Giga-I(3) Research, Hematology Unit, CHU of Liege and University of Liege, Liege, Belgium.
Cytotherapy. 2013 Nov;15(11):1385-94. doi: 10.1016/j.jcyt.2013.06.012.
The therapeutic potential of human mesenchymal stromal cells (MSCs) has generated considerable interest in a wide variety of areas. MSC banking is feasible, but the optimal technique of cryopreservation remains to be determined.
To reduce dimethyl sulfoxide (DMSO) concentration in cryopreservation medium, DMSO was replaced with sucrose or trehalose. To increase cell survival and proliferation rates after thawing and to eliminate the need for fetal bovine serum (FBS), neuropeptides of the vasoactive intestinal peptide/glucose-dependent insulinotropic peptide/pituitary adenylate cyclase activating polypeptide family were added to the cryopreservation medium. Cell survival was analyzed by a trypan blue dye exclusion assay. Cell proliferation of cryopreserved MSCs was determined after 7 days of culture.
No significant differences in cell survival rates were detected between cryopreservation solutions with 5% and 10% DMSO, independently of the addition of trehalose or sucrose. Cell proliferation rates tended to be highest when MSCs were frozen in 5% DMSO + trehalose. FBS could be replaced by human albumin (HA) without loss in cell survival and proliferation potential. With FBS, the addition of neuropeptides could increase cell survival and proliferation rates. Without FBS or HA, cell survival and proliferation rates in the presence of neuropeptides were comparable to rates achieved with FBS or HA.
Classic cryopreservation with 10% DMSO could be replaced by 5% DMSO + 30 mmol/L trehalose. FBS could be replaced by HA or neuropeptides without loss in cell survival and proliferation potential. The addition of neuropeptides in the cryopreservation medium containing FBS could increase the cell proliferation rate and consequently cellular output.
人类间充质基质细胞（MSC）的治疗潜力在许多领域引起了极大的兴趣。MSC 银行是可行的，但最佳的冷冻保存技术仍有待确定。
为了降低冷冻保存介质中二甲基亚砜（DMSO）的浓度，用蔗糖或海藻糖替代 DMSO。为了提高解冻后的细胞存活率和增殖率，并消除对胎牛血清（FBS）的需求，将血管活性肠肽/葡萄糖依赖性胰岛素释放肽/垂体腺苷酸环化酶激活肽家族的神经肽添加到冷冻保存介质中。通过台盼蓝排斥试验分析细胞存活率。培养 7 天后测定冷冻保存 MSC 的细胞增殖率。
无论是否添加海藻糖或蔗糖，5%和 10%DMSO 冷冻保存溶液之间的细胞存活率无显著差异。当 MSCs 在 5%DMSO+海藻糖中冷冻时，细胞增殖率倾向于最高。可以用人白蛋白（HA）代替 FBS 而不损失细胞的存活和增殖潜力。有 FBS 时，添加神经肽可以提高细胞存活率和增殖率。没有 FBS 或 HA 的情况下，神经肽存在时的细胞存活率和增殖率与使用 FBS 或 HA 时相当。
经典的 10%DMSO 冷冻保存可以用 5%DMSO+30mmol/L 海藻糖代替。可以用人白蛋白（HA）代替 FBS 或神经肽，而不损失细胞的存活和增殖潜力。在含有 FBS 的冷冻保存介质中添加神经肽可以提高细胞增殖率，从而增加细胞产量。
