Criocenter SRL, Buenos Aires, Argentina.
Centro de Investigaciones Endocrinológicas "Dr. César Bergadá" (CEDIE), CONICET - FEI - División de Endocrinología, Hospital de Niños R. Gutiérrez, Buenos Aires, Argentina.
Mol Biol Rep. 2021 Feb;48(2):1559-1565. doi: 10.1007/s11033-020-06079-x. Epub 2021 Jan 4.
Mesenchymal stem cells (MSC) differentiate into different cell types and have immunomodulatory and paracrine effects. Cryopreservation of umbilical cord tissue as a source of MSC is very promising for regenerative medicine. We aim to evaluate a protocol for cryopreserving this tissue sectioned into small fragments with viable MSC. A total of 723 samples were frozen, thawed and cultured to obtain primary cultures of MSC. These were followed until 90-100% confluence and flow cytometric analysis were performed to confirm the mesenchymal phenotype. Samples in which protocol alterations at the collection of the samples were reported, were excluded for microbial contamination analysis leaving a total of 634 samples composed of 181 vaginal and 453 cesarean births. All cultures reach confluence with a media of 22.57 days and 97% in 28 or fewer days. Evaluated cultures showed low percentage of CD45+ and high of CD73 and CD90. Eight samples were subcultured 4 or 5 times and differentiated to chondrocytes and osteocytes to test differentiation potential with positive results. Umbilical cord tissue collections showed similar microbial profile and risk factors to those reported of umbilical cord blood collections, but with higher contamination frequencies. Cryopreserved tissue samples had viable cells that can be expanded without losing differentiation potential. Higher contamination frequencies compared to umbilical cord blood collection are not surprising, however, microbial load and survival of microorganisms to cryopreservation are expected to be lower.
间充质干细胞（MSC）可分化为不同的细胞类型，具有免疫调节和旁分泌作用。冷冻保存脐带组织作为 MSC 的来源，对于再生医学非常有前途。我们旨在评估一种方案，即将含有活 MSC 的小块脐带组织进行冷冻保存。总共冷冻了 723 个样本，然后解冻并培养以获得 MSC 的原代培养物。这些培养物一直培养到 90-100%融合，并进行流式细胞术分析以确认间充质表型。对于在采集样本时报告了方案改变的样本，由于存在微生物污染分析，因此将其排除在外，总共得到了 634 个样本，其中 181 个为阴道分娩，453 个为剖宫产。所有培养物在 22.57 天的培养基中达到融合，97%在 28 天或更短的时间内达到融合。评估的培养物显示 CD45+的百分比较低，CD73 和 CD90 的百分比较高。8 个样本进行了 4 或 5 次传代培养，并分化为软骨细胞和成骨细胞，以测试分化潜能，结果为阳性。脐带组织采集的微生物谱和危险因素与脐带血采集相似，但污染频率更高。冷冻保存的组织样本中有可扩增的活细胞，而不会失去分化潜能。与脐带血采集相比，污染频率较高并不奇怪，但是预计微生物的负荷和对冷冻保存的微生物的存活能力会较低。
