Influence of temperature fluctuations during cryopreservation on vital parameters, plasticity, and transgene expression of placental multipotent stromal cells

In this study we performed comprehensive analysis of changes in cell survival, vital parameters, plasticity, as well as transgene expression of placental MSCs after temperature fluctuations within the liquid nitrogen steam storage, mimicking long-term preservation in practical biobanking, transportation, and temporal storage. It was shown that viability and metabolic parameters of placental MSCs did not significantly differ after temperature fluctuations in the range from -196ºC to -100ºC in less than 20 cycles in comparison to constant temperature storage. However, increasing of the temperature range to -80ºC as well as increasing the number of cycles, leads to significant lowering of these parameters after thawing. The number of apoptotic changes increases depending on the number of cycles of temperature fluctuations. Besides, adhesive properties of the cells after thawing are significantly compromised in the samples subjected to temperature fluctuations during storage. Plasticity of placental MSCs was not compromised neither after cryopreservation with constant end temperatures nor with temperature fluctuations. However, regulation of various genes after cryopreservation procedures significantly varies. Alterations in structural and functional parameters of placental MSCs after long-term preservation should be considered in practical biobanking due to potential temperature fluctuations in samples. At the same time, plasticity and transgene expression are not compromised during studied storage conditions, while certain gene regulation is observed. Part of the samples was cryopreserved with the constant end temperature, while the other part of the samples was subjected to repeated temperature fluctuations between -80ºC and -196ºC during cryopreservation. After thawing the samples were differentiated into three mesenchymal lineages with the following comparative evaluation of gene regulation. Five dual-colour microarrays were processed. Each sample was tested once.