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Improvement of Hepatocyte Viability After Cryopreservation by Supplementation of Long-Chain Oligosaccharide in the Freezing Medium in Rats and Humans

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Abstract:

Factors affecting cell viability, plating efficiency, and survival of hepatocytes after cryopreservation have been investigated. We focused especially on the effect of including trehalose and related oligosaccharides in the cryopreservation fluid. This was supplemented with either glucose, trehalose, maltotriose, or other sugars, in addition to dimethyl sulfoxide (10%) and first tested with primary rat hepatocytes cooled in a controlled rate freezer. After thawing, viability by trypan blue exclusion of cells frozen in oligosaccharide-supplemented medium was significantly higher than for those cryopreserved without oligosaccharides. Use of oligosaccharides with higher molecular weights resulted in greatest improvement in viability. Moreover, attachment and survival rates in plastic dishes were approximately 1.2–1.8-fold greater after freezing in the presence of di-, tri-, and tetrasaccharides. Human hepatocytes isolated from untransplantable liver showed the same tendency regarding viability, but cell adherence was not similarly improved by the addition of oligosaccharides. Possible reasons for these differences may be prior cell damage during extended cold ischemia of the human liver, donor age, or cell degradation caused by progression of fatty liver in humans, and/or species differences.

Keywords: Freeze; Hepatocytes; Oligosaccharide; Preservation; Sugar

Document Type: Research Article

DOI: http://dx.doi.org/10.3727/000000006783981404

Affiliations: 1: Department of Innovative Surgery, National Research Institute for Child Health and Development, Tokyo 157-8535, Japan 2: Research Laboratories, HAB Research Organization, Ichikawa General Hospital, Chiba 272-8523, Japan 3: Department of Animal Science, Tokyo University of Agriculture, Kanagawa 243-0034, Japan

Publication date: October 1, 2006

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  • Cell Transplantation publishes original, peer-reviewed research and review articles on the subject of cell transplantation and its application to human diseases. To ensure high-quality contributions from all areas of transplantation, separate section editors and editorial boards have been established. Articles deal with a wide range of topics including physiological, medical, preclinical, tissue engineering, and device-oriented aspects of transplantation of nervous system, endocrine, growth factor-secreting, bone marrow, epithelial, endothelial, and genetically engineered cells, among others. Basic clinical studies and immunological research papers are also featured. To provide complete coverage of this revolutionary field, Cell Transplantation will report on relevant technological advances, and ethical and regulatory considerations of cell transplants. Cell Transplantation is now an Open Access journal starting with volume 18 in 2009, and therefore there will be an inexpensive publication charge, which is dependent on the number of pages, in addition to the charge for color figures. This will allow work to be disseminated to a wider audience and also entitle the corresponding author to a free PDF, as well as prepublication of an unedited version of the manuscript.
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