Effects of various medium formulations and attachment substrata on the performance of cultured ruminant hepatocytes in biotransformation studies.
1. A procedure for the isolation and primary culture of hepatocytes from goat and cattle is described. Hepatocyte culture performance was monitored for 51 h by measuring viability, cytochrome P-450 maintenance, dealkylation of scoparone and ethylmorphine, and glucuronidation of phenol red. 2. Culture medium composition is discussed in relation to differences between splanchnic blood composition of ruminant and monogastric animal species. Main differences are in glucose and volatile fatty acid concentrations. Modified Williams' E culture medium did not yield higher culture performance than non-modified Williams' E. 3. Coating of culture dishes with either collagen or fibronectin did not improve culture performance. 4. Williams' E, although developed for rodent cells, proves to be a suitable basal medium for ruminant hepatocytes. In this medium, culture quality is high for at least several days. 5. In cultured goat hepatocytes, biotransformation rate for scoparone amounted to 20 nmol/mg protein per h, for ethylmorphine 96 nmol/mg protein per h and for phenol red 2 nmol/mg protein per h. Biotransformation activity in cow hepatocytes is approximately half that in goat hepatocytes.
van 't Klooster
G A
GA
University of Utrecht, Department of Veterinary Basic Sciences, The Netherlands.
Woutersen-van Nijnanten
F M
FM
Klein
W R
WR
Blaauboer
B J
BJ
Noordhoek
J
J
van Miert
A S
AS
D000818:Animals; D001711:Biotransformation; D002417:Cattle; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000494:pharmacology*; D003374:Coumarins / Q000378:metabolism / Q000493:pharmacokinetics*; D003470:Culture Media; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D005036:Ethylmorphine / Q000378:metabolism / Q000493:pharmacokinetics*; D005227:Fatty Acids / Q000494:pharmacology; D005260:Female; D005353:Fibronectins / Q000494:pharmacology*; D005947:Glucose / Q000494:pharmacology; D006041:Goats; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D008297:Male; D008954:Models, Biological; D010637:Phenolsulfonphthalein / Q000378:metabolism / Q000493:pharmacokinetics*; D011506:Proteins / Q000378:metabolism; D013997:Time Factors
D003374:Coumarins; D003470:Culture Media; D005227:Fatty Acids; D005353:Fibronectins; D011506:Proteins; D003094:Collagen; D003577:Cytochrome P-450 Enzyme System; D007770:L-Lactate Dehydrogenase; C018145:scoparone; D010637:Phenolsulfonphthalein; D005947:Glucose; D005036:Ethylmorphine