SV40‐immortalization of rabbit articular chondrocytes: Alteration of differentiated functions
SV40‐immortalization of rabbit articular chondrocytes: Alteration of differentiated functions
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DOI：
10.1002/jcp.1041500121
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发表时间：
1992-01
影响因子：
5.6
通讯作者：
S. Thenet;P. Benya;S. Demignot;J. Feunteun;M. Adolphe
中科院分区：
文献类型：
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作者：
S. Thenet;P. Benya;S. Demignot;J. Feunteun;M. Adolphe
用编码SV 40早期功能基因的质粒转染兔关节软骨细胞，建立细胞系。这导致细胞永生化（最老的细胞系已完成130代），并获得部分转化的特征，例如降低正常和克隆生长的血清需求。永生化的软骨细胞，称为SVRAC，并没有形成多层病灶时，保持在融合培养。与正常软骨细胞相比，它们在软琼脂中形成集落的能力没有增加，但它们在裸鼠中具有弱致瘤性。SVRAC失去了合成II型胶原和阿辛蓝-胶原基质的能力，这是分化的软骨细胞表型的标志物，并主要合成I型胶原。胶原基因表达的研究表明，proα1（II）mRNA是检测不到的，而proα1（I）胶原mRNA的表达，即使在后期传代培养。与正常的去分化软骨细胞不同，SVRAC不能在三维培养或微丝解聚反应中重新表达分化的表型。在原代培养中或刚从软骨释放细胞后从转染的软骨细胞获得的细胞系显示出相同的行为。因此，SV 40早期基因能够使兔关节软骨细胞永生化，但所得细胞系显示出明显不可逆的去分化表型。这些细胞系可用作模型以鉴定维持或重新表达软骨细胞中分化功能所需的调节途径。
Cell lines were established from rabbit articular chondrocytes following transfection with a plasmid encoding SV40 early function genes. This resulted in cell immortalization (130 passages have been completed for the oldest cell line) with acquisition of characteristics of partial transformation such as reduced serum requirements for normal and clonal growth. The immortalized chondrocytes, called SVRAC, did not form multilayer foci when maintained in postconfluent culture. Their ability to form colonies in soft agar was not increased in comparison with normal chondrocytes, but they were weakly tumorigenic in nude mice. SVRAC lost the ability to synthesize type II collagen and Alcian blue–stainable matrix, which are markers of the differentiated chondrocyte phenotype, and synthesized predominantly type I collagen. Studies of collagen gene expression showed that proα1 (II) mRNA was undetectable, whereas proα1 (I) collagen mRNA was expressed even in late passage cultures. Unlike normal dedifferentiated chondrocytes, SVRAC were unable to re‐express the differentiated phenotype in response to tridimensional culture or microfilament depolymerization. Cell lines obtained from chondrocytes transfected either in primary culture or just after release of cells from cartilage displayed the same behaviour. Thus SV40 early genes were able to immortalize rabbit articular chondrocytes, but the resulting cell lines displayed an apparently irreversibly dedifferentiated phenotype. These cell lines can be used as models to identify regulatory pathways that are required for the maintenance or reexpression of differentiated function in chondrocytes.