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Cytochrome P450 in highly purified suspension of nonparenchymal liver cells
Author(s) -
Rich K.,
Lodola A.
Publication year - 1989
Publication title -
cell biochemistry and function
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.933
H-Index - 61
eISSN - 1099-0844
pISSN - 0263-6484
DOI - 10.1002/cbf.290070406
Subject(s) - hepatocyte , biochemistry , chemistry , microbiology and biotechnology , cytochrome p450 , peroxidase , enzyme , alkaline phosphatase , western blot , liver cell , phenobarbital , biology , in vitro , medicine , endocrinology , gene
Rat liver nonparenchymal cells (NPC) were prepared by pronase digestion and purified on discontinuous gradients on Nycodenz. Morphological and biochemical characterization of cell suspensions showed that they were free of contamination by hepatocytes. We have confirmed the usefulness of pyruvate kinase activity in monitoring the degree of hepatocyte contamination of NPC and we have derived an equation which allows this carry‐over to be calculated. Using highly purified suspensions of NPC we have shown that they contain glucose‐6‐phosphatase in low but detectable levels. Spectrophotometric studies showed that they contain cytochrome P450, with a specific content of 24 +/− 5 pmole mg −1 cell protein. A potential source of error in previous studies was recognized; namely that peroxidase, present in NPC in high concentration, is able to mask the absorption due to cytochrome P450. Both the presence and inducibility of this enzyme in NPC prepared from rats pretreated with phenobarbital or 3‐methylcholanthrene have been confirmed using Western blot analysis.