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Loic Blanchon, Aureli Comptour, Marion Rouzaire, corinne Belville, frederic chiambaretta, Vincent Sapin; The cornea wound healing properties of retinoic acid passed by the action of the LOXL4 protein. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1250.
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© ARVO (1962-2015); The Authors (2016-present)
Epithelial wound healing is a multistep mechanism implying a combination of molecular and cellular events. Following alkali burn traumatisms, cell migration, proliferation and differentiation were demonstrated to be required to recover an intact corneal epithelium and a good visual acuity. Already studied for its pro-healing properties, vitamin A and its active derivatives (i.e. retinoic acid / RA) appear to be good candidates to better understand the cornea wound healing. After having previously demonstrated in-vitro and in-vivo the effect of RA on wound corneal epithelium by acting on cell migration, we studied here the lysyl oxidase (LOX) gene family regulated by RA and the importance of such proteins for corneal epithelium recovery
HCE cells were treated by RA for LOX family members’ characterization and induction by PCR /qPCR and immunocytochemistry. Promotology experiments were done to study LOX(s) induction by RA and the RAR (Retinoic Acid Receptor) isoforms implied. Scratch assays were done after RA treatment in combination with βAPN (β-aminopropionytrile, a LOX family enzymatic inhibitor) or after transfection with siRNA against LOX family members. Cell proliferation and migration were determined after scratch assay. The right eye of 7 male CD1 per group was burned with NaOH and treated with RA and w/o βAPN 6 times/day (d) during 7d. To evaluate wound healing, wounds areas were quantified from slit lamp photographs at 0 and 7d using imageJ
Among the 5 members of LOX family, only LOXL4 expression (mRNA and protein levels) is increased in HCE treated with RA from 12 to 60 hours. This regulation passed through the RARα/RXRα heterodimer fixation on a RARE-DR5 type (Retinoic Acid Response Element) located in the LOXL4 promoter. In the in-vitro model, the use of βAPN or siRNA directed against LOXL4 showed a decrease in the wound healing by inhibiting cell migration promoted by RA treatment. In the in-vivo model the use of βAPN during RA treatment also inhibits such positive action on wound healing
We confirmed the importance of RA in cornea epithelium wound repair. Then, we demonstrated that LOXL4, a gene implied in the ECM dynamic and induced by RA, is essential for such phenomenon. This study is one of the first demonstrating a direct link between vitamin A regulated genes and cornea wound healing and could constitute a cornerstone of future clinical therapies
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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