Protective effect of paeoniflorin on H₂O₂ induced Schwann cells injury based on network pharmacology and experimental validation

This study was to investigate the protective effect of paeoniflorin (PF) on hydrogen peroxide-induced injury. Firstly, “SMILES” of PF was searched in Pubchem and further was used for reverse molecular docking in Swiss Target Prediction database to obtain potential targets. Injury-related molecules were obtained from GeenCards database, and the predicted targets of PF for injury treatment were selected by Wayne diagram. For mechanism analysis, the protein-protein interactions were constructed by String, and the KEGG analysis was conducted in Webgestalt. Then, cell viability and cytotoxicity assay were established by CCK8 assay. Also, the experimental cells were allocated to control, model (200 μmol·L⁻¹ H₂O₂), SB203580 10 μmol·L⁻¹ (200 μmol·L⁻¹ H₂O₂ + SB203580 10 μmol·L⁻¹), PF 50 μmol·L⁻¹ (200 μmol·L⁻¹ H₂O₂ + PF 50 μmol·L⁻¹), and PF 100 μmol·L⁻¹ (200 μmol·L⁻¹ H₂O₂ + PF 100 μmol·L⁻¹) groups. We measured the intracellular ROS, Hoechst 33258 staining, cell apoptosis, the levels of Bcl-xl, Bcl-2, Caspase-3, Cleaved-caspase3, Cleaved-caspase7, TRPA1, TRPV1, and the phosphorylation expression of p38MAPK. There are 96 potential targets that may be associated with PF for injury treatment. Then, we chose the “Inflammatory mediator regulation of TRP channels” pathway for the experimental verification from the first 10 KEGG pathway. In experimental verification, H₂O₂ decreased the cell viability moderately (P < 0.05), and 100 μmol·L⁻¹ PF increased the cell viability significantly (P < 0.05). Depending on the difference of intracellular ROS fluorescence intensity, PF inhibited H₂O₂-induced reactive oxygen species production in Schwann cells. In Hoechst 33258 staining, PF reversed the condensed chromatin and apoptotic nuclei following H₂O₂ treatment. Moreover, Flow cytometry results showed that PF could substantially inhibit H₂O₂ induced apoptosis (P < 0.05). Pretreatment with PF obviously reduced the levels of Caspase3, Cleaved-caspase3, Cleaved-caspase7, TRPA1, TRPV1, and the phosphorylation expression of p38MAPK after H₂O₂ treatment (P < 0.05), increased the levels of Bcl-2, and Bcl-xl (P < 0.05). PF inhibited Schwann cell injury and apoptosis induced by hydrogen peroxide, which mechanism was linked to the inhibition of phosphorylation of p38MAPK.