The Effect of Decursin on Chondrocyte Apoptosis in Knee Osteoarthritis Rats by Regulating the HMGB1-RAGE Signaling Pathway
LIU Jiawei1, XIN Zhifeng1, Saiyinduleng2, ZHANG Fei1*
This study aimed to investigate the effects of DE (decursin) on regulating the HMGB1-RAGE signaling pathway to modulate apoptosis in chondrocytes from KOA (knee osteoarthritis) in rats. First, KOA rat chondrocytes were isolated and cultured and randomly assigned to the Control group, Model group, L-DE, M-DE, and H-DE groups (1, 5, and 10 μmol/L DE, respectively), and the H-DE+HMGB1 group (10 μmol/L DE+3 μg/mL re- combinant HMGB1 protein). Chondrocytes were identified by toluidine blue staining and type II collagen immunofluorescence; cell proliferation was measured by the CCK-8 assay; apoptosis was assessed by flow cytometry; IL-6 and TNF-α levels in the culture supernatant were detected by ELISA; mitochondrial membrane potential changes were evaluated by JC-1 assay; and HMGB1-RAGE pathway proteins as well as C-Caspase-3 (cleaved caspase-3) and Pro-Caspase-3 expression were analyzed by Western blot. The results showed that the isolated rat chondrocytes were correctly identified by toluidine blue staining and immunofluorescence for type II collagen. In the Model group, chondrocyte viability decreased compared with the Control group, while apoptosis, supernatant IL-6 and TNF-α levels, JC-1 monomers/JC-1 aggregates ratio, and expression of HMGB1, RAGE, C-Caspase-3, and the C- Caspase-3/Pro-Caspase-3 ratio increased, whereas Pro-Caspase-3 expression decreased (P<0.05). In the L-DE, M-DE, and H-DE groups, chondrocyte viability was higher than in the Model group, apoptosis and levels of IL-6 and TNF-α in the supernatant, JC-1 monomer/JC-1 aggregate ratio, HMGB1, RAGE, C-Caspase-3 expression, and the C-Caspase-3/Pro-Caspase-3 ratio were reduced, and Pro-Caspase-3 expression increased (P<0.05). In the H-DE+HMGB1 group, chondrocyte viability was lower than in the H-DE group, while apoptosis and levels of IL-6 and TNF-α in the supernatant, JC-1 monomer/JC-1 aggregate ratio, HMGB1, RAGE, C-Caspase-3 expression, and the C-Caspase-3/Pro-Caspase-3 ratio were increased, and Pro-Caspase-3 expression decreased (P<0.05). In summary, DE may reduce mitochondrial-associated apoptosis of KOA rat chondrocytes by inhibiting the HMGB1/RAGE signaling pathway.
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