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  • Title: Regulation of gamma-glutamyltranspeptidase in rat hepatocyte monolayer cultures.
    Author: Edwards AM.
    Journal: Cancer Res; 1982 Mar; 42(3):1107-15. PubMed ID: 6120759.
    Abstract:
    Hepatocytes isolated as a relatively pure population from normal adult rats were maintained in primary monolayer culture for 4 to 7 days on plastic dishes. Factors affecting activity of gamma-glutamyltranspeptidase (GGT) in culture were investigated to establish a basis for in vitro studies on carcinogen-induced changes in GGT. Freshly plated cultures contained few GGT-positive cells (0.4%) and very low total GGT activity, but with all media tested, activity increased progressively with time in culture, the extent of increase depending on medium composition. For hepatocytes in modified Waymouth medium alone (control cultures), there was a slow rise in activity after a 1- to 2-day lag period. This increase was enhanced up to 10-fold in the presence of 3 microM dexamethasone for cells on plastic but only 2- to 3-fold for cells on collagen gels. The time- and dose-dependent action of dexamethasone was prevented by cycloheximide, partly blocked by actinomycin D, and reversed after several hr by removing the steroid, when GGT activity decreased with a maximum half-life of 60 to 80 hr. These observations suggest that dexamethasone caused reversible enzyme induction dependent on continuing RNA and protein synthesis. Cell maintenance with fetal calf serum or with N6,O2'-dibutyryl adenosine 3':5'-monophosphate or glucagon markedly reduced the extent of induction by dexamethasone and slightly lowered activity in control cultures. Maintenance at lower pH within the range of 7.2 to 7.8 or at higher glucose concentrations within the range of 2.8 to 28 mM also resulted in markedly lower GGT activities in control or dexamethasone-induced cultures. Glucose repression was independent on insulin concentration. The reversible induction and repression of GGT may reflect broad changes in hepatocyte gene expression rather than specific controls of GGT function. Since GGT in cultured hepatocytes is subject to regulation by a variety of noncarcinogens, caution is required in its use as a preneoplastic marker. Nevertheless, the identification of culture media which preserve low activity may allow studies on carcinogen-induced changes in GGT as a probe of the early events in vitro hepatocarcinogenesis.
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