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KMID : 0614620030410010041
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Korean Journal of Gastroenterology
2003 Volume.41 No. 1 p.41 ~ p.48
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Cryopreservation of Rat Hepatocytes for the Use of Primary Culture
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Jung Yong-Hee
Kim Byung-Ho
Lee Byoung-Wook
Han Yo-Seb
Dong Seok-Ho
Kim Hyo-Jong
Chang Young-Woon
Lee Joung-Il
Chang Rin
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Abstract
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Background/Aims:The cryopreservation of primary hepatocytes could avoid unnecessary isolation of hepatocytes and supply them on demand. We compared two programs of computer-controlled freezing methods on the efficacy of hepatocyte cryopreservation.
Methods: Rat hepatocytes were cryopreserved by computer-controlled freezing methods. In program I, the overall cooling rate was 1.33℃/min and single-step shock cooling was used to reduce the latent heat of fusion. In program
II, the cooling rate of 2℃/min and two-step shock cooling were used. Two programs were compared in regard to hepatocyte viability and long-term cryopreservation and thawing temperature were also evaluated.
Results: The hepatocyte viability showed the highest value of 74.5±2.5% when cryopreserved using the program II and at 2×106/㎖ cell concentration in cryopreservation medium. The hepatocyte attachment rate on culture was similar in every occasion, more or less 50%. The hepatocyte viability was improved by 10% when thawed at 40℃, compared to the value at 37℃ in the program I. The hepatocyte viability was decreased to 38.2±5.5% in the program I and 45.4±1.6% in the program II after long-term cryopreservation.
Conclusion: The program II showed better survival of hepatocytes at 2×106/㎖ cell concentration. However, overall efficacy of hepatocyte cryopreservation was less than 35% and decreased more after long-term cryopreservation. Further studies are needed to develop a more effective program for hepatocyte cryopreservation.
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