Integrated Network Pharmacology and Transcriptomics to Explore the Mechanism of Compound Dihuang Granule (CDG) Protects Dopaminergic Neurons by Regulating the Nrf2/HMOX1 Pathway in the 6-OHDA/MPP+ -Induced Model of Parkinson's Disease

Abstract Background: Parkinson's disease (PD) is a degenerative neurological disease that worsens over time. Ferroptosis has been proven to contribute to PD pathogenesis. CDG exhibits neuroprotective effects. However, CDG's potential mechanism in PD therapy remains uncertain. Purpose: The purpose of this investigation is to ascertain the specific molecular mechanisms of CDG against neuronal ferroptosis and present an alternative option for PD management. Methods: Network pharmacology along with LC-MS were used to identify possible targets and candidate pathways. Then RNA-sequencing combined in vitro and in vivo experiments were utilized to validate these findings. Results: According to network pharmacology prediction, NFE2L2, HMOX1, PTGS2 may be the key genes for ferroptosis in PD. In vivo experiments, CDG improved the neurobehavior of PD rats, alleviated the damage of dopamine neurons, decreased the levels of MDA, ROS and Fe2+, increased the GSH level, and inhibited ferroptosis by decreasing ACSL4, TF, and PTGS2 expression levels, increased the GPX4, FTH, Nrf2, and HMOX1 levels. RNA-seq analysis showed the differential genes in Model and CDG group were all enriched in Nrf2 and HMOX1, and the enrichment analysis of these differential genes showed they were closely related to the ferroptosis. Subsequently, in vitro experiments, the CDG, OE-Nrf2 and OE-HMOX1 group showed more active cell vitality, decreased MDA, ROS, Fe2+, ACSL4, TF, PTGS2 levels, and increased GSH, GPX4, FTH, Nrf2, HMOX1 levels. Conclusion: CDG has a neuroprotective involvement in alleviating ferroptosis by regulating the Nrf2/HMOX1 pathway. Moreover, this research offers pharmacological evidence supporting the applications of CDG for treating PD.