In this study a combined analysis of osmotic injury and cytotoxic effect useful for the optimization of the cryopreservation process of a cell suspension is carried out. The case of human Mesenchymal Stem Cells (hMSCs) from Umbilical Cord Blood (UCB) in contact with dimethyl sulfoxide (DMSO) acting as Cryo-Protectant Agent (CPA) is investigated from the experimental as well as the theoretical perspective. The experimental runs are conducted by suspending the cells in hypertonic solutions of DMSO at varying osmolality, system temperature, and contact times; then, at room temperature, cells are pelleted by centrifugation and suspended back to isotonic conditions. Eventually, cell count and viability are measured by means of a Coulter counter and flow-cytometer, respectively. Overall, a decrease in cell count and viability results when DMSO concentration, temperature, and contact time increase. A novel mathematical model is developed and proposed to interpret measured data by dividing the cell population between viable and nonviable cells. The decrease of cell count is ascribed exclusively to the osmotic injury caused by expansion lysis: excessive swelling causes the burst of both viable as well as nonviable cells. On the other hand, the reduction of cell viability is ascribed only to cytotoxicity which gradually transforms viable cells into nonviable ones. A chemical reaction engineering approach is adopted to describe the dynamics of both phenomena: by following the kinetics of two chemical reactions during cell osmosis inside a closed system it is shown that the simultaneous reduction of cell count and viability may be successfully interpreted. The use of the Surface Area Regulation (SAR) model recently proposed by the authors allows one to avoid the setting in advance of fixed cell Osmotic Tolerance Limits (OTLs), as traditionally done in cryopreservation literature to circumvent the mathematical simulation of osmotic injury. Comparisons between experimental data and theoretical simulations are provided: first, a nonlinear regression analysis is performed to evaluate unknown model parameters through a best-fitting procedure carried out in a sequential fashion; then, the proposed model is validated by full predictions of system behavior measured at operating conditions different from those used during the best-fit procedure.

中文翻译：

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与 DMSO 接触进行冷冻保存的 hMSCs：渗透损伤和细胞毒作用的实验和建模

在这项研究中，对渗透性损伤和细胞毒性作用进行了组合分析，可用于优化细胞悬液的冷冻保存过程。从实验和理论角度研究了来自脐带血 (UCB) 的人间充质干细胞 (hMSCs) 与作为冷冻保护剂 (CPA) 的二甲基亚砜 (DMSO) 接触的情况。实验运行是通过将细胞悬浮在不同渗透压、系统温度和接触时间的 DMSO 高渗溶液中进行的；然后，在室温下，细胞通过离心沉淀并悬浮回等渗条件。最终，细胞计数和活力分别通过库尔特计数器和流式细胞仪进行测量。全面的，当 DMSO 浓度、温度和接触时间增加时，细胞计数和活力会降低。开发并提出了一种新的数学模型，通过将细胞群划分为活细胞和非活细胞来解释测量数据。细胞计数的减少完全归因于膨胀裂解引起的渗透性损伤：过度肿胀会导致活细胞和非活细胞的爆裂。另一方面，细胞活力的降低仅归因于细胞毒性，细胞毒性逐渐将有活力的细胞转化为无活力的细胞。采用化学反应工程方法来描述这两种现象的动力学：通过跟踪封闭系统内细胞渗透过程中两种化学反应的动力学，可以成功地解释细胞数量和活力的同时减少。作者最近提出的表面积调节 (SAR) 模型的使用允许人们避免预先设置固定细胞渗透耐受极限 (OTL)，如传统的冷冻保存文献中所做的那样，以规避渗透损伤的数学模拟。提供了实验数据和理论模拟之间的比较：首先，进行非线性回归分析，通过以顺序方式执行的最佳拟合程序来评估未知模型参数；然后，