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Alexandra Pietraszkiewicz, Christopher Hampton, Ling Lei, Paola Bargagna-Mohan, Royce Mohan; Desmin is a Novel Druggable Regulator of Corneal Fibrosis. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1283. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The intermediate filaments (IFs) vimentin and desmin are coordinately upregulated in myofibroblasts during corneal fibrosis and their downregulation by the potent IF-targeting drug withaferin A (WFA) is protective against fibrosis and haze (JBC 2012, 287: 989-1006). In this study, we have employed mice deficient for desmin to unravel how desmin contributes to corneal fibrosis.
Wild-type (WT) 129Svev litter mates, desmin hemizygous (Des+/-) and homozygous (Des-/-) knockout mice were employed. Mice were anesthetized by intraperitoneal injection of ketamine and xylazine, and corneas were anesthetized with proparacain eye drop. A 1 μl drop of dilute 0.15 M NaOH was applied for 1 minute, the cornea was washed with saline solution, and the corneal and limbal epithelium was removed by scraping with a blunt Tooke corneal knife. After 7, 14 or 30 days of recovery, mouse corneas were assessed for corneal opacity and mice sacrificed. The enucleated eyes were either cryosectioned for immunohistochemistry (IHC) or corneas dissected and subjected to western blot (WB) analysis. In other studies, injured Des-/- mice were also treated with vehicle or 2 mg/kg/d WFA for 14 days.
We discovered that the injured corneas of Des-/- mice like those of WT mice lose corneal transparency and become fibrotic. Surprisingly, Des+/- corneas were protected against corneal fibrosis and revealed absence of a-smooth muscle actin and vimentin upregulation in repairing stromal cells. Recently, we showed that vimentin becomes phosphorylated at serine 38 residue (pSer38Vim) and the expression of this biomarker is correlated with corneal fibrosis (PlosOne 2015, 10(7):e0133399). Therefore, we investigated the expression of pSer38Vim in corneas of desmin-deficient mice. We discovered that the healing Des+/- corneas, unlike fibrotic Des-/- or WT corneas, show very low expression levels of pSer38Vim. The expression of pSer38vim was also altered in Des-/- corneas by treatment with WFA.
Vimentin and desmin play overlapping roles in wound repair and fibrosis, however, depending of gene dosage, the degree of desmin loss can have protective or pathological consequences. Our studies, for the first time, illuminates that the phosphorylation status of vimentin is also governed by desmin during wound repair, and that targeting of pSer38vim by WFA in desmin deficiency can prevent corneal fibrosis.
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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