Necessity of enzymatic activity of alkaline phosphatase for mineralization of osteoblastic cells
Necessity of enzymatic activity of alkaline phosphatase for mineralization of osteoblastic cells
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DOI：
10.1254/jjp.88.262
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发表时间：
2002-03-01
期刊：
影响因子：
--
通讯作者：
Iida, J
中科院分区：
文献类型：
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作者：
Sugawara, Y;Suzuki, K;Iida, J
碱性磷酸酶（ALP）被认为对骨形成很重要；然而，其作用尚不清楚。在这项研究中，我们研究了 ALP 酶活性和 ALP 蛋白锚定到细胞上对于成骨细胞系 MC3T3-E1 矿化的重要性。当我们在 ALP 活性抑制剂四咪唑存在的情况下培养细胞时，ALP 蛋白的表达水平与对照相似。尽管四咪唑对细胞生长和增加羟脯氨酸积累没有影响，但它减少了骨钙素的产生以及基质中钙和磷酸盐的积累。四咪唑还抑制矿化结节的形成，这可以通过光学显微镜观察并通过 Von Kossa 染色检测。另一方面，当使用磷脂酰肌醇特异性磷脂酶C从细胞膜释放ALP蛋白时，钙化后期基质中羟脯氨酸、钙和磷酸盐的积累没有检测到明显变化，这与形态学发现一致。这些结果清楚地表明，ALP 的酶活性对于 MC3T3-E1 细胞的矿化是必需的，但 ALP 蛋白的存在或 ALP 锚定到细胞上并不是必需的。
Alkaline phosphatase (ALP) is supposed to be important for bone formation; however, its role is not clear. In this study, we examined the importance of enzymatic activity of ALP and anchoring of ALP protein to the cells for mineralization of an osteoblastic cell line, MC3T3-E1. While we cultured the cells in the presence of tetramisole, an inhibitor of ALP activity, ALP protein was expressed at a similar level to that in the control. Although tetramisole showed no effect on cell growth and increased hydroxyproline accumulation, it decreased the osteocalcin production and the accumulation of calcium and phosphate in the matrices. Tetramisole also inhibited mineralized nodule formation, which was observed by optical microscopy and detected by Von Kossa staining. On the other hand, when ALP protein was released from the cell membranes with the use of phosphatidylinositol-specific phospholipase C, no marked changes were detected in hydroxyproline, calcium and phosphate accumulations in the matrices at late calcification stage, which was consistent with the morphological findings. These results clearly show that enzymatic activity of ALP is necessary for mineralization of MC3T3-E1 cells, but not the presence of ALP protein or anchoring of ALP to the cells.