Chondrogenic differentiation of murine C3H10T1/2 multipotential mesenchymal cells: I. Stimulation by bone morphogenetic protein-2 in high-density micromass cultures
Chondrogenic differentiation of murine C3H10T1/2 multipotential mesenchymal cells: I. Stimulation by bone morphogenetic protein-2 in high-density micromass cultures
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DOI：
10.1046/j.1432-0436.1999.6420067.x
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发表时间：
1999-01-01
期刊：
影响因子：
2.9
通讯作者：
Tuan, RS
中科院分区：
文献类型：
--
作者：
Denker, AE;Haas, AR;Tuan, RS
间充质细胞的成软骨分化通常被认为是由特定生长因子的诱导作用启动的，并且依赖于密切的细胞-细胞相互作用。本研究以小鼠C3 H10 T1/2多能细胞为材料，探讨骨形态发生蛋白2（BMP-2）对软骨形成的影响及其作用机制。C3 H10 T1/2细胞先前已显示经历多种分化途径。虽然已经观察到软骨形成、骨形成、肌形成和脂肪形成，但软骨细胞出现的频率明显低于其他细胞类型，并且没有观察到除了其他细胞类型之外的软骨细胞的出现。我们在这里报告，C3 H10 T1/2细胞中的软骨细胞的外观显着增强作为培养条件下有利于软骨形成，即平板接种高密度微团和BMP-2处理的结果。这种培养物含有软骨细胞样细胞，精心制作阿辛蓝染色的软骨样基质，表达连接蛋白和II型胶原蛋白，这两种软骨基质标志物，并显示增加的[S-35]硫酸盐掺入。阿尔新蓝阳性物质的出现和硫酸盐掺入的增加取决于BMP-2的剂量、培养时间和微团培养物的细胞平板接种密度。微团内细胞的分化是软骨形成谱系特异性的，因为碱性磷酸酶染色显示在最高BMP-2浓度下微团中仅有微弱的染色。在BMP-2对高密度C3 H10 T1/2培养物的软骨诱导作用中增强的细胞-细胞相互作用的重要性进一步受到在聚阳离子化合物聚-L-赖氨酸存在下软骨形成的额外促进的影响，聚-L-赖氨酸先前已报道增强胚胎肢体间充质细胞中的细胞相互作用和软骨形成。总之，这些发现表明，C3 H10 T1/2细胞中的软骨形成是由BMP-2诱导的，需要细胞-细胞相互作用。
Chondrogenic differentiation of mesenchymal cells is generally thought to be initiated by the inductive action of specific growth factors and depends on intimate cell-cell interactions. In this study, we have used multipotential murine C3H10T1/2 cells to analyze the effect and mechanism of action of bone morphogenetic protein 2 (BMP-2) on chondrogenesis. C3H10T1/2 cells have been previously shown to undergo multiple differentiation pathways. While chondrogenesis, osteogenesis, myogenesis and adipogenesis have been observed, chondrocytes appear significantly less frequently than the other cell types, and the appearance of chondrocytes exclusive of the other cell types has not been observed. We report here that the appearance of chondrocytes in C3H10T1/2 cells is markedly enhanced as a result of culture under conditions favorable for chondrogenesis, i.e. plating as high-density micromass and treatment with BMP-2. Such cultures contain chondrocyte-like cells, elaborate an Alcian blue stained cartilage-like matrix, express link protein and type II collagen, both cartilage matrix markers, and show increased [S-35]sulfate incorporation. The appearance of Alcian blue positive material and increased sulfate incorporation are dependent on the dose of BMP-2, culture time, and cell plating density of the micromass cultures. Differentiation of cells within the micromass was specific to the chondrogenic lineage, as alkaline phosphatase staining revealed only faint staining in the micromass at the highest BMP-2 concentration. The importance of enhanced cell-cell interaction in the chondroinductive effects of BMP-2 on high-density C3H10T1/2 cultures was further implicated by the additional promotion of chondrogenesis in the presence of the polycationic compound, poly-l-lysine, which has been previously reported to enhance cellular interactions and chondrogenesis in embryonic limb mesenchymal cells. Taken together, these findings suggest that chondrogenesis in C3H10T1/2 cells is inducible by BMP-2 and requires cell-cell interaction.