1988 Fiscal Year Final Research Report Summary
Tissue culture of fish hepatocytes
| Project/Area Number |
62560202
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
General fisheries
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| Research Institution | Nihon University |
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Principal Investigator |
WATANABE Tasuku College of Agriculture and Veterinary Medicine, Nihon University, 農獣医学部, 助教授 (30095517)
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| Project Period (FY) |
1987 – 1988
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| Keywords | Tissue Culture / hepatocyte(s) / Salmonid fish / Growth factor(s) / Rainbow trout Yamame / Liver / Embryo / 胚 / 血清蛋白 |
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| Research Abstract |
The primary cultures of rainbow trout liver cells were performed in TC199 medium supplemented with 10% fetal bovine serum and with 10ng/ml of epidermal growth factor (EGF) and/or 40nM of insulin. Epithelial cells grown in a flask supplemented with EGF and insulin, designated RTL-4, could be transferred at the 40th day of the primary culture and have been subcultured more than 100 times. The cells got fast doubling time gradually, and were able to be transferred at intervals of 7-10 days. ETL-4 grow in the manner of contact inhibited monolayer and consist of large epithelial cells with a round nucleus and abundant granular organella. Electron microscopically, most of the cells were large and round with a round nucleus having a large electron dense nucleolus at the center of nucleus and had well developed rough surfaced endoplasmic reticulum and mitochondria. RTL-4 had as high a susceptibility to IPNV as that of SE-A10 and had a tenth or less susceptibility to IHNV than SE-A10, when examined by plaque assay.
The cell culture of Yamame liver was also performed using the embryonated eggs. A continuous cell line designated YEL-13 was established and has been subcultured more than 120 times over the period of 3 years. YEL-13 grow in the manner of contact inhibited monolayer and consisted of epithelial cells and a few fibroblastic cells. Electron microscopically, most of the cells had a large and round nucleus and well developed rough surfaced endoplasmic reticulum in cytoplasm and smooth cell surface with poorly developed microvilli. Chromosome number of YEL-13 ranged widely and most of them were between 53 and 83. The cells with 56 chromosomes occupied 24% of those examined. YEL-13 cells have permissible temperature range of 15-20C for growth. The cell line has been producing and releasing a Yamame serum protein constantly which is one of the main functions of liver parenchymal cells.
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