- Article
Corneal stromal fibrosis is characterized by the transition of quiescent keratocytes into α-smooth muscle actin (α-SMA)-positive myofibroblasts and excessive extracellular matrix deposition, processes largely driven by transforming growth factor-beta 1 (TGF-β1) signaling. Atractylenolide III (ATL-III), a naturally derived sesquiterpene lactone, has demonstrated antifibrotic activity in several experimental models; however, its effects on human corneal keratocytes remain unclear. In this study, we investigated the effects of ATL-III on TGF-β1-induced myofibroblast differentiation in primary human corneal keratocytes. Cells were treated with TGF-β1 (10 ng/mL) alone or in combination with ATL-III (50 or 100 µM). Cell viability, fibrosis-associated gene expression, α-SMA expression and cytoskeletal organization, and intracellular signaling were evaluated using a cell viability assay, quantitative real-time PCR, immunofluorescence staining, and Western blot analysis. ATL-III did not significantly affect cell viability at concentrations up to 120 µM after 24 h, and cell viability remained unchanged under the 48 h treatment conditions used for the phenotypic analyses. TGF-β1 markedly increased the expression of ACTA2, COL1A1, CTGF, and Fibronectin (FN1). ATL-III significantly reduced ACTA2 expression at both 50 and 100 µM and reduced COL1A1 expression at 100 µM, whereas CTGF and FN1 showed decreasing trends without statistical significance. Immunofluorescence analysis showed attenuation of TGF-β1-induced α-SMA-associated filamentous organization following ATL-III treatment. Western blot analysis further demonstrated a significant reduction in α-SMA protein expression at 100 µM ATL-III. At the signaling level, ATL-III significantly reduced TGF-β1-induced SMAD2/3 phosphorylation at 50 µM, whereas AMPK, ERK1/2, and NF-κB phosphorylation were not significantly altered by ATL-III. Notably, p38 MAPK phosphorylation was further increased at 100 µM ATL-III despite attenuation of selected myofibroblast-associated features. These findings provide preliminary evidence that ATL-III attenuates selected features of TGF-β1-induced myofibroblast differentiation in primary human corneal keratocytes. The reduction in SMAD2/3 phosphorylation observed at 50 µM was associated with, but does not establish a causal relationship with, the phenotypic changes. In addition, p38 MAPK phosphorylation was increased at 100 µM, indicating that the signaling response was not uniformly concentration-dependent. Further protein-level, pathway-specific, and in vivo studies are required to determine the potential relevance of ATL-III to pathological corneal stromal remodeling.
9 November 2026






![Reduction of branched polyethylenimine-derived carbamates to CO catalyzed by [SiCu2IIGaIII(H2O)3W9O37]9−. W: dark grey; Cu: blue; Ga: turquoise; O: red. For clarity the branched polyethylenimine is presented as a linear polymer.](https://mdpi-res.com/molecules/molecules-31-03276/article_deploy/html/images/molecules-31-03276-g001-550.jpg)



