Health & Environmental Research Online (HERO)


Print Feedback Export to File
2278530 
Journal Article 
Primary cultures of adult mouse and rat hepatocytes for studying the metabolism of foreign chemicals 
Dougherty, KK; Spilman, SD; Green, CE; Steward, AR; Byard, JL 
1980 
Yes 
Biochemical Pharmacology
ISSN: 0006-2952
EISSN: 1873-2968 
HEEP/81/02540 
29 
15 
2117-2124 
English 
7406924 
WOS:A1980KD30000006 
HEEP COPYRIGHT: BIOL ABS. A primary hepatocyte culture was developed as a model system to investigate the metabolism of foreign chemicals. Hepatocytes were prepared from adult male Charles River CD-1 mice and adult male Sprague-Dawley rats by in situ pre-perfusion of the liver with ethyleneglycol-bis(beta-amino-ethyl ether) N,N'-tetraacetic acid (EGTA) followed by perfusion with Ca and collagenase. The digested liver was dispersed, and hepatocytes were isolated by filtration and differential centrifugation yielding 108 hepatocytes per mouse liver and 5hepatocytes per rat liver. More than 90% of the hepatocytes excluded trypan blue. Hepatocytes were prepared aseptically, plated on tissue culture dishes coated with rodent tail collagen (2.5Waymouth's medium. Within 4 h the hepatocytes attached to the collagen, and by 24 h they had flattened and formed a monolayer. A non-metabolizable Ala analog, alpha-aminoisobutyric acid, accumulated in mouse hepatocytes with peak incorporation occurring at 24 h. Cultured mouse and rat hepatocytes were able to N-demethylate p-chloro-N-methylaniline (PCMA). An NADPH-generating system stimulated N-demethylation 2.75-fold in freshly isolated mouse hepatocytes, but did not stimulate metabolism in cultured mouse hepatocytes. SKF 525-A (proadifen hydrochloride) inhibited PCMA N-demethylation in cultured mouse hepatocytes with a median inhibitory concentration of 3.75 | 10-5 M. Hormonal supplementation of the culture medium stimulated PCMA metabolism measured in 24 and 48 h cultures. These studies demonstrate the utility of rodent hepatocyte cultures as models of hepatic metabolism of foreign chemicals.