1 Department of Orthopedics, University Medical Center Utrecht , Utrecht, The Netherlands .
2 Institute of Biological Chemistry, Department of Biophysics and Bioengineering, School of Engineering and Physical Sciences, Heriot-Watt University , Edinburgh, United Kingdom .
Tissue Eng Part C Methods. 2018 Apr;24(4):222-232. doi: 10.1089/ten.TEC.2017.0226. Epub 2018 Apr 2.
Hydrogels can facilitate nucleus pulposus (NP) regeneration, either for clinical application or research into mechanisms of regeneration. However, many different hydrogels and culture conditions for human degenerated NP have been employed, making literature data difficult to compare. Therefore, we compared six different hydrogels of natural polymers and investigated the role of serum in the medium and of osmolarity during expansion or redifferentiation in an attempt to provide comparators for future studies. Human NP cells of Thompson grade III discs were cultured in alginate, agarose, fibrin, type II collagen, gelatin methacryloyl (gelMA), and hyaluronic acid-poly(ethylene glycol) hydrogels. Medium containing fetal bovine serum and a serum-free (SF) medium were compared in agarose, gelMA, and type II collagen hydrogels. Isolation and expansion of NP cells in low compared to high osmolarity medium were performed before culture in agarose and type II collagen hydrogels in media of varying osmolarity. NP cells in agarose produced the highest amounts of proteoglycans, followed by cells in type II collagen hydrogels. The absence of serum reduced the total amount of proteoglycans produced by the cells, although incorporation efficiency was higher in type II collagen hydrogels in the absence than in the presence of serum. Isolation and expansion of NP cells in high osmolarity medium improved proteoglycan production during culture in hydrogels, but variation in osmolarity during redifferentiation did not have any effect. Agarose hydrogels seem to be the best option for in vitro culture of human NP cells, but for clinical application, type II collagen hydrogels may be better because, as opposed to agarose, it degrades in time. Although culture in SF medium reduces the amount of proteoglycans produced during redifferentiation culture, isolating and expanding the cells in high osmolarity medium can largely compensate for this loss.
水凝胶可以促进髓核（NP）再生，无论是用于临床应用还是研究再生机制。然而，已经使用了许多不同的水凝胶和人退变 NP 的培养条件，使得文献数据难以比较。因此，我们比较了六种不同的天然聚合物水凝胶，并研究了血清在培养基中的作用以及渗透压在扩张或再分化过程中的作用，试图为未来的研究提供比较器。人 NP 细胞取自 Thompson 分级 III 椎间盘，在藻酸盐、琼脂糖、纤维蛋白、II 型胶原、明胶甲基丙烯酰（gelMA）和透明质酸-聚（乙二醇）水凝胶中进行培养。在琼脂糖、gelMA 和 II 型胶原水凝胶中比较了含有胎牛血清的培养基和无血清（SF）培养基。在琼脂糖和 II 型胶原水凝胶中，在不同渗透压的培养基中培养之前，先在低渗透压培养基中分离和扩增 NP 细胞。琼脂糖中的 NP 细胞产生的蛋白聚糖最多，其次是 II 型胶原水凝胶中的细胞。无血清减少了细胞产生的蛋白聚糖总量，但在无血清存在时，II 型胶原水凝胶中的掺入效率更高。在高渗透压培养基中分离和扩增 NP 细胞可改善水凝胶培养过程中的蛋白聚糖产生，但再分化过程中渗透压的变化没有任何影响。琼脂糖水凝胶似乎是体外培养人 NP 细胞的最佳选择，但对于临床应用，II 型胶原水凝胶可能更好，因为与琼脂糖不同，它会随时间降解。虽然 SF 培养基中的培养会减少再分化培养过程中产生的蛋白聚糖量，但在高渗透压培养基中分离和扩增细胞可以在很大程度上弥补这种损失。
