Establishment of an osteoid preosteocyte-like cell MLO-A5 that spontaneously mineralizes in culture
Establishment of an osteoid preosteocyte-like cell MLO-A5 that spontaneously mineralizes in culture
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DOI：
10.1359/jbmr.2001.16.9.1622
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发表时间：
2001-09-01
影响因子：
6.2
通讯作者：
Bonewald, LF
中科院分区：
文献类型：
--
作者：
Kato, Y;Boskey, K;Bonewald, LF
控制骨基质矿化起始的机制尚不清楚。为了检验该过程，我们建立了称为NMO-A5的细胞系，其在β-甘油磷酸盐（β-GP）和抗坏血酸存在下在培养3天内矿化成薄片，而不是结节，并且在β-GP和抗坏血酸不存在下在培养7天内矿化。在这两种情况下形成的矿物被证明是骨样磷灰石的傅立叶变换红外光谱（FTIR）。从FTIR数据计算的与灰分重量直接相关的矿物-基质比（min/基质）在不存在β-GP和抗坏血酸的情况下约为0.4，在存在β-GP和抗坏血酸的情况下约为1.2。相比之下，在没有β-GP的胎鼠头盖骨细胞中这些比率等于0，而在有β-GP的胎鼠头盖骨细胞中这些比率为1.9.从表达由骨钙蛋白启动子驱动的大T抗原的转基因小鼠的长骨中分离出该细胞系和其他三个细胞系（MLO-A2、-D1和-D 6），所述转基因小鼠与分离出骨细胞样细胞系NMO-Y 4的小鼠相同。((1))基于树突状或星状形态选择细胞系。NMO-A5细胞表达高碱性磷酸酶、1型胶原蛋白、甲状旁腺激素/甲状旁腺激素相关肽（PTH/PTHrP）受体、骨唾液蛋白（BSP）和骨钙素（767 ng/10（6）细胞，而原代小鼠成骨细胞和5种成骨细胞系< 1-2.2 ng/10（6）细胞）。与其他三种非矿化细胞系相比，MLO-A5细胞的唯一独特特征是BSP信使RNA（mRNA）的高表达。这些细胞系可能代表骨细胞分化的阶段，MLO-A5细胞代表后成骨细胞，负责触发类骨质矿化的前骨细胞。
The mechanisms controlling the initiation of mineralization of bone matrix are not clear. To examine this process, we established a cell line called NMO-A5 that mineralizes in sheets, not nodules, within 3 days of culture in the presence of beta -glycerophosphate (beta -GP) and ascorbic acid and within 7 days in the absence of beta -GP and ascorbic acid. The mineral formed in both cases was shown to be bonelike apatite by Fourier transformed infrared (FTIR) spectroscopy. Mineral-to-matrix ratios (min/matrix) calculated from the FTIR data, which are related directly to ash weight, were approximately 0.4 in the absence of beta -GP and ascorbic acid and approximately 1.2 in the presence of beta -GP and ascorbic acid. By comparison, these ratios in fetal rat calvarial cells without beta -GP equal 0 and with beta -GP 1.9. This cell line and three others (MLO-A2, -D1, and -D6) were isolated from the long bones of transgenic mice expressing the large T-antigen driven by the osteocalcin promoter, the same mice from which the osteocyte-like cell line NMO-Y4 was isolated.((1)) The cell lines were selected based on a dendritic or stellate morphology. NMO-A5 cells express high alkaline phosphatase, collagen type 1, parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) receptor, bone sialoprotein (BSP), and osteocalcin (767 ng/10(6) cells compared with < 1-2.2 ng/10(6) cell for primary mouse osteoblasts and five osteoblast cell lines). The single unique feature of the MLO-A5 cells compared with the other three nonmineralizing cell lines is the high expression of messenger RNA (mRNA) for BSP. These cell lines may represent stages of osteocyte differentiation and the MLO-A5 cells represent the postosteoblast, preosteocyte responsible for triggering mineralization of osteoid.