Lin Zhen, Fitzgerald Jonathan B, Xu Jiake, Willers Craig, Wood David, Grodzinsky Alan J, Zheng Ming H
Centre of Orthopaedic Surgery, University of Western Australia, M Block, QEII Medical Centre, Monash Avenue, Nedlands, Western Australia 6009, Australia.
J Orthop Res. 2008 Sep;26(9):1230-7. doi: 10.1002/jor.20523.
Chondrocyte phenotype has been shown to dedifferentiate during passaged monolayer cultivation. Hence, we have investigated the expression profile of 27 chondrocyte-associated genes from both osteoarthritic cartilage tissue and healthy passaged human articular chondrocytes by quantitative real-time PCR. Our results indicate that the gene expression levels of matrix proteins and proteases in chondrocytes from monolayer culture decrease compared with those from cartilage tissue, while monolayer cultured chondrocytes from normal and osteoarthritic cartilage exhibit similar gene expression patterns. However, chondrocytic gene expression profiles were differentially altered at various stages of passage. The expression of the matrix proteins aggrecan, type II collagen, and fibromodulin inversely correlated with increasing passage number, while fibronectin and link protein exhibited a marked increase with passage. The expression of matrix proteinases MMP-3/9/13 and ADAMTS-4/5 decreased with passage, whereas proteinase inhibitors TIMP-2/3 were elevated. The cytokine IL-1 also showed increased expression with monolayer chondrocyte culture, while IGF-1 expression levels were diminished. No significant changes in TGF-beta, or the chondrogenic transcription factors Sox-9, c-fos, or c-jun were observed. Our data indicates that cultured chondrocytes undergo dedifferentiation during monolayer culture, although the gene expression level of transcription factors necessary for chondrogenesis remains unchanged. This data may prove important for the future development of more specific and efficacious cultivation techniques for human articular chondrocyte-based therapies.
软骨细胞表型已被证明在传代单层培养过程中会去分化。因此，我们通过定量实时PCR研究了骨关节炎软骨组织和健康传代人关节软骨细胞中27个软骨细胞相关基因的表达谱。我们的结果表明，与软骨组织中的软骨细胞相比，单层培养的软骨细胞中基质蛋白和蛋白酶的基因表达水平降低，而来自正常和骨关节炎软骨的单层培养软骨细胞表现出相似的基因表达模式。然而，软骨细胞基因表达谱在传代的不同阶段有不同的变化。基质蛋白聚集蛋白聚糖、II型胶原蛋白和纤调蛋白的表达与传代次数增加呈负相关，而纤连蛋白和连接蛋白则随传代显著增加。基质蛋白酶MMP-3/9/13和ADAMTS-4/5的表达随传代而降低，而蛋白酶抑制剂TIMP-2/3则升高。细胞因子IL-1在单层软骨细胞培养中也显示出表达增加，而IGF-1表达水平则降低。未观察到TGF-β或软骨形成转录因子Sox-9、c-fos或c-jun有显著变化。我们的数据表明，培养的软骨细胞在单层培养过程中会发生去分化，尽管软骨形成所需转录因子的基因表达水平保持不变。这些数据可能对基于人关节软骨细胞的疗法更具特异性和有效性的培养技术的未来发展具有重要意义。
