Materials Processing Research Centre, Dublin City University, Glasnevin, Dublin, Ireland.
J Tissue Eng Regen Med. 2012 Apr;6(4):280-90. doi: 10.1002/term.431. Epub 2011 Jun 27.
It is desirable to produce cryopreservable cell-laden tissue-engineering scaffolds whose final properties can be adjusted during the thawing process immediately prior to use. Polyvinyl alcohol (PVA)-based solutions provide platforms in which cryoprotected cell suspensions can be turned into a ready-to-use, cell-laden scaffold by a process of cryogelation. In this study, such a PVA system, with DMSO as the cryoprotectant, was successfully developed. Vascular smooth muscle cell (vSMC)-encapsulated cryogels were investigated under conditions of cyclic strain and in co-culture with vascular endothelial cells to mimic the environment these cells experience in vivo in a vascular tissue-engineering setting. In view of the cytotoxicity DMSO imposes with respect to the production procedure, carboxylated poly-L-lysine (COOH-PLL) was substituted as a non-cytotoxic cryoprotectant to allow longer, slower thawing periods to generate more stable cryogels. Encapsulated vSMC with DMSO as a cryoprotectant responded to 10% cyclic strain with increased alignment and proliferation. Cells were stored frozen for 1 month without loss of viability compared to immediate thawing. SMC-encapsulated cryogels also successfully supported functional endothelial cell co-culture. Substitution of COOH-PLL in place of DMSO resulted in a significant increase in cell viability in encapsulated cryogels for a range of thawing periods. We conclude that incorporation of COOH-PLL during cryogelation preserved cell functionality while retaining fundamental cryogel physical properties, thereby making it a promising platform for tissue-engineering scaffolds, particularly for vascular tissue engineering, or cell preservation within microgels.
理想情况下，应生产可低温保存的细胞负载组织工程支架，其最终性能可在使用前的解冻过程中立即进行调整。聚乙烯醇（PVA）基溶液为可保护细胞的悬浮液提供了平台，可通过冷冻凝胶化过程将其转化为即用型细胞负载支架。在这项研究中，成功开发了一种含有 DMSO 的 PVA 系统作为保护剂。研究了包封血管平滑肌细胞（vSMC）的冷冻凝胶在循环应变下的情况，并与血管内皮细胞共培养，以模拟这些细胞在血管组织工程环境中体内经历的环境。鉴于 DMSO 对生产过程造成的细胞毒性，用羧基化聚赖氨酸（COOH-PLL）替代作为非细胞毒性保护剂，以允许更长、更缓慢的解冻过程来生成更稳定的冷冻凝胶。用 DMSO 作为保护剂包封的 vSMC 对 10%的循环应变表现出更高的对齐和增殖。与立即解冻相比，细胞在冷冻状态下可储存 1 个月而不会失去活力。包封的 vSMC 冷冻凝胶还成功支持功能性内皮细胞共培养。用 COOH-PLL 替代 DMSO 可显著提高包封冷冻凝胶中细胞的活力，延长解冻时间。我们得出结论，在冷冻凝胶化过程中加入 COOH-PLL 可在保留基本冷冻凝胶物理性质的同时保持细胞功能，从而使其成为组织工程支架的有前途的平台，特别是用于血管组织工程或微凝胶内细胞保存。
