2022, Vol. 26 ›› Issue (20): 3236-3242

Astragalus polysaccharides repair articular cartilage damage in a mouse osteoarthritis model

Shi Guirong1, Ren Bowen2, Zhang Zhongbo2, Wang Lisha2, Zhang Qiwei2, Shi Dongliang2   

Abstract: BACKGROUND: Astragalus polysaccharide has a definite effect on osteoarthritis, but its mechanism of action is still unclear.  
OBJECTIVE: To investigate the effects of astragalus polysaccharide on osteoarthritis through regulating ubiquitination of Notch receptor 1 mediated by WW domain containing E3 ubiquitin protein ligase 2 (WWP2).
METHODS: In the in vivo experiment, 30 C57BL/6 mice were randomly divided into 3 groups (n=10): a sham operation group, a model group, and an astragalus polysaccharide group. A mouse osteoarthritis model was established in the model and astragalus polysaccharide groups. Mice in the astragalus polysaccharide group were treated with astragalus polysaccharide. After 4 weeks of intervention, the knee joint tissue was taken under anesthesia. Safranin O staining method and Osteoarthritis Research Society International score were used to evaluate the damage of articular cartilage in mice. In the in vitro experiment, C28/12 cells were cultured with 5 μg/L interleukin 1β for 24 hours to establish osteoarthritis cell models, which were divided into a control group, an interleukin 1β group, a 50 mg/L astragalus polysaccharide group, and an astragalus polysaccharide-Notch receptor 1 overexpression and a WWP2 interference group. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to detect cell proliferation. Flow cytometry was used to detect apoptosis. Co-immunoprecipitation assay was used to verify the binding of WWP2 and Notch receptor 1. Ubiquitination experiment was used to analyze the effect of astragalus polysaccharides on the ubiquitination of Notch receptor 1 mediated by WWP2. Enzyme-linked immunosorbent assay was used to detect the levels of proteoglycans, type II collagen, matrix metallopeptidase 3, and matrix metallopeptidase 13 in mouse knee joint tissue and C28/12 cells. Western blot assay was used to detect the protein expression of WWP2, Notch receptor 1, jagged canonical Notch ligand 1, and Notch intracellular domain 1 in mouse knee joint tissue and C28/12 cells.  
RESULTS AND CONCLUSION: The results of in vivo experiments confirmed that intervention of astragalus polysaccharides alleviated cartilage damage in mice, inhibited the levels of matrix metallopeptidase 3 and matrix metallopeptidase 13, and promoted the expression of proteoglycans and type II collagen in mouse knee joint tissue. The results of in vitro experiments confirmed that the intervention of astragalus polysaccharides promoted cell proliferation and the expression of proteoglycan, type II collagen, and WWP2, and inhibited cell apoptosis and the expression of matrix metallopeptidase 3 and matrix metallopeptidase 13, Notch receptor 1, jagged canonical Notch ligand 1, and Notch intracellular domain 1, while Notch receptor 1 overexpression and WWP2 interference reversed the effect of astragalus polysaccharides. Astragalus polysaccharides promoted the ubiquitination of Notch receptor 1 mediated by WWP2. To conclude, astragalus polysaccharides can play a certain therapeutic effect on mouse osteoarthritis by increasing the WWP2-mediated ubiquitination level of Notch receptor 1 and inhibiting the Notch signaling pathway.
Key words: astragalus polysaccharide, WW domain containing E3 ubiquitin protein ligase 2, ubiquitination, Notch receptor 1, osteoarthritis