Bicuculline from Forsythia suspensa attenuates liver fibrosis via TLR4/MD2 inhibition and suppression of MyD88/NF-κB signaling.

Liver fibrosis, a cardinal feature of chronic liver injury, involves hepatic stellate cells (HSCs) activation and excessive extracellular matrix (ECM) deposition. Upon liver fibrosis, TLR4 is highly expressed on HSCs and Kupffer cells. TLR4 activation induces liver inflammation followed by exacerbated fibrosis. Inhibition of TLR4 signaling represents a validated anti-liver fibrosis therapeutic approach. In the realm of liver fibrosis treatment, traditional Chinese medicine (TCM) has demonstrated certain anti-fibrotic effects in clinical practice after long-term application. However, due to the complex composition of TCM, understanding the underlying biological mechanisms and controlling drug quality remain challenging. To address these issues, this study developed a database of potential active ingredients from TCM against liver fibrosis and conducted virtual screening based on TLR4/MD2. It is worth noting that bicuculline, a known GABAA receptor antagonist, is a newly identified MD2 ligand that confers protection against liver fibrosis via disruption of TLR4 signaling. Bicuculline exhibits prominent anti-liver fibrosis activity in activated HSCs and effectively reduces fibrosis in bile duct ligation (BDL) and CCl4-induced mouse liver fibrosis models. Concomitantly, the upregulation of MyD88, phosphorylated NF-κB (p-NF-κB), and downstream cytokines (TNF-α, IL-1β, and IL-6) in activated HSCs and liver fibrosis models could be effectively attenuated by bicuculline treatment. Molecular dynamics simulations further revealed that bicuculline binds to the LPS-binding pocket of MD2, which may inhibit TLR4 dimerization and subsequent production of pro-inflammatory factors. This study endeavors to overcome the limitations of TCM treatment and explore novel anti-liver fibrosis drugs.