title: Influence of temperature fluctuations during cryopreservation on vital parameters, differentiation potential, and transgene expression of placental multipotent stromal cells.
abstract: Successful implementation of rapidly advancing regenerative medicine approaches has led to high demand for readily available cellular suspensions. In particular, multipotent stromal cells (MSCs) of placental origin have shown therapeutic efficiency in the treatment of numerous pathologies of varied etiology. Up to now, cryopreservation is the only effective way to preserve the viability and unique properties of such cells in the long term. However, practical biobanking is often associated with repeated temperature fluctuations or interruption of a cold chain due to various technical, transportation, and stocking events. While biochemical processes are expected to be suspended during cryopreservation, such temperature fluctuations may lead to accumulation of stress as well as to periodic release of water fractions in the samples, possibly leading to damage during long-term storage.
  
  
  
  In this study, we performed a comprehensive analysis of changes in cell survival, vital parameters, and differentiation potential, 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 -196 °C to -100 °C in less than 20 cycles in comparison to constant temperature storage. However, increasing the temperature range to -80 °C 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. Differentiation potential of placental MSCs was not compromised after cryopreservation with constant end temperatures or with temperature fluctuations. However, regulation of various genes after cryopreservation procedures significantly varies. Interestingly, transgene expression was not compromised in any of the studied samples.
  
  
  
  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, differentiation potential and transgene expression are not compromised during studied storage conditions, while variation in gene regulation is observed.
authors.affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany. pogozhykh.denys@mh-hannover.de.|Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Pereyaslavskaya Str. 23, 61015, Kharkiv, Ukraine. pogozhykh.denys@mh-hannover.de.
authors.forename: Denys
authors.identifier: 0000-0001-7298-5425
authors.initials: D
authors.lastname: Pogozhykh
authors.affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.|Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Pereyaslavskaya Str. 23, 61015, Kharkiv, Ukraine.
authors.forename: Olena
authors.initials: O
authors.lastname: Pogozhykh
authors.affiliation: Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Pereyaslavskaya Str. 23, 61015, Kharkiv, Ukraine.
authors.forename: Volodymyr
authors.initials: V
authors.lastname: Prokopyuk
authors.affiliation: Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Pereyaslavskaya Str. 23, 61015, Kharkiv, Ukraine.
authors.forename: Larisa
authors.initials: L
authors.lastname: Kuleshova
authors.affiliation: Institute for Problems of Cryobiology and Cryomedicine, National Academy of Sciences of Ukraine, Pereyaslavskaya Str. 23, 61015, Kharkiv, Ukraine.
authors.forename: Anatoliy
authors.initials: A
authors.lastname: Goltsev
authors.affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Blasczyk
authors.affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany.|Synlab Medical Care Center Weiden Ltd., Zur Kesselschmiede 4, 92637, Weiden in der Oberpfalz, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Mueller
mesh_terms: D017667:Adipocytes / Q000166:cytology / Q000378:metabolism*; D000650:Amnion / Q000166:cytology / Q000378:metabolism; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D018070:Biological Specimen Banks / Q000592:standards; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation; D002470:Cell Survival; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D015870:Gene Expression; D006801:Humans; D046228:Microarray Analysis; D039902:Multipotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D010920:Placenta / Q000166:cytology / Q000378:metabolism*; D011247:Pregnancy; D013696:Temperature; D019076:Transgenes
chemical_list: D015703:Antigens, CD; D015415:Biomarkers
keywords: Amnion; Biobanking; Cryomicroscopy; Cryopreservation; MSC; Mimicking long-term storage; Placenta; Temperature fluctuations; Viability