Ahmed Nazish, Gan Lu, Nagy Andras, Zheng Jianing, Wang Chen, Kandel Rita A
CIHR-BioEngineering of Skeletal Tissues Team, Mount Sinai Hospital, University of Toronto, Toronto, Canada.
Tissue Eng Part A. 2009 Mar;15(3):665-73. doi: 10.1089/ten.tea.2008.0004.
Articular cartilage has limited ability for repair when damaged by trauma or degenerative disease, such as osteoarthritis, which can result in pain and compromised quality of life. Biological surface replacements developed using tissue engineering methods are a promising approach for cartilage repair, which would avoid the need for total joint replacement with the synthetic implants used currently. A basic requirement of in vitro tissue generation is a supply of sufficient number of cells, which are difficult to acquire from sparsely cellular cartilage tissue. Previously, we have shown that coculture of in vitro-expanded dedifferentiated chondrocytes (P2) with small numbers of primary chondrocytes (P0) induces redifferentiation in passaged (P2) cells. In this study we show that this redifferentiation is not a transient change. After 4 weeks of coculture, the P0 and P2 cells were separated by flow-associated cell sorting, and the redifferentiated P2 (dP2) were cultured alone for a further 4 weeks. The redifferentiated dP2 cells formed thicker cartilage tissue compared to the tissue generated by P2 cells. The newly formed tissue contained type II collagen as demonstrated by immunohistochemical staining and accumulated more proteoglycan per cell than the tissue formed by P2 cells. The dP2 cells also exhibited higher type II collagen and lower type I collagen gene expression than the P2 cells. Interestingly, dP2 cells were able to exert the same effect as P0 cells when cocultured with P2 cells. In conclusion, under proper culture conditions, redifferentiated passaged chondrocytes behave similarly to primary chondrocytes. This coculture system approach can be used to increase the number of differentiated chondrocytes that can be obtained by classical monolayer cell expansion and represents a novel way to acquire sufficient cell numbers for cartilage tissue engineering.
关节软骨在受到创伤或退行性疾病(如骨关节炎)损伤时,修复能力有限,这可能导致疼痛并降低生活质量。利用组织工程方法开发的生物表面替代物是软骨修复的一种有前景的方法,可避免使用目前的合成植入物进行全关节置换。体外组织生成的一个基本要求是提供足够数量的细胞,而从细胞稀少的软骨组织中获取这些细胞很困难。此前,我们已经表明,体外扩增的去分化软骨细胞(P2)与少量原代软骨细胞(P0)共培养可诱导传代(P2)细胞重新分化。在本研究中,我们表明这种重新分化不是一种短暂的变化。共培养4周后,通过流式细胞分选将P0和P2细胞分离,重新分化的P2(dP2)细胞再单独培养4周。与P2细胞生成的组织相比,重新分化的dP2细胞形成了更厚的软骨组织。免疫组织化学染色显示,新形成的组织含有II型胶原蛋白,且每个细胞积累的蛋白聚糖比P2细胞形成的组织更多。dP2细胞的II型胶原蛋白基因表达也高于P2细胞,而I型胶原蛋白基因表达则低于P2细胞。有趣的是,当与P2细胞共培养时,dP2细胞能够发挥与P0细胞相同的作用。总之,在适当的培养条件下,重新分化的传代软骨细胞的行为与原代软骨细胞相似。这种共培养系统方法可用于增加通过经典单层细胞扩增获得的分化软骨细胞数量,代表了一种获取足够细胞数量用于软骨组织工程的新方法。