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Gregory S. Schultz, Yuanjing Liu, Daniel Gibson, Paulette Robinson, Sonal Tuli; Histone Deacetylase Inhihitor SAHA Inhibits TGFb-Induced Transformation of Rabbit Corneal Fibroblasts to Myofibroblast. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2537.
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To assess the effects of the histone deacetylase inhibitor, suberoylanilide hydroxymic acid (SAHA) on TGFb-induced differentiation of cultures of rabbit corneal fibroblasts into myofibroblasts.
Early passage (≤3) of rabbit corneal fibroblasts (RCF) were grown to ~80% confluence in chemically defined medium (DMEM) supplemented with 10% FBS then cultured under serum free medium for an additional 2 days. RCF were then cultured for 48 hours in DMEM alone or supplemented with TGFβ1 (1 ng/ml) and SAHA in doses of 0, 1.25, 2.5, 5, 7.5, or 10 µM. Cell viability was measured by MTS assay. Transformation of rabbit corneal fibroblasts into myofibroblasts was evaluated by immunocytochemical staining of a-smooth muscle actin (SMA) protein, a marker of myofibroblasts.
SAHA concentrations of 10 µM or less were noncytotoxic and did not alter normal RCF morphology. RCF treated with TGFβ1 had approximately an increase of 80% SMA immunostaining levels when compared to control RCF maintained in DMEM. The combination of SAHA and TGFβ1 treatment of RCF reduced the intensity of SMA immunostaining compared to TGFβ1 alone at all levels of SAHA tested. Addition of SAHA at 10 µM reduced SMA protein staining intensity to levels comparable to SMA staining observed in control cells without TGFβ1 treatment. Cell viability of RCF incubated with SAHA at all concentrations tested was not different than control RCF cultured in DMEM or RCF cultured in TGFβ1.
At non-toxic levels, SAHA inhibited TGFβ1-induced myofibroblast transformation of rabbit corneal fibroblasts in vitro.
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