OBJECTIVE To parameterize the theory of “Ro Nus Zhurjes”, assign weight values to medicinal flavors, construct a weighted network, combine AlphaFold2-based reverse docking, molecular dynamics simulation and in vitro experiments, and explore the material basis and molecular mechanism of Jingzhu Gantaishu in the treatment of hepatitis.

METHODS Chemical constituents of Jingzhu Gantaishu were collected through literature retrieval and the TCMSP database, and their potential targets were predicted using SwissTargetPrediction. Hepatitis-related targets were retrieved from OMIM, GeneCards, DrugBank, and TTD databases. The Tibetan medicine theory of “Ro Nus Zhurjes” was parameterized and assigned medicinal flavor weights, and a “medicinal flavor-component-target” weighted network was constructed using the adjacency matrix method to intuitively correlate components with targets and displayed multi-component, multi-target synergistic characteristics. Based on this network, core components and core targets with Degree values exceeding the median were screened and subjected to AlphaFold2-based reverse docking with hepatitis targets. The representative component with the best binding capacity was identified, and molecular dynamics simulations were performed to explore the dynamic interaction mechanisms between ingredients and targets. In vitro verification adopted the CCK-8 method to detect cell viability, utilized Hoechst staining and flow cytometry to detect cell apoptosis, and applied the JC-10 staining method to detect mitochondrial membrane potential.

RESULTS The weighted network clearly illustrated the many-to-many correlations between constituents and targets of Jingzhu Gantaishu, uncovered the synergistic characteristics of this compound prescription with multiple components acting on multiple targets, and accordingly screened 80 key potential active chemical constituents and 80 key targets whose Degree values were higher than the median. Reverse docking revealed that glycyrol obtained the highest docking score with stearoyl-CoA desaturase(SCD), serving as the core representative component with the strongest binding affinity. It stably inserted into the active pocket of SCD via hydrogen bonds, π-π interactions and hydrophobic interactions. Molecular dynamics simulations verified that glycyrol formed hydrogen bonds and π-π interactions with the key residue His-171 of SCD. It might disrupt the normal catalytic function of the enzyme by altering the conformation of the active center, thus suppressing the activity of SCD. In vitro experiments demonstrated that glycyrol could elevate the viability of hepatic stellate cells induced by lipopolysaccharide(P<0.05), suppress cell apoptosis(P<0.05), and upregulate mitochondrial membrane potential.

CONCLUSION Based on the “Ro Nus Zhurjes” theory, this study adopts a weighted network to reveal the overall multi-component, multi-target action characteristics of Jingzhu Gantaishu. It predictes that glycyrrhizin may exert its anti-hepatitis activity by targeting SCD, and preliminarily validates its protective effect on injured hepatic stellate cells. This research strategy provides a new paradigm for investigating the material basis of Tibetan medicine compound prescriptions and modernizing Tibetan medicine theory.