Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype
Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype
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DOI：
10.1007/s00441-010-1120-3
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发表时间：
2011-01
影响因子：
3.6
通讯作者：
Li‐an Wu;Junsheng Feng;Lynn Wang;Y. Mu;Andrew Baker;K. Donly;S. Harris;M. MacDougall;Shuo Chen
中科院分区：
文献类型：
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作者：
Li‐an Wu;Junsheng Feng;Lynn Wang;Y. Mu;Andrew Baker;K. Donly;S. Harris;M. MacDougall;Shuo Chen
骨形态发生蛋白2（Bmp 2）是成骨细胞分化和成骨所必需的。Bmp 2成骨细胞系的建立为研究Bmp 2对成骨细胞分化的影响及其在骨骼代谢过程中的信号通路提供了有价值的工具。由于原代成骨细胞的来源相对有限，我们已经开发了细胞系，作为良好的替代模型的成骨细胞分化和骨矿化的研究。在这项研究中，我们建立了永生化的小鼠成骨细胞系Bmp 2。用pSV 3-neo转染原代小鼠floxedBmp 2成骨细胞并进行克隆选择。经PCR和免疫组化证实转染细胞。为了确定永生化细胞的基因型和表型，分析细胞形态、增殖、分化和矿化。采用定量RT-PCR和免疫组织化学方法检测成骨细胞相关基因Runx 2、Osx、ATF 4、Dlx 3、骨涎蛋白、牙本质基质蛋白1、骨连接素、骨钙素和骨桥蛋白的表达。这些结果表明，永生化的Bmp 2成骨细胞具有更高的增殖率，但保留了其与原代细胞相似的基因型和表型特征。因此，我们首次描述了永生化小鼠成骨细胞系Bmp 2的发展，并为研究BMP 2介导的成骨细胞生物学及其下游信号转导通路提供了一个有用的模型。
Bone morphogenetic protein 2 (Bmp2) is essential for osteoblast differentiation and osteogenesis. Generation of floxedBmp2osteoblast cell lines is a valuable tool for studying the effects ofBmp2on osteoblast differentiation and its signaling pathways during skeletal metabolism. Due to relatively limited sources of primary osteoblasts, we have developed cell lines that serve as good surrogate models for the study of osteoblast cell differentiation and bone mineralization. In this study, we established and characterized immortalized mouse floxedBmp2osteoblast cell lines. Primary mouse floxedBmp2osteoblasts were transfected with pSV3-neo and clonally selected. These transfected cells were verified by PCR and immunohistochemistry. To determine the genotype and phenotype of the immortalized cells, cell morphology, proliferation, differentiation and mineralization were analyzed. Also, expression of osteoblast-related gene markers includingRunx2,Osx,ATF4,Dlx3, bone sialoprotein, dentin matrix protein 1, osteonectin, osteocalcin and osteopontin were examined by quantitative RT-PCR and immunohistochemistry. These results showed that immortalized floxedBmp2osteoblasts had a higher proliferation rate but preserved their genotypic and phenotypic characteristics similar to the primary cells. Thus, we, for the first time, describe the development of immortalized mouse floxedBmp2osteoblast cell lines and present a useful model to study osteoblast biology mediated byBMP2and its downstream signaling transduction pathways.