Tougu Xiaotong Capsule alleviates cartilage degeneration in mice with knee osteoarthritis by modulating Nav1.7

doi:10.12122/j.issn.1673-4254.2024.11.03

Abstract

Abstract:

Objective To investigate the mechanism by which Tougu Xiaotong Capsule (TGXTC) alleviates chondrocyte degeneration in knee osteoarthritis (KOA). Methods Thirty 2-month-old C57BL/6 mouse models of KOA established using the Hulth method were randomized into model group, TGXTC group, and diclofenac sodium group and received treatment with saline, TGXTC (368 mg/kg), and diclofenac sodium (10 mg/kg) by gavage, respectively, with another 10 untreated mice as the blank control group. All interventions were administered 6 times a week for 4 weeks. After the treatments, structural changes in the cartilage tissue were observed with morphological staining, and Nav1.7 mRNA expression and the protein expression levels of Nav1.7, MMP-3, ADAMTS-5, and COX-2 were detected using RT-qPCR and Western blotting. Fluorescence in situ hybridization (FISH) was used to detect Nav1.7 expression in the chondrocytes. In cultured KOA chondrocytes, the effect of TGXTC and lentivirus-mediated Nav1.7 knockdown on MMP-3, MMP-13, ADAMTS-4, ADAMTS-5, and COX-2 protein expressions were assessed with Western blotting. Results In KOA mice treatments with TGXTC and diclofenac sodium both significantly alleviated structural damage of the cartilage layer, reduced Nav1.7 protein and mRNA expressions and lowered the expressions of MMP-3, ADAMTS-5, and COX-2 proteins in the cartilage tissues. FISH results indicated that TGXTC treatment significantly reduced IL-1β-induced Nav1.7 expression in the chondrocytes. In Nav1.7 knockdown experiment, Nav1.7 levels were significantly lower in IL-1β+sh-Nav1.7 group than in IL-1β group, and also lower in IL-1β+TGXTC group than in IL-1β+sh-Nav1.7+TGXTC group. TGXTC treatment significantly inhibited IL-1β-induced elevation of MMP-3, MMP-13, ADAMTS-4, ADAMTS-5 and COX-2 protein expressions in the chondrocytes, but its effects were strongly weakened by Nav1.7 knockdown. Conclusion TGXTC alleviates extracellular matrix metabolic disorder in KOA chondrocytes by regulating Nav1.7, thereby mitigating chondrocyte degeneration in KOA mice.

Key words: Tougu Xiaotong Capsule, Nav1.7, chondrocyte degeneration, knee osteoarthritis

Changlong FU, Yanming LIN, Shujie LAN, Yue CHEN, Chao LI, Shiyu LU, Qing LIN. Tougu Xiaotong Capsule alleviates cartilage degeneration in mice with knee osteoarthritis by modulating Nav1.7[J]. Journal of Southern Medical University, 2024, 44(11): 2074-2081.

Figures/Tables 5

Fig.1 HE, safranin O and toluidine blue staining of the articular cartilage in different groups (Original magnification: ×100).

Fig.2 Nav1.7, MMP-3, ADAMTS-5, and COX-2 protein expressions in the cartilage tissue in different groups. A-E: Western blotting and quantitative data of Nav1.7, MMP-3, ADAMTS-5, and COX-2 expressions in each group (n=3). *P<0.05 vs Sham group, #P<0.05 vs Model group.

Fig.3 Multiplicity of infection conditions of Nav1.7 transfection into the chondrocytes.

Fig.4 Fluorescence in situ hybridization reveals Nav1.7 expression in the cytoplasm of the chondrocytes (×100). *P<0.05 vs Sham group; #P<0.05 vs IL-1β group.

Fig.5 MMP-3, MMP-13, ADAMTS-4, ADAMTS-5, and COX-2 proteins expressions in the chondrocytes with TGXTC treatment and lentivirus-mediated Nav1.7 knockdown. A-F: Western blotting and quantitative data of MMP-3, MMP-13, ADAMTS-4, ADAMTS-5 and COX-2 expressions in each group (n=3). *P<0.05 vs Sham group; #P<0.05 vs IL-1β group; &P<0.05 vs IL-1β+sh-Nav1.7 group; △P<0.05 vs IL-1β+TGXTC group.

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