水飞蓟宾对白细胞介素-1β诱导的软骨细胞损伤的影响

Effect of silybin on chondrocyte injury induced by interleukin-1β

目的 探讨水飞蓟宾对白细胞介素-1β(IL-1β)诱导的软骨细胞损伤的影响及其机制。方法 将体外培养的软骨细胞分为对照组、诱导组(以10 ng·mL^(-1)IL-1β诱导处理)和低、中、高剂量实验组(在10 ng·mL^(-1)IL-1β诱导基础上分别以40,80,160μmol·L^(-1)水飞蓟宾处理)。用流式细胞术检测细胞凋亡率,用酶联免疫吸附(ELISA)法检测基质金属蛋白酶-13(MMP-13)、肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)水平,用蛋白质印迹(Western blot)法检测细胞中微管相关蛋白1(Beclin-1)、无翅型小鼠乳房肿瘤病毒整合位点1(Wnt1)和β-连环蛋白(β-catenin)表达水平。结果 对照组、诱导组、低剂量实验组、中剂量实验组和高剂量实验组细胞凋亡率分别为(3.24±0.60)%,(31.18±2.51)%,(28.04±2.10)%,(22.03±1.79)%和(17.48±0.69)%;MMP-13水平分别为(21.18±3.09),(84.05±5.12),(75.16±4.15),(59.71±3.31)和(40.43±2.72)μg·L^(-1);TNF-α水平分别为(4.66±0.55),(57.22±3.53),(50.38±2.16),(38.75±2.36)和(15.64±0.73)ng·L^(-1);IL-6水平分别为(7.38±0.69),(68.76±4.73),(60.88±3.21),(49.74±2.86)和(32.35±2.14)ng·L^(-1);Beclin-1蛋白表达水平分别为0.85±0.06,0.26±0.02,0.35±0.03,0.48±0.03和0.61±0.04;Wnt1蛋白表达水平分别为0.26±0.03,0.85±0.06,0.66±0.03,0.45±0.04和0.35±0.03;β-catenin蛋白表达水平分别为0.13±0.02,0.76±0.05,0.66±0.05,0.53±0.04和0.42±0.03。诱导组和对照组相比,各剂量实验组与诱导组比较,差异均有统计学意义(均P<0.05)。结论 水飞蓟宾可改善IL-1β诱导的软骨细胞损伤,其作用机制可能与抑制炎症反应和Wnt/β-catenin信号通路活化有关。

Objective To investigate the effect of silybin on interleukin 1β(IL-1β)-induced chondrocyte injury and its mechanism. Methods The chondrocytes cultured in vitro were divided into control group, induction group(10 ng·mL^(-1)IL-1β) and low, medium, high dose experimental groups(40, 80 and 160 μmol·L^(-1)silybin on the basis of 10 ng· mL^(-1)IL-1β). Cell apoptosis rate was measured by flow cytometry. The levels of matrix metalloproteinase-13(MMP-13), tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) were detected by enzyme-linked immunosorbent assay(ELISA), and the protein expression levels of Beclin-1, Wnt1 and β-catenin were detected by Western blot. Results The cell apoptosis rates of control group,induction group and low,medium,high dose experimental groups were( 3. 24 ± 0. 60) %,( 31. 18 ± 2. 51) %,( 28. 04 ± 2. 10) %,( 22. 03 ± 1. 79) % and( 17. 48 ± 0. 69) %,respectively;MMP-13 levels were( 21. 18 ± 3. 09),( 84. 05 ± 5. 12),( 75. 16 ± 4. 15),( 59. 71 ± 3. 31) and( 40. 43 ± 2. 72) μg·L^(-1),respectively;TNF-α levels were( 4. 66 ± 0. 55),( 57. 22 ± 3. 53),( 50. 38 ± 2. 16),( 38. 75 ± 2. 36) and( 15. 64 ± 0. 73) ng·L^(-1),respectively;IL-6 levels were( 7. 38 ± 0. 69),( 68. 76 ± 4. 73),( 60. 88 ± 3. 21),( 49. 74 ± 2. 86) and( 32. 35 ± 2. 14) ng·L^(-1);Beclin-1 protein expression levels were 0. 85 ± 0. 06,0. 26 ± 0. 02,0. 35 ± 0. 03,0. 48 ± 0. 03 and 0. 61 ± 0. 04,respectively;the expression levels of Wnt1 protein were 0. 26 ± 0. 03,0. 85 ± 0. 06,0. 66 ± 0. 03,0. 45 ± 0. 04 and 0. 35 ± 0. 03,respectively;β-catenin protein expression levels were 0. 13 ± 0. 02,0. 76 ± 0. 05,0. 66 ± 0. 05,0. 53 ± 0. 04 and 0. 42 ± 0. 03,respectively. There were statistically significant differences in the above indexes between induction group and control group or between experimental groups at each dose and the induction group( all P < 0. 05). Conclusion Silybin can improve IL-1β-induced chondrocyte injury,and its mechanism may be related to the inhibition of inflammatory response and the activation of Wnt/β-catenin signaling pathway.