Isolation and characterization of calcium-accumulating matrix vesicles from chondrocytes of chicken epiphyseal growth plate cartilage in primary culture.
Isolation and characterization of calcium-accumulating matrix vesicles from chondrocytes of chicken epiphyseal growth plate cartilage in primary culture.
复制标题
DOI：
--
复制
发表时间：
1985-12
期刊：
影响因子：
--
通讯作者：
R. Wuthier;J. Chin;J. Hale;T. Register;L. Hale;Y. Ishikawa
中科院分区：
文献类型：
--
作者：
R. Wuthier;J. Chin;J. Hale;T. Register;L. Hale;Y. Ishikawa
从原代培养的鸡生长板肥大软骨细胞的培养液中可以很容易地分离到基质小泡(MV)。在整个培养过程中，软骨细胞保持正常形态并合成II型胶原。培养得到的MV在形态上与原位观察到的MV难以区分，并且富含碱性磷酸酶。富含碱性磷酸酶的MV的形成受培养阶段的影响很大：细胞接种后不久大量释放；在细胞扩展和快速分裂过程中显著下降；当细胞达到融合时，富碱性磷酸酶MV的产生显著回升。增加初始软骨细胞种植密度会成比例地增加MV的产量。肥大区的细胞比增殖区的细胞更能形成富含碱性磷酸酶的MV，说明MV的形成依赖于细胞的分化。将培养的软骨细胞释放的MV与从相同组织中获得的质膜部分和胶原酶释放的MV进行比较。蛋白质的电泳图和磷脂图谱表明，在MV形成过程中，质膜发生了显著的变化。当供应碱性磷酸酶底物时，这些囊泡能够从亚稳态合成软骨淋巴中积累大量矿物质离子。因此，该培养系统似乎是分离天然MV和鉴定囊泡形成和矿化所需因素的有用模型。
Matrix vesicles (MV) can be readily isolated from culture media of chicken growth plate hypertrophic chondrocytes grown in primary culture. The chondrocytes maintain normal morphology and synthesize type II collagen throughout the culture period. The culture-derived MV are morphologically indistinguishable from MV seen in situ and are rich in alkaline phosphatase. Formation of alkaline phosphatase-rich MV is strongly influenced by the stage of culture: large numbers are released shortly after cell seeding; marked decline is seen during cell spreading and rapid cell division; notable resurgence in alkaline phosphatase-rich MV production occurs as the cells attain confluency. Increasing the initial chondrocyte seeding density proportionately increases MV production. Cells derived from the hypertrophic region are much more capable of forming alkaline phosphatase-rich MV than those from the proliferating zone, indicating that MV formation is dependent on cellular differentiation. MV released by the cultured chondrocytes were compared in protein and phospholipid composition and in their ability to accumulate mineral ions, with plasma membrane fractions and collagenase-released MV obtained from the same tissue. Electrophoretic patterns of proteins, and the phospholipid profiles, suggest that significant modification of the plasma membrane occurs during MV formation. The vesicles are capable of accumulating large amounts of mineral ions from a metastable synthetic cartilage lymph when supplied with alkaline phosphatase substrates. This culture system thus appears to be a useful model for isolating native MV and characterizing factors required for vesicle formation and mineralization.