Evaluation of the effect of culture matrices on induction of CYP3A isoforms in cultured porcine hepatocytes.
Several bioartificial liver devices have been developed as temporary therapy for patients suffering from fulminant hepatic failure. Some of these devices contain porcine hepatocytes entrapped in collagen matrices. In order to improve the function of these BAL devices, there exists a need to optimize metabolic function of cultured hepatocytes. The goal of these investigations was to evaluate the effect of altering culture conditions on rifampin-mediated induction of CYP3A isoforms in cultured porcine hepatocytes. Midazolam metabolism was compared in porcine hepatocytes cultured in a monolayer configuration on collagen gels, in a sandwich configuration between collagen gels and a Matrigel overlay, and in spheroidal cultures. The effect of culture conditions was evaluated, by measuring CYP3A-mediated metabolism of midazolam and by immunoblotting to detect CYP3A proteins, in control cultures and in rifampin-treated cultures. Results obtained by normalizing the metabolism rate data to cell numbers (based on DNA content) present at the end of the culture experiment, showed that there was no difference between the different culture conditions tested. Our results suggest that culturing porcine hepatocytes as spheroids or in a sandwich configuration between collagen and Matrigel, offers no advantage in terms of CYP3A-mediated metabolic function on a per cell basis compared to culturing on collagen gels.
Hosagrahara
V P
VP
Department of Medicinal Chemistry, University of Minnesota, 8-101 WDH 308 Harvard Street SE, Minneapolis, MN 55455, USA.
Hansen
L K
LK
Beilman
G J
GJ
Remmel
R P
RP
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D015153:Blotting, Western; D018929:Cell Culture Techniques / Q000379:methods*; D048429:Cell Size; D002478:Cells, Cultured; D003094:Collagen; D003470:Culture Media; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D004790:Enzyme Induction / Q000187:drug effects; D015151:Immunoblotting; D007527:Isoenzymes / Q000096:biosynthesis*; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D008874:Midazolam / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000096:biosynthesis*; D012293:Rifampin / Q000494:pharmacology; D013552:Swine
D003470:Culture Media; D007527:Isoenzymes; D003094:Collagen; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; D051544:Cytochrome P-450 CYP3A; D010089:Oxidoreductases, N-Demethylating; D008874:Midazolam; D012293:Rifampin