Department of Orthopaedic Surgery, Shiga University of Medical Science, Otsu, Shiga, Japan.
Acta Orthop. 2009 Dec;80(6):724-33. doi: 10.3109/17453670903413111.
Three-dimensionally (3D-) embedded chondrocytes have been suggested to maintain the chondrocytic phenotype. Furthermore, mechanical stress and growth factors have been found to be capable of enhancing cell proliferation and ECM synthesis. We investigated the effect of mechanical loading and growth factors on reactivation of the 3D-embedded chondrocytes.
Freshly isolated chondrocytes from rat articular cartilage were grown in monolayer cultures and then in collagen gel. Real-time RT-PCR and histological analysis for aggrecan and type II and type I collagen was performed to evaluate their chondrocytic activity. Then, the 3D-embedded chondrocytes were cultured under either mechanical loading alone or in combination with growth factor. The dynamic compression (5% compression, 0.33 Hz) was loaded for 4 durations: 0, 10, 60, and 120 min/day. The growth factor administered was either basic fibroblast growth factor (bFGF) or bone morphogenetic protein-2 (BMP-2).
Mechanical loading statistically significantly reactivated the aggrecan and type II collagen expression with loading of 60 min/day as compared to the other durations. The presence of BMP-2 and bFGF clearly enhanced the aggrecan and type II collagen expression of 3D-embedded chondrocytes. Unlike previous reports using monolayer chondrocytes, however, BMP-2 or bFGF did not augment the chondrocytic phenotype when applied together with mechanical loading.
Dynamic compression effectively reactivated the dedifferentiated chondrocytes in 3D culture. However, the growth factors did not play any synergistic role when applied with dynamic compressive loading, suggesting that growth factors should be administered at different time points during regeneration of the transplantation-ready cartilage.
三维（3D）嵌入的软骨细胞被认为可以维持软骨细胞表型。此外，已发现机械应力和生长因子能够增强细胞增殖和 ECM 合成。我们研究了机械加载和生长因子对 3D 嵌入软骨细胞再激活的影响。
从大鼠关节软骨中分离的新鲜软骨细胞在单层培养物中培养，然后在胶原凝胶中培养。通过实时 RT-PCR 和组织学分析对聚集蛋白聚糖和 II 型和 I 型胶原进行分析，以评估其软骨细胞活性。然后，在单独或结合生长因子的情况下对 3D 嵌入的软骨细胞进行培养。动态压缩（5%压缩，0.33 Hz）加载持续 4 个时间段：0、10、60 和 120 分钟/天。给予的生长因子为碱性成纤维细胞生长因子（bFGF）或骨形态发生蛋白-2（BMP-2）。
与其他时间段相比，60 分钟/天的机械加载可显著重新激活聚集蛋白聚糖和 II 型胶原的表达。BMP-2 和 bFGF 的存在明显增强了 3D 嵌入软骨细胞的聚集蛋白聚糖和 II 型胶原表达。然而，与使用单层软骨细胞的先前报告不同，当与机械加载一起应用时，BMP-2 或 bFGF 并未增强软骨细胞表型。
动态压缩可有效重新激活 3D 培养中的去分化软骨细胞。然而，当应用于动态压缩加载时，生长因子没有发挥任何协同作用，这表明在移植准备好的软骨再生过程中，应在不同时间点给予生长因子。
