Department of Orthopaedics, Biomechanics and Implant Technology Research Laboratory, University of Rostock, Rostock, Germany.
Connect Tissue Res. 2011;52(6):503-11. doi: 10.3109/03008207.2011.593673. Epub 2011 Jul 25.
Healing capacity of cartilage is low. Thus, cartilage defects do not regenerate as hyaline but mostly as fibrous cartilage which is a major drawback since this tissue is not well adapted to the mechanical loading within the joint. During in vitro cultivation in monolayers, chondrocytes proliferate and de-differentiate to fibroblasts. In three-dimensional cell cultures, de-differentiated chondrocytes could re-differentiate toward the chondrogenic lineage and re-express the chondrogenic phenotype. The objective of this study was to characterize the mesenchymal stem cell (MSC) potential of human chondrocytes isolated from articular cartilage. Furthermore, the differentiation capacity of human chondrocytes in three-dimensional cell cultures was analyzed to target differentiation direction into hyaline cartilage. After isolation and cultivation of chondrogenic cells, the expression of the MSC-associated markers: cluster of differentiation (CD)166, CD44, CD105, and CD29 was performed by flow cytometry. The differentiation capacity of human chondrocytes was analyzed in alginate matrix cultured in Dulbecco?s modified eagle medium with (chondrogenic stimulation) and without (control) chondrogenic growth factors. Additionally, the expression of collagen type II, aggrecan, and glycosaminoglycans was determined. Cultivated chondrocytes showed an enhanced expression of the MSC-associated markers with increasing passages. After chondrogenic stimulation in alginate matrix, the chondrocytes revealed a significant increase of cell number compared with unstimulated cells. Further, a higher synthesis rate of glycosaminoglycans and a positive collagen type II and aggrecan immunostaining was detected in stimulated alginate beads. Human chondrocytes showed plasticity whilst cells were encapsulated in alginate and stimulated by growth factors. Stimulated cells demonstrated characteristics of chondrogenic re-differentiation due to collagen type II and aggrecan synthesis.
软骨的自我修复能力很低。因此，软骨损伤不会像透明软骨那样再生，而大多会变成纤维软骨，这是一个主要的缺点，因为这种组织不能很好地适应关节内的机械负荷。在单层培养的体外培养中，软骨细胞增殖并去分化为成纤维细胞。在三维细胞培养中，去分化的软骨细胞可以重新向软骨谱系分化，并重新表达软骨细胞表型。本研究的目的是表征从关节软骨分离的人软骨细胞的间充质干细胞（MSC）潜能。此外，还分析了三维细胞培养中人类软骨细胞的分化能力，以确定向透明软骨分化的方向。在软骨细胞分离和培养后，通过流式细胞术检测 MSC 相关标志物：分化簇（CD）166、CD44、CD105 和 CD29 的表达。在含（软骨生成刺激）和不含（对照）软骨生成生长因子的海藻酸钠基质中培养人软骨细胞，分析其分化能力。此外，还测定了胶原 II 型、聚集蛋白聚糖和糖胺聚糖的表达。培养的软骨细胞随着传代次数的增加，MSC 相关标志物的表达增强。在海藻酸钠基质中的软骨生成刺激后，与未刺激的细胞相比，软骨细胞的细胞数量显著增加。此外，在刺激的海藻酸珠中检测到糖胺聚糖和胶原 II 型和聚集蛋白聚糖的合成率更高的阳性免疫染色。当细胞被包裹在海藻酸盐中并受到生长因子的刺激时，人软骨细胞表现出可塑性。刺激的细胞由于胶原 II 型和聚集蛋白聚糖的合成而表现出软骨再分化的特征。
