scholarly journals Maintenance of total cytochrome P-450 content in rat hepatocyte culture and the abundance of CYP1A2 and CYP2B1/2 mRNAs

1992 ◽  
Vol 285 (3) ◽  
pp. 929-932 ◽  
Author(s):  
C R W Padgham ◽  
A J Paine ◽  
I R Phillips ◽  
E A Shephard
Keyword(s):  
Rat Hepatocytes ◽  
Culture Conditions ◽  
Dimethyl Sulphoxide ◽  
Hepatocyte Culture ◽  
Rat Hepatocyte ◽  
Aminolaevulinic Acid ◽  
Rat Hepatocyte Culture ◽  
Cytochrome P 450 ◽  
In Cells

mRNAs encoding cytochrome P-450s CYP1A2 and CYP2B1/2 have been quantified in rat hepatocytes cultured for periods up to 72 h under several different culture conditions that maintain total cytochrome P-450 content. When hepatocytes were cultured at either 37 or 30 degrees C in Williams E media, both CYP1A2 and CYP2B1/2 mRNAs declined dramatically. However, when cultured at 30 degrees C for 24 h, the decline in these mRNAs was not as great as that observed in cells grown at 37 degrees C. The addition of dimethyl sulphoxide to cells grown at 37 degrees C did not affect the rate of disappearance of the CYP1A2 or CYP2B1/2 mRNAs. These mRNAs also declined rapidly in cells grown in ‘P-450 medium’ i.e. RPMI 1640 medium without cyst(e)ine but supplemented with 0.1 mM-delta-aminolaevulinic acid. However, the levels of CYP2B1/2 mRNAs were maintained when hepatocytes were cultured in Williams E medium supplemented with 0.5 mM-metyrapone. These conditions did not, however, maintain the levels of CYP1A2 mRNA.

scholarly journals Immunochemical quantification of cytochrome P450IA and IIB subfamilies in the livers of metyrapone-treated rats. Relevance to the ability of metyrapone to prevent the loss of cytochrome P-450 in rat hepatocyte culture

1990 ◽  
Vol 267 (3) ◽  
pp. 715-719 ◽  
Author(s):  
K Shean ◽  
A J Paine
Keyword(s):  
Polyclonal Antibodies ◽  
Rat Hepatocytes ◽  
Microsomal Protein ◽  
Hepatocyte Culture ◽  
Rat Hepatocyte ◽  
Rat Hepatocyte Culture ◽  
Rat Livers ◽  
Cytochrome P 450 ◽  
Hepatic Cytochrome

Polyclonal antibodies to the major beta-naphthoflavone (BNF)-inducible form of cytochrome P-450 (P450IA) and to the major phenobarbitone (PB)-inducible form (P450IIB) have been used to quantify the contribution of these subfamilies to the total amount of cytochrome P-450 in rat livers and rat hepatocyte cultures treated with PB, BNF and metyrapone for 24 and 72 h. The P450IA and IIB subfamilies were not detectable (less than 5 pmol/mg of microsomal protein) in the livers of control rats, but administration of BNF resulted in the P450IA subfamily comprising more than 80% of the total hepatic cytochrome P-450. Administration of PB and metyrapone to rats did not elevate the level of this subfamily but elevated the levels of the P450IIB subfamily to 60% and 30% respectively of the total. Thus metyrapone is a ‘PB-like’ inducer. However, in contrast with their effects in vivo, treatment with PB and metyrapone of rat hepatocytes did not elevate the proportion of the P450IIB subfamily relative to that in untreated cells but rather, like BNF, increased the P450IA subfamily. This would account for the ability of metyrapone to produce in hepatocyte culture, like BNF, a pronounced induction of ethoxyresorufin O-de-ethylase activity, but it does not account for why of all inducers studied only metyrapone can maintain the total cytochrome P-450 content of cultured hepatocytes, or the activity of ethylmorphine N-demethylase. This activity is generally considered to be associated with the P450IIB subfamily, but the lack of effect of metyrapone on this subfamily in hepatocyte culture must suggest that metyrapone is able to prevent the loss of the total amount of the cytochrome by increasing the expression of other cytochromes P-450.

scholarly journals Dehydroepiandrosterone 3β-sulphate is an endogenous activator of the peroxisome-proliferation pathway: induction of cytochrome P-450 4A and acyl-CoA oxidase mRNAs in primary rat hepatocyte culture and inhibitory effects of Ca2+-channel blockers

1994 ◽  
Vol 301 (3) ◽  
pp. 753-758 ◽  
Author(s):  
P A Ram ◽  
D J Waxman
Keyword(s):  
Clofibric Acid ◽  
Hepatocyte Culture ◽  
Peroxisome Proliferator ◽  
Channel Blockers ◽  
Peroxisome Proliferation ◽  
Mrna Levels ◽  
Rat Hepatocyte ◽  
Acyl Coa Oxidase ◽  
Rat Hepatocyte Culture ◽  
Cytochrome P 450

The role of steroids related to the adrenal androgen dehydroepiandrosterone (5-androstene-3 beta-ol-17-one; DHEA) in regulating the expression of peroxisomal and cytochrome P-450 4A (CYP4A) enzymes active in fatty acid metabolism was assessed using a primary rat hepatocyte culture system. Exposure of hepatocytes to the peroxisome proliferator, clofibric acid (10-250 microM), for 48-96 h led to substantial increases in CYP4A protein, CYP4A1, CYP4A2 and CYP4A3 mRNAs, and the mRNAs encoding both forms of peroxisomal acyl-CoA oxidase (ACOX-I and ACOX-II), as judged by Northern-blot analysis using gene-specific oligonucleotide probes. Although DHEA treatment in vivo is effective in inducing these mRNAs in rat liver, it had no effect in the cultured hepatocytes. In contrast, treatment of the cells with DHEA 3 beta-sulphate (DHEA-S; 10-250 microM) stimulated major increases in CYP4A and ACOX mRNA levels. Examination of several analogues indicated a preference for 3 beta-sulphate over 17 beta-sulphated steroids and the inactivity of a 3 alpha-hydroxy-17 beta-sulphate derivative (DHEA-S > 5-androstene-3 beta,17 beta-diol 3-sulphate approximately 5 alpha-androstene-3 beta-ol-17-one 3-sulphate > 5-androstene-3 beta, 17 beta,17 beta-diol 17-sulphate approximately 5 beta-androstane-3 alpha-ol-17-one 3-sulphate >> 5 alpha-androstane-3 alpha, 17 beta-diol 17-sulphate). Induction of CYP4A mRNAs by either DHEA-S or clofibric acid was partially blocked by structurally diverse Ca(2+)-channel antagonists (nicardipine, nifedipine and diltiazem; 50 microM), suggesting that both the steroidal and fibrate classes of CYP4A inducers stimulate peroxisomal-proliferative responses via a Ca(2+)-dependent pathway. Retinoic acid alone slightly induced CYP4A mRNAs but did not enhance the induction by clofibrate or DHEA-S. As DHEA-S corresponds to a physiologically important major circulating androgen, these findings suggest that it may serve as an endogenous regulator of hepatic peroxisome enzyme levels. They further suggest that Ca(2+)-channel blockers may be useful pharmacological tools for the further study of the underlying cellular mechanism whereby endogenous steroids and fibrate drugs induce peroxisome proliferation, and the relationship of these events to activation of the peroxisome proliferator-activated receptor.

Warifteine and milonine, alkaloids isolated from Cissampelos sympodialis Eichl: cytotoxicity on rat hepatocyte culture and in V79 cells

2003 ◽  
Vol 142 (1-2) ◽  
pp. 143-151 ◽  
Author(s):  
Patricia Silva Melo ◽  
Horacinna Maria de Medeiros Cavalcante ◽  
José Maria Barbosa-Filho ◽  
Margareth de Fátima Formiga Melo Diniz ◽  
Isac Almeida de Medeiros ◽  
...  
Keyword(s):  
Hepatocyte Culture ◽  
V79 Cells ◽  
Rat Hepatocyte ◽  
Rat Hepatocyte Culture

Influence of Tumor Necrosis Factor-α and Silibin on the Cytotoxic Action of α-Amanitin in Rat Hepatocyte Culture

1999 ◽  
Vol 158 (3) ◽  
pp. 253-260 ◽  
Author(s):  
Claudia El-Bahay ◽  
Elisabeth Gerber ◽  
Martina Horbach ◽  
Quynh-Hoa Tran-Thi ◽  
Elke Röhrdanz ◽  
...  
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Tumor Necrosis Factor Α ◽  
Hepatocyte Culture ◽  
Cytotoxic Action ◽  
Rat Hepatocyte ◽  
Rat Hepatocyte Culture ◽  
Factor Α ◽  
Necrosis Factor
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