低强度脉冲超声波对软骨细胞中金属蛋白酶-13与Ⅱ型胶原的影响

Effects of low intensity pulsed ultrasound on MMP-13 and type Ⅱ collagen of chondrocytes

目的:研究低强度脉冲超声波(LIPUS)对体外培养软骨细胞中金属蛋白酶-13(MMP-13)与Ⅱ型胶原的影响。方法:选用6只1月龄的新西兰大白兔膝关节软骨进行体外培养,传至第二代,分为对照组与4个LIPUS辐射组,LI-PUS强度分别是20mW/cm2、30mW/cm2、40mW/cm2、50mW/cm2。每天辐射20min,连续10d,每天细胞计数;分别在第5天及第10天收集细胞,采用Western-blot技术、qRT-PCR技术检测软骨细胞中MMP-13、Ⅱ型胶原的含量,并分析两者之间的相关性。结果:①与对照组相比,各LIPUS辐射组软骨细胞数均明显增高(P<0.05),其中40mW/cm2组软骨细胞数最高,与其他各辐射组比较有显著性差异(P<0.05);②细胞培养5d和10d后,MMP-13含量均较第0天明显升高(P<0.05)。但各辐射组MMP-13含量均较其对照组显著降低(P<0.05);其中40mW/cm2组MMP-13含量最低(P<0.05);③细胞培养5d和10d后,Ⅱ型胶原表达量均较第0天明显降低(P<0.05)。但各辐射组Ⅱ型胶原表达量均较其对照组显著升高(P<0.05)。其中40mW/cm2组Ⅱ型胶原表达量最高(P<0.05);④MMP-13与Ⅱ型胶原含量呈负相关(r=-0.757,P<0.05)。结论:不同强度的LIPUS体外辐射可促进软骨细胞增殖,抑制MMP-13的表达,延缓II型胶原的降解。

Objective： To study the effects of low intensity pulsed ultrasound（LIPUS） on matrix metalloproteinase（MMP-13） and type II collagen of chondrocytes. Method： The chondrocytes were obtained from knee joints of six one-month old rabbits. Chondrocytes were divid- ed into 5 groups：control group and 4 LIPUS groups with intensity of 20mW/cm2,30mW/cm2,40mW/cmZ,50mW/cm2 respectively.During the next cultured 10d, the chondrocytes were irradiated 20rain and cells were counted every day.On the 5th d and 10th d, cells were collected and the MMP-13 and type II collagen content were detected by Western-blot and qRT-PCR. The correlation between MMP-13 and type II collagen was analyzed. Result： @Contrasting with control group, the chondrocytes number was higher in each LIPUS group（P〈0.05） and the number was significantly higher in 40mW/cm2 group （P〈0.05）.@MMP-13 contents were higher at the 5th d and 10th d than that at the 0d（P〈0.05）.After irradiated 5d and 10d, the content of MMP-13 in each LIPUS group was remarkable lower than that in control group（P〈0.05） and in 40mW/cm2 group the content of MMP-13 was the lowest （P〈0.05）.（1）After cultured 5d and 10d, type II collagen content was lower than that at the 0d（P〈0.05）. The content of type II collagen in each LIPUS groups was remarkable higher than that in control group（P〈 0.05） and in 40mW/cm2 group the content of type II collagen was the highest （P〈0.05）.（4）The correlation between MMP-13 and type II collagen content was negative correlation （r=-0.757,P〈0.05）. Conclusion： Different intensity of LIPUS could promote chondrocyte proliferation and inhibit the expression of MMP-13 and postpone the degradation of typell collagen.