Regulation of glutathione S‐transferase subunits 3 and 4 in cultured rat hepatocytes

Morel, Franck; Vandenberghe, Yves; Pemble, Sally; Taylor, John B.; Ratanasavanh, Damrong; Rogiers, Vera; Ketterer, Brian

doi:10.1016/0014-5793(89)81624-2

Cited by 17 publications

(13 citation statements)

References 22 publications

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“…In particular, changes in cytochrome P-450.dependent enzymatic activities and correlated gene expression have been frequently studied during recent years [26][27][28]. Some previous investigations demonstrate that the ghttathione S-transferase isoenzyme profile and RNA transcripts of the different subunlts also change when hepatocytes ale removed from their natural environment and that variations occur depending on the type of culture and media conditions used [15][16][17]19].…”

Section: Discussionmentioning

confidence: 99%

“…The aim of the present work was to study GST 1/2, 3/4 and 7 gene transcription in conventionally and cocultured adult rat hepatocytes using medium conditions investigated in previous studies where GST activity, protein levels and steady-state levels of mRNA were reported [15][16][17]. Transcription of the albumin gene was also assessed in isolated nuclei at different stages to serve as a reference.…”

Section: Discussionmentioning

confidence: 99%

“…Comparison with earlier studies [16,17] should identify whetlaer any correlation exist~ between the c~anging mRNA levels coding for the GST subunits 1/2, 3/4 and 7 and the transcriptional activity of the respective genes in cultured hepatocytes, or if the steady-state mRNA levels are the consequence of RNA stabilizat:on. A nuclear 'run-on' experiment has been performed on nuclei isolated from conventionally and co-cultured adult rat hepatocytes, maintained in standard medium or medium containing 10% fetal calf serum (FCS), 25 mM nicotinamide (Nie) or 2% dimethylsulphoxide (DMSO).…”

Section: Introductionmentioning

confidence: 94%

“…It has been demonstrated that OST enzymatic activity [14], subunit composition [15], de hOVe synthesis ofGST subunits in vitro [! 5] and steadystate mRNA levels, using complementary DNA sequences for GST subunits 1/2, 3/4 and 7 [16,17], vary depending on the culture and media conditions used.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptional‐ and post‐transcriptional‐dependent regulation of glutathione S‐transferase expression in rat hepatocytes as a function of culture conditions

et al. 1992

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Transcriptional activity of the glutathlone S-transfemse (GST) ~t (subumt~ 1 and 2),/~ (.~ubunit~ 3 and 4) :rod rt (~ubumt 7) gent fam~hes has been analyzed u~mg the nuclear "run-on' technique on adult rat hepatoeyte~ ,naintained for 4 days m convenuonal culture and for 4 and 12 days in co-culture with rat liver epithehal cells. Several medium conditions are included m tlus utudy, namely with or without fetal calf ~rum and with meotmamide or dlmethylnulphoxlde. Hepatocyte~ co-cultured for 4 days maintain approximatdy 30-70% of the a t~ene family transcriptional acuvity, whatever the medium conditions, when compared to freshly isolated hepatocytes. A marked decrease is observed after 12 days ofeo-culture or when hepatoeytes are maintained in convenuonal culture. The tmn~npuonal activity of the/~ gene family is saammiugd at 40-160% when hepatocytes are cultured with or without fetal calf serum, and is inducible by nicotinamidc (approximately 4-fold) and d:meth]llsulphoxlde (approximately 2-fold) m conventional culture and/or in co-culture. In contrast to freshly ~solated hepatocyte.% GST ~ gene transcriptional activity is observed in eouventLonal and go-cultured hepatoeytes, trrespeetive of the medium conditions. Dlmcthylsulphoxidc treatment however, represses the expression of GST 7 in vitro. These rehultn demonbtrate that the expression of GST 0~,/~ and rt genes m conventional and eo.eultured rat hepatoeytgs is controlled primarily at the level of transerq)tion. It cannot be excluded, however, that dimethylsulphoxide ntablllze~ the GST mRNA levels m vitro,

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 94%

Section: Introductionmentioning

confidence: 99%

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Transcriptional‐ and post‐transcriptional‐dependent regulation of glutathione S‐transferase expression in rat hepatocytes as a function of culture conditions

et al. 1992

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“…The increase in GST activity in vitro is due to an increase of GST subunits 1, 3 and especially 7, in both culture systems [2 11. Northern blot experiments indicate that induction of individual GST subunits by PB treatment in vitro, results from a corresponding increase in the amount of mRNA encoding the GST subunits [21,25]. Nuclear 'run-on' experiments were done to establish whether the elevation in GST l/2,3/4 and 7 mRNA levels is due to transcriptional activation or post-transcriptional changes.…”

Section: Discussionmentioning

confidence: 99%

Regulation of glutathione S‐transferase gene expression by phenobarbital in cultured adult rat hepatocytes

et al. 1991

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Previous studies, by using Northern blotting analyses, showed that phenobarbital (PB) affects the steady-state mRNA levels of glutathione S-transferase (GST) subunits l/2, 314 and 7 in both conventional cultures of adult rat hepatocytes and cotuttures, with rat liver epithelial cells [Vandenberghe ct al., 1989, FEW Lett. 2.51, 59-64;Morel et al., 1989, FEBS Lett. 258, 99-1021. To determine whether PB acts at the transcriptional level, nuclear 'run-on' experiments using cDNA probes hybridizing to GST subunits l/2, 3/4 and 7 mRNA were performed on purified nuclei isolated from control and PI3 treated hepatocytes seeded under conventional and co-culture conditions. Data from this study demonstrate that the increase in steady-state mRNA levels observed in both conventionai cuIture and co-culture after 4 days PB exposure results from an increased tmns~ptional activity of the GST genes. However, a substantial increase in steady-state mRNA levels in the absence of a commensurate increase in transcriptional activity at 12 days of co-culture, indicates that the barbiturate has also a skbilizing effect in vitro on the GST mRNAs.Glutathione S-transferase; Gene transcription; Rat hepatocyte; Primary culture; Phenobarbital 1, INTRODUCTIONPhenobarbital (PB) has previously been reported to alter the expression, regulation and activity of several liver drug metabolizing enzymes [l-3]. Most attention has been paid to its effect on cytochrome P-450 [4-61, UDP-glucuronyltransferase [7,8] and glutathione Stransferase enzymes [9-l 11, However, it is yet not clear how this compound affects the regulation of expression of these enzymes. It is not known whether PB induction of transcriptional and translational activities involves one or more receptors [12,13].It has been shown that PB does not elevate alf cytochrome P-450 enzymes to the same extent. Cytochrome P-450 IIBI and III32 are the major liver enzymes to be induced [ 14,151. The rapid increase in the rates of transcription and the appearance of elevated Ievels of nuclear pre-mRNA and cytoplasmi~ mRNAs unequivocaily demonstrated that PB elevates levels of cytochrome P-450 III& and IIBz primarily by augmenting th: rate of transcription of the respective genes Rat bcpatocytes were isolated from Sprague-Dawley rats (ISO-ZOO 11, 1348 Mont-St-Guibert, Belgium. Fax: (32) (10) 450290. g) as described by Gugucn et al. [23). I'hcy were sccdcd in convcnm result from an increased transcriptional activity as found for the intact liver. Liver glutathione ~-transferase subunits 1 and 3 especially are also induced by PB treatment Ill]. Recently, Pickett et al. [20] reported that the increase of GST mRNA in total liver after PB addition results from an elevated GST gene transcription. In cultured hepatocytes, PB treatment induces the GST enzymatic activity 121,223. It was shown in previous study [21] that, by supplementing the culture medium with 3.2 mM PB, concentrations of individual GST subunits and GST steady-state mRNA levels were changed. However, no information is available on the re...

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Reduced and oxidized glutathione contents in adult rat hepatocytes under various culture conditions

et al. 1991

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Reduced and oxidized glutathione contents of adult rat hepatocytes in pure culture and in co-culture with rat epithelial cells were measured under various medium conditions. To the standard medium fetal calf serum, nicotinamide, H2SeO3, dimethylsulphoxide or no supplements were added. For freshly isolated hepatocytes, intracellular contents of 24 +/- 7 nmol reduced and 0.7 +/- 0.2 nmol oxidized glutathione/mg cellular protein were obtained, respectively. In pure culture as well as in co-culture and regardless of the medium conditions involved, the protein content stays constant during the culture time with the exception of a decrease in protein content after 6 days of pure culture, caused by deterioration and loss of the hepatocytes. In both culture systems, an initial increase in intracellular reduced glutathione levels was observed, followed by a decrease and a quick normalisation in co-culture. On the contrary, in pure culture, the decrease was slower, but not transient and a stabilized situation was never reached. The various supplementations of the culture media had no significant effect on the intracellular reduced glutathione contents of both culture systems. As far as the intra- and the extracellular oxidized glutathione contents and the extracellular reduced form are concerned, these were only present in small amounts.

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Regulation of glutathione S‐transferase subunits 3 and 4 in cultured rat hepatocytes

Cited by 17 publications

References 22 publications

Transcriptional‐ and post‐transcriptional‐dependent regulation of glutathione S‐transferase expression in rat hepatocytes as a function of culture conditions

Transcriptional‐ and post‐transcriptional‐dependent regulation of glutathione S‐transferase expression in rat hepatocytes as a function of culture conditions

Regulation of glutathione S‐transferase gene expression by phenobarbital in cultured adult rat hepatocytes

Reduced and oxidized glutathione contents in adult rat hepatocytes under various culture conditions

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