title: Successful mouse hepatocyte culture with sandwich collagen gel formation.
abstract: OBJECTIVE
  Primary mammalian hepatocytes largely retain their liver-specific functions when they are freshly derived from donors. However, long-term cultures of functional hepatocytes are difficult to establish. To increase the longevity and maintain the differentiated functions of hepatocytes in primary culture, cells can be cultured in a sandwich configuration of collagen. In such a configuration, hepatocytes can be cultured for longer periods compared with cultures on single layers of collagen. However, research regarding mouse hepatocytes in sandwich culture is lacking.
  
  
  METHODS
  Primary mouse hepatocytes were sandwiched between two layers of collagen to maintain the stability of their liver-specific functions. After gelation, 2 mL of hepatocyte culture medium was applied.
  
  
  RESULTS
  After 24 hours, 5, 10 days of culture, the collagen gel sandwich maintained the cellular border and numbers of bile canaliculi more efficiently than a single collagen coating in both high and low density culture dishes. Reverse transcription-polymerase chain reaction analysis of alpha-1-antitrypsin (AAT), hepatocyte nuclear factor 4 alpha (HNF4A), alphafetoprotein, albumin, tryptophan oxygenase (TO), the tyrosine aminotransferase gene, glucose-6-phosphatase, glyceraldehyde-3-phosphate dehydrogenase for mouse primary hepatocytes cultured on collagen coated dishes and collagen gels showed superior hepatocyte-related gene expression in cells grown using the collagen gel sandwich culture system. AAT, HNF4A, albumin, TO were found to be expressed in mouse hepatocytes cultured on collagen gels for 5 and 10 days. In contrast, mouse hepatocytes grown on collagen-coated dishes did not express these genes after 5 and 10 days of culture.
  
  
  CONCLUSIONS
  The collagen gel sandwich method is suitable for primary culture system of adult mouse hepatocytes.
authors.affiliation: Department of Surgery, Soonchunhyang University Seoul Hospital, Soonchunhyang University College of Medicine, Seoul, Korea.
authors.forename: Hyun Jung
authors.initials: HJ
authors.lastname: Choi
authors.forename: Dongho
authors.initials: D
authors.lastname: Choi
keywords: Collagen; Culture; Hepatocyte