Objective To study the expression of the pyrophosphoric acid loop's key enzymes (ENPP1, TNAP and ANK proteins) in the degeneration of rat endplate chondrocyte. Methods The method of enzyme digestion plus natural subculture was employed. And the primary to forth-generation cells were collected. Western blot was used to analyze the expression of ENPP1, TNAP and ANK proteins in each generation of endplate chondrocytes. Results The expression of ENPP1, TNAP and ANK proteins could be detected in each generation of endplate chondrocytes. With the continuing passage of cells, the level of protein expression declined gradually and that of the third and forth-generation decreased. Conclusion While studying the correlation between the degeneration of rat endplate chondrocytes and pyrophosphoric acid loop,it is more significant to choose the cells before the third generation. Meanwhile it is possible to lower the incidence of intervertebral discs degeneration by controlling the expression of the above-mentioned proteins in endplate cartilage.%目的 观察大鼠终板软骨细胞体外自然退变过程中焦磷酸环路关键酶(ENPP1、TNAP、ANK蛋白)表达的变化.方法 采用连续酶消化及自然传代法,选取原代至第四代终板软骨细胞,用免疫印迹方法分析各代终板软骨细胞中ENPP1、TNAP、ANK蛋白的表达.结果 ENPP1、TNAP、ANK蛋白在各代终板软骨细胞中均有表达;随着细胞的传代,蛋白表达量逐渐减少,第三、四代的表达微弱,ENPP1、TNAP、ANK蛋白在第三、四代的光密度值和内参Actin表达条带光密度值相比差异有统计学意义(P＜0.05).结论 该实验提示我们对终板细胞自然退变过程中焦磷酸环路关键蛋白进行研究时,选取第三代细胞以前的意义较大;从分子水平对软骨终板退变的发生机制进行研究,通过调控软骨终板中上述蛋白的表达,可能有助于遏制椎间盘退变的发生,为防治椎间盘退变确立新的有效靶点.
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