Chromone scaffold–mediated reprogramming of the epithelial–mesenchymal transition prevents fibrosis

Abstract

Fibrosis is an intractable human disease for which no curative drugs have yet been developed.Recent work revealed that renal fibrosis is a manifestation of the epithelial-mesenchymal transition (EMT). Chromones and chromone derivatives (CDs) are compounds based on a chemical scaffold found ubiquitously throughout the plant kingdom. We recently showed that eupatilin, a CD from an Artemisia species, efficiently shuts down osteoclastogenesis, concomitant with global downregulation of EMT-associated genes. For this study, we established the mouse hepatic stellate cell (HSC) line ONGHEPA1.Real-time imaging revealed that upon stimulation with TGF, ONGHEPA1 cells uniformly differentiated into myofibroblasts, but fibrosis was completely inhibited by 50 M eupatilin. Concomitant with these anti-fibrotic effects, multiple EMT-inducible genes, such as Col11a1, periostin and Slit3, were significantly downregulated. Although pretreatment with eupatilin did not affect the antifibrotic effect by TGF, stimulation with eupatilin following pretreatment with TGF completely blocked fibrosis, suggesting that the anti-fibrotic capacity of eupatilin is tightly coupled to TGF signaling components or TGF-inducible molecules. This observation led us to hypothesize that eupatilin could reverse fibrosis in ONGHEPA1 cells by reprogramming the EMT. When ONGHEPA1 cells were sequentially stimulated and differentiated into fibroblasts with TGF and eupatilin or selected CDs, we observed dramatic morphological changes along with reversal of the EMT. RNA-Seq analyses and interactome studies revealed that multiple genes involved in the EMT were induced by TGF, but repressed by eupatilin. We identified 103 genes whose expression reached the highest levels following stimulation with TGF, but dropped to basal levels in cells treated with eupatilin. The majority of these genes were EMTassociated. A defined set of 34 CDs was tested in the fibrosis assay, leading to the not peer-reviewed)