硝普钠诱导猪软骨细胞凋亡

Sodium nitroprusside induced apoptosis of porcine chondrocytes

[目的]研究硝普钠(Sodium nitroprusside, SNP)诱导猪软骨细胞凋亡的适宜剂量和作用时间,构建猪软骨细胞的凋亡模型。[方法]体外培养并鉴定猪踝关节软骨细胞。分别用0、0.05、0.1、0.2、0.4、0.8、1.6 mmol/L的SNP处理软骨细胞。于12、24、48 h后分别检测软骨细胞增殖、凋亡和线粒体膜电位。[结果]随SNP浓度增加,细胞增殖呈曲线改变,SNP在0.1 mmol/L时OD值达峰值,而后下降,不同浓度SNP组间差异有统计学意义(P<0.05)。相同SNP不同时间点的差异亦有统计学意义(P<0.05)。凋亡检测显示0.2、0.4、0.8、1.6 mmol/L SNP处理的猪软骨细胞发生了凋亡,且凋亡率随SNP浓度的增大而加大。线粒体膜电位染色显示随着SNP浓度的增大,猪软骨细胞的红色荧光强度与绿色荧光强度的相对比值逐渐缩小。[结论] 0.2、0.4、0.8、1.6 mmol/L SNP均引发了猪软骨细胞凋亡,可以用于构建猪软骨细胞凋亡模型。且当SNP的浓度逐渐增大,猪软骨细胞的线粒体膜电位逐渐减小。

[Objective]To investigate the appropriate dose and duration of sodium nitroprusside(SNP)for inducing apoptosis of porcine chondrocytes.[Methods]The chondrocytes isolated from porcine ankle cartilage were cultured and identified in vitro,and then treated with different concentrations of SNP,including 0,0.05,0.1,0.2,0.4,0.8,1.6 mmol/L.At 12,24,and 48 h,proliferation,apoptosis and mitochondrial membrane potential of the chondrocytes were detected.[Results]As increase of SNP concentration,the optical density(OD)of CCK-8 assay varied in a cure,which raised to the peak at 0.01 mmol/L and then declined continuously with a statistically significant difference among the concentrations(P<0.05).In addition,the OD value changed significantly at same concentration among different time points(P<0.05).Flow cytometry showed that chondrocyte apoptosis occurred in 0.2,0.4,0.8,1.6 mmol/L of SNP,which proved dosage dependent.In term of mitochondrial membrane potential assay,the relative ratio of the red fluorescence intensity to the green fluorescence intensity in the chondrocytes significantly decreased as the increase of SNP concentration.[Conclusion]The 0.2,0.4,0.8,1.6 mmol/L of SNP do induce apoptosis of chondrocytes.In addition,as the SNP concentration increases,the mitochondrial membrane potential of chondrocytes gradually decreases.