Solanas Estela, Sostres Carlos, Serrablo Alejandro, García-Gil Agustín, Aranguren Francisco, Jimenez Pilar, Serrano M Trinidad
1 IIS Aragón, CIBER Enfermedades Hepáticas y Digestivas (CIBERehd). Molecular Research Laboratory , Zaragoza, Spain .
Biopreserv Biobank. 2012 Oct;10(5):446-53. doi: 10.1089/bio.2012.0015.
Efficient cryopreservation of human hepatocytes is essential for their use in cell therapy. This study investigated the effects of adding melatonin and/or dimethyl sulfoxide (DMSO) to pre-incubation and/or freezing solutions on the viability and function of thawed human hepatocytes.
Isolated human hepatocytes were pre-incubated for 90 min at 4°C in Williams' Medium E (WEM), WEM containing 5 mM melatonin dissolved in DMSO, or WEM containing the equivalent amount of DMSO (1%). The hepatocytes were frozen in University of Wisconsin solution (UW) and 10% DMSO, with or without 5 mM melatonin. After thawing, viability, plating efficiency, mitochondrial dehydrogenase activity (MTT), and albumin and urea production were analyzed.
Viability and plating efficiency were not affected by melatonin or DMSO in pre-incubation media. Unexpectedly, hepatocytes pre-incubated with DMSO had significantly higher MTT (29.7% vs. control, p<0.01), albumin (82.8% vs. control, p<0.05), and urea amounts (26.2% vs. control, p=0.06) than those incubated only with WEM. Hepatocytes pre-incubated in media containing melatonin had amounts between those of cells incubated with DMSO or only with WEM (p<0.05 for MTT and p>0.05 for albumin and urea values). Also, the addition of melatonin to the freezing media did not significantly improve any of the studied parameters (p>0.05).
Adding 1% DMSO to pre-incubation media prior to the cryopreservation of human hepatocytes preserves hepatocyte function after thawing. These findings could be considered in current hepatocyte cryopreservation protocols.
