Phenotypic stability of bovine articular chondrocytes after long-term culture in alginate beads.
Phenotypic stability of bovine articular chondrocytes after long-term culture in alginate beads.
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发表时间：
1994
影响因子：
4
通讯作者：
Hans J. Häuselmann;Russell J. Fernandes;Su S. Mok;Thomas M. Schmid;Joel A. Block;M. Aydelotte;Klaus E. Kuettner;E. Thonar
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作者：
Hans J. Häuselmann;Russell J. Fernandes;Su S. Mok;Thomas M. Schmid;Joel A. Block;M. Aydelotte;Klaus E. Kuettner;E. Thonar
嵌入藻酸盐凝胶的关节软骨细胞会重新生成富含胶原和蛋白聚糖的基质。这种培养系统的一个主要优点是细胞可以通过螯合钙来恢复，否则会使海藻酸盐保持凝胶状态。因此释放的软骨细胞被紧密结合的细胞相关基质包围，这似乎与电子显微镜下软骨中发现的细胞周基质和区域基质相对应。通过轻度离心可以很容易地将细胞及其相关基质从主要来自“区域间”基质的更可溶性的基质成分中分离出来。因此，这种新的细胞培养系统使得研究存在于两个不同基质池中的分子的组装和周转成为可能。重要的是，可以使用非变性溶剂提取这两个池中的大部分聚集蛋白分子，从而使研究天然蛋白多糖聚集体的代谢和周转成为可能。我们在这篇报告中表明，从成年牛关节软骨的全深度中分离出来的软骨细胞，在海藻酸盐凝胶中维持8个月后，仍然具有代谢活性，并继续合成软骨特异性的II型胶原蛋白和聚集蛋白。当软骨细胞失去其表型稳定性时，细胞不会合成大量的I型胶原蛋白或小的非聚集性蛋白聚糖。经过长时间的培养后，细胞呈现为两个群体，在大小，形状和周围细胞外基质的数量上表现出差异。第一个群体只在头部表面附近发现：这些细胞被扁平，周围是稀疏的蛋白聚糖和胶原原纤维基质。第二个种群分布在头部的剩余深度：细胞更圆，几乎总是被一个篮子状的网状结构包围，网状结构由密集排列的原纤维组成，与表面相切。
Articular chondrocytes embedded in alginate gel produce de novo a matrix rich in collagens and proteoglycans. A major advantage of this culture system is that the cells can be recovered by chelating the calcium, which otherwise maintains the alginate in its gel state. Chondrocytes thus released are surrounded by tightly bound cell-associated matrix, which seems to correspond to the pericellular and territorial matrices identified in cartilage by electron microscopy. The cells and their associated matrix can be easily separated by mild centrifugation from more soluble matrix components derived principally from the 'interterritorial' matrix. This new cell culture system thus makes it possible to study the assembly and turnover of molecules present in two distinct matrix pools. Importantly, a significant proportion of the aggrecan molecules in each of these two pools can be extracted using a non-denaturing solvent, thereby making possible studies of the metabolism and turnover of native proteoglycan aggregates. We show in this report that chondrocytes isolated from the full depth of adult bovine articular cartilage and maintained for 8 months in alginate gel are still metabolically active and continue to synthesize cartilage-specific type II collagen and aggrecan. The cells did not synthesize large amounts of type I collagen or of the small nonaggregating proteoglycans as usually occurs when chondrocytes lose their phenotypic stability. After this extended period of time in culture, the cells were present as two populations exhibiting differences in size, shape and amount of extracellular matrix surrounding them. The first population was found only near the surface of the bead: these cells were flattened and surrounded by a matrix sparse in proteoglycans and collagen fibrils. The second population was found throughout the remaining depth of the bead: the cells were more round and almost always surrounded by a basket-like meshwork consisting of densely packed fibrils running tangential to the surface.