Ent-abietane diterpenoid Enantiomers from Euphorbia helioscopia inhibits liver fibrosis via suppressing the TGF-β1/smad pathway

Abstract

Despite abietane diterpenoids exhibit various biological activities, their anti-fibrotic activity has rarely been investigated. In the study, 19 diterpenoids (twelve abietanes and seven other types) were isolated from E. helioscopia , comprising five undescribed ent -abietane type diterpenoids ( 1a , 1b , and 2−4 ). Among them, (+)-euphonane A ( 1a ) and (−)-euphonane A ( 1b ) represent previously unreported ent -abietane enantiomers. The structures of unreported diterpenes were mainly assigned through comprehensive nuclear magnetic resonance (NMR) data analysis, in combination with computational techniques involving NMR and electronic circular dichroism (ECD) calculations. In addition, single crystal diffraction analysis precisely defined the absolute stereochemistry of euphonane C ( 3 ). The effect of these 19 diterpenoids to inhibit fibronectin (FN) expression was evaluated in transforming growth factor-β (TGF-β1)-stimulated human hepatic stellate cell line (LX-2 cells), and three compounds ( 1a , 1b , and 3 ) showed significant inhibitory activity at micromolar level, being stronger than the positive control silymarin. A dose-dependent suppression of fibrotic marker expression, particularly that of FN, collagen I, and α-smooth muscle actin (α-SMA), was observed upon the treatment with compounds 1a and 1b . Mechanistic investigations, including transcriptomics analysis and western blotting, revealed that the anti-fibrotic action of 1b in hepatic cells is probably mediated through the TGF-β/Smad signaling cascade. The above results both extend our knowledge regarding the bioactivities of diterpenes isolated from E. helioscopia , and position abietane-type diterpenoids as prospective molecular structures for the innovation of new therapeutics against liver fibrosis.