALDH3A1) and vimentin (VIM). Conversely, serpin peptidase inhibitor clade A member 1 (SERPINA1) and transferrin (TF) were down-regulated. Upon adding S100A8, S100A9 or both, the inflammatory chemokine CXCL1, matrix proteins vimentin, biglycan, and gelsolin, as well as annexin-A2, thymosin-β4, chymase (CMA1), member of Ras oncogene family RAB10 and SERPINA1 were found to be up-regulated.


CONCLUSIONS
We identified 3 up-regulated and 2 down-regulated proteins by using a stringent approach comparing microarray and proteomic data. On stimulating cells with S100A8/9, a repertoire of key genes found to be up-regulated in pterygium tissue, were induced in these cells. S100A8/9 may be an upstream trigger for inflammation and other disease pathways in pterygium.
Ocular Surface Research Group, Singapore Eye Research Institute, Singapore, Singapore.
Aihua
A
Hou
Ocular Surface Research Group, Singapore Eye Research Institute, Singapore, Singapore.
Wanwen
W
Lan
Department of Biochemistry, National University of Singapore, Singapore, Singapore; Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Kai Pong
KP
Law
Ocular Surface Research Group, Singapore Eye Research Institute, Singapore, Singapore.
Ser Chin Jasmine
SC
Khoo
Ocular Surface Research Group, Singapore Eye Research Institute, Singapore, Singapore.
Min Qi
MQ
Tin
Department of Biochemistry, National University of Singapore, Singapore, Singapore; Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Yoon Pin
YP
Lim
Ocular Surface Research Group, Singapore Eye Research Institute, Singapore, Singapore; Department of Biochemistry, National University of Singapore, Singapore, Singapore; Duke-NUS Graduate Medical School, Singapore, Singapore, Singapore; Singapore National Eye Center, Singapore, Singapore; Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Louis
L
Tong
D000444:Aldehyde Dehydrogenase / Q000378:metabolism; D058578:Biglycan / Q000378:metabolism; D015153:Blotting, Western; D040501:Calgranulin A / Q000235:genetics / Q000378:metabolism*; D040502:Calgranulin B / Q000235:genetics / Q000378:metabolism*; D054360:Chemokine CXCL1 / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D018260:Gelsolin / Q000378:metabolism; D005786:Gene Expression Regulation / Q000235:genetics*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D040901:Proteomics / Q000379:methods; D011625:Pterygium / Q000235:genetics* / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics*; D014168:Transferrin / Q000378:metabolism; D014746:Vimentin / Q000378:metabolism; D000515:alpha 1-Antitrypsin / Q000378:metabolism
C546884:BGN protein, human; D058578:Biglycan; D040501:Calgranulin A; D040502:Calgranulin B; D054360:Chemokine CXCL1; D018260:Gelsolin; C043392:SERPINA1 protein, human; D014168:Transferrin; D014746:Vimentin; D000515:alpha 1-Antitrypsin; C480021:ALDH3A1 protein, human; D000444:Aldehyde Dehydrogenase