Department of Plastic and Reconstructive Surgery, Shanghai 9th People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Tissue Engineering, 639 Zhi Zao Ju Road, Shanghai 200011, PR China.
Biomaterials. 2011 Nov;32(33):8426-35. doi: 10.1016/j.biomaterials.2011.07.008. Epub 2011 Sep 1.
Tissue-engineered epidermal membranes are useful for clinical wound healing. To facilitate these products in the clinic, optimized storage methods need to be developed. We studied the efficiency of extracellular trehalose at various concentrations for cryopreserving human tissue-engineered epidermal membranes compared with that of dimethyl-sulfoxide (DMSO) used by most organ banks for cryopreserving skin grafts and artificial skin substitutes. Keratinocyte (KC) viability, proliferation and marker expression following cryopreservation in trehalose were examined with similar results to those using DMSO. Trehalose concentration (0.4m) was optimized according to the described cellular activities following cryopreservation. Artificial epidermal substitutes were then cryopreserved in trehalose at the optimized concentration. Cell viability, growth factor secretion and wound healing properties of cryopreserved artificial epidermal substitutes using nude mice were examined and compared with those of DMSO cryopreservation. Cryopreservation with trehalose enhanced human KC viability in suspension and artificial skin substitutes. In addition, trehalose cryopreservation provided fast recovery of EGF and TGF-β1 secretion by KCs after thawing. When transplanted into nude mice, trehalose-cryopreserved artificial skin repaired skin defects in a similar manner to that of a non-cryopreserved control. Moreover, trehalose-cryopreserved artificial skin resulted in engraftment and wound closure that was significantly enhanced compared with that of DMSO-cryopreserved epidermal membranes. The results indicate that the use of trehalose improves cryopreservation of tissue-engineered epithelial sheets.
组织工程表皮膜在临床伤口愈合中很有用。为了在临床上应用这些产品，需要开发优化的储存方法。我们研究了不同浓度的细胞外海藻糖在冷冻保存人组织工程表皮膜方面的效率，与大多数器官库用于冷冻保存皮肤移植物和人工皮肤替代品的二甲亚砜（DMSO）相比。通过冷冻保存海藻糖，检查了角质形成细胞（KC）的活力、增殖和标记物表达，结果与使用 DMSO 相似。根据冷冻保存后描述的细胞活性，优化了海藻糖的浓度（0.4m）。然后将人工表皮替代品在优化浓度的海藻糖中冷冻保存。在裸鼠中检查并比较了冷冻保存的人工表皮替代品的细胞活力、生长因子分泌和伤口愈合特性与 DMSO 冷冻保存的特性。海藻糖冷冻保存增强了悬浮液中和人工皮肤替代品中的人 KC 活力。此外，海藻糖冷冻保存可使 KC 解冻后快速恢复 EGF 和 TGF-β1 的分泌。移植到裸鼠后，海藻糖冷冻保存的人工皮肤以与非冷冻保存对照相似的方式修复皮肤缺陷。此外，与 DMSO 冷冻保存的表皮膜相比，海藻糖冷冻保存的人工皮肤的植入和伤口闭合明显增强。结果表明，海藻糖的使用可改善组织工程上皮片的冷冻保存。
