Apoptosis occurs in isolated and banked primary mouse hepatocytes

T. Fu, D. Guo, X. Huang, M. R.G. O'Gorman, L. Huang, S. E. Crawford, H. E. Soriano*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

73 Scopus citations

Abstract

Isolation and cryopreservation of freshly isolated hepatocytes is considered a standard procedure for the long-term storage of liver cells. However, most existing methods for banking hepatocytes do not allow sufficient recovery of viable cells to meet the needs of basic research or clinical trials of hepatocyte transplantation. The mechanisms underlying this poor rate of hepatocyte recovery are unknown. Although much of the cellular damage in freezing is caused by formation of ice crystals within the cells, this is largely prevented by the use of dimethyl sulfoxide (DMSO) and controlled rate freezing. As we demonstrated recently, necrosis does occur in primary hepatocytes following isolation and cryopreservation. In the present study, we explored the contribution of apoptosis, another form of cell death, in primary hepatocytes banked for transplantation. We evaluated apoptosis of C57BL/6J mouse primary hepatocytes using several different methods. Annexin binding and the TUNEL assay, in conjunction with flow cytometry and confocal laser scanning microscopy, revealed that the percentage of apoptotic cells was dramatically elevated in cryopreserved cells compared with that in the control group of unfrozen cells. DNA laddering detected by DNA electrophoresis in agarose gel also supported the presence of apoptosis in isolated and banked liver cells. Moreover, we found that the addition of glucose (from 10 to 20 mM) into the freezing solution (University of Wisconsin Solution) decreased the rate of apoptosis by 84% and improved the cell attachment at least fourfold in cryopreserved cells. These results suggest that apoptosis might contribute to cell death in isolated and banked primary hepatocytes.

Original languageEnglish (US)
Pages (from-to)59-66
Number of pages8
JournalCell transplantation
Volume10
Issue number1
DOIs
StatePublished - 2001

Keywords

  • Apoptosis
  • Cell attachment
  • Cryostorage
  • Lactate dehydrogenase release
  • Liver
  • Necrosis

ASJC Scopus subject areas

  • Biomedical Engineering
  • Cell Biology
  • Transplantation

Fingerprint

Dive into the research topics of 'Apoptosis occurs in isolated and banked primary mouse hepatocytes'. Together they form a unique fingerprint.

Cite this