Regulation of type-II collagen gene expression during human chondrocyte de-differentiation and recovery of chondrocyte-specific phenotype in culture involves Sry-type high-mobility-group box (SOX) transcription factors
Regulation of type-II collagen gene expression during human chondrocyte de-differentiation and recovery of chondrocyte-specific phenotype in culture involves Sry-type high-mobility-group box (SOX) transcription factors
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DOI：
10.1042/0264-6021:3600461
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发表时间：
2001-12-01
影响因子：
4.1
通讯作者：
Jimenez, SA
中科院分区：
文献类型：
--
作者：
Stokes, DG;Liu, G;Jimenez, SA
在作为单层培养物的离体生长期间，软骨细胞增殖并经历去分化过程。这个过程涉及形态学的变化和从软骨细胞特异性基因表达到成纤维细胞中正常表达的基因表达的变化。将单层软骨细胞培养物转移到三维培养系统诱导细胞重新获得软骨细胞特异性表型并在体外产生软骨样组织。我们研究了体外去分化和再分化过程中的控制机制。通过形态学、软骨细胞特异性mRNA的逆转录酶PCR、Western-blot分析和软骨细胞特异性启动子活性测定，去分化软骨细胞在聚甲基丙烯酸2-羟乙酯（polyHEMA）上培养时重新获得其软骨细胞特异性表型。基本上，当将在塑料上培养4周的细胞转移到polyHEMA上培养时，观察到软骨细胞特异性表型的完全恢复。然而，在随后的塑料上传代后，表型恢复不完全或没有发生。含有来自人II型胶原蛋白基因（COL 2A 1）的启动子和增强子的基因报告构建体的活性通过培养条件进行调节，使得其转录活性在单层培养物中被抑制，并且当细胞切换到polyHEMA培养物时在一定程度上被拯救。Sry型高迁移率族盒（SOX）转录因子与增强子区域的结合受到培养条件的调节，SOX 9的mRNA水平也受到调节。一种被感染的人类类型。-通过SOX转录因子的异位表达在去分化细胞中激活II型胶原报告基因构建体。这些结果强调了软骨细胞在组织培养塑料基质上单层培养时表型的明显变化。
During ex vivo growth as monolayer cultures, chondrocytes proliferate and undergo a process of de-differentiation. This process involves a change in morphology and a change from expression of chondrocyte-specific genes to that of genes that are normally expressed in fibroblasts. Transfer of the monolayer chondrocyte culture to three-dimensional culture systems induces the cells to re-acquire a chondrocyte-specific phenotype and produce a cartilaginous-like tissue in vitro. We investigated mechanisms involved in the control of the de-differentiation and re-differentiation process in vitro. De-differentiated chondrocytes re-acquired their chondrocyte-specific phenotype when cultured on poly-(2-hydroxyethyl methacrylate) (polyHEMA) as assayed by morphology, reverse transcriptase PCR of chondrocyte-specific mRNA, Western-blot analysis and chondrocyte-specific promoter activity. Essentially, full recovery of the chondrocyte-specific phenotype was observed when cells that had been cultured for 4 weeks on plastic were transferred to culture on polyHEMA. However, after subsequent passages on plastic, the phenotype recovery was incomplete or did not occur. The activity of a gene reporter construct containing the promoter and enhancer from the human type-II collagen gene (COL2A1) was modulated by the culture conditions, so that its transcriptional activity was repressed in monolayer cultures and rescued to some extent when the cells were switched to polyHEMA cultures. The binding of Sry-type high-mobility-group box (SOX) transcription factors to the enhancer region was modulated by the culture conditions, as were the mRNA levels for SOX9. A transfected human type.-Il collagen reporter construct was activated in dedifferentiated cells by ectopic expression of SOX transcription factors. These results underscore the overt change in phenotype that occurs when chondrocytes are cultured as monolayers on tissue-culture plastic substrata.