Similarities in DSG1 and KRT3 Downregulation through Retinoic Acid Treatment and PAX6 Knockdown Related Expression Profiles: Does PAX6 Affect RA Signaling in Limbal Epithelial Cells?

Lorenz Latta,Claire Zussy,Fabian Norbert Fries,Priya Katiyar,Berthold Seitz,Nóra Szentmáry,Barbara Käsmann-Kellner,Igor Knebel,Tanja Stachon,Constanze Bleil

doi:10.3390/biom11111651

Abstract

Congenital PAX6-aniridia is a rare panocular disease resulting from limbal stem cell deficiency. In PAX6-aniridia, the downregulation of the retinol-metabolizing enzymes ADH7 (All-trans-retinol dehydrogenase 7) and ALDH1A1/A3 (Retinal dehydrogenase 1, Aldehyde dehydrogenase family 1 member A3) have been described in limbal epithelial cells (LECs) and conjunctival epithelial cells. The aim of this study was to identify the role of retinol derivates in the differentiation of human LEC and its potential impact on aniridia-associated keratopathy development. Human LEC were isolated from healthy donor corneas and were cultured with retinol, retinoic acid, or pan-retinoic acid receptor antagonist (AGN 193109) acting on RARα, β, γ (NR1B1, NR1B2 NR1B3) or were cultured with pan-retinoid X receptor antagonist (UVI 3003) acting on RXR α, β, γ (retinoid X receptor, NR2B1, NR2B2, BR2B3). Using qPCR, differentiation marker and retinoid-/fatty acid metabolism-related mRNA expression was analysed. DSG1 (Desmoglein 1), KRT3 (Keratin 3), and SPINK7 (Serine Peptidase Inhibitor Kazal Type 7) mRNA expression was downregulated when retinoid derivates were used. AGN 193109 treatment led to the upregulation of ADH7, KRT3, and DSG1 mRNA expression and to the downregulation of KRT12 (Keratin 12) and KRT19 (Keratin 19) mRNA expression. Retinol and all-trans retinoic acid affect some transcripts of corneal LEC in a similar way to what has been observed in the LEC of PAX6-aniridia patients with the altered expression of differentiation markers. An elevated concentration of retinol derivatives in LEC or an altered response to retinoids may contribute to this pattern. These initial findings help to explain ocular surface epithelia differentiation disorders in PAX6-aniridia and should be investigated in patient cells or in cell models in the future in more detail.

Highlights

Since the effect of retinoids at the transcription level has not been studied on limbal epithelial cells so far, we provided the first profile of a set of transcripts to generate an idea as to what extent retinoic acid metabolism and signaling could influence AAK-related expression profiles or which transcripts are sensitive to all trans-retinoic acid (at-RA) treatment
At the corneal surface, these cells would further differentiate into mature corneal epithelial cells
Changes in differentiation marker expression are already measurable in limbal epithelial cells (LECs) cultures, and these can be useful in identifying factors influencing differentiation at the gene expression level

Summary

Introduction

Retinoids act through nuclear receptors at the transcriptional level; the nonreceptor-mediated functions of retinoids have not been extensively studied as of yet [1]. Besides their important role in development, retinoids have the capability to affect apoptosis, the differentiation and proliferation of skin epidermal cells [2], as well as to affect the apoptosis, proliferation, surface wound healing, and keratinization of corneal epithelial cells [1,3]. A severe vitamin A deficiency leads to xerophthalmia (dry eye) and keratopathy [4].

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