Induction of catalase, alpha, and microsomal glutathione S-transferase in CYP2E1 overexpressing HepG2 cells and protection against short-term oxidative stress

Abstract

Induction of cytochrome P450 2E1 (CYP2E1) and the formation of reactive oxygen species (ROS) appear to be one of the mechanisms by which ethanol is hepatotoxic. Glutathione peroxidase and catalase detoxify H 2 O 2 . Glutathione Stransferases (GST) provide protection from membrane lipid peroxidation, have GSH peroxidase activity, and reduce lipid hydroperoxides. Previous studies showed an up-regulation of GSH synthesis in CYP2E1 expressing HepG2 cells; this finding prompted an evaluation of the levels of other antioxidant exzymes. In CYP2E1 expressing cells, the alpha and microsomal GST messenger RNA (mRNA) are increased by 4-and 2-fold, respectively, and catalase protein and mRNA is increased by 2-fold. The increase in alpha and microsomal GST mRNA correlates with increased total enzymatic activity and is caused by increased transcription as evidenced by run-on transcription assays. In HepG2 cells transfected to express a different cytochrome P450, CYP3A4, there was an increase in alpha GST. However, in contrast to the CYP2E1 expressing cells, neither microsomal GST nor catalase were induced, suggesting some specificity for CYP2E1. In agreement with an increased antioxidant defense system, the sensitivity to added prooxidants such as menadione, antimycin A, H 2 O 2 , and 4-hydroxynonenal was lower in the CYP2E1 expressing cells as compared with control cells. In conclusion, overexpression of CYP2E1 in HepG2 cells, besides elevating total GSH levels, also induces expression of catalase and alpha and microsomal GST. This induction confers resistance to the cells against several prooxidants and is suggested to reflect an adaptive response by the cells against CYP2E1-mediated oxidative stress. (HEPATOLOGY 2001;33:652-661.)The glutathione S-transferases (GST; EC 2.5.1.18) are a family of enzymes that catalyze the conjugation of glutathione (GSH) to a wide variety of hydrophobic molecules of both exogenous and endogenous origin. 1-3 The cytosolic GSTs are dimeric proteins of subunits having a molecular mass in the 22-to 28-kd range. On the basis of their amino acid sequence, 5 classes have been described, namely alpha, mu, pi, theta, and sigma. [4][5][6] In humans the alpha class, subunits A1 and A2, is the predominant GST accounting for about 85% of the total GST protein. The major GST subunits expressed in rat liver are the alpha class, subunits A1 and A2, and the mu class, subunits 3 and 4, 7,8 whereas most of the cytosolic GST in HepG2 cells is the alpha isozyme. 9 ␣,␤, unsaturated aldehydes, such as 4-hydroxy-2,3-nonenal (4-HNE) and other endogenous or exogenous aldehydes, are generated in response to a variety of oxidative stimuli such as free intracellular iron, 10 chronic ethanol consumption, 11,12 and treatment with anticancer drugs. 13 GST-mediated conjugation of 4-HNE to GSH is the major pathway for 4-HNE metabolism. 14,15 Among the numerous cytosolic GST isozymes, the alpha-class displays the highest activity toward lipid hydroperoxides and has been implicated in the protection against oxidative stress. [1...

Decreased expression of cytochrome P450 2E1 is associated with poor prognosis of hepatocellular carcinoma

“…Furthermore, an elevated level of CYP2E1 may also generate excessive reactive oxygen intermediates that cause damage to the surrounding cells. It has been shown that an increased CYP2E1 foci is associated with early state of transformation, 38 which is in accordance with our observation that high level of CYP2E1 in nontumor liver may promote the transformation of normal hepatocyte to tumor.…”

Mechanism of hepatoprotective action of methionine and composition “Metovitan” against a background of antituberculosis drugs administration to rats

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Cytochrome P450 2E1 participation in the pathogenesis of experimental metabolic syndrome in guinea pigs

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