Cryopreservation of primary human hepatocytes: The benefit of trehalose as an additional cryoprotective agent
Cryopreservation of primary human hepatocytes: The benefit of trehalose as an additional cryoprotective agent
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DOI：
10.1002/lt.20921
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发表时间：
2007-01-01
影响因子：
4.6
通讯作者：
Sauer, Igor Maximilian
中科院分区：
文献类型：
--
作者：
Katenz, Ekaterina;Vondran, Florian Wolfgang Rudolf;Sauer, Igor Maximilian
通过有效的冷冻保存技术和形成适当的细胞库，可以克服用于细胞移植的人肝细胞的有限可用性的问题。在这项研究中，我们研究了二糖海藻糖对人肝细胞冷冻保存的影响。为了进行分析，将肝细胞冷冻在含有10%二甲基亚砜（DMSO）并补充不同浓度海藻糖的培养基中。在解冻后培养期间，分析了活力、平板效率、总蛋白、细胞增殖、酶渗漏、白蛋白和尿素形成以及I相和II相代谢。在初步研究中，在研究的浓度中，0.2 M海藻糖显示出最佳的总体结果。与单独使用DMSO相比，我们发现海藻糖组的解冻后细胞活力（62.9 +/- 13 vs. 46.9 +/-11%，P < 0.01）和平板接种效率（41.5 +/- 18 vs. 17.6 +/-13%，P < 0.01）显著改善。使用海藻糖作为冷冻保存人肝细胞的添加剂导致解冻后贴壁细胞中总蛋白水平显著增加、白蛋白分泌增加和天冬氨酸转氨酶（AST）水平降低。总之，使用海藻糖作为冷冻保护剂显著改善了人肝细胞冷冻保存的结果。
Problems with the limited availability of human hepatocytes for cell transplantation may be overcome by efficient cryopreservation techniques and formation of appropriate cell banking. In this study we investigated the effect of the disaccharide trehalose on the cryopreservation of human hepatocytes. For analysis, liver cells were frozen in culture medium containing 10% dimethyl sulfoxide (DMSO) that was supplemented with varying concentrations of trehalose. During the postthawing culture period, viability, plating efficiency, total protein, cell proliferation, enzyme leakage, albumin and urea formation, as well as phase I and 11 metabolism were analyzed. In the pilot study, among the concentrations investigated, 0.2 M trehalose showed the best overall outcome. Compared to the use of DMSO alone, we found significant improvement in postthaw cell viability (62.9 +/- 13 vs. 46.9 +/- 11%, P < 0.01) and plating efficiency (41.5 +/- 18 vs. 17.6 +/- 13%, P < 0.01) in the trehalose group. The use of trehalose as an additive for cryopreserving human hepatocytes resulted in a significantly increased total protein level in the attached cells, higher secretion of albumin and a lower aspartate aminotransferase (AST) level after thawing. In conclusion, the use of trehalose as cryoprotective agent significantly improves the outcome of human hepatocyte cryopreservation.