Differentiation-inducing factor-1 (DIF-1), a morphogen produced by the cellular slime mold Dictyoste-Hum discoideum, is a natural product that has attracted considerable attention for its antitumor properties. Here, we report a novel inhibitory effect of DIF-1 on the activation of hepatic stellate cells (HSCs) responsible for liver fibrosis. DIF-1 drastically inhibited transdifferentiation of quiescent HSCs into myofibroblastic activated HSCs in a concentration-dependent manner, thus conferring an antifibrotic effect against in the liver. Neither SQ22536, an adenylate cyclase inhibitor, nor ODQ a guanylate cyclase inhibitor, showed any effect on the inhibition of HSC activation by DIF-1. In contrast, TWS119, a glycogen synthase kinase 3 beta (GSK3 beta) inhibitor, attenuated the inhibitory effect of DIF-1. Moreover, the level of inactive GSK3 beta (phosphorylated at Ser9) was significantly reduced by DIF-1. DIF-1 also inhibited nuclear translocation of beta-catenin and reduced the level of non-phospho (active)beta-catenin. These results suggest that DIF-1 inhibits HSC activation by disrupting the Wnt/beta-catenin signaling pathway through dephosphorylation of GSK3 beta. We propose that DIF-1 is a possible candidate as a therapeutic agent for preventing liver fibrosis. (C) 2019 Elsevier Inc. All rights reserved.