Viability and differential function of rainbow trout liver cells in primary culture: Coculture with two permanent fish cells |
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Authors: | S Scholz T Braunbeck H Segner |
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Institution: | (1) Institute of Zoology, University of Technology, D-01062 Dresden, FRG;(2) Present address: University of Technology, Institute of Zoology, D-01062 Dresden, FRG;(3) Department of Chemical Ecotoxicology, UFZ Centre of Environmental Research, D-04301 Leipzig, FRG;(4) Institute of Zoology, University of Heidelberg, D-69120 Heidelberg, FRG |
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Abstract: | Summary The study investigates the influence of different culture conditions on attachment, viability and functional status of rainbow
trout (Oncorhynchus mykiss) liver cells in primary culture. Cells were isolated by a two-step collagenase perfusion and incubated in serum-free, chemically
defined minimal essential medium (MEM), (a) as a monolayer on uncoated PRI-MARIA? dishes, (b) as a monolayer on culture dishes
coated with calf collagen type 1, and (c) in coculture with the established fish cell lines RTH-149 or RTG-2. Cell attachment was assessed from DNA and protein concentrations
per dish, viability was estimated from cellular lactate dehydrogenase release, and the metabolic status was investigated by
measuring activities of the phosphoenolpyruvate carboxykinase and biotransformation enzymes as well as the total cytochrome
P450 contents. Seeding of hepatocytes on collagen-coated dishes did not alter cell attachment or detachment from the culture
substrate, but had a small, but not significant effect on cell viability and metabolic parameters. Coculture of liver cells
and RTG-2 cells reduced hepatocyte detachment from the culture substrate, and it was associated with a significant elevation
of 7-ethoxyresorufin-O-deethylase activities in the hepatic cells. Cytochrome P450 contents, however, were not altered. The coculture effect on
liver cell physiology clearly depended on the type of cell line, because coculture with RTH-149 cells led to similar, but
much weaker effects than obtained in cocultures with RTG-2 cells. Electron microscopical observations revealed the existence
of gap junctions and possible exocytosis-like transport between cell lines and hepatocytes. The results point to the potential
of coculture systems to improve physiological parameters of trout liver cells in primary culture. |
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Keywords: | biotransformation coculture hepatocytes fish continuous cell lines RTG-2 RTH-149 |
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