Piperine inhibits AML-12 hepatocyte EMT and LX-2 HSC activation and alleviates mouse liver fibrosis provoked by CCl4: roles in the activation of the Nrf2 cascade and subsequent suppression of the TGF-β1/Smad axis

Piperine (PIP) is an alkaloid derived from peppercorns. Herein, we assessed its effects on hepatocyte EMT and HSC activation in vitro and CCl₄-elicited liver fibrosis in mice. Further experiments were performed to unveil the molecular mechanisms underlying the hepatoprotective activity of PIP. We found that PIP inhibited TGF-β1-provoked AML-12 hepatocyte EMT and LX-2 HSC activation. Mechanistically, in AML-12 and LX-2 cells, PIP evoked Nrf2 nuclear translocation and increased transcriptions of Nrf2-responsive antioxidative genes. These events decreased TGF-β1-induced production of ROS. Moreover, PIP increased the expression of Smad7, suppressed phosphorylation and nuclear translocation of Smad2/3, and decreased the transcriptions of Smad2/3-downstream genes. Knockdown of Nrf2 abrogated the protective activity of PIP against TGF-β1. Modulatory effects of PIP on the TGF-β1/Smad cascade were also crippled, which suggested that activation of Nrf2 played critical roles in the regulatory effects of PIP on TGF-β1/Smad signaling. Experiments in vivo unveiled that PIP ameliorated mouse liver fibrosis provoked by CCl₄. PIP modulated the intrahepatic contents of the markers of EMT and HSC activation. In mouse livers, PIP activated Nrf2 signaling and reduced Smad2/3-dependent gene transcriptions. Our findings collectively suggested PIP as a new chemical entity with the capacity of alleviating liver fibrosis. The activation of the Nrf2 cascade and subsequent suppression of the TGF-β1/Smad axis are implicated in the hepatoprotective activity of PIP.