Kolettas E, Buluwela L, Bayliss M T, Muir H I
Department of Biochemistry, Charing Cross and Westminster Medical School, University of London, UK.
J Cell Sci. 1995 May;108 ( Pt 5):1991-9. doi: 10.1242/jcs.108.5.1991.
Normal human adult articular chondrocytes were used to determine how the chondrocyte phenotype is modulated by culture conditions following long-term culture. We report here for the first time that human articular chondrocytes have a lifespan in the range of 34-37 population doublings. While chondrocytes cultured as monolayers displayed a fibroblastoid morphology and grew faster, those cultured as suspensions over agarose adopted a round morphology and formed clusters of cells reminiscent of chondrocyte differentiation in intact cartilage, with little or no DNA synthesis. These morphologies were independent of the age of the culture. Despite, these morphological differences, however, chondrocytes expressed markers at mRNA and protein levels characteristic of cartilage: namely, types II and IX collagens and the large aggregating proteoglycans, aggrecan, versican and link protein, but not syndecan, under both culture conditions. However, they also expressed type I collagen alpha 1(I) and alpha 2(I) chains. It has been suggested that expression of collagen alpha 1(I) by chondrocytes cultured as monolayers is a marker of the loss of the chondrocyte phenotype. However, we show here, using reverse transcriptase/polymerase chain reaction, that normal fresh intact human articular cartilage expresses collagen alpha 1(I). The data show that following long-term culture human articular chondrocytes retain their differentiated characteristics and that cell shape does not correlate with the expression of the chondrocyte phenotype. It is proposed that loss of the chondrocyte phenotype is marked by the loss of one or more cartilage-specific molecules rather than by the appearance of non-cartilage-specific molecules.
使用正常人类成年关节软骨细胞来确定长期培养后培养条件如何调节软骨细胞表型。我们首次在此报告，人类关节软骨细胞的寿命在34 - 37次群体倍增范围内。单层培养的软骨细胞呈现成纤维细胞样形态且生长更快，而在琼脂糖上悬浮培养的软骨细胞则呈现圆形形态并形成细胞簇，这让人联想到完整软骨中的软骨细胞分化，且几乎没有或没有DNA合成。这些形态与培养的代数无关。然而，尽管存在这些形态差异，但在两种培养条件下，软骨细胞在mRNA和蛋白质水平均表达软骨特征性标志物：即II型和IX型胶原蛋白以及大型聚集蛋白聚糖、聚集蛋白聚糖、多功能蛋白聚糖和连接蛋白，但不表达syndecan。然而，它们也表达I型胶原蛋白α1（I）和α2（I）链。有人提出，单层培养的软骨细胞表达胶原蛋白α1（I）是软骨细胞表型丧失的标志物。然而，我们在此使用逆转录酶/聚合酶链反应表明，正常新鲜完整的人类关节软骨表达胶原蛋白α1（I）。数据表明，长期培养后人类关节软骨细胞保留其分化特征，且细胞形状与软骨细胞表型的表达无关。有人提出，软骨细胞表型的丧失以一种或多种软骨特异性分子的丧失为标志，而非以非软骨特异性分子的出现为标志。
