Effect of Danshen injection on expression of TNF-α, IL-6 and MMP-3 in articular cartilage of rabbits with knee osteoarthritis

Abstract

Objective To study the expression of tumor necrosis factor-α(TNF-α), interleukin(IL)-6, matrix metalloproteinase-3 (MMP-3) in the articular cartilage of rabbits with knee osteoarthritis (KOA) treated by Danshen injection, and to explore the molecular mechanism of Danshen injection in the treatment of knee osteoarthritis. Methods Forty healthy white rabbits were randomly divided into four groups: sham operation group, KOA model(KOA) group, KOA+ sodium hyaluronate (SH) group and KOA+ Danshen group.Sham operation group only made incision without cutting meniscus.The other groups all used cutting meniscus of rabbit right knee joint to establish an experimental animal model of joint instability.The KOA group was given 0.9% normal saline (0.7 mL/d), the KOA+ SH treatment group was given intra-articular injection of SH (0.4 mL/d), and the KOA+ Danshen treatment group was given Danshen injection (0.7 mL/d) for 5 weeks.The field experiment was used to verify whether the instability model of rabbits could affect the motor ability of rabbits.The mRNA content and the protein expression of TNF-α, IL-6 and MMP-3 were detected separately by polymerase chain reaction(PCR) and Western blot.The contents of TNF-α, IL-6 and MMP-3 in knee joint fluid were detected by enzyme linked immunosorbent assay(ELISA) kit. Results Before the establishment of KOA model, there was no significant difference in motor ability among the groups. After the establishment of the model, the movement distance of rabbits in the KOA group was significantly lower than that in the sham operation group[(150±5) vs (580±9), t=60.610, P<0.05]. The walking distance in the KOA+ Danshen group was significantly increased than that in the KOA group[(438±27) vs (150±5), t=16.730, P<0.05]. In rabbit joint fluid, the contents of TNF-α, IL-6, MMP-3 in KOA group were significantly higher than those in sham operation group [(22.62±2.18)mg/mL vs (11.74±2.09)mg/mL, (4.01±0.14)mg/mL vs (1.76±0.11)mg/mL, (0.57±0.05)mg/mL vs(0.27±0.03)mg/mL, t values were 4.413, 17.620 and 6.495 respectively, all P values<0.05], while, compared with KOA group, the content of TNF-α, IL-6, MMP-3 in KOA+ Danshen group were significantly lower [(22.62±2.18)mg/mL vs (15.01±2.37)mg/mL, (4.01±0.14)mg/mL vs (2.47±0.19)mg/mL, (0.57±0.05)mg/mL vs (0.40 ±0.02)mg/mL, t values were 3.725, 8.803 and 9.185 respectively, all P values<0.05]. In the mRNA detection of rabbit knee cartilage, the mRNA expression of TNF-α, IL-6, MMP-3 in KOA group was significantly higher than that in the sham operation group [(1.90±0.02) vs (1.00±0.15), (1.70±0.02) vs (1.00±0.07), (1.60±0.20) vs (1.00±0.13), t values were 11.050, 24.250 and 14.850 respectively, all P values< 0.05], and the mRNA expression of TNF-α, IL-6, MMP-3 in KOA+ Danshen group was significantly lower than that in KOA group [(1.09±0.05) vs (1.90±0.02), (1.13±0.09) vs (1.70±0.02), (1.04±0.08) vs (1.60±0.20), t values were 32.190, 8.822 and 5.868 respectively, all P values<0.05]. In the protein detection of rabbit knee cartilage, the protein expression of TNF-α, IL-6, MMP-3 in KOA group was increased significantly compared with sham operation group (t values were 65.280, 12.320 and 22.280 respectively, all P values<0.05), and the protein expression of TNF-α, IL-6, MMP-3 in KOA+ Danshen group was decreased significantly compared with KOA group (t values were 26.15, 9.053 and 24.670 respectively, all P values<0.05). Conclusion Danshen injection could obviously improve the motor ability of rabbits in KOA group, and also reduce the levels of TNF-α, IL-6 and MMP-3.Therefore, Danshen injection may inhibit the release of osteoarthritis matrix protease and the degradation of articular cartilage in the treatment of knee osteoarthritis, which can control the degenerative lesion of articular cartilage, and ultimately improve the symptoms of knee osteoarthritis. Key words: Knee osteoarthritis; Danshen injection; Tumor necrosis factor-α; Interleukin-6; Matrix metalloproteinase-3