Department of Cartilage & Bone Regeneration, Graduate School of Medicine, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-8655, Japan.
Biomaterials. 2012 Jan;33(2):444-54. doi: 10.1016/j.biomaterials.2011.09.056. Epub 2011 Oct 19.
To promote clinical application of cartilage tissue engineering, we should establish a serum-free chondrocyte growth medium. The serum-free medium would increase the cell numbers by more than 20-fold within one week, which proliferation ability almost matches that of serum-based one. For that, we examined the combinations of growth factors and the methods to enhance their effects by making use of the interaction with biomaterials. From various growth factors that are contained within the serum, we made the cocktail of FGF-2 (100 ng/mL), insulin (5 μg/mL), EGF (10 pg/mL), PDGF (625 pg/mL) and TGF-β (5 pg/mL), which increased the chondrocyte numbers by approximately 3-fold for 7 days. Moreover, we used the biomaterials including albumin and hyaluronan as the carrier of those factors. By direct mixing of those factors with biomaterials before the administration to the medium, the medium containing those mixture showed the chondrocyte growth of approximately a 25-fold increase by day 10. In this medium, the FGF-2 or insulin concentration hardly decreased, and rather enhanced the activation of ERK. Due to the optimal usage of biomaterials, this serum-free medium will realize a constant harvest of chondrocytes and could contribute to the safety and quality in regenerative medicine.
为了促进软骨组织工程的临床应用，我们应该建立一个无血清的软骨细胞生长培养基。该无血清培养基可使细胞数量在一周内增加 20 多倍，其增殖能力几乎与基于血清的培养基相当。为此，我们研究了生长因子的组合以及通过利用与生物材料的相互作用来增强其效果的方法。从血清中包含的各种生长因子中，我们制作了 FGF-2（100ng/mL）、胰岛素（5μg/mL）、EGF（10pg/mL）、PDGF（625pg/mL）和 TGF-β（5pg/mL）的鸡尾酒，该鸡尾酒可使软骨细胞数量在 7 天内增加约 3 倍。此外，我们还使用包括白蛋白和透明质酸在内的生物材料作为这些因子的载体。通过在将这些因子直接混合到培养基中之前与生物材料混合，含有这些混合物的培养基在第 10 天显示出约 25 倍的软骨细胞生长增加。在该培养基中，FGF-2 或胰岛素浓度几乎没有下降，反而增强了 ERK 的激活。由于生物材料的最佳使用，这种无血清培养基将实现持续收获软骨细胞，并有助于再生医学的安全性和质量。
