A study on the network pharmacology of HQGZWWT that prevents CINP by inhibiting ferroptosis through regulation of the p38/c-FOS/NF-kB pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Chemotherapy-induced neuropathic pain (CINP) is a frequent side effect of chemotherapy, and the mechanisms by which Huangqi Guizhi Wuwu Tang (HQGZWWT) prevents and treats CINP remain to be further elucidated.

BACKGROUND: Oxaliplatin is a widely used chemotherapy drug with proven efficacy against multiple malignancies; however, its significant side effects severely limit patient dosing. One such effect is CINP, which involves multiple factors. Ferroptosis, a form of cell death, plays a crucial role in maintaining chronic neuropathic pain. HQGZWWT effectively prevents and treats CINP and is recommended by the Expert Consensus on Diagnosis and Treatment of Chemotherapy-Induced Peripheral Neuropathic Pain (A + Level). However, the specific mechanism remains unclear.

OBJECTIVE: In this study, the preventive and therapeutic effects of HQGZWWT and its major monomeric component paeoniflorin on CINP were investigated, and the effects of HQGZWWT on CINP through the regulation of ferroptosis mediated by the p38/c-FOS/NF-κB pathway were explored.

METHODS: In this study, a CINP rat model was established to evaluate the protective efficacy of astragalus-cinnamon five-ingredient decoction against neural injury in CINP rats. Further investigations focused on the pharmacological effects of its primary monomer, paeoniflorin. Through in vitro experiments using rat dorsal root ganglion (DRG) cells (CPR126) and in vivo experiments with SD rats, the potential mechanism through which paeoniflorin modulates ferroptosis by regulating the p38/c-FOS/NF-κB pathway was investigated. The corresponding targets were screened via network pharmacology and validated through molecular docking, molecular dynamics simulations, and immunofluorescence colocalization.

RESULTS: HQGZWWT effectively alleviated oxaliplatin-induced neuropathic pain. In vivo experiments demonstrated that HQGZWWT improved mechanical pain sensitization and lowered thermal pain thresholds in oxaliplatin-treated rats. Hematoxylin and eosin (HE) staining revealed that HQGZWWT mitigated oxaliplatin-induced abnormalities in dorsal root ganglion neurons, including indistinct nuclei, reduced proliferative activity, and abnormal neuronal axons, as well as a disorganized sciatic nerve arrangement and increased intercellular spacing. Transmission electron microscopy revealed uniform mitochondrial structure and density in the HQGZWWT high-concentration group, counteracting oxaliplatin-induced reductions in mitochondrial density, vacuolation and rupture of mitochondrial membranes, and structural blurring. The HQGZWWT-high group exhibited reduced oxaliplatin-induced increases in Malondialdehyde(MDA)levels. The total tissue iron and Fe2+ levels indicated that HQGZWWT mitigated oxaliplatin-induced iron accumulation. Through network pharmacology screening, bioactive component analysis, molecular docking, and molecular dynamics simulations, in vitro experiments identified paeoniflorin as the optimal active monomer in HQGZWWT. CCK-8 assays confirmed 50 μM as the optimal paeoniflorin concentration and demonstrated that oxaliplatin induces DRG cell death via ferroptosis. Paeoniflorin effectively protected rat DRG cells against oxaliplatin-induced damage and reduced oxaliplatin-induced reactive oxygen species (ROS) production. Mitochondrial electron microscopy revealed that 50 μM paeoniflorin improved oxaliplatin-induced alterations in DRG cells, including reduced mitochondrial density, mitochondrial vacuolation, and cristae damage, while decreasing oxidative stress levels. These findings provide strong evidence for the inhibitory effect of 50 μM paeoniflorin on ferroptosis. Additionally, network pharmacology identified the p38/c-Fos/NF-κB signaling pathway, validating the hypothesis that HQGZWWT and paeoniflorin inhibit ferroptosis via this pathway to prevent CINP.

CONCLUSION: HQGZWWT significantly ameliorated CINP by increasing sciatic nerve conduction velocity, reducing oxidative stress and inflammatory responses, and inhibiting ferroptosis. Both HQGZWWT and its active monomeric component paeoniflorin protect neurons by suppressing ferroptosis, thereby exerting preventive and therapeutic effects against CINP. These findings provide new insights into the clinical treatment of CINP with HQGZWWT. Inhibiting ferroptosis by regulating the p38/c-FOS/NF-κB pathway through HQGZWWT and its primary monomeric component paeoniflorin represents a promising therapeutic strategy for preventing and treating oxaliplatin-induced CINP.