Effect of retinoic acid on gene expression in human conjunctival epithelium: secretory phospholipase A2 mediates retinoic acid induction of MUC16.

Abstract:

PURPOSE:How vitamin A contributes to the maintenance of the wet-surfaced phenotype at the ocular surface is not well understood. This study sought to identify vitamin A-responsive genes in ocular surface epithelia using gene microarray analysis of cultures of a human conjunctival epithelial (HCjE) cell line grown with all-trans-retinoic acid (RA). The analysis showed that secretory phospholipase A(2) group IIA (sPLA(2)-IIA) was the gene most upregulated by RA, followed by the membrane-associated mucin MUC16 at a later time point. Since eicosanoids, the product of arachidonic acid generated by the PLA(2) family, have been shown to increase mucin production, this study sought to determine whether sPLA(2) mediates the RA induction of MUC16. METHODS:HCjE cells were cultured with or without RA for 3, 6, 24, and 48 hours. Complementary RNA prepared from RNA of the HCjE cells was hybridized to human gene chips and analyzed using commercial software. Microarray data on mucin expression were validated by real-time PCR. To investigate whether sPLA(2) is associated with RA-induced MUC16 upregulation, HCjE cells were incubated with RA and the broad-spectrum PLA(2) inhibitor aristolochic acid (ArA) or the specific sPLA(2)-IIA inhibitor LY315920, followed by analysis of MUC16 mRNA and protein by real-time PCR and Western blot analysis. RESULTS:After RA addition, 28 transcripts were upregulated and 6 downregulated by more than twofold (P < 0.01) at both 3 and 6 hours (early phase). Eighty gene transcripts were upregulated and 45 downregulated at both 24 and 48 hours (late phase). Group IIA sPLA(2), significantly upregulated by 24 hours, and MUC16 were the most upregulated RNAs by RA at 48 hours. sPLA(2) upregulation by RA was confirmed by Western blot analysis. When HCjE cells were incubated with RA plus ArA or specific inhibitor of sPLA(2)-IIA, LY315920, the RA-induced MUC16 mRNA was significantly reduced (P < 0.01). CONCLUSIONS:The RA-associated upregulation of membrane-associated mucin MUC16 at late phase appears to be through sPLA(2)-IIA. Upregulation of this hydrophilic membrane-associated mucin may be one of the important mechanisms by which vitamin A facilitates maintenance of the wet-surfaced phenotype on the ocular surface.

authors

Hori Y,Spurr-Michaud SJ,Russo CL,Argüeso P,Gipson IK

doi

10.1167/iovs.05-0627

keywords:

subject

Has Abstract

pub_date

2005-11-01 00:00:00

pages

4050-61

issue

11

eissn

0146-0404

issn

1552-5783

pii

46/11/4050

journal_volume

46

pub_type

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