Successful mouse hepatocyte culture with sandwich collagen gel formation.
10.4174/jkss.2013.84.4.202
- Author:
Hyun Jung CHOI
1
;
Dongho CHOI
Author Information
1. Department of Surgery, Soonchunhyang University Seoul Hospital, Soonchunhyang University College of Medicine, Seoul, Korea. dhchoi@schmc.ac.kr
- Publication Type:Original Article
- Keywords:
Collagen;
Culture;
Hepatocyte
- MeSH:
Adult;
Animals;
Bile Canaliculi;
Collagen;
Gels;
Gene Expression;
Glucose-6-Phosphatase;
Hepatocyte Nuclear Factor 4;
Hepatocytes;
Humans;
Longevity;
Mice;
Oxidoreductases;
Tissue Donors;
Tryptophan Oxygenase;
Tyrosine Transaminase
- From:Journal of the Korean Surgical Society
2013;84(4):202-208
- CountryRepublic of Korea
- Language:English
-
Abstract:
PURPOSE: 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. CONCLUSION: The collagen gel sandwich method is suitable for primary culture system of adult mouse hepatocytes.
