Hederagenin's uric acid-lowering effects in hyperuricemic mice: Mechanistic insights from molecular docking and in vivo analysis

Abstract

This study explored the uric acid-lowering effects of hederagenin (HD) through molecular docking analysis and a chronic hyperuricemia (HUA) mouse model. Molecular docking was performed to evaluate HD's interactions key urate-regulating proteins, including xanthine oxidase (XOD), ABCG2, OAT1, URAT1, and GLUT9. To establish a chronic HUA model, mice were fed a yeast-adenine diet supplemented with potassium oxonate. The mice were randomly assigned to six groups: normal control, HUA model control, benzbromarone (BEN) group, and three HD treatment groups at doses of 50, 100, and 200 mg/kg. Serum uric acid (UA) levels, liver and kidney function indicators, XOD activity, and oxidative stress markers were assessed. Histopathological analyses of the liver and kidney were also conducted. In addition, gene and protein expression levels of urate transporters and inflammatory markers were assessed using RT-PCR and Western blotting. The results showed that HD interacts with XOD and urate transporters, significantly reducing serum UA levels and inhibiting XOD activity in HUA model. It also modulated the expression of urate transporter to enhance UA excretion. Moreover, HD protected liver and kidney function by reducing pro-inflammatory cytokine levels and inhibiting the TLR4/Myd88/NF-κB and NLRP3 signaling pathways. These findings suggest HD may serve as a promising therapeutic agent for lowing uric acid and preventing organ damage associated with HUA.

Fig 1. The modelled structure and Ramachandran plot of OAT1.

(A), URAT1 (B), GLUT9 (C) models, and docked complex of hederagenin (2D and 3D) with XOD (D).

Fig 2. Docked complex of hederagenin (2D and 3D) with ABCG2.

(A), OAT1 (B), URAT1 (C), and GLUT9 (D).

Fig 3. Effects of HD on serum uric acid levels in mice (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 4. Effects of HD on serum Cr.

(A), BUN (B), ALT(C), and AST(D) in mice (**p < 0.01 compare with NC group; #p < 0.05, ##p < 0.01 compare with MC group).

Fig 5. Effects of HD on organ index in mice.

(A), Kidney index (B), Liver index (C), and Thymus index (D), and Spleen index (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 6. Effects of HD on liver XOD activities in mice (Note: *p < 0.05, **p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 7. Effects of HD on oxidative stress indices of kidney and liver in mice on Kidney SOD activities.

(A), Kidney GSH-PX activities (B), Kidney MDA contents (C), Liver SOD activities (D), Liver GSH-PX activities (E), and Liver MDA contents (F).

Fig 8. Histological analyses micrograpahs of liver tissue stained with HE (HE, scale bar 100 µm).

(A), and results of semi-quantitative analysis(**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group) (B).

Fig 9. Histological analyses micrograpahs of renal tissue stained with HE (HE, scale bar 100 µm).

(A), and results of semi-quantitative analysis (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group) (B).

Fig 10. Effects of HD on mRNA of OAT1, OAT3 and ABCG2 of kidney in mice.

(A), RT-PCR was used to detect the gene expression of OAT1, OAT3 and ABCG2; Results of semi-quantitative analysis of OAT1 (B), OAT3 (C), and ABCG2 (D).

Fig 11. Effects of HD on mRNA of URAT1 and GLUT9 of kidney in mice as follows: RT-PCR was used to detect the gene expressions of URAT1 and GLUT9.

(A), results of semi-quantitative analysis of URAT1 and GLUT9 (B-C) (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 12. Effects of HD on the protein expressionsof uric acid transporters of kidney in mice as follows: western blot was used to detect the protein expressions of OAT1, OAT3, ABCG2, URAT1 and GLUT9.

(A), and results of semi-quantitative analysis of OAT1, OAT3, ABCG2, URAT1 and GLUT9 (B-F) (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 13. Effects of HD on inflammatory cytokines in renal tissue homogenatein mice as follows: western blot was used to detect the protein expressions of TNF-

α, IL-6 and IL-1β. (A), and results of semi-quantitative analysis of TNF-α, IL-6 and IL-1β (B-D) (**p < 0.01 compare with NC group; #p < 0.05, ##p < 0.01 compare with MC group).

Fig 14. Expressions of TLR4 pathway related proteins of kidney in mice as follows: western blot was used to detect the protein expressions of TLR4, Myd88 and NF-

κB p65. (A), and results of semi-quantitative analysis of TLR4, Myd88 and NF-κB p65 (B-D) (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).

Fig 15. Expressions of NLRP3 pathway related proteins of kidney in mice as follows: western blot was used to detect the protein expressions of NLRP3, ASC and Casepase-1.

(A), and results of semi-quantitative analysis of NLRP3, ASC and Casepase-1 (B-D) (**p < 0.01 compare with NC group; ##p < 0.01 compare with MC group).