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Fadhilah Zainal Abidin, Dimitrios Karamichos, Francisco C Figueiredo, Che John Connon; The effects of retinoic acid on human keratoconic fibroblast in vitro under serum-free conditions. Invest. Ophthalmol. Vis. Sci. 201657(12):.
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© ARVO (1962-2015); The Authors (2016-present)
Keratoconus is a relatively common, often bilateral, non-inflammatory and progressive corneal ectasia. The aetiology of this condition is complex and multifactorial. Stromal thinning is associated with significant reduction of fibrillar collagens and proteoglycans within the matrix organization. Our previous studies on normal human corneal keratocytes have shown that retinoic acid (RA) supplementation in serum-free medium is able to enhance both collagens and keratocyte-characteristic proteoglycans productions both in 2D and 3D environment. The purpose of this study was to investigate the effects of RA on the phenotype of keratoconic fibroblast maintained in vitro under serum free conditions.
Keratoconic fibroblasts (KF) isolated from human keratoconic corneas (n=3) were cultured in serum containing medium until passage 5. The cells then underwent serum-starvation for 3 days, and later divided into different groups: 1) serum containing medium, 2) serum-free medium with DMSO vehicle, and 3) serum-free media supplemented with RA (1, 5, 10, 15 and 25 µM) for culture up to 14 days. The effect of RA on cell proliferation and morphology were evaluated. In addition, the expressions of keratocyte markers as well as matrix metalloprotease were measured using quantitative PCR.
The RA did not significantly enhance cell proliferation at lower concentrations but did affect the morphology of the KF cells. More KF cells with dendritic morphology appeared in cultures supplemented with RA at 5 and 10 µM within the first week of culture as compared to spindle-shaped KF cells in media not containing RA. KF cultured in lower RA concentrations also expressed significantly higher (P ≤ 0.05) keratocyte-characteristic proteoglycans, such as keratocan, lumican and decorin, while significantly reducing the expression of matrix metalloprotease 1.
These results suggest that the supplementation of RA at low concentrations appears to promote keratoconic fibroblasts into a more non-diseased keratocytes in terms of morphology and keratocyte marker expression, and may be of potential use in the treatment of keratoconus.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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