Gene-expression analysis of polyI:C-stimulated primary human conjunctival epithelial cells.
BACKGROUND
The authors previously reported that human ocular surface epithelium expressed TLR3 and that its ligand polyI:C stimulated the secretion of IL-6, IL-8 and IFN-β. In this study, to examine comprehensive effects of polyI:C stimulation of primary human conjunctival epithelial cells (PHCjECs), the authors performed a gene-expression analysis of the polyI:C-stimulated PHCjECs using oligonucleotide microarrays, GeneChip.


METHODS
The transcripts upregulated upon polyI:C stimulation in PHCjECs from two individuals were examined using GeneChip. Eleven new upregulated transcripts of interest were confirmed by quantitative real-time PCR (RT-PCR), and seven proteins produced by those transcripts were examined by ELISA or immunoblot analysis in PHCjECs from three other individuals, respectively.


RESULTS
GeneChip analysis showed that 150 transcripts were upregulated more than threefold and that 47 transcripts were upregulated more than 10-fold upon polyI:C stimulation in the PHCjECs. Eleven of the 47 upregulated transcripts (CXCL11, RIG-I, IL28A, CXCL10, CCL5, CCL4, MDA5, IL7R, TSLP, CCL20 and ICAM-1) were significantly upregulated upon polyI:C stimulation by quantitative RT-PCR, and the levels of seven proteins of the transcripts CXCL11, CXCL10, CCL5, CCL20, TSLP, RIG-I and MDA5 were confirmed by ELISA or immunoblot analysis to increase significantly in polyI:C-stimulated PHCjECs.


CONCLUSIONS
Our results might show that TLR3 of conjunctival epithelium could not only induce antiviral innate immune responses but also regulate the allergic reactions.
Ueta
Mayumi
M
Research Center for Inﬂammation and Regenerative Medicine, Faculty of Life and Medical Sciences, Doshisha University, Doshisha, Kyoto, Japan. mueta@koto.kpu-m.ac.jp.
Mizushima
Katsura
K
Yokoi
Norihiko
N
Naito
Yuji
Y
Kinoshita
Shigeru
S
D003228:Conjunctiva / Q000378:metabolism*; D016207:Cytokines / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells; D015870:Gene Expression; D006801:Humans; D007369:Interferon Inducers / Q000494:pharmacology*; D008565:Membrane Proteins / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D011070:Poly I-C / Q000494:pharmacology*; D015854:Up-Regulation
D016207:Cytokines; D007369:Interferon Inducers; D008565:Membrane Proteins; D011070:Poly I-C