Glycogene Expression in Conjunctiva of Patients with Dry Eye: Downregulation of Notch Signaling

Abstract

Purpose Glycoconjugates regulate a variety of biological events in mucosal surfaces, such as differentiation of postmitotic epithelial cells and maintenance of the wet-surfaced phenotype. This study aimed to identify the repertoire of genes (glycogenes) involved in biosynthesis of glycoconjugates in conjunctiva of normal subjects and dry eye patients. Methods RNA from conjunctival impression cytology samples was amplified and hybridized to a custom-designed glycogene microarray. Intensity data were converted to expression values and analyzed by ANOVA. Microarray data for selected Notch glycogenes were confirmed by quantitative real-time PCR. Notch receptors and ligands were immunolocalized on conjunctival biopsies by confocal microscopy. Results By microarray, 424 glycogenes were identified in normal conjunctival epithelium; galectins, glycosyltransferases, mucins, Notch signaling molecules, and proteoglycans were among the most highly expressed. In dry eye, 46 glycogenes were significantly downregulated, and included five members of the Notch signaling pathway (Notch1, -2, -3, Jagged1, Delta1), four Wnt signaling molecules (Wnt4, -5A, Frizzled6, -7), and three heparan sulfate glycotransferases (HS2ST1, HS3ST6, EXTL2). Only interferon-induced transmembrane protein 1 was upregulated. By real-time PCR, expression ratios of Notch1, -3, and Jagged1 in dry eye were 0.43, 0.56 and 0.50, respectively, compared to controls (p<0.05). Notch1, -3, and Jagged1 immunolocalized throughout the conjunctival epithelium, whereas Notch2 and Delta1 were distributed apically. Conclusions This study revealed the differential glycogene expression profiles in normal subjects and dry eye patients. Downregulation of Notch signaling in dry eye may result in abnormal differentiation of the conjunctival epithelium and have implications in the pathogenesis of the disease.

Association of Cell Surface Mucins with Galectin-3 Contributes to the Ocular Surface Epithelial Barrier

“…Saccharide inhibitors used in the current study may also release other galectins. However, galectin-3 is the most abundant galectin present in the ocular surface epithelia (ϳ80% of the total galectin content (27)), and therefore, it is reasonable to expect that the effect of the inhibitors observed in the current (21). On the other hand, the dye penetrated into cells that do not express cell surface mucins, such as corneal fibroblasts and brain astrocytes grown under the same conditions.…”

Mastermind-like transcriptional co-activator-mediated Notch signaling is indispensable for maintaining conjunctival epithelial identity

“…1C), suggesting that Notch signaling has an important role in conjunctival morphogenesis. The expression of multiple Notch family members suggests possible redundancy among different Notch ligands and receptors in the ocular surface tissues (Ma et al, 2007;Djalilian et al, 2008;Mantelli et al, 2009). Therefore, we employed a dnMaml1 overexpression strategy using triple-transgenic K14-rtTA/TC/Rosa LSL-dnMaml1 mice, in which excess dnMaml1 competes with endogenous Maml for binding to NICD/RBP-J complexes in K14-positive stratified epithelia, including the ocular surface.…”

“…It has also been suggested that Notch activation contributes to the maintenance of the stratified and non-keratinized corneal (Ma et al, 2007;Vauclair et al, 2007;Djalilian et al, 2008;Nakamura et al, 2008) and conjunctival type epithelial differentiation (Fukushima et al, 2008;Mantelli et al, 2009;Xiong et al, 2011). Notch signaling is a central mediator of short-range intercellular communication in metazoans (Kovall and Blacklow, 2010;Andersson et al, 2011).…”

“…Notch1, Notch 2, Notch 3, Jagged1 and Delta1, are significantly downregulated (Mantelli et al, 2009), albeit the specific function of the canonical Notch signaling pathway in ocular surface goblet cell differentiation remains largely unknown. We hypothesize that Notch signaling plays a pivotal role in the differentiation and maintenance of goblet cells in order to establish the conjunctival epithelial integrity necessary for ocular surface health.…”

Adhesion/growth-regulatory galectins in the human eye: localization profiles and tissue reactivities as a standard to detect disease-associated alterations

“…Another functional aspect concerns the involvement of Gal-3 in maintaining the integrity of the ocular surface epithelium by mucin cross-linking [33]. The presence of Gal-3, along with Gal-7 and Gal-8, was also indicated by microarray analysis for conjunctival epithelium [34]. Its gene belongs to the most highly expressed glycogenes, whereas Gal-7 takes a prominent place in this class, since its expression is downregulated in specimens of dryeye patients [34].…”