This study aims to investigate the therapeutic effects of eriodictyol on rheumatoid arthritis and its underlying molecular mechanisms. By establishing in vitro cell models and in vivo animal models, the intervention effects of eriodictyol on inflammatory responses, cartilage destruction, and bone erosion will be evaluated. Furthermore, the interaction patterns between eriodictyol and potential target proteins will be explored, aiming to provide a novel candidate drug and theoretical basis for the treatment of rheumatoid arthritis.

This study employed a variety of experimental techniques to systematically investigate the mechanism by which eriodictyol alleviates rheumatoid arthritis. First, an in vitro inflammation model was established using TNF-α-induced MH7A cells. The CCK-8 assay was used to determine the safe concentration range of the drug, flow cytometry was employed to detect the cell apoptosis rate, and wound healing assays and Transwell chambers were utilized to assess cell migration ability. Concurrently, a tube formation assay was performed on human umbilical vein endothelial cells using Matrigel to observe the inhibitory effect of eriodictyol on angiogenesis. In in vivo experiments, a collagen-induced arthritis mouse model was established. Following intervention with different doses of eriodictyol, joint tissues were collected for Hematoxylin-Eosin staining to observe inflammatory infiltration and synovial hyperplasia, and Safranin O-Fast Green staining to assess cartilage destruction and proteoglycan loss. Furthermore, MicroCT imaging technology was used for three-dimensional reconstruction of bone microstructure and quantitative analysis of parameters related to bone erosion. Additionally, synovial tissue samples from the control, model, and eriodictyol treatment groups were collected for proteomic analysis. Liquid chromatography-tandem mass spectrometry was performed to identify differentially expressed proteins, followed by bioinformatics methods for GO functional enrichment and KEGG pathway enrichment analysis. These methods, spanning the cellular, molecular, and whole-animal levels, systematically elucidated the multi-target mechanism of eriodictyol in regulating inflammatory responses, inhibiting abnormal synovial cell activation, protecting cartilage, and reducing bone destruction.

Compared with the untreated group, eriodictyol and MTX showed no significant effects on the cell cycle or apoptosis of MH7A cells. Wound healing and Transwell migration assays demonstrated that eriodictyol inhibited the migration of MH7A cells in a dose-dependent manner. When MH7A cells were indirectly co-cultured with human umbilical vein endothelial cells (HUVECs), the HUVECs exhibited excessive proliferation, high migration capacity, strong adhesion, and formed dense tube networks on Matrigel, with increased numbers of junction points and branches. Following treatment with eriodictyol, HUVEC viability returned to normal levels, tube formation was inhibited, and the number of junction points decreased. H&E staining revealed extensive inflammatory cell infiltration, significant synovitis, and marked synovial hyperplasia in the CIA group, while both the MTX and eriodictyol treatment groups showed significant improvement in synovial inflammation. Safranin O-Fast Green staining indicated that articular cartilage erosion was markedly ameliorated in the eriodictyol group. MicroCT imaging was used to observe and measure joint bone erosion. CIA mice exhibited irregular bone surfaces and significant erosion, whereas eriodictyol effectively reduced bone erosion in these mice. To investigate the anti-inflammatory and anti-proliferative effects of eriodictyol on MH7A cells, cells were treated with 80 μM eriodictyol. Collected cells were subjected to proteomic analysis, and the results of KEGG enrichment analysis revealed that eriodictyol regulated carbohydrate metabolism, lipid metabolism, and amino acid metabolism. Among the significantly downregulated proteins, SLC7A2, a key protein involved in arginine metabolism, was identified as a critical target in the treatment of rheumatoid arthritis with eriodictyol.

This study confirms that eriodictyol can alleviate rheumatoid arthritis through a multi-target mechanism. In vitro experiments showed that although eriodictyol did not affect the apoptosis or cell cycle of MH7A cells, it inhibited their migration in a dose-dependent manner and suppressed angiogenesis by blocking the synovial-endothelial cell interaction. In vivo experiments demonstrated that eriodictyol significantly reduced synovial inflammation, cartilage erosion, and bone destruction in CIA mice. Proteomic analysis suggested that its mechanism of action may involve the regulation of SLC7A2, a key protein in arginine metabolism, and related metabolic pathways. In conclusion, eriodictyol has potential value for the treatment of rheumatoid arthritis.