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The effects of over expressing aquaporins on the cryopreservation of hepatocytes

Kumar, Balasubramanian K., Coger, Robin N., Schrum, Laura W., Lee, Charles Y.
Cryobiology 2015 v.71 no.2 pp. 273-278
aquaporins, cell membranes, cell viability, cryopreservation, cyclic AMP, fluorescence microscopy, fluorescent antibody technique, freezers, glucagon, hepatocytes, rats, shrinkage
During cryopreservation, aquaporins are critical in regulating water transport across cellular membranes and preventing osmotic damages. Hepatocytes express aquaporin (AQP) 0, 8, 9, 11, and 12; this study investigates whether increasing the localization of AQP8 on the cellular membrane would improve cell viability by increasing water transport during cryopreservation. Primary rat hepatocytes were cultured and treated with dibutyryl cAMP (Bt2cAMP) or glucagon to increase the expression of AQP8 at the cellular membrane via translocation. This phenomenon is verified through two experiments – confocal immunofluorescence microscopy and cell shrinkage analysis. The immunofluorescence results showed increase in AQP8 on the cellular membrane of treated cells, and cell shrinkage analysis showed an increase in water transport of treated cells compared to controls. Primary rat hepatocytes were treated with Bt2cAMP or glucagon and cryopreserved using standard protocols in a controlled rate freezer. This resulted in a significant increase in the cell viability on warming. These results indicate that Bt2cAMP or glucagon treated hepatocytes had increased expression of aquaporin in the cellular membrane, increased water transport during cryopreservation, and increased post-thaw viability.